Reference Type:  Journal Article
Record Number: 470
Author: Adamany, A. M., Blumenfeld, O. O., Sabo, B. and McCreary, J.
Year: 1983
Title: A carbohydrate structural variant of MM glycoprotein (glycophorin A)
Journal: J Biol Chem
Volume: 258
Issue: 19
Pages: 11537-45
Epub Date: 1983/10/10
Date: Oct 10
Short Title: A carbohydrate structural variant of MM glycoprotein (glycophorin A)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6619126
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/analysis
Erythrocyte Membrane/*analysis
Female
*Genetic Variation
Glycophorins/*genetics/isolation & purification
Hemagglutination
Humans
Oligosaccharides/*isolation & purification
Peptide Fragments/analysis
Sialic Acids/analysis
Sialoglycoproteins/*genetics
Abstract: A variant of the MM glycoprotein (glycophorin A) was isolated from erythrocyte membranes of two individual donors, a mother (L.G.) and daughter (V.W.). This glycoprotein was found to be a carbohydrate variant in which, for both donors, certain O-glycosidically linked saccharides retained the core structure consisting of NeuAc(alpha 2,3)Gal(beta 1,3)GalNAc that is common to all O-linked saccharides of the MN glycoproteins, and, in addition, contained substituents, of varying chain lengths, on the primary carbinol of GalNAc. These saccharides were released from the polypeptide by beta-elimination in the presence of sodium borohydride, and aspects of their structure were investigated by glycosidase digestion and periodate oxidation. Thus, the smallest variant structure was deduced to be NeuAc(alpha 2,3)Gal(beta 1,3)[GlcNAc(beta 1,6)]H2GalNAc. The 6-O-linked GlcNAc appears to serve as the focus of further chain elongation reactions, involving alternate additions of Gal and GlcNAc residues and leading to the formation of several homologous structures. Two such structures, NeuAc(alpha 2,3)Gal(beta 1,3)[GlcNAc(beta 1,?) Gal(beta 1,3/4)GlcNAc(beta 1,6)]H2GalNAc and NeuAc(alpha 2,3) Gal(beta 1,3)[Gal(beta 1,3/4)GlcNAc(beta 1,6)]H2GalNAc were the predominant species present. A larger saccharide was also isolated and its partial sequence was determined to be Gal(beta 1,3/4)GlcNAc(beta 1,?)[Gal(beta 1,3/4)Glc-NAc(beta 1,?)] Gal(beta 1,3/4)GlcNAc(beta 1,6)[NeuAc(alpha 2,3)Gal-(beta 1,3)]H2GalNAc. Because the peptide portion of these glycoproteins contains two methionine residues, it was possible to isolate two CNBr glycopeptides from separate regions of the molecule, and to assess the distribution of these variant structures in the polypeptide. The saccharides were linked to about 2-3 Ser and/or Thr residues in the donor LG glycoprotein and one of the attachment sites was located within the CNBr glycooctapeptide representing the NH2 terminus. Considerable heterogeneity in saccharide structure was documented for this site, and it is likely that such heterogeneity occurs also at other sites. The variant saccharides bear structural similarities to the core region of O-linked saccharides of certain blood group-active mucins and ovarian cyst secretions, and to the outer sequences of N-linked carbohydrate units (I-, i-active) of the major glycoprotein of human erythrocytes, band 3. The structures of the variant saccharides suggest that they may be potential precursors of H blood group-active carbohydrates, present in varying degrees of maturity, and attached to an integral protein of erythrocytes.
Notes: Adamany, A M
Blumenfeld, O O
Sabo, B
McCreary, J
GM 16389/GM/NIGMS NIH HHS/United States
HL 19011/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1983 Oct 10;258(19):11537-45.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 307
Author: Adolf, G. R., Kalsner, I., Ahorn, H., Maurer-Fogy, I. and Cantell, K.
Year: 1991
Title: Natural human interferon-alpha 2 is O-glycosylated
Journal: Biochem J
Volume: 276 ( Pt 2)
Issue: Pt 2
Pages: 511-8
Epub Date: 1991/06/01
Date: Jun 1
Short Title: Natural human interferon-alpha 2 is O-glycosylated
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2760511
PMCID: PMC1151121
Accession Number: 2049076
Keywords: Amino Acid Sequence
Antibodies, Monoclonal
Chromatography, Affinity
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Enzyme-Linked Immunosorbent Assay
Glycoside Hydrolases
Glycosylation
Humans
Interferon Type I/*chemistry/isolation & purification
Molecular Sequence Data
Molecular Weight
Monosaccharides/analysis
Peptide Fragments/isolation & purification
Peptide Mapping
Abstract: Natural human interferon alpha 2 (IFN-alpha 2) was isolated from a preparation of partially purified human leucocyte IFN by monoclonal-antibody immunoaffinity chromatography. The purified protein had a specific activity of 1.5 x 10(8) i.u./mg; it was estimated to constitute 10-20% of the total antiviral activity of leucocyte IFN. N-Terminal amino-acid-sequence analysis identified the subspecies IFN-alpha 2b and/or IFN-alpha 2c, whereas IFN-alpha 2a was not detectable. The structure of natural IFN-alpha 2 was found to differ from that of its recombinant (Escherichia coli-derived) equivalent. First, reverse-phase h.p.l.c. showed that natural IFN-alpha 2 was significantly more hydrophilic then expected. Secondly, the apparent molecular mass of the natural protein determined by SDS/PAGE was higher than that of recombinant IFN-alpha 2; incubation under mild alkaline conditions known to eliminate O-linked carbohydrates resulted in a reduction of the apparent molecular mass to that of the recombinant protein. On sequence analysis of proteolytic peptides, Thr-106 was found to be modified. These results suggested that Thr-106 of natural IFN-alpha 2 carries O-linked carbohydrates. Reverse-phase h.p.l.c. as well as SDS/PAGE of natural IFN-alpha 2 showed that glycosylation is heterogeneous. For characterization of the carbohydrate moieties, the protein was treated with neuraminidase and/or O-glycanase and analysed by gel electrophoresis; in addition, glycopeptides obtained by proteinase digestion and separated by h.p.l.c. were characterized by sequence analysis and m.s. Further information on the composition of the glycans was obtained by monosaccharide analysis. The results indicate that natural IFN-alpha 2 contains the disaccharide galactosyl-N-acetylgalactosamine (Gal-GalNAc) linked to Thr-106. In part of the molecules, this core carbohydrate carries (alpha-)N-acetylneuraminic acid, whereas a disaccharide, probably N-acetyl-lactosamine, is bound to Gal-GalNAc in another proportion of the protein. Further glycosylation isomers are present in small amounts. As IFN-alpha 2 is the only IFN-alpha species with a threonine residue at position 106, it may represent the only O-glycosylated human IFN-alpha protein.
Notes: 1470-8728
Adolf, G R
Kalsner, I
Ahorn, H
Maurer-Fogy, I
Cantell, K
Journal Article
Biochem J. 1991 Jun 1;276 ( Pt 2)(Pt 2):511-8. doi: 10.1042/bj2760511.
Author Address: Ernst-Boehringer-Institut für Arzneimittelforschung, Bender + Co Ges mbH, Vienna, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 270
Author: Adolf, G. R., Maurer-Fogy, I., Kalsner, I. and Cantell, K.
Year: 1990
Title: Purification and characterization of natural human interferon omega 1. Two alternative cleavage sites for the signal peptidase
Journal: J Biol Chem
Volume: 265
Issue: 16
Pages: 9290-5
Epub Date: 1990/06/05
Date: Jun 5
Short Title: Purification and characterization of natural human interferon omega 1. Two alternative cleavage sites for the signal peptidase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1693148
Keywords: Amino Acid Sequence
Antibodies, Monoclonal
Chromatography, Affinity
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Endopeptidases/*metabolism
Glycosylation
Humans
Immunoassay
Interferon Type I/*isolation & purification/metabolism
Interferons/*analysis
Isoelectric Point
Leukocytes/*analysis
*Membrane Proteins
Molecular Sequence Data
Molecular Weight
Parainfluenza Virus 1, Human/physiology
Protein Sorting Signals/metabolism
*Serine Endopeptidases
Abstract: Human interferon omega 1 (IFN-omega 1 = IFN-alpha II1) is a recently discovered protein structurally related to IFN-alpha and -beta; the biological activities of IFN-omega 1 and its physiological role are not known to date. We have purified IFN-omega 1 from preparations of human leukocyte IFN, derived from peripheral blood leukocytes induced with Sendai virus, by two sequential cycles of monoclonal antibody affinity chromatography. The resulting protein was at least 95% pure as determined by reverse-phase high performance liquid chromatography and showed an Mr of 24,500 upon sodium dodecyl sulfate-gel electrophoresis (theoretical Mr, 19,984). Amino acid sequence analysis revealed that only about 40% of the molecules have the NH2 terminus expected on the basis of the sequence similarity to IFN-alpha, whereas the others contain two additional amino acids. This difference probably results from variable cleavage of the pre-protein by the signal peptidase. No evidence for COOH-terminal heterogeneity was found. Essentially all IFN-omega 1 molecules are glycosylated; enzymatic deglycosylation resulted in a reduction of the Mr to 20,500. Experiments using several plant lectins indicated the presence of biantennary complex oligosaccharides containing neuraminic acid. Two major peaks were observed upon chromatofocusing, with isoelectric points of 8.1 and 8.5. The specific antiviral activity of purified IFN-omega 1 assayed on human cells was determined to be 2.7 x 10(8) IU/mg, similar to that of other human class I IFNs; potent antiviral activity was also observed on cells of bovine and ovine but not of equine or murine origin.
Notes: Adolf, G R
Maurer-Fogy, I
Kalsner, I
Cantell, K
Comparative Study
Journal Article
United States
J Biol Chem. 1990 Jun 5;265(16):9290-5.
Author Address: Ernst Boehringer-Institut für Arzneimittelforschung, Bender & Co Ges mbH, Vienna, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 553
Author: Aeed, P. A. and Elhammer, A. P.
Year: 1994
Title: Glycosylation of recombinant prorenin in insect cells: the insect cell line Sf9 does not express the mannose 6-phosphate recognition signal
Journal: Biochemistry
Volume: 33
Issue: 29
Pages: 8793-7
Epub Date: 1994/07/26
Date: Jul 26
Short Title: Glycosylation of recombinant prorenin in insect cells: the insect cell line Sf9 does not express the mannose 6-phosphate recognition signal
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00195a022
Accession Number: 8038170
Keywords: Animals
Carbohydrate Sequence
Cell Line
Enzyme Precursors/*metabolism
Glycosylation
Humans
In Vitro Techniques
Lysosomes/metabolism
Molecular Sequence Data
Moths/*enzymology
Protein Precursors/metabolism
Protein Processing, Post-Translational
Receptor, IGF Type 2/metabolism
Recombinant Proteins/*metabolism
Renin/*metabolism
Transferases (Other Substituted Phosphate Groups)/metabolism
Abstract: Sf9 cells infected with a recombinant baculovirus containing the gene for human prorenin were cultured in the presence of [3H]mannose. In vivo labeled prorenin was isolated by immunoprecipitation from the culture medium and digested with Pronase. The oligosaccharide structures on the resulting glycopeptides were analyzed by a combination of lectin, ion-exchange, paper, and high-pressure liquid chromatography. Of the N-linked oligosaccharides isolated from the Sf9-produced prorenin, 98% were of a truncated (trimannosyl) high-mannose type, approximately two-thirds of which contained a fucose residue linked to the reducing N-acetylglucosamine. The remaining 2% constituted a mixture of high-mannose-type structures containing six, seven, or eight mannose residues; none of these structures were core-fucosylated. None of the oligosaccharide structures recovered from recombinant prorenin synthesized by Sf9 cells were phosphorylated or contained any other form of charge. Furthermore, assays for UDP-GlcNAc-lysosomal-enzyme N-acetylglucosamine phosphotransferase demonstrated no activity above background in lysates prepared from Sf9 cells. Blotting of Sf9 cell lysates with an 125I-labeled, soluble form of the cation-independent mannose 6-phosphate receptor failed to detect any proteins carrying the mannose 6-phosphate recognition signal. Taken together, the data suggest that Sf9 cells do not synthesize high-mannose-type oligosaccharides containing mannose 6-phosphate, and consequently it appears unlikely that these cells utilize the mannose 6-phosphate receptor mediated pathway for targeting of lysosomal enzymes.
Notes: Aeed, P A
Elhammer, A P
Journal Article
United States
Biochemistry. 1994 Jul 26;33(29):8793-7. doi: 10.1021/bi00195a022.
Author Address: Department of Biochemistry, Upjohn Company, Kalamazoo, Michigan 49001.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 267
Author: Aeed, P. A., Guido, D. M., Mathews, W. R. and Elhammer, A. P.
Year: 1992
Title: Characterization of the oligosaccharide structures on recombinant human prorenin expressed in Chinese hamster ovary cells
Journal: Biochemistry
Volume: 31
Issue: 30
Pages: 6951-61
Epub Date: 1992/08/04
Date: Aug 4
Short Title: Characterization of the oligosaccharide structures on recombinant human prorenin expressed in Chinese hamster ovary cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00145a013
Accession Number: 1637829
Keywords: Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography
Chromatography, Affinity
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Cricetinae
DNA/genetics
Enzyme Precursors/*chemistry/isolation & purification
Glycoside Hydrolases/metabolism
Humans
Mannose/analysis/chemistry
Methylation
Molecular Sequence Data
Oligosaccharides/analysis/*chemistry
Recombinant Proteins/chemistry/isolation & purification
Renin/*chemistry/isolation & purification
Sialic Acids/analysis/chemistry
Transfection
Abstract: Prorenin was isolated by immunoprecipitation from the culture medium of Chinese hamster ovary cells transfected with a human prorenin cDNA. The N-linked oligosaccharide structures on the in vivo [3H]mannose-labeled, purified protein were characterized using a combination of serial lectin affinity chromatography, high-pressure liquid chromatography, ion-exchange chromatography, and size-exclusion chromatography and treatment with specific glycosidases and methylation analysis. Approximately 61% of the oligosaccharides on the molecule are complex type, in the form of tetraantennary (2%), 2,6-branched triantennary (13%), 2,4-branched triantennary (3%), and biantennary (43%) structures. The majority of all complex type structures are core-fucosylated. Sialic acids are linked at the C-3 position of terminal galactose, and the degree of sialylation of the bi- and triantennary structures varies between nonsialylated and fully sialylated; no tetraatennary structure contains more than three sialic acid residues. Recombinant prorenin contains 4% hybrid-type structures, all of which carry a terminal sialic acid residue. The remaining 35% of the structures on the molecule are high mannose type, composed of 5, 6, or 7 mannose residues. Approximately 6% of the high mannose type structures and 10% of the hybrid structures are phosphorylated, as judged by their susceptibility to treatment with alkaline phosphatase. Compositional analysis of an unlabeled preparation of the protein suggested the presence of approximately 1.4 oligosaccharide units per molecule.
Notes: Aeed, P A
Guido, D M
Mathews, W R
Elhammer, A P
Journal Article
United States
Biochemistry. 1992 Aug 4;31(30):6951-61. doi: 10.1021/bi00145a013.
Author Address: Upjohn Laboratories, Upjohn Company, Kalamazoo, Michigan 49001.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 982
Author: Aertgeerts, K., Levin, I., Shi, L., Snell, G. P., Jennings, A., Prasad, G. S., Zhang, Y., Kraus, M. L., Salakian, S., Sridhar, V., Wijnands, R. and Tennant, M. G.
Year: 2005
Title: Structural and kinetic analysis of the substrate specificity of human fibroblast activation protein alpha
Journal: J Biol Chem
Volume: 280
Issue: 20
Pages: 19441-4
Epub Date: 2005/04/06
Date: May 20
Short Title: Structural and kinetic analysis of the substrate specificity of human fibroblast activation protein alpha
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.C500092200
Accession Number: 15809306
Keywords: Amino Acid Motifs
Catalytic Domain/genetics
Crystallography, X-Ray
Dipeptides/chemistry/metabolism
Dipeptidyl Peptidase 4/chemistry/genetics/metabolism
Fluorescent Dyes
Gelatinases/chemistry/genetics/*metabolism
Humans
In Vitro Techniques
Kinetics
Membrane Proteins/chemistry/genetics/*metabolism
Models, Molecular
Mutagenesis, Site-Directed
Protein Conformation
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Serine Endopeptidases/chemistry/genetics/*metabolism
Substrate Specificity
Abstract: Fibroblast activation protein alpha (FAPalpha) is highly expressed in epithelial cancers and has been implicated in extracellular matrix remodeling, tumor growth, and metastasis. We present the first high resolution structure for the apoenzyme as well as kinetic data toward small dipeptide substrates. FAPalpha exhibits a dipeptidyl peptidase IV (DPPIV)-like fold, featuring an alpha/beta-hydrolase domain and an eight-bladed beta-propeller domain. Known DPPIV dipeptides are cleaved by FAPalpha with an approximately 100-fold decrease in catalytic efficiency compared with DPPIV. Moreover, FAPalpha, but not DPPIV, possesses endopeptidase activity toward N-terminal benzyloxycarbonyl (Z)-blocked peptides. Comparison of the crystal structures of FAPalpha and DPPIV revealed one major difference in the vicinity of the Glu motif (Glu(203)-Glu(204) for FAPalpha; Glu(205)-Glu(206) for DPPIV) within the active site of the enzyme. Ala(657) in FAPalpha, instead of Asp(663) as in DP-PIV, reduces the acidity in this pocket, and this change could explain the lower affinity for N-terminal amines by FAPalpha. This hypothesis was tested by kinetic analysis of the mutant FAPalpha/A657D, which shows on average an approximately 60-fold increase in the catalytic efficiency, as measured by k(cat)/K(m), for the cleavage of dipeptide substrates. Furthermore, the catalytic efficiency of the mutant is reduced by approximately 350-fold for cleavage of Z-Gly-Pro-7-amino-4-methylcoumarin. Our data provide a clear understanding of the molecular determinants responsible for the substrate specificity and endopeptidase activity of FAPalpha.
Notes: Aertgeerts, Kathleen
Levin, Irena
Shi, Lihong
Snell, Gyorgy P
Jennings, Andy
Prasad, G Sridhar
Zhang, Yanming
Kraus, Michelle L
Salakian, Shant
Sridhar, Vandana
Wijnands, Robert
Tennant, Mike G
Comparative Study
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Biol Chem. 2005 May 20;280(20):19441-4. doi: 10.1074/jbc.C500092200. Epub 2005 Apr 4.
Author Address: Takeda San Diego, Inc., California 92121, USA. kaertgeerts@takedasd.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 563
Author: Agarwala, K. L., Kawabata, S., Takao, T., Murata, H., Shimonishi, Y., Nishimura, H. and Iwanaga, S.
Year: 1994
Title: Activation peptide of human factor IX has oligosaccharides O-glycosidically linked to threonine residues at 159 and 169
Journal: Biochemistry
Volume: 33
Issue: 17
Pages: 5167-71
Epub Date: 1994/05/03
Date: May 3
Short Title: Activation peptide of human factor IX has oligosaccharides O-glycosidically linked to threonine residues at 159 and 169
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00183a021
Accession Number: 8172892
Keywords: Amino Acid Sequence
Amino Acids/analysis
Animals
Carbohydrate Sequence
Cattle
Chromatography, High Pressure Liquid
Dogs
Factor IX/*chemistry/*metabolism
Factor IXa/*chemistry/metabolism
Glycopeptides/*chemistry/isolation & purification
Humans
Mice
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/*chemistry/isolation & purification
Rabbits
Sequence Homology, Amino Acid
Spectrometry, Mass, Fast Atom Bombardment
*Threonine
Abstract: O-Linked oligosaccharide chains were identified in the activation peptide (AP) of human blood coagulation factor IX. The peptide obtained from human factor IX was separated into three molecular species (AP alpha, AP beta, and AP gamma) by reversed-phase high-performance liquid chromatography. Amino acid analysis showed that AP alpha, but not AP beta and AP gamma, contained galactosamine in addition to glucosamine, thereby suggesting the presence of an O-linked sugar chain(s) in the molecule of AP alpha. A nonapeptide (AP alpha-D4, residues 157-165) and an undecapeptide (AP alpha-D5, 166-176) derived from AP alpha contained Thr-159 and Thr-169, neither of which could be identified using a gas-phase protein sequencer. All other serine and threonine residues present in AP alpha were identified by peptide sequencing. Component sugar and sialic acid analyses of AP alpha-D4 and AP alpha-D5 revealed that they contained 1 mol each of N-acetyl-D-galactosamine (GalNAc), D-galactose (Gal), and sialic acid. Fast atom bombardment tandem mass spectrometric analysis of AP alpha-D4 suggested the existence of Gal-GalNAc-Thr, NeuNAc-(Gal-)GalNAc-Thr, and NeuNAc-Gal-GalNAc-Thr structures. On the basis of amino acid analysis after the isolation of AP alpha, it accounted for approximately 35% of the total activation peptide obtained. From these results, it was concluded that a part of the activation peptide of human factor IX in circulating blood has tri- and tetrasaccharides O-glycosidically linked to the threonine residues at 159 and 169.
Notes: Agarwala, K L
Kawabata, S
Takao, T
Murata, H
Shimonishi, Y
Nishimura, H
Iwanaga, S
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1994 May 3;33(17):5167-71. doi: 10.1021/bi00183a021.
Author Address: Department of Molecular Biology, Graduate School of Medical Science, Kyushu University, Fukuoka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1629
Author: Aguilar, H., Fricovsky, E., Ihm, S., Schimke, M., Maya-Ramos, L., Aroonsakool, N., Ceballos, G., Dillmann, W., Villarreal, F. and Ramirez-Sanchez, I.
Year: 2014
Title: Role for high-glucose-induced protein O-GlcNAcylation in stimulating cardiac fibroblast collagen synthesis
Journal: Am J Physiol Cell Physiol
Volume: 306
Issue: 9
Pages: C794-804
Epub Date: 2014/02/21
Date: May 1
Short Title: Role for high-glucose-induced protein O-GlcNAcylation in stimulating cardiac fibroblast collagen synthesis
Alternate Journal: American journal of physiology. Cell physiology
ISSN: 0363-6143 (Print)
0363-6143
DOI: 10.1152/ajpcell.00251.2013
PMCID: PMC4010805
Accession Number: 24553187
Keywords: Acetylglucosaminidase/genetics/metabolism
Animals
Arginase/metabolism
Cells, Cultured
Collagen/*biosynthesis
Diabetic Cardiomyopathies/*metabolism/pathology
Fibroblasts/*metabolism/pathology
Glucose/*metabolism
Glycosylation
Male
Myocardium/*metabolism/pathology
*Protein Processing, Post-Translational
Rats
Rats, Sprague-Dawley
Signal Transduction
Smad Proteins/metabolism
Sp1 Transcription Factor/metabolism
Time Factors
Transfection
Transforming Growth Factor beta1/metabolism
Up-Regulation
cardiomyopathy
diabetes
fibroblast
fibrosis
Abstract: Excess enzyme-mediated protein O-GlcNAcylation is known to occur with diabetes mellitus. A characteristic of diabetic cardiomyopathy is the development of myocardial fibrosis. The role that enhanced protein O-GlcNAcylation plays in modulating the phenotype of cardiac fibroblasts (CF) is unknown. To address this issue, rat CF were cultured in normal glucose (NG; 5 mM glucose) or high-glucose (HG; 25 mM) media for 48 h. Results demonstrate that CF cultured in HG have higher levels (~50%) of overall protein O-GlcNAcylation vs. NG cells. Key regulators of collagen synthesis such as transforming-growth factor-β1 (TGF-β1), SMADs 2/3, and SMAD 7 protein levels, including those of arginase I and II, were altered, leading to increases in collagen levels. The nuclear transcription factor Sp1 and arginase II evidence excess O-GlcNAcylation in HG cells. Expression in CF of an adenovirus coding for the enzyme N-acetylglucosaminidase, which removes O-GlcNAc moieties from proteins, decreased Sp1 and arginase II O-GlcNAcylation and restored HG-induced perturbations in CF back to NG levels. These findings may have important pathophysiological implications for the development of diabetes-induced cardiac fibrosis.
Notes: 1522-1563
Aguilar, Hugo
Fricovsky, Eduardo
Ihm, Sang
Schimke, Magdalena
Maya-Ramos, Lisandro
Aroonsakool, Nakon
Ceballos, Guillermo
Dillmann, Wolfgang
Villarreal, Francisco
Ramirez-Sanchez, Israel
P60-MD-00220/MD/NIMHD NIH HHS/United States
R01-HL-43617/HL/NHLBI NIH HHS/United States
R24-DK-092154/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Am J Physiol Cell Physiol. 2014 May 1;306(9):C794-804. doi: 10.1152/ajpcell.00251.2013. Epub 2014 Feb 19.
Author Address: Seccion de Posgrado, Escuela Superior de Medicina, Instituto Politecnico Nacional, Ciudad de Mexico, Mexico.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1435
Author: Ahn, V. E., Chu, M. L., Choi, H. J., Tran, D., Abo, A. and Weis, W. I.
Year: 2011
Title: Structural basis of Wnt signaling inhibition by Dickkopf binding to LRP5/6
Journal: Dev Cell
Volume: 21
Issue: 5
Pages: 862-73
Epub Date: 2011/10/18
Date: Nov 15
Short Title: Structural basis of Wnt signaling inhibition by Dickkopf binding to LRP5/6
Alternate Journal: Developmental cell
ISSN: 1534-5807 (Print)
1534-5807
DOI: 10.1016/j.devcel.2011.09.003
PMCID: PMC3215855
NIHMSID: NIHMS326472
Accession Number: 22000856
Keywords: Binding Sites
Humans
Intercellular Signaling Peptides and Proteins/chemistry/*metabolism
Low Density Lipoprotein Receptor-Related Protein-5/chemistry/*metabolism
Low Density Lipoprotein Receptor-Related Protein-6/chemistry/*metabolism
Models, Molecular
Protein Conformation
Wnt Proteins/antagonists & inhibitors/chemistry/*metabolism
*Wnt Signaling Pathway/drug effects
Abstract: LDL receptor-related proteins 5 and 6 (LRP5/6) are coreceptors for Wnt growth factors, and also bind Dkk proteins, secreted inhibitors of Wnt signaling. The LRP5/6 ectodomain contains four β-propeller/EGF-like domain repeats. The first two repeats, LRP6(1-2), bind to several Wnt variants, whereas LRP6(3-4) binds other Wnts. We present the crystal structure of the Dkk1 C-terminal domain bound to LRP6(3-4), and show that the Dkk1 N-terminal domain binds to LRP6(1-2), demonstrating that a single Dkk1 molecule can bind to both portions of the LRP6 ectodomain and thereby inhibit different Wnts. Small-angle X-ray scattering analysis of LRP6(1-4) bound to a noninhibitory antibody fragment or to full-length Dkk1 shows that in both cases the ectodomain adopts a curved conformation that places the first three repeats at a similar height relative to the membrane. Thus, Wnts bound to either portion of the LRP6 ectodomain likely bear a similar spatial relationship to Frizzled coreceptors.
Notes: 1878-1551
Ahn, Victoria E
Chu, Matthew Ling-Hon
Choi, Hee-Jung
Tran, Denise
Abo, Arie
Weis, William I
R01 AG039420-01/AG/NIA NIH HHS/United States
R21 AG033596-01/AG/NIA NIH HHS/United States
R21 AG033596-02/AG/NIA NIH HHS/United States
R01 AG039420/AG/NIA NIH HHS/United States
Canadian Institutes of Health Research/Canada
R01AG39420/AG/NIA NIH HHS/United States
R21 AG033596-01S1/AG/NIA NIH HHS/United States
R21 AG033596/AG/NIA NIH HHS/United States
R01 AG039420-02/AG/NIA NIH HHS/United States
R21AG33596/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Dev Cell. 2011 Nov 15;21(5):862-73. doi: 10.1016/j.devcel.2011.09.003. Epub 2011 Oct 13.
Author Address: Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 768
Author: Ailor, E., Takahashi, N., Tsukamoto, Y., Masuda, K., Rahman, B. A., Jarvis, D. L., Lee, Y. C. and Betenbaugh, M. J.
Year: 2000
Title: N-glycan patterns of human transferrin produced in Trichoplusia ni insect cells: effects of mammalian galactosyltransferase
Journal: Glycobiology
Volume: 10
Issue: 8
Pages: 837-47
Epub Date: 2000/08/06
Date: Aug
Short Title: N-glycan patterns of human transferrin produced in Trichoplusia ni insect cells: effects of mammalian galactosyltransferase
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/10.8.837
Accession Number: 10929010
Keywords: Animals
Base Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
DNA Primers
Galactosyltransferases/*metabolism
Humans
Molecular Sequence Data
Moths/*cytology
Polysaccharides/*metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Transferrin/chemistry/genetics/*metabolism
Tunicamycin/pharmacology
Abstract: The N-glycans of human serum transferrin produced in Trichopulsia ni cells were analyzed to examine N-linked oligosaccharide processing in insect cells. Metabolic radiolabeling of the intra- and extracellular protein fractions revealed the presence of multiple transferrin glycoforms with molecular weights lower than that observed for native human transferrin. Consequently, the N-glycan structures of transferrin in the culture medium were determined using three-dimensional high performance liquid chromatography. The attached oligosaccharides included high mannose, paucimannosidic, and hybrid structures with over 50% of these structures containing one fucose, alpha(1,6)-, or two fucoses, alpha(1,6)- and alpha(1,3)-, linked to the Asn-linked N-acetylglucosamine. Neither sialic acid nor galactose was detected on any of the N-glycans. However, when transferrin was coexpressed with beta(1,4)-galactosyltransferase three additional galactose-containing hybrid oligosaccharides were obtained. The galactose attachments were exclusive to the alpha(1, 3)-mannose branch and the structures varied by the presence of zero, one, or two attached fucose residues. Furthermore, the presence of the galactosyltransferase appeared to reduce the number of paucimannosidic structures, which suggests that galactose attachment inhibits the ability of hexosaminidase activity to remove the terminal N-acetylglucosamine. The ability to promote galactosylation and reduce paucimannosidic N-glycans suggests that the oligosaccharide processing pathway in insect cells may be manipulated to mimic more closely that of mammalian cells.
Notes: Ailor, E
Takahashi, N
Tsukamoto, Y
Masuda, K
Rahman, B A
Jarvis, D L
Lee, Y C
Betenbaugh, M J
R01 GM049734/GM/NIGMS NIH HHS/United States
Journal Article
England
Glycobiology. 2000 Aug;10(8):837-47. doi: 10.1093/glycob/10.8.837.
Author Address: Department of Chemical Engineering, The Johns Hopkins University, Baltimore, MD 21218, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1004
Author: Airenne, T. T., Nymalm, Y., Kidron, H., Smith, D. J., Pihlavisto, M., Salmi, M., Jalkanen, S., Johnson, M. S. and Salminen, T. A.
Year: 2005
Title: Crystal structure of the human vascular adhesion protein-1: unique structural features with functional implications
Journal: Protein Sci
Volume: 14
Issue: 8
Pages: 1964-74
Epub Date: 2005/07/28
Date: Aug
Short Title: Crystal structure of the human vascular adhesion protein-1: unique structural features with functional implications
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.051438105
PMCID: PMC2279308
Accession Number: 16046623
Keywords: Amine Oxidase (Copper-Containing)/*chemistry/metabolism
Amino Acid Sequence
Binding Sites
Cell Adhesion Molecules/*chemistry/metabolism
Crystallography, X-Ray
Dimerization
Glycosylation
Humans
*Models, Molecular
Molecular Sequence Data
Protein Structure, Tertiary
Abstract: The expression of human vascular adhesion protein-1 (hVAP-1) is induced at sites of inflammation where extravasation of lymphocytes from blood to the peripheral tissue occurs. We have solved the X-ray structure of hVAP-1, a human copper amine oxidase (CAO), which is distinguished from other CAOs in being membrane-bound. The dimer structure reveals some intriguing features that may have fundamental roles in the adhesive and enzymatic functions of hVAP-1, especially regarding the role of hVAP-1 in inflammation, lymphocyte attachment, and signaling. Firstly, Leu469 at the substrate channel may play a key role in controlling the substrate entry; depending on its conformation, it either blocks or gives access to the active site. Secondly, sugar units are clearly observed at two of the six predicted N-glycosylation sites. Moreover, mutagenesis analysis showed that all of the predicted sites were glycosylated in the protein used for crystallization. Thirdly, the existence of a solvent-exposed RGD motif at the entrance to each active site in hVAP-1 suggests that it may have a functional role.
Notes: 1469-896x
Airenne, Tomi T
Nymalm, Yvonne
Kidron, Heidi
Smith, David J
Pihlavisto, Marjo
Salmi, Marko
Jalkanen, Sirpa
Johnson, Mark S
Salminen, Tiina A
Journal Article
Research Support, Non-U.S. Gov't
Protein Sci. 2005 Aug;14(8):1964-74. doi: 10.1110/ps.051438105.
Author Address: Department of Biochemistry and Pharmacy, Abo Akademi University, FIN-20520 Turku, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1713
Author: Airola, M. V., Allen, W. J., Pulkoski-Gross, M. J., Obeid, L. M., Rizzo, R. C. and Hannun, Y. A.
Year: 2015
Title: Structural Basis for Ceramide Recognition and Hydrolysis by Human Neutral Ceramidase
Journal: Structure
Volume: 23
Issue: 8
Pages: 1482-1491
Epub Date: 2015/07/21
Date: Aug 4
Short Title: Structural Basis for Ceramide Recognition and Hydrolysis by Human Neutral Ceramidase
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2015.06.013
PMCID: PMC4830088
NIHMSID: NIHMS705988
Accession Number: 26190575
Keywords: Amino Acid Sequence
Animals
Catalytic Domain
Ceramides/*chemistry/metabolism
Crystallography, X-Ray
Escherichia coli/chemistry
Humans
Hydrolysis
Hydrophobic and Hydrophilic Interactions
Kinetics
Ligands
Lysophospholipids/*chemistry/metabolism
Molecular Docking Simulation
Molecular Sequence Data
Neutral Ceramidase/*chemistry/genetics/metabolism
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Alignment
Sf9 Cells
Species Specificity
Sphingosine/*analogs & derivatives/*chemistry/metabolism
Spodoptera
Abstract: Neutral ceramidase (nCDase) catalyzes conversion of the apoptosis-associated lipid ceramide to sphingosine, the precursor for the proliferative factor sphingosine-1-phosphate. As an enzyme regulating the balance of ceramide and sphingosine-1-phosphate, nCDase is emerging as a therapeutic target for cancer. Here, we present the 2.6-Å crystal structure of human nCDase in complex with phosphate that reveals a striking, 20-Å deep, hydrophobic active site pocket stabilized by a eukaryotic-specific subdomain not present in bacterial ceramidases. Utilizing flexible ligand docking, we predict a likely binding mode for ceramide that superimposes closely with the crystallographically observed transition state analog phosphate. Our results suggest that nCDase uses a new catalytic strategy for Zn(2+)-dependent amidases, and generates ceramide specificity by sterically excluding sphingolipids with bulky headgroups and specifically recognizing the small hydroxyl head group of ceramide. Together, these data provide a foundation to aid drug development and establish common themes for how proteins recognize the bioactive lipid ceramide.
Notes: 1878-4186
Airola, Michael V
Allen, William J
Pulkoski-Gross, Michael J
Obeid, Lina M
Rizzo, Robert C
Hannun, Yusuf A
R01 GM097741/GM/NIGMS NIH HHS/United States
R01 CA172517/CA/NCI NIH HHS/United States
F32GM100679/GM/NIGMS NIH HHS/United States
T32GM007518/GM/NIGMS NIH HHS/United States
R37 GM043825/GM/NIGMS NIH HHS/United States
T32 GM007518/GM/NIGMS NIH HHS/United States
P01 CA097132/CA/NCI NIH HHS/United States
F32 GM100679/GM/NIGMS NIH HHS/United States
I01 BX000156/BX/BLRD VA/United States
R01GM097741/GM/NIGMS NIH HHS/United States
R01CA172517/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Structure. 2015 Aug 4;23(8):1482-1491. doi: 10.1016/j.str.2015.06.013. Epub 2015 Jul 16.
Author Address: Department of Medicine, Stony Brook University, Stony Brook, NY 11794, USA; Department of Medicine, Stony Brook Cancer Center, Stony Brook, NY 11794, USA.
Department of Applied Mathematics and Statistics, Stony Brook University, Stony Brook, NY 11794, USA.
Department of Pharmacological Sciences, Stony Brook University, Stony Brook, NY 11794, USA.
Department of Medicine, Stony Brook University, Stony Brook, NY 11794, USA; Department of Medicine, Stony Brook Cancer Center, Stony Brook, NY 11794, USA; Department of Medicine, Northport Veterans Affairs Medical Center, Northport, NY 11768, USA.
Department of Medicine, Stony Brook University, Stony Brook, NY 11794, USA; Department of Medicine, Stony Brook Cancer Center, Stony Brook, NY 11794, USA. Electronic address: yusuf.hannun@stonybrookmedicine.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 475
Author: Akiyama, K., Simons, E. R., Bernasconi, P., Schmid, K., van Halbeek, H., Vliegenthart, J. F., Haupt, H. and Schwick, H. G.
Year: 1984
Title: The structure of the carbohydrate units of human plasma galactoglycoprotein determined by 500-megahertz 1H NMR spectroscopy
Journal: J Biol Chem
Volume: 259
Issue: 11
Pages: 7151-4
Epub Date: 1984/06/10
Date: Jun 10
Short Title: The structure of the carbohydrate units of human plasma galactoglycoprotein determined by 500-megahertz 1H NMR spectroscopy
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6725284
Keywords: Carbohydrate Sequence
*Glycoproteins
Humans
Magnetic Resonance Spectroscopy
Abstract: Human plasma galactoglycoprotein (Mr = 81,000) which was recently identified and characterized (Schmid, K., Mao, S. K. Y., Kimura, A., Hayashi, S., and Binette , J. P. (1980) J. Biol. Chem. 255, 3221-3226) was found to possess an unusually high carbohydrate content (76%) comprised of NeuAc, Gal, Man, Fuc, GalNAc, and GlcNAc. The aim of the present investigation was to elucidate the primary structure of the oligosaccharide units of this protein. For the study of the O-glycosidic oligosaccharide chains, the protein was subjected to beta-elimination and the resulting oligosaccharide preparations were analyzed by 500-MHz 1H NMR spectroscopy. The structure of the predominant glycan, a hexasaccharide: (formula; see text) and that of a tetrasaccharide: NeuAc alpha(2----3)Gal beta(1----3)[NeuAc alpha (2----6)]GalNAc-ol, were determined. The protein possesses approximately 40 hexasaccharides and 3 tetrasaccharides/mol. For the isolation of its N-glycosidic oligosaccharide chains, the protein was exhaustively digested with proteases followed by chromatography of the desialyzed resulting glycopeptide fraction on concanavalin A-Sepharose. 500-MHz 1H NMR spectroscopy of the obtained preparations revealed the presence of 3 diantennary N-glycosidic chains that are extended with a Fuc residue at the Asn-linked GlcNAc.
Notes: Akiyama, K
Simons, E R
Bernasconi, P
Schmid, K
van Halbeek, H
Vliegenthart, J F
Haupt, H
Schwick, H G
GM-10374/GM/NIGMS NIH HHS/United States
HL-15335/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1984 Jun 10;259(11):7151-4.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1276
Author: Akiyama, M., Takeda, S., Kokame, K., Takagi, J. and Miyata, T.
Year: 2009
Title: Crystal structures of the noncatalytic domains of ADAMTS13 reveal multiple discontinuous exosites for von Willebrand factor
Journal: Proc Natl Acad Sci U S A
Volume: 106
Issue: 46
Pages: 19274-9
Epub Date: 2009/11/03
Date: Nov 17
Short Title: Crystal structures of the noncatalytic domains of ADAMTS13 reveal multiple discontinuous exosites for von Willebrand factor
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0909755106
PMCID: PMC2780749
Accession Number: 19880749
Keywords: ADAM Proteins/*chemistry/genetics/metabolism
ADAMTS13 Protein
Crystallography, X-Ray
Humans
*Protein Interaction Domains and Motifs
von Willebrand Factor/*metabolism
Abstract: ADAMTS13 specifically cleaves plasma von Willebrand factor (VWF) and thereby controls VWF-mediated platelet thrombus formation. Severe deficiencies in ADAMTS13 can cause life-threatening thrombotic thrombocytopenic purpura. Here, we determined 2 crystal structures of ADAMTS13-DTCS (residues 287-685), an exosite-containing human ADAMTS13 fragment, at 2.6-A and 2.8-A resolution. The structures revealed folding similarities between the disintegrin-like (D) domain and the N-terminal portion of the cysteine-rich domain (designated the C(A) domain). The spacer (S) domain forms a globular functional unit with a 10-stranded beta-sandwich fold that has multiple interaction sites with the C(A) domain. We expressed 25 structure-based mutants of ADAMTS13-MDTCS (residues 75-685) and measured their enzymatic activity. We identified 3 VWF-binding exosites on the linearly aligned discontinuous surfaces of the D, C(A), and S domains traversing the W-shaped molecule. Since the MDTCS domains are conserved among ADAMTS family proteins, the structural framework of the multiple enzyme-substrate interactions identified in the ADAMTS13-VWF system provides the basis for a common substrate recognition mode in this class of proteinases.
Notes: 1091-6490
Akiyama, Masashi
Takeda, Soichi
Kokame, Koichi
Takagi, Junichi
Miyata, Toshiyuki
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2009 Nov 17;106(46):19274-9. doi: 10.1073/pnas.0909755106. Epub 2009 Oct 30.
Author Address: aNational Cardiovascular Center Research Institute, Suita, Osaka 565-8565, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 837
Author: Alam, M., Vance, D. E. and Lehner, R.
Year: 2002
Title: Structure-function analysis of human triacylglycerol hydrolase by site-directed mutagenesis: identification of the catalytic triad and a glycosylation site
Journal: Biochemistry
Volume: 41
Issue: 21
Pages: 6679-87
Epub Date: 2002/05/23
Date: May 28
Short Title: Structure-function analysis of human triacylglycerol hydrolase by site-directed mutagenesis: identification of the catalytic triad and a glycosylation site
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi0255625
Accession Number: 12022871
Keywords: Animals
Catalytic Domain
Cells, Cultured
Escherichia coli/genetics/metabolism
Glutamic Acid/genetics/*metabolism
Glycosylation
Histidine/genetics/*metabolism
Humans
Insecta/genetics/metabolism
Lipase/*chemistry/genetics/metabolism
Models, Molecular
Mutagenesis, Site-Directed
Recombinant Proteins/chemistry/genetics/metabolism
Serine/genetics/*metabolism
Structure-Activity Relationship
Abstract: Triacylglycerol hydrolase is a microsomal enzyme that hydrolyzes stored cytoplasmic triacylglycerol in the liver and participates in the lipolysis/re-esterification cycle during the assembly of very-low-density lipoproteins. The structure-activity relationship of the enzyme was investigated by site-directed mutagenesis and heterologous expression. Expression of human TGH in Escherichia coli yields a protein without enzymatic activity, which suggests that posttranslational processing is necessary for the catalytic activity. Expression in baculovirus-infected Sf-9 cells resulted in correct processing of the N-terminal signal sequence and yielded a catalytically active enzyme. A putative catalytic triad consisting of a nucleophilic serine (S221), glutamic acid (E354), and histidine (H468) was identified. Site-directed mutagenesis of the residues (S221A, E354A, and H468A) yielded a catalytically inactive enzyme. CD spectra of purified mutant proteins were very similar to that of the wild-type enzyme, which suggests that the mutations did not affect folding. Human TGH was glycosylated in the insect cells. Mutagenesis of the putative N-glycosylation site (N79A) yielded an active nonglycosylated enzyme. Deletion of the putative C-terminal endoplasmic reticulum retrieval signal (HIEL) did not result in secretion of the mutant protein. A model of human TGH structure suggested a lipase alpha/beta hydrolase fold with a buried active site and two disulfide bridges (C87-C116 and C274-C285).
Notes: Alam, Mustafa
Vance, Dennis E
Lehner, Richard
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2002 May 28;41(21):6679-87. doi: 10.1021/bi0255625.
Author Address: Department of Pediatrics, CIHR Group on Molecular and Cell Biology of Lipids, University of Alberta, 328 Heritage Medical Centre, Edmonton, Alberta, Canada T6G 2S2.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 913
Author: Albach, C., Damoc, E., Denzinger, T., Schachner, M., Przybylski, M. and Schmitz, B.
Year: 2004
Title: Identification of N-glycosylation sites of the murine neural cell adhesion molecule NCAM by MALDI-TOF and MALDI-FTICR mass spectrometry
Journal: Anal Bioanal Chem
Volume: 378
Issue: 4
Pages: 1129-35
Epub Date: 2003/12/06
Date: Feb
Short Title: Identification of N-glycosylation sites of the murine neural cell adhesion molecule NCAM by MALDI-TOF and MALDI-FTICR mass spectrometry
Alternate Journal: Analytical and bioanalytical chemistry
ISSN: 1618-2642 (Print)
1618-2642
DOI: 10.1007/s00216-003-2383-2
Accession Number: 14658030
Keywords: Animals
Glycosylation
Mice
Mice, Inbred ICR
Mice, Inbred Strains
Neural Cell Adhesion Molecules/*analysis/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/*methods
Spectroscopy, Fourier Transform Infrared/methods
Abstract: Mass spectrometry has been shown in recent years to be a powerful tool to determine accurate molecular masses and sequences of peptides and proteins and post-translational modifications such as glycosylation, phosphorylation, and sulfation. For glycosylation, it has been increasingly recognized to be of pivotal importance to identify whether potential glycosylation sites are actually modified by glycans, because functions of proteins may be modulated or depend on the presence of glycans at specific sites. Several recent reports have established that mass spectrometric techniques such as matrix-assisted laser desorption/ionization or electrospray ionization mass spectrometry (MALDI-TOF or ESI-MS, respectively) with or without preceding HPLC and in combination with PNGase F treatment are suited to analyze whether consensus sequences for N-glycosylation are glycosylated or not. Here we report the mass spectrometric analysis of the six potential N-glycosylation sites of the neural cell adhesion molecule NCAM from adult mouse brain. Unmodified peptides and glycopeptides each carrying a single glycosylation site were generated from NCAM by AspN and trypsin treatment and submitted to reversed-phase HPLC with or without prior enzymatic release of N-glycans. The resulting peptides were analyzed by MALDI-TOF-MS. In addition, high-resolution Fourier transform-ion cyclotron resonance (MALDI-FTICR) mass spectrometry was performed after in-gel deglycosylation and subsequent trypsin digestion. By using these procedures all six consensus sequences were shown to be glycosylated; the observation of an unmodified peptide with the consensus sequence N-1 indicates only partial glycosylation at this site.
Notes: Albach, Claus
Damoc, Eugen
Denzinger, Thomas
Schachner, Melitta
Przybylski, Michael
Schmitz, Brigitte
Journal Article
Research Support, Non-U.S. Gov't
Germany
Anal Bioanal Chem. 2004 Feb;378(4):1129-35. doi: 10.1007/s00216-003-2383-2. Epub 2003 Dec 5.
Author Address: Institute for Physiology, Biochemistry and Animal Health, University of Bonn, Katzenburgweg 7-9, 53115 Bonn, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1390
Author: Albers, J. J., Day, J. R., Wolfbauer, G., Kennedy, H., Vuletic, S. and Cheung, M. C.
Year: 2011
Title: Impact of site-specific N-glycosylation on cellular secretion, activity and specific activity of the plasma phospholipid transfer protein
Journal: Biochim Biophys Acta
Volume: 1814
Issue: 7
Pages: 908-11
Epub Date: 2011/04/26
Date: Jul
Short Title: Impact of site-specific N-glycosylation on cellular secretion, activity and specific activity of the plasma phospholipid transfer protein
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbapap.2011.04.004
PMCID: PMC3112057
NIHMSID: NIHMS294708
Accession Number: 21515415
Keywords: Alanine/chemistry/genetics/metabolism
Amino Acid Substitution
Animals
Binding Sites/genetics
Blotting, Western
CHO Cells
Carbohydrates/*chemistry
Cricetinae
Cricetulus
Culture Media, Conditioned/metabolism
Electrophoresis, Polyacrylamide Gel
Enzyme-Linked Immunosorbent Assay
Glycosylation
Humans
Mutagenesis, Site-Directed
Mutant Proteins/chemistry/genetics/metabolism
*Mutation
Phospholipid Transfer Proteins/*chemistry/*metabolism
Serine/chemistry/genetics/metabolism
Abstract: The plasma phospholipid transfer protein (PLTP) plays a key role in lipid and lipoprotein metabolism. It has six potential N-glycosylation sites. To study the impact of these sites on PLTP secretion and activity, six variants containing serine to alanine point mutations were prepared by site-directed mutagenesis and expressed in Chinese hamster ovary Flp-In cells. The apparent size of each of the six PLTP mutants was slightly less than that of wild type by Western blot, indicating that all six sites are glycosylated or utilized. The size of the carbohydrate at each N-glycosylation site ranged from 3.14 to 4.2kDa. The effect of site-specific N-glycosylation removal on PLTP secretion varied from a modest enhancement (15% and 60%), or essentially no effect, to a reduction in secretion (8%, 14% and 32%). Removal of N-glycosylation at any one of the six glycosylation sites resulted in a significant 35-78% decrease in PLTP activity, and a significant 29-80% decrease in PLTP specific activity compared to wild type. These data indicate that although no single N-linked carbohydrate chain is a requirement for secretion or activity, the removal of the carbohydrate chains had a quantitative impact on cellular secretion of PLTP and its phospholipid transfer activity.
Notes: Albers, John J
Day, Joseph R
Wolfbauer, Gertrud
Kennedy, Hal
Vuletic, Simona
Cheung, Marian C
P01 HL030086/HL/NHLBI NIH HHS/United States
P01 HL030086-25/HL/NHLBI NIH HHS/United States
HL030086/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Biochim Biophys Acta. 2011 Jul;1814(7):908-11. doi: 10.1016/j.bbapap.2011.04.004. Epub 2011 Apr 16.
Author Address: Northwest Lipid Metrobolism and Diabetes Research Laboratories, Division of Metabolism, Endocrinology, and Nutrition, Department of Medicine, University of Washington, Seattle, Washington 98109-4517, USA. jja@uw.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1616
Author: Albright, R. A., Ornstein, D. L., Cao, W., Chang, W. C., Robert, D., Tehan, M., Hoyer, D., Liu, L., Stabach, P., Yang, G., De La Cruz, E. M. and Braddock, D. T.
Year: 2014
Title: Molecular basis of purinergic signal metabolism by ectonucleotide pyrophosphatase/phosphodiesterases 4 and 1 and implications in stroke
Journal: J Biol Chem
Volume: 289
Issue: 6
Pages: 3294-306
Epub Date: 2013/12/18
Date: Feb 7
Short Title: Molecular basis of purinergic signal metabolism by ectonucleotide pyrophosphatase/phosphodiesterases 4 and 1 and implications in stroke
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.505867
PMCID: PMC3916532
Accession Number: 24338010
Keywords: Adenosine Triphosphate/*chemistry/genetics/metabolism
Blood Platelets/enzymology/pathology
Brain/enzymology/pathology
Dinucleoside Phosphates/*chemistry/genetics/metabolism
Endothelium, Vascular/enzymology/pathology/physiology
Humans
Phosphoric Diester Hydrolases/*chemistry/genetics/metabolism
Platelet Aggregation/genetics
Polymorphism, Genetic
Protein Structure, Tertiary
Pyrophosphatases/*chemistry/genetics/metabolism
Stroke/*enzymology/genetics/pathology
Substrate Specificity
Bone
Enzyme Catalysis
Platelets
Purinergic Metabolism
Stroke
X-ray Crystallography
Abstract: NPP4 is a type I extracellular membrane protein on brain vascular endothelium inducing platelet aggregation via the hydrolysis of Ap3A, whereas NPP1 is a type II extracellular membrane protein principally present on the surface of chondrocytes that regulates tissue mineralization. To understand the metabolism of purinergic signals resulting in the physiologic activities of the two enzymes, we report the high resolution crystal structure of human NPP4 and explore the molecular basis of its substrate specificity with NPP1. Both enzymes cleave Ap3A, but only NPP1 can hydrolyze ATP. Comparative structural analysis reveals a tripartite lysine claw in NPP1 that stabilizes the terminal phosphate of ATP, whereas the corresponding region of NPP4 contains features that hinder this binding orientation, thereby inhibiting ATP hydrolysis. Furthermore, we show that NPP1 is unable to induce platelet aggregation at physiologic concentrations reported in human blood, but it could stimulate platelet aggregation if localized at low nanomolar concentrations on vascular endothelium. The combined studies expand our understanding of NPP1 and NPP4 substrate specificity and range and provide a rational mechanism by which polymorphisms in NPP1 confer stroke resistance.
Notes: 1083-351x
Albright, Ronald A
Ornstein, Deborah L
Cao, Wenxiang
Chang, William C
Robert, Donna
Tehan, Martin
Hoyer, Denton
Liu, Lynn
Stabach, Paul
Yang, Guangxiao
De La Cruz, Enrique M
Braddock, Demetrios T
P30 CA016359/CA/NCI NIH HHS/United States
R25 GM096161/GM/NIGMS NIH HHS/United States
T32 GM007223/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2014 Feb 7;289(6):3294-306. doi: 10.1074/jbc.M113.505867. Epub 2013 Dec 12.
Author Address: From the Department of Pathology, Yale University School of Medicine, New Haven, Connecticut 06510.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1452
Author: Aleshin, A. E., Schraufstatter, I. U., Stec, B., Bankston, L. A., Liddington, R. C. and DiScipio, R. G.
Year: 2012
Title: Structure of complement C6 suggests a mechanism for initiation and unidirectional, sequential assembly of membrane attack complex (MAC)
Journal: J Biol Chem
Volume: 287
Issue: 13
Pages: 10210-10222
Epub Date: 2012/01/24
Date: Mar 23
Short Title: Structure of complement C6 suggests a mechanism for initiation and unidirectional, sequential assembly of membrane attack complex (MAC)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.327809
PMCID: PMC3323040
Accession Number: 22267737
Keywords: Complement C6/*chemistry/metabolism
*Complement Membrane Attack Complex
Complement System Proteins/chemistry/metabolism
Crystallography, X-Ray
Humans
*Models, Biological
*Models, Molecular
Protein Structure, Secondary
Protein Structure, Tertiary
Structure-Activity Relationship
Abstract: The complement membrane attack complex (MAC) is formed by the sequential assembly of C5b with four homologous proteins as follows: one copy each of C6, C7, and C8 and 12-14 copies of C9. Together these form a lytic pore in bacterial membranes. C6 through C9 comprise a MAC-perforin domain flanked by 4-9 "auxiliary" domains. Here, we report the crystal structure of C6, the first and longest of the pore proteins to be recruited by C5b. Comparisons with the structures of the C8αβγ heterodimer and perforin show that the central domain of C6 adopts a "closed" (perforin-like) state that is distinct from the "open" conformations in C8. We further show that C6, C8α, and C8β contain three homologous subdomains ("upper," "lower," and "regulatory") related by rotations about two hinge points. In C6, the regulatory segment includes four auxiliary domains that stabilize the closed conformation, inhibiting release of membrane-inserting elements. In C8β, rotation of the regulatory segment is linked to an opening of the central β-sheet of its clockwise partner, C8α. Based on these observations, we propose a model for initiation and unidirectional propagation of the MAC in which the auxiliary domains play key roles: in the assembly of the C5b-8 initiation complex; in driving and regulating the opening of the β-sheet of the MAC-performin domain of each new recruit as it adds to the growing pore; and in stabilizing the final pore. Our model of the assembled pore resembles those of the cholesterol-dependent cytolysins but is distinct from that recently proposed for perforin.
Notes: 1083-351x
Aleshin, Alexander E
Schraufstatter, Ingrid U
Stec, Boguslaw
Bankston, Laurie A
Liddington, Robert C
DiScipio, Richard G
R21 HL094878/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2012 Mar 23;287(13):10210-10222. doi: 10.1074/jbc.M111.327809. Epub 2012 Jan 20.
Author Address: Infectious and Inflammatory Disease Center, Sanford-Burnham Medical Research Institute, La Jolla, California 92037 and.
Torrey Pines Institute for Molecular Studies, San Diego, California 92121.
Infectious and Inflammatory Disease Center, Sanford-Burnham Medical Research Institute, La Jolla, California 92037 and. Electronic address: rlidding@sanfordburnham.org.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1423
Author: Alhamidi, M., Kjeldsen Buvang, E., Fagerheim, T., Brox, V., Lindal, S., Van Ghelue, M. and Nilssen, Ø
Year: 2011
Title: Fukutin-related protein resides in the Golgi cisternae of skeletal muscle fibres and forms disulfide-linked homodimers via an N-terminal interaction
Journal: PLoS One
Volume: 6
Issue: 8
Pages: e22968
Epub Date: 2011/09/03
Short Title: Fukutin-related protein resides in the Golgi cisternae of skeletal muscle fibres and forms disulfide-linked homodimers via an N-terminal interaction
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0022968
PMCID: PMC3160285
Accession Number: 21886772
Keywords: Animals
Biomarkers/metabolism
Cell Line
Cell Survival/drug effects
Cysteine/metabolism
Disulfides/*metabolism
Dithiothreitol/pharmacology
Glycosylation/drug effects
Golgi Apparatus/drug effects/*metabolism/ultrastructure
Humans
Immunoprecipitation
Mannose/metabolism
Models, Biological
Multiprotein Complexes/metabolism
Muscle Fibers, Skeletal/drug effects/*metabolism/ultrastructure
Myofibrils/drug effects/metabolism/ultrastructure
Oligosaccharides/metabolism
Oxidation-Reduction/drug effects
Pentosyltransferases
Protein Binding/drug effects
*Protein Multimerization/drug effects
Protein Transport/drug effects
Proteins/*metabolism
Receptors, Fibroblast Growth Factor/metabolism
Recombinant Proteins/metabolism
Sialoglycoproteins/metabolism
Two-Hybrid System Techniques
Abstract: Limb-Girdle Muscular Dystrophy type 2I (LGMD2I) is an inheritable autosomal, recessive disorder caused by mutations in the FuKutin-Related Protein (FKRP) gene (FKRP) located on chromosome 19 (19q13.3). Mutations in FKRP are also associated with Congenital Muscular Dystrophy (MDC1C), Walker-Warburg Syndrome (WWS) and Muscle Eye Brain disease (MEB). These four disorders share in common an incomplete/aberrant O-glycosylation of the membrane/extracellular matrix (ECM) protein α-dystroglycan. However, further knowledge on the FKRP structure and biological function is lacking, and its intracellular location is controversial. Based on immunogold electron microscopy of human skeletal muscle sections we demonstrate that FKRP co-localises with the middle-to-trans-Golgi marker MG160, between the myofibrils in human rectus femoris muscle fibres. Chemical cross-linking experiments followed by pairwise yeast 2-hybrid experiments, and co-immune precipitation, demonstrate that FKRP can exist as homodimers as well as in large multimeric protein complexes when expressed in cell culture. The FKRP homodimer is kept together by a disulfide bridge provided by the most N-terminal cysteine, Cys6. FKRP contains N-glycan of high mannose and/or hybrid type; however, FKRP N-glycosylation is not required for FKRP homodimer or multimer formation. We propose a model for FKRP which is consistent with that of a Golgi resident type II transmembrane protein.
Notes: 1932-6203
Alhamidi, Maisoon
Kjeldsen Buvang, Elisabeth
Fagerheim, Toril
Brox, Vigdis
Lindal, Sigurd
Van Ghelue, Marijke
Nilssen, Øivind
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2011;6(8):e22968. doi: 10.1371/journal.pone.0022968. Epub 2011 Aug 23.
Author Address: Division of Child and Adolescent Health, Department of Medical Genetics, University Hospital of North-Norway, Tromsø, Norway.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1996
Author: Ali, A., Kim, M. J., Kim, M. Y., Lee, H. J., Roh, G. S., Kim, H. J., Cho, G. J. and Choi, W. S.
Year: 2018
Title: Quercetin induces cell death in cervical cancer by reducing O-GlcNAcylation of adenosine monophosphate-activated protein kinase
Journal: Anat Cell Biol
Volume: 51
Issue: 4
Pages: 274-283
Epub Date: 2019/01/15
Date: Dec
Short Title: Quercetin induces cell death in cervical cancer by reducing O-GlcNAcylation of adenosine monophosphate-activated protein kinase
Alternate Journal: Anatomy & cell biology
ISSN: 2093-3665 (Print)
2093-3665
DOI: 10.5115/acb.2018.51.4.274
PMCID: PMC6318463
Accession Number: 30637162
Keywords: Ampk
O-GlcNAcylation
Quercetin
Sterol regulatory element binding protein 1
Uterine cervical neoplasms
Abstract: Hyper-O-GlcNAcylation is a general feature of cancer which contributes to various cancer phenotypes, including cell proliferation and cell growth. Quercetin, a naturally occurring dietary flavonoid, has been reported to reduce the proliferation and growth of cancer. Several reports of the anticancer effect of quercetin have been published, but there is no study regarding its effect on O-GlcNAcylation. The aim of this study was to investigate the anticancer effect of quercetin on HeLa cells and compare this with its effect on HaCaT cells. Cell viability and cell death were determined by MTT and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labelling assays. O-GlcNAcylation of AMP-activated protein kinase (AMPK) was examined by succinylated wheat germ agglutinin pulldown and immunoprecipitation. Immunofluorescence staining was used to detect the immunoreactivitiy of O-linked N-acetylglucosamine transferase (OGT) and sterol regulatory element binding protein 1 (SREBP-1). Quercetin decreased cell proliferation and induced cell death, but its effect on HaCaT cells was lower than that on HeLa cells. O-GlcNAcylation level was higher in HeLa cells than in HaCaT cells. Quercetin decreased the expression of global O-GlcNAcylation and increased AMPK activation by reducing the O-GlcNAcylation of AMPK. AMPK activation due to reduced O-GlcNAcylation of AMPK was confirmed by treatment with 6-diazo-5-oxo-L-norleucine. Our results also demonstrated that quercetin regulated SREBP-1 and its transcriptional targets. Furthermore, immunofluorescence staining showed that quercetin treatment decreased the immunoreactivities of OGT and SREBP-1 in HeLa cells. Our findings demonstrate that quercetin exhibited its anticancer effect by decreasing the O-GlcNAcylation of AMPK. Further studies are needed to explore how quercetin regulates O-GlcNAcylation in cancer.
Notes: 2093-3673
Ali, Akhtar
Kim, Min Jun
Kim, Min Young
Lee, Han Ju
Roh, Gu Seob
Kim, Hyun Joon
Cho, Gyeong Jae
Choi, Wan Sung
Journal Article
Anat Cell Biol. 2018 Dec;51(4):274-283. doi: 10.5115/acb.2018.51.4.274. Epub 2018 Dec 29.
Author Address: Department of Anatomy and Convergence Medical Science, Institute of Health Sciences, Gyeongsang National University School of Medicine, Jinju, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1870
Author: Ali, A., Kim, S. H., Kim, M. J., Choi, M. Y., Kang, S. S., Cho, G. J., Kim, Y. S., Choi, J. Y. and Choi, W. S.
Year: 2017
Title: O-GlcNAcylation of NF-κB Promotes Lung Metastasis of Cervical Cancer Cells via Upregulation of CXCR4 Expression
Journal: Mol Cells
Volume: 40
Issue: 7
Pages: 476-484
Epub Date: 2017/07/07
Date: Jul 31
Short Title: O-GlcNAcylation of NF-κB Promotes Lung Metastasis of Cervical Cancer Cells via Upregulation of CXCR4 Expression
Alternate Journal: Molecules and cells
ISSN: 1016-8478 (Print)
1016-8478
DOI: 10.14348/molcells.2017.2309
PMCID: PMC5547217
Accession Number: 28681591
Keywords: Acetylglucosamine/metabolism
Animals
Female
Gene Knockdown Techniques
Glycosylation
HeLa Cells
Humans
Lung Neoplasms/metabolism/*secondary
Mice, Nude
N-Acetylglucosaminyltransferases/metabolism
NF-kappa B/*metabolism
Protein Binding
Receptors, CXCR4/*metabolism
Transcription Factor RelA/metabolism
*Up-Regulation
Uterine Cervical Neoplasms/*pathology
Cxcr4
NF-κB p65
O-GlcNAcylation
cervical cancer
lung metastasis
Abstract: C-X-C chemokine receptor 4 (CXCR4) stimulates cancer metastasis. NF-κB regulates CXCR4 expression in cancer cells, and O-GlcNAc modification of NF-κB promotes its transcriptional activity. Here, we determined whether CXCR4 expression is affected by O-GlcNAcylation of NF-κB in lung metastasis of cervical cancer. We found elevated levels of O-linked-N-actylglucosamine transferase (OGT) and O-GlcNAcylation in cervical cancer cells compared to those in non-malignant epithelial cells and detected increased expression of NF-κB p65 (p65) and CXCR4 in cervical cancer cells. Knockdown of OGT inhibited the O-GlcNAcylation of p65 and decreased CXCR4 expression levels in HeLa cells. Thiamet G treatment increased O-GlcNAcylated p65, which subsequently enhanced CXCR4 expression levels. Inhibition of O-GlcNAcylation by 6-Diazo-5-oxo-L-norleucine (DON) treatment decreased p65 activation, eventually inhibiting CXCR4 expression in HeLa cells. Lung tissues from mice engrafted with OGT-knockdown HeLa cells (shOGT) exhibited lower expression of Ki-67 and HPV E6 and E7 oncogenes compared to lung tissues from mice engrafted with control HeLa cells (shCTL). In addition, lung tissues from mice engrafted with shOGT cells exhibited lower p65 and CXCR4 immunoreactivity compared to tissues from mice engrafted with shCTL cells. Taken together, our data suggest that p65 O-GlcNAcylation promotes lung metastasis of cervical cancer cells by activating CXCR4 expression.
Notes: 0219-1032
Ali, Akhtar
Kim, Sung Hwan
Kim, Min Jun
Choi, Mee Young
Kang, Sang Soo
Cho, Gyeong Jae
Kim, Yoon Sook
Choi, Jun-Young
Choi, Wan Sung
Journal Article
Mol Cells. 2017 Jul 31;40(7):476-484. doi: 10.14348/molcells.2017.2309. Epub 2017 Jul 6.
Author Address: Department of Anatomy and Convergence Medical Science, Institute of Health Sciences, Gyeongsang National University, School of Medicine, Jinju 52727, Korea.
Department of Thoriac and Cardiovascular Surgery, Gyeongsang National University, School of Medicine, Jinju 52727, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1662
Author: Ali, L., Flowers, S. A., Jin, C., Bennet, E. P., Ekwall, A. K. and Karlsson, N. G.
Year: 2014
Title: The O-glycomap of lubricin, a novel mucin responsible for joint lubrication, identified by site-specific glycopeptide analysis
Journal: Mol Cell Proteomics
Volume: 13
Issue: 12
Pages: 3396-409
Epub Date: 2014/09/05
Date: Dec
Short Title: The O-glycomap of lubricin, a novel mucin responsible for joint lubrication, identified by site-specific glycopeptide analysis
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M114.040865
PMCID: PMC4256492
Accession Number: 25187573
Keywords: Amino Acid Sequence
Arginine/chemistry/metabolism
Carbohydrate Sequence
Fibroblasts/*chemistry/cytology/metabolism
Gene Expression
Glycopeptides/*chemistry/genetics/metabolism
Glycoproteins/*chemistry/genetics/metabolism
Glycosylation
Humans
Isoenzymes/chemistry/genetics/metabolism
Lubrication
Lysine/chemistry/metabolism
Molecular Sequence Data
N-Acetylgalactosaminyltransferases/chemistry/genetics/metabolism
Peptide Mapping
Primary Cell Culture
Protein Structure, Tertiary
Static Electricity
Synovial Fluid/chemistry/cytology
Threonine/*chemistry/metabolism
Abstract: The lubricative, heavily glycosylated mucin-like synovial glycoprotein lubricin has previously been observed to contain glycosylation changes related to rheumatoid and osteoarthritis. Thus, a site-specific investigation of the glycosylation of lubricin was undertaken, in order to further understand the pathological mechanisms involved in these diseases. Lubricin contains an serine/threonine/proline (STP)-rich domain composed of imperfect tandem repeats (EPAPTTPK), the target for O-glycosylation. In this study, using a liquid chromatography-tandem mass spectrometry approach, employing both collision-induced and electron-transfer dissociation fragmentation methods, we identified 185 O-glycopeptides within the STP-rich domain of human synovial lubricin. This showed that adjacent threonine residues within the central STP-rich region could be simultaneously and/or individually glycosylated. In addition to core 1 structures responsible for biolubrication, core 2 O-glycopeptides were also identified, indicating that lubricin glycosylation may have other roles. Investigation of the expression of polypeptide N-acetylgalactosaminyltransferase genes was carried out using cultured primary fibroblast-like synoviocytes, a cell type that expresses lubricin in vivo. This analysis showed high mRNA expression levels of the less understood polypeptide N-acetylgalactosaminyltransferase 15 and 5 in addition to the ubiquitously expressed polypeptide N-acetylgalactosaminyltransferase 1 and 2 genes. This suggests that there is a unique combination of transferase genes important for the O-glycosylation of lubricin. The site-specific glycopeptide analysis covered 82% of the protein sequence and showed that lubricin glycosylation displays both micro- and macroheterogeneity. The density of glycosylation was shown to be high: 168 sites of O-glycosylation, predominately sialylated, were identified. These glycosylation sites were focused in the central STP-rich region, giving the domain a negative charge. The more positively charged lysine and arginine residues in the N and C termini suggest that synovial lubricin exists as an amphoteric molecule. The identification of these unique properties of lubricin may provide insight into the important low-friction lubricating functions of lubricin during natural joint movement.
Notes: 1535-9484
Ali, Liaqat
Flowers, Sarah A
Jin, Chunsheng
Bennet, Eric Paul
Ekwall, Anna-Karin H
Karlsson, Niclas G
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2014 Dec;13(12):3396-409. doi: 10.1074/mcp.M114.040865. Epub 2014 Sep 3.
Author Address: From the ‡Department of Medical Biochemistry, Institute of Biomedicine, University of Gothenburg, Box 440, Medicinaregatan 9A, 405 30, Gothenburg, Sweden;
§Department of Odontology, Copenhagen Center for Glycomics, University of Copenhagen, Norre Alle 20, DK-2200 Copenhagen N, Denmark;
¶Department of Rheumatology and Inflammation Research, Institute of Medicine, University of Gothenburg, Guldhedsgatan 10A, SE-41346, Gothenburg, Sweden.
From the ‡Department of Medical Biochemistry, Institute of Biomedicine, University of Gothenburg, Box 440, Medicinaregatan 9A, 405 30, Gothenburg, Sweden; niclas.karlsson@medkem.gu.se.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 394
Author: Alliel, P. M., Périn, J. P., Maillet, P., Bonnet, F., Rosa, J. P. and Jollès, P.
Year: 1988
Title: Complete amino acid sequence of a human platelet proteoglycan
Journal: FEBS Lett
Volume: 236
Issue: 1
Pages: 123-6
Epub Date: 1988/08/15
Date: Aug 15
Short Title: Complete amino acid sequence of a human platelet proteoglycan
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(88)80298-9
Accession Number: 3402609
Keywords: Amino Acid Sequence
Base Sequence
Blood Platelets/*analysis
Chromatography, Gel
Chromatography, High Pressure Liquid
Cloning, Molecular
DNA/genetics
Humans
Leukemia
Molecular Sequence Data
Proteoglycans/*blood/genetics
Repetitive Sequences, Nucleic Acid
Sequence Homology, Nucleic Acid
Tumor Cells, Cultured
Abstract: The primary structure of a human platelet proteoglycan (P.PG) core was established by a combination of amino acid sequence analysis and cDNA cloning. The deduced 131 amino acid long protein contains eight Ser-Gly repeats. The significance of homologies observed between P.PG and promyelocytic leukemia cell line proteoglycans is discussed.
Notes: Alliel, P M
Périn, J P
Maillet, P
Bonnet, F
Rosa, J P
Jollès, P
Journal Article
England
FEBS Lett. 1988 Aug 15;236(1):123-6. doi: 10.1016/0014-5793(88)80298-9.
Author Address: Laboratoire des Protéines, UA CNRS no. 1188, Université de Paris V, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1519
Author: Allison, D. F., Wamsley, J. J., Kumar, M., Li, D., Gray, L. G., Hart, G. W., Jones, D. R. and Mayo, M. W.
Year: 2012
Title: Modification of RelA by O-linked N-acetylglucosamine links glucose metabolism to NF-κB acetylation and transcription
Journal: Proc Natl Acad Sci U S A
Volume: 109
Issue: 42
Pages: 16888-93
Epub Date: 2012/10/03
Date: Oct 16
Short Title: Modification of RelA by O-linked N-acetylglucosamine links glucose metabolism to NF-κB acetylation and transcription
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1208468109
PMCID: PMC3479489
Accession Number: 23027940
Keywords: Acetylation
Acetylglucosamine/*metabolism
Chromatin Immunoprecipitation
DNA, Complementary/genetics
Enzyme-Linked Immunosorbent Assay
Etoposide/metabolism
Gene Expression Regulation/*physiology
Glucose/*metabolism
HEK293 Cells
Hexosamines/biosynthesis
Humans
Immunoblotting
Immunoprecipitation
Luciferases
Metabolic Networks and Pathways/genetics/*physiology
NF-kappa B/*metabolism
Plasmids/genetics
Real-Time Polymerase Chain Reaction
Transcription Factor RelA/genetics/*metabolism
Tumor Necrosis Factor-alpha/metabolism
Abstract: The molecular mechanisms linking glucose metabolism with active transcription remain undercharacterized in mammalian cells. Using nuclear factor-κB (NF-κB) as a glucose-responsive transcription factor, we show that cells use the hexosamine biosynthesis pathway and O-linked β-N-acetylglucosamine (O-GlcNAc) transferase (OGT) to potentiate gene expression in response to tumor necrosis factor (TNF) or etoposide. Chromatin immunoprecipitation assays demonstrate that, upon induction, OGT localizes to NF-κB-regulated promoters to enhance RelA acetylation. Knockdown of OGT abolishes p300-mediated acetylation of RelA on K310, a posttranslational mark required for full NF-κB transcription. Mapping studies reveal T305 as an important residue required for attachment of the O-GlcNAc moiety on RelA. Furthermore, p300 fails to acetylate a full-length RelA(T305A) mutant, linking O-GlcNAc and acetylation events on NF-κB. Reconstitution of RelA null cells with the RelA(T305A) mutant illustrates the importance of this residue for NF-κB-dependent gene expression and cell survival. Our work provides evidence for a unique regulation where attachment of the O-GlcNAc moiety to RelA potentiates p300 acetylation and NF-κB transcription.
Notes: 1091-6490
Allison, David F
Wamsley, J Jacob
Kumar, Manish
Li, Duo
Gray, Lisa G
Hart, Gerald W
Jones, David R
Mayo, Marty W
R01 CA132580/CA/NCI NIH HHS/United States
R01CA136705/CA/NCI NIH HHS/United States
R01CA104397/CA/NCI NIH HHS/United States
T32 GM008136/GM/NIGMS NIH HHS/United States
R01DK61671/DK/NIDDK NIH HHS/United States
R01 CA104397/CA/NCI NIH HHS/United States
R01CA132580/CA/NCI NIH HHS/United States
R01 CA136705/CA/NCI NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
P01HL107153/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2012 Oct 16;109(42):16888-93. doi: 10.1073/pnas.1208468109. Epub 2012 Oct 1.
Author Address: Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, VA 22908, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2060
Author: Alteen, M. G., Gros, C., Meek, R. W., Cardoso, D. A., Busmann, J. A., Sangouard, G., Deen, M. C., Tan, H. Y., Shen, D. L., Russell, C. C., Davies, G. J., Robinson, P. J., McCluskey, A. and Vocadlo, D. J.
Year: 2020
Title: A Direct Fluorescent Activity Assay for Glycosyltransferases Enables Convenient High-Throughput Screening: Application to O-GlcNAc Transferase
Journal: Angew Chem Int Ed Engl
Volume: 59
Issue: 24
Pages: 9601-9609
Epub Date: 2020/02/25
Date: Jun 8
Short Title: A Direct Fluorescent Activity Assay for Glycosyltransferases Enables Convenient High-Throughput Screening: Application to O-GlcNAc Transferase
Alternate Journal: Angewandte Chemie (International ed. in English)
ISSN: 1433-7851
DOI: 10.1002/anie.202000621
Accession Number: 32092778
Keywords: Enzyme Assays/*methods
Glycosylation
Kinetics
N-Acetylglucosaminyltransferases/*metabolism
Spectrometry, Fluorescence/*methods
Substrate Specificity
*enzymes
*fluorescent probes
*glycosylation
*high-throughput screening
*inhibitors
Abstract: Glycosyltransferases carry out important cellular functions in species ranging from bacteria to humans. Despite their essential roles in biology, simple and robust activity assays that can be easily applied to high-throughput screening for inhibitors of these enzymes have been challenging to develop. Herein, we report a bead-based strategy to measure the group-transfer activity of glycosyltransferases sensitively using simple fluorescence measurements, without the need for coupled enzymes or secondary reactions. We validate the performance and accuracy of the assay using O-GlcNAc transferase (OGT) as a model system through detailed Michaelis-Menten kinetic analysis of various substrates and inhibitors. Optimization of this assay and application to high-throughput screening enabled screening for inhibitors of OGT, leading to a novel inhibitory scaffold. We believe this assay will prove valuable not only for the study of OGT, but also more widely as a general approach for the screening of glycosyltransferases and other group-transfer enzymes.
Notes: 1521-3773
Alteen, Matthew G
Orcid: 0000-0002-9639-7668
Gros, Christina
Orcid: 0000-0001-8869-9028
Meek, Richard W
Orcid: 0000-0002-1370-0896
Cardoso, David A
Orcid: 0000-0003-3359-7926
Busmann, Jil A
Sangouard, Gontran
Deen, Matthew C
Tan, Hong-Yee
Shen, David L
Russell, Cecilia C
Orcid: 0000-0001-8401-5856
Davies, Gideon J
Orcid: 0000-0002-7343-776x
Robinson, Phillip J
Orcid: 0000-0002-7878-0313
McCluskey, Adam
Orcid: 0000-0001-7125-863x
Vocadlo, David J
Orcid: 0000-0001-6897-5558
Journal Article
Research Support, Non-U.S. Gov't
Germany
Angew Chem Int Ed Engl. 2020 Jun 8;59(24):9601-9609. doi: 10.1002/anie.202000621. Epub 2020 Apr 2.
Author Address: Department of Chemistry, Simon Fraser University, Burnaby, BC, V5A 1S6, Canada.
York Structural Biology Laboratory, Department of Chemistry, University of York, York, YO10 5DD, UK.
Children's Medical Research Institute, The University of Sydney, Sydney, NSW, 2145, Australia.
Department of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, BC, V5A 1S6, Canada.
Chemistry, School of Environmental and Life Sciences, The University of Newcastle, University Drive, Callaghan, NSW, 2308, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1268
Author: Alterio, V., Hilvo, M., Di Fiore, A., Supuran, C. T., Pan, P., Parkkila, S., Scaloni, A., Pastorek, J., Pastorekova, S., Pedone, C., Scozzafava, A., Monti, S. M. and De Simone, G.
Year: 2009
Title: Crystal structure of the catalytic domain of the tumor-associated human carbonic anhydrase IX
Journal: Proc Natl Acad Sci U S A
Volume: 106
Issue: 38
Pages: 16233-8
Epub Date: 2009/10/07
Date: Sep 22
Short Title: Crystal structure of the catalytic domain of the tumor-associated human carbonic anhydrase IX
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0908301106
PMCID: PMC2752527
Accession Number: 19805286
Keywords: Amino Acid Sequence
Antigens, Neoplasm/*chemistry/genetics/metabolism
Bicarbonates/chemistry/metabolism
Carbon Dioxide/chemistry/metabolism
Carbonic Anhydrase IX
Carbonic Anhydrases/*chemistry/genetics/metabolism
Catalytic Domain
Crystallization
Crystallography, X-Ray
Disulfides/chemistry
Drug Design
Enzyme Inhibitors/chemistry/metabolism/pharmacology
Humans
Hydrogen-Ion Concentration
Kinetics
Models, Molecular
Molecular Sequence Data
Neoplasms/drug therapy/*enzymology/pathology
Protein Conformation
Protein Multimerization
*Protein Structure, Tertiary
Protons
Recombinant Proteins/chemistry/metabolism
Sequence Homology, Amino Acid
Structure-Activity Relationship
Water/chemistry/metabolism
Abstract: Carbonic anhydrase (CA) IX is a plasma membrane-associated member of the alpha-CA enzyme family, which is involved in solid tumor acidification. It is a marker of tumor hypoxia and a prognostic factor in several human cancers. An aberrant increase in CA IX expression in chronic hypoxia and during development of various carcinomas contributes to tumorigenesis through at least two mechanisms: pH regulation and cell adhesion control. Here we report the X-ray structure of the catalytic domain of CA IX in complex with a classical, clinically used sulfonamide inhibitor, acetazolamide. The structure reveals a typical alpha-CA fold, which significantly differs from the other CA isozymes when the protein quaternary structure is considered. Thus, two catalytic domains of CA IX associate to form a dimer, which is stabilized by the formation of an intermolecular disulfide bond. The active site clefts and the PG domains are located on one face of the dimer, while the C-termini are located on the opposite face to facilitate protein anchoring to the cell membrane. A correlation between the three-dimensional structure and the physiological role of the enzyme is here suggested, based on the measurement of the pH profile of the catalytic activity for the physiological reaction, CO(2) hydration to bicarbonate and protons. On the basis of the structural differences observed between CA IX and the other membrane-associated alpha-CAs, further prospects for the rational drug design of isozyme-specific CA inhibitors are proposed, given that inhibition of this enzyme shows antitumor activity both in vitro and in vivo.
Notes: 1091-6490
Alterio, Vincenzo
Hilvo, Mika
Di Fiore, Anna
Supuran, Claudiu T
Pan, Peiwen
Parkkila, Seppo
Scaloni, Andrea
Pastorek, Jaromir
Pastorekova, Silvia
Pedone, Carlo
Scozzafava, Andrea
Monti, Simona Maria
De Simone, Giuseppina
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2009 Sep 22;106(38):16233-8. doi: 10.1073/pnas.0908301106. Epub 2009 Sep 14.
Author Address: Istituto di Biostrutture e Bioimmagini-CNR, via Mezzocannone 16, 80134 Naples, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1619
Author: Alterio, V., Pan, P., Parkkila, S., Buonanno, M., Supuran, C. T., Monti, S. M. and De Simone, G.
Year: 2014
Title: The structural comparison between membrane-associated human carbonic anhydrases provides insights into drug design of selective inhibitors
Journal: Biopolymers
Volume: 101
Issue: 7
Pages: 769-78
Epub Date: 2014/01/01
Date: Jul
Short Title: The structural comparison between membrane-associated human carbonic anhydrases provides insights into drug design of selective inhibitors
Alternate Journal: Biopolymers
ISSN: 0006-3525
DOI: 10.1002/bip.22456
Accession Number: 24374484
Keywords: Amino Acid Sequence
Animals
Carbonic Anhydrases/*chemistry/genetics/metabolism
Chromatography, Gel
Chromatography, Liquid
Crystallography, X-Ray
*Drug Design
Enzyme Inhibitors/*chemistry/metabolism/pharmacology
Humans
Isoenzymes/antagonists & inhibitors/chemistry/genetics
Kinetics
Membrane Proteins/antagonists & inhibitors/*chemistry/genetics
Models, Molecular
Molecular Sequence Data
Protein Structure, Secondary
Protein Structure, Tertiary
Recombinant Proteins/chemistry/metabolism
Sequence Homology, Amino Acid
Sf9 Cells
Spectrometry, Mass, Electrospray Ionization
X-ray crystallography
carbonic anhydrase
drug design
inhibitors
Abstract: Carbonic anhydrase isoform XIV (CA XIV) is the last member of the human (h) CA family discovered so far, being localized in brain, kidneys, colon, small intestine, urinary bladder, liver, and spinal cord. It has recently been described as a possible drug target for treatment of epilepsy, some retinopathies as well as some skin tumors. Human carbonic anhydrase (hCA) XIV is a membrane-associated protein consisting of an N-terminal extracellular domain, a putative transmembrane region, and a small cytoplasmic tail. In this article, we report the expression, purification, and the crystallographic structure of the entire extracellular domain of this enzyme. The analysis of the structure revealed the typical α-CA fold, in which a 10-stranded β-sheet forms the core of the molecule, while the comparison with all the other membrane associated isoforms (hCAs IV, IX, and XII) allowed to identify the diverse oligomeric arrangement and the sequence and structural differences observed in the region 127-136 as the main factors to consider in the design of selective inhibitors for each one of the membrane associated α-CAs.
Notes: 1097-0282
Alterio, Vincenzo
Pan, Peiwen
Parkkila, Seppo
Buonanno, Martina
Supuran, Claudiu T
Monti, Simona M
De Simone, Giuseppina
Comparative Study
Journal Article
United States
Biopolymers. 2014 Jul;101(7):769-78. doi: 10.1002/bip.22456.
Author Address: Institute of Biostructures and Bioimaging, CNR, Via Mezzocannone 16, 80134, Naples, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 387
Author: Amano, J., Nishimura, R., Mochizuki, M. and Kobata, A.
Year: 1988
Title: Comparative study of the mucin-type sugar chains of human chorionic gonadotropin present in the urine of patients with trophoblastic diseases and healthy pregnant women
Journal: J Biol Chem
Volume: 263
Issue: 3
Pages: 1157-65
Epub Date: 1988/01/25
Date: Jan 25
Short Title: Comparative study of the mucin-type sugar chains of human chorionic gonadotropin present in the urine of patients with trophoblastic diseases and healthy pregnant women
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3335536
Keywords: Acetylation
Carbohydrate Conformation
Choriocarcinoma/*urine
Chorionic Gonadotropin/*urine
Chromatography, Paper
Electrophoresis, Paper
Female
Glycoside Hydrolases/metabolism
Humans
Hydatidiform Mole/*urine
Mucins/*analysis
Neuraminidase/metabolism
Oligosaccharides/analysis
Pregnancy/*urine
Uterine Neoplasms/*urine
Abstract: Human chorionic gonadotropins (hCGs) highly purified from the urine of patients with trophoblastic diseases and of healthy pregnant women contain approximately four mucin-type sugar chains in one molecule. The structures of these sugar chains were studied comparatively by using a new sensitive method to obtain mucin-type sugar chains quantitatively as radioactive oligosaccharides from a small amount of glycoproteins. The mucin-type sugar chains of all hCGs include sialylated and nonsialylated Gal beta 1----3GalNAc and Gal beta 1----4GlcNAc beta 1----6(Gal beta 1----3)GalNAc. In the case of normal hCG and hydatidiform mole hCG, oligosaccharides containing the tetrasaccharide core occupy approximately 10% of the total mucin-type sugar chains. The ratio of the tetrasaccharide containing oligosaccharides is increased prominently to approximately 60% in choriocarcinoma hCG. The proportion in invasive mole hCG was also increased, but less than the proportion of choriocarcinoma hCG.
Notes: Amano, J
Nishimura, R
Mochizuki, M
Kobata, A
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1988 Jan 25;263(3):1157-65.
Author Address: Department of Biochemistry, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 296
Author: Amano, J., Nishimura, R., Sato, S. and Kobata, A.
Year: 1990
Title: Altered glycosylation of human chorionic gonadotropin decreases its hormonal activity as determined by cyclic-adenosine 3',5'-monophosphate production in MA-10 cells
Journal: Glycobiology
Volume: 1
Issue: 1
Pages: 45-50
Epub Date: 1990/09/01
Date: Sep
Short Title: Altered glycosylation of human chorionic gonadotropin decreases its hormonal activity as determined by cyclic-adenosine 3',5'-monophosphate production in MA-10 cells
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/1.1.45
Accession Number: 1967025
Keywords: Animals
Carbohydrate Sequence
Cell Line
Choriocarcinoma/pathology/*urine
Chorionic Gonadotropin/chemistry/*pharmacology/urine
Cyclic AMP/*metabolism
Enzyme-Linked Immunosorbent Assay
Female
Genetic Variation
Glycosylation
Humans
Hydatidiform Mole/pathology/*urine
Leydig Cell Tumor
Mice
Molecular Sequence Data
Neoplasm Invasiveness
Nucleic Acid Conformation
Oligosaccharides/*chemistry/isolation & purification
Pregnancy
Reference Values
Uterine Neoplasms/pathology/*urine
Abstract: Human chorionic gonadotropin (hCG) purified from pooled urine of normal pregnant women contains four asparagine-linked sugar chains and four mucin-type sugar chains. The structures of asparagine-linked sugar chains of this hormone are constant and site-specific. hCGs obtained from the urine of patients with invasive mole or choriocarcinoma have quite different sets of oligosaccharides although the primary structures of the polypeptides and the numbers of the sugar chains are the same as those of normal hCG. In order to examine the biological activities of these hCGs with altered glycosylation, we measured the amount of cAMP produced in a murine Leydig tumour cell line, MA-10, after incubation with the hCG samples. The extent of sialylation of oligosaccharides in the three hCG samples used in this study were 88% in normal hCG, 82% in invasive mole hCG and 63% in choriocarcinoma hCG. The hormonal activity of invasive mole hCG was slightly lower while that of choriocarcinoma hCG was significantly (P less than 0.01) lower than that of normal hCG. Complete desialylation induced remarkable loss of full activities in all the samples. However, the hormonal activities of the three samples were different even after desialylation. The full activities of the desialylated samples of invasive mole hCG and choriocarcinoma hCG were 78 and 65% of that of desialylated normal hCG. These results indicated that the structures of the neutral oligosaccharide portion are also important for the expression of full hormonal activity.
Notes: Amano, J
Nishimura, R
Sato, S
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1990 Sep;1(1):45-50. doi: 10.1093/glycob/1.1.45.
Author Address: Department of Biochemistry, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1227
Author: Ande, S. R., Moulik, S. and Mishra, S.
Year: 2009
Title: Interaction between O-GlcNAc modification and tyrosine phosphorylation of prohibitin: implication for a novel binary switch
Journal: PLoS One
Volume: 4
Issue: 2
Pages: e4586
Epub Date: 2009/02/25
Short Title: Interaction between O-GlcNAc modification and tyrosine phosphorylation of prohibitin: implication for a novel binary switch
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0004586
PMCID: PMC2642629
Accession Number: 19238206
Keywords: Acetylglucosamine/*metabolism
Humans
Insulin
N-Acetylglucosaminyltransferases/*metabolism
Phosphorylation
*Protein Processing, Post-Translational
Repressor Proteins/*metabolism
Signal Transduction
Tyrosine/metabolism
Abstract: Prohibitin (PHB or PHB1) is an evolutionarily conserved, multifunctional protein which is present in various cellular compartments including the plasma membrane. However, mechanisms involved in various functions of PHB are not fully explored yet. Here we report for the first time that PHB interacts with O-linked beta-N-acetylglucosamine transferase (O-GlcNAc transferase, OGT) and is O-GlcNAc modified; and also undergoes tyrosine phosphorylation in response to insulin. Tyrosine 114 (Tyr114) and tyrosine 259 (Tyr259) in PHB are in the close proximity of potential O-GlcNAc sites serine 121 (Ser121) and threonine 258 (Thr258) respectively. Substitution of Tyr114 and Tyr259 residues in PHB with phenylalanine by site-directed mutagenesis results in reduced tyrosine phosphorylation as well as reduced O-GlcNAc modification of PHB. Surprisingly, this also resulted in enhanced tyrosine phosphorylation and activity of OGT. This is attributed to the presence of similar tyrosine motifs in PHB and OGT. Substitution of Ser121 and Thr258 with alanine and isoleucine respectively resulted in attenuation of O-GlcNAc modification and increased tyrosine phosphorylation of PHB suggesting an association between these two dynamic modifications. Sequence analysis of O-GlcNAc modified proteins having known O-GlcNAc modification site(s) or known tyrosine phosphorylation site(s) revealed a strong potential association between these two posttranslational modifications in various proteins. We speculate that O-GlcNAc modification and tyrosine phosphorylation of PHB play an important role in tyrosine kinase signaling pathways including insulin, growth factors and immune receptors signaling. In addition, we propose that O-GlcNAc modification and tyrosine phosphorylation is a novel previously unidentified binary switch which may provide new mechanistic insights into cell signaling pathways and is open for direct experimental examination.
Notes: 1932-6203
Ande, Sudharsana R
Moulik, Saby
Mishra, Suresh
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2009;4(2):e4586. doi: 10.1371/journal.pone.0004586. Epub 2009 Feb 24.
Author Address: Department of Internal Medicine, University of Manitoba, Winnipeg, Manitoba, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1585
Author: Ande, S. R., Padilla-Meier, G. P. and Mishra, S.
Year: 2013
Title: Mutually exclusive acetylation and ubiquitylation among enzymes involved in glucose metabolism
Journal: Adipocyte
Volume: 2
Issue: 4
Pages: 256-61
Epub Date: 2013/09/21
Date: Oct 1
Short Title: Mutually exclusive acetylation and ubiquitylation among enzymes involved in glucose metabolism
Alternate Journal: Adipocyte
ISSN: 2162-3945 (Print)
2162-3945
DOI: 10.4161/adip.26070
PMCID: PMC3774702
Accession Number: 24052902
Keywords: TCA cycle
gluconeogenesis
glycogen synthesis
glycolysis
phosphorylation
posttranslational modification
Abstract: The posttranslational modification (PTM) in protein occurs in a regiospecific manner. In addition, the most commonly occurring PTMs involve similar residues in proteins such as acetylation, ubiquitylation, methylation and sumoylation at the lysine residue and phosphorylation and O-GlcNAc modification at serine/threonine residues. Thus, the possibility of modification sites where two such PTMs may occur in a mutually exclusive manner (ME-PTM) is much higher than known. A recent surge in the identification and the mapping of the commonly occurring PTMs in proteins has revealed that this is indeed the case. However, in what way such ME-PTM sites are regulated and what could be their relevance in the coordinated network of protein function remains to be known. To gain such potential insights in a biological context, we analyzed two most prevalent PTMs on the lysine residue by acetylation and ubiquitylation along with the most abundant PTM in proteins by phosphorylation among enzymes involved in glucose metabolism, a fundamental process in biology. The analysis of the PTM data sets has revealed two important clues that may be intrinsically associated with their regulation and function. First, the most commonly occurring PTMs by phosphorylation, acetylation and ubiquitylation are widespread and clustered in most of the enzymes involved in glucose metabolism; and the prevalence of phosphorylation sites correlates with the number of acetylation and ubiquitylation sites including the ME-modification sites. Second, the prevalence of ME-acetylation/ubiquitylation sites is exceptionally high among enzymes involved in glucose metabolism and have distinct pattern among the subset of enzymes of glucose metabolism such as glycolysis, tricarboxylic acid (TCA) cycle, glycogen synthesis, and the irreversible steps of gluconeogenesis. We hypothesize that phosphorylation including tyrosine phosphorylation plays an important role in the regulation of ME-acetylation/ubiquitylation sites and their similar pattern among the subset of functionally related proteins allows their coordinated regulation in the normal physiology. Similarly their coordinated dysregulation may underlie the disease processes such as reprogrammed metabolism in cancer, obesity, type 2 diabetes, and cardiovascular diseases. Our hypothesis provides an opportunity to understand the regulation of ME-PTMs in proteins and their relevance at the network level and is open for experimental validation.
Notes: 2162-397x
Ande, Sudharsana Rao
Padilla-Meier, G Pauline
Mishra, Suresh
Journal Article
Adipocyte. 2013 Oct 1;2(4):256-61. doi: 10.4161/adip.26070. Epub 2013 Aug 13.
Author Address: Department of Internal Medicine; University of Manitoba; Winnipeg, MB Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1299
Author: Andersen, C. B., Madsen, M., Storm, T., Moestrup, S. K. and Andersen, G. R.
Year: 2010
Title: Structural basis for receptor recognition of vitamin-B(12)-intrinsic factor complexes
Journal: Nature
Volume: 464
Issue: 7287
Pages: 445-8
Epub Date: 2010/03/20
Date: Mar 18
Short Title: Structural basis for receptor recognition of vitamin-B(12)-intrinsic factor complexes
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature08874
Accession Number: 20237569
Keywords: Aspartic Acid/metabolism
Binding Sites
Calcium/metabolism
Crystallography, X-Ray
Glutamic Acid/metabolism
Humans
Intrinsic Factor/*chemistry/*metabolism
Ligands
Models, Molecular
Protein Binding
Protein Structure, Tertiary
Receptors, Cell Surface/*chemistry/*metabolism
Static Electricity
Vitamin B 12/*chemistry/*metabolism
Abstract: Cobalamin (Cbl, vitamin B(12)) is a bacterial organic compound and an essential coenzyme in mammals, which take it up from the diet. This occurs by the combined action of the gastric intrinsic factor (IF) and the ileal endocytic cubam receptor formed by the 460-kilodalton (kDa) protein cubilin and the 45-kDa transmembrane protein amnionless. Loss of function of any of these proteins ultimately leads to Cbl deficiency in man. Here we present the crystal structure of the complex between IF-Cbl and the cubilin IF-Cbl-binding-region (CUB(5-8)) determined at 3.3 A resolution. The structure provides insight into how several CUB (for 'complement C1r/C1s, Uegf, Bmp1') domains collectively function as modular ligand-binding regions, and how two distant CUB domains embrace the Cbl molecule by binding the two IF domains in a Ca(2+)-dependent manner. This dual-point model provides a probable explanation of how Cbl indirectly induces ligand-receptor coupling. Finally, the comparison of Ca(2+)-binding CUB domains and the low-density lipoprotein (LDL) receptor-type A modules suggests that the electrostatic pairing of a basic ligand arginine/lysine residue with Ca(2+)-coordinating acidic aspartates/glutamates is a common theme of Ca(2+)-dependent ligand-receptor interactions.
Notes: 1476-4687
Andersen, Christian Brix Folsted
Madsen, Mette
Storm, Tina
Moestrup, Søren K
Andersen, Gregers R
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2010 Mar 18;464(7287):445-8. doi: 10.1038/nature08874.
Author Address: Department of Medical Biochemistry, Aarhus University, 8000 Aarhus C, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1138
Author: André, M., Morelle, W., Planchon, S., Milhiet, P. E., Rubinstein, E., Mollicone, R., Chamot-Rooke, J. and Le Naour, F.
Year: 2007
Title: Glycosylation status of the membrane protein CD9P-1
Journal: Proteomics
Volume: 7
Issue: 21
Pages: 3880-95
Epub Date: 2007/10/27
Date: Nov
Short Title: Glycosylation status of the membrane protein CD9P-1
Alternate Journal: Proteomics
ISSN: 1615-9853 (Print)
1615-9853
DOI: 10.1002/pmic.200700355
Accession Number: 17960739
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Cell Line
Cell Line, Tumor
Electrophoresis, Gel, Two-Dimensional
Gas Chromatography-Mass Spectrometry
Glycoside Hydrolases
Glycosylation
Humans
Immunoprecipitation
Molecular Sequence Data
Neoplasm Proteins/*chemistry/genetics/isolation & purification
Polysaccharides/chemistry
Proteomics/*methods
Recombinant Proteins/chemistry/genetics/isolation & purification
Spectrometry, Mass, Electrospray Ionization
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Spectroscopy, Fourier Transform Infrared
Transfection
Abstract: The membrane protein CD9P-1 is a major component of the tetraspanin web, a network of molecular interactions in the plasma membrane, in which it specifically associates with tetraspanins CD9 and CD81. The various functional effects of CD9 and CD81 may be related to their partners. Thus, we have addressed the characterization of the CD9P-1 glycosylation using stably transfected HEK-293 cells. After immunoprecipitation, CD9P-1 was subjected to enzymatic PNGase F cleavage of N-glycans, resulting in Asn to Asp conversion and increase in 1 mass unit. Thus, following protease digestion, deglycosylated peptides were selectively identified by high mass accuracy FTICR-MS, using this conversion as a signature. This has demonstrated that all nine potential N-glycosylation sites were actually engaged. On the other hand, the N-glycan structures were determined combining chemical derivatization and exoglycosidase digestions followed by MALDI-TOF MS, ESI-MS/MS, and GC-MS analysis. CD9P-1 was shown to exhibit more than 40 different N-glycans, essentially composed of complex and high mannose-type structures. Finally, 2-D PAGE and lectino-blot analyses have revealed the presence of at least 17 glycosylated isoforms of CD9P-1 at cell surface. All CD9P-1 isoforms associate with CD9 leading to additional level of complexity of this primary complex in the tetraspanin web.
Notes: André, Magali
Morelle, Willy
Planchon, Sébastien
Milhiet, Pierre-Emmanuel
Rubinstein, Eric
Mollicone, Rosella
Chamot-Rooke, Julia
Le Naour, François
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2007 Nov;7(21):3880-95. doi: 10.1002/pmic.200700355.
Author Address: Inserm U602, Hôpital Paul Brousse, Villejuif, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1266
Author: Andrew, A. J., Miyagi, E., Kao, S. and Strebel, K.
Year: 2009
Title: The formation of cysteine-linked dimers of BST-2/tetherin is important for inhibition of HIV-1 virus release but not for sensitivity to Vpu
Journal: Retrovirology
Volume: 6
Pages: 80
Epub Date: 2009/09/10
Date: Sep 8
Short Title: The formation of cysteine-linked dimers of BST-2/tetherin is important for inhibition of HIV-1 virus release but not for sensitivity to Vpu
Alternate Journal: Retrovirology
ISSN: 1742-4690
DOI: 10.1186/1742-4690-6-80
PMCID: PMC2754425
Accession Number: 19737401
Keywords: Amino Acid Substitution/genetics
Antigens, CD/*immunology/metabolism
Cell Line
Cell Membrane/chemistry
Cysteine/genetics
Dimerization
Disulfides
GPI-Linked Proteins
Glycosylation
HIV-1/*immunology/*physiology
Human Immunodeficiency Virus Proteins/*physiology
Humans
Membrane Glycoproteins/antagonists & inhibitors/*immunology/metabolism
Mutagenesis, Site-Directed
Viral Regulatory and Accessory Proteins/*physiology
*Virus Replication
Abstract: BACKGROUND: The Human Immunodeficiency virus type 1 (HIV-1) Vpu protein enhances virus release from infected cells and induces proteasomal degradation of CD4. Recent work identified BST-2/CD317 as a host factor that inhibits HIV-1 virus release in a Vpu sensitive manner. A current working model proposes that BST-2 inhibits virus release by tethering viral particles to the cell surface thereby triggering their subsequent endocytosis. RESULTS: Here we defined structural properties of BST-2 required for inhibition of virus release and for sensitivity to Vpu. We found that BST-2 is modified by N-linked glycosylation at two sites in the extracellular domain. However, N-linked glycosylation was not important for inhibition of HIV-1 virus release nor did it affect surface expression or sensitivity to Vpu. Rodent BST-2 was previously found to form cysteine-linked dimers. Analysis of single, double, or triple cysteine mutants revealed that any one of three cysteine residues present in the BST-2 extracellular domain was sufficient for BST-2 dimerization, for inhibition of virus release, and sensitivity to Vpu. In contrast, BST-2 lacking all three cysteines in its ectodomain was unable to inhibit release of wild type or Vpu-deficient HIV-1 virions. This defect was not caused by a gross defect in BST-2 trafficking as the mutant protein was expressed at the cell surface of transfected 293T cells and was down-modulated by Vpu similar to wild type BST-2. CONCLUSION: While BST-2 glycosylation was functionally irrelevant, formation of cysteine-linked dimers appeared to be important for inhibition of virus release. However lack of dimerization did not prevent surface expression or Vpu sensitivity of BST-2, suggesting Vpu sensitivity and inhibition of virus release are separable properties of BST-2.
Notes: 1742-4690
Andrew, Amy J
Miyagi, Eri
Kao, Sandra
Strebel, Klaus
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Retrovirology. 2009 Sep 8;6:80. doi: 10.1186/1742-4690-6-80.
Author Address: Laboratory of Molecular Microbiology, Viral Biochemistry Section, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, Maryland 20892-0460, USA. andrewa@niaid.nih.gov
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 791
Author: Angata, K., Yen, T. Y., El-Battari, A., Macher, B. A. and Fukuda, M.
Year: 2001
Title: Unique disulfide bond structures found in ST8Sia IV polysialyltransferase are required for its activity
Journal: J Biol Chem
Volume: 276
Issue: 18
Pages: 15369-77
Epub Date: 2001/03/30
Date: May 4
Short Title: Unique disulfide bond structures found in ST8Sia IV polysialyltransferase are required for its activity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M100576200
Accession Number: 11279095
Keywords: Amino Acid Sequence
Animals
Base Sequence
Chromatography, High Pressure Liquid
Cysteine/metabolism
DNA Primers
DNA, Complementary
Disulfides/chemistry/*metabolism
Insecta
Molecular Sequence Data
Mutagenesis, Site-Directed
Sequence Homology, Amino Acid
Sialyltransferases/chemistry/genetics/*metabolism
Spectrometry, Mass, Electrospray Ionization
Abstract: NCAM polysialylation plays a critical role in neuronal development and regeneration. Polysialylation of the neural cell adhesion molecule (NCAM) is catalyzed by two polysialyltransferases, ST8Sia II (STX) and ST8Sia IV (PST), which contain sialylmotifs L and S conserved in all members of the sialyltransferases. The members of the ST8Sia gene family, including ST8Sia II and ST8Sia IV are unique in having three cysteines in sialylmotif L, one cysteine in sialylmotif S, and one cysteine at the COOH terminus. However, structural information, including how disulfide bonds are formed, has not been determined for any of the sialyltransferases. To obtain insight into the structure/function of ST8Sia IV, we expressed human ST8Sia IV in insect cells, Trichoplusia ni, and found that the enzyme produced in the insect cells catalyzes NCAM polysialylation, although it cannot polysialylate itself ("autopolysialylation"). We also found that ST8Sia IV does not form a dimer through disulfide bonds. By using the same enzyme preparation and performing mass spectrometric analysis, we found that the first cysteine in sialylmotif L and the cysteine in sialylmotif S form a disulfide bridge, whereas the second cysteine in sialylmotif L and the cysteine at the COOH terminus form a second disulfide bridge. Site-directed mutagenesis demonstrated that mutation at cysteine residues involved in the disulfide bridges completely inactivated the enzyme. Moreover, changes in the position of the COOH-terminal cysteine abolished its activity. By contrast, the addition of green fluorescence protein at the COOH terminus of ST8Sia IV did not render the enzyme inactive. These results combined indicate that the sterical structure formed by intramolecular disulfide bonds, which bring the sialylmotifs and the COOH terminus within close proximity, is critical for the catalytic activity of ST8Sia IV.
Notes: Angata, K
Yen, T Y
El-Battari, A
Macher, B A
Fukuda, M
P20 RR11805/RR/NCRR NIH HHS/United States
R01 CA33895/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2001 May 4;276(18):15369-77. doi: 10.1074/jbc.M100576200. Epub 2001 Feb 7.
Author Address: Glycobiology Program, Cancer Research Center, The Burnham Institute, La Jolla, California 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 295
Author: Angel, A. S., Grönberg, G., Krotkiewski, H., Lisowska, E. and Nilsson, B.
Year: 1991
Title: Structural analysis of the N-linked oligosaccharides from murine glycophorin
Journal: Arch Biochem Biophys
Volume: 291
Issue: 1
Pages: 76-88
Epub Date: 1991/11/15
Date: Nov 15
Short Title: Structural analysis of the N-linked oligosaccharides from murine glycophorin
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1016/0003-9861(91)90107-t
Accession Number: 1929437
Keywords: Animals
Carbohydrate Sequence
Chromatography, Ion Exchange
Fabaceae/metabolism
Glycophorins/*chemistry
Magnetic Resonance Spectroscopy
Mass Spectrometry
Mice
Mice, Inbred BALB C
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Plants, Medicinal
Abstract: Glycophorins, isolated from BALB/c mouse erythrocytes, were degraded under mild and strong reductive alkaline conditions and the N-linked oligosaccharides were isolated as alditols. The oligosaccharide alditols were fractionated and purified using gel filtration, concanavalin A-Sepharose affinity chromatography, and high-performance ion-exchange chromatography. Structural analysis was carried out by chemical analyses, periodate oxidation in combination with fast atom bombardment mass spectrometry, and 500-MHz 1H NMR spectroscopy. The results revealed the presence of sialylated biantennary, triantennary, and tetraantennary complex type oligosaccharides, all fucosylated at the innermost N-acetylglucosamine residue. The tri- and tetraantennary oligosaccharide-containing fractions also contained species elongated by one and/or two N-acetyllactosamine (-3Gal beta 1-4GlcNAc beta 1-) sequences. The N-linked oligosaccharides were shown to be combined only with one (the low molecular weight) of the two mouse glycophorins.
Notes: Angel, A S
Grönberg, G
Krotkiewski, H
Lisowska, E
Nilsson, B
Journal Article
Research Support, Non-U.S. Gov't
United States
Arch Biochem Biophys. 1991 Nov 15;291(1):76-88. doi: 10.1016/0003-9861(91)90107-t.
Author Address: BioCarb Technology AB, Lund, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1280
Author: Anthonisen, E. H., Berven, L., Holm, S., Nygård, M., Nebb, H. I. and Grønning-Wang, L. M.
Year: 2010
Title: Nuclear receptor liver X receptor is O-GlcNAc-modified in response to glucose
Journal: J Biol Chem
Volume: 285
Issue: 3
Pages: 1607-15
Epub Date: 2009/11/26
Date: Jan 15
Short Title: Nuclear receptor liver X receptor is O-GlcNAc-modified in response to glucose
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.082685
PMCID: PMC2804318
Accession Number: 19933273
Keywords: Acetylglucosamine/*metabolism
Animals
Cell Line, Tumor
Diabetes Mellitus, Experimental/genetics/metabolism
Glucose/metabolism/*pharmacology
Glycosylation
Humans
Lipid Metabolism/drug effects
Liver/drug effects/metabolism
Liver X Receptors
Male
Mice
Orphan Nuclear Receptors/*metabolism
Promoter Regions, Genetic/genetics
Retinoid X Receptors/metabolism
Sterol Regulatory Element Binding Protein 1/genetics
Streptozocin/pharmacology
Transcriptional Activation/drug effects
Abstract: Post-translational modification of nucleocytoplasmic proteins by O-linked beta-N-acetylglucosamine (O-GlcNAc) has for the last 25 years emerged as an essential glucose-sensing mechanism. The liver X receptors (LXRs) function as nutritional sensors for cholesterol-regulating lipid metabolism, glucose homeostasis, and inflammation. LXRs are shown to be post-translationally modified by phosphorylation, acetylation, and sumoylation, affecting their target gene specificity, stability, and transactivating and transrepressional activity, respectively. In the present study, we show for the first time that LXRalpha and LXRbeta are targets for glucose-hexosamine-derived O-GlcNAc modification in human Huh7 cells. Furthermore, we observed increased hepatic LXRalpha O-GlcNAcylation in vivo in refed mice and in streptozotocin-induced refed diabetic mice. Importantly, induction of LXRalpha O-GlcNAcylation in both mouse models was concomitant with increased expression of the lipogenic gene SREBP-1c (sterol regulatory element-binding protein 1c). Furthermore, glucose increased LXR/retinoic acid receptor-dependent activation of luciferase reporter activity driven by the mouse SREBP-1c promoter via the hexosamine biosynthetic pathway in Huh7 cells. Altogether, our results suggest that O-GlcNAcylation of LXR is a novel mechanism by which LXR acts as a glucose sensor affecting LXR-dependent gene expression, substantiating the crucial role of LXR as a nutritional sensor in lipid and glucose metabolism.
Notes: 1083-351x
Anthonisen, Elin Holter
Berven, Lise
Holm, Sverre
Nygård, Maria
Nebb, Hilde I
Grønning-Wang, Line M
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Jan 15;285(3):1607-15. doi: 10.1074/jbc.M109.082685. Epub 2009 Nov 20.
Author Address: Department of Nutrition, Institute of Basic Medical Sciences, University of Oslo, P. O. Box 1046, Blindern, 0316 Oslo, Norway.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 903
Author: Antipenko, A., Himanen, J. P., van Leyen, K., Nardi-Dei, V., Lesniak, J., Barton, W. A., Rajashankar, K. R., Lu, M., Hoemme, C., Püschel, A. W. and Nikolov, D. B.
Year: 2003
Title: Structure of the semaphorin-3A receptor binding module
Journal: Neuron
Volume: 39
Issue: 4
Pages: 589-98
Epub Date: 2003/08/20
Date: Aug 14
Short Title: Structure of the semaphorin-3A receptor binding module
Alternate Journal: Neuron
ISSN: 0896-6273 (Print)
0896-6273
DOI: 10.1016/s0896-6273(03)00502-6
Accession Number: 12925274
Keywords: Amino Acid Sequence
Animals
Binding Sites
COS Cells
Cell Adhesion Molecules/metabolism
Mice
Models, Theoretical
Molecular Sequence Data
Nerve Tissue Proteins/metabolism
Neuropilins/metabolism
Protein Structure, Tertiary
Semaphorin-3A/*chemistry/metabolism
Signal Transduction
Structural Homology, Protein
Abstract: The semaphorins are a large group of extracellular proteins involved in a variety of processes during development, including neuronal migration and axon guidance. Their distinctive feature is a conserved 500 amino acid semaphorin domain, a ligand-receptor interaction module also present in plexins and scatter-factor receptors. We report the crystal structure of a secreted 65 kDa form of Semaphorin-3A (Sema3A), containing the full semaphorin domain. Unexpectedly, the semaphorin fold is a variation of the beta propeller topology. Analysis of the Sema3A structure and structure-based mutagenesis data identify the neuropilin binding site and suggest a potential plexin interaction site. Based on the structure, we present a model for the initiation of semaphorin signaling and discuss potential similarities with the signaling mechanisms of other beta propeller cell surface receptors, such as integrins and the LDL receptor.
Notes: Antipenko, Alexander
Himanen, Juha-Pekka
van Leyen, Klaus
Nardi-Dei, Vincenzo
Lesniak, Jacob
Barton, William A
Rajashankar, Kanagalaghatta R
Lu, Min
Hoemme, Claudia
Püschel, Andreas W
Nikolov, Dimitar B
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Neuron. 2003 Aug 14;39(4):589-98. doi: 10.1016/s0896-6273(03)00502-6.
Author Address: Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 950
Author: Aono, S., Tokita, Y., Shuo, T., Yamauchi, S., Matsui, F., Nakanishi, K., Hirano, K., Sano, M. and Oohira, A.
Year: 2004
Title: Glycosylation site for chondroitin sulfate on the neural part-time proteoglycan, neuroglycan C
Journal: J Biol Chem
Volume: 279
Issue: 45
Pages: 46536-41
Epub Date: 2004/08/28
Date: Nov 5
Short Title: Glycosylation site for chondroitin sulfate on the neural part-time proteoglycan, neuroglycan C
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M403263200
Accession Number: 15331613
Keywords: Age Factors
Amino Acid Sequence
Animals
Astrocytes/metabolism
Binding Sites
Biotinylation
Blotting, Western
COS Cells
Cell Line
Cell Line, Tumor
Central Nervous System
Cerebellum/metabolism
Chondroitin Sulfates/*chemistry
DNA, Complementary/metabolism
Electrophoresis, Polyacrylamide Gel
Glycosylation
Membrane Proteins/*chemistry
Mice
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutation
Neurons/metabolism
PC12 Cells
Plasmids/metabolism
Protein Binding
Proteoglycans/*chemistry
Rats
Reverse Transcriptase Polymerase Chain Reaction
Sequence Homology, Amino Acid
Telencephalon/metabolism
Time Factors
Transfection
Abstract: Neuroglycan C (NGC) is a membrane-spanning chondroitin sulfate (CS) proteoglycan that is expressed predominantly in the central nervous system (CNS). NGC dramatically changed its structure from a proteoglycan to a nonproteoglycan form with cerebellar development, whereas a small portion of NGC molecules existed in a nonproteoglycan form in the other areas of the mature CNS, suggesting that the CS glycosylation of NGC is developmentally regulated in the whole CNS. As primary cultured neurons and astrocytes from cerebral cortices expressed NGC in a proteoglycan form and in a nonproteoglycan form, respectively, CS glycosylation seems to be regulated differently depending on cell type. To investigate the glycosylation process, cell lines expressing a proteoglycan form of NGC would be favorable experimental models. When a mouse NGC cDNA was transfected into COS 1, PC12D, and Neuro 2a cells, only Neuro 2a cells, a mouse neuroblastoma cell line, expressed NGC bearing CS chains. In PC12D cells, although three intrinsic CS proteoglycans were detected, exogenously expressed NGC did not bear any short CS chains just like NGC in the mature cerebellum. This suggests that the addition of CS chains to the NGC core protein is regulated in a manner different from that of other CS proteoglycans. As the first step in investigating the CS glycosylation mechanism using Neuro 2a cells, we determined the CS attachment site as Ser-123 on the NGC core protein by site-directed mutagenesis. The CS glycosylation was not necessary for intracellular trafficking of NGC to the cell surface at least in Neuro 2a cells.
Notes: Aono, Sachiko
Tokita, Yoshihito
Shuo, Takuya
Yamauchi, Shinobu
Matsui, Fumiko
Nakanishi, Keiko
Hirano, Kanako
Sano, Mamoru
Oohira, Atsuhiko
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2004 Nov 5;279(45):46536-41. doi: 10.1074/jbc.M403263200. Epub 2004 Aug 24.
Author Address: Department of Perinatology, Institute for Developmental Research, Aichi Human Service Center, Kasugai, Aichi 480-0392, Japan. saono@inst-hsc.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 523
Author: Aoyagi, Y., Suzuki, Y., Igarashi, K., Saitoh, A., Oguro, M., Yokota, T., Mori, S., Suda, T., Isemura, M. and Asakura, H.
Year: 1993
Title: Carbohydrate structures of human alpha-fetoprotein of patients with hepatocellular carcinoma: presence of fucosylated and non-fucosylated triantennary glycans
Journal: Br J Cancer
Volume: 67
Issue: 3
Pages: 486-92
Epub Date: 1993/03/01
Date: Mar
Short Title: Carbohydrate structures of human alpha-fetoprotein of patients with hepatocellular carcinoma: presence of fucosylated and non-fucosylated triantennary glycans
Alternate Journal: British journal of cancer
ISSN: 0007-0920 (Print)
0007-0920
DOI: 10.1038/bjc.1993.91
PMCID: PMC1968279
Accession Number: 7679920
Keywords: Carbohydrate Conformation
Carcinoma, Hepatocellular/*chemistry
Chromatography, High Pressure Liquid
Humans
Liver Neoplasms/*chemistry
Polysaccharides/analysis
alpha-Fetoproteins/*chemistry/isolation & purification
Abstract: Chemical structures of the sugar chains of various human alpha-fetoprotein (AFP) species with different affinity for Concanavalin A (Con A) and Lens culinaris agglutinin (LCA) were examined by pyridylamination of their oligosaccharides and stepwise exoglycosidase digestion. Using reversed-phase and size-fractionation high performance liquid chromatography systems we identified six pyridylamino-sugar chains. The Con A-reactive and LCA-nonreactive species of AFP from patients with hepatocellular carcinoma contained a biantennary sugar chain, and the Con A-reactive and LCA-reactive species had a biantennary one with a fucose residue at the innermost N-acetylglucosamine residue. The Con A-nonreactive and LCA-reactive species contained a biantennary sugar chain both with a bisecting-N-acetylglucosamine residue at the trimannosyl core and with a focus residue at the innermost N-acetylglucosamine residue. The Con A-nonreactive and LCA-nonreactive species contained a fucosylated triantennary sugar chain as a major component, and two minor components: a triantennary sugar chain and a biantennary sugar chain with a bisecting-N-acetylglucosamine residue at the trimannosyl core. Thus, the fucosylated and non-fucosylated triantennary sugar chains were newly identified in human AFP. Essentially identical results were obtained for AFP from the patient with gallbladder carcinoma which metastasizes to the liver. These results indicate that the increment in fucosylation and branching to form new antennae is a characteristic feature of the carbohydrate chains of AFP from patients with neoplastic diseases of the liver.
Notes: 1532-1827
Aoyagi, Y
Suzuki, Y
Igarashi, K
Saitoh, A
Oguro, M
Yokota, T
Mori, S
Suda, T
Isemura, M
Asakura, H
Journal Article
Research Support, Non-U.S. Gov't
Br J Cancer. 1993 Mar;67(3):486-92. doi: 10.1038/bjc.1993.91.
Author Address: Department of Internal Medicine, Niigata University School of Medicine, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1140
Author: Appleton, B. A., Wu, P., Maloney, J., Yin, J., Liang, W. C., Stawicki, S., Mortara, K., Bowman, K. K., Elliott, J. M., Desmarais, W., Bazan, J. F., Bagri, A., Tessier-Lavigne, M., Koch, A. W., Wu, Y., Watts, R. J. and Wiesmann, C.
Year: 2007
Title: Structural studies of neuropilin/antibody complexes provide insights into semaphorin and VEGF binding
Journal: Embo j
Volume: 26
Issue: 23
Pages: 4902-12
Epub Date: 2007/11/09
Date: Nov 28
Short Title: Structural studies of neuropilin/antibody complexes provide insights into semaphorin and VEGF binding
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7601906
PMCID: PMC2099469
Accession Number: 17989695
Keywords: Amino Acid Sequence
Antibodies/chemistry
Binding Sites
Crystallography, X-Ray/methods
Dimerization
Molecular Conformation
Molecular Sequence Data
Neuropilins/*chemistry/physiology
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Semaphorin-3A/*chemistry/metabolism
Semaphorins/metabolism
Sequence Homology, Amino Acid
Vascular Endothelial Growth Factor A/*chemistry/metabolism
Abstract: Neuropilins (Nrps) are co-receptors for class 3 semaphorins and vascular endothelial growth factors and important for the development of the nervous system and the vasculature. The extracellular portion of Nrp is composed of two domains that are essential for semaphorin binding (a1a2), two domains necessary for VEGF binding (b1b2), and one domain critical for receptor dimerization (c). We report several crystal structures of Nrp1 and Nrp2 fragments alone and in complex with antibodies that selectively block either semaphorin or vascular endothelial growth factor (VEGF) binding. In these structures, Nrps adopt an unexpected domain arrangement in which the a2, b1, and b2 domains form a tightly packed core that is only loosely connected to the a1 domain. The locations of the antibody epitopes together with in vitro experiments indicate that VEGF and semaphorin do not directly compete for Nrp binding. Based upon our structural and functional data, we propose possible models for ligand binding to neuropilins.
Notes: 1460-2075
Appleton, Brent A
Wu, Ping
Maloney, Janice
Yin, JianPing
Liang, Wei-Ching
Stawicki, Scott
Mortara, Kyle
Bowman, Krista K
Elliott, J Michael
Desmarais, William
Bazan, J Fernando
Bagri, Anil
Tessier-Lavigne, Marc
Koch, Alexander W
Wu, Yan
Watts, Ryan J
Wiesmann, Christian
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
EMBO J. 2007 Nov 28;26(23):4902-12. doi: 10.1038/sj.emboj.7601906. Epub 2007 Nov 8.
Author Address: Department of Protein Engineering, Genentech, Inc., South San Francisco, CA 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1458
Author: Araç, D., Boucard, A. A., Bolliger, M. F., Nguyen, J., Soltis, S. M., Südhof, T. C. and Brunger, A. T.
Year: 2012
Title: A novel evolutionarily conserved domain of cell-adhesion GPCRs mediates autoproteolysis
Journal: Embo j
Volume: 31
Issue: 6
Pages: 1364-78
Epub Date: 2012/02/16
Date: Mar 21
Short Title: A novel evolutionarily conserved domain of cell-adhesion GPCRs mediates autoproteolysis
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2012.26
PMCID: PMC3321182
Accession Number: 22333914
Keywords: Amino Acid Sequence
Animals
Cell Adhesion/genetics
Cells, Cultured
*Conserved Sequence
*Evolution, Molecular
HEK293 Cells
Humans
Hydrolysis
Mice
Models, Molecular
Molecular Sequence Data
Mutation/genetics
Nerve Tissue Proteins/genetics/metabolism
Polycystic Kidney Diseases/genetics/metabolism
Protein Folding
Protein Structure, Tertiary
Proteolysis
Rats
Receptors, G-Protein-Coupled/*chemistry/*genetics/metabolism
Receptors, Peptide/genetics/metabolism
TRPP Cation Channels/genetics/metabolism
Abstract: The G protein-coupled receptor (GPCR) Proteolysis Site (GPS) of cell-adhesion GPCRs and polycystic kidney disease (PKD) proteins constitutes a highly conserved autoproteolysis sequence, but its catalytic mechanism remains unknown. Here, we show that unexpectedly the ∼40-residue GPS motif represents an integral part of a much larger ∼320-residue domain that we termed GPCR-Autoproteolysis INducing (GAIN) domain. Crystal structures of GAIN domains from two distantly related cell-adhesion GPCRs revealed a conserved novel fold in which the GPS motif forms five β-strands that are tightly integrated into the overall GAIN domain. The GAIN domain is evolutionarily conserved from tetrahymena to mammals, is the only extracellular domain shared by all human cell-adhesion GPCRs and PKD proteins, and is the locus of multiple human disease mutations. Functionally, the GAIN domain is both necessary and sufficient for autoproteolysis, suggesting an autoproteolytic mechanism whereby the overall GAIN domain fine-tunes the chemical environment in the GPS to catalyse peptide bond hydrolysis. Thus, the GAIN domain embodies a unique, evolutionarily ancient and widespread autoproteolytic fold whose function is likely relevant for GPCR signalling and for multiple human diseases.
Notes: 1460-2075
Araç, Demet
Boucard, Antony A
Bolliger, Marc F
Nguyen, Jenna
Soltis, S Michael
Südhof, Thomas C
Brunger, Axel T
R37 MH052804/MH/NIMH NIH HHS/United States
R37 MH52804-08/MH/NIMH NIH HHS/United States
Howard Hughes Medical Institute/United States
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
EMBO J. 2012 Mar 21;31(6):1364-78. doi: 10.1038/emboj.2012.26. Epub 2012 Feb 14.
Author Address: Howard Hughes Medical Institute, Stanford, CA, USA. demeta@stanford.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1150
Author: Araç, D., Boucard, A. A., Ozkan, E., Strop, P., Newell, E., Südhof, T. C. and Brunger, A. T.
Year: 2007
Title: Structures of neuroligin-1 and the neuroligin-1/neurexin-1 beta complex reveal specific protein-protein and protein-Ca2+ interactions
Journal: Neuron
Volume: 56
Issue: 6
Pages: 992-1003
Epub Date: 2007/12/21
Date: Dec 20
Short Title: Structures of neuroligin-1 and the neuroligin-1/neurexin-1 beta complex reveal specific protein-protein and protein-Ca2+ interactions
Alternate Journal: Neuron
ISSN: 0896-6273 (Print)
0896-6273
DOI: 10.1016/j.neuron.2007.12.002
Accession Number: 18093522
Keywords: Alternative Splicing
Amino Acid Sequence
Animals
Calcium/*metabolism
Cell Adhesion Molecules, Neuronal
Cells, Cultured
Crystallography/methods
Membrane Proteins/*chemistry/*metabolism
Models, Biological
Models, Molecular
Molecular Sequence Data
Nerve Tissue Proteins/*chemistry/*metabolism
Protein Binding
Protein Conformation
Protein Folding
Rats
Recombinant Proteins
Spectrum Analysis/methods
Surface Plasmon Resonance
Synapses/*physiology
Abstract: Neurexins and neuroligins provide trans-synaptic connectivity by the Ca2+-dependent interaction of their alternatively spliced extracellular domains. Neuroligins specify synapses in an activity-dependent manner, presumably by binding to neurexins. Here, we present the crystal structures of neuroligin-1 in isolation and in complex with neurexin-1 beta. Neuroligin-1 forms a constitutive dimer, and two neurexin-1 beta monomers bind to two identical surfaces on the opposite faces of the neuroligin-1 dimer to form a heterotetramer. The neuroligin-1/neurexin-1 beta complex exhibits a nanomolar affinity and includes a large binding interface that contains bound Ca2+. Alternatively spliced sites in neurexin-1 beta and in neuroligin-1 are positioned nearby the binding interface, explaining how they regulate the interaction. Structure-based mutations of neuroligin-1 at the interface disrupt binding to neurexin-1 beta, but not the folding of neuroligin-1 and confirm the validity of the binding interface of the neuroligin-1/neurexin-1 beta complex. Our results provide molecular insights for understanding the role of cell-adhesion proteins in synapse function.
Notes: Araç, Demet
Boucard, Antony A
Ozkan, Engin
Strop, Pavel
Newell, Evan
Südhof, Thomas C
Brunger, Axel T
R37 MH52804-08/MH/NIMH NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
Neuron. 2007 Dec 20;56(6):992-1003. doi: 10.1016/j.neuron.2007.12.002.
Author Address: Howard Hughes Medical Institute, Stanford University, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 440
Author: Arima, T. and Spiro, R. G.
Year: 1972
Title: Studies on the carbohydrate units of thyroglobulin. Structure of the mannose-N-acetylglucosamine unit (unit A) of the human and calf proteins
Journal: J Biol Chem
Volume: 247
Issue: 6
Pages: 1836-48
Epub Date: 1972/03/25
Date: Mar 25
Short Title: Studies on the carbohydrate units of thyroglobulin. Structure of the mannose-N-acetylglucosamine unit (unit A) of the human and calf proteins
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4335100
Keywords: Amino Acids
Animals
Borohydrides
Cattle
Chemical Phenomena
Chemistry
Chitin/analysis
Chromatography, Paper
Chromatography, Thin Layer
Disaccharides
Ethers
*Glucosamine
Glycopeptides/analysis
Glycoside Hydrolases
Hexosaminidases
Humans
*Mannose
Methylation
Monosaccharides
Oligosaccharides
Oxidation-Reduction
Periodic Acid
Plants/enzymology
Pronase
*Thyroglobulin
Notes: Arima, T
Spiro, R G
Journal Article
United States
J Biol Chem. 1972 Mar 25;247(6):1836-48.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 729
Author: Aritomi, M., Kunishima, N., Okamoto, T., Kuroki, R., Ota, Y. and Morikawa, K.
Year: 1999
Title: Atomic structure of the GCSF-receptor complex showing a new cytokine-receptor recognition scheme
Journal: Nature
Volume: 401
Issue: 6754
Pages: 713-7
Epub Date: 1999/10/28
Date: Oct 14
Short Title: Atomic structure of the GCSF-receptor complex showing a new cytokine-receptor recognition scheme
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/44394
Accession Number: 10537111
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Granulocyte Colony-Stimulating Factor/*chemistry/metabolism
Humans
Mice
Models, Molecular
Molecular Sequence Data
Protein Conformation
Receptors, Granulocyte Colony-Stimulating Factor/*chemistry/metabolism
Sequence Homology, Amino Acid
Abstract: Granulocyte colony-stimulating factor (GCSF) is the principal growth factor regulating the maturation, proliferation and differentiation of the precursor cells of neutrophilic granulocytes and is used to treat neutropenia. GCSF is a member of the long-chain subtype of the class 1 cytokine superfamily, which includes growth hormone, erythropoietin, interleukin 6 and oncostatin M. Here we have determined the crystal structure of GCSF complexed to the BN-BC domains, the principal ligand-binding region of the GCSF receptor (GCSFR). The two receptor domains form a complex in a 2:2 ratio with the ligand, with a non-crystallographic pseudo-twofold axis through primarily the interdomain region and secondarily the BC domain. This structural view of a gp130-type receptor-ligand complex presents a new molecular basis for cytokine-receptor recognition.
Notes: Aritomi, M
Kunishima, N
Okamoto, T
Kuroki, R
Ota, Y
Morikawa, K
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 1999 Oct 14;401(6754):713-7. doi: 10.1038/44394.
Author Address: Biomolecular Engineering Research Institute, Suita-city, Osaka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1515
Author: Arolas, J. L., Broder, C., Jefferson, T., Guevara, T., Sterchi, E. E., Bode, W., Stöcker, W., Becker-Pauly, C. and Gomis-Rüth, F. X.
Year: 2012
Title: Structural basis for the sheddase function of human meprin β metalloproteinase at the plasma membrane
Journal: Proc Natl Acad Sci U S A
Volume: 109
Issue: 40
Pages: 16131-6
Epub Date: 2012/09/19
Date: Oct 2
Short Title: Structural basis for the sheddase function of human meprin β metalloproteinase at the plasma membrane
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1211076109
PMCID: PMC3479590
Accession Number: 22988105
Keywords: Cell Membrane/*metabolism
Crystallography
Dimerization
Humans
Metalloendopeptidases/*chemistry/*metabolism
*Models, Molecular
Protein Binding
*Protein Conformation
Protein Structure, Tertiary
Abstract: Ectodomain shedding at the cell surface is a major mechanism to regulate the extracellular and circulatory concentration or the activities of signaling proteins at the plasma membrane. Human meprin β is a 145-kDa disulfide-linked homodimeric multidomain type-I membrane metallopeptidase that sheds membrane-bound cytokines and growth factors, thereby contributing to inflammatory diseases, angiogenesis, and tumor progression. In addition, it cleaves amyloid precursor protein (APP) at the β-secretase site, giving rise to amyloidogenic peptides. We have solved the X-ray crystal structure of a major fragment of the meprin β ectoprotein, the first of a multidomain oligomeric transmembrane sheddase, and of its zymogen. The meprin β dimer displays a compact shape, whose catalytic domain undergoes major rearrangement upon activation, and reveals an exosite and a sugar-rich channel, both of which possibly engage in substrate binding. A plausible structure-derived working mechanism suggests that substrates such as APP are shed close to the plasma membrane surface following an "N-like" chain trace.
Notes: 1091-6490
Arolas, Joan L
Broder, Claudia
Jefferson, Tamara
Guevara, Tibisay
Sterchi, Erwin E
Bode, Wolfram
Stöcker, Walter
Becker-Pauly, Christoph
Gomis-Rüth, F Xavier
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2012 Oct 2;109(40):16131-6. doi: 10.1073/pnas.1211076109. Epub 2012 Sep 17.
Author Address: Proteolysis Laboratory, Department of Structural Biology, Molecular Biology Institute of Barcelona, Consejo Superior de Investigaciones Cientificas, Barcelona Science Park, E-08028 Barcelona, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1534
Author: Arshad, N., Ballal, S. and Visweswariah, S. S.
Year: 2013
Title: Site-specific N-linked glycosylation of receptor guanylyl cyclase C regulates ligand binding, ligand-mediated activation and interaction with vesicular integral membrane protein 36, VIP36
Journal: J Biol Chem
Volume: 288
Issue: 6
Pages: 3907-17
Epub Date: 2012/12/28
Date: Feb 8
Short Title: Site-specific N-linked glycosylation of receptor guanylyl cyclase C regulates ligand binding, ligand-mediated activation and interaction with vesicular integral membrane protein 36, VIP36
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.413906
PMCID: PMC3567644
Accession Number: 23269669
Keywords: Cell Line
Cell Membrane/genetics/*metabolism
Endoplasmic Reticulum/genetics/*metabolism
Gastrointestinal Hormones/genetics/metabolism
Glycosylation
Humans
Ligands
Mannose-Binding Lectins/genetics/*metabolism
Membrane Transport Proteins/genetics/*metabolism
Natriuretic Peptides/genetics/metabolism
Protein Binding
*Protein Folding
Protein Structure, Tertiary
Receptors, Enterotoxin
Receptors, Guanylate Cyclase-Coupled/genetics/*metabolism
Receptors, Peptide/genetics/*metabolism
Abstract: Guanylyl cyclase C (GC-C) is a multidomain, membrane-associated receptor guanylyl cyclase. GC-C is primarily expressed in the gastrointestinal tract, where it mediates fluid-ion homeostasis, intestinal inflammation, and cell proliferation in a cGMP-dependent manner, following activation by its ligands guanylin, uroguanylin, or the heat-stable enterotoxin peptide (ST). GC-C is also expressed in neurons, where it plays a role in satiation and attention deficiency/hyperactive behavior. GC-C is glycosylated in the extracellular domain, and differentially glycosylated forms that are resident in the endoplasmic reticulum (130 kDa) and the plasma membrane (145 kDa) bind the ST peptide with equal affinity. When glycosylation of human GC-C was prevented, either by pharmacological intervention or by mutation of all of the 10 predicted glycosylation sites, ST binding and surface localization was abolished. Systematic mutagenesis of each of the 10 sites of glycosylation in GC-C, either singly or in combination, identified two sites that were critical for ligand binding and two that regulated ST-mediated activation. We also show that GC-C is the first identified receptor client of the lectin chaperone vesicular integral membrane protein, VIP36. Interaction with VIP36 is dependent on glycosylation at the same sites that allow GC-C to fold and bind ligand. Because glycosylation of proteins is altered in many diseases and in a tissue-dependent manner, the activity and/or glycan-mediated interactions of GC-C may have a crucial role to play in its functions in different cell types.
Notes: 1083-351x
Arshad, Najla
Ballal, Suhas
Visweswariah, Sandhya S
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2013 Feb 8;288(6):3907-17. doi: 10.1074/jbc.M112.413906. Epub 2012 Dec 26.
Author Address: Department of Molecular Reproduction, Development, and Genetics, Indian Institute of Science, Bangalore 560012, India.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 818
Author: Assil, I. Q. and Abou-Samra, A. B.
Year: 2001
Title: N-glycosylation of CRF receptor type 1 is important for its ligand-specific interaction
Journal: Am J Physiol Endocrinol Metab
Volume: 281
Issue: 5
Pages: E1015-21
Epub Date: 2001/10/12
Date: Nov
Short Title: N-glycosylation of CRF receptor type 1 is important for its ligand-specific interaction
Alternate Journal: American journal of physiology. Endocrinology and metabolism
ISSN: 0193-1849 (Print)
0193-1849
DOI: 10.1152/ajpendo.2001.281.5.E1015
Accession Number: 11595658
Keywords: 1-Methyl-3-isobutylxanthine/pharmacology
Amphibian Proteins
Animals
Binding Sites
Binding, Competitive
Cell Line
Consensus Sequence
Corticotropin-Releasing Hormone/metabolism
Cyclic AMP/metabolism/pharmacology
Electrophoresis, Polyacrylamide Gel
Glycosylation
Ligands
Mice
Molecular Weight
Mutagenesis, Site-Directed
Peptide Hormones
Peptides/metabolism
Radioligand Assay
Receptors, Corticotropin-Releasing Hormone/chemistry/genetics/*metabolism
Structure-Activity Relationship
Tunicamycin/pharmacology
Urotensins/metabolism
Abstract: The corticotropin-releasing factor (CRF) receptor type 1 (CRFR1) contains five potential N-glycosylation sites: N38, N45, N78, N90, and N98. Cells expressing CRFR1 were treated with tunicamycin to block receptor glycosylation. The nonglycosylated receptor did not bind the radioligand and had a decreased cAMP stimulation potency in response to CRF. To determine which of the polysaccharide chain(s) is/are involved in ligand interaction, the polysaccharide chains were deleted using site-directed mutagenesis of the glycosylation consensus, N-X-S/T. Two sets of mutations were performed for each glycosylation site: N to Q and S/T to A, respectively. The single mutants Q38, Q45, Q78, Q90, Q98, A40, A47, A80, A92, and A100 and the double mutants A40/A47 and A80/A100 were well expressed, bound CRF, sauvagine (SVG), and urotensin-I (UTS-I) with a normal affinity, and increased cAMP accumulation with a high efficiency. In contrast, the combined mutations A80/A92/A100, A40/A80/A92/A100, and A40/A47/A80/A92/A100 had low levels of expression, did not bind the radioligand, and had a decreased cAMP stimulation. These data indicate the requirement for three or more polysaccharide chains for normal CRFR1 function.
Notes: Assil, I Q
Abou-Samra, A B
2R01 DK 45020/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Am J Physiol Endocrinol Metab. 2001 Nov;281(5):E1015-21. doi: 10.1152/ajpendo.2001.281.5.E1015.
Author Address: Endocrine Unit, Department of Medicine, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1066
Author: Attrill, H., Imamura, A., Sharma, R. S., Kiso, M., Crocker, P. R. and van Aalten, D. M.
Year: 2006
Title: Siglec-7 undergoes a major conformational change when complexed with the alpha(2,8)-disialylganglioside GT1b
Journal: J Biol Chem
Volume: 281
Issue: 43
Pages: 32774-83
Epub Date: 2006/08/10
Date: Oct 27
Short Title: Siglec-7 undergoes a major conformational change when complexed with the alpha(2,8)-disialylganglioside GT1b
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M601714200
Accession Number: 16895906
Keywords: Amino Acid Sequence
Animals
Antigens, CD/chemistry/metabolism
Binding, Competitive
CHO Cells
COS Cells
Chlorocebus aethiops
Cricetinae
Crystallography, X-Ray
Gangliosides/chemical synthesis/chemistry/*metabolism
Humans
Hydrogen Bonding
Hydrophobic and Hydrophilic Interactions
Lectins/chemistry/genetics/*metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Molecular Structure
Mutagenesis, Site-Directed
Oligosaccharides/chemical synthesis/chemistry/metabolism
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Receptors, Cell Surface/metabolism
Sequence Homology, Amino Acid
Sialic Acid Binding Immunoglobulin-like Lectins
Abstract: The siglecs are a group of mammalian sialic acid binding receptors expressed predominantly in the immune system. The CD33-related siglecs show complex recognition patterns for sialylated glycans. Siglec-7 shows a preference for alpha(2,8)-disialylated ligands and provides a structural template for studying the key interactions that drive this selectivity. We have co-crystallized Siglec-7 with a synthetic oligosaccharide corresponding to the alpha(2,8)-disialylated ganglioside GT1b. The crystal structure of the complex offers a first glimpse into how this important family of lectins binds the structurally diverse gangliosides. The structure reveals that the C-C' loop, a region implicated in previous studies as driving siglec specificity, undergoes a dramatic conformational shift, allowing it to interact with the underlying neutral glycan core of the ganglioside. The structural data in combination with mutagenesis studies show that binding of the ganglioside is driven by extensive hydrophobic contacts together with key polar interactions and that the binding site structure is complementary to preferred solution conformations of GT1b.
Notes: Attrill, Helen
Imamura, Akihiro
Sharma, Ritu S
Kiso, Makoto
Crocker, Paul R
van Aalten, Daan M F
GM 62116/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Oct 27;281(43):32774-83. doi: 10.1074/jbc.M601714200. Epub 2006 Aug 8.
Author Address: Division of Biological Chemistry and Molecular Microbiology, School of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1040
Author: Attrill, H., Takazawa, H., Witt, S., Kelm, S., Isecke, R., Brossmer, R., Ando, T., Ishida, H., Kiso, M., Crocker, P. R. and van Aalten, D. M.
Year: 2006
Title: The structure of siglec-7 in complex with sialosides: leads for rational structure-based inhibitor design
Journal: Biochem J
Volume: 397
Issue: 2
Pages: 271-8
Epub Date: 2006/04/21
Date: Jul 15
Short Title: The structure of siglec-7 in complex with sialosides: leads for rational structure-based inhibitor design
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20060103
PMCID: PMC1513286
Accession Number: 16623661
Keywords: Animals
Antigens, Differentiation, Myelomonocytic/*chemistry/*physiology
CHO Cells
COS Cells
Chlorocebus aethiops
Cricetinae
Crystallography, X-Ray
Drug Design
Erythrocytes/metabolism
Humans
Lectins/*chemistry/*physiology
Leukocytes/metabolism
Molecular Conformation
N-Acetylneuraminic Acid/chemistry
Sialic Acids/chemistry
Abstract: Siglecs (sialic acid binding Ig-like lectins) are transmembrane receptors for sialylated glycoconjugates that modulate cellular interactions and signalling events in the haematopoietic, immune and nervous systems. Siglec-7 is a structural prototype for the recently described family of immune inhibitory CD33-related siglecs and is predominantly expressed on natural killer cells and monocytes, as well as subsets of CD8 T-cells. Siglec-specific inhibitors are desired for the detection of masked and unmasked forms of siglecs, to aid in dissection of signalling pathways and as tools to investigate siglecs as potential therapeutic targets. As a first step towards this end, we present the crystal structure of siglec-7 in complex with a sialylated ligand, the ganglioside analogue DSLc4 [alpha(2,3)/alpha(2,6) disialyl lactotetraosyl 2-(trimethylsilyl)ethyl], which allows for a detailed description of the binding site, required for structure-guided inhibitor design. Mutagenesis and binding assays were used to demonstrate a key structural role for Lys131, a residue that changes conformation upon sialic acid binding. Differences between the binding sites of siglec family members were then exploited using alpha-methyl Neu5Ac (N-acetylneuraminic acid) as a basic scaffold. A co-crystal of siglec-7 in complex with the sialoside inhibitor, oxamido-Neu5Ac [methyl alpha-9-(amino-oxalyl-amino)-9-deoxy-Neu5Ac] and inhibition data for the sialosides gives clear leads for future inhibitor design.
Notes: 1470-8728
Attrill, Helen
Takazawa, Hirokazu
Witt, Simone
Kelm, Soerge
Isecke, Rainer
Brossmer, Reinhard
Ando, Takayuki
Ishida, Hideharu
Kiso, Makoto
Crocker, Paul R
van Aalten, Daan M F
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2006 Jul 15;397(2):271-8. doi: 10.1042/BJ20060103.
Author Address: Division of Biological Chemistry, School of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1898
Author: Autzen, H. E., Myasnikov, A. G., Campbell, M. G., Asarnow, D., Julius, D. and Cheng, Y.
Year: 2018
Title: Structure of the human TRPM4 ion channel in a lipid nanodisc
Journal: Science
Volume: 359
Issue: 6372
Pages: 228-232
Epub Date: 2017/12/09
Date: Jan 12
Short Title: Structure of the human TRPM4 ion channel in a lipid nanodisc
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.aar4510
PMCID: PMC5898196
NIHMSID: NIHMS955925
Accession Number: 29217581
Keywords: Binding Sites
Calcium/chemistry/metabolism
Cryoelectron Microscopy
Humans
Hydrophobic and Hydrophilic Interactions
Lipids
Models, Molecular
Nanostructures
Protein Conformation
Protein Domains
Protein Structure, Secondary
Recombinant Proteins/chemistry/metabolism/ultrastructure
TRPM Cation Channels/*chemistry/metabolism/ultrastructure
Abstract: Transient receptor potential (TRP) melastatin 4 (TRPM4) is a widely expressed cation channel associated with a variety of cardiovascular disorders. TRPM4 is activated by increased intracellular calcium in a voltage-dependent manner but, unlike many other TRP channels, is permeable to monovalent cations only. Here we present two structures of full-length human TRPM4 embedded in lipid nanodiscs at ~3-angstrom resolution, as determined by single-particle cryo-electron microscopy. These structures, with and without calcium bound, reveal a general architecture for this major subfamily of TRP channels and a well-defined calcium-binding site within the intracellular side of the S1-S4 domain. The structures correspond to two distinct closed states. Calcium binding induces conformational changes that likely prime the channel for voltage-dependent opening.
Notes: 1095-9203
Autzen, Henriette E
Orcid: 0000-0003-3691-5010
Myasnikov, Alexander G
Orcid: 0000-0003-2607-7121
Campbell, Melody G
Orcid: 0000-0003-1909-5751
Asarnow, Daniel
Orcid: 0000-0001-7870-5308
Julius, David
Orcid: 0000-0002-6365-4867
Cheng, Yifan
Orcid: 0000-0001-9535-0369
HHMI/Howard Hughes Medical Institute/United States
S10 OD021741/OD/NIH HHS/United States
R01 GM098672/GM/NIGMS NIH HHS/United States
S10 OD020054/OD/NIH HHS/United States
R01 NS047723/NS/NINDS NIH HHS/United States
T32 GM008284/GM/NIGMS NIH HHS/United States
R35 NS105038/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2018 Jan 12;359(6372):228-232. doi: 10.1126/science.aar4510. Epub 2017 Dec 7.
Author Address: Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94143, USA.
Department of Molecular Biology and Genetics, University of Aarhus, 8000 Aarhus, Denmark.
Department of Physiology, University of California, San Francisco, CA 94143, USA.
Howard Hughes Medical Institute, University of California, San Francisco, CA 94143, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 473
Author: Avvakumov, G. V., Matveentseva, I. V., Akhrem, L. V., Strel'chyonok, O. A. and Akhrem, A. A.
Year: 1983
Title: Study of the carbohydrate moiety of human serum sex hormone-binding globulin
Journal: Biochim Biophys Acta
Volume: 760
Issue: 1
Pages: 104-10
Epub Date: 1983/10/04
Date: Oct 4
Short Title: Study of the carbohydrate moiety of human serum sex hormone-binding globulin
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/0304-4165(83)90130-7
Accession Number: 6684483
Keywords: Amino Acids/analysis
Carbohydrates/analysis
Chromatography, Affinity
Concanavalin A
Glycopeptides/analysis
Humans
Methylation
Sex Hormone-Binding Globulin/*isolation & purification
Abstract: Sex hormone-binding globulin from human blood serum contains two biantennary N-linked oligosaccharide chains of the N-acetyllactosamine type and one O-linked oligosaccharide per one molecule of the glycoprotein. These conclusions have been based on the results of methylation analysis of the whole glycoprotein and investigation of the structures of its glycopeptides prepared using pronase digestion.
Notes: Avvakumov, G V
Matveentseva, I V
Akhrem, L V
Strel'chyonok, O A
Akhrem, A A
Journal Article
Netherlands
Biochim Biophys Acta. 1983 Oct 4;760(1):104-10. doi: 10.1016/0304-4165(83)90130-7.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1143
Author: Azuma, Y., Miura, K., Higai, K. and Matsumoto, K.
Year: 2007
Title: Protein O-N-acetylglucosaminylation modulates promoter activities of cyclic AMP response element and activator protein 1 and enhances E-selectin expression on HuH-7 human hepatoma cells
Journal: Biol Pharm Bull
Volume: 30
Issue: 12
Pages: 2284-9
Epub Date: 2007/12/07
Date: Dec
Short Title: Protein O-N-acetylglucosaminylation modulates promoter activities of cyclic AMP response element and activator protein 1 and enhances E-selectin expression on HuH-7 human hepatoma cells
Alternate Journal: Biological & pharmaceutical bulletin
ISSN: 0918-6158 (Print)
0918-6158
DOI: 10.1248/bpb.30.2284
Accession Number: 18057713
Keywords: Acetylglucosamine/*metabolism
CREB-Binding Protein/metabolism
Carcinoma, Hepatocellular/*metabolism
Cell Line, Tumor
Cyclic AMP Response Element Modulator/*biosynthesis/genetics
E-Selectin/*biosynthesis
Glucose/pharmacology
Humans
Immunoblotting
Interleukin-1beta/pharmacology
Liver Neoplasms/*metabolism
Luciferases/metabolism
Promoter Regions, Genetic/genetics/*physiology
Receptors, Cell Surface
Transcription Factor AP-1/*biosynthesis/genetics
Abstract: High glucose accelerates O-N-acetylglucosaminylation (O-GlcNAcylation) of proteins and causes diabetic complications. In the present study, we found that treatment of HuH-7 human hepatoma cells with high glucose or the protein O-N-acetylglucosaminidase (O-GlcNAcase) inhibitor O-(2-acetoamide-2-deoxy-D-glucopyranosylidene)amino-N-phenylcarbamate (PUGNAc) increased the cell surface expression of E-selectin. A dual luciferase reporter assay indicated that high glucose and PUGNAc suppressed promoter activities of the cyclic AMP response element (CRE) and enhanced those of activator protein 1 (AP-1). Enhanced CRE promoter activities in HuH-7 cells treated with dibutyryl cAMP or co-transfected with a protein kinase A expression vector pFC-PKA that enhances the phosphorylation of CRE binding protein (CREB) were suppressed by PUGNAc. In contrast, PUGNAc further increased the enhanced AP-1 promoter activity in cells transfected with a mitogen-activated protein kinase kinase kinase expression vector pFC-MEKK that enhances c-Jun phosphorylation. Immuno-blotting using an anti-O-GlcNAc antibody revealed that high glucose and PUGNAc accelerated protein O-GlcNAcylation and that there were substantial differences in the O-GlcNAcylated proteins in the cytoplasmic and nuclear fractions. In addition, PUGNAc increased the nuclear import of O-GlcNAcylated CREB. These results suggest that protein O-GlcNAcylation modulates the promoter activities of E-selectin gene, suppression of CRE and enhancement of AP-1, and enhances E-selectin protein expression on hepatocytes.
Notes: Azuma, Yutaro
Miura, Kana
Higai, Koji
Matsumoto, Kojiro
Journal Article
Research Support, Non-U.S. Gov't
Japan
Biol Pharm Bull. 2007 Dec;30(12):2284-9. doi: 10.1248/bpb.30.2284.
Author Address: Department of Clinical Chemistry, School of Pharmaceutical Sciences, Toho University, 2-2-1 Miyama, Funabashi, Chiba247-8510, Japan. yutaro@phar.toho-u.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1517
Author: Bader, H. L., Wang, L. W., Ho, J. C., Tran, T., Holden, P., Fitzgerald, J., Atit, R. P., Reinhardt, D. P. and Apte, S. S.
Year: 2012
Title: A disintegrin-like and metalloprotease domain containing thrombospondin type 1 motif-like 5 (ADAMTSL5) is a novel fibrillin-1-, fibrillin-2-, and heparin-binding member of the ADAMTS superfamily containing a netrin-like module
Journal: Matrix Biol
Volume: 31
Issue: 7-8
Pages: 398-411
Epub Date: 2012/09/27
Date: Sep-Oct
Short Title: A disintegrin-like and metalloprotease domain containing thrombospondin type 1 motif-like 5 (ADAMTSL5) is a novel fibrillin-1-, fibrillin-2-, and heparin-binding member of the ADAMTS superfamily containing a netrin-like module
Alternate Journal: Matrix biology : journal of the International Society for Matrix Biology
ISSN: 0945-053X (Print)
0945-053x
DOI: 10.1016/j.matbio.2012.09.003
PMCID: PMC3546522
NIHMSID: NIHMS413290
Accession Number: 23010571
Keywords: ADAM Proteins/genetics/*metabolism
ADAMTS Proteins
Alternative Splicing/genetics
Amino Acid Sequence
Animals
Base Sequence
Blotting, Western
Fibrillin-1
Fibrillin-2
Fibrillins
Fluorescent Antibody Technique
HEK293 Cells
Heparin/*metabolism
Humans
Immunohistochemistry
In Situ Hybridization
Mice
Microfibrils/*metabolism
Microfilament Proteins/*metabolism
Molecular Sequence Data
Muscle, Skeletal/metabolism
Nerve Growth Factors/genetics/metabolism
Netrin-1
Recombinant Proteins/*metabolism
Reverse Transcriptase Polymerase Chain Reaction
Sequence Analysis, DNA
Thrombospondin 1/genetics/*metabolism
Tumor Suppressor Proteins/genetics/metabolism
Abstract: ADAMTS-like proteins are related to ADAMTS metalloproteases by their similarity to ADAMTS ancillary domains. Here, we have characterized ADAMTSL5, a novel member of the superfamily with a unique modular organization that includes a single C-terminal netrin-like (NTR) module. Alternative splicing of ADAMTSL5 at its 5' end generates two transcripts that encode different signal peptides, but the same mature protein. These transcripts differ in their translational efficiency. Recombinant ADAMTSL5 is a secreted, N-glycosylated 60kDa glycoprotein located in the subcellular matrix, on the cell-surface, and in the medium of transfected cells. RT-PCR and western blot analysis of adult mouse tissues showed broad expression. Western blot analysis suggested proteolytic release of the NTR module in transfected cells as well as in some mouse tissues. Immunostaining during mouse organogenesis identified ADAMTSL5 in musculoskeletal tissues such as skeletal muscle, cartilage and bone, as well as in many epithelia. Affinity-chromatography demonstrated heparin-binding of ADAMTSL5 through its NTR-module. Recombinant ADAMTSL5 bound to both fibrillin-1 and fibrillin-2, and co-localized with fibrillin microfibrils in the extracellular matrix of cultured fibroblasts, but without discernible effect on microfibril assembly. ADAMTSL5 is the first family member shown to bind both fibrillin-1 and fibrillin-2. Like other ADAMTS proteins implicated in microfibril biology through identification of human and animal mutations, ADAMTSL5 could have a role in modulating microfibril functions.
Notes: 1569-1802
Bader, Hannah L
Wang, Lauren W
Ho, Jason C
Tran, Thu
Holden, Paul
Fitzgerald, Jamie
Atit, Radhika P
Reinhardt, Dieter P
Apte, Suneel S
102821-1/Canadian Institutes of Health Research/Canada
R01AR055957/AR/NIAMS NIH HHS/United States
T32 HL007914/HL/NHLBI NIH HHS/United States
AR53890/AR/NIAMS NIH HHS/United States
EY021151/EY/NEI NIH HHS/United States
R01DE01870/DE/NIDCR NIH HHS/United States
UL1 TR000439/TR/NCATS NIH HHS/United States
R01 AR055957/AR/NIAMS NIH HHS/United States
T32HL007914-08/HL/NHLBI NIH HHS/United States
R01 EY021151/EY/NEI NIH HHS/United States
R01 AR053890/AR/NIAMS NIH HHS/United States
106494-1/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, N.I.H., Extramural
Matrix Biol. 2012 Sep-Oct;31(7-8):398-411. doi: 10.1016/j.matbio.2012.09.003. Epub 2012 Sep 23.
Author Address: Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic, Cleveland, OH 44195, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 443
Author: Baenziger, J. and Kornfeld, S.
Year: 1974
Title: Structure of the carbohydrate units of IgA1 immunoglobulin. II. Structure of the O-glycosidically linked oligosaccharide units
Journal: J Biol Chem
Volume: 249
Issue: 22
Pages: 7270-81
Epub Date: 1974/11/25
Date: Nov 25
Short Title: Structure of the carbohydrate units of IgA1 immunoglobulin. II. Structure of the O-glycosidically linked oligosaccharide units
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4373463
Keywords: Amino Acid Sequence
Amino Acids/analysis
Chromatography, Gas
Chromatography, Gel
Chromatography, Ion Exchange
Chromatography, Paper
Galactosamine/analysis
Galactose/analysis
Glycopeptides/analysis
Humans
Hydrogen-Ion Concentration
Hydrolysis
*Immunoglobulin A
Mass Spectrometry
*Myeloma Proteins
Oligosaccharides/analysis
Oxidation-Reduction
Periodic Acid
Sugar Alcohols/analysis
Time Factors
Notes: Baenziger, J
Kornfeld, S
Journal Article
United States
J Biol Chem. 1974 Nov 25;249(22):7270-81.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 444
Author: Baenziger, J. and Kornfeld, S.
Year: 1974
Title: Structure of the carbohydrate units of IgA1 immunoglobulin. I. Composition, glycopeptide isolation, and structure of the asparagine-linked oligosaccharide units
Journal: J Biol Chem
Volume: 249
Issue: 22
Pages: 7260-9
Epub Date: 1974/11/25
Date: Nov 25
Short Title: Structure of the carbohydrate units of IgA1 immunoglobulin. I. Composition, glycopeptide isolation, and structure of the asparagine-linked oligosaccharide units
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4436308
Keywords: Amino Acid Sequence
Amino Acids/analysis
Asparagine/analysis
Chromatography, DEAE-Cellulose
Chromatography, Gas
Chromatography, Gel
Chromatography, Ion Exchange
Fucose/analysis
Galactosamine/analysis
Galactose/analysis
Glucosamine/analysis
Glycopeptides/isolation & purification
Glycoside Hydrolases
Humans
Immunoelectrophoresis
*Immunoglobulin A
Mannose/analysis
Mass Spectrometry
Molecular Weight
*Myeloma Proteins
Oligosaccharides/analysis
Oxidation-Reduction
Pronase
Sialic Acids/analysis
Spectrophotometry, Ultraviolet
Notes: Baenziger, J
Kornfeld, S
Journal Article
United States
J Biol Chem. 1974 Nov 25;249(22):7260-9.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 441
Author: Baenziger, J., Kornfeld, S. and Kochwa, S.
Year: 1974
Title: Structure of the carbohydrate units of IgE immunoglobulin. I. Over-all composition, glycopeptide isolation, and structure of the high mannose oligosaccharide unit
Journal: J Biol Chem
Volume: 249
Issue: 6
Pages: 1889-96
Epub Date: 1974/03/25
Date: Mar 25
Short Title: Structure of the carbohydrate units of IgE immunoglobulin. I. Over-all composition, glycopeptide isolation, and structure of the high mannose oligosaccharide unit
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4361830
Keywords: Acetamides/analysis
Amino Acids/analysis
Chemical Phenomena
Chemistry
Fucose/analysis
Galactose/analysis
Galactosidases
Glucosamine/analysis
Glycopeptides/analysis/*isolation & purification
Glycoside Hydrolases
Hexosaminidases
Humans
Immunoglobulin E/*analysis
Immunoglobulin Fragments/analysis
Mannose/analysis
Methylation
Molecular Weight
Myeloma Proteins/*analysis
Neuraminic Acids/analysis
Neuraminidase
Oligosaccharides/*analysis
Oxidation-Reduction
Peptide Hydrolases
Periodic Acid
Notes: Baenziger, J
Kornfeld, S
Kochwa, S
Journal Article
United States
J Biol Chem. 1974 Mar 25;249(6):1889-96.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 442
Author: Baenziger, J., Kornfeld, S. and Kochwa, S.
Year: 1974
Title: Structure of the carbohydrate units of IgE immunoglobulin. II. Sequence of the sialic acid-containing glycopeptides
Journal: J Biol Chem
Volume: 249
Issue: 6
Pages: 1897-903
Epub Date: 1974/03/25
Date: Mar 25
Short Title: Structure of the carbohydrate units of IgE immunoglobulin. II. Sequence of the sialic acid-containing glycopeptides
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4361831
Keywords: Acetates/analysis
Asparagine/analysis
Borohydrides
Chemical Phenomena
Chemistry
Fucose/analysis
Galactose/analysis
Glucosamine/analysis
Glycopeptides/*analysis/isolation & purification
Glycoside Hydrolases
Humans
Hydrogen-Ion Concentration
Immunoglobulin E/*analysis
Immunoglobulins/analysis
Mannose/analysis
Methylation
Molecular Weight
Myeloma Proteins/*analysis
Neuraminic Acids/*analysis
Oligosaccharides/*analysis
Periodic Acid
Notes: Baenziger, J
Kornfeld, S
Kochwa, S
Journal Article
United States
J Biol Chem. 1974 Mar 25;249(6):1897-903.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 218
Author: Baenziger, J. U.
Year: 1979
Title: Structure of the oligosaccharide of human J chain
Journal: J Biol Chem
Volume: 254
Issue: 10
Pages: 4063-71
Epub Date: 1979/05/25
Date: May 25
Short Title: Structure of the oligosaccharide of human J chain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 108268
Keywords: Amino Acids/analysis
Carbohydrates/analysis
Humans
*Immunoglobulin J-Chains
*Immunoglobulin M
Molecular Conformation
Myeloma Proteins
Oligosaccharides/analysis
Abstract: The complete structure of the oligosaccharide moiety of J chain isolated from a Waldenströms macroglobulin Wa has been established. The oligosaccharide is present in three forms differing in the amount of sialic acid and designated J-A, J-B, and J-C. The structure and proportion of each of these is: formula : (see text) : Removal of the oligosaccharide moiety with glycosidases results in an increased mobility of J chain in sodium dodecyl sulfate polyacrylamide gel electrophoresis corresponding to a shift in apparent molecular weight from 23,500 to 19,500. The preparation and utilization of iodinated glycopeptides for sequence analysis is presented.
Notes: Baenziger, J U
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1979 May 25;254(10):4063-71.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 847
Author: Baglin, T. P., Carrell, R. W., Church, F. C., Esmon, C. T. and Huntington, J. A.
Year: 2002
Title: Crystal structures of native and thrombin-complexed heparin cofactor II reveal a multistep allosteric mechanism
Journal: Proc Natl Acad Sci U S A
Volume: 99
Issue: 17
Pages: 11079-84
Epub Date: 2002/08/10
Date: Aug 20
Short Title: Crystal structures of native and thrombin-complexed heparin cofactor II reveal a multistep allosteric mechanism
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.162232399
PMCID: PMC123213
Accession Number: 12169660
Keywords: Allosteric Regulation
Amino Acid Substitution
Antithrombins/chemistry
Binding Sites
Crystallography, X-Ray
Heparin Cofactor II/*chemistry
Kinetics
Models, Molecular
Mutagenesis, Site-Directed
Protein Conformation
Recombinant Proteins/chemistry
Serpins/chemistry
Thrombin/*chemistry/genetics
Abstract: The serine proteases sequentially activated to form a fibrin clot are inhibited primarily by members of the serpin family, which use a unique beta-sheet expansion mechanism to trap and destroy their targets. Since the discovery that serpins were a family of serine protease inhibitors there has been controversy as to the role of conformational change in their mechanism. It now is clear that protease inhibition depends entirely on rapid serpin beta-sheet expansion after proteolytic attack. The regulatory advantage afforded by the conformational mobility of serpins is demonstrated here by the structures of native and S195A thrombin-complexed heparin cofactor II (HCII). HCII inhibits thrombin, the final protease of the coagulation cascade, in a glycosaminoglycan-dependent manner that involves the release of a sequestered hirudin-like N-terminal tail for interaction with thrombin. The native structure of HCII resembles that of native antithrombin and suggests an alternative mechanism of allosteric activation, whereas the structure of the S195A thrombin-HCII complex defines the molecular basis of allostery. Together, these structures reveal a multistep allosteric mechanism that relies on sequential contraction and expansion of the central beta-sheet of HCII.
Notes: 1091-6490
Baglin, Trevor P
Carrell, Robin W
Church, Frank C
Esmon, Charles T
Huntington, James A
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2002 Aug 20;99(17):11079-84. doi: 10.1073/pnas.162232399. Epub 2002 Aug 8.
Author Address: Department of Haematology, Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 2XY, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1723
Author: Bai, X. C., Yan, C., Yang, G., Lu, P., Ma, D., Sun, L., Zhou, R., Scheres, S. H. W. and Shi, Y.
Year: 2015
Title: An atomic structure of human γ-secretase
Journal: Nature
Volume: 525
Issue: 7568
Pages: 212-217
Epub Date: 2015/08/19
Date: Sep 10
Short Title: An atomic structure of human γ-secretase
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature14892
PMCID: PMC4568306
NIHMSID: EMS64147
Accession Number: 26280335
Keywords: Alzheimer Disease/genetics
Amyloid Precursor Protein Secretases/*chemistry/genetics/metabolism/*ultrastructure
Binding Sites
*Cryoelectron Microscopy
Humans
Membrane Glycoproteins/*chemistry/metabolism/*ultrastructure
Models, Molecular
Mutation
Presenilin-1/*chemistry/genetics/*ultrastructure
Protein Structure, Tertiary
Protein Subunits/chemistry/genetics/metabolism
Abstract: Dysfunction of the intramembrane protease γ-secretase is thought to cause Alzheimer's disease, with most mutations derived from Alzheimer's disease mapping to the catalytic subunit presenilin 1 (PS1). Here we report an atomic structure of human γ-secretase at 3.4 Å resolution, determined by single-particle cryo-electron microscopy. Mutations derived from Alzheimer's disease affect residues at two hotspots in PS1, each located at the centre of a distinct four transmembrane segment (TM) bundle. TM2 and, to a lesser extent, TM6 exhibit considerable flexibility, yielding a plastic active site and adaptable surrounding elements. The active site of PS1 is accessible from the convex side of the TM horseshoe, suggesting considerable conformational changes in nicastrin extracellular domain after substrate recruitment. Component protein APH-1 serves as a scaffold, anchoring the lone transmembrane helix from nicastrin and supporting the flexible conformation of PS1. Ordered phospholipids stabilize the complex inside the membrane. Our structure serves as a molecular basis for mechanistic understanding of γ-secretase function.
Notes: 1476-4687
Bai, Xiao-Chen
Yan, Chuangye
Yang, Guanghui
Lu, Peilong
Ma, Dan
Sun, Linfeng
Zhou, Rui
Scheres, Sjors H W
Shi, Yigong
MC_UP_A025_1013/Medical Research Council/United Kingdom
MC_UP_A025_101/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Nature. 2015 Sep 10;525(7568):212-217. doi: 10.1038/nature14892. Epub 2015 Aug 17.
Author Address: MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge CB2 0QH, UK.
Ministry of Education Key Laboratory of Protein Science, Tsinghua-Peking Joint Center for Life Sciences, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1780
Author: Baldini, S. F., Wavelet, C., Hainault, I., Guinez, C. and Lefebvre, T.
Year: 2016
Title: The Nutrient-Dependent O-GlcNAc Modification Controls the Expression of Liver Fatty Acid Synthase
Journal: J Mol Biol
Volume: 428
Issue: 16
Pages: 3295-3304
Epub Date: 2016/05/18
Date: Aug 14
Short Title: The Nutrient-Dependent O-GlcNAc Modification Controls the Expression of Liver Fatty Acid Synthase
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2016.04.035
Accession Number: 27185461
Keywords: Animals
Cell Line
Fatty Acid Synthases/*metabolism
Food
Lipogenesis/physiology
Liver/*metabolism
Mice
Mice, Inbred C57BL
N-Acetylglucosaminyltransferases/*metabolism
Obesity/metabolism
Protein Processing, Post-Translational/physiology
*Fas
*O-GlcNAcylation
*lipogenesis
*liver
*ob/ob mice
Abstract: Liver Fatty Acid Synthase (FAS) is pivotal for de novo lipogenesis. Loss of control of this metabolic pathway contributes to the development of liver pathologies ranging from steatosis to nonalcoholic steatohepatitis (NASH) which can lead to cirrhosis and, less frequently, to hepatocellular carcinoma. Therefore, deciphering the molecular mechanisms governing the expression and function of key enzymes such as FAS is crucial. Herein, we link the availability of this lipogenic enzyme to the nutrient-dependent post-translational modification O-GlcNAc that is thought to be deregulated in metabolic diseases (diabetes, obesity, and metabolic syndrome). We demonstrate that expression and activity of liver FAS correlate with O-GlcNAcylation contents in ob/ob mice and in mice fed with a high-carbohydrate diet both in a transcription-dependent and -independent manner. More importantly, inhibiting the removal of O-GlcNAc residues in mice intraperitoneally injected with the selective and potent O-GlcNAcase (OGA) inhibitor Thiamet-G increases FAS expression. FAS and O-GlcNAc transferase (OGT) physically interact, and FAS is O-GlcNAc modified. Treatment of a liver cell line with drugs or nutrients that elevate the O-GlcNAcylation interferes with FAS expression. Inhibition of OGA increases the interaction between FAS and the deubiquitinase Ubiquitin-specific protease-2a (USP2A) in vivo and ex vivo, providing mechanistic insights into the control of FAS expression through O-GlcNAcylation. Together, these results reveal a new type of regulation of FAS, linked to O-GlcNAcylation status, and advance our knowledge on deregulation of lipogenesis in diverse forms of liver diseases.
Notes: 1089-8638
Baldini, Steffi F
Wavelet, Cindy
Hainault, Isabelle
Guinez, Céline
Lefebvre, Tony
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2016 Aug 14;428(16):3295-3304. doi: 10.1016/j.jmb.2016.04.035. Epub 2016 May 13.
Author Address: Univ. Lille, CNRS, UMR 8576, UGSF, Unité de Glycobiologie Structurale et Fonctionnelle, 59000 Lille, France.
Institute of Cardiometabolism and Nutrition, Université Pierre et Marie Curie, INSERM, UMR_S1138, Centre de Recherche des Cordeliers, 75006 Paris, France.
Univ. Lille, EA4489, Environnement Périnatal et Croissance, Equipe dénutritions maternelles périnatales, 59000 Lille, France.
Univ. Lille, CNRS, UMR 8576, UGSF, Unité de Glycobiologie Structurale et Fonctionnelle, 59000 Lille, France. Electronic address: tony.lefebvre@univ-lille1.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 586
Author: Baldwin, E. T., Bhat, T. N., Gulnik, S., Hosur, M. V., Sowder, R. C., 2nd, Cachau, R. E., Collins, J., Silva, A. M. and Erickson, J. W.
Year: 1993
Title: Crystal structures of native and inhibited forms of human cathepsin D: implications for lysosomal targeting and drug design
Journal: Proc Natl Acad Sci U S A
Volume: 90
Issue: 14
Pages: 6796-800
Epub Date: 1993/07/15
Date: Jul 15
Short Title: Crystal structures of native and inhibited forms of human cathepsin D: implications for lysosomal targeting and drug design
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.90.14.6796
PMCID: PMC47019
Accession Number: 8393577
Keywords: Amino Acid Sequence
Aspartic Acid Endopeptidases/chemistry
Biological Transport
Cathepsin D/*antagonists & inhibitors/*chemistry
Drug Design
Glycosylation
Humans
Lysosomes
Models, Molecular
Molecular Sequence Data
Pepstatins/*chemistry
Phosphotransferases
Protein Conformation
Renin/chemistry
X-Ray Diffraction
Abstract: Cathepsin D (EC 3.4.23.5) is a lysosomal protease suspected to play important roles in protein catabolism, antigen processing, degenerative diseases, and breast cancer progression. Determination of the crystal structures of cathepsin D and a complex with pepstatin at 2.5 A resolution provides insights into inhibitor binding and lysosomal targeting for this two-chain, N-glycosylated aspartic protease. Comparison with the structures of a complex of pepstatin bound to rhizopuspepsin and with a human renin-inhibitor complex revealed differences in subsite structures and inhibitor-enzyme interactions that are consistent with affinity differences and structure-activity relationships and suggest strategies for fine-tuning the specificity of cathepsin D inhibitors. Mutagenesis studies have identified a phosphotransferase recognition region that is required for oligosaccharide phosphorylation but is 32 A distant from the N-domain glycosylation site at Asn-70. Electron density for the crystal structure of cathepsin D indicated the presence of an N-linked oligosaccharide that extends from Asn-70 toward Lys-203, which is a key component of the phosphotransferase recognition region, and thus provides a structural explanation for how the phosphotransferase can recognize apparently distant sites on the protein surface.
Notes: 1091-6490
Baldwin, E T
Bhat, T N
Gulnik, S
Hosur, M V
Sowder, R C 2nd
Cachau, R E
Collins, J
Silva, A M
Erickson, J W
N01-CO-74102/CO/NCI NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1993 Jul 15;90(14):6796-800. doi: 10.1073/pnas.90.14.6796.
Author Address: Structural Biochemistry Program, Program Resources Inc./DynCorp, National Cancer Institute-Frederick Cancer Research and Development Center, MD 21702.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 766
Author: Baldwin, T. A., Gogela-Spehar, M. and Ostergaard, H. L.
Year: 2000
Title: Specific isoforms of the resident endoplasmic reticulum protein glucosidase II associate with the CD45 protein-tyrosine phosphatase via a lectin-like interaction
Journal: J Biol Chem
Volume: 275
Issue: 41
Pages: 32071-6
Epub Date: 2000/08/02
Date: Oct 13
Short Title: Specific isoforms of the resident endoplasmic reticulum protein glucosidase II associate with the CD45 protein-tyrosine phosphatase via a lectin-like interaction
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M003088200
Accession Number: 10921916
Keywords: Alternative Splicing/genetics
Animals
Binding, Competitive
Catalysis
Catalytic Domain
Endoplasmic Reticulum/*enzymology/metabolism
Exons/genetics
Glycosylation
Isoenzymes/chemistry/genetics/metabolism
Lectins/chemistry/genetics/*metabolism
Leukocyte Common Antigens/*metabolism
Mannose/pharmacology
Mice
Molecular Weight
Protein Binding/drug effects
Protein Subunits
Transfection
Tumor Cells, Cultured
alpha-Glucosidases/chemistry/genetics/*metabolism
Abstract: We have previously demonstrated that CD45 physically associates with the endoplasmic reticulum processing enzyme glucosidase II (GII). GII consists of the catalytic alpha-chain and an associated beta-chain. To gain insight into the basis of the association between CD45 and GII, we examined the biochemical requirements for the interaction. We show that the alpha-subunit is essential for the interaction. Interestingly, only a higher molecular weight form of GIIalpha is capable of associating with CD45 in a competitive situation where multiple GIIalpha isoforms are expressed. Further, transfection studies demonstrate that only isoforms containing the alternatively spliced sequence Box A1 are capable of binding CD45, although all isoforms are catalytically active. The interaction between CD45 and GII is dependent on the active site of GII, is mediated through the carbohydrate on CD45, and can be inhibited with mannose. Taken together, these results suggest that GIIalpha acts as a lectin and binds to CD45 in an exon-dependent manner. This lectin activity of GII may be a novel mechanism for the regulation of CD45 biology and play a role in immune function, possibly by regulating CD45 glycosylation.
Notes: Baldwin, T A
Gogela-Spehar, M
Ostergaard, H L
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2000 Oct 13;275(41):32071-6. doi: 10.1074/jbc.M003088200.
Author Address: Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, T6G 2S2 Alberta, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1047
Author: Balkhi, M. Y., Trivedi, A. K., Geletu, M., Christopeit, M., Bohlander, S. K., Behre, H. M. and Behre, G.
Year: 2006
Title: Proteomics of acute myeloid leukaemia: Cytogenetic risk groups differ specifically in their proteome, interactome and post-translational protein modifications
Journal: Oncogene
Volume: 25
Issue: 53
Pages: 7041-58
Epub Date: 2006/05/30
Date: Nov 9
Short Title: Proteomics of acute myeloid leukaemia: Cytogenetic risk groups differ specifically in their proteome, interactome and post-translational protein modifications
Alternate Journal: Oncogene
ISSN: 0950-9232 (Print)
0950-9232
DOI: 10.1038/sj.onc.1209689
Accession Number: 16732326
Keywords: Cell Shape
*Cytogenetics
Female
Gene Regulatory Networks
Humans
Leukemia, Myeloid, Acute/*genetics/*metabolism/pathology
Male
Mass Spectrometry
Methylation
*Protein Processing, Post-Translational
Proteome/chemistry/*genetics/*metabolism
Proteomics
Risk Factors
Abstract: Acute myeloid leukaemia (AML) is characterized by specific cytogenetic aberrations that are strong determinants of prognostic outcome and therapeutic response. Because the pathological outcome of AML patients with cytogenetic abnormalities differs considerably, we hypothesized that their proteome may also differ specifically in their expression pattern, protein interaction pathways and post-translational modifications (PTM). We performed this study using 42 AML patients diagnosed for various cytogenetic abnormalities based on two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MS) and MSMS tandem MS. We could identify significant differences in the proteome and PTM of peptides, later confirmed by other methods, between cytogenetic groups. The interactome analysis based on computational bioinformatics reveals major regulating networks: MAPK8 and MYC for complex aberrant karyotype, TP53 for t(8;21), TP53-MYC-PRKAC for 11q23 and JUN and MYC for Inv(16). Further, we analysed 42 MS spectra representative of hnRNPH1, calreticulin and hnRNPA2/B1 in a peak explorer, which reveals a cytogenetic-specific PTM of beta-O-linked N-acetyl glucosamine (O-GlcNAc) of hnRNPH1 in AML patients with 11q23 translocation, an acetylation of calreticulin in t(8;21) translocation and methylation of hnRNPA2/B1 in patients with translocations of t(8;21) and inv(16). This report may lead to a new thinking about AML pathogenesis, as differences at PTM level could be used to distinguish different subtypes of AML.
Notes: Balkhi, M Y
Trivedi, A K
Geletu, M
Christopeit, M
Bohlander, S K
Behre, H M
Behre, G
Journal Article
England
Oncogene. 2006 Nov 9;25(53):7041-58. doi: 10.1038/sj.onc.1209689. Epub 2006 May 29.
Author Address: Bone Marrow Transplantation Section, State Center for Cell and Gene Therapy, Department of Internal Medicine IV-Hematology and Oncology, Martin-Luther-University, Halle, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1426
Author: Bandaranayake, A. D., Correnti, C., Ryu, B. Y., Brault, M., Strong, R. K. and Rawlings, D. J.
Year: 2011
Title: Daedalus: a robust, turnkey platform for rapid production of decigram quantities of active recombinant proteins in human cell lines using novel lentiviral vectors
Journal: Nucleic Acids Res
Volume: 39
Issue: 21
Pages: e143
Epub Date: 2011/09/14
Date: Nov
Short Title: Daedalus: a robust, turnkey platform for rapid production of decigram quantities of active recombinant proteins in human cell lines using novel lentiviral vectors
Alternate Journal: Nucleic acids research
ISSN: 0305-1048 (Print)
0305-1048
DOI: 10.1093/nar/gkr706
PMCID: PMC3241668
Accession Number: 21911364
Keywords: Acute-Phase Proteins/biosynthesis/chemistry/genetics
Animals
Cell Line
Crystallography
Cytokines/genetics/metabolism
Genetic Techniques
Genetic Vectors
Humans
Lentivirus/*genetics
Lipocalin-2
Lipocalins/biosynthesis/chemistry/genetics
Mice
Oncogene Proteins/biosynthesis/chemistry/genetics
Recombinant Proteins/*biosynthesis/chemistry/isolation & purification
Transduction, Genetic
Abstract: A key challenge for the academic and biopharmaceutical communities is the rapid and scalable production of recombinant proteins for supporting downstream applications ranging from therapeutic trials to structural genomics efforts. Here, we describe a novel system for the production of recombinant mammalian proteins, including immune receptors, cytokines and antibodies, in a human cell line culture system, often requiring <3 weeks to achieve stable, high-level expression: Daedalus. The inclusion of minimized ubiquitous chromatin opening elements in the transduction vectors is key for preventing genomic silencing and maintaining the stability of decigram levels of expression. This system can bypass the tedious and time-consuming steps of conventional protein production methods by employing the secretion pathway of serum-free adapted human suspension cell lines, such as 293 Freestyle. Using optimized lentiviral vectors, yields of 20-100 mg/l of correctly folded and post-translationally modified, endotoxin-free protein of up to ~70 kDa in size, can be achieved in conventional, small-scale (100 ml) culture. At these yields, most proteins can be purified using a single size-exclusion chromatography step, immediately appropriate for use in structural, biophysical or therapeutic applications.
Notes: 1362-4962
Bandaranayake, Ashok D
Correnti, Colin
Ryu, Byoung Y
Brault, Michelle
Strong, Roland K
Rawlings, David J
HD037091/HD/NICHD NIH HHS/United States
HD075453/HD/NICHD NIH HHS/United States
AI084803/AI/NIAID NIH HHS/United States
R01 AI084457/AI/NIAID NIH HHS/United States
HL092557/HL/NHLBI NIH HHS/United States
DK073462/DK/NIDDK NIH HHS/United States
DK56465/DK/NIDDK NIH HHS/United States
AI59432/AI/NIAID NIH HHS/United States
AI48675/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nucleic Acids Res. 2011 Nov;39(21):e143. doi: 10.1093/nar/gkr706. Epub 2011 Sep 12.
Author Address: Department of Immunology, University of Washington, Seattle, WA 98195, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1598
Author: Banerjee, S., Sangwan, V., McGinn, O., Chugh, R., Dudeja, V., Vickers, S. M. and Saluja, A. K.
Year: 2013
Title: Triptolide-induced cell death in pancreatic cancer is mediated by O-GlcNAc modification of transcription factor Sp1
Journal: J Biol Chem
Volume: 288
Issue: 47
Pages: 33927-33938
Epub Date: 2013/10/17
Date: Nov 22
Short Title: Triptolide-induced cell death in pancreatic cancer is mediated by O-GlcNAc modification of transcription factor Sp1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.500983
PMCID: PMC3837133
Accession Number: 24129563
Keywords: Acetylglucosamine/genetics/*metabolism
Active Transport, Cell Nucleus/drug effects/genetics
Antineoplastic Agents, Alkylating/*pharmacology
Cell Death/drug effects/genetics
Cell Line, Tumor
Cell Nucleus/genetics/*metabolism/pathology
Cell Survival/drug effects/genetics
DNA-Binding Proteins/genetics/metabolism
Diterpenes/*pharmacology
Down-Regulation/drug effects/genetics
Epoxy Compounds/pharmacology
Gene Expression Regulation, Neoplastic/drug effects/genetics
Glycosylation/drug effects
HSP70 Heat-Shock Proteins/biosynthesis/genetics
Heat Shock Transcription Factors
Humans
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/metabolism
NF-kappa B/genetics/metabolism
Neoplasm Proteins/genetics/*metabolism
Pancreatic Neoplasms/*drug therapy/genetics/metabolism/pathology
Phenanthrenes/*pharmacology
Signal Transduction/drug effects/genetics
Sp1 Transcription Factor/genetics/*metabolism
Transcription Factors/genetics/metabolism
Heat Shock Protein
Minnelide
O-GlcNAc
Pancreas
Pancreatic Cancer
Sp1
Triptolide
Abstract: Pancreatic cancer, the fourth most prevalent cancer-related cause of death in the United States, is a disease with a dismal survival rate of 5% 5 years after diagnosis. One of the survival proteins responsible for its extraordinary ability to evade cell death is HSP70. A naturally derived compound, triptolide, and its water-soluble prodrug, Minnelide, down-regulate the expression of this protein in pancreatic cancer cells, thereby causing cell death. However, the mechanism of action of triptolide has not been elucidated. Our study shows that triptolide-induced down-regulation of HSP70 expression is associated with a decrease in glycosylation of the transcription factor Sp1. We further show that triptolide inhibits glycosylation of Sp1, inhibiting the hexosamine biosynthesis pathway, particularly the enzyme O-GlcNAc transferase. Inhibition of O-GlcNAc transferase prevents nuclear localization of Sp1 and affects its DNA binding activity. This in turn down-regulates prosurvival pathways like NF-κB, leading to inhibition of HSF1 and HSP70 and eventually to cell death. In this study, we evaluated the mechanism by which triptolide affects glycosylation of Sp1, which in turn affects downstream pathways controlling survival of pancreatic cancer cells.
Notes: 1083-351x
Banerjee, Sulagna
Sangwan, Veena
McGinn, Olivia
Chugh, Rohit
Dudeja, Vikas
Vickers, Selwyn M
Saluja, Ashok K
T32CA132715/CA/NCI NIH HHS/United States
CA124723/CA/NCI NIH HHS/United States
R01 CA170946/CA/NCI NIH HHS/United States
P50 CA101955/CA/NCI NIH HHS/United States
R01 CA124723/CA/NCI NIH HHS/United States
T32 CA132715/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2013 Nov 22;288(47):33927-33938. doi: 10.1074/jbc.M113.500983. Epub 2013 Oct 15.
Author Address: Division of Basic and Translational Research, Department of Surgery, University of Minnesota, Minneapolis, Minnesota 55455.
Division of Basic and Translational Research, Department of Surgery, University of Minnesota, Minneapolis, Minnesota 55455; Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455.
Division of Basic and Translational Research, Department of Surgery, University of Minnesota, Minneapolis, Minnesota 55455; Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455. Electronic address: asaluja@umn.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 276
Author: Bank, R. A., Hettema, E. H., Arwert, F., Amerongen, A. V. and Pronk, J. C.
Year: 1991
Title: Electrophoretic characterization of posttranslational modifications of human parotid salivary alpha-amylase
Journal: Electrophoresis
Volume: 12
Issue: 1
Pages: 74-9
Epub Date: 1991/01/01
Date: Jan
Short Title: Electrophoretic characterization of posttranslational modifications of human parotid salivary alpha-amylase
Alternate Journal: Electrophoresis
ISSN: 0173-0835 (Print)
0173-0835
DOI: 10.1002/elps.1150120114
Accession Number: 1710976
Keywords: Amino Acid Sequence
Densitometry
Humans
Isoenzymes/chemistry/genetics
Lasers
Molecular Sequence Data
Parotid Gland/*enzymology
*Protein Processing, Post-Translational
Saliva/*enzymology
Staining and Labeling
alpha-Amylases/*chemistry/genetics
Abstract: Human salivary alpha-amylase displays multiple bands upon native polyacrylamide gel electrophoresis. In fresh saliva, due to posttranslational modifications, a pattern of 5-6 isozymes is observed. The isozymes are designated 1-6, in the order of increasing anodal mobility. As a result of the development of a rapid and sensitive electrophoresis system, with markedly higher resolution than previously reported, we concluded that a previously proposed model (Karn et al., Biochem. Genet. 1973, 10, 341-350) is inadequate to explain the origin of the various bands. We propose an alternative model that fits in with our new and previously made observations. According to this model, band 2 is the primary gene product and band 1 is its glycosylated counterpart--with only one neutral oligosaccharide present on each molecule. Band 3 originates from band 1 by the transialidase-catalyzed incorporation of sialic acid into the biantennary chain. Bands 4 and 6 originate from bands 2 and 4, respectively, by deamidation; band 5 is the deamidation product of amylase with an acidic oligosaccharide (band 3). Only a minor part of band 3 consists of the deamidation product of band 1. Peptide Asn-Gly-Ser (residues 427-429) is the most probable candidate for glycosylation; literature data suggests that deamidation occurs in the stretch Glu-Asn-Gly-Lys-Asp (residues 364-368) and Asn-Gly-Asn-Cys (residues 474-477). Both glycosylation and deamidation might play a role in the clearance of amylase from the systemic circulation. The electrophoresis system described is a powerful tool to determine amylase isozyme distributions in health and disease, especially for the screening of alterations seen in ectopically produced amylase.
Notes: Bank, R A
Hettema, E H
Arwert, F
Amerongen, A V
Pronk, J C
Journal Article
Germany
Electrophoresis. 1991 Jan;12(1):74-9. doi: 10.1002/elps.1150120114.
Author Address: Institute of Human Genetics, Faculty of Medicine, Vrije Universiteit, Amsterdam, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 827
Author: Bannert, N., Craig, S., Farzan, M., Sogah, D., Santo, N. V., Choe, H. and Sodroski, J.
Year: 2001
Title: Sialylated O-glycans and sulfated tyrosines in the NH2-terminal domain of CC chemokine receptor 5 contribute to high affinity binding of chemokines
Journal: J Exp Med
Volume: 194
Issue: 11
Pages: 1661-73
Epub Date: 2001/12/26
Date: Dec 3
Short Title: Sialylated O-glycans and sulfated tyrosines in the NH2-terminal domain of CC chemokine receptor 5 contribute to high affinity binding of chemokines
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.194.11.1661
PMCID: PMC2193526
Accession Number: 11733580
Keywords: Amino Acid Sequence
Animals
Binding Sites
CHO Cells
Cell Line
Cell Line, Transformed
Cells, Cultured
Chemokine CCL4
Cricetinae
Dogs
Gene Expression
Glycosylation
HIV-1/metabolism/physiology
HeLa Cells
Humans
Macrophage Inflammatory Proteins/*metabolism
Macrophages
Molecular Sequence Data
N-Acetylneuraminic Acid/*metabolism
Polysaccharides/*metabolism
Protein Binding
Receptors, CCR5/genetics/*metabolism
Simian Immunodeficiency Virus/metabolism/physiology
Sulfates/*metabolism
Tyrosine/*metabolism
Abstract: The chemokine receptor CCR5 plays an important role in leukocyte chemotaxis and activation, and also acts as a coreceptor for human and simian immunodeficiency viruses (HIV-1, HIV-2, and SIV). We provide evidence that CCR5 is O-glycosylated on serine 6 in the NH2 terminus. The O-linked glycans, particularly sialic acid moieties, significantly contribute to binding of the chemokine ligands. By contrast, removal of O-linked oligosaccharide exerted little effect on HIV-1 infection. Sulfation of specific tyrosine residues in the CCR5 NH2 terminus was important for efficient beta-chemokine binding. Thus, as has been observed for the binding of selectins and their ligands, O-linked carbohydrates and tyrosine sulfates play major roles in promoting the interaction of chemokines with CCR5. The resulting flexible arrays of negative charges on the CCR5 surface may allow specific, high-affinity interactions with diverse chemokine ligands. Although this is the first example of O-linked oligosaccharides and tyrosine sulfates playing a role in chemokine binding, the high density of serines, threonines and tyrosines in the N-termini of many CC chemokine receptors suggests that these posttranslational modifications may commonly contribute to chemokine binding.
Notes: 1540-9538
Bannert, N
Craig, S
Farzan, M
Sogah, D
Santo, N V
Choe, H
Sodroski, J
R01 AI043891/AI/NIAID NIH HHS/United States
AI28691/AI/NIAID NIH HHS/United States
AI41851/AI/NIAID NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
J Exp Med. 2001 Dec 3;194(11):1661-73. doi: 10.1084/jem.194.11.1661.
Author Address: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1213
Author: Barinka, C., Byun, Y., Dusich, C. L., Banerjee, S. R., Chen, Y., Castanares, M., Kozikowski, A. P., Mease, R. C., Pomper, M. G. and Lubkowski, J.
Year: 2008
Title: Interactions between human glutamate carboxypeptidase II and urea-based inhibitors: structural characterization
Journal: J Med Chem
Volume: 51
Issue: 24
Pages: 7737-43
Epub Date: 2008/12/05
Date: Dec 25
Short Title: Interactions between human glutamate carboxypeptidase II and urea-based inhibitors: structural characterization
Alternate Journal: Journal of medicinal chemistry
ISSN: 0022-2623 (Print)
0022-2623
DOI: 10.1021/jm800765e
PMCID: PMC5516903
NIHMSID: NIHMS875185
Accession Number: 19053759
Keywords: Binding Sites
Chemistry, Pharmaceutical/methods
Drug Design
Glutamate Carboxypeptidase II/*chemistry
Glutamic Acid/chemistry
Humans
Ions
Ligands
Models, Molecular
Molecular Conformation
Protein Conformation
Protein Structure, Tertiary
Tyrosine/chemistry
Urea/chemistry/*pharmacology
Zinc/*chemistry
Abstract: Urea-based, low molecular weight ligands of glutamate carboxypeptidase II (GCPII) have demonstrated efficacy in various models of neurological disorders and can serve as imaging agents for prostate cancer. To enhance further development of such compounds, we determined X-ray structures of four complexes between human GCPII and urea-based inhibitors at high resolution. All ligands demonstrate an invariant glutarate moiety within the S1' pocket of the enzyme. The ureido linkage between P1 and P1' inhibitor sites interacts with the active-site Zn(1)(2+) ion and the side chains of Tyr552 and His553. Interactions within the S1 pocket are defined primarily by a network of hydrogen bonds between the P1 carboxylate group of the inhibitors and the side chains of Arg534, Arg536, and Asn519. Importantly, we have identified a hydrophobic pocket accessory to the S1 site that can be exploited for structure-based design of novel GCPII inhibitors with increased lipophilicity.
Notes: 1520-4804
Barinka, Cyril
Byun, Youngjoo
Dusich, Crystal L
Banerjee, Sangeeta R
Chen, Ying
Castanares, Mark
Kozikowski, Alan P
Mease, Ronnie C
Pomper, Martin G
Lubkowski, Jacek
CA1114111/CA/NCI NIH HHS/United States
CA111982/CA/NCI NIH HHS/United States
R21 MH080580-02/MH/NIMH NIH HHS/United States
R21 MH080580/MH/NIMH NIH HHS/United States
Intramural NIH HHS/United States
R21 CA111982-02/CA/NCI NIH HHS/United States
MH080580/MH/NIMH NIH HHS/United States
R24 CA092871/CA/NCI NIH HHS/United States
R21 CA114111-02/CA/NCI NIH HHS/United States
R21 CA111982/CA/NCI NIH HHS/United States
EB005423/EB/NIBIB NIH HHS/United States
R33 MH080580/MH/NIMH NIH HHS/United States
CA92871/CA/NCI NIH HHS/United States
R21 EB005324-02/EB/NIBIB NIH HHS/United States
U24 CA092871/CA/NCI NIH HHS/United States
R21 EB005324/EB/NIBIB NIH HHS/United States
R21 CA114111/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, U.S. Gov't, Non-P.H.S.
J Med Chem. 2008 Dec 25;51(24):7737-43. doi: 10.1021/jm800765e.
Author Address: Center for Cancer Research, National Cancer Institute at Frederick, Frederick, Maryland 21702, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1156
Author: Barinka, C., Hlouchova, K., Rovenska, M., Majer, P., Dauter, M., Hin, N., Ko, Y. S., Tsukamoto, T., Slusher, B. S., Konvalinka, J. and Lubkowski, J.
Year: 2008
Title: Structural basis of interactions between human glutamate carboxypeptidase II and its substrate analogs
Journal: J Mol Biol
Volume: 376
Issue: 5
Pages: 1438-50
Epub Date: 2008/02/01
Date: Mar 7
Short Title: Structural basis of interactions between human glutamate carboxypeptidase II and its substrate analogs
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2007.12.066
PMCID: PMC2753231
NIHMSID: NIHMS42351
Accession Number: 18234225
Keywords: Antigens, Surface/*chemistry/metabolism
Arginine/metabolism
Binding Sites
Crystallography, X-Ray
Glutamate Carboxypeptidase II/antagonists & inhibitors/*chemistry/metabolism
Humans
Models, Molecular
Recombinant Proteins/chemistry/metabolism
Substrate Specificity
Zinc/chemistry
Abstract: Human glutamate carboxypeptidase II (GCPII) is involved in neuronal signal transduction and intestinal folate absorption by means of the hydrolysis of its two natural substrates, N-acetyl-aspartyl-glutamate and folyl-poly-gamma-glutamates, respectively. During the past years, tremendous efforts have been made toward the structural analysis of GCPII. Crystal structures of GCPII in complex with various ligands have provided insight into the binding of these ligands, particularly to the S1' site of the enzyme. In this article, we have extended structural characterization of GCPII to its S1 site by using dipeptide-based inhibitors that interact with both S1 and S1' sites of the enzyme. To this end, we have determined crystal structures of human GCPII in complex with phosphapeptide analogs of folyl-gamma-glutamate, aspartyl-glutamate, and gamma-glutamyl-glutamate, refined at 1.50, 1.60, and 1.67 A resolution, respectively. The S1 pocket of GCPII could be accurately defined and analyzed for the first time, and the data indicate the importance of Asn519, Arg463, Arg534, and Arg536 for recognition of the penultimate (i.e., P1) substrate residues. Direct interactions between the positively charged guanidinium groups of Arg534 and Arg536 and a P1 moiety of a substrate/inhibitor provide mechanistic explanation of GCPII preference for acidic dipeptides. Additionally, observed conformational flexibility of the Arg463 and Arg536 side chains likely regulates GCPII affinity toward different inhibitors and modulates GCPII substrate specificity. The biochemical experiments assessing the hydrolysis of several GCPII substrate derivatives modified at the P1 position, also included in this report, further complement and extend conclusions derived from the structural analysis. The data described here form an a solid foundation for the structurally aided design of novel low-molecular-weight GCPII inhibitors and imaging agents.
Notes: 1089-8638
Barinka, Cyril
Hlouchova, Klara
Rovenska, Miroslava
Majer, Pavel
Dauter, Miroslawa
Hin, Niyada
Ko, Yao-Sen
Tsukamoto, Takashi
Slusher, Barbara S
Konvalinka, Jan
Lubkowski, Jacek
NIH0012079625/Intramural NIH HHS/United States
Z01 BC010760-01/Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Mol Biol. 2008 Mar 7;376(5):1438-50. doi: 10.1016/j.jmb.2007.12.066. Epub 2008 Jan 5.
Author Address: Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA. cyril@ncifcrf.gov
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1113
Author: Barinka, C., Rovenská, M., Mlcochová, P., Hlouchová, K., Plechanovová, A., Majer, P., Tsukamoto, T., Slusher, B. S., Konvalinka, J. and Lubkowski, J.
Year: 2007
Title: Structural insight into the pharmacophore pocket of human glutamate carboxypeptidase II
Journal: J Med Chem
Volume: 50
Issue: 14
Pages: 3267-73
Epub Date: 2007/06/15
Date: Jul 12
Short Title: Structural insight into the pharmacophore pocket of human glutamate carboxypeptidase II
Alternate Journal: Journal of medicinal chemistry
ISSN: 0022-2623 (Print)
0022-2623
DOI: 10.1021/jm070133w
Accession Number: 17567119
Keywords: Crystallography, X-Ray
Glutamate Carboxypeptidase II/*chemistry
Humans
Models, Molecular
Molecular Mimicry
Protein Conformation
Recombinant Proteins/chemistry
Abstract: Inhibition of glutamate carboxypeptidase II (GCPII) has been shown to be neuroprotective in multiple preclinical models in which dysregulated glutamatergic transmission is implicated. Herein, we report crystal structures of the human GCPII complexed with three glutamate mimetics/derivatives, 2-(phosphonomethyl)pentanedioic acid (2-PMPA), quisqualic acid (QA), and L-serine O-sulfate (L-SOS), at 1.72, 1.62, and 2.10 A resolution, respectively. Despite the structural differences between the distal parts of the inhibitors, all three compounds share similar binding modes in the pharmacophore (i.e., S1') pocket of GCPII, where they are stabilized by a combination of polar and van der Waals interactions. The structural diversity of the distal parts of the inhibitors leads to rearrangements of the S1' site that are necessary for efficient interactions between the enzyme and an inhibitor. The set of structures presented here, in conjunction with the available biochemical data, illustrates a flexibility of the GCPII pharmacophore pocket and highlights the structural features required for potent GCPII inhibition. These findings could facilitate the rational structure-based drug design of new GCPII inhibitors in the future.
Notes: Barinka, Cyril
Rovenská, Miroslava
Mlcochová, Petra
Hlouchová, Klára
Plechanovová, Anna
Majer, Pavel
Tsukamoto, Takashi
Slusher, Barbara S
Konvalinka, Jan
Lubkowski, Jacek
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Med Chem. 2007 Jul 12;50(14):3267-73. doi: 10.1021/jm070133w. Epub 2007 Jun 14.
Author Address: Center for Cancer Research, National Cancer Institute at Frederick, Frederick, Maryland 21702, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 939
Author: Barinka, C., Sácha, P., Sklenár, J., Man, P., Bezouska, K., Slusher, B. S. and Konvalinka, J.
Year: 2004
Title: Identification of the N-glycosylation sites on glutamate carboxypeptidase II necessary for proteolytic activity
Journal: Protein Sci
Volume: 13
Issue: 6
Pages: 1627-35
Epub Date: 2004/05/21
Date: Jun
Short Title: Identification of the N-glycosylation sites on glutamate carboxypeptidase II necessary for proteolytic activity
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.04622104
PMCID: PMC2279971
Accession Number: 15152093
Keywords: Animals
Antigens, Surface/*chemistry/genetics/*metabolism
Binding Sites
Catalysis
Cell Line
Gene Expression
Glutamate Carboxypeptidase II/*chemistry/genetics/*metabolism
Glycosylation
Humans
Hydrolysis
Insecta
Kinetics
Mutagenesis, Site-Directed/genetics
Abstract: Glutamate carboxypeptidase II (GCPII) is a membrane peptidase expressed in the prostate, central and peripheral nervous system, kidney, small intestine, and tumor-associated neovasculature. The GCPII form expressed in the central nervous system, termed NAALADase, is responsible for the cleavage of N-acetyl-L-aspartyl-L-glutamate (NAAG) yielding free glutamate in the synaptic cleft, and is implicated in various pathologic conditions associated with glutamate excitotoxicity. The prostate form of GCPII, termed prostate-specific membrane antigen (PSMA), is up-regulated in cancer and used as an effective prostate cancer marker. Little is known about the structure of this important pharmaceutical target. As a type II membrane protein, GCPII is heavily glycosylated. In this paper we show that N-glycosylation is vital for proper folding and subsequent secretion of human GCPII. Analysis of the predicted N-glycosylation sites also provides evidence that these sites are critical for GCPII carboxypeptidase activity. We confirm that all predicted N-glycosylation sites are occupied by an oligosaccharide moiety and show that glycosylation at sites distant from the putative catalytic domain is critical for the NAAG-hydrolyzing activity of GCPII calling the validity of previously described structural models of GCPII into question.
Notes: 1469-896x
Barinka, Cyril
Sácha, Pavel
Sklenár, Jan
Man, Petr
Bezouska, Karel
Slusher, Barbara S
Konvalinka, Jan
Journal Article
Research Support, Non-U.S. Gov't
Protein Sci. 2004 Jun;13(6):1627-35. doi: 10.1110/ps.04622104.
Author Address: Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Praha 6, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2049
Author: Barnes, J. W., Tian, L., Krick, S., Helton, E. S., Denson, R. S., Comhair, S. A. A. and Dweik, R. A.
Year: 2019
Title: O-GlcNAc Transferase Regulates Angiogenesis in Idiopathic Pulmonary Arterial Hypertension
Journal: Int J Mol Sci
Volume: 20
Issue: 24
Epub Date: 2019/12/19
Date: Dec 13
Short Title: O-GlcNAc Transferase Regulates Angiogenesis in Idiopathic Pulmonary Arterial Hypertension
Alternate Journal: International journal of molecular sciences
ISSN: 1422-0067
DOI: 10.3390/ijms20246299
PMCID: PMC6941156
Accession Number: 31847126
Keywords: Adult
Animals
Coculture Techniques
Enzyme Inhibitors/pharmacology
Familial Primary Pulmonary Hypertension/*enzymology/pathology
Female
Gene Expression Regulation/drug effects
Humans
Male
Mice
Mice, Inbred NOD
Mice, SCID
Middle Aged
N-Acetylglucosaminyltransferases/antagonists & inhibitors/*metabolism
Neovascularization, Pathologic/*enzymology/pathology
Vascular Endothelial Growth Factor A/biosynthesis
Ipah
O-GlcNAc
Ogt
Vegf
angiogenesis
Abstract: Idiopathic pulmonary arterial hypertension (IPAH) is considered a vasculopathy characterized by elevated pulmonary vascular resistance due to vasoconstriction and/or lung remodeling such as plexiform lesions, the hallmark of the PAH, as well as cell proliferation and vascular and angiogenic dysfunction. The serine/threonine hydroxyl-linked N-Acetylglucosamine (O-GlcNAc) transferase (OGT) has been shown to drive pulmonary arterial smooth muscle cell (PASMC) proliferation in IPAH. OGT is a cellular nutrient sensor that is essential in maintaining proper cell function through the regulation of cell signaling, proliferation, and metabolism. The aim of this study was to determine the role of OGT and O-GlcNAc in vascular and angiogenic dysfunction in IPAH. Primary isolated human control and IPAH patient PASMCs and pulmonary arterial endothelial cells (PAECs) were grown in the presence or absence of OGT inhibitors and subjected to biochemical assessments in monolayer cultures and tube formation assays, in vitro vascular sprouting 3D spheroid co-culture models, and de novo vascularization models in NODSCID mice. We showed that knockdown of OGT resulted in reduced vascular endothelial growth factor (VEGF) expression in IPAH primary isolated vascular cells. In addition, specificity protein 1 (SP1), a known stimulator of VEGF expression, was shown to have higher O-GlcNAc levels in IPAH compared to control at physiological (5 mM) and high (25 mM) glucose concentrations, and knockdown resulted in decreased VEGF protein levels. Furthermore, human IPAH PAECs demonstrated a significantly higher degree of capillary tube-like structures and increased length compared to control PAECs. Addition of an OGT inhibitor, OSMI-1, significantly reduced the number of tube-like structures and tube length similar to control levels. Assessment of vascular sprouting from an in vitro 3D spheroid co-culture model using IPAH and control PAEC/PASMCs and an in vivo vascularization model using control and PAEC-embedded collagen implants demonstrated higher vascularization in IPAH compared to control. Blocking OGT activity in these experiments, however, altered the vascular sprouting and de novo vascularization in IPAH similar to control levels when compared to controls. Our findings in this report are the first to describe a role for the OGT/O-GlcNAc axis in modulating VEGF expression and vascularization in IPAH. These findings provide greater insight into the potential role that altered glucose uptake and metabolism may have on the angiogenic process and the development of plexiform lesions. Therefore, we believe that the OGT/O-GlcNAc axis may be a potential therapeutic target for treating the angiogenic dysregulation that is present in IPAH.
Notes: 1422-0067
Barnes, Jarrod W
Tian, Liping
Krick, Stefanie
Helton, E Scott
Denson, Rebecca S
Comhair, Suzy A A
Dweik, Raed A
R00 HL131866/HL/NHLBI NIH HHS/United States
R01 HL130209/HL/NHLBI NIH HHS/United States
R00HL131866/HL/NHLBI NIH HHS/United States
R01HL130209/HL/NHLBI NIH HHS/United States
Journal Article
Int J Mol Sci. 2019 Dec 13;20(24):6299. doi: 10.3390/ijms20246299.
Author Address: Division of Pulmonary, Allergy and Critical Care Medicine, Department of Medicine, The University of Alabama at Birmingham, THT 422, 1720 2nd Ave S, Birmingham, AL 35294-0006, USA.
Department of Inflammation & Immunity, Cleveland Clinic, 9500 Euclid Ave, Cleveland, OH 44195, USA.
Respiratory Institute, Cleveland Clinic, 9500 Euclid Ave, Cleveland, OH 44195, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1062
Author: Barnes, K., Dobrzynski, H., Foppolo, S., Beal, P. R., Ismat, F., Scullion, E. R., Sun, L., Tellez, J., Ritzel, M. W., Claycomb, W. C., Cass, C. E., Young, J. D., Billeter-Clark, R., Boyett, M. R. and Baldwin, S. A.
Year: 2006
Title: Distribution and functional characterization of equilibrative nucleoside transporter-4, a novel cardiac adenosine transporter activated at acidic pH
Journal: Circ Res
Volume: 99
Issue: 5
Pages: 510-9
Epub Date: 2006/07/29
Date: Sep 1
Short Title: Distribution and functional characterization of equilibrative nucleoside transporter-4, a novel cardiac adenosine transporter activated at acidic pH
Alternate Journal: Circulation research
ISSN: 0009-7330
DOI: 10.1161/01.res.0000238359.18495.42
Accession Number: 16873718
Keywords: Acids/*metabolism
Adenosine/metabolism
Animals
Biological Transport/drug effects
Cells, Cultured
Equilibrative Nucleoside Transport Proteins
Glycosylation
Humans
Hydrogen-Ion Concentration
Kinetics
Membrane Transport Proteins/*metabolism
Mice
Myocardium/*metabolism
Nerve Tissue Proteins/antagonists & inhibitors/*metabolism
Nucleosides/metabolism
Oocytes
Serotonin/metabolism
Subcellular Fractions/metabolism
Tissue Distribution
Xenopus
Abstract: Adenosine plays multiple roles in the efficient functioning of the heart by regulating coronary blood flow, cardiac pacemaking, and contractility. Previous studies have implicated the equilibrative nucleoside transporter family member equilibrative nucleoside transporter-1 (ENT1) in the regulation of cardiac adenosine levels. We report here that a second member of this family, ENT4, is also abundant in the heart, in particular in the plasma membranes of ventricular myocytes and vascular endothelial cells but, unlike ENT1, is virtually absent from the sinoatrial and atrioventricular nodes. Originally described as a monoamine/organic cation transporter, we found that both human and mouse ENT4 exhibited a novel, pH-dependent adenosine transport activity optimal at acidic pH (apparent K(m) values 0.78 and 0.13 mmol/L, respectively, at pH 5.5) and absent at pH 7.4. In contrast, serotonin transport by ENT4 was relatively insensitive to pH. ENT4-mediated nucleoside transport was adenosine selective, sodium independent and only weakly inhibited by the classical inhibitors of equilibrative nucleoside transport, dipyridamole, dilazep, and nitrobenzylthioinosine. We hypothesize that ENT4, in addition to playing roles in cardiac serotonin transport, contributes to the regulation of extracellular adenosine concentrations, in particular under the acidotic conditions associated with ischemia.
Notes: 1524-4571
Barnes, Kay
Dobrzynski, Halina
Foppolo, Sophie
Beal, Paul R
Ismat, Fouzia
Scullion, Elspeth R
Sun, Lijie
Tellez, James
Ritzel, Mabel W L
Claycomb, William C
Cass, Carol E
Young, James D
Billeter-Clark, Rudi
Boyett, Mark R
Baldwin, Stephen A
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Circ Res. 2006 Sep 1;99(5):510-9. doi: 10.1161/01.RES.0000238359.18495.42. Epub 2006 Jul 27.
Author Address: Institute of Membrane and Systems Biology, University of Leeds, UK. K.Barnes@leeds.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 891
Author: Barton, W. A., Liu, B. P., Tzvetkova, D., Jeffrey, P. D., Fournier, A. E., Sah, D., Cate, R., Strittmatter, S. M. and Nikolov, D. B.
Year: 2003
Title: Structure and axon outgrowth inhibitor binding of the Nogo-66 receptor and related proteins
Journal: Embo j
Volume: 22
Issue: 13
Pages: 3291-302
Epub Date: 2003/07/04
Date: Jul 1
Short Title: Structure and axon outgrowth inhibitor binding of the Nogo-66 receptor and related proteins
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/cdg325
PMCID: PMC165649
Accession Number: 12839991
Keywords: Amino Acid Sequence
Animals
*Axons
Crystallization
GPI-Linked Proteins
Mice
Models, Molecular
Molecular Sequence Data
Myelin Proteins/chemistry/*metabolism
Nogo Receptor 1
Protein Binding
Protein Conformation
Receptors, Cell Surface/chemistry/*metabolism
Sequence Homology, Amino Acid
Abstract: The myelin-derived proteins Nogo, MAG and OMgp limit axonal regeneration after injury of the spinal cord and brain. These cell-surface proteins signal through multi-subunit neuronal receptors that contain a common ligand-binding glycosylphosphatidylinositol-anchored subunit termed the Nogo-66 receptor (NgR). By deletion analysis, we show that the binding of soluble fragments of Nogo, MAG and NgR to cell-surface NgR requires the entire leucine-rich repeat (LRR) region of NgR, but not other portions of the protein. Despite sharing extensive sequence similarity with NgR, two related proteins, NgR2 and NgR3, which we have identified, do not bind Nogo, MAG, OMgp or NgR. To investigate NgR specificity and multi-ligand binding, we determined the crystal structure of the biologically active ligand-binding soluble ectodomain of NgR. The molecule is banana shaped with elongation and curvature arising from eight LRRs flanked by an N-terminal cap and a small C-terminal subdomain. The NgR structure analysis, as well as a comparison of NgR surface residues not conserved in NgR2 and NgR3, identifies potential protein interaction sites important in the assembly of a functional signaling complex.
Notes: 1460-2075
Barton, William A
Liu, Betty P
Tzvetkova, Dorothea
Jeffrey, Philip D
Fournier, Alyson E
Sah, Dinah
Cate, Richard
Strittmatter, Stephen M
Nikolov, Dimitar B
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
EMBO J. 2003 Jul 1;22(13):3291-302. doi: 10.1093/emboj/cdg325.
Author Address: Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1046
Author: Barton, W. A., Tzvetkova-Robev, D., Miranda, E. P., Kolev, M. V., Rajashankar, K. R., Himanen, J. P. and Nikolov, D. B.
Year: 2006
Title: Crystal structures of the Tie2 receptor ectodomain and the angiopoietin-2-Tie2 complex
Journal: Nat Struct Mol Biol
Volume: 13
Issue: 6
Pages: 524-32
Epub Date: 2006/05/30
Date: Jun
Short Title: Crystal structures of the Tie2 receptor ectodomain and the angiopoietin-2-Tie2 complex
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb1101
Accession Number: 16732286
Keywords: Amino Acid Sequence
Angiopoietin-2/*chemistry
Calcium/chemistry/metabolism
Chromatography, Gel
Crystallography, X-Ray
Epidermal Growth Factor/chemistry
Fibrinogen/chemistry
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Conformation
Receptor, TIE-2/*chemistry/metabolism
Sequence Homology, Amino Acid
Abstract: The Tie receptor tyrosine kinases and their angiopoietin (Ang) ligands play central roles in developmental and tumor-induced angiogenesis. Here we present the crystal structures of the Tie2 ligand-binding region alone and in complex with Ang2. In contrast to prediction, Tie2 contains not two but three immunoglobulin (Ig) domains, which fold together with the three epidermal growth factor domains into a compact, arrowhead-shaped structure. Ang2 binds at the tip of the arrowhead utilizing a lock-and-key mode of ligand recognition-unique for a receptor kinase-where two complementary surfaces interact with each other with no domain rearrangements and little conformational change in either molecule. Ang2-Tie2 recognition is similar to antibody-protein antigen recognition, including the location of the ligand-binding site within the Ig fold. Analysis of the structures and structure-based mutagenesis provide insight into the mechanism of receptor activation and support the hypothesis that all angiopoietins interact with Tie2 in a structurally similar manner.
Notes: Barton, William A
Tzvetkova-Robev, Dorothea
Miranda, Edward P
Kolev, Momchil V
Rajashankar, Kanagalaghatta R
Himanen, Juha P
Nikolov, Dimitar B
R01-HL077249/HL/NHLBI NIH HHS/United States
RR-15301/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
United States
Nat Struct Mol Biol. 2006 Jun;13(6):524-32. doi: 10.1038/nsmb1101. Epub 2006 May 28.
Author Address: Structural Biology Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, New York 10021, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1404
Author: Bas, T., Gao, G. Y., Lvov, A., Chandrasekhar, K. D., Gilmore, R. and Kobertz, W. R.
Year: 2011
Title: Post-translational N-glycosylation of type I transmembrane KCNE1 peptides: implications for membrane protein biogenesis and disease
Journal: J Biol Chem
Volume: 286
Issue: 32
Pages: 28150-9
Epub Date: 2011/06/17
Date: Aug 12
Short Title: Post-translational N-glycosylation of type I transmembrane KCNE1 peptides: implications for membrane protein biogenesis and disease
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.235168
PMCID: PMC3151060
Accession Number: 21676880
Keywords: Animals
CHO Cells
Cricetinae
Cricetulus
Glycosylation
HEK293 Cells
HeLa Cells
Humans
Long QT Syndrome/genetics/metabolism
Peptides/genetics/metabolism
Point Mutation
Potassium Channels, Voltage-Gated/genetics/*metabolism
Protein Processing, Post-Translational/*physiology
Abstract: N-Glycosylation of membrane proteins is critical for their proper folding, co-assembly and subsequent matriculation through the secretory pathway. Here, we examine the kinetics of N-glycan addition to type I transmembrane KCNE1 K(+) channel β-subunits, where point mutations that prevent N-glycosylation at one consensus site give rise to disorders of the cardiac rhythm and congenital deafness. We show that KCNE1 has two distinct N-glycosylation sites: a typical co-translational site and a consensus site ∼20 residues away that unexpectedly acquires N-glycans after protein synthesis (post-translational). Mutations that ablate the co-translational site concomitantly reduce glycosylation at the post-translational site, resulting in unglycosylated KCNE1 subunits that cannot reach the cell surface with their cognate K(+) channel. This long range inhibition is highly specific for post-translational N-glycosylation because mutagenic conversion of the KCNE1 post-translational site into a co-translational site restored both monoglycosylation and anterograde trafficking. These results directly explain how a single point mutation can prevent N-glycan attachment at multiple sites, providing a new biogenic mechanism for human disease.
Notes: 1083-351x
Bas, Tuba
Gao, Grace Y
Lvov, Anatoli
Chandrasekhar, Kshama D
Gilmore, Reid
Kobertz, William R
R01 GM043768/GM/NIGMS NIH HHS/United States
R01 DC007669/DC/NIDCD NIH HHS/United States
DC-007669/DC/NIDCD NIH HHS/United States
R01 DC007669-05/DC/NIDCD NIH HHS/United States
R01 GM070650/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2011 Aug 12;286(32):28150-9. doi: 10.1074/jbc.M111.235168. Epub 2011 Jun 15.
Author Address: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605-2324, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 263
Author: Baussant, T., Alonso, C., Wieruszeski, J. M., Strecker, G., Montreuil, J., Alhenc-Gelas, F. and Michalski, J. C.
Year: 1992
Title: Comparative study of asparagine-linked glycans of plasma T-kininogen in normal rats and during acute inflammation
Journal: Biochem J
Volume: 283 ( Pt 2)
Issue: Pt 2
Pages: 531-5
Epub Date: 1992/04/15
Date: Apr 15
Short Title: Comparative study of asparagine-linked glycans of plasma T-kininogen in normal rats and during acute inflammation
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2830531
PMCID: PMC1131068
Accession Number: 1575697
Keywords: Animals
Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Chromatography, Gel
Chromatography, High Pressure Liquid
Inflammation/*blood
Kininogens/chemistry/isolation & purification/*metabolism
Magnetic Resonance Spectroscopy
Male
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Rats
Rats, Inbred Strains
Reference Values
Abstract: Rat T-kininogen has been separated into two molecular variants by affinity chromatography on concanavalin A (ConA): a ConA-reactive (ConA+) and a ConA-non-reactive (ConA-) fraction, from which carbohydrate chains were quantitatively released by hydrazinolysis. On the basis of high-resolution 400 MHz 1H-n.m.r. spectroscopy of the re-N-acetylated hydrazinolysates, the carbohydrate structures of the two ConA molecular variants of rat T-kininogen were established. The ConA-non-reactive species contains a single type of carbohydrate chain with the following structure: [formula: see text] The ConA-reactive fraction contains the same structure and the following additional one: [formula: see text] The relative abundance of the two molecular forms is profoundly affected during inflammation (ratio ConA+/ConA-: 44% in normal and 95% in inflamed T-kininogen), but no structural modification of the carbohydrate chains was observed.
Notes: 1470-8728
Baussant, T
Alonso, C
Wieruszeski, J M
Strecker, G
Montreuil, J
Alhenc-Gelas, F
Michalski, J C
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1992 Apr 15;283 ( Pt 2)(Pt 2):531-5. doi: 10.1042/bj2830531.
Author Address: Unité 36 I.N.S.E.R.M., Paris, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 323
Author: Beaudry, G. A., Mehta, N. M., Ray, M. L. and Bertelsen, A. H.
Year: 1990
Title: Purification and characterization of functional recombinant alpha-amidating enzyme secreted from mammalian cells
Journal: J Biol Chem
Volume: 265
Issue: 29
Pages: 17694-9
Epub Date: 1990/10/15
Date: Oct 15
Short Title: Purification and characterization of functional recombinant alpha-amidating enzyme secreted from mammalian cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2211657
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cell Line
Genetic Vectors
Kinetics
Mice
Mixed Function Oxygenases/genetics/*isolation & purification/metabolism
Molecular Sequence Data
*Multienzyme Complexes
Mutagenesis, Site-Directed
Oligonucleotide Probes
Rats
Recombinant Proteins/biosynthesis/isolation & purification/metabolism
Transfection
Abstract: A rat alpha-amidating enzyme (alpha-AE) cDNA has been expressed in mouse C127 cells using a bovine papilloma virus vector in which transcription was regulated by the mouse metallothionein 1 promoter. The cDNA encoding the full length alpha-AE protein was modified to terminate translation at a site preceding the transmembrane and cytoplasmic domains, thereby enabling functional enzyme to be secreted into the medium. Purification of recombinant alpha-AE to homogeneity indicated that the enzyme was synthesized and secreted as two proteins of 75-77 kDA. The observed heterogeneity was due to inefficient glycosylation at Asn660, as demonstrated by glycopeptidase F digestion. Using the synthetic peptide, dansyl-Tyr-Val-Gly, the specific activity of the recombinant enzyme at pH 7.0 was found to be 1.4 mumol/min/mg and the Km of the enzyme was determined to be 3 microM. The purified recombinant enzyme has maximal activity at pH 4.5-5.5; however, a rapid inactivation of the enzyme occurs in acidic solutions in vitro. This inactivation is diminished when activity is measured at pH 7.0-10.0. The availability of large amounts of readily purified, active recombinant alpha-AE should allow detailed probing of reaction mechanism, copper coordination chemistry, and turn-over-based inactivation events.
Notes: Beaudry, G A
Mehta, N M
Ray, M L
Bertelsen, A H
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1990 Oct 15;265(29):17694-9.
Author Address: Department of Molecular Biology, Unigene Laboratories, Inc., Fairfield, New Jersey 07004.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 781
Author: Becamel, C., Figge, A., Poliak, S., Dumuis, A., Peles, E., Bockaert, J., Lubbert, H. and Ullmer, C.
Year: 2001
Title: Interaction of serotonin 5-hydroxytryptamine type 2C receptors with PDZ10 of the multi-PDZ domain protein MUPP1
Journal: J Biol Chem
Volume: 276
Issue: 16
Pages: 12974-82
Epub Date: 2001/01/21
Date: Apr 20
Short Title: Interaction of serotonin 5-hydroxytryptamine type 2C receptors with PDZ10 of the multi-PDZ domain protein MUPP1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M008089200
Accession Number: 11150294
Keywords: Amino Acid Sequence
Animals
Brain/metabolism
COS Cells
Carrier Proteins/*chemistry/*metabolism
Cell Membrane/metabolism
Chlorocebus aethiops
Choroid Plexus/metabolism
Consensus Sequence
Epitopes/chemistry
Intracellular Signaling Peptides and Proteins
Molecular Sequence Data
Rats
Receptor, Serotonin, 5-HT2A
Receptor, Serotonin, 5-HT2B
Receptor, Serotonin, 5-HT2C
Receptors, Serotonin/*chemistry/genetics/*metabolism
Recombinant Proteins/chemistry/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Transcription, Genetic
Transfection
Vesicular stomatitis Indiana virus/chemistry
Abstract: By using the yeast two-hybrid system, we previously isolated a cDNA clone encoding a novel member of the multivalent PDZ protein family called MUPP1 containing 13 PDZ domains. Here we report that the C terminus of the 5-hydroxytryptamine type 2C (5-HT(2C)) receptor selectively interacts with the 10th PDZ domain of MUPP1. Mutations in the extreme C-terminal SSV sequence of the 5-HT(2C) receptor confirmed that the SXV motif is critical for the interaction. Co-immunoprecipitations of MUPP1 and 5-HT(2C) receptors from transfected COS-7 cells and from rat choroid plexus verified this interaction in vivo. Immunocytochemistry revealed an SXV motif-dependent co-clustering of both proteins in transfected COS-7 cells as well as a colocalization in rat choroid plexus. A 5-HT(2C) receptor-dependent unmasking of a C-terminal vesicular stomatitis virus epitope of MUPP1 suggests that the interaction triggers a conformational change within the MUPP1 protein. Moreover, 5-HT(2A) and 5-HT(2B), sharing the C-terminal EX(V/I)SXV sequence with 5-HT(2C) receptors, also bind MUPP1 PDZ domains in vitro. The highest MUPP1 mRNA levels were found in all cerebral cortical layers, the hippocampus, the granular layer of the dentate gyrus, as well as the choroid plexus, where 5-HT(2C) receptors are highly enriched. We propose that MUPP1 may serve as a multivalent scaffold protein that selectively assembles and targets signaling complexes.
Notes: Becamel, C
Figge, A
Poliak, S
Dumuis, A
Peles, E
Bockaert, J
Lubbert, H
Ullmer, C
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2001 Apr 20;276(16):12974-82. doi: 10.1074/jbc.M008089200. Epub 2001 Jan 9.
Author Address: Biofrontera Pharmaceuticals AG, Hemmelratherweg 201, 51377 Leverkusen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1639
Author: Béguin, P., Nagashima, K., Mahalakshmi, R. N., Vigot, R., Matsunaga, A., Miki, T., Ng, M. Y., Ng, Y. J., Lim, C. H., Tay, H. S., Hwang, L. A., Firsov, D., Tang, B. L., Inagaki, N., Mori, Y., Seino, S., Launey, T. and Hunziker, W.
Year: 2014
Title: BARP suppresses voltage-gated calcium channel activity and Ca2+-evoked exocytosis
Journal: J Cell Biol
Volume: 205
Issue: 2
Pages: 233-49
Epub Date: 2014/04/23
Date: Apr 28
Short Title: BARP suppresses voltage-gated calcium channel activity and Ca2+-evoked exocytosis
Alternate Journal: The Journal of cell biology
ISSN: 0021-9525 (Print)
0021-9525
DOI: 10.1083/jcb.201304101
PMCID: PMC4003244
Accession Number: 24751537
Keywords: Animals
Binding Sites
COS Cells
Calcium/*metabolism
Calcium Channels, L-Type/genetics/*metabolism
Chlorocebus aethiops
Cricetinae
Humans
Membrane Glycoproteins/genetics/*metabolism
Mice
Nerve Tissue Proteins/genetics/*metabolism
Neuroendocrine Cells/cytology/*metabolism
Neurons/cytology/*metabolism
PC12 Cells
Protein Binding
Protein Structure, Tertiary
Rats
Abstract: Voltage-gated calcium channels (VGCCs) are key regulators of cell signaling and Ca(2+)-dependent release of neurotransmitters and hormones. Understanding the mechanisms that inactivate VGCCs to prevent intracellular Ca(2+) overload and govern their specific subcellular localization is of critical importance. We report the identification and functional characterization of VGCC β-anchoring and -regulatory protein (BARP), a previously uncharacterized integral membrane glycoprotein expressed in neuroendocrine cells and neurons. BARP interacts via two cytosolic domains (I and II) with all Cavβ subunit isoforms, affecting their subcellular localization and suppressing VGCC activity. Domain I interacts at the α1 interaction domain-binding pocket in Cavβ and interferes with the association between Cavβ and Cavα1. In the absence of domain I binding, BARP can form a ternary complex with Cavα1 and Cavβ via domain II. BARP does not affect cell surface expression of Cavα1 but inhibits Ca(2+) channel activity at the plasma membrane, resulting in the inhibition of Ca(2+)-evoked exocytosis. Thus, BARP can modulate the localization of Cavβ and its association with the Cavα1 subunit to negatively regulate VGCC activity.
Notes: 1540-8140
Béguin, Pascal
Nagashima, Kazuaki
Mahalakshmi, Ramasubbu N
Vigot, Réjan
Matsunaga, Atsuko
Miki, Takafumi
Ng, Mei Yong
Ng, Yu Jin Alvin
Lim, Chiaw Hwee
Tay, Hock Soon
Hwang, Le-Ann
Firsov, Dmitri
Tang, Bor Luen
Inagaki, Nobuya
Mori, Yasuo
Seino, Susumu
Launey, Thomas
Hunziker, Walter
Journal Article
Research Support, Non-U.S. Gov't
J Cell Biol. 2014 Apr 28;205(2):233-49. doi: 10.1083/jcb.201304101. Epub 2014 Apr 21.
Author Address: Epithelial Cell Biology Laboratory and 2 Monoclonal Antibody Unit, Institute of Molecular and Cell Biology, Agency for Science, Technology and Research, Singapore 138673.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1620
Author: Behnken, H. N., Ruthenbeck, A., Schulz, J. M. and Meyer, B.
Year: 2014
Title: Glycan analysis of Prostate Specific Antigen (PSA) directly from the intact glycoprotein by HR-ESI/TOF-MS
Journal: J Proteome Res
Volume: 13
Issue: 2
Pages: 997-1001
Epub Date: 2014/01/08
Date: Feb 7
Short Title: Glycan analysis of Prostate Specific Antigen (PSA) directly from the intact glycoprotein by HR-ESI/TOF-MS
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr400999y
Accession Number: 24393138
Keywords: Chromatography, Liquid
Glycoproteins/*chemistry
Polysaccharides/*analysis
Prostate-Specific Antigen/*chemistry
Spectrometry, Mass, Electrospray Ionization/*methods
Abstract: Glycans are important modulators of the biological function of proteins and are normally characterized from proteolytic glycopeptides or from (N-)glycans released enzymatically by glycosidase treatment or chemically by hydrazinolysis. We demonstrate that glycan compositions can easily be determined directly by LC-ESI/TOF-MS from intact glycoproteins even with a very complex glycosylation pattern. Interpretation of isotopically resolved mass spectra of prostate specific antigen (PSA) using bioinformatics tools gives within a few hours the glycan compositions of 38 glycoforms.
Notes: 1535-3907
Behnken, Henning N
Ruthenbeck, Alexandra
Schulz, Jan-Mirco
Meyer, Bernd
Journal Article
United States
J Proteome Res. 2014 Feb 7;13(2):997-1001. doi: 10.1021/pr400999y. Epub 2014 Jan 21.
Author Address: Organic Chemistry, Department of Chemistry, University of Hamburg , Martin-Luther-King Platz 6, 20144 Hamburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1160
Author: Beigneux, A. P., Gin, P., Davies, B. S., Weinstein, M. M., Bensadoun, A., Ryan, R. O., Fong, L. G. and Young, S. G.
Year: 2008
Title: Glycosylation of Asn-76 in mouse GPIHBP1 is critical for its appearance on the cell surface and the binding of chylomicrons and lipoprotein lipase
Journal: J Lipid Res
Volume: 49
Issue: 6
Pages: 1312-21
Epub Date: 2008/03/15
Date: Jun
Short Title: Glycosylation of Asn-76 in mouse GPIHBP1 is critical for its appearance on the cell surface and the binding of chylomicrons and lipoprotein lipase
Alternate Journal: Journal of lipid research
ISSN: 0022-2275 (Print)
0022-2275
DOI: 10.1194/jlr.M700593-JLR200
PMCID: PMC3055742
Accession Number: 18340083
Keywords: Amino Acid Sequence
Animals
Asparagine/*metabolism
Base Sequence
Blotting, Western
CHO Cells
Cell Membrane/metabolism
Chylomicrons/*metabolism
Cricetinae
Cricetulus
DNA Primers
Glycosylation
Humans
Lipoprotein Lipase/*metabolism
Mice
Microscopy, Fluorescence
Protein Binding
Protein Transport
Receptors, Lipoprotein/chemistry/*metabolism
Sequence Homology, Amino Acid
Abstract: GPIHBP1 is a glycosylphosphatidylinositol-anchored protein in the lymphocyte antigen 6 (Ly-6) family that recently was identified as a platform for the lipolytic processing of triglyceride-rich lipoproteins. GPIHBP1 binds both LPL and chylomicrons and is expressed on the luminal face of microvascular endothelial cells. Here, we show that mouse GPIHBP1 is N-glycosylated at Asn-76 within the Ly-6 domain. Human GPIHBP1 is also glycosylated. The N-linked glycan could be released from mouse GPIHBP1 with N-glycosidase F, endoglycosidase H, or endoglycosidase F1. The glycan was marginally sensitive to endoglycosidase F2 digestion but resistant to endoglycosidase F3 digestion, suggesting that the glycan on GPIHBP1 is of the oligomannose type. Mutating the N-glycosylation site in mouse GPIHBP1 results in an accumulation of GPIHBP1 in the endoplasmic reticulum and a markedly reduced amount of the protein on the cell surface. Consistent with this finding, cells expressing a nonglycosylated GPIHBP1 lack the ability to bind LPL or chylomicrons. Eliminating the N-glycosylation site in a truncated soluble version of GPIHBP1 causes a modest reduction in the secretion of the protein. These studies demonstrate that N-glycosylation of GPIHBP1 is important for the trafficking of GPIHBP1 to the cell surface.
Notes: Beigneux, Anne P
Gin, Peter
Davies, Brandon S J
Weinstein, Michael M
Bensadoun, André
Ryan, Robert O
Fong, Loren G
Young, Stephen G
HL66621/HL/NHLBI NIH HHS/United States
R01 HL094732-01S1/HL/NHLBI NIH HHS/United States
HL66600/HL/NHLBI NIH HHS/United States
R01 HL094732-01/HL/NHLBI NIH HHS/United States
R01 HL094732-02/HL/NHLBI NIH HHS/United States
HL073061/HL/NHLBI NIH HHS/United States
R01 HL087228/HL/NHLBI NIH HHS/United States
R01 HL094732/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Lipid Res. 2008 Jun;49(6):1312-21. doi: 10.1194/jlr.M700593-JLR200. Epub 2008 Mar 13.
Author Address: Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095, USA. abeigneux@mednet.ucla.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1103
Author: Beinrohr, L., Harmat, V., Dobó, J., Lörincz, Z., Gál, P. and Závodszky, P.
Year: 2007
Title: C1 inhibitor serpin domain structure reveals the likely mechanism of heparin potentiation and conformational disease
Journal: J Biol Chem
Volume: 282
Issue: 29
Pages: 21100-9
Epub Date: 2007/05/10
Date: Jul 20
Short Title: C1 inhibitor serpin domain structure reveals the likely mechanism of heparin potentiation and conformational disease
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M700841200
Accession Number: 17488724
Keywords: Alleles
Angioedema/pathology/therapy
Anions
Complement C1 Inactivator Proteins/*chemistry
Complement C1 Inhibitor Protein
Glycosylation
Heparin/*chemistry
Humans
Kinetics
Models, Molecular
Mutagenesis
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Recombinant Proteins/chemistry
Serpins/*chemistry
Surface Properties
Abstract: C1 inhibitor, a member of the serpin family, is a major down-regulator of inflammatory processes in blood. Genetic deficiency of C1 inhibitor results in hereditary angioedema, a dominantly inheritable, potentially lethal disease. Here we report the first crystal structure of the serpin domain of human C1 inhibitor, representing a previously unreported latent form, which explains functional consequences of several naturally occurring mutations, two of which are discussed in detail. The presented structure displays a novel conformation with a seven-stranded beta-sheet A. The unique conformation of the C-terminal six residues suggests its potential role as a barrier in the active-latent transition. On the basis of surface charge pattern, heparin affinity measurements, and docking of a heparin disaccharide, a heparin binding site is proposed in the contact area of the serpin-proteinase encounter complex. We show how polyanions change the activity of the C1 inhibitor by a novel "sandwich" mechanism, explaining earlier reaction kinetic and mutagenesis studies. These results may help to improve therapeutic C1 inhibitor preparations used in the treatment of hereditary angioedema, organ transplant rejection, and heart attack.
Notes: Beinrohr, László
Harmat, Veronika
Dobó, József
Lörincz, Zsolt
Gál, Péter
Závodszky, Péter
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2007 Jul 20;282(29):21100-9. doi: 10.1074/jbc.M700841200. Epub 2007 May 8.
Author Address: Institute of Enzymology, Biological Research Center, Hungarian Academy of Sciences, Karolina út 29, H-1113 Budapest, Hungary. lbeinrohr@enzim.hu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 382
Author: Beintema, J. J., Blank, A., Schieven, G. L., Dekker, C. A., Sorrentino, S. and Libonati, M.
Year: 1988
Title: Differences in glycosylation pattern of human secretory ribonucleases
Journal: Biochem J
Volume: 255
Issue: 2
Pages: 501-5
Epub Date: 1988/10/15
Date: Oct 15
Short Title: Differences in glycosylation pattern of human secretory ribonucleases
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
PMCID: PMC1135256
Accession Number: 3202829
Keywords: Amino Acid Sequence
Carbohydrates/analysis
Glycopeptides/analysis
Glycosylation
Humans
Male
Molecular Sequence Data
Pancreas/enzymology
Ribonucleases/analysis/*metabolism/urine
Semen/enzymology
Abstract: The major secretory ribonuclease (RNase) of human urine (RNase HUA) was isolated and sequenced by automatic Edman degradation and analysis of peptides and glycopeptides. The isolated enzyme was shown to be free of other urine RNase activities by SDS/polyacrylamide-gel electrophoresis and activity staining. It is a glycoprotein 128 amino acids long, differing from human pancreatic RNase in the presence of an additional threonine residue at the C-terminus. It differs from the pancreatic enzyme in its glycosylation pattern as well, and contains about 45 sugar residues. Each of the three Asn-Xaa-Ser/Thr sequences (Asn-34, Asn-76, Asn-88) is glycosylated with a complex-type oligosaccharide chain. Glycosylation at Asn-88 has not been observed previously in mammalian secretory RNases. Preliminary sequence data on the major RNase of human seminal plasma have revealed no difference between it and the major urinary enzyme; their similarities include the presence of threonine at the C-terminus. The glycosylation pattern of human seminal RNase is very similar to that of the pancreatic enzyme. The structural differences between the secretory RNases from human pancreas, urine and seminal plasma must originate from organ-specific post-translational modifications of the one primary gene product. Detailed characterization of peptides and the results of gel filtration of tryptic and tryptic/chymotryptic digests of performic acid-oxidized RNase have been deposited as Supplementary Publication SUP 50146 (4 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1988) 249, 5.
Notes: 1470-8728
Beintema, J J
Blank, A
Schieven, G L
Dekker, C A
Sorrentino, S
Libonati, M
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1988 Oct 15;255(2):501-5.
Author Address: Biochemisch Laboratorium, Rijksuniversiteit Groningen, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1003
Author: Bell, J. K., Botos, I., Hall, P. R., Askins, J., Shiloach, J., Segal, D. M. and Davies, D. R.
Year: 2005
Title: The molecular structure of the Toll-like receptor 3 ligand-binding domain
Journal: Proc Natl Acad Sci U S A
Volume: 102
Issue: 31
Pages: 10976-80
Epub Date: 2005/07/27
Date: Aug 2
Short Title: The molecular structure of the Toll-like receptor 3 ligand-binding domain
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0505077102
PMCID: PMC1182468
Accession Number: 16043704
Keywords: Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Humans
In Vitro Techniques
Ligands
Membrane Glycoproteins/*chemistry/genetics/metabolism
Models, Molecular
Molecular Sequence Data
Protein Conformation
Protein Structure, Tertiary
RNA, Double-Stranded/chemistry/metabolism
Receptors, Cell Surface/*chemistry/genetics/metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Toll-Like Receptor 3
Toll-Like Receptors
Abstract: Innate immunity is the first line of defense against invading pathogens. Toll-like receptors (TLRs) act as sentinels of the innate immune system, sensing a variety of ligands from lipopolysaccharide to flagellin to dsRNA through their ligand-binding domain that is composed of leucine-rich repeats (LRRs). Ligand binding initiates a signaling cascade that leads to the up-regulation of inflammation mediators. In this study, we have expressed and crystallized the ectodomain (ECD) of human TLR3, which recognizes dsRNA, a molecular signature of viruses, and have determined the molecular structure to 2.4-A resolution. The overall horseshoe-shaped structure of the TLR3-ECD is formed by 23 repeating LRRs that are capped at each end by specialized non-LRR domains. The extensive beta-sheet on the molecule's concave surface forms a platform for several modifications, including insertions in the LRRs and 11 N-linked glycans. The TLR3-ECD structure indicates how LRR loops can establish distinct pathogen recognition receptors.
Notes: 1091-6490
Bell, Jessica K
Botos, Istvan
Hall, Pamela R
Askins, Janine
Shiloach, Joseph
Segal, David M
Davies, David R
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2005 Aug 2;102(31):10976-80. doi: 10.1073/pnas.0505077102. Epub 2005 Jul 25.
Author Address: Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 670
Author: Bella, J., Kolatkar, P. R., Marlor, C. W., Greve, J. M. and Rossmann, M. G.
Year: 1998
Title: The structure of the two amino-terminal domains of human ICAM-1 suggests how it functions as a rhinovirus receptor and as an LFA-1 integrin ligand
Journal: Proc Natl Acad Sci U S A
Volume: 95
Issue: 8
Pages: 4140-5
Epub Date: 1998/05/16
Date: Apr 14
Short Title: The structure of the two amino-terminal domains of human ICAM-1 suggests how it functions as a rhinovirus receptor and as an LFA-1 integrin ligand
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.95.8.4140
PMCID: PMC22455
Accession Number: 9539703
Keywords: Amino Acid Sequence
Animals
Antigens, CD/chemistry
Binding Sites
Cell Adhesion Molecules/chemistry
Computer Simulation
Crystallography, X-Ray
Dimerization
Humans
Intercellular Adhesion Molecule-1/*chemistry/*physiology/ultrastructure
Ligands
Lymphocyte Function-Associated Antigen-1/*metabolism
Mice
Microscopy, Electron
Models, Molecular
Molecular Sequence Data
*Protein Structure, Secondary
Receptors, Virus/*chemistry/*physiology/ultrastructure
Recombinant Proteins/chemistry/metabolism/ultrastructure
Rhinovirus/*physiology
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: The normal function of human intercellular adhesion molecule-1 (ICAM-1) is to provide adhesion between endothelial cells and leukocytes after injury or stress. ICAM-1 binds to leukocyte function-associated antigen (LFA-1) or macrophage-1 antigen (Mac-1). However, ICAM-1 is also used as a receptor by the major group of human rhinoviruses and is a catalyst for the subsequent viral uncoating during cell entry. The three-dimensional atomic structure of the two amino-terminal domains (D1 and D2) of ICAM-1 has been determined to 2.2-A resolution and fitted into a cryoelectron microscopy reconstruction of a rhinovirus-ICAM-1 complex. Rhinovirus attachment is confined to the BC, CD, DE, and FG loops of the amino-terminal Ig-like domain (D1) at the end distal to the cellular membrane. The loops are considerably different in structure to those of human ICAM-2 or murine ICAM-1, which do not bind rhinoviruses. There are extensive charge interactions between ICAM-1 and human rhinoviruses, which are mostly conserved in both major and minor receptor groups of rhinoviruses. The interaction of ICAMs with LFA-1 is known to be mediated by a divalent cation bound to the insertion (I)-domain on the alpha chain of LFA-1 and the carboxyl group of a conserved glutamic acid residue on ICAMs. Domain D1 has been docked with the known structure of the I-domain. The resultant model is consistent with mutational data and provides a structural framework for the adhesion between these molecules.
Notes: 1091-6490
Bella, J
Kolatkar, P R
Marlor, C W
Greve, J M
Rossmann, M G
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1998 Apr 14;95(8):4140-5. doi: 10.1073/pnas.95.8.4140.
Author Address: Department of Biological Sciences, Purdue University, West Lafayette, IN 47907-1392, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1068
Author: Benjannet, S., Rhainds, D., Hamelin, J., Nassoury, N. and Seidah, N. G.
Year: 2006
Title: The proprotein convertase (PC) PCSK9 is inactivated by furin and/or PC5/6A: functional consequences of natural mutations and post-translational modifications
Journal: J Biol Chem
Volume: 281
Issue: 41
Pages: 30561-72
Epub Date: 2006/08/17
Date: Oct 13
Short Title: The proprotein convertase (PC) PCSK9 is inactivated by furin and/or PC5/6A: functional consequences of natural mutations and post-translational modifications
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M606495200
Accession Number: 16912035
Keywords: Amino Acid Sequence
Cell Line, Tumor
Endoplasmic Reticulum/metabolism
Female
Furin/*chemistry
Humans
Lipoproteins, LDL/metabolism
Male
Molecular Sequence Data
*Mutation
Proprotein Convertase 5/*chemistry
Proprotein Convertase 9
Proprotein Convertases
*Protein Processing, Post-Translational
Serine Endopeptidases/*genetics/metabolism/*physiology
trans-Golgi Network/metabolism
Abstract: PCSK9 is the ninth member of the proprotein convertase (PC) family. Some of its natural mutations have been genetically associated with the development of a dominant form of familial hyper- or hypocholesterolemia. The exact mechanism of action of PCSK9 is not clear, although it is known to enhance the intracellular degradation of the low density lipoprotein (LDL) receptor in acidic compartments, likely the endosomes/lysosomes. We analyzed the post-translational modifications of PCSK9 and show that it is sulfated within its prosegment at Tyr38. We also examined the susceptibility of PCSK9 to proteolytic cleavage by the other members of the PC family. The data show that the natural gain-of-function mutations R218S, F216L, and D374Y associated with hypercholesterolemia result in total or partial loss of furin/PC5/6A processing at the motif RFHR218 downward arrow. In contrast, the loss-of-function mutations A443T and C679X lead either to the lack of trans-Golgi network/recycling endosome localization and an enhanced susceptibility to furin cleavage (A443T) or to the inability of PCSK9 to exit the endoplasmic reticulum (C679X). Furthermore, we report the presence of both native and furin-like cleaved forms of PCSK9 in circulating human plasma. Thus, we propose that PCSK9 levels are finely regulated by the basic amino acid convertases furin and PC5/6A. The latter may reduce the lifetime of this proteinase and its ability to degrade the cell-surface LDL receptor, thereby regulating the levels of circulating LDL cholesterol.
Notes: Benjannet, Suzanne
Rhainds, David
Hamelin, Josée
Nassoury, Nasha
Seidah, Nabil G
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Oct 13;281(41):30561-72. doi: 10.1074/jbc.M606495200. Epub 2006 Aug 15.
Author Address: Laboratory of Biochemical Neuroendocrinology, Clinical Research Institute of Montreal, Montreal, Quebec H2W 1R7, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1836
Author: Benoit, R. M., Schärer, M. A., Wieser, M. M., Li, X., Frey, D. and Kammerer, R. A.
Year: 2017
Title: Crystal structure of the BoNT/A2 receptor-binding domain in complex with the luminal domain of its neuronal receptor SV2C
Journal: Sci Rep
Volume: 7
Pages: 43588
Epub Date: 2017/03/03
Date: Mar 2
Short Title: Crystal structure of the BoNT/A2 receptor-binding domain in complex with the luminal domain of its neuronal receptor SV2C
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/srep43588
PMCID: PMC5333631
Accession Number: 28252640
Keywords: Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Botulinum Toxins, Type A/*chemistry/metabolism
Humans
Membrane Glycoproteins/*chemistry
Models, Molecular
Nerve Tissue Proteins/*chemistry
Protein Binding
Protein Conformation
*Protein Interaction Domains and Motifs
Abstract: A detailed molecular understanding of botulinum neurotoxin (BoNT)/host-cell-receptor interactions is fundamental both for developing strategies against botulism and for generating improved BoNT variants for medical applications. The X-ray crystal structure of the receptor-binding domain (H(C)) of BoNT/A1 in complex with the luminal domain (LD) of its neuronal receptor SV2C revealed only few specific side-chain - side-chain interactions that are important for binding. Notably, two BoNT/A1 residues, Arg 1156 and Arg 1294, that are crucial for the interaction with SV2, are not conserved among subtypes. Because it has been suggested that differential receptor binding of subtypes might explain their differences in biological activity, we determined the crystal structure of BoNT/A2-H(C) in complex with SV2C-LD. Although only few side-chain interactions are conserved between the two BoNT/A subtypes, the overall binding mode of subtypes A1 and A2 is virtually identical. In the BoNT/A2-H(C) - SV2C complex structure, a missing cation-π stacking is compensated for by an additional salt bridge and an anion-π stacking interaction, which explains why the binding of BoNT/A subtypes to SV2C tolerates variable side chains. These findings suggest that motif extensions and a shallow binding cleft in BoNT/A-H(C) contribute to binding specificity.
Notes: 2045-2322
Benoit, Roger M
Schärer, Martin A
Wieser, Mara M
Li, Xiaodan
Frey, Daniel
Kammerer, Richard A
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2017 Mar 2;7:43588. doi: 10.1038/srep43588.
Author Address: Laboratory of Biomolecular Research, Division of Biology and Chemistry, Paul Scherrer Institute, CH-5232 Villigen PSI, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 248
Author: Bergwerff, A. A., Thomas-Oates, J. E., van Oostrum, J., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1992
Title: Human urokinase contains GalNAc beta (1-4)[Fuc alpha (1-3)]GlcNAc beta (1-2) as a novel terminal element in N-linked carbohydrate chains
Journal: FEBS Lett
Volume: 314
Issue: 3
Pages: 389-94
Epub Date: 1992/12/21
Date: Dec 21
Short Title: Human urokinase contains GalNAc beta (1-4)[Fuc alpha (1-3)]GlcNAc beta (1-2) as a novel terminal element in N-linked carbohydrate chains
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(92)81512-k
Accession Number: 1468573
Keywords: Carbohydrate Sequence
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Spectrometry, Mass, Fast Atom Bombardment
Urokinase-Type Plasminogen Activator/*chemistry
Abstract: Structural analysis of enzymically released N-linked carbohydrate chains of human urokinase (urinary-type plasminogen activator) by 1H NMR spectroscopy and FAB-MS demonstrated that the N-linked oligosaccharides on the only N-glycosylation site contain diantennary structures with the novel GalNAc beta (1-4) [Fuc alpha (1-3)]GlcNAc beta (1-2) element in the upper or the lower branch.
Notes: Bergwerff, A A
Thomas-Oates, J E
van Oostrum, J
Kamerling, J P
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1992 Dec 21;314(3):389-94. doi: 10.1016/0014-5793(92)81512-k.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 595
Author: Bergwerff, A. A., van Oostrum, J., Asselbergs, F. A., Bürgi, R., Hokke, C. H., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1993
Title: Primary structure of N-linked carbohydrate chains of a human chimeric plasminogen activator K2tu-PA expressed in Chinese hamster ovary cells
Journal: Eur J Biochem
Volume: 212
Issue: 3
Pages: 639-56
Epub Date: 1993/03/15
Date: Mar 15
Short Title: Primary structure of N-linked carbohydrate chains of a human chimeric plasminogen activator K2tu-PA expressed in Chinese hamster ovary cells
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb17702.x
Accession Number: 8462541
Keywords: Amino Acid Sequence
Animals
Asparagine
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chimera
Cricetinae
Genetic Variation
Glycopeptides/isolation & purification
Glycosylation
Humans
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Mapping
Protein Structure, Secondary
Recombinant Proteins/biosynthesis/chemistry/isolation & purification
Tissue Plasminogen Activator/biosynthesis/*chemistry/genetics
Transfection
Urokinase-Type Plasminogen Activator/biosynthesis/*chemistry/genetics
Abstract: A recombinant human plasminogen activator hybrid variant K2tu-PA, expressed in Chinese hamster ovary cells, is partially glycosylated at Asn12 (A chain, kringle-2 domain) and completely glycosylated at Asn247 (B chain, protease domain). After release of the N-linked carbohydrate chains by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F, the oligosaccharides were separated from the protein by gel permeation chromatography, then fractionated by FPLC on Mono Q, followed by HPLC on Lichrosorb-NH2, and analysed by 500-MHz 1H-NMR spectroscopy. The following types of carbohydrates occur: monosialylated diantennary (8%), disialylated diantennary (45%), disialylated tri- and tri'-antennary (1%), trisialylated tri- and tri'-antennary (28%), and tetrasialylated tetra-antennary (18%) structures, all having fucose in alpha(1-6)-linkage at the Asn-bound N-acetylglucosamine. Sialic acid occurred exclusively in alpha(2-3)-linkage to galactose, and consisted of N-acetylneuraminic acid (94%), N-glycolylneuraminic acid (3%), and N-acetyl-9-O-acetylneuraminic acid (3%). In addition, glycopeptide fragments corresponding with the A or B chain of K2tu-PA were analysed. The oligosaccharides attached to Asn12 are less processed than those attached to Asn247. Comparison of the glycosylation pattern of K2tu-PA with that of tissue-type plasminogen activator from different biological sources showed significant differences. Profiling studies on different K2tu-PA production batches demonstrated that the structures of N-linked oligosaccharides were identical, but that relative amounts vary with the applied isolation procedure of the chimeric glycoprotein.
Notes: Bergwerff, A A
van Oostrum, J
Asselbergs, F A
Bürgi, R
Hokke, C H
Kamerling, J P
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1993 Mar 15;212(3):639-56. doi: 10.1111/j.1432-1033.1993.tb17702.x.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1581
Author: Berk, J. M., Maitra, S., Dawdy, A. W., Shabanowitz, J., Hunt, D. F. and Wilson, K. L.
Year: 2013
Title: O-Linked β-N-acetylglucosamine (O-GlcNAc) regulates emerin binding to barrier to autointegration factor (BAF) in a chromatin- and lamin B-enriched "niche"
Journal: J Biol Chem
Volume: 288
Issue: 42
Pages: 30192-30209
Epub Date: 2013/09/10
Date: Oct 18
Short Title: O-Linked β-N-acetylglucosamine (O-GlcNAc) regulates emerin binding to barrier to autointegration factor (BAF) in a chromatin- and lamin B-enriched "niche"
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.503060
PMCID: PMC3798487
Accession Number: 24014020
Keywords: Acetylglucosamine
Acylation/physiology
Amino Acid Substitution
Chromatin/genetics/*metabolism
DNA-Binding Proteins/genetics/*metabolism
HeLa Cells
Histones/genetics/metabolism
Humans
Laminin/genetics/*metabolism
Membrane Proteins/genetics/*metabolism
Mutation, Missense
N-Acetylglucosaminyltransferases/genetics/*metabolism
Nuclear Lamina/genetics/*metabolism
Nuclear Proteins/genetics/*metabolism
Phosphorylation/physiology
Cardiomyopathy
Chromosomes/Non-histone Chromosomal Proteins
Emerin
Lamin
Laminopathy
Muscular Dystrophy
Nuclear Matrix
Nuclear Membrane
Nucleoskeleton
O-GlcNAc
Abstract: Emerin, a membrane component of nuclear "lamina" networks with lamins and barrier to autointegration factor (BAF), is highly O-GlcNAc-modified ("O-GlcNAcylated") in mammalian cells. Mass spectrometry analysis revealed eight sites of O-GlcNAcylation, including Ser-53, Ser-54, Ser-87, Ser-171, and Ser-173. Emerin O-GlcNAcylation was reduced ~50% by S53A or S54A mutation in vitro and in vivo. O-GlcNAcylation was reduced ~66% by the triple S52A/S53A/S54A mutant, and S173A reduced O-GlcNAcylation of the S52A/S53A/S54A mutant by ~30%, in vivo. We separated two populations of emerin, A-type lamins and BAF; one population solubilized easily, and the other required sonication and included histones and B-type lamins. Emerin and BAF associated only in histone- and lamin-B-containing fractions. The S173D mutation specifically and selectively reduced GFP-emerin association with BAF by 58% and also increased GFP-emerin hyper-phosphorylation. We conclude that β-N-acetylglucosaminyltransferase, an essential enzyme, controls two regions in emerin. The first region, defined by residues Ser-53 and Ser-54, flanks the LEM domain. O-GlcNAc modification at Ser-173, in the second region, is proposed to promote emerin association with BAF in the chromatin/lamin B "niche." These results reveal direct control of a conserved LEM domain nuclear lamina component by β-N-acetylglucosaminyltransferase, a nutrient sensor that regulates cell stress responses, mitosis, and epigenetics.
Notes: 1083-351x
Berk, Jason M
Maitra, Sushmit
Dawdy, Andrew W
Shabanowitz, Jeffrey
Hunt, Donald F
Wilson, Katherine L
R01 GM037537/GM/NIGMS NIH HHS/United States
R01 GM048646/GM/NIGMS NIH HHS/United States
T32 GM007445/GM/NIGMS NIH HHS/United States
R01 GM37537/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2013 Oct 18;288(42):30192-30209. doi: 10.1074/jbc.M113.503060. Epub 2013 Sep 6.
Author Address: From the Department of Cell Biology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 and.
the Department of Chemistry, University of Virginia, Charlottesville, Virginia 22904.
From the Department of Cell Biology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 and. Electronic address: klwilson@jhmi.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1095
Author: Bernhard, O. K., Kapp, E. A. and Simpson, R. J.
Year: 2007
Title: Enhanced analysis of the mouse plasma proteome using cysteine-containing tryptic glycopeptides
Journal: J Proteome Res
Volume: 6
Issue: 3
Pages: 987-95
Epub Date: 2007/03/03
Date: Mar
Short Title: Enhanced analysis of the mouse plasma proteome using cysteine-containing tryptic glycopeptides
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr0604559
Accession Number: 17330941
Keywords: Animals
Blood Proteins/*analysis
Chromatography, Liquid
Cysteine
*Glycopeptides
Glycosylation
Mice
Mice, Inbred C57BL
Proteomics/*methods
Tandem Mass Spectrometry
Trypsin
Abstract: A comprehensive understanding of the mouse plasma proteome is important for studies using mouse models to identify protein markers of human disease. To enhance our analysis of the mouse plasma proteome, we have developed a method for isolating low-abundance proteins using a cysteine-containing glycopeptide strategy. This method involves two orthogonal affinity capture steps. First, glycoproteins are coupled to an azlactone copolymer gel using hydrazide chemistry and cysteine residues are then biotinylated. After trypsinization and extensive washing, tethered N-glycosylated tryptic peptides are released from the gel using PNGase F. Biotinylated cysteinyl-containing glycopeptides are then affinity selected using a monomeric avidin gel and analyzed by LC-MS/MS. We have applied the method to a proteome analysis of mouse plasma. In two independent analyses using 200 muL each of C57BL mouse plasma, 51 proteins were detected. Only 42 proteins were seen when the same plasma sample was analyzed by glycopeptides only. A total of 104 N-glycosylation sites were identified. Of these, 17 sites have hitherto not been annotated in the Swiss-Prot database whereas 48 were considered probable, potential, or by similarity - i.e., based on little or no experimental evidence. We show that analysis by cysteine-containing glycopeptides allows detection of low-abundance proteins such as the epidermal growth factor receptor, the Vitamin K-dependent protein Z, the hepatocyte growth factor activator, and the lymphatic endothelium-specific hyaluronan receptor as these proteins were not detected in the glycopeptide control analysis.
Notes: Bernhard, Oliver K
Kapp, Eugene A
Simpson, Richard J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2007 Mar;6(3):987-95. doi: 10.1021/pr0604559.
Author Address: Joint ProteomicS Laboratory, The Ludwig Institute for Cancer Research and the Walter and Eliza Hall Instititute, Royal Melbourne Hospital, Parkville, Victoria 3050, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1980
Author: Berthier, A., Vinod, M., Porez, G., Steenackers, A., Alexandre, J., Yamakawa, N., Gheeraert, C., Ploton, M., Maréchal, X., Dubois-Chevalier, J., Hovasse, A., Schaeffer-Reiss, C., Cianférani, S., Rolando, C., Bray, F., Duez, H., Eeckhoute, J., Lefebvre, T., Staels, B. and Lefebvre, P.
Year: 2018
Title: Combinatorial regulation of hepatic cytoplasmic signaling and nuclear transcriptional events by the OGT/REV-ERBα complex
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 47
Pages: E11033-e11042
Epub Date: 2018/11/07
Date: Nov 20
Short Title: Combinatorial regulation of hepatic cytoplasmic signaling and nuclear transcriptional events by the OGT/REV-ERBα complex
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1805397115
PMCID: PMC6255172
Accession Number: 30397120
Keywords: Animals
Cell Line, Tumor
Circadian Clocks/physiology
Gene Expression Regulation/genetics
Glucose/metabolism
HEK293 Cells
Hep G2 Cells
Humans
Insulin/*metabolism
Lipid Metabolism/physiology
Liver/metabolism
Mice
Mice, Inbred C57BL
Mice, Knockout
N-Acetylglucosaminyltransferases/genetics/*metabolism
Nuclear Receptor Subfamily 1, Group D, Member 1/genetics/*metabolism
Phosphorylation
Proto-Oncogene Proteins c-akt/metabolism
Signal Transduction
Sterol Regulatory Element Binding Protein 1/*biosynthesis/genetics
*O-GlcNAcylation
*Rev-erbα
*epigenomics
*metabolism
*signal transduction
Abstract: The nuclear receptor REV-ERBα integrates the circadian clock with hepatic glucose and lipid metabolism by nucleating transcriptional comodulators at genomic regulatory regions. An interactomic approach identified O-GlcNAc transferase (OGT) as a REV-ERBα-interacting protein. By shielding cytoplasmic OGT from proteasomal degradation and favoring OGT activity in the nucleus, REV-ERBα cyclically increased O-GlcNAcylation of multiple cytoplasmic and nuclear proteins as a function of its rhythmically regulated expression, while REV-ERBα ligands mostly affected cytoplasmic OGT activity. We illustrate this finding by showing that REV-ERBα controls OGT-dependent activities of the cytoplasmic protein kinase AKT, an essential relay in insulin signaling, and of ten-of-eleven translocation (TET) enzymes in the nucleus. AKT phosphorylation was inversely correlated to REV-ERBα expression. REV-ERBα enhanced TET activity and DNA hydroxymethylated cytosine (5hmC) levels in the vicinity of REV-ERBα genomic binding sites. As an example, we show that the REV-ERBα/OGT complex modulates SREBP-1c gene expression throughout the fasting/feeding periods by first repressing AKT phosphorylation and by epigenomically priming the Srebf1 promoter for a further rapid response to insulin. Conclusion: REV-ERBα regulates cytoplasmic and nuclear OGT-controlled processes that integrate at the hepatic SREBF1 locus to control basal and insulin-induced expression of the temporally and nutritionally regulated lipogenic SREBP-1c transcript.
Notes: 1091-6490
Berthier, Alexandre
Orcid: 0000-0003-4153-4810
Vinod, Manjula
Porez, Geoffrey
Steenackers, Agata
Alexandre, Jérémy
Yamakawa, Nao
Gheeraert, Céline
Ploton, Maheul
Orcid: 0000-0002-1569-0070
Maréchal, Xavier
Dubois-Chevalier, Julie
Hovasse, Agnès
Schaeffer-Reiss, Christine
Cianférani, Sarah
Rolando, Christian
Bray, Fabrice
Orcid: 0000-0002-4723-8206
Duez, Hélène
Eeckhoute, Jérôme
Lefebvre, Tony
Staels, Bart
Lefebvre, Philippe
Orcid: 0000-0002-9366-5129
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2018 Nov 20;115(47):E11033-E11042. doi: 10.1073/pnas.1805397115. Epub 2018 Nov 5.
Author Address: University of Lille, Inserm, Centre Hospitalier Universitaire de Lille, Institut Pasteur de Lille, European Genomic Institute for Diabetes, U1011, Lille F-59045, France.
University of Lille, CNRS, Unité de Glycobiologie Structurale et Fonctionnelle, UMR 8576, Villeneuve d'Ascq F-59655, France.
Laboratoire de Spectrométrie de Masse BioOrganique, University of Strasbourg, CNRS, Institut Pluridisciplinaire Hubert Curien, UMR 7178, Strasbourg F-67037, France.
Miniaturisation pour la Synthèse, l'Analyse & la Protéomique, CNRS, Unité de Service et de Recherche (USR) 3290, University of Lille, Villeneuve d'Ascq F-59655, France.
Fédération de Recherche Biochimie Structurale et Fonctionnelle des Assemblages Biomoléculaires FRABio, FR 3688 CNRS, University of Lille, Villeneuve d'Ascq F-59655, France.
Institut M.-E. Chevreul, CNRS, FR 2638, University of Lille, Villeneuve d'Ascq F-59655, France.
University of Lille, Inserm, Centre Hospitalier Universitaire de Lille, Institut Pasteur de Lille, European Genomic Institute for Diabetes, U1011, Lille F-59045, France; philippe-claude.lefebvre@inserm.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 702
Author: Bewley, M. C., Tam, B. M., Grewal, J., He, S., Shewry, S., Murphy, M. E., Mason, A. B., Woodworth, R. C., Baker, E. N. and MacGillivray, R. T.
Year: 1999
Title: X-ray crystallography and mass spectroscopy reveal that the N-lobe of human transferrin expressed in Pichia pastoris is folded correctly but is glycosylated on serine-32
Journal: Biochemistry
Volume: 38
Issue: 8
Pages: 2535-41
Epub Date: 1999/02/25
Date: Feb 23
Short Title: X-ray crystallography and mass spectroscopy reveal that the N-lobe of human transferrin expressed in Pichia pastoris is folded correctly but is glycosylated on serine-32
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi9824543
Accession Number: 10029548
Keywords: Animals
Cell Line
Cricetinae
Crystallization
Crystallography, X-Ray
Ferric Compounds/chemistry
Glycosylation
Humans
Kidney/cytology
Mass Spectrometry
Models, Molecular
Peptide Fragments/*chemistry/genetics/metabolism
Pichia/*genetics
*Protein Folding
Recombinant Proteins/biosynthesis/*chemistry/metabolism
Serine/genetics/*metabolism
Transferrin/*chemistry/genetics/metabolism
Abstract: The ferric form of the N-lobe of human serum transferrin (Fe(III)-hTF/2N) has been expressed at high levels in Pichia pastoris. The Fe(III)-hTF/2N was crystallized in the space group P41212, and X-ray crystallography was used to solve the structure of the recombinant protein at 2.5 A resolution. This represents only the second P. pastoris-derived protein structure determined to date, and allows the comparison of the structures of recombinant Fe(III)-hTF/2N expressed in P. pastoris and mammalian cells with serum-derived transferrin. The polypeptide folding pattern is essentially identical in all of the three proteins. Mass spectroscopic analyses of P. pastoris- hTF/2N and proteolytically derived fragments revealed glycosylation of Ser-32 with a single hexose. This represents the first localization of an O-linked glycan in a P. pastoris-derived protein. Because of its distance from the iron-binding site, glycosylation of Ser-32 should not affect the iron-binding properties of hTF/2N expressed in P. pastoris, making this an excellent expression system for the production of hTF/2N.
Notes: Bewley, M C
Tam, B M
Grewal, J
He, S
Shewry, S
Murphy, M E
Mason, A B
Woodworth, R C
Baker, E N
MacGillivray, R T
R01 DK 21739/DK/NIDDK NIH HHS/United States
R01 DK 35533/DK/NIDDK NIH HHS/United States
R01 HD 20859/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1999 Feb 23;38(8):2535-41. doi: 10.1021/bi9824543.
Author Address: Institute of Molecular Biosciences, College of Sciences, Massey University, Palmerston North, New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 666
Author: Bhavanandan, V. P., Zhu, Q., Yamakami, K., Dilulio, N. A., Nair, S., Capon, C., Lemoine, J. and Fournet, B.
Year: 1998
Title: Purification and characterization of the MUC1 mucin-type glycoprotein, epitectin, from human urine: structures of the major oligosaccharide alditols
Journal: Glycoconj J
Volume: 15
Issue: 1
Pages: 37-49
Epub Date: 1998/04/08
Date: Jan
Short Title: Purification and characterization of the MUC1 mucin-type glycoprotein, epitectin, from human urine: structures of the major oligosaccharide alditols
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1023/a:1006987315827
Accession Number: 9530955
Keywords: Amino Acids/analysis
Antigens, Tumor-Associated, Carbohydrate/chemistry/*urine
Carbohydrate Sequence
Centrifugation, Density Gradient
Chromatography/methods
Glycoconjugates/analysis
Humans
Laryngeal Neoplasms/immunology
Male
Mass Spectrometry
Molecular Sequence Data
Molecular Weight
Monosaccharides/analysis
Mucin-1/chemistry
Tumor Cells, Cultured
Abstract: The MUC1 glycoprotein, epitectin, a component of the human bladder epithelium, was purified from human urine. Sedimentation equilibrium analysis and gel filtration using polysaccharide or protein standards revealed a polydisperse preparation with molecular weights ranging from about 0.9 to 1.3 x 10(6). This suggests that in the native state epitectin exists as aggregates of three or four monomer units of 350-400 kDa. Epitectin was found to have significant affinity to hexyl-, octyl- or phenyl agarose indicating that hydrophobic interactions and possibly carbohydrate-carbohydrate interactions may be responsible for the self-association. Chemical and enzymic deglycosylation of [125I]-labeled urine epitectin and metabolically labeled H.Ep.2 epitectin resulted in extremely polydisperse products. The buoyant densities of epitectin purified from urine and H.Ep.2 cells were found to be 1.39-1.40 g ml(-1), suggesting that the total carbohydrate content of these preparations is not significantly different. The O-linked saccharides of epitectin were fractionated by HPLC and analyzed by permethylation and FAB-MS. The neutral saccharides from both sources contain three common structures, namely Gal1 --> 3GalNAc, GlcNAc1 --> 6 (Gal1 --> 3) GalNAc and Gal1 --> 4GlcNAc --> 6 (Gal1 --> 3)GalNAc. The sialic acid of urine epitectin consisted entirely of N-acetylneuraminic acid. The two sources of epitectin, in vitro labeled on sialic acid, were found to have the same sialyl oligosaccharides but in different proportions. Metabolic labeling and N-glycanase susceptibility experiments firmly established the presence of N-linked saccharides in epitectin as minor components. The remarkable similarities in the total carbohydrate content, the carbohydrate composition and structures of saccharides between epitectin from urine, a non-malignant source, and H.Ep.2 cells is surprising in view of the prevailing view that MUC1 glycoproteins of cancer cells are underglycosylated compared to those produced by non-malignant cells.
Notes: Bhavanandan, V P
Zhu, Q
Yamakami, K
Dilulio, N A
Nair, S
Capon, C
Lemoine, J
Fournet, B
CA38797/CA/NCI NIH HHS/United States
DK47511/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Glycoconj J. 1998 Jan;15(1):37-49. doi: 10.1023/a:1006987315827.
Author Address: Department of Biochemistry and Molecular Biology, Penn State University College of Medicine, Hershey 17033, USA. vbhavana@bcmic.hmc.psu.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1726
Author: Bhuiyan, T., Waridel, P., Kapuria, V., Zoete, V. and Herr, W.
Year: 2015
Title: Distinct OGT-Binding Sites Promote HCF-1 Cleavage
Journal: PLoS One
Volume: 10
Issue: 8
Pages: e0136636
Epub Date: 2015/08/26
Short Title: Distinct OGT-Binding Sites Promote HCF-1 Cleavage
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0136636
PMCID: PMC4549301
Accession Number: 26305326
Keywords: Binding Sites
Cytokinesis/genetics
Glutamic Acid/metabolism
HeLa Cells
Host Cell Factor C1/genetics/*metabolism
Humans
N-Acetylglucosaminyltransferases/genetics/*metabolism
Protein Subunits/genetics/*metabolism
*Proteolysis
Repetitive Sequences, Amino Acid/genetics
*Transcription, Genetic
Abstract: Human HCF-1 (also referred to as HCFC-1) is a transcriptional co-regulator that undergoes a complex maturation process involving extensive O-GlcNAcylation and site-specific proteolysis. HCF-1 proteolysis results in two active, noncovalently associated HCF-1N and HCF-1C subunits that regulate distinct phases of the cell-division cycle. HCF-1 O-GlcNAcylation and site-specific proteolysis are both catalyzed by O-GlcNAc transferase (OGT), which thus displays an unusual dual enzymatic activity. OGT cleaves HCF-1 at six highly conserved 26 amino acid repeat sequences called HCF-1PRO repeats. Here we characterize the substrate requirements for OGT cleavage of HCF-1. We show that the HCF-1PRO-repeat cleavage signal possesses particular OGT-binding properties. The glutamate residue at the cleavage site that is intimately involved in the cleavage reaction specifically inhibits association with OGT and its bound cofactor UDP-GlcNAc. Further, we identify a novel OGT-binding sequence nearby the first HCF-1PRO-repeat cleavage signal that enhances cleavage. These results demonstrate that distinct OGT-binding sites in HCF-1 promote proteolysis, and provide novel insights into the mechanism of this unusual protease activity.
Notes: 1932-6203
Bhuiyan, Tanja
Waridel, Patrice
Kapuria, Vaibhav
Zoete, Vincent
Herr, Winship
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2015 Aug 25;10(8):e0136636. doi: 10.1371/journal.pone.0136636. eCollection 2015.
Author Address: Center for Integrative Genomics, University of Lausanne, Génopode, Lausanne, Switzerland.
Protein Analysis Facility, Center for Integrative Genomics, Faculty of Biology and Medicine, University of Lausanne, Génopode, Lausanne, Switzerland.
Molecular Modeling Group, Swiss Institute of Bioinformatics, Génopode, Lausanne, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1583
Author: Bie, H., Yin, J., He, X., Kermode, A. R., Goddard-Borger, E. D., Withers, S. G. and James, M. N.
Year: 2013
Title: Insights into mucopolysaccharidosis I from the structure and action of α-L-iduronidase
Journal: Nat Chem Biol
Volume: 9
Issue: 11
Pages: 739-45
Epub Date: 2013/09/17
Date: Nov
Short Title: Insights into mucopolysaccharidosis I from the structure and action of α-L-iduronidase
Alternate Journal: Nature chemical biology
ISSN: 1552-4450 (Print)
1552-4450
DOI: 10.1038/nchembio.1357
PMCID: PMC4954775
NIHMSID: CAMS3970
Accession Number: 24036510
Keywords: Crystallography, X-Ray
Humans
Iduronidase/*chemistry/*metabolism
Models, Molecular
Mucopolysaccharidosis I/*enzymology/metabolism
Protein Conformation
Abstract: Mucopolysaccharidosis type I (MPS I), caused by mutations in the gene encoding α-L-iduronidase (IDUA), is one of approximately 70 genetic disorders collectively known as the lysosomal storage diseases. To gain insight into the basis for MPS I, we crystallized human IDUA produced in an Arabidopsis thaliana cgl mutant. IDUA consists of a TIM barrel domain containing the catalytic site, a β-sandwich domain and a fibronectin-like domain. Structures of IDUA bound to iduronate analogs illustrate the Michaelis complex and reveal a (2,5)B conformation in the glycosyl-enzyme intermediate, which suggest a retaining double displacement reaction involving the nucleophilic Glu299 and the general acid/base Glu182. Unexpectedly, the N-glycan attached to Asn372 interacts with iduronate analogs in the active site and is required for enzymatic activity. Finally, these IDUA structures and biochemical analysis of the disease-relevant P533R mutation have enabled us to correlate the effects of mutations in IDUA to clinical phenotypes.
Notes: 1552-4469
Bie, Haiying
Yin, Jiang
He, Xu
Kermode, Allison R
Goddard-Borger, Ethan D
Withers, Stephen G
James, Michael N G
123222-1/Canadian Institutes of Health Research/Canada
MOP123222/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
Nat Chem Biol. 2013 Nov;9(11):739-45. doi: 10.1038/nchembio.1357. Epub 2013 Sep 11.
Author Address: 1] Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada. [2] These authors contributed equally to this work.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1977
Author: Bilyard, M. K., Bailey, H. J., Raich, L., Gafitescu, M. A., Machida, T., Iglésias-Fernández, J., Lee, S. S., Spicer, C. D., Rovira, C., Yue, W. W. and Davis, B. G.
Year: 2018
Title: Palladium-mediated enzyme activation suggests multiphase initiation of glycogenesis
Journal: Nature
Volume: 563
Issue: 7730
Pages: 235-240
Epub Date: 2018/10/26
Date: Nov
Short Title: Palladium-mediated enzyme activation suggests multiphase initiation of glycogenesis
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/s41586-018-0644-7
Accession Number: 30356213
Keywords: Biocatalysis
Enzyme Activation
Galactose/metabolism
Glucose/*biosynthesis
Glucosyltransferases/metabolism
Glycoproteins/metabolism
Glycosylation
Humans
Kinetics
Palladium/*metabolism
Uridine Diphosphate/metabolism
Abstract: Biosynthesis of glycogen, the essential glucose (and hence energy) storage molecule in humans, animals and fungi(1), is initiated by the glycosyltransferase enzyme, glycogenin (GYG). Deficiencies in glycogen formation cause neurodegenerative and metabolic disease(2-4), and mouse knockout(5) and inherited human mutations(6) of GYG impair glycogen synthesis. GYG acts as a 'seed core' for the formation of the glycogen particle by catalysing its own stepwise autoglucosylation to form a covalently bound gluco-oligosaccharide chain at initiation site Tyr 195. Precise mechanistic studies have so far been prevented by an inability to access homogeneous glycoforms of this protein, which unusually acts as both catalyst and substrate. Here we show that unprecedented direct access to different, homogeneously glucosylated states of GYG can be accomplished through a palladium-mediated enzyme activation 'shunt' process using on-protein C-C bond formation. Careful mimicry of GYG intermediates recapitulates catalytic activity at distinct stages, which in turn allows discovery of triphasic kinetics and substrate plasticity in GYG's use of sugar substrates. This reveals a tolerant but 'proof-read' mechanism that underlies the precision of this metabolic process. The present demonstration of direct, chemically controlled access to intermediate states of active enzymes suggests that such ligation-dependent activation could be a powerful tool in the study of mechanism.
Notes: 1476-4687
Bilyard, Matthew K
Bailey, Henry J
Raich, Lluís
Gafitescu, Maria A
Machida, Takuya
Iglésias-Fernández, Javier
Lee, Seung Seo
Spicer, Christopher D
Rovira, Carme
Yue, Wyatt W
Davis, Benjamin G
092809/Z/10/Z/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2018 Nov;563(7730):235-240. doi: 10.1038/s41586-018-0644-7. Epub 2018 Oct 24.
Author Address: Department of Chemistry, University of Oxford, Oxford, UK.
Structural Genomics Consortium, University of Oxford, Oxford, UK.
Departament de Química Inorgànica i Orgànica (Secció de Química Orgànica) and Institut de Química Teòrica i Computacional (IQTC), Universitat de Barcelona, Barcelona, Spain.
Institut de Química Computacional i Catalisi and Departament de Química, Universitat de Girona, Girona, Spain.
School of Chemistry, University of Southampton, Southampton, UK.
Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain.
Structural Genomics Consortium, University of Oxford, Oxford, UK. wyatt.yue@sgc.ox.ac.uk.
Department of Chemistry, University of Oxford, Oxford, UK. ben.davis@chem.ox.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1382
Author: Bimboese, P., Gibson, C. J., Schmidt, S., Xiang, W. and Ehrlich, B. E.
Year: 2011
Title: Isoform-specific regulation of the inositol 1,4,5-trisphosphate receptor by O-linked glycosylation
Journal: J Biol Chem
Volume: 286
Issue: 18
Pages: 15688-97
Epub Date: 2011/03/09
Date: May 6
Short Title: Isoform-specific regulation of the inositol 1,4,5-trisphosphate receptor by O-linked glycosylation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.206482
PMCID: PMC3091177
Accession Number: 21383013
Keywords: Animals
Calcium/*metabolism
Calcium Signaling/*physiology
Cell Line, Tumor
Endoplasmic Reticulum/genetics/*metabolism
Glycosylation
Humans
Inositol 1,4,5-Trisphosphate/genetics/*metabolism
Inositol 1,4,5-Trisphosphate Receptors/genetics/*metabolism
Mice
N-Acetylglucosaminyltransferases/genetics/*metabolism
Protein Isoforms/genetics/metabolism
Rats
Abstract: The inositol 1,4,5-trisphosphate receptor (InsP(3)R), an intracellular calcium channel, has three isoforms with >65% sequence homology, yet the isoforms differ in their function and regulation by post-translational modifications. We showed previously that InsP(3)R-1 is functionally modified by O-linked β-N-acetylglucosamine glycosylation (O-GlcNAcylation) (Rengifo, J., Gibson, C. J., Winkler, E., Collin, T., and Ehrlich, B. E. (2007) J. Neurosci. 27, 13813-13821). We now report the effect of O-GlcNAcylation on InsP(3)R-2 and InsP(3)R-3. Analysis of AR4-2J cells, a rat pancreatoma cell line expressing predominantly InsP(3)R-2, showed no detectable O-GlcNAcylation of InsP(3)R-2 and no significant functional changes despite the presence of the enzymes for addition (O-β-N-acetylglucosaminyltransferase) and removal (O-β-N-acetylglucosaminidase) of the monosaccharide. In contrast, InsP(3)R-3 in Mz-ChA-1 cells, a human cholangiocarcinoma cell line expressing predominantly InsP(3)R-3, was functionally modified by O-GlcNAcylation. Interestingly, the functional impact of O-GlcNAcylation on the InsP(3)R-3 channel was opposite the effect measured with InsP(3)R-1. Addition of O-GlcNAc by O-β-N-acetylglucosaminyltransferase increased InsP(3)R-3 single channel open probability. Incubation of Mz-ChA-1 cells in hyperglycemic medium caused an increase in the InsP(3)-dependent calcium release from the endoplasmic reticulum. The dynamic and inducible nature of O-GlcNAcylation and the InsP(3)R isoform specificity suggest that this form of modification of InsP(3)R and subsequent changes in intracellular calcium transients are important in physiological and pathophysiological processes.
Notes: 1083-351x
Bimboese, Patricia
Gibson, Craig J
Schmidt, Stefan
Xiang, Wanqing
Ehrlich, Barbara E
R01 DK061747/DK/NIDDK NIH HHS/United States
DK57751/DK/NIDDK NIH HHS/United States
DK61747/DK/NIDDK NIH HHS/United States
P01 DK057751/DK/NIDDK NIH HHS/United States
R01 DK061747-08/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 May 6;286(18):15688-97. doi: 10.1074/jbc.M110.206482. Epub 2011 Mar 7.
Author Address: Department of Pharmacology, Yale University, New Haven, Connecticut 06520-8066, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1681
Author: Birkinshaw, R. W., Pellicci, D. G., Cheng, T. Y., Keller, A. N., Sandoval-Romero, M., Gras, S., de Jong, A., Uldrich, A. P., Moody, D. B., Godfrey, D. I. and Rossjohn, J.
Year: 2015
Title: αβ T cell antigen receptor recognition of CD1a presenting self lipid ligands
Journal: Nat Immunol
Volume: 16
Issue: 3
Pages: 258-66
Epub Date: 2015/02/03
Date: Mar
Short Title: αβ T cell antigen receptor recognition of CD1a presenting self lipid ligands
Alternate Journal: Nature immunology
ISSN: 1529-2908 (Print)
1529-2908
DOI: 10.1038/ni.3098
PMCID: PMC7103088
NIHMSID: NIHMS1571368
Accession Number: 25642819
Keywords: Antigen Presentation/*immunology
Antigens, CD1/*immunology
Autoantigens/*immunology
Cell Line
Cell Line, Tumor
HEK293 Cells
Humans
Jurkat Cells
Ligands
Lipids/*immunology
Protein Binding
Receptors, Antigen, T-Cell/*immunology
Receptors, Antigen, T-Cell, alpha-beta/*immunology
T-Lymphocytes/*immunology
Abstract: A central paradigm in αβ T cell-mediated immunity is the simultaneous co-recognition of antigens and antigen-presenting molecules by the αβ T cell antigen receptor (TCR). CD1a presents a broad repertoire of lipid-based antigens. We found that a prototypical autoreactive TCR bound CD1a when it was presenting a series of permissive endogenous ligands, while other lipid ligands were nonpermissive to TCR binding. The structures of two TCR-CD1a-lipid complexes showed that the TCR docked over the A' roof of CD1a in a manner that precluded direct contact with permissive ligands. Nonpermissive ligands indirectly inhibited TCR binding by disrupting the TCR-CD1a contact zone. The exclusive recognition of CD1a by the TCR represents a previously unknown mechanism whereby αβ T cells indirectly sense self antigens that are bound to an antigen-presenting molecule.
Notes: 1529-2916
Birkinshaw, Richard W
Pellicci, Daniel G
Cheng, Tan-Yun
Keller, Andrew N
Sandoval-Romero, Maria
Gras, Stephanie
de Jong, Annemieke
Uldrich, Adam P
Moody, D Branch
Godfrey, Dale I
Rossjohn, Jamie
R01 AI049313/AI/NIAID NIH HHS/United States
R01 AR048632/AR/NIAMS NIH HHS/United States
AI049313/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Immunol. 2015 Mar;16(3):258-66. doi: 10.1038/ni.3098. Epub 2015 Feb 2.
Author Address: 1] Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Australia. [2] ARC Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, Australia.
1] Department of Microbiology and Immunology, Peter Doherty Institute for Infection and Immunity, University of Melbourne, Parkville, Australia. [2] ARC Centre of Excellence in Advanced Molecular Imaging, University of Melbourne, Parkville, Australia.
Brigham and Women's Hospital Division of Rheumatology, Immunology and Allergy and Harvard Medical School, Boston, Massachusetts, USA.
Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Australia.
Department of Dermatology, Columbia University, New York, New York, USA.
1] Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Australia. [2] ARC Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, Australia. [3] Institute of Infection and Immunity, Cardiff University, School of Medicine, Heath Park, Cardiff, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1991
Author: Birrane, G., Beigneux, A. P., Dwyer, B., Strack-Logue, B., Kristensen, K. K., Francone, O. L., Fong, L. G., Mertens, H. D. T., Pan, C. Q., Ploug, M., Young, S. G. and Meiyappan, M.
Year: 2019
Title: Structure of the lipoprotein lipase-GPIHBP1 complex that mediates plasma triglyceride hydrolysis
Journal: Proc Natl Acad Sci U S A
Volume: 116
Issue: 5
Pages: 1723-1732
Epub Date: 2018/12/19
Date: Jan 29
Short Title: Structure of the lipoprotein lipase-GPIHBP1 complex that mediates plasma triglyceride hydrolysis
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1817984116
PMCID: PMC6358717
Accession Number: 30559189
Keywords: Animals
CHO Cells
Capillaries/metabolism
Cell Line
Cricetulus
Crystallography, X-Ray/methods
Endothelial Cells/metabolism
Humans
Hydrolysis
Hypertriglyceridemia/metabolism
Lipoprotein Lipase/*metabolism
Plasma/*metabolism
Receptors, Lipoprotein/*metabolism
Triglycerides/*blood/*metabolism
*Gpihbp1
*X-ray crystallography
*lipase
*lipoproteins
*triglycerides
Shire and hold stock and stock options in Shire. R.Z. and S.G.Y. are coordinators on
a Leducq Transatlantic Network Grant. They have not collaborated directly on this
project.
Abstract: Lipoprotein lipase (LPL) is responsible for the intravascular processing of triglyceride-rich lipoproteins. The LPL within capillaries is bound to GPIHBP1, an endothelial cell protein with a three-fingered LU domain and an N-terminal intrinsically disordered acidic domain. Loss-of-function mutations in LPL or GPIHBP1 cause severe hypertriglyceridemia (chylomicronemia), but structures for LPL and GPIHBP1 have remained elusive. Inspired by our recent discovery that GPIHBP1's acidic domain preserves LPL structure and activity, we crystallized an LPL-GPIHBP1 complex and solved its structure. GPIHBP1's LU domain binds to LPL's C-terminal domain, largely by hydrophobic interactions. Analysis of electrostatic surfaces revealed that LPL contains a large basic patch spanning its N- and C-terminal domains. GPIHBP1's acidic domain was not defined in the electron density map but was positioned to interact with LPL's large basic patch, providing a likely explanation for how GPIHBP1 stabilizes LPL. The LPL-GPIHBP1 structure provides insights into mutations causing chylomicronemia.
Notes: 1091-6490
Birrane, Gabriel
Orcid: 0000-0002-1759-5499
Beigneux, Anne P
Dwyer, Brian
Strack-Logue, Bettina
Kristensen, Kristian Kølby
Francone, Omar L
Fong, Loren G
Mertens, Haydyn D T
Orcid: 0000-0002-5664-744x
Pan, Clark Q
Ploug, Michael
Orcid: 0000-0003-2215-4265
Young, Stephen G
Meiyappan, Muthuraman
Orcid: 0000-0002-0466-0458
R01 HL087228/HL/NHLBI NIH HHS/United States
P01 HL146358/HL/NHLBI NIH HHS/United States
R01 HL125335/HL/NHLBI NIH HHS/United States
P01 HL090553/HL/NHLBI NIH HHS/United States
R35 HL139725/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2019 Jan 29;116(5):1723-1732. doi: 10.1073/pnas.1817984116. Epub 2018 Dec 17.
Author Address: Division of Experimental Medicine, Beth Israel Deaconess Medical Center, Boston, MA 02215.
Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095.
Discovery Therapeutics, US Drug Discovery, Shire Pharmaceuticals, Cambridge, MA 02142.
Finsen Laboratory, Rigshospitalet, DK-2200 Copenhagen, Denmark.
Biotech Research and Innovation Centre, University of Copenhagen, DK-2200 Copenhagen, Denmark.
BIOSAXS Group, European Molecular Biology Laboratory Hamburg, D-22607 Hamburg, Germany.
Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095; sgyoung@mednet.ucla.edu mmeiyappan@shire.com.
Discovery Therapeutics, US Drug Discovery, Shire Pharmaceuticals, Cambridge, MA 02142; sgyoung@mednet.ucla.edu mmeiyappan@shire.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1441
Author: Birtley, J. R., Saridakis, E., Stratikos, E. and Mavridis, I. M.
Year: 2012
Title: The crystal structure of human endoplasmic reticulum aminopeptidase 2 reveals the atomic basis for distinct roles in antigen processing
Journal: Biochemistry
Volume: 51
Issue: 1
Pages: 286-95
Epub Date: 2011/11/24
Date: Jan 10
Short Title: The crystal structure of human endoplasmic reticulum aminopeptidase 2 reveals the atomic basis for distinct roles in antigen processing
Alternate Journal: Biochemistry
ISSN: 0006-2960
DOI: 10.1021/bi201230p
Accession Number: 22106953
Keywords: Aminopeptidases/*chemistry/metabolism/physiology
Antigen Presentation/*immunology
Crystallography, X-Ray
Endoplasmic Reticulum/*enzymology/*immunology
Glycosylation
HLA Antigens/chemistry/physiology
Histocompatibility Antigens Class I/chemistry/physiology
Humans
Minor Histocompatibility Antigens
Models, Molecular
Peptides/chemistry/physiology
Protein Conformation
Protein Precursors/chemistry/physiology
Protein Structure, Tertiary/physiology
Abstract: Endoplasmic reticulum aminopeptidases ERAP1 and ERAP2 cooperate to trim a vast variety of antigenic peptide precursors to generate mature epitopes for binding to major histocompatibility class I molecules. We report here the first structure of ERAP2 determined at 3.08 Å by X-ray crystallography. On the basis of residual electron density, a lysine residue has been modeled in the active site of the enzyme; thus, the structure corresponds to an enzyme-product complex. The overall domain organization is highly similar to that of the recently determined structure of ERAP1 in its closed conformation. A large internal cavity adjacent to the catalytic site can accommodate large peptide substrates. The ERAP2 structure provides a structural explanation for the different peptide N-terminal specificities between ERAP1 and ERAP2 and suggests that such differences extend throughout the whole peptide sequence. A noncrystallographic dimer observed may constitute a model for a proposed ERAP1-ERAP2 heterodimer. Overall, the structure helps explain how two homologous aminopeptidases cooperate to process a large variety of sequences, a key property of their biological role.
Notes: 1520-4995
Birtley, James R
Saridakis, Emmanuel
Stratikos, Efstratios
Mavridis, Irene M
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2012 Jan 10;51(1):286-95. doi: 10.1021/bi201230p. Epub 2011 Dec 9.
Author Address: Structural and Supramolecular Chemistry Laboratory, Institute of Physical Chemistry, National Center for Scientific Research Demokritos, Aghia Paraskevi 15310, Athens, Greece.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1251
Author: Bishop, B., Aricescu, A. R., Harlos, K., O'Callaghan, C. A., Jones, E. Y. and Siebold, C.
Year: 2009
Title: Structural insights into hedgehog ligand sequestration by the human hedgehog-interacting protein HHIP
Journal: Nat Struct Mol Biol
Volume: 16
Issue: 7
Pages: 698-703
Epub Date: 2009/06/30
Date: Jul
Short Title: Structural insights into hedgehog ligand sequestration by the human hedgehog-interacting protein HHIP
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb.1607
PMCID: PMC2709225
NIHMSID: UKMS4576
Accession Number: 19561611
Keywords: Animals
Binding Sites
Calcium/metabolism
Carrier Proteins/*chemistry/genetics/*metabolism
Hedgehog Proteins/*chemistry/genetics/metabolism
Humans
Ligands
Membrane Glycoproteins/*chemistry/genetics/*metabolism
Models, Molecular
Molecular Sequence Data
Multiprotein Complexes/metabolism
*Protein Structure, Secondary
*Protein Structure, Tertiary
Zinc/metabolism
Abstract: Hedgehog (Hh) morphogens have fundamental roles in development, whereas dysregulation of Hh signaling leads to disease. Multiple cell-surface receptors are responsible for transducing and/or regulating Hh signals. Among these, the Hedgehog-interacting protein (Hhip) is a highly conserved, vertebrate-specific inhibitor of Hh signaling. We have solved a series of crystal structures for the human HHIP ectodomain and Desert hedgehog (DHH) in isolation, as well as HHIP in complex with DHH (HHIP-DHH) and Sonic hedgehog (Shh) (HHIP-Shh), with and without Ca2+. The interaction determinants, confirmed by biophysical studies and mutagenesis, reveal previously uncharacterized and distinct functions for the Hh Zn2+ and Ca2+ binding sites--functions that may be common to all vertebrate Hh proteins. Zn2+ makes a key contribution to the Hh-HHIP interface, whereas Ca2+ is likely to prevent electrostatic repulsion between the two proteins, suggesting an important modulatory role. This interplay of several metal binding sites suggests a tuneable mechanism for regulation of Hh signaling.
Notes: 1545-9985
Bishop, Benjamin
Aricescu, A Radu
Harlos, Karl
O'Callaghan, Chris A
Jones, E Yvonne
Siebold, Christian
082301/Wellcome Trust/United Kingdom
Cancer Research UK/United Kingdom
G0700232/Medical Research Council/United Kingdom
Wellcome Trust/United Kingdom
G0500365/Medical Research Council/United Kingdom
G116/165/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Nat Struct Mol Biol. 2009 Jul;16(7):698-703. doi: 10.1038/nsmb.1607. Epub 2009 Jun 28.
Author Address: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2032
Author: Biwi, J., Clarisse, C., Biot, C., Kozak, R. P., Madunic, K., Mortuaire, M., Wuhrer, M., Spencer, D. I. R., Schulz, C., Guerardel, Y., Lefebvre, T. and Vercoutter-Edouart, A. S.
Year: 2019
Title: OGT Controls the Expression and the Glycosylation of E-cadherin, and Affects Glycosphingolipid Structures in Human Colon Cell Lines
Journal: Proteomics
Volume: 19
Issue: 21-22
Pages: e1800452
Epub Date: 2019/08/03
Date: Nov
Short Title: OGT Controls the Expression and the Glycosylation of E-cadherin, and Affects Glycosphingolipid Structures in Human Colon Cell Lines
Alternate Journal: Proteomics
ISSN: 1615-9853
DOI: 10.1002/pmic.201800452
Accession Number: 31373757
Keywords: Cadherins/*genetics
Colorectal Neoplasms/*genetics/pathology
Epithelial-Mesenchymal Transition/genetics
Gene Expression Regulation, Neoplastic/genetics
Gene Silencing
Glycosphingolipids/biosynthesis/genetics
Glycosylation
HCT116 Cells
HT29 Cells
Humans
N-Acetylglucosaminyltransferases/*genetics
Polysaccharides/genetics
*E-cadherin
*O-GlcNAcylation
*colon cancer
*glycomics
*glycosphingolipids
Abstract: Colorectal cancer (CRC) affects both women and men living in societies with a high sedentary lifestyle. Amongst the phenotypic changes exhibited by tumor cells, a wide range of glycosylation has been reported for colon cancer-derived cell lines and CRC tissues. These aberrant modifications affect different aspects of glycosylation, including an increase in core fucosylation and GlcNAc branching on N-glycans, alteration of O-glycans, upregulated sialylation, and O-GlcNAcylation. Although O-GlcNAcylation and complex glycosylations differ in many aspects, sparse evidences report on the interference of O-GlcNAcylation with complex glycosylation. Nevertheless, this relationship is still a matter of debate. Combining different approaches on three human colon cell lines (HT29, HCT116 and CCD841CoN), it is herein reported that silencing O-GlcNAc transferase (OGT, the sole enzyme driving O-GlcNAcylation), only slightly affects overall N- and O-glycosylation patterns. Interestingly, silencing of OGT in HT29 cells upregulates E-cadherin (a major actor of epithelial-to-mesenchymal transition) and changes its glycosylation. On the other hand, OGT silencing perturbs biosynthesis of glycosphingolipids resulting in a decrease in gangliosides and an increase in globosides. Together, these results provide novel insights regarding the selective regulation of complex glycosylations by O-GlcNAcylation in colon cancer cells.
Notes: 1615-9861
Biwi, James
Clarisse, Charlotte
Biot, Christophe
Kozak, Radoslaw Pawel
Madunic, Katarina
Mortuaire, Marlène
Wuhrer, Manfred
Spencer, Daniel Ian Richard
Schulz, Céline
Guerardel, Yann
Lefebvre, Tony
Orcid: 0000-0001-9883-2240
Vercoutter-Edouart, Anne-Sophie
676421/MCCC_/Marie Curie/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2019 Nov;19(21-22):e1800452. doi: 10.1002/pmic.201800452. Epub 2019 Aug 20.
Author Address: Université de Lille, CNRS, UMR 8576, UGSF, Unité de Glycobiologie Structurale et Fonctionnelle, F-59000, Lille, France.
Ludger Ltd, Culham Science Centre, OX14 3EB, Abingdon, Oxfordshire, United Kingdom.
Leiden University Medical Centre, Centre for Proteomics and Metabolomics, 2333ZA, Leiden, Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 292
Author: Bjoern, S., Foster, D. C., Thim, L., Wiberg, F. C., Christensen, M., Komiyama, Y., Pedersen, A. H. and Kisiel, W.
Year: 1991
Title: Human plasma and recombinant factor VII. Characterization of O-glycosylations at serine residues 52 and 60 and effects of site-directed mutagenesis of serine 52 to alanine
Journal: J Biol Chem
Volume: 266
Issue: 17
Pages: 11051-7
Epub Date: 1991/06/15
Date: Jun 15
Short Title: Human plasma and recombinant factor VII. Characterization of O-glycosylations at serine residues 52 and 60 and effects of site-directed mutagenesis of serine 52 to alanine
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1904059
Keywords: *Alanine
Amino Acid Sequence
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Factor VII/*chemistry/genetics/metabolism
Factor X/metabolism
Glycopeptides/isolation & purification
Glycosylation
Humans
Kinetics
Mass Spectrometry
Molecular Sequence Data
Molecular Weight
*Mutagenesis, Site-Directed
Protein Conformation
Recombinant Proteins/chemistry/metabolism
*Serine
Abstract: Factor VII is a multidomain, vitamin K-dependent plasma glycoprotein that participates in the extrinsic pathway of blood coagulation. Earlier studies demonstrated a novel disaccharide (Xyl-Glc) or trisaccharide (Xyl2-Glc) O-glycosidically linked to serine 52 in human plasma factor VII (Nishimura, H., Kawabata, S., Kisiel, W., Hase, S., Ikenaka, T., Shimonishi, Y., and Iwanaga, S. (1989) J. Biol. Chem. 264, 20320-20325). In the present study, human plasma and recombinant factor VII were isolated and subjected to enzymatic fragmentation. Peptides comprising residues 48-62 of the first epidermal growth factor-like domain of each factor VII preparation were isolated for comparative analysis. Using a combined strategy of amino acid sequencing, carbohydrate and amino acid composition analysis, and mass spectrometry, three different glycan structures consisting of either glucose, glucose-xylose, or glucose-(xylose)2 were detected O-glycosidically linked to serine 52 in plasma and recombinant factor VII. Approximately equal amounts of the three glycan structures were observed in plasma factor VII, whereas in recombinant factor VII the glucose and the glucose-(xylose)2 structures predominated. In addition to the O-linked glycan structures observed at serine 52, a single fucose was found to be covalently linked at serine 60 in both human plasma and recombinant factor VII. Carbohydrate and mass spectrometry analyses indicated that the fucosylation of serine 60 was virtually quantitative. Metabolic labeling studies using [14C]fucose confirmed the presence of O-linked fucose at serine 60. In order to assess whether the carbohydrate moiety at serine 52 contributes to the biological activity of factor VII, we have constructed a site-specific mutant of recombinant factor VII in which serine 52 has been replaced with an alanine residue. Mutant factor VIIa exhibited approximately 60% of the coagulant activity of wild-type factor VIIa in a clotting assay. The amidolytic activity of mutant factor VIIa was indistinguishable from that observed for recombinant wild-type factor VIIa. In addition, the ability of mutant factor VIIa in complex with either purified relipidated tissue factor apoprotein or tissue factor on the surface of a human bladder carcinoma cell line (J82) to activate either factor X or factor IX was virtually identical to that observed for wild-type factor VIIa. These results indicate that the carbohydrate moiety O-glycosidically linked to serine 52 does not appear to be involved either in the interaction of factor VIIa with tissue factor, or the expression of its proteolytic activity toward factor X or factor IX following complex formation with tissue factor.(ABSTRACT TRUNCATED AT 400 WORDS)
Notes: Bjoern, S
Foster, D C
Thim, L
Wiberg, F C
Christensen, M
Komiyama, Y
Pedersen, A H
Kisiel, W
HL 35246/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1991 Jun 15;266(17):11051-7.
Author Address: Bioscience Corporate Research, Novo Nordisk A/S, Bagsvaerd, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 437
Author: Blanchard, D., Capon, C., Leroy, Y., Cartron, J. P. and Fournet, B.
Year: 1985
Title: Comparative study of glycophorin A derived O-glycans from human Cad, Sd(a+) and Sd(a-) erythrocytes
Journal: Biochem J
Volume: 232
Issue: 3
Pages: 813-8
Epub Date: 1985/12/15
Date: Dec 15
Short Title: Comparative study of glycophorin A derived O-glycans from human Cad, Sd(a+) and Sd(a-) erythrocytes
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2320813
PMCID: PMC1152955
Accession Number: 4091823
Keywords: *Blood Group Antigens
Borohydrides
Carbohydrates/analysis
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Erythrocyte Membrane/*analysis
*Glycophorins/metabolism
Humans
Oligosaccharides/*analysis
*Sialoglycoproteins/metabolism
Sugar Alcohols/*analysis
Abstract: Glycophorin A was purified from the erythrocyte membranes of blood group Cad, Sd(a+) and Sd(a-) donors and the oligosaccharide alditols, obtained after alkaline borohydride degradation, separated by h.p.l.c. on an alkylamine silica gel column, were characterized by sugar analysis. Structure determination of the major acid components by methylation analysis, g.l.c.-m.s. and 1H-n.m.r. indicated that the three blood group Cad red cells under study (samples Cad., Bui. and Des.) carry the same pentasaccharide GalNAc(beta 1-4)[NeuAc(alpha 2-3)]Gal(beta 1-3)[NeuAc(alpha 2-6)]GalNAc -ol(Cad determinant) but in different amounts. This pentasaccharide, however, was absent from glycophorin A of Sd(a+) and Sd (a-) donors, suggesting that the Sda determinant is not associated with glycophorins. It was calculated that glycophorin A from the original Cad donor (Cad.) carries about 12 O-glycosidically linked pentasaccharide chains per molecule whereas only 2-3 of these chains were present in the samples from the two other unrelated Cad individuals (Bui. and Des.) It is well known from quantitative agglutination studies that the proportion of red cells which can be agglutinated by the Dolichos biflorus lectin varies from one Cad blood sample to another. Some are completely agglutinated (Cad. donor) whereas others are only partially agglutinated (Bui. and Des. donors) suggesting that some red cells might not carry the Cad determinants. From the results presented above and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis studies it is suggested that Cad red cells from Bui. and Des. do not carry a mixture of glycophorin A molecules with or without the Cad pentasaccharides but a spectrum of glycoprotein molecules with varying amounts of Cad determinants.
Notes: 1470-8728
Blanchard, D
Capon, C
Leroy, Y
Cartron, J P
Fournet, B
Comparative Study
Journal Article
Biochem J. 1985 Dec 15;232(3):813-8. doi: 10.1042/bj2320813.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 452
Author: Blanchard, D., Cartron, J. P., Fournet, B., Montreuil, J., van Halbeek, H. and Vliegenthart, J. F.
Year: 1983
Title: Primary structure of the oligosaccharide determinant of blood group Cad specificity
Journal: J Biol Chem
Volume: 258
Issue: 12
Pages: 7691-5
Epub Date: 1983/06/25
Date: Jun 25
Short Title: Primary structure of the oligosaccharide determinant of blood group Cad specificity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6190803
Keywords: Blood Group Antigens/*genetics
Carbohydrate Conformation
Carbohydrate Sequence
Epitopes/*analysis
Erythrocyte Membrane/*immunology
Erythrocytes/*immunology
Glycophorins/*immunology
Humans
Magnetic Resonance Spectroscopy
Oligosaccharides/*immunology
Sialoglycoproteins/*immunology
Abstract: Glycophorin A and B from Cad erythrocyte membranes are the carriers of the blood group Cad determinants. They are characterized by a significant increase in molecular mass, as compared to the corresponding glycophorins from control erythrocytes (Cartron, J.-P., and Blanchard, D. (1982) Biochem. J. 207, 497-504). Lipid-free glycophorin A, purified from Cad red cells, showed an increased GalNAc content in comparison to blood group B, Cad-negative, control cells. Alkaline-borohydride treatment of this Cad glycophorin A released as a predominant species a pentasaccharide; its structure was determined, by methylation analysis and by 500-MHz 1H-NMR studies, to be: (formula; see text) This novel oligosaccharide inhibited strongly the hemagglutination of Cad erythrocytes by the Dolichos biflorus lectin. It shares with the blood group Sda determinant a terminal GalNAc beta (1 leads to 4)Gal beta (1 leads to .) sequence.
Notes: Blanchard, D
Cartron, J P
Fournet, B
Montreuil, J
van Halbeek, H
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1983 Jun 25;258(12):7691-5.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 413
Author: Blanchard, D., Dahr, W., Hummel, M., Latron, F., Beyreuther, K. and Cartron, J. P.
Year: 1987
Title: Glycophorins B and C from human erythrocyte membranes. Purification and sequence analysis
Journal: J Biol Chem
Volume: 262
Issue: 12
Pages: 5808-11
Epub Date: 1987/04/25
Date: Apr 25
Short Title: Glycophorins B and C from human erythrocyte membranes. Purification and sequence analysis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3571235
Keywords: Amino Acid Sequence
DNA/analysis
Erythrocyte Membrane/analysis
Glycophorins/genetics/*isolation & purification
Humans
Polymorphism, Genetic
Sialoglycoproteins/*isolation & purification
Abstract: We have developed methods for the preparative purification of two sialoglycoproteins (glycophorins B and C) from human erythrocyte membranes by high-performance ion exchange and gel permeation chromatography in the presence of Triton X-100. Glycophorin B was obtained without any detectable contaminants, and glycophorin C exhibited a purity of about 90-95%. The amino acid sequence of the intramembranous domain (residues 36-71) of glycophorin B was determined and found to be similar to that of the hydrophobic region of the major sialoglycoprotein (glycophorin A). The amino acid sequence of the hydrophobic domain (residues 49-88) of glycophorin C, that was also determined, agreed completely with the structure recently deduced from cDNA sequencing.
Notes: Blanchard, D
Dahr, W
Hummel, M
Latron, F
Beyreuther, K
Cartron, J P
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1987 Apr 25;262(12):5808-11.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 349
Author: Blochberger, T. C., Sabatine, J. M., Lee, Y. C. and Hughey, R. P.
Year: 1989
Title: O-linked glycosylation of rat renal gamma-glutamyltranspeptidase adjacent to its membrane anchor domain
Journal: J Biol Chem
Volume: 264
Issue: 34
Pages: 20718-22
Epub Date: 1989/12/05
Date: Dec 5
Short Title: O-linked glycosylation of rat renal gamma-glutamyltranspeptidase adjacent to its membrane anchor domain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2573604
Keywords: Amino Acid Sequence
Animals
Binding Sites
Carbohydrates/analysis
Glycosylation
Hydrolases/genetics
Kidney/*enzymology
Microvilli/*enzymology
Molecular Sequence Data
Papain
Peptide Fragments/isolation & purification
Rats
Wheat Germ Agglutinins
*gamma-Glutamyltransferase/genetics
Abstract: Large domains rich in serine and threonine, that are likely to exhibit clusters of O-linked oligosaccharides, have been reported adjacent to the anchor of several cell surface proteins. No such domain is evident in the primary sequence of rat renal gamma-glutamyltranspeptidase. However, papain treatment of the amphipathic enzyme (Triton-purified gamma-glutamyltranspeptidase, T gamma GT), pretreated with galactose oxidase and NaB3H4 (Frielle, T., and Curthoys, N. P. (1983) Biochemistry 22, 5709-5714), yields the hydrophilic enzyme (papain-treated Triton-purified gamma-glutamyltranspeptidase, PT gamma GT) and a labeled peptide which contains both the amino-terminal membrane anchor and the sequence Pro27-Thr28-Thr29-Ser30. Since [3H]galactose was identified in this peptide, the presence of O-linked oligosaccharides was investigated. Carbohydrate analysis is consistent with the presence of two simple O-linked oligosaccharides on T gamma GT and one on PT gamma GT. Lectin blot analysis of T gamma GT and PT gamma GT was carried out after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The small subunits of both T gamma GT and PT gamma GT and the large amphipathic subunit of T gamma GT all react with the peanut agglutinin lectin, but the large subunit of PT gamma GT exhibits no such reactivity. The reactivity with PNA is consistent with the presence of one oligosaccharide with the structure galactose beta 1-3N-acetylgalactosamine alpha 1-Ser/Thr attached to each subunit of T gamma GT. The papain-sensitivity of the oligosaccharide from the larger subunit is consistent with O-glycosylation at the Thr28-Thr29-Ser30 sequence. The results of lectin blot analysis with wheat germ agglutinin imply that the content of N-linked oligosaccharides is unaffected by papain treatment of the transpeptidase. These data represent the first direct evidence for O-glycosylation of a microvillar hydrolase at a site immediately adjacent to the membrane anchor and indicates that even small clusters of Thr and Ser can be O-glycosylated. Isolated O-linked oligosaccharides may have functional significance since single Ser and Thr residues are consistently found near the membrane anchor of many cell surface proteins.
Notes: Blochberger, T C
Sabatine, J M
Lee, Y C
Hughey, R P
DK-09970/DK/NIDDK NIH HHS/United States
DK-33211/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1989 Dec 5;264(34):20718-22.
Author Address: Department of Microbiology, University of Pittsburgh School of Medicine, Pennsylvania 15261.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 404
Author: Blumenfeld, O. O., Adamany, A. M., Kikuchi, M., Sabo, B. and McCreary, J.
Year: 1986
Title: Membrane glycophorins in Sta blood group erythrocytes
Journal: J Biol Chem
Volume: 261
Issue: 12
Pages: 5544-52
Epub Date: 1986/04/25
Date: Apr 25
Short Title: Membrane glycophorins in Sta blood group erythrocytes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3514617
Keywords: Amino Acids/analysis
Antibodies, Monoclonal
Carbohydrates/analysis
Chromatography, Affinity
Chromatography, Gel
Cyanogen Bromide/pharmacology
Electrophoresis, Polyacrylamide Gel
Erythrocyte Membrane/*analysis
Glycophorins/*analysis
Homozygote
Humans
Immunosorbent Techniques
*MNSs Blood-Group System
Oligosaccharides/analysis
Peptide Fragments/analysis
Phenotype
Sialoglycoproteins/*analysis
Abstract: Structural and immunochemical studies of glycophorins isolated from erythrocytes of an individual homozygous for the M Sta blood group phenotype are described. Reactivities with specific monoclonal antibodies indicated that two major M and N glycophorins were present. The M and N Sta glycophorins were resolved by Lens culinaris lectin affinity chromatography. The N species was not held on the lectin but the M species, like control alpha glycophorins, was retained and could be eluted with alpha-methylmannoside. The two proteins were present in almost equimolar amounts. Studies of the CNBr fragments provided evidence that the structure of M Sta glycophorin is the same as that of the usual M alpha glycophorin but that the N Sta glycophorin is a variant. The amino-terminal octapeptides of the M and N species were similar in amino acid and carbohydrate composition to those isolated, respectively, from M and N alpha glycophorins. The studies focused on CNBr glycopeptide B that, in control alpha glycophorins, extends from amino acid residues 9 to 81. The fragment from the M species exhibited properties identical to those of the corresponding fragment of control alpha glycophorins in terms of size, chromatographic behavior, amino acid and carbohydrate contents and compositions, the presence of O-glycosidically linked saccharides and a single Asn-linked carbohydrate unit. The structures of the O-linked units were inferred experimentally to be NeuAc(alpha 2,3)Gal-(beta 1,3)GalNAc and NeuAc(alpha 2,3)Gal(beta 1,3) [NeuAc(alpha 2,6)]GalNAc, present in a ratio similar to that found in controls; and the Asn-linked unit also appeared to be as in the control. The tryptic glycopeptide pattern of the M Sta glycophorin CNBr fragment B was identical to the pattern of the corresponding control fragment, and the composition of the tryptic peptides suggested sequence identity with the control fragment. In contrast, the N Sta glycophorin yielded two CNBr glycopeptides B; both contained fewer amino acid residues and virtually lacked Man and GlcNAc, indicating the absence of the Asn-linked carbohydrate. The much decreased levels of these carbohydrates in the intact N protein, corroborated the latter finding. The O-glycosidic saccharides appeared similar to those found in control alpha glycophorins. However, the tryptic glycopeptide pattern of the variant differed from control M or N alpha glycophorins, suggesting a deletion of a large segment of the molecule near residues 40-61 and/or a substitution of methionine for a residue upstream from residue 40.(ABSTRACT TRUNCATED AT 400 WORDS)
Notes: Blumenfeld, O O
Adamany, A M
Kikuchi, M
Sabo, B
McCreary, J
16389/PHS HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1986 Apr 25;261(12):5544-52.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 237
Author: Blumenfeld, O. O., Lalezari, P., Khorshidi, M., Puglia, K. and Fukuda, M.
Year: 1992
Title: O-linked oligosaccharides of glycophorins A and B in erythrocytes of two individuals with the Tn polyagglutinability syndrome
Journal: Blood
Volume: 80
Issue: 9
Pages: 2388-95
Epub Date: 1992/11/01
Date: Nov 1
Short Title: O-linked oligosaccharides of glycophorins A and B in erythrocytes of two individuals with the Tn polyagglutinability syndrome
Alternate Journal: Blood
ISSN: 0006-4971 (Print)
0006-4971
Accession Number: 1421410
Keywords: Antibodies, Monoclonal
Carbohydrate Sequence
Chromatography, Gel
Electrophoresis, Polyacrylamide Gel
Erythrocytes/chemistry/*physiology
Glycophorins/*chemistry/isolation & purification
*Hemagglutination
Hematologic Diseases/*blood
Humans
Molecular Sequence Data
Oligosaccharides/blood/*chemistry/isolation & purification
Syndrome
Abstract: Tn polyagglutinability syndrome is an acquired condition where erythrocytes express Tn neo-antigen and become susceptible to hemagglutination by the naturally occurring anti-Tn present in normal sera. Early studies had indicated that O-linked N-acetyl galactosamine was the sole serologic Tn determinant, but more recently O-linked NeuNAc alpha 2, 6GalNAc also has been implicated as a Tn antigen (sialosyl-Tn). However, none of these studies were performed on purified glycoproteins. In this report we examine oligosaccharides of glycophorins A and B purified from Tn erythrocytes of two affected individuals to establish how N- and O-linked saccharides differ from normal. Analysis of carbohydrate composition and treatment with N-glycanase showed that the Asn-linked unit of glycophorin A was not affected. O-linked oligosaccharides were obtained by beta-elimination in the presence of tritiated sodium borohydride. The reduced radiolabeled products were fractionated by Bio-Gel P-2 chromatography, and their structures were investigated by comparison with standards, by monosaccharide quantification, and by neuraminidases of known specificities. The results show that Tn glycophorins from both donors contain intact and truncated forms of trisaccharide and tetrasaccharide NeuNAc alpha 2,3Gal beta 1,3GalNAc and NeuNAc alpha 2,3Gal beta 1,3- (NeuNAc alpha 2,6)GalNAc usually present in glycophorins A and B. The truncated forms include the protein O-linked monosaccharide, GalNAc and disaccharide, NeuNAc alpha 2,6GalNAc (major isomer). The presence of intact glycans in the total population of Tn erythrocytes was confirmed by their susceptibility to T activation after treatment with neuraminidase. The proportion of the four species was not identical in glycophorins of these two donors but, in both, the truncated units predominated and the amount of the disaccharide was approximately one half of that of the monosaccharide. The data are consistent with alterations in UDPGal:GalNAc beta 1,3galactosyl transferase that may have multiple molecular origins and with induction of a specific GalNAc protein alpha 2,6 sialosyl transferase in Tn hematopoietic precursor cells. The molecular basis for these alterations awaits further study.
Notes: Blumenfeld, O O
Lalezari, P
Khorshidi, M
Puglia, K
Fukuda, M
CA 33000/CA/NCI NIH HHS/United States
R01 GM16389/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Blood. 1992 Nov 1;80(9):2388-95.
Author Address: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 385
Author: Blumenfeld, O. O., Smith, A. J. and Moulds, J. J.
Year: 1987
Title: Membrane glycophorins of Dantu blood group erythrocytes
Journal: J Biol Chem
Volume: 262
Issue: 24
Pages: 11864-70
Epub Date: 1987/08/25
Date: Aug 25
Short Title: Membrane glycophorins of Dantu blood group erythrocytes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3305497
Keywords: Amino Acid Sequence
*Blood Group Antigens
Chromatography, Affinity
Chromatography, Gel
Erythrocyte Membrane/*analysis
Glycophorins/*blood
Hexosamines/analysis
Humans
Immunosorbent Techniques
Sialic Acids/analysis
Sialoglycoproteins/*blood
Abstract: Glycophorins of erythrocytes of two unrelated individuals who exhibit the Dantu blood group phenotype were studied. Immunoblots indicated that erythrocytes of each individual contained a complement of a normal alpha-glycophorin (glycophorin A) and a variant N-glycophorin. delta-Glycophorin (glycophorin B) was present in one donor's cells but not the other's; the s and N phenotypes of the latter's erythrocytes may derive from the variant glycophorin. The variant glycophorin is of a smaller size, does not bind to Lens culinaris lectin agarose, and lacks residues approximately 40-60 of alpha-glycophorin and its single asparagine-linked carbohydrate; it contains approximately 2 less O-glycosidically bound units whose structures are identical to those found in alpha-glycophorins. All these properties are characteristic of delta-glycophorin. The variant is related to alpha-glycophorin in the carboxyl-terminal region as shown by reaction with a specific antiserum. Sequence analyses of a mixture of chymotryptic peptides of a CNBr fragment of the variant glycophorin identified the sequence Val-His-Arg-Phe-Thr-Val-Pro-Glu-Ile-Thr-Leu-Ile-Ile that contains the junction point of delta- and alpha-glycophorins spanning residues 33-38/39 of delta-glycophorin and residues 71/72-77 of alpha-glycophorin. Sequence analysis of a mixture of CNBr fragments allowed us to conclude that the variant originates from delta-s- rather than delta-S-glycophorin. The quantity of the variant Dantu glycophorin when compared to alpha-glycophorin differed in the two individuals, the ratio being 2/1 in one individual's cells and 0.5/1 in the other's. This may reflect that the two donors belong to different varieties of Dantu phenotypes. Together, the evidence indicates that both donors' erythrocytes contain a (delta-alpha) variant glycophorin, whose amino terminus originates from delta-s-glycophorin and the carboxyl end from alpha-glycophorin with a junction point around residues 39 of delta- and 71 of alpha-glycophorins. The results suggest that the unique junction region may be characteristic of the Dantu phenotype.
Notes: Blumenfeld, O O
Smith, A J
Moulds, J J
GM 16389/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1987 Aug 25;262(24):11864-70.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1128
Author: Boassa, D., Ambrosi, C., Qiu, F., Dahl, G., Gaietta, G. and Sosinsky, G.
Year: 2007
Title: Pannexin1 channels contain a glycosylation site that targets the hexamer to the plasma membrane
Journal: J Biol Chem
Volume: 282
Issue: 43
Pages: 31733-43
Epub Date: 2007/08/24
Date: Oct 26
Short Title: Pannexin1 channels contain a glycosylation site that targets the hexamer to the plasma membrane
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M702422200
Accession Number: 17715132
Keywords: Asparagine/metabolism
Biotinylation
Cell Line
Cell Membrane/*metabolism/ultrastructure
Connexins/*chemistry/genetics/*metabolism/ultrastructure
Cross-Linking Reagents/pharmacology
Dose-Response Relationship, Drug
Electrophysiology
Glycosylation
Humans
Immunohistochemistry
Ion Channels/*ultrastructure
Kidney/cytology
Models, Biological
Mutation
Nerve Tissue Proteins
Protein Processing, Post-Translational
Protein Structure, Secondary
Succinimides/pharmacology
Abstract: Pannexins are newly discovered channel proteins expressed in many different tissues and abundantly in the vertebrate central nervous system. Based on membrane topology, folding and secondary structure prediction, pannexins are proposed to form gap junction-like structures. We show here that Pannexin1 forms a hexameric channel and reaches the cell surface but, unlike connexins, is N-glycosylated. Using site-directed mutagenesis we analyzed three putative N-linked glycosylation sites and examined the effects of each mutation on channel expression. We show for the first time that Pannexin1 is glycosylated at Asn-254 and that this residue is important for plasma membrane targeting. The glycosylation of Pannexin1 at its extracellular surface makes it unlikely that two oligomers could dock to form an intercellular channel. Ultrastructural analysis by electron microscopy confirmed that Pannexin1 junctional areas do not appear as canonical gap junctions. Rather, Pannexin1 channels are distributed throughout the plasma membrane. We propose that N-glycosylation of Pannexin1 could be a significant mechanism for regulating the trafficking of these membrane proteins to the cell surface in different tissues.
Notes: Boassa, Daniela
Ambrosi, Cinzia
Qiu, Feng
Dahl, Gerhard
Gaietta, Guido
Sosinsky, Gina
RR04050/RR/NCRR NIH HHS/United States
P41 RR004050/RR/NCRR NIH HHS/United States
R01 GM072881/GM/NIGMS NIH HHS/United States
GM072881/GM/NIGMS NIH HHS/United States
P41 RR002250/RR/NCRR NIH HHS/United States
R01 GM048610-14/GM/NIGMS NIH HHS/United States
GM065937/GM/NIGMS NIH HHS/United States
GM48610/GM/NIGMS NIH HHS/United States
R01 GM048610/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Biol Chem. 2007 Oct 26;282(43):31733-43. doi: 10.1074/jbc.M702422200. Epub 2007 Aug 22.
Author Address: National Center for Microscopy and Imaging Research, Center for Research in Biological Systems, Department of Neurosciences, University of California, San Diego, La Jolla, California 92093-0608, USA. dboassa@gmail.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 419
Author: Bock, S. C., Skriver, K., Nielsen, E., Thøgersen, H. C., Wiman, B., Donaldson, V. H., Eddy, R. L., Marrinan, J., Radziejewska, E., Huber, R. and et al.
Year: 1986
Title: Human C1 inhibitor: primary structure, cDNA cloning, and chromosomal localization
Journal: Biochemistry
Volume: 25
Issue: 15
Pages: 4292-301
Epub Date: 1986/07/29
Date: Jul 29
Short Title: Human C1 inhibitor: primary structure, cDNA cloning, and chromosomal localization
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00363a018
Accession Number: 3756141
Keywords: Amino Acid Sequence
Base Sequence
Carbohydrates/analysis
Chromosome Mapping
*Chromosomes, Human, 6-12 and X
*Cloning, Molecular
Complement C1 Inactivator Proteins/*genetics/isolation & purification
DNA/*metabolism
*Genes
Humans
Hybrid Cells/cytology
Models, Molecular
Nucleic Acid Hybridization
Polymorphism, Genetic
Protein Conformation
Abstract: The primary structure of human C1 inhibitor was determined by peptide and DNA sequencing. The single-chain polypeptide moiety of the intact inhibitor is 478 residues (52,869 Da), accounting for only 51% of the apparent molecular mass of the circulating protein (104,000 Da). The positions of six glucosamine-based and five galactosamine-based oligosaccharides were determined. Another nine threonine residues are probably also glycosylated. Most of the carbohydrate prosthetic groups (probably 17) are located at the amino-terminal end (residues 1-120) of the protein and are particularly concentrated in a region where the tetrapeptide sequence Glx-Pro-Thr-Thr, and variants thereof, is repeated 7 times. No phosphate was detected in C1 inhibitor. Two disulfide bridges connect cysteine-101 to cysteine-406 and cysteine-108 to cysteine-183. Comparison of the amino acid and cDNA sequences indicates that secretion is mediated by a 22-residue signal peptide and that further proteolytic processing does not occur. C1 inhibitor is a member of the large serine protease inhibitor (serpin) gene family. The homology concerns residues 120 through the C-terminus. The sequence was compared with those of nine other serpins, and conserved and nonconserved regions correlated with elements in the tertiary structure of alpha 1-antitrypsin. The C1 inhibitor gene maps to chromosome 11, p11.2-q13. C1 inhibitor genes of patients from four hereditary angioneurotic edema kindreds do not have obvious deletions or rearrangements in the C1 inhibitor locus. A HgiAI DNA polymorphism, identified following the observation of sequence variants, will be useful as a linkage marker in studies of mutant C1 inhibitor genes.
Notes: Bock, S C
Skriver, K
Nielsen, E
Thøgersen, H C
Wiman, B
Donaldson, V H
Eddy, R L
Marrinan, J
Radziejewska, E
Huber, R
HL-15690/HL/NHLBI NIH HHS/United States
HL-16238/HL/NHLBI NIH HHS/United States
HL-30712/HL/NHLBI NIH HHS/United States
etc.
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1986 Jul 29;25(15):4292-301. doi: 10.1021/bi00363a018.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 549
Author: Bodian, D. L., Jones, E. Y., Harlos, K., Stuart, D. I. and Davis, S. J.
Year: 1994
Title: Crystal structure of the extracellular region of the human cell adhesion molecule CD2 at 2.5 A resolution
Journal: Structure
Volume: 2
Issue: 8
Pages: 755-66
Epub Date: 1994/08/15
Date: Aug 15
Short Title: Crystal structure of the extracellular region of the human cell adhesion molecule CD2 at 2.5 A resolution
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/s0969-2126(94)00076-x
Accession Number: 7994575
Keywords: Amino Acid Sequence
Animals
Binding Sites
CD2 Antigens/*chemistry
Cell Adhesion Molecules/*chemistry
Crystallography, X-Ray
Glycoproteins/*chemistry
Humans
Immunoglobulins/chemistry
Ligands
Models, Molecular
Molecular Sequence Data
Protein Conformation
Rats
Recombinant Proteins/chemistry
Sequence Homology, Amino Acid
Solubility
T-Lymphocytes/chemistry
Abstract: BACKGROUND: The T-lymphocyte antigen CD2 is an adhesion molecule implicated in immune responses in vivo. The extracellular regions of the human and rat homologues of CD2 share only 45% sequence identity and bind different protein ligands. Comparison of the human and rat soluble CD2 (sCD2) structures should provide insights into the structural basis of cell surface recognition. RESULTS: We therefore determined the crystal structure of a form of human sCD2 with single N-acetylglucosamine residues at each glycosylation site to 2.5 A resolution with an R-factor of 19.3%. It is composed of two immunoglobulin superfamily domains similar to those of rat sCD2, but the relative orientation of the domains in the two homologues differs by up to 20 degrees. An interaction involving the flat, highly charged, ligand binding GFCC'C" faces of crystallographically related human sCD2 molecules duplicates, in a different lattice, that observed in the rat sCD2 crystals. CONCLUSIONS: Intramolecular flexibility appears to be a conserved feature of CD2. The head-to-head interaction between molecules represents a general model for interactions between adhesion molecules of this structural class. Ligand specificity may be influenced by the distribution of charged residues on the binding face.
Notes: Bodian, D L
Jones, E Y
Harlos, K
Stuart, D I
Davis, S J
Journal Article
Research Support, Non-U.S. Gov't
United States
Structure. 1994 Aug 15;2(8):755-66. doi: 10.1016/s0969-2126(94)00076-x.
Author Address: Laboratory of Molecular Biophysics, Oxford, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1525
Author: Boersema, P. J., Geiger, T., Wisniewski, J. R. and Mann, M.
Year: 2013
Title: Quantification of the N-glycosylated secretome by super-SILAC during breast cancer progression and in human blood samples
Journal: Mol Cell Proteomics
Volume: 12
Issue: 1
Pages: 158-71
Epub Date: 2012/10/24
Date: Jan
Short Title: Quantification of the N-glycosylated secretome by super-SILAC during breast cancer progression and in human blood samples
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M112.023614
PMCID: PMC3536897
Accession Number: 23090970
Keywords: Biomarkers, Tumor/blood
Breast Neoplasms/*blood
Cell Line, Tumor
Cell Membrane
Culture Media, Conditioned
Disease Progression
Female
Glycopeptides/*blood
Glycoproteins/*blood
Glycosylation
Humans
Isotope Labeling
Neoplasm Proteins/*blood/metabolism
Proteomics
Tandem Mass Spectrometry
Abstract: Cells secrete a large number of proteins to communicate with their surroundings. Furthermore, plasma membrane proteins and intracellular proteins can be released into the extracellular space by regulated or non-regulated processes. Here, we profiled the supernatant of 11 cell lines that are representative of different stages of breast cancer development by specifically capturing N-glycosylated peptides using the N-glyco FASP technology. For accurate quantification we developed a super-SILAC mix from several labeled breast cancer cell lines and used it as an internal standard for all samples. In total, 1398 unique N-glycosylation sites were identified and quantified. Enriching for N-glycosylated peptides focused the analysis on classically secreted and membrane proteins. N-glycosylated secretome profiles correctly clustered the different cell lines to their respective cancer stage, suggesting that biologically relevant differences were detected. Five different profiles of glycoprotein dynamics during cancer development were detected, and they contained several proteins with known roles in breast cancer. We then used the super-SILAC mix in plasma, which led to the quantification of a large number of the previously identified N-glycopeptides in this important body fluid. The combination of quantifying the secretome of cancer cell lines and of human plasma with a super-SILAC approach appears to be a promising new approach for finding markers of disease.
Notes: 1535-9484
Boersema, Paul J
Geiger, Tamar
Wisniewski, Jacek R
Mann, Matthias
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2013 Jan;12(1):158-71. doi: 10.1074/mcp.M112.023614. Epub 2012 Oct 22.
Author Address: Department of Proteomics and Signal Transduction, Max Planck Institute of Biochemistry, Am Klopferspitz 18, D-82152 Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 889
Author: Boja, E. S., Hoodbhoy, T., Fales, H. M. and Dean, J.
Year: 2003
Title: Structural characterization of native mouse zona pellucida proteins using mass spectrometry
Journal: J Biol Chem
Volume: 278
Issue: 36
Pages: 34189-202
Epub Date: 2003/06/12
Date: Sep 5
Short Title: Structural characterization of native mouse zona pellucida proteins using mass spectrometry
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M304026200
Accession Number: 12799386
Keywords: Amino Acid Motifs
Amino Acid Sequence
Animals
Blotting, Western
Carbohydrates/chemistry
Cell Membrane/metabolism
Chromatography, Liquid
DNA, Complementary/metabolism
Disulfides
Egg Proteins/*chemistry
Electrophoresis, Polyacrylamide Gel
Glycosylation
Mass Spectrometry
Membrane Glycoproteins/*chemistry
Mice
Molecular Sequence Data
Peptides/chemistry
Protein Binding
Protein Conformation
Protein Processing, Post-Translational
Protein Sorting Signals
Protein Structure, Tertiary
*Receptors, Cell Surface
Spectrometry, Mass, Electrospray Ionization
Zona Pellucida/*metabolism
Zona Pellucida Glycoproteins
Abstract: The zona pellucida is an extracellular matrix consisting of three glycoproteins that surrounds mammalian eggs and mediates fertilization. The primary structures of mouse ZP1, ZP2, and ZP3 have been deduced from cDNA. Each has a predicted signal peptide and a transmembrane domain from which an ectodomain must be released. All three zona proteins undergo extensive co- and post-translational modifications important for secretion and assembly of the zona matrix. In this report, native zonae pellucidae were isolated and structural features of individual zona proteins within the mixture were determined by high resolution electrospray mass spectrometry. Complete coverage of the primary structure of native ZP3, 96% of ZP2, and 56% of ZP1, the least abundant zona protein, was obtained. Partial disulfide bond assignments were made for each zona protein, and the size of the processed, native protein was determined. The N termini of ZP1 and ZP3, but not ZP2, were blocked by cyclization of glutamine to pyroglutamate. The C termini of ZP1, ZP2, and ZP3 lie upstream of a dibasic motif, which is part of, but distinct from, a proprotein convertase cleavage site. The zona proteins are highly glycosylated and 4/4 potential N-linkage sites on ZP1, 6/6 on ZP2, and 5/6 on ZP3 are occupied. Potential O-linked carbohydrate sites are more ubiquitous, but less utilized.
Notes: Boja, Emily S
Hoodbhoy, Tanya
Fales, Henry M
Dean, Jurrien
Journal Article
United States
J Biol Chem. 2003 Sep 5;278(36):34189-202. doi: 10.1074/jbc.M304026200. Epub 2003 Jun 10.
Author Address: Laboratory of Biophysical Chemistry, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA. bojae@nhlbi.nih.gov
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1020
Author: Boja, E. S., Hoodbhoy, T., Garfield, M. and Fales, H. M.
Year: 2005
Title: Structural conservation of mouse and rat zona pellucida glycoproteins. Probing the native rat zona pellucida proteome by mass spectrometry
Journal: Biochemistry
Volume: 44
Issue: 50
Pages: 16445-60
Epub Date: 2005/12/14
Date: Dec 20
Short Title: Structural conservation of mouse and rat zona pellucida glycoproteins. Probing the native rat zona pellucida proteome by mass spectrometry
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi051883f
Accession Number: 16342937
Keywords: Amino Acid Sequence
Animals
Glycoproteins/*chemistry
Glycosylation
Mass Spectrometry/*methods
Mice
Molecular Sequence Data
Protein Conformation
*Proteome
Rats
Sequence Homology, Amino Acid
Zona Pellucida/*chemistry
Abstract: The mammalian zona pellucida is an egg extracellular matrix to which sperm bind. Mouse zonae are composed of three glycoproteins (ZP1, ZP2, and ZP3), while rat zonae contain four (ZP1, ZP2, ZP3, and ZP4/ZPB). Mouse sperm bind to zonae comprised solely of mouse ZP2 and ZP3. In this report, we show that rat sperm also bind to these zonae, indicating that ZP2 and ZP3 contain a "minimum structure(s)" to which rodent sperm can bind, and ZP1 and ZP4/ZPB are dispensable in these two rodents. These data are consistent with our mass spectrometric analysis of the native rat zona pellucida proteome (defined as the fraction of the total rat proteome to which the zonae glycoproteins contribute) demonstrating that the rat zonae glycoproteins share a high degree of conservation of structural features with respect to their mouse counterparts. The primary sequences of the rat zonae proteins have been deduced from cDNA. Each zona protein undergoes extensive co- and post-translational modification prior to its secretion and incorporation into an extracellular zona matrix. Each has a predicted N-terminal signal peptide that is cleaved off once protein translation begins and an anchoring C-terminal transmembrane domain from which the mature protein is released. Mass spectrometric analysis with a limited amount of native material allowed determination of the mature N-termini of rat ZP1 and ZP3, both of which are characterized by cyclization of glutamine to pyroglutamate; the N-terminus of ZP2 was identified by Edman degradation. The mature C-termini of ZP1 and ZP3 end two amino acids upstream of a conserved dibasic residue that is part of, but distinct from, the consensus furin cleavage sequence, while the C-terminus of ZP2 was not determined. Each zona protein contains a "zona domain" with eight conserved cysteine residues that is thought to play a role in the polymerization of the zona proteins into matrix filaments. Partial disulfide bond assignment indicates that the intramolecular disulfide patterns in rat ZP1, ZP2, and ZP3 are identical to those of their corresponding mouse counterparts. Last, nearly all potential N-glycosylation sites are occupied in the rat zonae glycoproteins (three of three for ZP1, six or seven of seven for ZP2, and four or five of six for ZP3). In comparison, potential O-glycosylation sites are numerous (59-83 Ser/Thr residues), but only two regions were observed to carry O-glycans in rat ZP3.
Notes: Boja, Emily S
Hoodbhoy, Tanya
Garfield, Mark
Fales, Henry M
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
United States
Biochemistry. 2005 Dec 20;44(50):16445-60. doi: 10.1021/bi051883f.
Author Address: Laboratory of Applied Mass Spectrometry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1753
Author: Bokhove, M., Nishimura, K., Brunati, M., Han, L., de Sanctis, D., Rampoldi, L. and Jovine, L.
Year: 2016
Title: A structured interdomain linker directs self-polymerization of human uromodulin
Journal: Proc Natl Acad Sci U S A
Volume: 113
Issue: 6
Pages: 1552-7
Epub Date: 2016/01/27
Date: Feb 9
Short Title: A structured interdomain linker directs self-polymerization of human uromodulin
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1519803113
PMCID: PMC4760807
Accession Number: 26811476
Keywords: Amino Acid Sequence
Animals
Blotting, Western
Crystallography, X-Ray
Disulfides/metabolism
Dogs
Extracellular Matrix Proteins/genetics
Fluorescent Antibody Technique
GPI-Linked Proteins/genetics
HEK293 Cells
Humans
Madin Darby Canine Kidney Cells
Maltose-Binding Proteins/metabolism
Mice
Models, Molecular
Molecular Sequence Data
Mutation, Missense/genetics
*Polymerization
Protein Multimerization
Protein Structure, Tertiary
Recombinant Fusion Proteins/metabolism
Sequence Alignment
Structural Homology, Protein
Uromodulin/*chemistry/ultrastructure
X-ray crystallography
Zp2
polymerization
uromodulin
zona pellucida domain
Abstract: Uromodulin (UMOD)/Tamm-Horsfall protein, the most abundant human urinary protein, plays a key role in chronic kidney diseases and is a promising therapeutic target for hypertension. Via its bipartite zona pellucida module (ZP-N/ZP-C), UMOD forms extracellular filaments that regulate kidney electrolyte balance and innate immunity, as well as protect against renal stones. Moreover, salt-dependent aggregation of UMOD filaments in the urine generates a soluble molecular net that captures uropathogenic bacteria and facilitates their clearance. Despite the functional importance of its homopolymers, no structural information is available on UMOD and how it self-assembles into filaments. Here, we report the crystal structures of polymerization regions of human UMOD and mouse ZP2, an essential sperm receptor protein that is structurally related to UMOD but forms heteropolymers. The structure of UMOD reveals that an extensive hydrophobic interface mediates ZP-N domain homodimerization. This arrangement is required for filament formation and is directed by an ordered ZP-N/ZP-C linker that is not observed in ZP2 but is conserved in the sequence of deafness/Crohn's disease-associated homopolymeric glycoproteins α-tectorin (TECTA) and glycoprotein 2 (GP2). Our data provide an example of how interdomain linker plasticity can modulate the function of structurally similar multidomain proteins. Moreover, the architecture of UMOD rationalizes numerous pathogenic mutations in both UMOD and TECTA genes.
Notes: 1091-6490
Bokhove, Marcel
Nishimura, Kaoru
Brunati, Martina
Han, Ling
de Sanctis, Daniele
Rampoldi, Luca
Jovine, Luca
Orcid: 0000-0002-2679-6946
260759/European Research Council/International
GGP14263/Telethon/Italy
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2016 Feb 9;113(6):1552-7. doi: 10.1073/pnas.1519803113. Epub 2016 Jan 25.
Author Address: Department of Biosciences and Nutrition & Center for Innovative Medicine, Karolinska Institutet, SE-141 83 Huddinge, Sweden;
Molecular Genetics of Renal Disorders Unit, Division of Genetics and Cell Biology, San Raffaele Scientific Institute, I-20132 Milan, Italy;
European Synchrotron Radiation Facility - The European Synchrotron, Grenoble 38000, France.
Department of Biosciences and Nutrition & Center for Innovative Medicine, Karolinska Institutet, SE-141 83 Huddinge, Sweden; luca.jovine@ki.se.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1911
Author: Bollineni, R. C., Koehler, C. J., Gislefoss, R. E., Anonsen, J. H. and Thiede, B.
Year: 2018
Title: Large-scale intact glycopeptide identification by Mascot database search
Journal: Sci Rep
Volume: 8
Issue: 1
Pages: 2117
Epub Date: 2018/02/03
Date: Feb 1
Short Title: Large-scale intact glycopeptide identification by Mascot database search
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/s41598-018-20331-2
PMCID: PMC5795011
Accession Number: 29391424
Keywords: Case-Control Studies
*Databases, Factual
Glycopeptides/*blood
Glycoproteins/*blood
Glycosylation
Humans
Male
Prostatic Neoplasms/*blood
*Search Engine
Abstract: Workflows capable of determining glycopeptides in large-scale are missing in the field of glycoproteomics. We present an approach for automated annotation of intact glycopeptide mass spectra. The steps in adopting the Mascot search engine for intact glycopeptide analysis included: (i) assigning one letter codes for monosaccharides, (ii) linearizing glycan sequences and (iii) preparing custom glycoprotein databases. Automated annotation of both N- and O-linked glycopeptides was proven using standard glycoproteins. In a large-scale study, a total of 257 glycoproteins containing 970 unique glycosylation sites and 3447 non-redundant N-linked glycopeptide variants were identified in 24 serum samples. Thus, a single tool was developed that collectively allows the (i) elucidation of N- and O-linked glycopeptide spectra, (ii) matching glycopeptides to known protein sequences, and (iii) high-throughput, batch-wise analysis of large-scale glycoproteomics data sets.
Notes: 2045-2322
Bollineni, Ravi Chand
Koehler, Christian Jeffrey
Gislefoss, Randi Elin
Anonsen, Jan Haug
Thiede, Bernd
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2018 Feb 1;8(1):2117. doi: 10.1038/s41598-018-20331-2.
Author Address: Department of Biosciences, University of Oslo, Oslo, Norway.
Cancer Registry of Norway, Institute of Population-based Cancer Research, Oslo, Norway.
Department of Biosciences, University of Oslo, Oslo, Norway. bernd.thiede@ibv.uio.no.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1116
Author: Borg, N. A., Wun, K. S., Kjer-Nielsen, L., Wilce, M. C., Pellicci, D. G., Koh, R., Besra, G. S., Bharadwaj, M., Godfrey, D. I., McCluskey, J. and Rossjohn, J.
Year: 2007
Title: CD1d-lipid-antigen recognition by the semi-invariant NKT T-cell receptor
Journal: Nature
Volume: 448
Issue: 7149
Pages: 44-9
Epub Date: 2007/06/22
Date: Jul 5
Short Title: CD1d-lipid-antigen recognition by the semi-invariant NKT T-cell receptor
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature05907
Accession Number: 17581592
Keywords: Animals
Antigen Presentation
Antigens, CD1/chemistry/*immunology
Antigens, CD1d
Carbohydrate Conformation
Crystallography, X-Ray
Galactosylceramides/chemistry/immunology
Humans
Killer Cells, Natural/*immunology
Mice
Protein Conformation
Receptors, Antigen, T-Cell, alpha-beta/chemistry/*immunology
Species Specificity
T-Lymphocyte Subsets/immunology
T-Lymphocytes/*immunology
Abstract: The CD1 family is a large cluster of non-polymorphic, major histocompatibility complex (MHC) class-I-like molecules that bind distinct lipid-based antigens that are recognized by T cells. The most studied group of T cells that interact with lipid antigens are natural killer T (NKT) cells, which characteristically express a semi-invariant T-cell receptor (NKT TCR) that specifically recognizes the CD1 family member, CD1d. NKT-cell-mediated recognition of the CD1d-antigen complex has been implicated in microbial immunity, tumour immunity, autoimmunity and allergy. Here we describe the structure of a human NKT TCR in complex with CD1d bound to the potent NKT-cell agonist alpha-galactosylceramide, the archetypal CD1d-restricted glycolipid. In contrast to T-cell receptor-peptide-antigen-MHC complexes, the NKT TCR docked parallel to, and at the extreme end of the CD1d-binding cleft, which enables a lock-and-key type interaction with the lipid antigen. The structure provides a basis for the interaction between the highly conserved NKT TCR alpha-chain and the CD1d-antigen complex that is typified in innate immunity, and also indicates how variability of the NKT TCR beta-chain can impact on recognition of other CD1d-antigen complexes. These findings provide direct insight into how a T-cell receptor recognizes a lipid-antigen-presenting molecule of the immune system.
Notes: 1476-4687
Borg, Natalie A
Wun, Kwok S
Kjer-Nielsen, Lars
Wilce, Matthew C J
Pellicci, Daniel G
Koh, Ruide
Besra, Gurdyal S
Bharadwaj, Mandvi
Godfrey, Dale I
McCluskey, James
Rossjohn, Jamie
G0400421/Medical Research Council/United Kingdom
Wellcome Trust/United Kingdom
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
England
Nature. 2007 Jul 5;448(7149):44-9. doi: 10.1038/nature05907. Epub 2007 Jun 20.
Author Address: The Protein Crystallography Unit, ARC Centre of Excellence in Structural and Functional Microbial Genomics, Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1607
Author: Borodkin, V. S., Schimpl, M., Gundogdu, M., Rafie, K., Dorfmueller, H. C., Robinson, D. A. and van Aalten, D. M.
Year: 2014
Title: Bisubstrate UDP-peptide conjugates as human O-GlcNAc transferase inhibitors
Journal: Biochem J
Volume: 457
Issue: 3
Pages: 497-502
Epub Date: 2013/11/22
Date: Feb 1
Short Title: Bisubstrate UDP-peptide conjugates as human O-GlcNAc transferase inhibitors
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20131272
PMCID: PMC3927924
Accession Number: 24256146
Keywords: Adaptor Proteins, Signal Transducing/chemistry/genetics/metabolism
Binding Sites
Crystallography, X-Ray
*Drug Design
Enzyme Inhibitors/chemical synthesis/chemistry/metabolism/*pharmacology
Glycosylation/drug effects
Humans
Interferometry
Kinetics
*Models, Molecular
N-Acetylglucosaminyltransferases/*antagonists & inhibitors/chemistry/metabolism
Oligopeptides/chemical synthesis/chemistry/metabolism/*pharmacology
Peptide Fragments/chemistry/genetics/metabolism
Protein Conformation
Protein Processing, Post-Translational/drug effects
Recombinant Proteins/chemistry/metabolism
Serine/chemistry
Uridine Diphosphate/*analogs & derivatives/chemistry/metabolism/pharmacology
Abstract: Inhibitors of OGT (O-GlcNAc transferase) are valuable tools to study the cell biology of protein O-GlcNAcylation. We report OGT bisubstrate-linked inhibitors (goblins) in which the acceptor serine in the peptide VTPVSTA is covalently linked to UDP, eliminating the GlcNAc pyranoside ring. Goblin1 co-crystallizes with OGT, revealing an ordered C₃ linker and retained substrate-binding modes, and binds the enzyme with micromolar affinity, inhibiting glycosyltransfer on to protein and peptide substrates.
Notes: 1470-8728
Borodkin, Vladimir S
Schimpl, Marianne
Gundogdu, Mehmet
Rafie, Karim
Dorfmueller, Helge C
Robinson, David A
van Aalten, Daan M F
087590/Wellcome Trust/United Kingdom
G0900138/Medical Research Council/United Kingdom
WT087590MA/Wellcome Trust/United Kingdom
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2014 Feb 1;457(3):497-502. doi: 10.1042/BJ20131272.
Author Address: *MRC Protein Phosphorylation und Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, U.K.
‡Drug Discovery Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, U.K.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1606
Author: Boskovski, M. T., Yuan, S., Pedersen, N. B., Goth, C. K., Makova, S., Clausen, H., Brueckner, M. and Khokha, M. K.
Year: 2013
Title: The heterotaxy gene GALNT11 glycosylates Notch to orchestrate cilia type and laterality
Journal: Nature
Volume: 504
Issue: 7480
Pages: 456-9
Epub Date: 2013/11/15
Date: Dec 19
Short Title: The heterotaxy gene GALNT11 glycosylates Notch to orchestrate cilia type and laterality
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature12723
PMCID: PMC3869867
NIHMSID: NIHMS528287
Accession Number: 24226769
Keywords: ADAM Proteins/metabolism
ADAM17 Protein
Amino Acid Sequence
Animals
*Body Patterning
Cilia/metabolism/*physiology
Embryo, Nonmammalian/embryology/metabolism
Glycosylation
Heterotaxy Syndrome/*genetics
Humans
Mice
Molecular Sequence Data
N-Acetylgalactosaminyltransferases/deficiency/genetics/*metabolism
Peptide Fragments/chemistry/metabolism
Receptor, Notch1/chemistry/deficiency/genetics/*metabolism
*Signal Transduction
Xenopus/embryology/genetics
Xenopus Proteins/deficiency/genetics/*metabolism
Abstract: Heterotaxy is a disorder of left-right body patterning, or laterality, that is associated with major congenital heart disease. The aetiology and mechanisms underlying most cases of human heterotaxy are poorly understood. In vertebrates, laterality is initiated at the embryonic left-right organizer, where motile cilia generate leftward flow that is detected by immotile sensory cilia, which transduce flow into downstream asymmetric signals. The mechanism that specifies these two cilia types remains unknown. Here we show that the N-acetylgalactosamine-type O-glycosylation enzyme GALNT11 is crucial to such determination. We previously identified GALNT11 as a candidate disease gene in a patient with heterotaxy, and now demonstrate, in Xenopus tropicalis, that galnt11 activates Notch signalling. GALNT11 O-glycosylates human NOTCH1 peptides in vitro, thereby supporting a mechanism of Notch activation either by increasing ADAM17-mediated ectodomain shedding of the Notch receptor or by modification of specific EGF repeats. We further developed a quantitative live imaging technique for Xenopus left-right organizer cilia and show that Galnt11-mediated Notch1 signalling modulates the spatial distribution and ratio of motile and immotile cilia at the left-right organizer. galnt11 or notch1 depletion increases the ratio of motile cilia at the expense of immotile cilia and produces a laterality defect reminiscent of loss of the ciliary sensor Pkd2. By contrast, Notch overexpression decreases this ratio, mimicking the ciliopathy primary ciliary dyskinesia. Together our data demonstrate that Galnt11 modifies Notch, establishing an essential balance between motile and immotile cilia at the left-right organizer to determine laterality, and reveal a novel mechanism for human heterotaxy.
Notes: 1476-4687
Boskovski, Marko T
Yuan, Shiaulou
Pedersen, Nis Borbye
Goth, Christoffer Knak
Makova, Svetlana
Clausen, Henrik
Brueckner, Martina
Khokha, Mustafa K
DE018824/DE/NIDCR NIH HHS/United States
R01 DE018824/DE/NIDCR NIH HHS/United States
R01 HL093280/HL/NHLBI NIH HHS/United States
5T32HD00709436/HD/NICHD NIH HHS/United States
R01 DE018825/DE/NIDCR NIH HHS/United States
R01HL093280/HL/NHLBI NIH HHS/United States
DE018825/DE/NIDCR NIH HHS/United States
TL 1 RR024137/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2013 Dec 19;504(7480):456-9. doi: 10.1038/nature12723. Epub 2013 Nov 13.
Author Address: 1] Program in Vertebrate Developmental Biology, Department of Pediatrics and Genetics, Yale University School of Medicine, 333 Cedar Street, New Haven, Connecticut 06520, USA [2] [3] Department of Surgery, Brigham and Women's Hospital, Harvard Medical School, 75 Francis Street, Boston, Massachusetts 02115, USA.
1] Program in Vertebrate Developmental Biology, Department of Pediatrics and Genetics, Yale University School of Medicine, 333 Cedar Street, New Haven, Connecticut 06520, USA [2].
Copenhagen Center for Glycomics, Department of Cellular and Molecular Medicine, Faculty of Health Sciences, University of Copenhagen, Building 24.6.30, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark.
Program in Vertebrate Developmental Biology, Department of Pediatrics and Genetics, Yale University School of Medicine, 333 Cedar Street, New Haven, Connecticut 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1231
Author: Bostrom, J., Yu, S. F., Kan, D., Appleton, B. A., Lee, C. V., Billeci, K., Man, W., Peale, F., Ross, S., Wiesmann, C. and Fuh, G.
Year: 2009
Title: Variants of the antibody herceptin that interact with HER2 and VEGF at the antigen binding site
Journal: Science
Volume: 323
Issue: 5921
Pages: 1610-4
Epub Date: 2009/03/21
Date: Mar 20
Short Title: Variants of the antibody herceptin that interact with HER2 and VEGF at the antigen binding site
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1165480
Accession Number: 19299620
Keywords: Animals
Antibodies, Bispecific/chemistry/genetics/*immunology/therapeutic use
Antibodies, Monoclonal/chemistry/genetics/*immunology/therapeutic use
Antibodies, Monoclonal, Humanized
Antibody Affinity
Antibody Specificity
Binding Sites, Antibody/genetics
Cell Proliferation/drug effects
Complementarity Determining Regions/genetics/immunology
Crystallography, X-Ray
Epitopes/immunology/metabolism
Genetic Engineering
Humans
Mice
Models, Molecular
Mutagenesis
Neoplasms, Experimental/drug therapy
Protein Conformation
Protein Structure, Tertiary
Receptor, ErbB-2/chemistry/*immunology/metabolism
Thermodynamics
Trastuzumab
Vascular Endothelial Growth Factor A/chemistry/*immunology/metabolism
Xenograft Model Antitumor Assays
Abstract: The interface between antibody and antigen is often depicted as a lock and key, suggesting that an antibody surface can accommodate only one antigen. Here, we describe an antibody with an antigen binding site that binds two distinct proteins with high affinity. We isolated a variant of Herceptin, a therapeutic monoclonal antibody that binds the human epidermal growth factor receptor 2 (HER2), on the basis of its ability to simultaneously interact with vascular endothelial growth factor (VEGF). Crystallographic and mutagenesis studies revealed that distinct amino acids of this antibody, called bH1, engage HER2 and VEGF energetically, but there is extensive overlap between the antibody surface areas contacting the two antigens. An affinity-improved version of bH1 inhibits both HER2- and VEGF-mediated cell proliferation in vitro and tumor progression in mouse models. Such "two-in-one" antibodies challenge the monoclonal antibody paradigm of one binding site, one antigen. They could also provide new opportunities for antibody-based therapy.
Notes: 1095-9203
Bostrom, Jenny
Yu, Shang-Fan
Kan, David
Appleton, Brent A
Lee, Chingwei V
Billeci, Karen
Man, Wenyan
Peale, Franklin
Ross, Sarajane
Wiesmann, Christian
Fuh, Germaine
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
United States
Science. 2009 Mar 20;323(5921):1610-4. doi: 10.1126/science.1165480.
Author Address: Department of Protein Engineering, Genentech, 1 DNA Way, South San Francisco, CA 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1451
Author: Boström, P., Wu, J., Jedrychowski, M. P., Korde, A., Ye, L., Lo, J. C., Rasbach, K. A., Boström, E. A., Choi, J. H., Long, J. Z., Kajimura, S., Zingaretti, M. C., Vind, B. F., Tu, H., Cinti, S., Højlund, K., Gygi, S. P. and Spiegelman, B. M.
Year: 2012
Title: A PGC1-α-dependent myokine that drives brown-fat-like development of white fat and thermogenesis
Journal: Nature
Volume: 481
Issue: 7382
Pages: 463-8
Epub Date: 2012/01/13
Date: Jan 11
Short Title: A PGC1-α-dependent myokine that drives brown-fat-like development of white fat and thermogenesis
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature10777
PMCID: PMC3522098
NIHMSID: NIHMS383119
Accession Number: 22237023
Keywords: Adipocytes/cytology/drug effects/metabolism
Adipose Tissue, Brown/*cytology/drug effects/metabolism
Adipose Tissue, White/*cytology/drug effects/metabolism
Animals
Cell Respiration/drug effects
Cells, Cultured
Culture Media, Conditioned/pharmacology
Energy Metabolism/drug effects/genetics/physiology
Exercise/physiology
Gene Expression Regulation/drug effects/genetics
Hormones/metabolism
Humans
Insulin Resistance/physiology
Intracellular Signaling Peptides and Proteins/genetics/metabolism
Ion Channels/metabolism
Mice
Mice, Inbred BALB C
Mice, Transgenic
Mitochondrial Proteins/metabolism
Models, Animal
Muscle Cells/metabolism
Obesity/blood/chemically induced/prevention & control
Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha
Physical Conditioning, Animal/physiology
Plasma/chemistry
Subcutaneous Fat/cytology/drug effects/metabolism
*Thermogenesis/drug effects/genetics
Trans-Activators/deficiency/genetics/*metabolism
Transcription Factors
Uncoupling Protein 1
Abstract: Exercise benefits a variety of organ systems in mammals, and some of the best-recognized effects of exercise on muscle are mediated by the transcriptional co-activator PPAR-γ co-activator-1 α (PGC1-α). Here we show in mouse that PGC1-α expression in muscle stimulates an increase in expression of FNDC5, a membrane protein that is cleaved and secreted as a newly identified hormone, irisin. Irisin acts on white adipose cells in culture and in vivo to stimulate UCP1 expression and a broad program of brown-fat-like development. Irisin is induced with exercise in mice and humans, and mildly increased irisin levels in the blood cause an increase in energy expenditure in mice with no changes in movement or food intake. This results in improvements in obesity and glucose homeostasis. Irisin could be therapeutic for human metabolic disease and other disorders that are improved with exercise.
Notes: 1476-4687
Boström, Pontus
Wu, Jun
Jedrychowski, Mark P
Korde, Anisha
Ye, Li
Lo, James C
Rasbach, Kyle A
Boström, Elisabeth Almer
Choi, Jang Hyun
Long, Jonathan Z
Kajimura, Shingo
Zingaretti, Maria Cristina
Vind, Birgitte F
Tu, Hua
Cinti, Saverio
Højlund, Kurt
Gygi, Steven P
Spiegelman, Bruce M
R01 DK054477/DK/NIDDK NIH HHS/United States
R01 DK061562/DK/NIDDK NIH HHS/United States
R56 DK054477/DK/NIDDK NIH HHS/United States
K99 DK087853/DK/NIDDK NIH HHS/United States
DK31405/DK/NIDDK NIH HHS/United States
R37 DK031405/DK/NIDDK NIH HHS/United States
DK54477/DK/NIDDK NIH HHS/United States
R01 DK031405/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2012 Jan 11;481(7382):463-8. doi: 10.1038/nature10777.
Author Address: Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 557
Author: Bourne, Y., Mazurier, J., Legrand, D., Rougé, P., Montreuil, J., Spik, G. and Cambillau, C.
Year: 1994
Title: Structures of a legume lectin complexed with the human lactotransferrin N2 fragment, and with an isolated biantennary glycopeptide: role of the fucose moiety
Journal: Structure
Volume: 2
Issue: 3
Pages: 209-19
Epub Date: 1994/03/15
Date: Mar 15
Short Title: Structures of a legume lectin complexed with the human lactotransferrin N2 fragment, and with an isolated biantennary glycopeptide: role of the fucose moiety
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/s0969-2126(00)00022-8
Accession Number: 8069634
Keywords: Binding Sites
Carbohydrate Conformation
Carbohydrate Sequence
*Fucose
Glycopeptides/*chemistry/metabolism
Humans
Lactoferrin/*chemistry/*metabolism
Lectins/*chemistry/*metabolism
Models, Molecular
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/*chemistry/metabolism
*Plant Lectins
*Protein Structure, Secondary
Abstract: BACKGROUND: Lectins mediate cell-cell interactions by specifically recognizing oligosaccharide chains. Legume lectins serve as mediators for the symbiotic interactions between plants and nitrogen-fixing microorganisms, an important process in the nitrogen cycle. Lectins from the Viciae tribe have a high affinity for the fucosylated biantennary N-acetyllactosamine-type glycans which are to be found in the majority of N-glycosylproteins. While the structures of several lectins complexed with incomplete oligosaccharides have been solved, no previous structure has included the complete glycoprotein. RESULTS: We have determined the crystal structures of Lathyrus ochrus isolectin II complexed with the N2 monoglycosylated fragment of human lactotransferrin (18 kDa) and an isolated glycopeptide (2.1 kDa) fragment of human lactotransferrin (at 3.3 A and 2.8 A resolution, respectively). Comparison between the two structures showed that the protein part of the glycoprotein has little influence on either the stabilization of the complex or the sugar conformation. In both cases the oligosaccharide adopts the same extended conformation. Besides the essential mannose moiety of the monosaccharide-binding site, the fucose-1' of the core has a large surface of interaction with the lectin. This oligosaccharide conformation differs substantially from that seen in the previously determined isolectin I-octasaccharide complex. Comparison of our structure with that of concanavalin A (ConA) suggests that the ConA binding site cannot accommodate this fucose. CONCLUSIONS: Our results explain the observation that Viciae lectins have a higher affinity for fucosylated oligosaccharides than for unfucosylated ones, whereas the affinity of ConA for these types of oligosaccharides is similar. This explanation is testable by mutagenesis experiments. Our structure shows a large complementary surface area between the oligosaccharide and the lectin, in contrast with the recently determined structure of a complex between the carbohydrate recognition domain of a C-type mammalian lectin and an oligomannoside, where only the non-reducing terminal mannose residue interacts with the lectin.
Notes: Bourne, Y
Mazurier, J
Legrand, D
Rougé, P
Montreuil, J
Spik, G
Cambillau, C
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Structure. 1994 Mar 15;2(3):209-19. doi: 10.1016/s0969-2126(00)00022-8.
Author Address: Laboratoire de Cristallographie et de Cristallisation des Macromolécules Biologiques, CNRS URA 1296, Faculté de Médecine Secteur-Nord, Marseille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1978
Author: Bourré, G., Cantrelle, F. X., Kamah, A., Chambraud, B., Landrieu, I. and Smet-Nocca, C.
Year: 2018
Title: Direct Crosstalk Between O-GlcNAcylation and Phosphorylation of Tau Protein Investigated by NMR Spectroscopy
Journal: Front Endocrinol (Lausanne)
Volume: 9
Pages: 595
Epub Date: 2018/11/06
Short Title: Direct Crosstalk Between O-GlcNAcylation and Phosphorylation of Tau Protein Investigated by NMR Spectroscopy
Alternate Journal: Frontiers in endocrinology
ISSN: 1664-2392 (Print)
1664-2392
DOI: 10.3389/fendo.2018.00595
PMCID: PMC6198643
Accession Number: 30386294
Keywords: NMR spectroscopy
O-GlcNAcylation
Tau protein
crosstalk
phosphorylation
Abstract: The formation of intraneuronal fibrillar inclusions of tau protein is associated with several neurodegenerative diseases referred to as tauopathies including Alzheimer's disease (AD). A common feature of these pathologies is hyperphosphorylation of tau, the main component of fibrillar assemblies such as Paired Helical Filaments (PHFs). O-β-linked N-acetylglucosaminylation (O-GlcNAcylation) is another important posttranslational modification involved in regulation of tau pathophysiology. Among the benefits of O-GlcNAcylation, modulation of tau phosphorylation levels and inhibition of tau aggregation properties have been described while decreased O-GlcNAcylation could be involved in the raise of tau phosphorylation associated with AD. However, the molecular mechanisms at the basis of these observations remain to be defined. In this study, we identify by NMR spectroscopy O-GlcNAc sites in the longest isoform of tau and investigate the direct role of O-GlcNAcylation on tau phosphorylation and conversely, the role of phosphorylation on tau O-GlcNAcylation. We show here by a systematic examination of the quantitative modification patterns by NMR spectroscopy that O-GlcNAcylation does not modify phosphorylation of tau by the kinase activity of ERK2 or a rat brain extract while phosphorylation slightly increases tau O-GlcNAcylation by OGT. Our data suggest that indirect mechanisms act in the reciprocal regulation of tau phosphorylation and O-GlcNAcylation in vivo involving regulation of the enzymes responsible of phosphate and O-GlcNAc dynamics.
Notes: 1664-2392
Bourré, Gwendoline
Cantrelle, François-Xavier
Kamah, Amina
Chambraud, Béatrice
Landrieu, Isabelle
Smet-Nocca, Caroline
Journal Article
Front Endocrinol (Lausanne). 2018 Oct 16;9:595. doi: 10.3389/fendo.2018.00595. eCollection 2018.
Author Address: Univ. Lille, CNRS UMR8576, Unité de Glycobiologie Structurale et Fonctionnelle, Lille, France.
Univ. Paris XI, UMR 1195 Inserm, Le Kremlin Bicêtre, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1013
Author: Bouyain, S., Longo, P. A., Li, S., Ferguson, K. M. and Leahy, D. J.
Year: 2005
Title: The extracellular region of ErbB4 adopts a tethered conformation in the absence of ligand
Journal: Proc Natl Acad Sci U S A
Volume: 102
Issue: 42
Pages: 15024-9
Epub Date: 2005/10/06
Date: Oct 18
Short Title: The extracellular region of ErbB4 adopts a tethered conformation in the absence of ligand
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0507591102
PMCID: PMC1257738
Accession Number: 16203964
Keywords: Adult
Amino Acid Sequence
Crystallography, X-Ray
ErbB Receptors/*chemistry/metabolism
Humans
Ligands
Models, Molecular
Molecular Sequence Data
Protein Binding
*Protein Conformation
Receptor, ErbB-4
Sequence Alignment
Abstract: The human ErbB family of receptor tyrosine kinases comprises the epidermal growth factor receptor (EGFR/ErbB1/HER1), ErbB2 (HER2/Neu), ErbB3 (HER3), and ErbB4 (HER4). ErbBs play fundamental roles in cell growth and differentiation events in embryonic and adult tissues, and inappropriate ErbB activity has been implicated in several human cancers. We report here the 2.4 A crystal structure of the extracellular region of human ErbB4 in the absence of ligand and show that it adopts a tethered conformation similar to inactive forms of ErbB1 and ErbB3. This structure completes the gallery of unliganded ErbB receptors and demonstrates that all human ligand-binding ErbBs adopt the autoinhibited conformation. We also show that the binding of neuregulin-1beta to ErbB4 and ErbB3 and the binding of betacellulin to both ErbB4 and ErbB1 does not decrease at low pH, unlike the binding of epidermal growth factor and transforming growth factor-alpha to ErbB1. These results indicate an important role for ligand in determining pH-dependent binding and may explain different responses observed when the same ErbB receptor is stimulated by different ligands.
Notes: 1091-6490
Bouyain, Samuel
Longo, Patti A
Li, Shiqing
Ferguson, Kathryn M
Leahy, Daniel J
R01-CA90466/CA/NCI NIH HHS/United States
T32-HL007027/HL/NHLBI NIH HHS/United States
T32 HL007027/HL/NHLBI NIH HHS/United States
K01-CA092246/CA/NCI NIH HHS/United States
R01 CA090466/CA/NCI NIH HHS/United States
Wellcome Trust/United Kingdom
K01 CA092246/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2005 Oct 18;102(42):15024-9. doi: 10.1073/pnas.0507591102. Epub 2005 Oct 3.
Author Address: Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1293
Author: Bouyain, S. and Watkins, D. J.
Year: 2010
Title: The protein tyrosine phosphatases PTPRZ and PTPRG bind to distinct members of the contactin family of neural recognition molecules
Journal: Proc Natl Acad Sci U S A
Volume: 107
Issue: 6
Pages: 2443-8
Epub Date: 2010/02/06
Date: Feb 9
Short Title: The protein tyrosine phosphatases PTPRZ and PTPRG bind to distinct members of the contactin family of neural recognition molecules
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0911235107
PMCID: PMC2823867
Accession Number: 20133774
Keywords: Amino Acid Sequence
Animals
Binding Sites/genetics
Cell Adhesion Molecules, Neuronal/*chemistry/genetics/metabolism
Cell Line
Chromatography, Gel
Contactin 1
Contactins
Crystallography, X-Ray
Humans
Mice
Models, Molecular
Molecular Sequence Data
*Multigene Family
Protein Binding
Protein Conformation
Protein Isoforms/chemistry/genetics/metabolism
Protein Structure, Secondary
*Protein Structure, Tertiary
Receptor-Like Protein Tyrosine Phosphatases, Class 5/*chemistry/genetics/metabolism
Sequence Homology, Amino Acid
Abstract: The receptor protein tyrosine phosphatases gamma (PTPRG) and zeta (PTPRZ) are expressed primarily in the nervous system and mediate cell adhesion and signaling events during development. We report here the crystal structures of the carbonic anhydrase-like domains of PTPRZ and PTPRG and show that these domains interact directly with the second and third immunoglobulin repeats of the members of the contactin (CNTN) family of neural recognition molecules. Interestingly, these receptors exhibit distinct specificities: PTPRZ binds only to CNTN1, whereas PTPRG interacts with CNTN3, 4, 5, and 6. Furthermore, we present crystal structures of the four N-terminal immunoglobulin repeats of mouse CNTN4 both alone and in complex with the carbonic anhydrase-like domain of mouse PTPRG. In these structures, the N-terminal region of CNTN4 adopts a horseshoe-like conformation found also in CNTN2 and most likely in all CNTNs. This restrained conformation of the second and third immunoglobulin domains creates a binding site that is conserved among CNTN3, 4, 5, and 6. This site contacts a discrete region of PTPRG composed primarily of an extended beta-hairpin loop found in both PTPRG and PTPRZ. Overall, these findings implicate PTPRG, PTPRZ and CNTNs as a group of receptors and ligands involved in the manifold recognition events that underlie the construction of neural networks.
Notes: 1091-6490
Bouyain, Samuel
Watkins, Dara J
Journal Article
Proc Natl Acad Sci U S A. 2010 Feb 9;107(6):2443-8. doi: 10.1073/pnas.0911235107. Epub 2010 Jan 21.
Author Address: Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri-Kansas City, Kansas City, MO 64110, USA. bouyains@umkc.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1167
Author: Bowden, T. A., Aricescu, A. R., Gilbert, R. J., Grimes, J. M., Jones, E. Y. and Stuart, D. I.
Year: 2008
Title: Structural basis of Nipah and Hendra virus attachment to their cell-surface receptor ephrin-B2
Journal: Nat Struct Mol Biol
Volume: 15
Issue: 6
Pages: 567-72
Epub Date: 2008/05/20
Date: Jun
Short Title: Structural basis of Nipah and Hendra virus attachment to their cell-surface receptor ephrin-B2
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.1435
Accession Number: 18488039
Keywords: Crystallography, X-Ray
Ephrin-B2/*chemistry/metabolism
Hendra Virus
Humans
Nipah Virus
Paramyxoviridae/*pathogenicity
Protein Binding
Protein Conformation
Receptors, Virus
Viral Envelope Proteins/*chemistry/metabolism
*Virus Attachment
Abstract: Nipah and Hendra viruses are emergent paramyxoviruses, causing disease characterized by rapid onset and high mortality rates, resulting in their classification as Biosafety Level 4 pathogens. Their attachment glycoproteins are essential for the recognition of the cell-surface receptors ephrin-B2 (EFNB2) and ephrin-B3 (EFNB3). Here we report crystal structures of both Nipah and Hendra attachment glycoproteins in complex with human EFNB2. In contrast to previously solved paramyxovirus attachment complexes, which are mediated by sialic acid interactions, the Nipah and Hendra complexes are maintained by an extensive protein-protein interface, including a crucial phenylalanine side chain on EFNB2 that fits snugly into a hydrophobic pocket on the viral protein. By analogy with the development of antivirals against sialic acid binding viruses, these results provide a structural template to target antiviral inhibition of protein-protein interactions.
Notes: 1545-9985
Bowden, Thomas A
Aricescu, A Radu
Gilbert, Robert J C
Grimes, Jonathan M
Jones, E Yvonne
Stuart, David I
G0500365/Medical Research Council/United Kingdom
G0700232/Medical Research Council/United Kingdom
Wellcome Trust/United Kingdom
Cancer Research UK/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2008 Jun;15(6):567-72. doi: 10.1038/nsmb.1435. Epub 2008 May 18.
Author Address: Division of Structural Biology, University of Oxford, Henry Wellcome Building of Genomic Medicine, Roosevelt Drive, Oxford OX3 7BN, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1270
Author: Bowden, T. A., Aricescu, A. R., Nettleship, J. E., Siebold, C., Rahman-Huq, N., Owens, R. J., Stuart, D. I. and Jones, E. Y.
Year: 2009
Title: Structural plasticity of eph receptor A4 facilitates cross-class ephrin signaling
Journal: Structure
Volume: 17
Issue: 10
Pages: 1386-97
Epub Date: 2009/10/20
Date: Oct 14
Short Title: Structural plasticity of eph receptor A4 facilitates cross-class ephrin signaling
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2009.07.018
PMCID: PMC2832735
Accession Number: 19836338
Keywords: Amino Acid Sequence
Binding Sites
Ephrins/*chemistry/*metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Phylogeny
Protein Conformation
Receptor, EphA4/*chemistry/*metabolism
*Signal Transduction
Structure-Activity Relationship
Abstract: The EphA4 tyrosine kinase cell surface receptor regulates an array of physiological processes and is the only currently known class A Eph receptor that binds both A and B class ephrins with high affinity. We have solved the crystal structure of the EphA4 ligand binding domain alone and in complex with (1) ephrinB2 and (2) ephrinA2. This set of structures shows that EphA4 has significant conformational plasticity in its ligand binding face. In vitro binding data demonstrate that it has a higher affinity for class A than class B ligands. Structural analyses, drawing on previously reported Eph receptor structures, show that EphA4 in isolation and in complex with ephrinA2 resembles other class A Eph receptors but on binding ephrinB2 assumes structural hallmarks of the class B Eph receptors. This interactive plasticity reveals EphA4 as a structural chameleon, able to adopt both A and B class Eph receptor conformations, and thus provides a molecular basis for EphA-type cross-class reactivity.
Notes: 1878-4186
Bowden, Thomas A
Aricescu, A Radu
Nettleship, Joanne E
Siebold, Christian
Rahman-Huq, Nahid
Owens, Raymond J
Stuart, David I
Jones, E Yvonne
G0701506/Medical Research Council/United Kingdom
G0700232/Medical Research Council/United Kingdom
C375/A3964/Cancer Research UK/United Kingdom
Wellcome Trust/United Kingdom
G0500365/Medical Research Council/United Kingdom
10976/Cancer Research UK/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Structure. 2009 Oct 14;17(10):1386-97. doi: 10.1016/j.str.2009.07.018.
Author Address: Division of Structural Biology, University of Oxford, Henry Wellcome Building of Genomic Medicine, Roosevelt Drive, Oxford, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1377
Author: Boyce, M., Carrico, I. S., Ganguli, A. S., Yu, S. H., Hangauer, M. J., Hubbard, S. C., Kohler, J. J. and Bertozzi, C. R.
Year: 2011
Title: Metabolic cross-talk allows labeling of O-linked beta-N-acetylglucosamine-modified proteins via the N-acetylgalactosamine salvage pathway
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 8
Pages: 3141-6
Epub Date: 2011/02/09
Date: Feb 22
Short Title: Metabolic cross-talk allows labeling of O-linked beta-N-acetylglucosamine-modified proteins via the N-acetylgalactosamine salvage pathway
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1010045108
PMCID: PMC3044403
Accession Number: 21300897
Keywords: Acetylgalactosamine/*metabolism
Acetylglucosamine/*metabolism
*Affinity Labels
Cell Line
Chromatography, Affinity
Glycosylation
Humans
*Metabolic Networks and Pathways
Methods
Protein Processing, Post-Translational
*Receptor Cross-Talk
Abstract: Hundreds of mammalian nuclear and cytoplasmic proteins are reversibly glycosylated by O-linked β-N-acetylglucosamine (O-GlcNAc) to regulate their function, localization, and stability. Despite its broad functional significance, the dynamic and posttranslational nature of O-GlcNAc signaling makes it challenging to study using traditional molecular and cell biological techniques alone. Here, we report that metabolic cross-talk between the N-acetylgalactosamine salvage and O-GlcNAcylation pathways can be exploited for the tagging and identification of O-GlcNAcylated proteins. We found that N-azidoacetylgalactosamine (GalNAz) is converted by endogenous mammalian biosynthetic enzymes to UDP-GalNAz and then epimerized to UDP-N-azidoacetylglucosamine (GlcNAz). O-GlcNAc transferase accepts UDP-GlcNAz as a nucleotide-sugar donor, appending an azidosugar onto its native substrates, which can then be detected by covalent labeling using azide-reactive chemical probes. In a proof-of-principle proteomics experiment, we used metabolic GalNAz labeling of human cells and a bioorthogonal chemical probe to affinity-purify and identify numerous O-GlcNAcylated proteins. Our work provides a blueprint for a wide variety of future chemical approaches to identify, visualize, and characterize dynamic O-GlcNAc signaling.
Notes: 1091-6490
Boyce, Michael
Carrico, Isaac S
Ganguli, Anjali S
Yu, Seok-Ho
Hangauer, Matthew J
Hubbard, Sarah C
Kohler, Jennifer J
Bertozzi, Carolyn R
R01 GM066047/GM/NIGMS NIH HHS/United States
GM066047/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
GM069157/GM/NIGMS NIH HHS/United States
F32 GM069157/GM/NIGMS NIH HHS/United States
PC080659/PC/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2011 Feb 22;108(8):3141-6. doi: 10.1073/pnas.1010045108. Epub 2011 Feb 7.
Author Address: Departments of Chemistry and Molecular and Cell Biology and Howard Hughes Medical Institute, University of California, Berkeley, CA 94720, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1373
Author: Brasch, J., Harrison, O. J., Ahlsen, G., Carnally, S. M., Henderson, R. M., Honig, B. and Shapiro, L.
Year: 2011
Title: Structure and binding mechanism of vascular endothelial cadherin: a divergent classical cadherin
Journal: J Mol Biol
Volume: 408
Issue: 1
Pages: 57-73
Epub Date: 2011/01/29
Date: Apr 22
Short Title: Structure and binding mechanism of vascular endothelial cadherin: a divergent classical cadherin
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2011.01.031
PMCID: PMC3084036
NIHMSID: NIHMS288748
Accession Number: 21269602
Keywords: Amino Acid Sequence
Animals
Antigens, CD/*chemistry/*metabolism
Cadherins/*chemistry/*metabolism
Cells, Cultured
Chickens
Chromatography, Gel
Crystallography, X-Ray
Endothelium, Vascular/*metabolism
Glycosylation
Humans
Mice
Microscopy, Atomic Force
Molecular Sequence Data
Protein Binding
Protein Conformation
Protein Multimerization
Sequence Homology, Amino Acid
Abstract: Vascular endothelial cadherin (VE-cadherin), a divergent member of the type II classical cadherin family of cell adhesion proteins, mediates homophilic adhesion in the vascular endothelium. Previous investigations with a bacterially produced protein suggested that VE-cadherin forms cell surface trimers that bind between apposed cells to form hexamers. Here we report studies of mammalian-produced VE-cadherin ectodomains suggesting that, like other classical cadherins, VE-cadherin forms adhesive trans dimers between monomers located on opposing cell surfaces. Trimerization of the bacterially produced protein appears to be an artifact that arises from a lack of glycosylation. We also present the 2.1-Å-resolution crystal structure of the VE-cadherin EC1-2 adhesive region, which reveals homodimerization via the strand-swap mechanism common to classical cadherins. In common with type II cadherins, strand-swap binding involves two tryptophan anchor residues, but the adhesive interface resembles type I cadherins in that VE-cadherin does not form a large nonswapped hydrophobic surface. Thus, VE-cadherin is an outlier among classical cadherins, with characteristics of both type I and type II subfamilies.
Notes: 1089-8638
Brasch, Julia
Harrison, Oliver J
Ahlsen, Goran
Carnally, Stewart M
Henderson, Robert M
Honig, Barry
Shapiro, Lawrence
R01 GM062270/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Mol Biol. 2011 Apr 22;408(1):57-73. doi: 10.1016/j.jmb.2011.01.031. Epub 2011 Jan 24.
Author Address: Department of Biochemistry and Molecular Biophysics, Columbia University, 701 West 168th Street, New York, NY 10032, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1439
Author: Brasch, J., Harrison, O. J., Ahlsen, G., Liu, Q. and Shapiro, L.
Year: 2011
Title: Crystal structure of the ligand binding domain of netrin G2
Journal: J Mol Biol
Volume: 414
Issue: 5
Pages: 723-34
Epub Date: 2011/11/02
Date: Dec 16
Short Title: Crystal structure of the ligand binding domain of netrin G2
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2011.10.030
PMCID: PMC4535689
NIHMSID: NIHMS689758
Accession Number: 22041449
Keywords: Amino Acid Sequence
Animals
Calcium/chemistry
Crystallography, X-Ray
GPI-Linked Proteins/*chemistry
Humans
Laminin/chemistry
Mice
Molecular Sequence Data
Nerve Tissue Proteins/*chemistry
Netrins
Protein Binding
Abstract: Netrin G proteins represent a small family of synaptic cell adhesion molecules related to netrins and to the polymerization domains of laminins. Two netrin G proteins are encoded in vertebrate genomes, netrins G1 and G2, which are known to bind the leucine-rich repeat proteins netrin G ligand (NGL)-1 and NGL-2, respectively. Netrin G proteins share a common multi-domain architecture comprising a laminin N-terminal (LN) domain followed by three laminin epidermal growth factor-like (LE) domains and a C' region containing a glycosylphosphatidylinositol anchor. Here, we use deletion analysis to show that the LN domain region of netrin Gs contains the binding site for NGLs to which they bind with 1:1 stoichiometry and sub-micromolar affinity. Netrin Gs are alternatively spliced in their LE domain regions, but the binding region, the LN domain, is identical in all splice forms. We determined the crystal structure for a fragment comprising the LN domain and domain LE1 of netrin G2 by sulfur single-wavelength anomalous diffraction phasing and refined it to 1.8 Å resolution. The structure reveals an overall architecture similar to that of laminin α chain LN domains but includes significant differences including a Ca(2+) binding site in the LN domain. These results reveal the minimal binding unit for interaction of netrin Gs with NGLs, define structural features specific to netrin Gs, and suggest that netrin G alternative splicing is not involved in NGL recognition.
Notes: 1089-8638
Brasch, Julia
Harrison, Oliver J
Ahlsen, Goran
Liu, Qun
Shapiro, Lawrence
R01 GM062270/GM/NIGMS NIH HHS/United States
GM62270/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Mol Biol. 2011 Dec 16;414(5):723-34. doi: 10.1016/j.jmb.2011.10.030. Epub 2011 Oct 25.
Author Address: Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1533
Author: Bräutigam, J., Scheidig, A. J. and Egge-Jacobsen, W.
Year: 2013
Title: Mass spectrometric analysis of hepatitis C viral envelope protein E2 reveals extended microheterogeneity of mucin-type O-linked glycosylation
Journal: Glycobiology
Volume: 23
Issue: 4
Pages: 453-74
Epub Date: 2012/12/18
Date: Apr
Short Title: Mass spectrometric analysis of hepatitis C viral envelope protein E2 reveals extended microheterogeneity of mucin-type O-linked glycosylation
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cws171
Accession Number: 23242014
Keywords: Amino Acid Sequence
Glycosylation
HEK293 Cells
Hepacivirus/*chemistry
Humans
Mass Spectrometry
Molecular Sequence Data
Mucins/*chemistry/metabolism
Viral Envelope Proteins/*chemistry/metabolism
Abstract: The infectious liver disease hepatitis C is caused by the small, enveloped, positive single-strand RNA hepatitis C virus (HCV). The HCV genome encodes for a single polyprotein precursor of ∼3010 amino acid residues. Host and cellular proteases co- and posttranslational process the precursor creating six nonstructural (NS) proteins and four structural components. Properly folded forms of the envelope proteins E1 and E2 form the associated E1-E2 complex. This complex represents a significant antigenic component at the viral surface that can interact with several target cell receptors. Extent and type of glycosylation is an important factor for virulence and escape from the immune system. Detailed characterization of the glycosylated sites is helpful for the understanding of different phenotypes as well as for the development of E1/E2-related treatments of HCV infection. Here, we have investigated in detail the O-linked glycosylation of the HCV envelope protein E2 expressed in and isolated from human embryonic kidney (HEK 293) cells. Using nano-liquid chromatography and tandem mass spectrometry approaches, we clearly have identified six residues for O-linked glycosylation within isolated glycopeptides (Ser393, Thr396, Ser401, Ser404, Thr473 and Thr518), carrying mainly Core 1 and Core 2 mucin-type structures. Based on our data, Thr385 is probably glycosylated as well. In addition, we could show that Ser479 within the hyper variable region (HVR) I is not O-glycosylated. For most of these sites, different degrees of microheterogeneity could be verified. Concerning HCV E2, this is the first case of experimentally proven O-linked glycosylation in detail via mass spectrometry.
Notes: 1460-2423
Bräutigam, Joachim
Scheidig, Axel J
Egge-Jacobsen, Wolfgang
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2013 Apr;23(4):453-74. doi: 10.1093/glycob/cws171. Epub 2012 Dec 14.
Author Address: Department of Structural Biology, Centre for Biochemistry and Molecular Biology, Christian-Albrechts Universität, 24118 Kiel, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 747
Author: Brayer, G. D., Sidhu, G., Maurus, R., Rydberg, E. H., Braun, C., Wang, Y., Nguyen, N. T., Overall, C. M. and Withers, S. G.
Year: 2000
Title: Subsite mapping of the human pancreatic alpha-amylase active site through structural, kinetic, and mutagenesis techniques
Journal: Biochemistry
Volume: 39
Issue: 16
Pages: 4778-91
Epub Date: 2000/04/19
Date: Apr 25
Short Title: Subsite mapping of the human pancreatic alpha-amylase active site through structural, kinetic, and mutagenesis techniques
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi9921182
Accession Number: 10769135
Keywords: Acarbose/chemistry/metabolism
Amino Acid Substitution/genetics
Animals
Binding Sites
Carbohydrate Sequence
Catalysis
Crystallography, X-Ray
Fluorides/metabolism
Glycosylation
Humans
Hydrolysis
Kinetics
Maltose/metabolism
Models, Chemical
Models, Molecular
Molecular Sequence Data
Mutagenesis/*genetics
Oligosaccharides/chemistry/metabolism
Pancreas/*enzymology
Protein Conformation
Structure-Activity Relationship
Swine
Thermodynamics
alpha-Amylases/antagonists & inhibitors/*chemistry/genetics/*metabolism
Abstract: We report a multifaceted study of the active site region of human pancreatic alpha-amylase. Through a series of novel kinetic analyses using malto-oligosaccharides and malto-oligosaccharyl fluorides, an overall cleavage action pattern for this enzyme has been developed. The preferred binding/cleavage mode occurs when a maltose residue serves as the leaving group (aglycone sites +1 and +2) and there are three sugars in the glycon (-1, -2, -3) sites. Overall it appears that five binding subsites span the active site, although an additional glycon subsite appears to be a significant factor in the binding of longer substrates. Kinetic parameters for the cleavage of substrates modified at the 2 and 4' ' positions also highlight the importance of these hydroxyl groups for catalysis and identify the rate-determining step. Further kinetic and structural studies pinpoint Asp197 as being the likely nucleophile in catalysis, with substitution of this residue leading to an approximately 10(6)-fold drop in catalytic activity. Structural studies show that the original pseudo-tetrasaccharide structure of acarbose is modified upon binding, presumably through a series of hydrolysis and transglycosylation reactions. The end result is a pseudo-pentasaccharide moiety that spans the active site region with its N-linked "glycosidic" bond positioned at the normal site of cleavage. Interestingly, the side chains of Glu233 and Asp300, along with a water molecule, are aligned about the inhibitor N-linked glycosidic bond in a manner suggesting that these might act individually or collectively in the role of acid/base catalyst in the reaction mechanism. Indeed, kinetic analyses show that substitution of the side chains of either Glu233 or Asp300 leads to as much as a approximately 10(3)-fold decrease in catalytic activity. Structural analyses of the Asp300Asn variant of human pancreatic alpha-amylase and its complex with acarbose clearly demonstrate the importance of Asp300 to the mode of inhibitor binding.
Notes: Brayer, G D
Sidhu, G
Maurus, R
Rydberg, E H
Braun, C
Wang, Y
Nguyen, N T
Overall, C M
Withers, S G
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2000 Apr 25;39(16):4778-91. doi: 10.1021/bi9921182.
Author Address: Department of Biochemistry, University of British Columbia, Vancouver V6T 1Z3, Canada. brayer@laue.biochem.ubc.ca
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 947
Author: Breuza, L., Halbeisen, R., Jenö, P., Otte, S., Barlowe, C., Hong, W. and Hauri, H. P.
Year: 2004
Title: Proteomics of endoplasmic reticulum-Golgi intermediate compartment (ERGIC) membranes from brefeldin A-treated HepG2 cells identifies ERGIC-32, a new cycling protein that interacts with human Erv46
Journal: J Biol Chem
Volume: 279
Issue: 45
Pages: 47242-53
Epub Date: 2004/08/17
Date: Nov 5
Short Title: Proteomics of endoplasmic reticulum-Golgi intermediate compartment (ERGIC) membranes from brefeldin A-treated HepG2 cells identifies ERGIC-32, a new cycling protein that interacts with human Erv46
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M406644200
Accession Number: 15308636
Keywords: Amino Acid Sequence
Animals
Brefeldin A/*pharmacology
Carrier Proteins/*biosynthesis/*chemistry
Cell Line
Cell Membrane/*metabolism
Cross-Linking Reagents/pharmacology
Cytoplasm/metabolism
Electrophoresis, Polyacrylamide Gel
Endoplasmic Reticulum/*metabolism
Golgi Apparatus/*metabolism
HeLa Cells
Humans
Immunoblotting
Immunoprecipitation
Mass Spectrometry
Membrane Proteins/*biosynthesis/*chemistry/*metabolism
Mice
Microscopy, Fluorescence
Models, Biological
Molecular Sequence Data
Phylogeny
Protein Binding
Protein Structure, Tertiary
Proteome/*chemistry
RNA Interference
Saccharomyces cerevisiae Proteins/chemistry/*metabolism
Sequence Homology, Amino Acid
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Subcellular Fractions/metabolism
Vesicular Transport Proteins/chemistry
Abstract: Cycling proteins play important roles in the organization and function of the early secretory pathway by participating in membrane traffic and selective transport of cargo between the endoplasmic reticulum (ER), the intermediate compartment (ERGIC), and the Golgi. To identify new cycling proteins, we have developed a novel procedure for the purification of ERGIC membranes from HepG2 cells treated with brefeldin A, a drug known to accumulate cycling proteins in the ERGIC. Membranes enriched 110-fold over the homogenate for ERGIC-53 were obtained and analyzed by mass spectrometry. Major proteins corresponded to established and putative cargo receptors and components mediating protein maturation and membrane traffic. Among the uncharacterized proteins, a 32-kDa protein termed ERGIC-32 is a novel cycling membrane protein with sequence homology to Erv41p and Erv46p, two proteins enriched in COPII vesicles of yeast. ERGIC-32 localizes to the ERGIC and partially colocalizes with the human homologs of Erv41p and Erv46p, which mainly localize to the cis-Golgi. ERGIC-32 interacts with human Erv46 (hErv46) as revealed by covalent cross-linking and mistargeting experiments, and silencing of ERGIC-32 by small interfering RNAs increases the turnover of hErv46. We propose that ERGIC-32 functions as a modulator of the hErv41-hErv46 complex by stabilizing hErv46. Our novel approach for the isolation of the ERGIC from BFA-treated cells may ultimately lead to the identification of all proteins rapidly cycling early in the secretory pathway.
Notes: Breuza, Lionel
Halbeisen, Regula
Jenö, Paul
Otte, Stefan
Barlowe, Charles
Hong, Wanjin
Hauri, Hans-Peter
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2004 Nov 5;279(45):47242-53. doi: 10.1074/jbc.M406644200. Epub 2004 Aug 11.
Author Address: Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1932
Author: Briggs, D. C. and Hohenester, E.
Year: 2018
Title: Structural Basis for the Initiation of Glycosaminoglycan Biosynthesis by Human Xylosyltransferase 1
Journal: Structure
Volume: 26
Issue: 6
Pages: 801-809.e3
Epub Date: 2018/04/24
Date: Jun 5
Short Title: Structural Basis for the Initiation of Glycosaminoglycan Biosynthesis by Human Xylosyltransferase 1
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2018.03.014
PMCID: PMC5992326
Accession Number: 29681470
Keywords: Catalytic Domain
Crystallography, X-Ray
Glycosaminoglycans/*biosynthesis
Glycosylation
Humans
Models, Molecular
Pentosyltransferases/*chemistry/*metabolism
Protein Binding
Protein Conformation
Serine/chemistry
Substrate Specificity
Uridine Diphosphate Xylose/*metabolism
*X-ray crystallography
*enzyme catalysis
*glycosyltransferase
*proteoglycan
Abstract: Proteoglycans (PGs) are essential components of the animal extracellular matrix and are required for cell adhesion, migration, signaling, and immune function. PGs are composed of a core protein and long glycosaminoglycan (GAG) chains, which often specify PG function. GAG biosynthesis is initiated by peptide O-xylosyltransferases, which transfer xylose onto selected serine residues in the core proteins. We have determined crystal structures of human xylosyltransferase 1 (XT1) in complex with the sugar donor, UDP-xylose, and various acceptor peptides. The structures reveal unique active-site features that, in conjunction with functional experiments, explain the substrate specificity of XT1. A constriction within the peptide binding cleft requires the acceptor serine to be followed by glycine or alanine. The remainder of the cleft can accommodate a wide variety of sequences, but with a general preference for acidic residues. These findings provide a framework for understanding the selectivity of GAG attachment.
Notes: 1878-4186
Briggs, David C
Hohenester, Erhard
Wellcome Trust/United Kingdom
BB/D524840/1/Biotechnology and Biological Sciences Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Structure. 2018 Jun 5;26(6):801-809.e3. doi: 10.1016/j.str.2018.03.014. Epub 2018 Apr 19.
Author Address: Department of Life Sciences, Imperial College London, London SW7 2AZ, UK. Electronic address: david.briggs@crick.ac.uk.
Department of Life Sciences, Imperial College London, London SW7 2AZ, UK. Electronic address: e.hohenester@imperial.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1479
Author: Brinkmalm, G., Portelius, E., Öhrfelt, A., Mattsson, N., Persson, R., Gustavsson, M. K., Vite, C. H., Gobom, J., Månsson, J. E., Nilsson, J., Halim, A., Larson, G., Rüetschi, U., Zetterberg, H., Blennow, K. and Brinkmalm, A.
Year: 2012
Title: An online nano-LC-ESI-FTICR-MS method for comprehensive characterization of endogenous fragments from amyloid β and amyloid precursor protein in human and cat cerebrospinal fluid
Journal: J Mass Spectrom
Volume: 47
Issue: 5
Pages: 591-603
Epub Date: 2012/05/12
Date: May
Short Title: An online nano-LC-ESI-FTICR-MS method for comprehensive characterization of endogenous fragments from amyloid β and amyloid precursor protein in human and cat cerebrospinal fluid
Alternate Journal: Journal of mass spectrometry : JMS
ISSN: 1076-5174
DOI: 10.1002/jms.2987
Accession Number: 22576872
Keywords: Amino Acid Sequence
Amino Acid Substitution
Amyloid beta-Peptides/*cerebrospinal fluid/chemistry
Amyloid beta-Protein Precursor/*cerebrospinal fluid/chemistry
Animals
Cats
Chromatography, Liquid/*methods
Glycosylation
Humans
Immunoprecipitation
Molecular Sequence Data
Sequence Alignment
Sequence Analysis, Protein/*methods
Spectrometry, Mass, Electrospray Ionization/*methods
Abstract: Amyloid precursor protein (APP) is the precursor protein to amyloid β (Aβ), the main constituent of senile plaques in Alzheimer's disease (AD). Endogenous Aβ peptides reflect the APP processing, and greater knowledge of different APP degradation pathways is important to understand the mechanism underlying AD pathology. When one analyzes longer Aβ peptides by low-energy collision-induced dissociation tandem mass spectrometry (MS/MS), mainly long b-fragments are observed, limiting the possibility to determine variations such as amino acid variants or post-translational modifications (PTMs) within the N-terminal half of the peptide. However, by using electron capture dissociation (ECD), we obtained a more comprehensive sequence coverage for several APP/Aβ peptide species, thus enabling a deeper characterization of possible variants and PTMs. Abnormal APP/Aβ processing has also been described in the lysosomal storage disease Niemann-Pick type C and the major large animal used for studying this disease is cat. By ECD MS/MS, a substitution of Asp7 → Glu in cat Aβ was identified. Further, sialylated core 1 like O-glycans at Tyr10, recently discovered in human Aβ (a previously unknown glycosylation type), were identified also in cat cerebrospinal fluid (CSF). It is therefore likely that this unusual type of glycosylation is common for (at least) species belonging to the magnorder Boreoeutheria. We here describe a detailed characterization of endogenous APP/Aβ peptide species in CSF by using an online top-down MS-based method.
Notes: 1096-9888
Brinkmalm, Gunnar
Portelius, Erik
Öhrfelt, Annika
Mattsson, Niklas
Persson, Rita
Gustavsson, Mikael K
Vite, Charles H
Gobom, Johan
Månsson, Jan-Eric
Nilsson, Jonas
Halim, Adnan
Larson, Göran
Rüetschi, Ulla
Zetterberg, Henrik
Blennow, Kaj
Brinkmalm, Ann
Journal Article
Research Support, Non-U.S. Gov't
England
J Mass Spectrom. 2012 May;47(5):591-603. doi: 10.1002/jms.2987.
Author Address: Clinical Neurochemistry Laboratory, Institute of Neuroscience and Physiology, Department of Psychiatry and Neurochemistry, Sahlgrenska Academy at University of Gothenburg, Mölndal, Sweden. gunnar.brinkmalm@neuro.gu.se
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 539
Author: Brown, J. C., Golbik, R., Mann, K. and Timpl, R.
Year: 1994
Title: Structure and stability of the triple-helical domains of human collagen XIV
Journal: Matrix Biol
Volume: 14
Issue: 4
Pages: 287-95
Epub Date: 1994/08/01
Date: Aug
Short Title: Structure and stability of the triple-helical domains of human collagen XIV
Alternate Journal: Matrix biology : journal of the International Society for Matrix Biology
ISSN: 0945-053X (Print)
0945-053x
DOI: 10.1016/0945-053x(94)90194-5
Accession Number: 7827751
Keywords: Amino Acid Sequence
Animals
Cattle
Chickens
Circular Dichroism
Collagen/*chemistry
Glycosylation
Hot Temperature
Humans
Hydroxylation
Molecular Sequence Data
*Protein Conformation
Protein Denaturation
Protein Folding
Protein Processing, Post-Translational
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: Two triple-helical domains, Col 1 and Col 2, were obtained from a pepsin digest of human placental collagen XIV and separated from each other under nondenaturing conditions. Edman degradation demonstrated 106 amino acids residues in the Col 1 and 149 residues in the Col 2 domain. All except one of the 37 prolines in the Yaa position of the Gly-Xaa-Yaa triplets were completely hydroxylated to 4-hydroxyproline, and there were three imperfections in the triplet repeat. Partial or complete hydroxylation and glycosylation were found for all seven lysines in the Yaa position. Domain Col 1 was joined by disulfide bonds into a trimer, while Col 2 appeared as a mixture of monomers and disulfide-linked dimers. Circular dichroic spectra were typical for the collagen triple helix and revealed relatively high melting temperatures for Col 1 (38 degrees C) and Col 2 (43 degrees C). An almost perfect refolding of the triple helix was observed for Col 1 but not for Col 2, emphasizing the importance of disulfide bonds for the folding kinetics and in part the stability of the triple helix. Circular dichroic spectra of the large nontriple helical domain, NC3, of collagen XIV indicated 11% alpha helix and 63% beta structure. Comparative melting profiles of NC3 and intact collagen XIV indicated that the triple helices in intact collagen XIV have a melting temperature of 44 degrees C.
Notes: Brown, J C
Golbik, R
Mann, K
Timpl, R
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Matrix Biol. 1994 Aug;14(4):287-95. doi: 10.1016/0945-053x(94)90194-5.
Author Address: Max-Planck-Institut für Biochemie, Martinsried, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1085
Author: Brumshtein, B., Wormald, M. R., Silman, I., Futerman, A. H. and Sussman, J. L.
Year: 2006
Title: Structural comparison of differently glycosylated forms of acid-beta-glucosidase, the defective enzyme in Gaucher disease
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 62
Issue: Pt 12
Pages: 1458-65
Epub Date: 2006/12/02
Date: Dec
Short Title: Structural comparison of differently glycosylated forms of acid-beta-glucosidase, the defective enzyme in Gaucher disease
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s0907444906038303
Accession Number: 17139081
Keywords: Amino Acid Sequence
Binding Sites
Catalysis
Crystallography, X-Ray
Endo-1,4-beta Xylanases/chemistry/metabolism
Gaucher Disease/*enzymology
Glucosylceramidase/*chemistry/*metabolism
Glycosylation
Humans
Isoenzymes/chemistry/metabolism
Molecular Sequence Data
Protein Structure, Secondary
Abstract: Gaucher disease is caused by mutations in the gene encoding acid-beta-glucosidase. A recombinant form of this enzyme, Cerezyme, is used to treat Gaucher disease patients by ;enzyme-replacement therapy'. Crystals of Cerezyme after its partial deglycosylation were obtained earlier and the structure was solved to 2.0 A resolution [Dvir et al. (2003), EMBO Rep. 4, 704-709]. The crystal structure of unmodified Cerezyme is now reported, in which a substantial number of sugar residues bound to three asparagines via N-glycosylation could be visualized. The structure of intact fully glycosylated Cerezyme is virtually identical to that of the partially deglycosylated enzyme. However, the three loops at the entrance to the active site, which were previously observed in alternative conformations, display additional variability in their structures. Comparison of the structure of acid-beta-glucosidase with that of xylanase, a bacterial enzyme from a closely related protein family, demonstrates a close correspondence between the active-site residues of the two enzymes.
Notes: Brumshtein, Boris
Wormald, Mark R
Silman, Israel
Futerman, Anthony H
Sussman, Joel L
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Acta Crystallogr D Biol Crystallogr. 2006 Dec;62(Pt 12):1458-65. doi: 10.1107/S0907444906038303. Epub 2006 Nov 23.
Author Address: Department of Structural Biology, Weizmann Institute of Science, Israel.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 304
Author: Buko, A. M., Kentzer, E. J., Petros, A., Menon, G., Zuiderweg, E. R. and Sarin, V. K.
Year: 1991
Title: Characterization of a posttranslational fucosylation in the growth factor domain of urinary plasminogen activator
Journal: Proc Natl Acad Sci U S A
Volume: 88
Issue: 9
Pages: 3992-6
Epub Date: 1991/05/01
Date: May 1
Short Title: Characterization of a posttranslational fucosylation in the growth factor domain of urinary plasminogen activator
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.88.9.3992
PMCID: PMC51579
Accession Number: 2023947
Keywords: Amino Acid Sequence
Animals
Cell Line
Fucose/metabolism
Glycosylation
Magnetic Resonance Spectroscopy
Mice
Molecular Sequence Data
Peptide Mapping
Protein Processing, Post-Translational
Recombinant Proteins/metabolism
Urokinase-Type Plasminogen Activator/*metabolism
Abstract: A posttranslational modification site in natural and recombinant urinary-type plasminogen activators (urokinases; EC 3.4.21.31) has been localized to Thr-18, in the growth factor domain of the molecule. This is the region of urinary plasminogen activator responsible for its specific receptor binding. An unusual carbohydrate-protein linkage, a single monosaccharide, fucose, covalently attached directly to threonine in the peptide, is described here. The glycan moiety and the site of modification have been identified with mass spectrometry and confirmed by carbohydrate composition analysis, Edman degradation, and one- and two-dimensional NMR studies. This type of modification is normally not detected without mass spectrometry because the fucose-threonine bond is hydrolyzed under standard acidic conditions of the amino acid analysis and Edman sequencing. This modification may be widely found in other proteins.
Notes: 1091-6490
Buko, A M
Kentzer, E J
Petros, A
Menon, G
Zuiderweg, E R
Sarin, V K
Journal Article
Proc Natl Acad Sci U S A. 1991 May 1;88(9):3992-6. doi: 10.1073/pnas.88.9.3992.
Author Address: Analytical Research Department, Abbott Laboratories, North Chicago, IL 60064.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1630
Author: Bullen, J. W., Balsbaugh, J. L., Chanda, D., Shabanowitz, J., Hunt, D. F., Neumann, D. and Hart, G. W.
Year: 2014
Title: Cross-talk between two essential nutrient-sensitive enzymes: O-GlcNAc transferase (OGT) and AMP-activated protein kinase (AMPK)
Journal: J Biol Chem
Volume: 289
Issue: 15
Pages: 10592-10606
Epub Date: 2014/02/25
Date: Apr 11
Short Title: Cross-talk between two essential nutrient-sensitive enzymes: O-GlcNAc transferase (OGT) and AMP-activated protein kinase (AMPK)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.523068
PMCID: PMC4036179
Accession Number: 24563466
Keywords: AMP-Activated Protein Kinases/*metabolism
Acetylglucosamine/metabolism
Animals
Cell Differentiation
Cell Line
Cell Nucleus/metabolism
Cell Proliferation
Cytoplasm/metabolism
*Gene Expression Regulation, Enzymologic
HEK293 Cells
HeLa Cells
Humans
Mass Spectrometry
Mice
Microscopy, Confocal
Muscle Fibers, Skeletal/metabolism
N-Acetylglucosaminyltransferases/*metabolism
Phosphorylation
Protein Processing, Post-Translational
Signal Transduction
Substrate Specificity
AMP-activated Kinase (AMPK)
Histones
Nuclear Translocation
Nutrient Sensing
O-GlcNAc
O-GlcNAc Transferase
O-GlcNAcylation
Skeletal Muscle
Abstract: Nutrient-sensitive pathways regulate both O-GlcNAc transferase (OGT) and AMP-activated protein kinase (AMPK), cooperatively connecting metabolic homeostasis to regulation of numerous intracellular processes essential for life. Similar to phosphorylation, catalyzed by kinases such as AMPK, O-GlcNAcylation is a highly dynamic Ser/Thr-specific post-translational modification of nuclear, cytoplasmic, and mitochondrial proteins catalyzed exclusively by OGT. OGT and AMPK target a multitude of intracellular proteins, with the net effect to protect cells from the damaging effects of metabolic stress. Despite hundreds of studies demonstrating significant overlap in upstream and downstream signaling processes, no study has investigated if OGT and AMPK can directly regulate each other. We show acute activation of AMPK alters the substrate selectivity of OGT in several cell lines and nuclear localization of OGT in C2C12 skeletal muscle myotubes. Nuclear localization of OGT affects O-GlcNAcylation of numerous nuclear proteins and acetylation of Lys-9 on histone 3 in myotubes. AMPK phosphorylates Thr-444 on OGT in vitro; phosphorylation of Thr-444 is tightly associated with AMPK activity and nuclear localization of OGT in myotubes, and phospho-mimetic T444E-OGT exhibits altered substrate selectivity. Conversely, the α- and γ-subunits of AMPK are O-GlcNAcylated, O-GlcNAcylation of the γ1-subunit increases with AMPK activity, and acute inhibition of O-GlcNAc cycling disrupts activation of AMPK. We have demonstrated significant cross-talk between the O-GlcNAc and AMPK systems, suggesting OGT and AMPK may cooperatively regulate nutrient-sensitive intracellular processes that mediate cellular metabolism, growth, proliferation, and/or tissue function.
Notes: 1083-351x
Bullen, John W
Balsbaugh, Jeremy L
Chanda, Dipanjan
Shabanowitz, Jeffrey
Hunt, Donald F
Neumann, Dietbert
Hart, Gerald W
R01CA42486/CA/NCI NIH HHS/United States
GM037537/GM/NIGMS NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
N01-HV-00240/HV/NHLBI NIH HHS/United States
R01 GM037537/GM/NIGMS NIH HHS/United States
R01DK61671/DK/NIDDK NIH HHS/United States
R24DK084949/DK/NIDDK NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
R24 DK084949/DK/NIDDK NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
P01HL107153/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2014 Apr 11;289(15):10592-10606. doi: 10.1074/jbc.M113.523068. Epub 2014 Feb 21.
Author Address: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Department of Chemistry, University of Virginia, Charlottesville, Virginia 22904.
Department of Molecular Genetics, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, 6200 MD Maastricht, The Netherlands.
Department of Chemistry, University of Virginia, Charlottesville, Virginia 22904; Department of Pathology, University of Virginia, Charlottesville, Virginia 22904.
Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205. Electronic address: gwhart@jhmi.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 796
Author: Bungert, S., Molday, L. L. and Molday, R. S.
Year: 2001
Title: Membrane topology of the ATP binding cassette transporter ABCR and its relationship to ABC1 and related ABCA transporters: identification of N-linked glycosylation sites
Journal: J Biol Chem
Volume: 276
Issue: 26
Pages: 23539-46
Epub Date: 2001/04/26
Date: Jun 29
Short Title: Membrane topology of the ATP binding cassette transporter ABCR and its relationship to ABC1 and related ABCA transporters: identification of N-linked glycosylation sites
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M101902200
Accession Number: 11320094
Keywords: ATP Binding Cassette Transporter 1
ATP-Binding Cassette Transporters/*chemistry/genetics/metabolism
Amino Acid Sequence
Animals
COS Cells
Cattle
Concanavalin A/metabolism
Consensus Sequence
Disulfides/metabolism
Glycoproteins/*chemistry
Glycosylation
Humans
Models, Biological
Molecular Sequence Data
Protein Structure, Tertiary
Rod Cell Outer Segment/metabolism
Sequence Deletion
Sequence Homology, Amino Acid
Transfection
Trypsin/chemistry
Abstract: ABCR is a member of the ABCA subclass of ATP binding cassette transporters that is responsible for Stargardt macular disease and implicated in retinal transport across photoreceptor disc membranes. It consists of a single polypeptide chain arranged in two tandem halves, each having a multi-spanning membrane domain followed by a nucleotide binding domain. To delineate between several proposed membrane topological models, we have identified the exocytoplasmic (extracellular/lumen) N-linked glycosylation sites on ABCR. Using trypsin digestion, site-directed mutagenesis, concanavalin A binding, and endoglycosidase digestion, we show that ABCR contains eight glycosylation sites. Four sites reside in a 600-amino acid exocytoplasmic domain of the N-terminal half between the first transmembrane segment H1 and the first multi-spanning membrane domain, and four sites are in a 275-amino acid domain of the C half between transmembrane segment H7 and the second multi-spanning membrane domain. This leads to a model in which each half has a transmembrane segment followed by a large exocytoplasmic domain, a multi-spanning membrane domain, and a nucleotide binding domain. Other ABCA transporters, including ABC1 linked to Tangier disease, are proposed to have a similar membrane topology based on sequence similarity to ABCR. Studies also suggest that the N and C halves of ABCR are linked through disulfide bonds.
Notes: Bungert, S
Molday, L L
Molday, R S
EY02422/EY/NEI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2001 Jun 29;276(26):23539-46. doi: 10.1074/jbc.M101902200. Epub 2001 Apr 24.
Author Address: Departments of Biochemistry & Molecular Biology and Ophthalmology, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 923
Author: Bunkenborg, J., Pilch, B. J., Podtelejnikov, A. V. and Wiśniewski, J. R.
Year: 2004
Title: Screening for N-glycosylated proteins by liquid chromatography mass spectrometry
Journal: Proteomics
Volume: 4
Issue: 2
Pages: 454-65
Epub Date: 2004/02/05
Date: Feb
Short Title: Screening for N-glycosylated proteins by liquid chromatography mass spectrometry
Alternate Journal: Proteomics
ISSN: 1615-9853 (Print)
1615-9853
DOI: 10.1002/pmic.200300556
Accession Number: 14760718
Keywords: Amino Acid Sequence
Blood Proteins/*analysis/chemistry
Chromatography, Liquid
Concanavalin A/*chemistry
Glycoproteins/*analysis/blood/chemistry
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/*analysis/blood/chemistry
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
Wheat Germ Agglutinins/*chemistry
Abstract: In the last few years mass spectrometry has become the method of choice for characterization of post-translationally modified proteins. Whereas most protein chemical modifications are binary in the sense that only one change can be associated with a given residue, many different oligosaccharides can be attached to a glycosylation site residue. The detailed characterization of glycoproteins in complex biological samples is extremely challenging. However, information on N-glycosylation can be gained at an intermediary level. Here we demonstrate a procedure for mapping N-glycosylation sites in complex mixtures by reducing sample complexity and enriching glycoprotein content. Glycosylated proteins are selected by an initial lectin chromatography step and digested with endoproteinase Lys-C. Glycosylated peptides are then selected from the digest mixture by a second lectin chromatography step. The glycan components are removed with N-glycosidase F and the peptides digested with trypsin before analysis by on-line reversed-phase liquid chromatography mass spectrometry. Using two different lectins, concanavalin A and wheat germ agglutinin, this procedure was applied to human serum and a total of 86 N-glycosylation sites in 77 proteins were identified.
Notes: Bunkenborg, Jakob
Pilch, Bartosz J
Podtelejnikov, Alexandre V
Wiśniewski, Jacek R
Journal Article
Germany
Proteomics. 2004 Feb;4(2):454-65. doi: 10.1002/pmic.200300556.
Author Address: MDS Proteomics Odense, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1801
Author: Burén, S., Gomes, A. L., Teijeiro, A., Fawal, M. A., Yilmaz, M., Tummala, K. S., Perez, M., Rodriguez-Justo, M., Campos-Olivas, R., Megías, D. and Djouder, N.
Year: 2016
Title: Regulation of OGT by URI in Response to Glucose Confers c-MYC-Dependent Survival Mechanisms
Journal: Cancer Cell
Volume: 30
Issue: 2
Pages: 290-307
Epub Date: 2016/08/10
Date: Aug 8
Short Title: Regulation of OGT by URI in Response to Glucose Confers c-MYC-Dependent Survival Mechanisms
Alternate Journal: Cancer cell
ISSN: 1535-6108
DOI: 10.1016/j.ccell.2016.06.023
Accession Number: 27505673
Keywords: Animals
Glucose/administration & dosage/*metabolism
Glucose Tolerance Test
HEK293 Cells
HeLa Cells
Humans
Intracellular Signaling Peptides and Proteins/*metabolism
Liver Neoplasms/genetics/*metabolism
Mice
N-Acetylglucosaminyltransferases/*metabolism
Proto-Oncogene Proteins c-myc/*metabolism
RNA, Small Interfering/genetics/metabolism
Repressor Proteins
Transfection
Abstract: Cancer cells can adapt and survive under low nutrient conditions, but underlying mechanisms remain poorly explored. We demonstrate here that glucose maintains a functional complex between the co-chaperone URI, PP1γ, and OGT, the enzyme catalyzing O-GlcNAcylation. Glucose deprivation induces the activation of PKA, which phosphorylates URI at Ser-371, resulting in PP1γ release and URI-mediated OGT inhibition. Low OGT activity reduces O-GlcNAcylation and promotes c-MYC degradation to maintain cell survival. In the presence of glucose, PP1γ-bound URI increases OGT and c-MYC levels. Accordingly, mice expressing non-phosphorylatable URI (S371A) in hepatocytes exhibit high OGT activity and c-MYC stabilization, accelerating liver tumorigenesis in agreement with c-MYC oncogenic functions. Our work uncovers that URI-regulated OGT confers c-MYC-dependent survival functions in response to glucose fluctuations.
Notes: 1878-3686
Burén, Stefan
Gomes, Ana L
Teijeiro, Ana
Fawal, Mohamad-Ali
Yilmaz, Mahmut
Tummala, Krishna S
Perez, Manuel
Rodriguez-Justo, Manuel
Campos-Olivas, Ramón
Megías, Diego
Djouder, Nabil
11-0242/Worldwide Cancer Research/United Kingdom
Journal Article
United States
Cancer Cell. 2016 Aug 8;30(2):290-307. doi: 10.1016/j.ccell.2016.06.023.
Author Address: Cancer Cell Biology Programme, Growth Factors, Nutrients and Cancer Group, Centro Nacional de Investigaciones Oncológicas, CNIO, Madrid 28029, Spain.
Biotechnology Programme, Confocal Microscopy Core Unit, Centro Nacional de Investigaciones Oncológicas, CNIO, Madrid 28029, Spain.
Department of Research Pathology, Cancer Institute, University College London, London WC1E 6JJ, UK.
Structural Biology and Biocomputing Programme, Spectroscopy and Nuclear Magnetic Resonance Unit, Centro Nacional de Investigaciones Oncológicas, CNIO, Madrid 28029, Spain.
Cancer Cell Biology Programme, Growth Factors, Nutrients and Cancer Group, Centro Nacional de Investigaciones Oncológicas, CNIO, Madrid 28029, Spain. Electronic address: ndjouder@cnio.es.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 503
Author: Burger, D., Pidoux, L. and Steck, A. J.
Year: 1993
Title: Identification of the glycosylated sequons of human myelin-associated glycoprotein
Journal: Biochem Biophys Res Commun
Volume: 197
Issue: 2
Pages: 457-64
Epub Date: 1993/12/15
Date: Dec 15
Short Title: Identification of the glycosylated sequons of human myelin-associated glycoprotein
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1006/bbrc.1993.2501
Accession Number: 7505568
Keywords: Amino Acid Sequence
Asparagine
Cell Adhesion Molecules, Neuronal/chemistry/metabolism
Chromatography, Affinity
Chromatography, Gel
Chromatography, High Pressure Liquid
Glycopeptides/chemistry/isolation & purification
Glycosylation
Humans
Molecular Sequence Data
Myelin Proteins/*chemistry/isolation & purification/metabolism
Myelin-Associated Glycoprotein
Abstract: Myelin-associated glycoprotein (MAG) is a neural cell adhesion molecule expressing the L2/HNK-1 carbohydrate epitope. MAG is heavily glycosylated containing 30% carbohydrate by weight. In this study, human MAG glycopeptides were isolated and sequenced. Of the 9 MAG sequons 7 were glycosylated and 1 was partially glycosylated at Asn106. Asn332 which was not recovered in the glycopeptide fractions was probably not glycosylated. Furthermore, preliminary data indicate that all MAG glycosylated sequons might bear the L2/HNK-1 epitope.
Notes: Burger, D
Pidoux, L
Steck, A J
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 1993 Dec 15;197(2):457-64. doi: 10.1006/bbrc.1993.2501.
Author Address: Department of Neurology, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1793
Author: Bürgi, J., Xue, B., Uversky, V. N. and van der Goot, F. G.
Year: 2016
Title: Intrinsic Disorder in Transmembrane Proteins: Roles in Signaling and Topology Prediction
Journal: PLoS One
Volume: 11
Issue: 7
Pages: e0158594
Epub Date: 2016/07/09
Short Title: Intrinsic Disorder in Transmembrane Proteins: Roles in Signaling and Topology Prediction
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0158594
PMCID: PMC4938508
Accession Number: 27391701
Keywords: Algorithms
Cell Adhesion/genetics/physiology
Computational Biology
Fluorescent Antibody Technique
HeLa Cells
Humans
Membrane Proteins/*chemistry
Phosphorylation
Plasmids/genetics
Protein Binding/genetics/physiology
Protein Conformation
Protein Processing, Post-Translational
Ubiquitination
Abstract: Intrinsically disordered regions (IDRs) are peculiar stretches of amino acids that lack stable conformations in solution. Intrinsic Disorder containing Proteins (IDP) are defined by the presence of at least one large IDR and have been linked to multiple cellular processes including cell signaling, DNA binding and cancer. Here we used computational analyses and publicly available databases to deepen insight into the prevalence and function of IDRs specifically in transmembrane proteins, which are somewhat neglected in most studies. We found that 50% of transmembrane proteins have at least one IDR of 30 amino acids or more. Interestingly, these domains preferentially localize to the cytoplasmic side especially of multi-pass transmembrane proteins, suggesting that disorder prediction could increase the confidence of topology prediction algorithms. This was supported by the successful prediction of the topology of the uncharacterized multi-pass transmembrane protein TMEM117, as confirmed experimentally. Pathway analysis indicated that IDPs are enriched in cell projection and axons and appear to play an important role in cell adhesion, signaling and ion binding. In addition, we found that IDP are enriched in phosphorylation sites, a crucial post translational modification in signal transduction, when compared to fully ordered proteins and to be implicated in more protein-protein interaction events. Accordingly, IDPs were highly enriched in short protein binding regions called Molecular Recognition Features (MoRFs). Altogether our analyses strongly support the notion that the transmembrane IDPs act as hubs in cellular signal events.
Notes: 1932-6203
Bürgi, Jérôme
Xue, Bin
Uversky, Vladimir N
van der Goot, F Gisou
Orcid: 0000-0002-8522-274x
Journal Article
PLoS One. 2016 Jul 8;11(7):e0158594. doi: 10.1371/journal.pone.0158594. eCollection 2016.
Author Address: Faculty of life science, Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.
Department of Cell Biology, Microbiology, and Molecular Biology, School of Natural Sciences and Mathematics, College of Arts and Sciences, University of South Florida, Tampa, FL, 33620, United States of America.
Department of Molecular Medicine and USF Health Byrd Alzheimer's Research Institute, Morsani College of Medicine, University of South Florida, Tampa, FL, 33647, United States of America.
Institute for Biological Instrumentation, Russian Academy of Sciences, Pushchino, Moscow Region, Russian Federation.
Department of Biology, Faculty of Science, King Abdulaziz University, P.O. Box 80203, Jeddah, 21589, Kingdom of Saudi Arabia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1265
Author: Bussink, A. P., Verhoek, M., Vreede, J., Ghauharali-van der Vlugt, K., Donker-Koopman, W. E., Sprenger, R. R., Hollak, C. E., Aerts, J. M. and Boot, R. G.
Year: 2009
Title: Common G102S polymorphism in chitotriosidase differentially affects activity towards 4-methylumbelliferyl substrates
Journal: Febs j
Volume: 276
Issue: 19
Pages: 5678-88
Epub Date: 2009/09/04
Date: Oct
Short Title: Common G102S polymorphism in chitotriosidase differentially affects activity towards 4-methylumbelliferyl substrates
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/j.1742-4658.2009.07259.x
Accession Number: 19725875
Keywords: Alleles
Amino Acid Substitution
Base Sequence
DNA Primers/genetics
Gaucher Disease/enzymology/genetics
Gene Frequency
Glycosylation
Hexosaminidases/*genetics/*metabolism
Humans
Hymecromone/*analogs & derivatives/metabolism
In Vitro Techniques
Models, Molecular
Netherlands
*Polymorphism, Single Nucleotide
Protein Conformation
Recombinant Proteins/genetics/metabolism
Substrate Specificity
Thermodynamics
Trisaccharides/metabolism
Umbelliferones/metabolism
Abstract: Chitotriosidase (CHIT1) is a chitinase that is secreted by activated macrophages. Plasma chitotriosidase activity reflects the presence of lipid-laden macrophages in patients with Gaucher disease. CHIT1 activity can be conveniently measured using fluorogenic 4-methylumbelliferyl (4MU)-chitotrioside or 4MU-chitobioside as the substrate, however, nonsaturating concentrations have to be used because of apparent substrate inhibition. Saturating substrate concentrations can, however, be used with the newly designed substrate 4MU-deoxychitobioside. We studied the impact of a known polymorphism, G102S, on the catalytic properties of CHIT1. The G102S allele was found to be common in type I Gaucher disease patients in the Netherlands ( approximately 24% of alleles). The catalytic efficiency of recombinant Ser102 CHIT1 was approximately 70% that of wild-type Gly102 CHIT1 when measured with 4MU-chitotrioside at a nonsaturating concentration. However, the activity was normal with 4MU-deoxychitobioside as the substrate at saturating concentrations, consistent with predictions from molecular dynamics simulations. In conclusion, interpretation of CHIT1 activity measurements with 4MU-chitotrioside with respect to CHIT1 protein concentrations depends on the presence of Ser102 CHIT1 in an individual, complicating estimation of the body burden of storage macrophages. Use of the superior 4MU-deoxychitobioside substrate avoids such complications because activity towards this substrate under saturating conditions is not affected by the G102S substitution.
Notes: 1742-4658
Bussink, Anton P
Verhoek, Marri
Vreede, Jocelyne
Ghauharali-van der Vlugt, Karen
Donker-Koopman, Wilma E
Sprenger, Richard R
Hollak, Carla E
Aerts, Johannes M F G
Boot, Rolf G
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2009 Oct;276(19):5678-88. doi: 10.1111/j.1742-4658.2009.07259.x. Epub 2009 Sep 2.
Author Address: Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 225
Author: Butkowski, R. J., Elion, J., Downing, M. R. and Mann, K. G.
Year: 1977
Title: Primary structure of human prethrombin 2 and alpha-thrombin
Journal: J Biol Chem
Volume: 252
Issue: 14
Pages: 4942-57
Epub Date: 1977/07/25
Date: Jul 25
Short Title: Primary structure of human prethrombin 2 and alpha-thrombin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 873923
Keywords: Amino Acid Sequence
Binding Sites
*Enzyme Precursors
Humans
Peptide Fragments
Prothrombin
*Thrombin
Notes: Butkowski, R J
Elion, J
Downing, M R
Mann, K G
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1977 Jul 25;252(14):4942-57.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 448
Author: Butler, W. T.
Year: 1970
Title: Chemical studies on the cyanogen bromide peptides of rat skin collagen. The covalent structure of alpha 1-CB5, the major hexose-containing cyanogen bromide peptide of alpha 1
Journal: Biochemistry
Volume: 9
Issue: 1
Pages: 44-50
Epub Date: 1970/01/06
Date: Jan 6
Short Title: Chemical studies on the cyanogen bromide peptides of rat skin collagen. The covalent structure of alpha 1-CB5, the major hexose-containing cyanogen bromide peptide of alpha 1
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00803a006
Accession Number: 5411206
Keywords: Amino Acid Sequence
Amino Acids/analysis
Animals
Chromatography, Gel
Chromatography, Ion Exchange
Chromatography, Paper
Collagen/*analysis
Cyanides
Electrophoresis
Galactose/analysis
Glucose/analysis
Hexoses/analysis
Imines/analysis
Lathyrism
Peptides/analysis
Rats
Skin/*analysis
Trypsin
Notes: Butler, W T
Journal Article
United States
Biochemistry. 1970 Jan 6;9(1):44-50. doi: 10.1021/bi00803a006.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 893
Author: Calero, G., Gupta, P., Nonato, M. C., Tandel, S., Biehl, E. R., Hofmann, S. L. and Clardy, J.
Year: 2003
Title: The crystal structure of palmitoyl protein thioesterase-2 (PPT2) reveals the basis for divergent substrate specificities of the two lysosomal thioesterases, PPT1 and PPT2
Journal: J Biol Chem
Volume: 278
Issue: 39
Pages: 37957-64
Epub Date: 2003/07/12
Date: Sep 26
Short Title: The crystal structure of palmitoyl protein thioesterase-2 (PPT2) reveals the basis for divergent substrate specificities of the two lysosomal thioesterases, PPT1 and PPT2
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M301225200
Accession Number: 12855696
Keywords: Amino Acid Sequence
Animals
Binding Sites
COS Cells
Catalysis
Crystallization
Humans
Molecular Sequence Data
Substrate Specificity
Thiolester Hydrolases/*chemistry/metabolism
Abstract: Mutations in palmitoyl protein thioesterase-1 (PPT1) have been found to cause the infantile form of neuronal ceroid lipofuscinosis, which is a lysosomal storage disorder characterized by impaired degradation of fatty acid-modified proteins with accumulation of amorphous granular deposits in cortical neurons, leading to mental retardation and death. Palmitoyl protein thioesterase-2 (PPT2) is a second lysosomal hydrolase that shares a 26% identity with PPT1. A previous study had suggested that palmitoyl-CoA was the preferred substrate of PPT2. Furthermore, PPT2 did not hydrolyze palmitate from the several S-palmitoylated protein substrates. Interestingly, PPT2 deficiency in a recent transgenic mouse model is associated with a form of neuronal ceroid lipofuscinosis, suggesting that PPT1 and -2 perform non-redundant roles in lysosomal thioester catabolism. In the current paper, we present the crystal structure of PPT2 at a resolution of 2.7 A. Comparisons of the structures of PPT1 and -2 show very similar architectural features; however, conformational differences in helix alpha4 lead to a solvent-exposed lipid-binding groove in PPT1. The limited space between two parallel loops (beta3-alphaA and beta8-alphaF) located immediately above the lipid-binding groove in PPT2 restricts the binding of fatty acids with bulky head groups, and this binding groove is significantly larger in PPT1. This structural difference accounts for the ability of PPT2 to hydrolyze an unbranched structure such as palmitoyl-CoA but not palmitoylcysteine or palmitoylated proteins. Furthermore, differences in fatty acid chain length specificity of PPT1 and -2, also reported here, are explained by the structure and may provide a biochemical basis for their non-redundant roles.
Notes: Calero, Guillermo
Gupta, Praveena
Nonato, M Cristina
Tandel, Sagun
Biehl, Edward R
Hofmann, Sandra L
Clardy, Jon
CA59021/CA/NCI NIH HHS/United States
NS35323/NS/NINDS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2003 Sep 26;278(39):37957-64. doi: 10.1074/jbc.M301225200. Epub 2003 Jul 10.
Author Address: Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853-1301, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 361
Author: Calvete, J. J., Henschen, A. and González-Rodríguez, J.
Year: 1989
Title: Complete localization of the intrachain disulphide bonds and the N-glycosylation points in the alpha-subunit of human platelet glycoprotein IIb
Journal: Biochem J
Volume: 261
Issue: 2
Pages: 561-8
Epub Date: 1989/07/15
Date: Jul 15
Short Title: Complete localization of the intrachain disulphide bonds and the N-glycosylation points in the alpha-subunit of human platelet glycoprotein IIb
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2610561
PMCID: PMC1138861
Accession Number: 2775232
Keywords: Amino Acid Sequence
Cysteine/blood
Disulfides/*blood
Glycosylation
Humans
Molecular Sequence Data
Peptide Fragments/blood
Platelet Membrane Glycoproteins/*isolation & purification
Abstract: Glycoprotein IIb (GPIIb), one of the two molecular components of the inducible receptor for fibrinogen on the platelet surface, is formed from two subunits, GPIIb alpha (114 kDa) and GPIIb beta (22.5 kDa), joined by a single disulphide bond. CNBr cleavage of GPIIb, together with tryptic or endoproteinase Lys-C digestion of some of the isolated CNBr peptides, followed by amino acid and N-terminal sequence analysis of the isolated fragments, allowed us to locate unambiguously all the unknown disulphide bonds and the N-glycosylation points in platelet GPIIb. It could be established that each cysteine residue in GPIIb, beginning at alpha-Cys-56, is disulphide-bonded to its nearest neighbour in the amino acid sequence. Given the extensive structural similarity among the two-chain alpha-subunits of Arg-Gly-Asp adhesion receptors and the conservative positions of cysteine residues in their amino acid sequences, the intrachain and interchain disulphide-bond pattern found here in GPIIb will most probably be conserved in all two-chain alpha-subunits of these receptors. The N-linked glycosylation points found here in platelet GPIIb are the same as the five N-glycosylated asparagine residues suggested after cDNA sequencing of human erythroleukaemic-cell GPIIb [Poncz, Eisman, Heindenreich, Silver, Vilaire, Surrey, Schwartz & Bennett (1987) J. Biol. Chem. 262, 8476-8482]. Some of the general features of the structure of GPIIb, such as the existence of distinct domains in the alpha- and beta-subunits, as well as the identification of well-defined points in its external topography, are discussed.
Notes: 1470-8728
Calvete, J J
Henschen, A
González-Rodríguez, J
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1989 Jul 15;261(2):561-8. doi: 10.1042/bj2610561.
Author Address: Max-Planck-Institut für Biochemie, Martinsried/München, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 528
Author: Calvete, J. J. and Muñiz-Diaz, E.
Year: 1993
Title: Localization of an O-glycosylation site in the alpha-subunit of the human platelet integrin GPIIb/IIIa involved in Baka (HPA-3a) alloantigen expression
Journal: FEBS Lett
Volume: 328
Issue: 1-2
Pages: 30-4
Epub Date: 1993/08/09
Date: Aug 9
Short Title: Localization of an O-glycosylation site in the alpha-subunit of the human platelet integrin GPIIb/IIIa involved in Baka (HPA-3a) alloantigen expression
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(93)80959-x
Accession Number: 7688323
Keywords: Amino Acid Sequence
Antigens, Human Platelet/*chemistry/immunology
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Enzyme-Linked Immunosorbent Assay
Epitopes/immunology
Glycosylation
Humans
Isoantigens/*analysis
Molecular Sequence Data
Platelet Membrane Glycoproteins/*chemistry
Serine/*chemistry
Abstract: The human alloantigen system Baka/b is associated with a Ile843-->Ser replacement on platelet glycoprotein IIb, the alpha-subunit of the integrin receptor for fibrinogen (GPIIb/IIIa). Recent immunological studies indicate that sialylated oligosaccharide chain(s) are also implicated in expression of the Baka determinant. Here we show that the GPIIb fragment 704-856 contains the whole Baka epitope, and that chemical cleavage of a single O-linked oligosaccharide chain within this GPIIb domain correlates with the loss of its anti-Baka antibodies binding ability. Ser847 was identified as the O-glycosylation site. Therefore, our results show that the Ser847 modification is responsible for the expression of the GPIIb-specific Baka alloantigen, and provide thus a link between the molecular biology and the immunologic observations.
Notes: Calvete, J J
Muñiz-Diaz, E
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1993 Aug 9;328(1-2):30-4. doi: 10.1016/0014-5793(93)80959-x.
Author Address: Instituto de Química-Física CSIC, Madrid, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 799
Author: Camara-Clayette, V., Rahuel, C., Lopez, C., Hattab, C., Verkarre, V., Bertrand, O. and Cartron, J. P.
Year: 2001
Title: Transcriptional regulation of the KEL gene and Kell protein expression in erythroid and non-erythroid cells
Journal: Biochem J
Volume: 356
Issue: Pt 1
Pages: 171-80
Epub Date: 2001/05/05
Date: May 15
Short Title: Transcriptional regulation of the KEL gene and Kell protein expression in erythroid and non-erythroid cells
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/0264-6021:3560171
PMCID: PMC1221825
Accession Number: 11336649
Keywords: *Antigens, Nuclear
Chloramphenicol O-Acetyltransferase
*DNA Helicases
DNA-Binding Proteins
Erythroid Precursor Cells
Gene Expression Regulation
Genes, Reporter
HeLa Cells
Humans
K562 Cells
Kell Blood-Group System/*genetics
Ku Autoantigen
Luciferases
Lymphoid Tissue
Male
Nuclear Proteins
Point Mutation
Promoter Regions, Genetic
Protein Binding
Testis
Tissue Distribution
Transcription Factors/metabolism
Abstract: The Kell blood-group antigen was originally reported to be a protein expressed in erythroid tissue only. Transcriptional analysis of the KEL promoter activity in human erythroleukaemia K562 and epithelial HeLa cells by electrophoretic mobility-shift and supershift assays, chloramphenicol acetyltransferase assays, co-transfection studies and site-directed mutagenesis provided the following results: (i) the KEL promoter exhibits a strong transcriptional activity in K562 cells and, unexpectedly, a basal non-erythroid activity in HeLa cells, (ii) up-regulation of the 5' distal promoter activity occurs only in the erythroid context, and (iii) two motifs localized in the exon 1 region, which bind the Sp1/Sp3 and the human GATA-1/Ku70/80 factors, were required for down-regulation of the promoter activity, but inhibition of the promoter activity by the repressing factors in HeLa cells was incomplete. KEL expression in HeLa cells was performed further by primer-extension analysis, which revealed the presence of a low amount of Kell transcript correlating with basal expression of the Kell protein in these cells, as shown by immunopurification and Western-blot analysis. DNA sequencing of the transcript revealed a sequence identical to that obtained from erythroid tissue. In human tissues, KEL expression was investigated by dot-blot analysis and revealed high levels of Kell mRNAs, particularly in brain tissues, testis and lymphoid tissues. Moreover, most tissues analysed exhibited low levels of Kell transcripts. The Kell protein was also detected by immunohistochemistry in the Sertoli cells of the testis and in lymphoid tissues like spleen and tonsil, specifically localized in the follicular dendritic cells. Altogether, the results indicated that KEL expression is not restricted to erythroid tissue.
Notes: 1470-8728
Camara-Clayette, V
Rahuel, C
Lopez, C
Hattab, C
Verkarre, V
Bertrand, O
Cartron, J P
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2001 May 15;356(Pt 1):171-80. doi: 10.1042/0264-6021:3560171.
Author Address: INSERM U76, Institut National de la Transfusion Sanguine, 6, rue Alexandre Cabanel, 75015 Paris, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1593
Author: Cameron, A., Giacomozzi, B., Joyce, J., Gray, A., Graham, D., Ousson, S., Neny, M., Beher, D., Carlson, G., O'Moore, J., Shearman, M. and Hering, H.
Year: 2013
Title: Generation and characterization of a rabbit monoclonal antibody site-specific for tau O-GlcNAcylated at serine 400
Journal: FEBS Lett
Volume: 587
Issue: 22
Pages: 3722-8
Epub Date: 2013/10/12
Date: Nov 15
Short Title: Generation and characterization of a rabbit monoclonal antibody site-specific for tau O-GlcNAcylated at serine 400
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1016/j.febslet.2013.09.042
Accession Number: 24113653
Keywords: Acetylglucosamine/metabolism
Animals
Antibodies, Monoclonal/*chemistry
Antibody Specificity
Glycosylation
HEK293 Cells
Humans
Mice
Mice, Knockout
Protein Binding
*Protein Processing, Post-Translational
Rabbits
Serine/metabolism
tau Proteins/chemistry/*immunology/metabolism
Monoclonal antibody
O-GlcNAc
Oga
Site-specific
Tau
Abstract: Aggregation of tau into paired helical filaments is a pathological process leading to neurotoxicity in Alzheimer's disease and other tauopathies. Tau is posttranslationally modified by O-linked N-acetylglucosamine (O-GlcNAc), and increasing tau O-GlcNAcylation may protect against its aggregation. Research tools to study the relationship between tau aggregation and tau O-GlcNAcylation have not been widely available. Here we describe the generation of a rabbit monoclonal antibody specific for tau O-GlcNAcylated at Ser400 (O-tau(S400)). We show the utility of this antibody for in vitro and in vivo experiments to investigate the function of O-GlcNAc modifications of tau at Ser400.
Notes: 1873-3468
Cameron, Andrew
Giacomozzi, Brandy
Joyce, John
Gray, Audrey
Graham, Danielle
Ousson, Solenne
Neny, Maud
Beher, Dirk
Carlson, George
O'Moore, Jill
Shearman, Mark
Hering, Heike
Journal Article
England
FEBS Lett. 2013 Nov 15;587(22):3722-8. doi: 10.1016/j.febslet.2013.09.042. Epub 2013 Oct 7.
Author Address: EMD Serono Research & Development Institute, Billerica, MA 01821, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 449
Author: Campbell, D. G., Gagnon, J., Reid, K. B. and Williams, A. F.
Year: 1981
Title: Rat brain Thy-1 glycoprotein. The amino acid sequence, disulphide bonds and an unusual hydrophobic region
Journal: Biochem J
Volume: 195
Issue: 1
Pages: 15-30
Epub Date: 1981/04/01
Date: Apr 1
Short Title: Rat brain Thy-1 glycoprotein. The amino acid sequence, disulphide bonds and an unusual hydrophobic region
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj1950015
PMCID: PMC1162851
Accession Number: 6118137
Keywords: Amino Acid Sequence
Amino Acids/analysis
Animals
*Antigens, Surface/isolation & purification
*Brain Chemistry
Chromatography, Gel
Disulfides
Endopeptidases
*Glycoproteins/isolation & purification
*Membrane Proteins/isolation & purification
Peptide Fragments/isolation & purification
Rats
Thy-1 Antigens
Trypsin
Abstract: The full sequence of the Thy-1 membrane glycoprotein of rat brain is reported. The sequence was determined from tryptic and V-8 proteinase peptides and consisted of 111 amino acids. The amino terminus was blocked and consisted of a pyroglutamic acid residue. The molecule contained two disulphide bonds, namely Cys-9--Cys-111 and Cys-19--Cys-85. Three N-linked amino sugars were located at Asn-23, Asn-74 and Asn-98. In each case the sequence on the C-terminal side of the attachment point was Asn-Xaa-Thr as would be expected for N-linkage. The C-terminal peptides were unusual, in that they were either obtained in a highly aggregated form, or could only be purified after binding to Brij 96 micelles. Thus they appeared to have hydrophobic properties, yet did not contain any extended sequence of hydrophobic amino acids. Other unusual features of the C-terminal peptides were the presence of unidentified ninhydrin-positive material and of glucosamine and galactosamine. The C-terminal residue has not been directly identified but Cys-111 is the last conventional amino acid. It is suggested that the hydrophobic properties of the C-terminal peptides may be due to the linkage of lipid. The sequence of the Thy-1 glycoprotein showed homologies with immunoglobulin domains. This relationship is examined in detail in the paper following [Cohen et al. (1981) Biochem. J. 193, 000--000].
Notes: 1470-8728
Campbell, D G
Gagnon, J
Reid, K B
Williams, A F
Journal Article
Biochem J. 1981 Apr 1;195(1):15-30. doi: 10.1042/bj1950015.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 426
Author: Campbell, D. J., Bouhnik, J., Coezy, E., Menard, J. and Corvol, P.
Year: 1985
Title: Processing of rat and human angiotensinogen precursors by microsomal membranes
Journal: Mol Cell Endocrinol
Volume: 43
Issue: 1
Pages: 31-40
Epub Date: 1985/11/01
Date: Nov
Short Title: Processing of rat and human angiotensinogen precursors by microsomal membranes
Alternate Journal: Molecular and cellular endocrinology
ISSN: 0303-7207 (Print)
0303-7207
DOI: 10.1016/0303-7207(85)90039-5
Accession Number: 3934016
Keywords: Acetylglucosaminidase
Angiotensinogen/*genetics
Angiotensins/*genetics
Animals
Cell Line
Cell-Free System
Humans
Intracellular Membranes/*enzymology
Liver Neoplasms, Experimental/*enzymology
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
Microsomes/*enzymology
Microsomes, Liver/*enzymology
*Protein Biosynthesis
*Protein Processing, Post-Translational
RNA, Messenger/genetics
Rats
Species Specificity
Abstract: We have studied the processing of rat and human angiotensinogen precursors by microsomal membranes as a means of determining the number of asparagine-linked oligosaccharide units per angiotensinogen molecule, and thus the utilization of potential sites of N-glycosylation. Glycosylated, processed forms of angiotensinogen were isolated by chromatography on lentil lectin-Sepharose 4B. 35S-Methionine-labeled precursor and processed forms of angiotensinogen were compared with glycosylated and nonglycosylated 35S-methionine-labeled mature forms of angiotensinogen secreted by hepatoma cells, using immunoprecipitation, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. N-Glycosylation of secreted angiotensinogen was inhibited using tunicamycin. For rat angiotensinogen, only 2 of 3 potential sites of N-glycosylation were utilized; in contrast, all 4 potential sites of N-glycosylation of human angiotensinogen were utilized. For neither rat or human angiotensinogen precursor was there any evidence for a prosequence.
Notes: Campbell, D J
Bouhnik, J
Coezy, E
Menard, J
Corvol, P
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Ireland
Mol Cell Endocrinol. 1985 Nov;43(1):31-40. doi: 10.1016/0303-7207(85)90039-5.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1694
Author: Campos, D., Freitas, D., Gomes, J., Magalhães, A., Steentoft, C., Gomes, C., Vester-Christensen, M. B., Ferreira, J. A., Afonso, L. P., Santos, L. L., Pinto de Sousa, J., Mandel, U., Clausen, H., Vakhrushev, S. Y. and Reis, C. A.
Year: 2015
Title: Probing the O-glycoproteome of gastric cancer cell lines for biomarker discovery
Journal: Mol Cell Proteomics
Volume: 14
Issue: 6
Pages: 1616-29
Epub Date: 2015/03/31
Date: Jun
Short Title: Probing the O-glycoproteome of gastric cancer cell lines for biomarker discovery
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M114.046862
PMCID: PMC4458724
Accession Number: 25813380
Keywords: Aged
Aged, 80 and over
Biomarkers, Tumor/blood/*metabolism
Cell Line, Tumor
Female
Glycoproteins/blood/*metabolism
Humans
Male
Middle Aged
N-Acetylgalactosaminyltransferases/blood/*metabolism
Proteome
Stomach Neoplasms/blood/*metabolism
Abstract: Circulating O-glycoproteins shed from cancer cells represent important serum biomarkers for diagnostic and prognostic purposes. We have recently shown that selective detection of cancer-associated aberrant glycoforms of circulating O-glycoprotein biomarkers can increase specificity of cancer biomarker assays. However, the current knowledge of secreted and circulating O-glycoproteins is limited. Here, we used the COSMC KO "SimpleCell" (SC) strategy to characterize the O-glycoproteome of two gastric cancer SimpleCell lines (AGS, MKN45) as well as a gastric cell line (KATO III) which naturally expresses at least partially truncated O-glycans. Overall, we identified 499 O-glycoproteins and 1236 O-glycosites in gastric cancer SimpleCells, and a total 47 O-glycoproteins and 73 O-glycosites in the KATO III cell line. We next modified the glycoproteomic strategy to apply it to pools of sera from gastric cancer and healthy individuals to identify circulating O-glycoproteins with the STn glycoform. We identified 37 O-glycoproteins in the pool of cancer sera, and only nine of these were also found in sera from healthy individuals. Two identified candidate O-glycoprotein biomarkers (CD44 and GalNAc-T5) circulating with the STn glycoform were further validated as being expressed in gastric cancer tissue. A proximity ligation assay was used to show that CD44 was expressed with the STn glycoform in gastric cancer tissues. The study provides a discovery strategy for aberrantly glycosylated O-glycoproteins and a set of O-glycoprotein candidates with biomarker potential in gastric cancer.
Notes: 1535-9484
Campos, Diana
Freitas, Daniela
Gomes, Joana
Magalhães, Ana
Steentoft, Catharina
Gomes, Catarina
Vester-Christensen, Malene B
Ferreira, José Alexandre
Afonso, Luis P
Santos, Lúcio L
Pinto de Sousa, João
Mandel, Ulla
Clausen, Henrik
Vakhrushev, Sergey Y
Reis, Celso A
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2015 Jun;14(6):1616-29. doi: 10.1074/mcp.M114.046862. Epub 2015 Mar 26.
Author Address: From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark; §IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
§IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark;
¶Experimental Pathology and Therapeutics Group, Portuguese Institute of Oncology, Rua Dr. António Bernardino de Almeida 4200-072 Porto, Portugal; ‖QOPNA, Department of Chemistry of the University of Aveiro, Campus Universitário de Santiago 3810-193 Aveiro, Portugal;
**Department of Pathology, Portuguese Institute of Oncology, Rua Dr. António Bernardino de Almeida 4200-072 Porto, Portugal;
¶Experimental Pathology and Therapeutics Group, Portuguese Institute of Oncology, Rua Dr. António Bernardino de Almeida 4200-072 Porto, Portugal;
‡‡Faculty of Medicine of the University of Porto, Al. Prof. Hernâni Monteiro, 4200-319 Porto, Portugal;
From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark; seva@sund.ku.dk celsor@ipatimup.pt.
§IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal; ‡‡Faculty of Medicine of the University of Porto, Al. Prof. Hernâni Monteiro, 4200-319 Porto, Portugal; §§Institute of Biomedical Sciences Abel Salazar, ICBAS, Rua de Jorge Viterbo Ferreira n.228, 4050-313 Porto, Portugal seva@sund.ku.dk celsor@ipatimup.pt.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1999
Author: Cao, B., Duan, M., Xing, Y., Liu, C., Yang, F., Li, Y., Yang, T., Wei, Y., Gao, Q. and Jiang, J.
Year: 2019
Title: O-GlcNAc transferase activates stem-like cell potential in hepatocarcinoma through O-GlcNAcylation of eukaryotic initiation factor 4E
Journal: J Cell Mol Med
Volume: 23
Issue: 4
Pages: 2384-2398
Epub Date: 2019/01/25
Date: Apr
Short Title: O-GlcNAc transferase activates stem-like cell potential in hepatocarcinoma through O-GlcNAcylation of eukaryotic initiation factor 4E
Alternate Journal: Journal of cellular and molecular medicine
ISSN: 1582-1838 (Print)
1582-1838
DOI: 10.1111/jcmm.14043
PMCID: PMC6433694
Accession Number: 30677218
Keywords: Acylation/genetics
Aged
Apoptosis/genetics
Carcinoma, Hepatocellular/*genetics/pathology
Cell Proliferation/genetics
Disease-Free Survival
Eukaryotic Initiation Factor-4E/*genetics
Female
Gene Expression Regulation, Neoplastic/genetics
Glucose/metabolism
Humans
Liver Neoplasms/*genetics/pathology
Male
Middle Aged
N-Acetylglucosaminyltransferases/*genetics
Neoplastic Stem Cells/pathology
Prognosis
Protein Processing, Post-Translational/genetics
RNA, Small Interfering/genetics
Signal Transduction/genetics
*O-GlcNAc transferase
*O-GlcNAcylation
*eukaryotic initiation factor 4E
*hepatocellular carcinoma
*stem-like cell potential
this article.
Abstract: O-GlcNAcylation catalysed by O-GlcNAc transferase (OGT) is a reversible post-translational modification. O-GlcNAcylation participates in transcription, epigenetic regulation, and intracellular signalling. Dysregulation of O-GlcNAcylation in response to high glucose or OGT expression has been implicated in metabolic diseases and cancer. However, the underlying mechanisms by which OGT regulates hepatoma development remain largely unknown. Here, we employed the lentiviral shRNA-based system to knockdown OGT to analyse the contribution of OGT in hepatoma cell proliferation and stem-like cell potential. The sphere-forming assay and western blot analysis of stem-related gene expression were used to evaluate stem-like cell potential of hepatoma cell. We found that the level of total O-GlcNAcylation or OGT protein was increased in hepatocellular carcinoma. OGT activated stem-like cell potential in hepatoma through eukaryotic initiation factor 4E (eIF4E) which bound to stem-related gene Sox2 5'-untranslated region. O-GlcNAcylation of eIF4E at threonine 168 and threonine 177 protected it from degradation through proteasome pathway. Expression of eIF4E in hepatoma was determined by immunostaining in 232 HCC patients, and Kaplan-Meier survival analysis was used to determine the correlation of eIF4E expression with prognosis. High glucose promoted stem-like cell potential of hepatoma cell through OGT-eIF4E axis. Collectively, our findings indicate that OGT promotes the stem-like cell potential of hepatoma cell through O-GlcNAcylation of eIF4E. These results provide a mechanism of HCC development and a cue between the pathogenesis of HCC and high glucose condition.
Notes: 1582-4934
Cao, Benjin
Orcid: 0000-0002-5085-9847
Duan, Meng
Xing, Yang
Liu, Chanjuan
Yang, Fan
Li, Yinan
Yang, Tianxiao
Wei, Yuanyan
Gao, Qiang
Jiang, Jianhai
Journal Article
Research Support, Non-U.S. Gov't
J Cell Mol Med. 2019 Apr;23(4):2384-2398. doi: 10.1111/jcmm.14043. Epub 2019 Jan 24.
Author Address: Key Laboratory of Glycoconjuates Research, Ministry of Public Health, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan, Shanghai, People's Republic of China.
Department of Liver Surgery and Transplantation, Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion of Ministry of Education, Shanghai, People's Republic of China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1224
Author: Cao, J., Shen, C., Wang, H., Shen, H., Chen, Y., Nie, A., Yan, G., Lu, H., Liu, Y. and Yang, P.
Year: 2009
Title: Identification of N-glycosylation sites on secreted proteins of human hepatocellular carcinoma cells with a complementary proteomics approach
Journal: J Proteome Res
Volume: 8
Issue: 2
Pages: 662-72
Epub Date: 2009/02/07
Date: Feb
Short Title: Identification of N-glycosylation sites on secreted proteins of human hepatocellular carcinoma cells with a complementary proteomics approach
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr800826u
Accession Number: 19196183
Keywords: Amino Acid Sequence
Animals
*Carcinoma, Hepatocellular/chemistry/metabolism
Cattle
Cell Line, Tumor
*Glycoproteins/chemistry/metabolism
Glycosylation
Humans
*Liver Neoplasms/chemistry/metabolism
Molecular Sequence Data
*Neoplasm Proteins/chemistry/metabolism
Proteomics/*methods
Abstract: N-linked glycosylation is prevalent in proteins destined for extracellular environments; nearly all secreted proteins are glycosylated. However, with respect to their glycosylation sites, little attention has been paid. Here, we report the analysis of N-glycosylation sites on secreted proteins of human hepatocellular carcinoma cells. For the enrichment of glycopeptides, capture methods with hydrophilic affinity (HA) and hydrazide chemistry (HC) were used complementarily. With the use of both methods in combination with nano-LC-ESI-MS/MS analysis, 300 different glycosylation sites within 194 unique glycoproteins were identified, and 172 glycosites have not been determined experimentally previously. A direct comparison between HA and HC methods was also investigated for the first time. In brief, in terms of selectivity for glycopeptides, HC is superior to HA (92.9% vs 51.3%); however, based on the number of glycosites identified, HA outweighs HC (265 vs 159). Furthermore, unavoidable contaminants such as actin and bovine serum albumin which are not N-glycosylated could be easily depleted by using this glycoproteomic strategy. As a consequence, more low-abundance and genuinely secreted proteins were identified. Among the glycoproteins identified, alpha-fetoprotein, CD44 and laminin have been reported to be implicated in HCC and its metastasis.
Notes: Cao, Jing
Shen, Chengping
Wang, Hong
Shen, Huali
Chen, Yaohan
Nie, Aiying
Yan, Guoquan
Lu, Haojie
Liu, Yinkun
Yang, Pengyuan
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2009 Feb;8(2):662-72. doi: 10.1021/pr800826u.
Author Address: Department of Chemistry, Fudan University, Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1591
Author: Cao, W., Cao, J., Huang, J., Yao, J., Yan, G., Xu, H. and Yang, P.
Year: 2013
Title: Discovery and confirmation of O-GlcNAcylated proteins in rat liver mitochondria by combination of mass spectrometry and immunological methods
Journal: PLoS One
Volume: 8
Issue: 10
Pages: e76399
Epub Date: 2013/10/08
Short Title: Discovery and confirmation of O-GlcNAcylated proteins in rat liver mitochondria by combination of mass spectrometry and immunological methods
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0076399
PMCID: PMC3788734
Accession Number: 24098488
Keywords: Acetylglucosamine/chemistry/metabolism
Animals
Glycosylation
Mass Spectrometry
Mitochondria, Liver/*metabolism
Mitochondrial Proteins/*chemistry/*metabolism
Protein Interaction Mapping
Protein Interaction Maps
*Protein Processing, Post-Translational
Rats
Reproducibility of Results
Signal Transduction
Abstract: O-linked β-N-acetylglucosamine (O-GlcNAc) is an important post-translational modification (PTM) consisting of a single N-acetylglucosamine moiety attached via an O-β-glycosidic linkage to serine and threonine residues. Glycosylation with O-GlcNAc occurs on myriad nuclear and cytosolic proteins from almost all functional classes. However, with respect to O-GlcNAcylated proteins special in mitochondria, little attention has been paid. In this study, we combined mass spectrometry and immunological methods to perform global exploration of O-GlcNAcylated proteins specific in mitochondria of rat liver. First, highly purified mitochondrial proteins were obviously shown to be O-GlcNAcylated by immunoblot profiling. Then, β-elimination followed by Michael Addition with Dithiothreitol (BEMAD) treatment and LC-MS/MS were performed to enrich and identify O-GlcNAcylated mitochondrial proteins, resulting in an unambiguous assignment of 14 O-GlcNAcylation sites, mapping to 11 O-GlcNAcylated proteins. Furthermore, the identified O-GlcNAcylated mitochondrial proteins were fully validated by both electron transfer dissociation tandem mass spectrometry (ETD/MS/MS) and western blot. Thus, for the first time, our study definitely not only identified but also validated that some mitochondrial proteins in rat liver are O-GlcNAcylated. Interestingly, all of these O-GlcNAcylated mitochondrial proteins are enzymes, the majority of which are involved in a wide variety of biological processes, such as urea cycle, tricarboxylic acid cycle and lipid metabolism, indicating a role for protein O-GlcNAcylation in mitochondrial function.
Notes: 1932-6203
Cao, Weiqian
Cao, Jing
Huang, Jiangming
Yao, Jun
Yan, Guoquan
Xu, Haoqi
Yang, Pengyuan
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2013 Oct 2;8(10):e76399. doi: 10.1371/journal.pone.0076399. eCollection 2013.
Author Address: Institutes of Biomedical Sciences and Department of Chemistry, Fudan University, Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 850
Author: Capellari, S., Zaidi, S. I., Long, A. C., Kwon, E. E. and Petersen, R. B.
Year: 2000
Title: The Thr183Ala Mutation, Not the Loss of the First Glycosylation Site, Alters the Physical Properties of the Prion Protein
Journal: J Alzheimers Dis
Volume: 2
Issue: 1
Pages: 27-35
Epub Date: 2002/09/06
Date: Mar
Short Title: The Thr183Ala Mutation, Not the Loss of the First Glycosylation Site, Alters the Physical Properties of the Prion Protein
Alternate Journal: Journal of Alzheimer's disease : JAD
ISSN: 1387-2877
DOI: 10.3233/jad-2000-2104
Accession Number: 12214108
Abstract: The abnormal form of the prion protein has increased resistance to protease digestion and is insoluble in non-ionic detergents. The normal prion protein is modified by the non-obligatory addition of two N-linked glycans. One pathogenic mutation, Thr to Ala at residue 183 of the human prion protein, blocks addition of the first glycan to the Asp residue 181. This mutation has been reported to result in intracellular retention of the mutant protein and its acquisition of pathogenic properties, presumably due to the lack of the glycan. We report that the lack of the N-linked glycan at residue 181 is not responsible for the block in transport or the acquisition of pathogen-like properties, rather, the Thr to Ala mutation is itself the probable cause of the pathogenic phenotype.
Notes: 1875-8908
Capellari, Sabina
Zaidi, Syed IA
Long, Amy C
Kwon, Eunice E
Petersen, Robert B
Journal Article
Netherlands
J Alzheimers Dis. 2000 Mar;2(1):27-35. doi: 10.3233/jad-2000-2104.
Author Address: Institute of Pathology, Case Western Reserve University, Cleveland, OH 44106-2622, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1375
Author: Capotosti, F., Guernier, S., Lammers, F., Waridel, P., Cai, Y., Jin, J., Conaway, J. W., Conaway, R. C. and Herr, W.
Year: 2011
Title: O-GlcNAc transferase catalyzes site-specific proteolysis of HCF-1
Journal: Cell
Volume: 144
Issue: 3
Pages: 376-88
Epub Date: 2011/02/08
Date: Feb 4
Short Title: O-GlcNAc transferase catalyzes site-specific proteolysis of HCF-1
Alternate Journal: Cell
ISSN: 0092-8674
DOI: 10.1016/j.cell.2010.12.030
Accession Number: 21295698
Keywords: Amino Acid Sequence
Cell Cycle
Glycosylation
Host Cell Factor C1/chemistry/genetics/*metabolism
Humans
Molecular Sequence Data
Mutation
N-Acetylglucosaminyltransferases/*metabolism
*Protein Processing, Post-Translational
Protein Subunits/metabolism
Sequence Alignment
Abstract: The human epigenetic cell-cycle regulator HCF-1 undergoes an unusual proteolytic maturation process resulting in stably associated HCF-1(N) and HCF-1(C) subunits that regulate different aspects of the cell cycle. Proteolysis occurs at six centrally located HCF-1(PRO)-repeat sequences and is important for activation of HCF-1(C)-subunit functions in M phase progression. We show here that the HCF-1(PRO) repeat is recognized by O-linked β-N-acetylglucosamine transferase (OGT), which both O-GlcNAcylates the HCF-1(N) subunit and directly cleaves the HCF-1(PRO) repeat. Replacement of the HCF-1(PRO) repeats by a heterologous proteolytic cleavage signal promotes HCF-1 proteolysis but fails to activate HCF-1(C)-subunit M phase functions. These results reveal an unexpected role of OGT in HCF-1 proteolytic maturation and an unforeseen nexus between OGT-directed O-GlcNAcylation and proteolytic maturation in HCF-1 cell-cycle regulation.
Notes: 1097-4172
Capotosti, Francesca
Guernier, Sophie
Lammers, Fabienne
Waridel, Patrice
Cai, Yong
Jin, Jingji
Conaway, Joan W
Conaway, Ronald C
Herr, Winship
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell. 2011 Feb 4;144(3):376-88. doi: 10.1016/j.cell.2010.12.030.
Author Address: Center for Integrative Genomics, University of Lausanne, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1800
Author: Caputo, A. T., Alonzi, D. S., Marti, L., Reca, I. B., Kiappes, J. L., Struwe, W. B., Cross, A., Basu, S., Lowe, E. D., Darlot, B., Santino, A., Roversi, P. and Zitzmann, N.
Year: 2016
Title: Structures of mammalian ER α-glucosidase II capture the binding modes of broad-spectrum iminosugar antivirals
Journal: Proc Natl Acad Sci U S A
Volume: 113
Issue: 32
Pages: E4630-8
Epub Date: 2016/07/28
Date: Aug 9
Short Title: Structures of mammalian ER α-glucosidase II capture the binding modes of broad-spectrum iminosugar antivirals
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1604463113
PMCID: PMC4987793
Accession Number: 27462106
Keywords: Animals
Antiviral Agents/*pharmacology
Catalysis
Crystallography, X-Ray
Endoplasmic Reticulum/*enzymology
Glycoside Hydrolase Inhibitors/*pharmacology
Mice
Protein Conformation
Protein Subunits
Scattering, Small Angle
Substrate Specificity
alpha-Glucosidases/*chemistry
*ER α-glucosidase II
*broad-spectrum antiviral
*eukaryotic secretion
*glycoprotein folding
*iminosugar
which describe iminosugars and methods of treating viral diseases:
WO/2001/010429(2001) and EP1210082(2002)
WO/1999/029321(1999), EP1037636(2000) and
KR1020010033028(2001)
WO/2010/096764(2010) and EP2398321(2011)
WO/2011/028775(2011) and EP2473482(2012)
WO/2011/028779(2011) and EP2473046(2012)
WO/2011/028781(2011) and EP2473493(2012)
WO/2010/144759(2010) and EP2440205(2012)
and WO/2010/099064(2010) and EP2400843(2012).
Abstract: The biosynthesis of enveloped viruses depends heavily on the host cell endoplasmic reticulum (ER) glycoprotein quality control (QC) machinery. This dependency exceeds the dependency of host glycoproteins, offering a window for the targeting of ERQC for the development of broad-spectrum antivirals. We determined small-angle X-ray scattering (SAXS) and crystal structures of the main ERQC enzyme, ER α-glucosidase II (α-GluII; from mouse), alone and in complex with key ligands of its catalytic cycle and antiviral iminosugars, including two that are in clinical trials for the treatment of dengue fever. The SAXS data capture the enzyme's quaternary structure and suggest a conformational rearrangement is needed for the simultaneous binding of a monoglucosylated glycan to both subunits. The X-ray structures with key catalytic cycle intermediates highlight that an insertion between the +1 and +2 subsites contributes to the enzyme's activity and substrate specificity, and reveal that the presence of d-mannose at the +1 subsite renders the acid catalyst less efficient during the cleavage of the monoglucosylated substrate. The complexes with iminosugar antivirals suggest that inhibitors targeting a conserved ring of aromatic residues between the α-GluII +1 and +2 subsites would have increased potency and selectivity, thus providing a template for further rational drug design.
Notes: 1091-6490
Caputo, Alessandro T
Orcid: 0000-0001-5007-6896
Alonzi, Dominic S
Marti, Lucia
Reca, Ida-Barbara
Kiappes, J L
Struwe, Weston B
Cross, Alice
Basu, Souradeep
Lowe, Edward D
Darlot, Benoit
Santino, Angelo
Roversi, Pietro
Orcid: 0000-0001-9280-9437
Zitzmann, Nicole
WT_/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2016 Aug 9;113(32):E4630-8. doi: 10.1073/pnas.1604463113. Epub 2016 Jul 26.
Author Address: Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom;
Institute of Sciences of Food Production, Consiglio Nazionale delle Ricerche Unit of Lecce, 73100 Lecce, Italy;
Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom; Ecole Nationale Supérieure de Chimie de Montpellier, 34296 Montpellier Cedex 5, France.
Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom; pietro.roversi@bioch.ox.ac.uk nicole.zitzmann@bioch.ox.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1473
Author: Carafoli, F., Mayer, M. C., Shiraishi, K., Pecheva, M. A., Chan, L. Y., Nan, R., Leitinger, B. and Hohenester, E.
Year: 2012
Title: Structure of the discoidin domain receptor 1 extracellular region bound to an inhibitory Fab fragment reveals features important for signaling
Journal: Structure
Volume: 20
Issue: 4
Pages: 688-97
Epub Date: 2012/04/10
Date: Apr 4
Short Title: Structure of the discoidin domain receptor 1 extracellular region bound to an inhibitory Fab fragment reveals features important for signaling
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2012.02.011
PMCID: PMC3320712
Accession Number: 22483115
Keywords: Antibodies, Neutralizing/*chemistry
Binding Sites
Collagen/*chemistry/genetics
Crystallography, X-Ray
Discoidin Domain Receptor 1
Discoidin Domain Receptors
Flow Cytometry
HEK293 Cells
Humans
Immunoglobulin Fab Fragments/chemistry
Models, Molecular
Protein Binding
Protein Multimerization
Protein Structure, Secondary
Protein Structure, Tertiary
Protein Subunits/antagonists & inhibitors/chemistry/genetics
Receptor Protein-Tyrosine Kinases/antagonists & inhibitors/*chemistry/genetics
Receptors, Mitogen/*chemistry/genetics
Recombinant Fusion Proteins/chemistry/genetics
Signal Transduction/genetics
Transfection
Abstract: The discoidin domain receptors, DDR1 and DDR2, are constitutively dimeric receptor tyrosine kinases that are activated by triple-helical collagen. Aberrant DDR signaling contributes to several human pathologies, including many cancers. We have generated monoclonal antibodies (mAbs) that inhibit DDR1 signaling without interfering with collagen binding. The crystal structure of the monomeric DDR1 extracellular region bound to the Fab fragment of mAb 3E3 reveals that the collagen-binding discoidin (DS) domain is tightly associated with the following DS-like domain, which contains the epitopes of all mAbs. A conserved surface patch in the DS domain outside the collagen-binding site is shown to be required for signaling. Thus, the active conformation of the DDR1 dimer involves collagen-induced contacts between the DS domains, in addition to the previously identified association of transmembrane helices. The mAbs likely inhibit signaling by sterically blocking the extracellular association of DDR1 subunits.
Notes: 1878-4186
Carafoli, Federico
Mayer, Marie Cathrin
Shiraishi, Kazushige
Pecheva, Mira Anguelova
Chan, Lai Yi
Nan, Ruodan
Leitinger, Birgit
Hohenester, Erhard
083942/Wellcome Trust/United Kingdom
BB/D524840/1/Biotechnology and Biological Sciences Research Council/United Kingdom
G0701121/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Structure. 2012 Apr 4;20(4):688-97. doi: 10.1016/j.str.2012.02.011. Epub 2012 Apr 3.
Author Address: Department of Life Sciences, Imperial College London, London SW7 2AZ, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1239
Author: Carletti, E., Aurbek, N., Gillon, E., Loiodice, M., Nicolet, Y., Fontecilla-Camps, J. C., Masson, P., Thiermann, H., Nachon, F. and Worek, F.
Year: 2009
Title: Structure-activity analysis of aging and reactivation of human butyrylcholinesterase inhibited by analogues of tabun
Journal: Biochem J
Volume: 421
Issue: 1
Pages: 97-106
Epub Date: 2009/04/17
Date: Jun 12
Short Title: Structure-activity analysis of aging and reactivation of human butyrylcholinesterase inhibited by analogues of tabun
Alternate Journal: The Biochemical journal
ISSN: 0264-6021
DOI: 10.1042/bj20090091
Accession Number: 19368529
Keywords: Butyrylcholinesterase/*metabolism
Catalytic Domain
Cholinesterase Inhibitors/chemistry/*pharmacology
Cholinesterase Reactivators/chemistry/*pharmacology
Humans
Kinetics
Models, Molecular
Molecular Structure
Organophosphates/*pharmacology
Oximes/chemistry/*pharmacology
Protein Conformation
Structure-Activity Relationship
Time Factors
Abstract: hBChE [human BChE (butyrylcholinesterase)] naturally scavenges OPs (organophosphates). This bioscavenger is currently in Clinical Phase I for pretreatment of OP intoxication. Phosphylated ChEs (cholinesterases) can undergo a spontaneous time-dependent process called 'aging' during which the conjugate is dealkylated, leading to creation of an enzyme that cannot be reactivated. hBChE inhibited by phosphoramidates such as tabun displays a peculiar resistance to oxime-mediated reactivation. We investigated the basis of oxime resistance of phosphoramidyl-BChE conjugates by determining the kinetics of inhibition, reactivation (obidoxime {1,1'-(oxybis-methylene) bis[4-(hydroxyimino) methyl] pyridinium dichloride}, TMB-4 [1,3-trimethylene-bis(4-hydroxyiminomethylpyridinium) dibromide], HLö 7 {1-[[[4-(aminocarbonyl) pyridinio]methoxy]methyl]-2,4-bis-[(hydroxyimino)methyl] pyridinium dimethanesulfonate)}, HI-6 {1-[[[4-(aminocarbonyl) pyridinio] methoxy] methyl]-2-[(hydroxyimino)methyl]pyridinium dichloride monohydrate} and aging, and the crystal structures of hBChE inhibited by different N-monoalkyl and N,N-dialkyl tabun analogues. The refined structures of aged hBChE conjugates show that aging proceeds through O-dealkylation of the P(R) enantiomer of N,N-diethyl and N-propyl analogues, with subsequent formation of a salt bridge preventing reactivation, similarly to a previous observation made on tabun-ChE conjugates. Interestingly, the N-methyl analogue projects its amino group towards the choline-binding pocket, so that aging proceeds through deamination. This orientation results from a preference of hBChE's acyl-binding pocket for larger than 2-atoms linear substituents. The correlation between the inhibitory potency and the N-monoalkyl chain length is related to increasingly optimized interactions with the acyl-binding pocket as shown by the X-ray structures. These kinetics and X-ray data lead to a structure-activity relationship that highlights steric and electronic effects of the amino substituent of phosphoramidate. This study provides the structural basis to design new oximes capable of reactivating phosphoramidyl-hBChE conjugates after intoxication, notably when hBChE is used as pretreatment, or to design BChE-based catalytic bioscavengers.
Notes: 1470-8728
Carletti, Eugénie
Aurbek, Nadine
Gillon, Emilie
Loiodice, Mélanie
Nicolet, Yvain
Fontecilla-Camps, Juan-Carlos
Masson, Patrick
Thiermann, Horst
Nachon, Florian
Worek, Franz
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem J. 2009 Jun 12;421(1):97-106. doi: 10.1042/BJ20090091.
Author Address: Unité d'Enzymologie, Département de Toxicologie, Centre de Recherches du Service de Santé des Armées (CRSSA), 24 av des Maquis du Grésivaudan, 38700 La Tronche, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1306
Author: Carletti, E., Colletier, J. P., Dupeux, F., Trovaslet, M., Masson, P. and Nachon, F.
Year: 2010
Title: Structural evidence that human acetylcholinesterase inhibited by tabun ages through O-dealkylation
Journal: J Med Chem
Volume: 53
Issue: 10
Pages: 4002-8
Epub Date: 2010/04/23
Date: May 27
Short Title: Structural evidence that human acetylcholinesterase inhibited by tabun ages through O-dealkylation
Alternate Journal: Journal of medicinal chemistry
ISSN: 0022-2623
DOI: 10.1021/jm901853b
Accession Number: 20408548
Keywords: Acetylcholinesterase/*chemistry
Catalytic Domain
Chemical Warfare Agents/*chemistry
Cholinesterase Inhibitors/*chemistry
Crystallography, X-Ray
Dealkylation
Elapid Venoms/chemistry
Enzyme Stability
Humans
Models, Molecular
Organophosphates/*chemistry
Protein Binding
Protein Conformation
Protein Denaturation
Solutions
Abstract: Tabun is a warfare agent that inhibits human acetylcholinesterase (hAChE) by rapid phosphylation of the catalytic serine. A time-dependent reaction occurs on the tabun adduct, leading to an "aged" enzyme, resistant to oxime reactivators. The aging reaction may proceed via either dealkylation or deamidation, depending on the stereochemistry of the phosphoramidyl adduct. We solved the X-ray structure of aged tabun-hAChE complexed with fasciculin II, and we show that aging proceeds through O-dealkylation, in agreement with the aging mechanism that we determined for tabun-inhibited human butyrylcholinesterase and mouse acetylcholinesterase. Noteworthy, aging and binding of fasciculin II lead to an improved thermostability, resulting from additional stabilizing interactions between the two subdomains that face each other across the active site gorge. This first structure of hAChE inhibited by a nerve agent provides structural insight into the inhibition and aging mechanisms and a structural template for the design of molecules capable of reactivating aged hAChE.
Notes: 1520-4804
Carletti, Eugénie
Colletier, Jacques-Philippe
Dupeux, Florine
Trovaslet, Marie
Masson, Patrick
Nachon, Florian
Journal Article
Research Support, Non-U.S. Gov't
United States
J Med Chem. 2010 May 27;53(10):4002-8. doi: 10.1021/jm901853b.
Author Address: Laboratoire de Biophysique Moléculaire, Institut de Biologie Structurale (CEA-CNRS-UJF), Grenoble, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1206
Author: Carletti, E., Li, H., Li, B., Ekström, F., Nicolet, Y., Loiodice, M., Gillon, E., Froment, M. T., Lockridge, O., Schopfer, L. M., Masson, P. and Nachon, F.
Year: 2008
Title: Aging of cholinesterases phosphylated by tabun proceeds through O-dealkylation
Journal: J Am Chem Soc
Volume: 130
Issue: 47
Pages: 16011-20
Epub Date: 2008/11/04
Date: Nov 26
Short Title: Aging of cholinesterases phosphylated by tabun proceeds through O-dealkylation
Alternate Journal: Journal of the American Chemical Society
ISSN: 0002-7863
DOI: 10.1021/ja804941z
Accession Number: 18975951
Keywords: Alkylation
Catalytic Domain
Cholinesterase Inhibitors/chemistry/pharmacology
Cholinesterases/*chemistry/*metabolism
Crystallography, X-Ray
Humans
Mass Spectrometry
Models, Molecular
Molecular Structure
Organophosphates/*chemistry
Phosphorylation
Abstract: Human butyrylcholinesterase (hBChE) hydrolyzes or scavenges a wide range of toxic esters, including heroin, cocaine, carbamate pesticides, organophosphorus pesticides, and nerve agents. Organophosphates (OPs) exert their acute toxicity through inhibition of acetylcholinesterase (AChE) by phosphorylation of the catalytic serine. Phosphylated cholinesterase (ChE) can undergo a spontaneous, time-dependent process called "aging", during which the OP-ChE conjugate is dealkylated. This leads to irreversible inhibition of the enzyme. The inhibition of ChEs by tabun and the subsequent aging reaction are of particular interest, because tabun-ChE conjugates display an extraordinary resistance toward most current oxime reactivators. We investigated the structural basis of oxime resistance for phosphoramidated ChE conjugates by determining the crystal structures of the non-aged and aged forms of hBChE inhibited by tabun, and by updating the refinement of non-aged and aged tabun-inhibited mouse AChE (mAChE). Structures for non-aged and aged tabun-hBChE were refined to 2.3 and 2.1 A, respectively. The refined structures of aged ChE conjugates clearly show that the aging reaction proceeds through O-dealkylation of the P(R) enantiomer of tabun. After dealkylation, the negatively charged oxygen forms a strong salt bridge with protonated His438N epsilon2 that prevents reactivation. Mass spectrometric analysis of the aged tabun-inhibited hBChE showed that both the dimethylamine and ethoxy side chains were missing from the phosphorus. Loss of the ethoxy is consistent with the crystallography results. Loss of the dimethylamine is consistent with acid-catalyzed deamidation during the preparation of the aged adduct for mass spectrometry. The reported 3D data will help in the design of new oximes capable of reactivating tabun-ChE conjugates.
Notes: 1520-5126
Carletti, Eugénie
Li, He
Li, Bin
Ekström, Fredrik
Nicolet, Yvain
Loiodice, Mélanie
Gillon, Emilie
Froment, Marie T
Lockridge, Oksana
Schopfer, Lawrence M
Masson, Patrick
Nachon, Florian
Journal Article
United States
J Am Chem Soc. 2008 Nov 26;130(47):16011-20. doi: 10.1021/ja804941z.
Author Address: Département de Toxicologie, Centre de Recherches du Service de Santé des Armées, 38700 La Tronche, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1011
Author: Carlson, C. B., Bernstein, D. A., Annis, D. S., Misenheimer, T. M., Hannah, B. L., Mosher, D. F. and Keck, J. L.
Year: 2005
Title: Structure of the calcium-rich signature domain of human thrombospondin-2
Journal: Nat Struct Mol Biol
Volume: 12
Issue: 10
Pages: 910-4
Epub Date: 2005/09/28
Date: Oct
Short Title: Structure of the calcium-rich signature domain of human thrombospondin-2
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb997
PMCID: PMC2219892
NIHMSID: NIHMS9908
Accession Number: 16186819
Keywords: Amino Acid Sequence
Bone Diseases/genetics
Calcium/*chemistry/metabolism
Crystallization
Humans
Joint Diseases/genetics
Molecular Sequence Data
Mutation
Protein Structure, Tertiary
Thrombospondins/*chemistry/genetics/metabolism
Abstract: Thrombospondins (THBSs) are secreted glycoproteins that have key roles in interactions between cells and the extracellular matrix. Here, we describe the 2.6-A-resolution crystal structure of the glycosylated signature domain of human THBS2, which includes three epidermal growth factor-like modules, 13 aspartate-rich repeats and a lectin-like module. These elements interact extensively to form three structural regions termed the stalk, wire and globe. The THBS2 signature domain is stabilized by these interactions and by a network of 30 bound Ca(2+) ions and 18 disulfide bonds. The structure suggests how genetic alterations of THBSs result in disease.
Notes: 1545-9985
Carlson, C Britt
Bernstein, Douglas A
Annis, Douglas S
Misenheimer, Tina M
Hannah, Blue-leaf A
Mosher, Deane F
Keck, James L
R01 HL054462/HL/NHLBI NIH HHS/United States
HL07899/HL/NHLBI NIH HHS/United States
GM08293/GM/NIGMS NIH HHS/United States
T32 GM008692/GM/NIGMS NIH HHS/United States
T32 GM008293/GM/NIGMS NIH HHS/United States
HL54462/HL/NHLBI NIH HHS/United States
T32 HL007899/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Nat Struct Mol Biol. 2005 Oct;12(10):910-4. doi: 10.1038/nsmb997. Epub 2005 Sep 25.
Author Address: Department of Medicine, 4285B Medical Sciences Center, University of Wisconsin-Madison, 1300 University Avenue, Madison, Wisconsin 53706, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 327
Author: Carlsson, S. R. and Fukuda, M.
Year: 1990
Title: The polylactosaminoglycans of human lysosomal membrane glycoproteins lamp-1 and lamp-2. Localization on the peptide backbones
Journal: J Biol Chem
Volume: 265
Issue: 33
Pages: 20488-95
Epub Date: 1990/11/25
Date: Nov 25
Short Title: The polylactosaminoglycans of human lysosomal membrane glycoproteins lamp-1 and lamp-2. Localization on the peptide backbones
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2243102
Keywords: Amino Acid Sequence
*Antigens, CD
Cell Line
Chromatography, Affinity
Chromatography, Gel
Chromatography, High Pressure Liquid
Glycopeptides/isolation & purification
Humans
Leukemia, Promyelocytic, Acute
Lysosome-Associated Membrane Glycoproteins
Membrane Glycoproteins/*chemistry/isolation & purification
Molecular Sequence Data
Molecular Weight
Oligosaccharides/chemistry/*isolation & purification
Polysaccharides/*analysis
Abstract: Lysosome membrane glycoproteins, lamp-1 and lamp-2, have been shown to contain 18 and 16 N-glycans, some of which are modified by poly-N-acetyl-lactosamine. We have localized the polylactosaminoglycans to specific sites on lamp-1 and lamp-2 purified from human chronic myelogenous leukemia cells. Polylactosaminoglycan-containing glycopeptides, obtained by trypsin, pepsin, and V8 protease digestion of the glycoproteins, were isolated by Datura stramonium agglutinin affinity chromatography, gel filtration, and reverse phase high performance liquid chromatography. The poly-N-acetyllactosaminyl structures of isolated glycopeptides were confirmed by the susceptibility of their released oligosaccharides to endo-beta-galactosidase. Amino acid analysis and sequencing demonstrated that polylactosaminoglycans were located at Asn-34, Asn-93 and/or Asn-102, and Asn-195 and/or Asn-200 in lamp-1, and at Asn-4 and/or Asn-10, and Asn-279 in lamp-2. These results indicated that only certain glycosylation sites can be selectively modified by poly-N-acetyllactosamine, and those sites may confer the requirement by beta 1----3-N-acetylglucosaminyl transferase.
Notes: Carlsson, S R
Fukuda, M
CA 48737/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1990 Nov 25;265(33):20488-95.
Author Address: Department of Medical Biochemistry and Biophysics, University of Umeå, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 573
Author: Carlsson, S. R., Lycksell, P. O. and Fukuda, M.
Year: 1993
Title: Assignment of O-glycan attachment sites to the hinge-like regions of human lysosomal membrane glycoproteins lamp-1 and lamp-2
Journal: Arch Biochem Biophys
Volume: 304
Issue: 1
Pages: 65-73
Epub Date: 1993/07/01
Date: Jul
Short Title: Assignment of O-glycan attachment sites to the hinge-like regions of human lysosomal membrane glycoproteins lamp-1 and lamp-2
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1993.1322
Accession Number: 8323299
Keywords: Amino Acid Sequence
*Antigens, CD
Circular Dichroism
Glycopeptides/chemistry
Glycoproteins/*chemistry
Humans
Lysosome-Associated Membrane Glycoproteins
Magnetic Resonance Spectroscopy
Membrane Glycoproteins/*chemistry/ultrastructure
Molecular Sequence Data
Polysaccharides/chemistry
Protein Structure, Secondary
Abstract: The lysosomal membrane glycoproteins lamp-1 and lamp-2 are extensively glycosylated with a variety of different carbohydrate structures of both N-linked and O-linked type. In the present paper, we report the localization of O-linked oligosaccharides exclusively to the hinge-like regions of lamp-1 and lamp-2 isolated from human chronic myelogenous leukemia cells. In both glycoproteins, the O-glycans appear in clusters. In lamp-1, Thr-171, Thr-172, Ser-179, Ser-181, and Ser-183 were fully glycosylated, whereas Ser-169 was partially glycosylated. In lamp-2, complete glycosylation was found at Ser-167, Thr-168, Thr-172, Thr-175, Thr-176, Thr-182, and Thr-183, and partial glycosylation at Ser-179 and Thr-181, and possibly also at Thr-185. The amino acid sequences of these O-glycosylation sites are consistent with the previous reports that residues at positions -1 and +3 may influence the glycosylation reaction. Circular dichroism and nuclear magnetic resonance spectroscopy was used for the structural characterization of a synthetic peptide corresponding to residues 167 to 190 of lamp-1. The results indicated that the proline-rich O-glycan acceptor region does not adopt any typical periodic structure but differs from random-coil structure. The circular dichroism spectrum of the peptide is, however, similar to that of porcine submaxillary apomucin. A significant conformational variability was observed in this region, presumably due to a slow (on the nuclear magnetic resonance time scale) cis-trans isomerization of several proline residues. These results, taken together, strongly suggest that a hinge region does not display any typical ordered structure. The presence of O-glycans thus likely protects this region from intralumenal lysosomal proteases.
Notes: Carlsson, S R
Lycksell, P O
Fukuda, M
CA 48737/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Arch Biochem Biophys. 1993 Jul;304(1):65-73. doi: 10.1006/abbi.1993.1322.
Author Address: Department of Medical Biochemistry and Biophysics, University of Umea, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 366
Author: Carlsson, S. R., Sasaki, H. and Fukuda, M.
Year: 1986
Title: Structural variations of O-linked oligosaccharides present in leukosialin isolated from erythroid, myeloid, and T-lymphoid cell lines
Journal: J Biol Chem
Volume: 261
Issue: 27
Pages: 12787-95
Epub Date: 1986/09/25
Date: Sep 25
Short Title: Structural variations of O-linked oligosaccharides present in leukosialin isolated from erythroid, myeloid, and T-lymphoid cell lines
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2943741
Keywords: *Antigens, CD
Carbohydrate Conformation
Cell Line
Chromatography, Gel
Electrophoresis, Polyacrylamide Gel
Erythrocytes/*analysis
Glucosamine/metabolism
Humans
Leukemia, Myeloid, Acute/*metabolism
Leukosialin
Oligosaccharides/*analysis
Pronase/metabolism
Sialoglycoproteins/*analysis
T-Lymphocytes/*analysis
Abstract: Structures of O-linked oligosaccharides of leukosialin isolated from K562 erythroid, HL-60 promyelocytic, and HSB-2 T-lymphoid cell lines were examined. Leukosialin was isolated by specific immunoprecipitation from cells which were metabolically labeled with [3H]glucosamine, and glycopeptides were isolated after Pronase digestion. O-Linked oligosaccharides were released by alkaline borohydride treatment, and the structures of purified oligosaccharides were elucidated by specific exoglycosidase digestion, Smith degradation, and methylation anaylsis. Oligosaccharides from K562 cells were found to be GalNAcOH, Gal beta 1----3GalNAcOH, NeuNAc alpha 2----6GalNAcOH, NeuNAc alpha 2----3Gal beta 1----3GalNAcOH, Gal beta 1----3(NeuNAc alpha 2----6)GalNAcOH, and NeuNAc alpha 2----3Gal beta 1----3(NeuNAc alpha 2----6)GalNAcOH. On the other hand, oligosaccharides from HL-60 and HSB-2 cells were found to be NeuNAc alpha 2----3Gal beta 1----3GalNAcOH, NeuNAc alpha 2----3Gal beta 1----4GlcNAc beta 1----6(Gal beta 1----3)GalNAcOH, Gal beta 1----4GlcNAc beta 1----6(NeuNAc alpha 2----3)Gal beta 1----3)GalNAcOH, and NeuNAc alpha 2----3Gal beta 1----4GlcNAc beta 1----6(NeuNAc alpha 2----3Gal beta 1----3)GalNAcOH. These results clearly indicate that leukosialin can be differently glycosylated with O-linked chains, and each erythroid or myeloid (and T-lymphoid) cell line expresses a characteristic set of O-linked oligosaccharides which differ in core structures as well as in sialylation.
Notes: Carlsson, S R
Sasaki, H
Fukuda, M
CA 33000/CA/NCI NIH HHS/United States
R01 CA 33895/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1986 Sep 25;261(27):12787-95.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 583
Author: Carlstedt, I., Herrmann, A., Karlsson, H., Sheehan, J., Fransson, L. A. and Hansson, G. C.
Year: 1993
Title: Characterization of two different glycosylated domains from the insoluble mucin complex of rat small intestine
Journal: J Biol Chem
Volume: 268
Issue: 25
Pages: 18771-81
Epub Date: 1993/09/05
Date: Sep 5
Short Title: Characterization of two different glycosylated domains from the insoluble mucin complex of rat small intestine
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8360170
Keywords: Amino Acids/analysis
Animals
Binding Sites
Carbohydrate Sequence
Centrifugation, Isopycnic
Glycopeptides/*chemistry/isolation & purification/metabolism
Glycosylation
Guanidine
Guanidines
Intestinal Mucosa/*chemistry
Intestine, Small/chemistry
Microscopy, Electron
Molecular Sequence Data
Molecular Weight
Mucins/*chemistry/metabolism
N-Acetylneuraminic Acid
Oligosaccharides/analysis/chemistry
Rats
Rats, Sprague-Dawley
Sialic Acids/analysis
Abstract: The highly glycosylated domains of rat small intestinal mucins were isolated after reduction and trypsin digestion and separated into two populations (A and B) by gel chromatography. The molecular mass values were 650 and 335 kDa, respectively, and the relative yields suggest that the two glycopeptides occur in equimolar proportions. Electron microscopy revealed linear structures with weight average lengths of 230 nm (A) and 110 nm (B) corresponding to a mass/unit length of about 3 kDa/nm. The protein cores (17-19%) contain large amounts of threonine (over 40%), serine (17-24%), and proline (18-19%). Carbohydrate and sulfate account for approximately 80 and 0.5%, respectively, and gas chromatography-mass spectrometry showed that the patterns of neutral and sialic acid-containing glycans are very similar in the two glycopeptides. Both contain a significant amount (7-10 mol %) of single GalNAc residues, the average oligosaccharide is about 4 sugar residues long, and the largest species observed are heptasaccharides. The major neutral and sialic acid-containing oligosaccharides are Fuc1-2Gal1-3GalNAcol and GlcNAc1-6(NeuGc2-Gal1-3)GalNAcol, respectively. Sialic acid is present as both N-acetyl- and N-glycoloyl-neuraminic acid. Repeated extractions of the tissue with guanidinium chloride left approximately 80% of the mucus glycoproteins as an insoluble glycoprotein complex whereas exposure to dithiothreitol or high speed homogenization accomplished complete solubilization. The "subunits" obtained after reduction with dithiothreitol are larger than glycopeptides A and B, and fragments corresponding in size to the latter are obtained after cleavage with trypsin. Most of the mucins from rat small intestine thus occurs as an insoluble glycoprotein complex composed of subunits joined with disulfide bonds. The subunits contain two highly glycosylated regions with different lengths substituted with very similar oligosaccharides.
Notes: Carlstedt, I
Herrmann, A
Karlsson, H
Sheehan, J
Fransson, L A
Hansson, G C
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1993 Sep 5;268(25):18771-81.
Author Address: Department of Physiological Chemistry 2, University of Lund, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 298
Author: Carr, C., Aykent, S., Kimack, N. M. and Levine, A. D.
Year: 1991
Title: Disulfide assignments in recombinant mouse and human interleukin 4
Journal: Biochemistry
Volume: 30
Issue: 6
Pages: 1515-23
Epub Date: 1991/02/12
Date: Feb 12
Short Title: Disulfide assignments in recombinant mouse and human interleukin 4
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00220a011
Accession Number: 1993171
Keywords: Amino Acid Sequence
Animals
Cell Division/drug effects
Cell Line
Cloning, Molecular
Disulfides/*analysis
Dithiothreitol/pharmacology
Escherichia coli/genetics
Humans
Interleukin-4/*chemistry/genetics/isolation & purification/pharmacology
Mice
Molecular Sequence Data
Protein Conformation
Recombinant Proteins/genetics/isolation & purification/pharmacology
Sequence Homology, Nucleic Acid
Abstract: The disulfide pairings of mouse and human interleukin 4 (IL-4) proteins have been determined. The purified proteins, synthesized by recombinant DNA technology, are fully active as judged by their ability to stimulate an appropriate biological response in a variety of functional assays. Peptide maps were produced by digesting the proteins with pepsin and separating the resulting fragments by reverse-phase HPLC using linear acetonitrile-TFA gradients. Cystine-containing peptides were identified by determining which reverse-phase peaks showed an altered elution pattern after reduction. These peptides were purified further and defined by composition and sequence analysis. Three sets of disulfide-linked peptides were consistently identified for each protein. For mouse IL-4, the first and fifth, second and fourth, and third and sixth cysteines are joined. The disulfide bonds in human IL-4 are between the first and sixth, second and fourth, and third and fifth cysteines. A large double-loop region within the central three-fifths of each protein is stabilized by these bonds. Sequence analysis of the peptides containing the third and fifth cysteines of human IL-4 also demonstrated that only one of the potential N-glycosylation sites is used by C127 mammary tumor cells. Complete alkylation of mouse IL-4 under mild conditions completely destroyed its biological activity in a hematopoietic precursor cell proliferation assay.
Notes: Carr, C
Aykent, S
Kimack, N M
Levine, A D
Comparative Study
Journal Article
United States
Biochemistry. 1991 Feb 12;30(6):1515-23. doi: 10.1021/bi00220a011.
Author Address: Department of Biological Sciences, Monsanto Company, St. Louis, Missouri 63198.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1050
Author: Carr, P. D., Conlan, F., Ford, S., Ollis, D. L. and Young, I. G.
Year: 2006
Title: An improved resolution structure of the human beta common receptor involved in IL-3, IL-5 and GM-CSF signalling which gives better definition of the high-affinity binding epitope
Journal: Acta Crystallogr Sect F Struct Biol Cryst Commun
Volume: 62
Issue: Pt 6
Pages: 509-13
Epub Date: 2006/06/07
Date: Jun 1
Short Title: An improved resolution structure of the human beta common receptor involved in IL-3, IL-5 and GM-CSF signalling which gives better definition of the high-affinity binding epitope
Alternate Journal: Acta crystallographica. Section F, Structural biology and crystallization communications
ISSN: 1744-3091
DOI: 10.1107/s1744309106016812
PMCID: PMC2243076
Accession Number: 16754968
Keywords: Amino Acids
Binding Sites
Cytokine Receptor Common beta Subunit
Epitopes/chemistry
Humans
Molecular Structure
Protein Conformation
Receptors, Cell Surface/*chemistry
X-Ray Diffraction
Abstract: X-ray diffraction has been used to produce and refine a model of the extracellular domains of the beta common cytokine receptor. A minor improvement in resolution has resulted in improved electron-density maps, which have given a clearer indication of the position and stabilization of the key residues Tyr15, Phe79, Tyr347, His349, Ile350 and Tyr403 in the elbow region between domain 1 and domain 4 of the dimer-related molecule.
Notes: 1744-3091
Carr, P D
Conlan, F
Ford, S
Ollis, D L
Young, I G
Journal Article
Research Support, Non-U.S. Gov't
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2006 Jun 1;62(Pt 6):509-13. doi: 10.1107/S1744309106016812. Epub 2006 May 31.
Author Address: Research School of Chemistry, Australian National University, Canberra, Australia. pdc@rsc.anu.edu.au
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 352
Author: Carr, S. A., Hemling, M. E., Folena-Wasserman, G., Sweet, R. W., Anumula, K., Barr, J. R., Huddleston, M. J. and Taylor, P.
Year: 1989
Title: Protein and carbohydrate structural analysis of a recombinant soluble CD4 receptor by mass spectrometry
Journal: J Biol Chem
Volume: 264
Issue: 35
Pages: 21286-95
Epub Date: 1989/12/15
Date: Dec 15
Short Title: Protein and carbohydrate structural analysis of a recombinant soluble CD4 receptor by mass spectrometry
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2592374
Keywords: Amino Acid Sequence
Animals
*CD4 Antigens/genetics/isolation & purification
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Chromatography, High Pressure Liquid
Gene Expression
Genes
Humans
Mass Spectrometry
Methylation
Molecular Sequence Data
Peptide Fragments/isolation & purification
Peptide Mapping
Recombinant Proteins
Abstract: The primary structure of a soluble form of the CD4 receptor (sCD4) expressed in Chinese hamster ovary cells has been confirmed by mass spectrometric peptide mapping and and tandem mass spectrometry. These studies corroborated 95% of the 369-amino acid-long sequence and established the fidelity of translation of the NH2 and COOH terminal including the absence of "ragged ends." The arrangement of the three disulfide bonds in recombinant sCD4 was also established by mass spectrometry and comparative high performance liquid chromatography mapping and shown to be identical to that expected from previous studies of intrachain disulfide bonding in T4 antigens derived from sheep and mouse. No other arrangements of disulfides were detected. Carbohydrate mapping by mass spectrometry was used to establish that both potential Asn-linked glycosylation sites in sCD4 (Asn271 and Asn300) have oligosaccharides attached. Structural characterization by mass spectrometry and methylation analysis of the heterogeneous family of oligosaccharides at each of the specific attachment sites indicates that the major components of both families of oligosaccharides have the following biantennary structures: (Formula, see text) where m + n = 0-2, and x = 0,1. Minor carbohydrate components having three N-acetylneuraminic acid (NeuAc) groups and an additional hexose-hexosamine unit were detected by high performance anion-exchange chromatography.
Notes: Carr, S A
Hemling, M E
Folena-Wasserman, G
Sweet, R W
Anumula, K
Barr, J R
Huddleston, M J
Taylor, P
GM-39526-02/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1989 Dec 15;264(35):21286-95.
Author Address: Department of Physical and Structural Chemistry, Smith Kline and French Laboratories, King of Prussia, Pennsylvania 19406.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1658
Author: Carvalho, A. S., Ribeiro, H., Voabil, P., Penque, D., Jensen, O. N., Molina, H. and Matthiesen, R.
Year: 2014
Title: Global mass spectrometry and transcriptomics array based drug profiling provides novel insight into glucosamine induced endoplasmic reticulum stress
Journal: Mol Cell Proteomics
Volume: 13
Issue: 12
Pages: 3294-307
Epub Date: 2014/08/17
Date: Dec
Short Title: Global mass spectrometry and transcriptomics array based drug profiling provides novel insight into glucosamine induced endoplasmic reticulum stress
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M113.034363
PMCID: PMC4256484
Accession Number: 25128556
Keywords: Acetylation
Acylation
Apoptosis/drug effects
Calnexin/genetics/metabolism
Cell Cycle/drug effects
Cell Line, Tumor
Cell Nucleus/drug effects/metabolism
Cytosol/drug effects/metabolism
Endoplasmic Reticulum Stress/*drug effects/genetics
Gene Expression Profiling
Glucosamine/*pharmacology
Heat-Shock Proteins/genetics/metabolism
Humans
Lymphocytes/*drug effects/metabolism/pathology
Mass Spectrometry
Methylation
Phosphorylation
Protein Folding
*Protein Processing, Post-Translational
Protein Transport
Tissue Array Analysis
*Transcriptome
Abstract: We investigated the molecular effects of glucosamine supplements, a popular and safe alternative to nonsteroidal anti-inflammatory drugs, for decreasing pain, inflammation, and maintaining healthy joints. Numerous studies have reported an array of molecular effects after glucosamine treatment. We questioned whether the differences in the effects observed in previous studies were associated with the focus on a specific subproteome or with the use of specific cell lines or tissues. To address this question, global mass spectrometry- and transcription array-based glucosamine drug profiling was performed on malignant cell lines from different stages of lymphocyte development. We combined global label-free MS-based protein quantitation with an open search for modifications to obtain the best possible proteome coverage. Our data were largely consistent with previous studies in a variety of cellular models. We mainly observed glucosamine induced O-GlcNAcylation/O-GalNAcylation (O-HexNAcylation); however, we also observed global and local changes in acetylation, methylation, and phosphorylation. For example, our data provides two additional examples of "yin-yang" between phosphorylation and O-HexNAcylation. Furthermore, we mapped novel O-HexNAc sites on GLU2B and calnexin. GLU2B and calnexin are known to be located in the endoplasmic reticulum (ER) and involved in protein folding and quality control. The O-HexNAc sites were regulated by glucosamine treatment and correlated with the up-regulation of the ER stress marker GRP78. The occupancy of O-HexNAc on GLU2B and calnexin sites differed between the cytosolic and nuclear fractions with a higher occupancy in the cytosolic fraction. Based on our data we propose the hypothesis that O-HexNAc either inactivates calnexin and/or targets it to the cytosolic fraction. Further, we hypothesize that O-HexNAcylation induced by glucosamine treatment enhances protein trafficking.
Notes: 1535-9484
Carvalho, Ana Sofia
Ribeiro, Helena
Voabil, Paula
Penque, Deborah
Jensen, Ole N
Molina, Henrik
Matthiesen, Rune
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2014 Dec;13(12):3294-307. doi: 10.1074/mcp.M113.034363. Epub 2014 Aug 15.
Author Address: From the ‡Proteolysis in Diseases, IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal; §Human Genetics Department, National Institute of Health Dr Ricardo Jorge, Av. Padre Cruz, 1649-016 Lisboa, Portugal;
From the ‡Proteolysis in Diseases, IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
§Human Genetics Department, National Institute of Health Dr Ricardo Jorge, Av. Padre Cruz, 1649-016 Lisboa, Portugal;
¶Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense;
‖Proteomics Resource Center, The Rockefeller University, 1230 York Avenue, New York, New York 10065-6399.
From the ‡Proteolysis in Diseases, IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal; §Human Genetics Department, National Institute of Health Dr Ricardo Jorge, Av. Padre Cruz, 1649-016 Lisboa, Portugal; runem2009@gmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 643
Author: Casasnovas, J. M., Springer, T. A., Liu, J. H., Harrison, S. C. and Wang, J. H.
Year: 1997
Title: Crystal structure of ICAM-2 reveals a distinctive integrin recognition surface
Journal: Nature
Volume: 387
Issue: 6630
Pages: 312-5
Epub Date: 1997/05/15
Date: May 15
Short Title: Crystal structure of ICAM-2 reveals a distinctive integrin recognition surface
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/387312a0
Accession Number: 9153399
Keywords: Amino Acid Sequence
Animals
Antigens, CD/*chemistry/genetics/metabolism
Binding Sites
CHO Cells
Cell Adhesion Molecules/*chemistry/genetics/metabolism
Cricetinae
Crystallography, X-Ray
Intercellular Adhesion Molecule-1/metabolism
Ligands
Lymphocyte Function-Associated Antigen-1/*metabolism
Models, Molecular
Molecular Sequence Data
Protein Conformation
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Homology, Amino Acid
Abstract: Recognition by integrin proteins on the cell surface regulates the adhesive interactions between cells and their surroundings. The structure of the 'I' domain that is found in some but not all integrins, has been determined. However, the only integrin ligands for which structures are known, namely fibronectin and VCAM-1, are recognized by integrins that lack I domains. The intercellular adhesion molecules ICAM-1, 2 and 3 are, like VCAM-1, members of the immunoglobulin superfamily (IgSF), but they are recognized by an I domain-containing integrin, lymphocyte-function-associated antigen 1 (LFA-1, or CD11a/CD18). Here we present the crystal structure of the extracellular region of ICAM-2. The glutamic acid residue at position 37 is critical for LFA-1 binding and is proposed to coordinate the Mg2+ ion in the I domain; this Glu 37 is surrounded by a relatively flat recognition surface and lies in a beta-strand, whereas the critical aspartic acid residue in VCAM-1 and fibronectin lie in protruding loops. This finding suggests that there are differences in the architecture of recognition sites between integrins that contain or lack I domains. A bend between domains 1 and 2 of ICAM-2 and a tripod-like arrangement of N-linked glycans in the membrane-proximal region of domain 2 may be important for presenting the recognition surface to LFA-1. A model of ICAM-1 based on the ICAM-2 structure provides a framework for understanding its recognition by pathogens.
Notes: Casasnovas, J M
Springer, T A
Liu, J H
Harrison, S C
Wang, J H
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Nature. 1997 May 15;387(6630):312-5. doi: 10.1038/387312a0.
Author Address: The Center for Blood Research, Harvard Medical School, Department of Pathology, Boston, Massachusetts 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 669
Author: Casasnovas, J. M., Stehle, T., Liu, J. H., Wang, J. H. and Springer, T. A.
Year: 1998
Title: A dimeric crystal structure for the N-terminal two domains of intercellular adhesion molecule-1
Journal: Proc Natl Acad Sci U S A
Volume: 95
Issue: 8
Pages: 4134-9
Epub Date: 1998/05/16
Date: Apr 14
Short Title: A dimeric crystal structure for the N-terminal two domains of intercellular adhesion molecule-1
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.95.8.4134
PMCID: PMC22454
Accession Number: 9539702
Keywords: Amino Acid Sequence
Animals
Antigens, CD/chemistry
Cell Adhesion Molecules/chemistry
Computer Simulation
Crystallography, X-Ray/methods
Dimerization
Erythrocytes/parasitology
Humans
Immunoglobulins/chemistry
Intercellular Adhesion Molecule-1/biosynthesis/*chemistry
Models, Molecular
Molecular Sequence Data
Peptide Fragments/chemistry
Plasmodium falciparum/physiology
*Protein Structure, Secondary
Recombinant Proteins/biosynthesis/chemistry
Rhinovirus/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Transfection
Vascular Cell Adhesion Molecule-1/chemistry
Abstract: The 3.0-A structure of a 190-residue fragment of intercellular adhesion molecule-1 (ICAM-1, CD54) reveals two tandem Ig-superfamily (IgSF) domains. Each of two independent molecules dimerizes identically with a symmetry-related molecule over a hydrophobic interface on the BED sheet of domain 1, in agreement with dimerization of ICAM-1 on the cell surface. The residues that bind to the integrin LFA-1 are well oriented for bivalent binding in the dimer, with the critical Glu-34 residues pointing away from each other on the periphery. Residues that bind to rhinovirus are in the flexible BC and FG loops at the tip of domain 1, and these and the upper half of domain 1 are well exposed in the dimer for docking to virus. By contrast, a residue important for binding to Plasmodium falciparum-infected erythrocytes is in the dimer interface. The presence of A' strands in both domains 1 and 2, conserved hydrogen bonds at domain junctions, and elaborate hydrogen bond networks around the key integrin binding residues in domain 1 make these domains suited to resist tensile forces during adhesive interactions. A subdivision of the intermediate (I) set of IgSF domains is proposed in which domain 1 of ICAM-1 and previously described I set domains belong to the I1 set and domain 2 of ICAM-1, ICAM-2, and vascular cell adhesion molecule-1 belong to the I2 set.
Notes: 1091-6490
Casasnovas, J M
Stehle, T
Liu, J H
Wang, J H
Springer, T A
P01 HL048675/HL/NHLBI NIH HHS/United States
AI31921/AI/NIAID NIH HHS/United States
HL-48675/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1998 Apr 14;95(8):4134-9. doi: 10.1073/pnas.95.8.4134.
Author Address: The Center for Blood Research and Department of Pathology, Harvard Medical School, 200 Longwood Avenue, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1440
Author: Cash, J. N., Angerman, E. B., Kattamuri, C., Nolan, K., Zhao, H., Sidis, Y., Keutmann, H. T. and Thompson, T. B.
Year: 2012
Title: Structure of myostatin·follistatin-like 3: N-terminal domains of follistatin-type molecules exhibit alternate modes of binding
Journal: J Biol Chem
Volume: 287
Issue: 2
Pages: 1043-53
Epub Date: 2011/11/05
Date: Jan 6
Short Title: Structure of myostatin·follistatin-like 3: N-terminal domains of follistatin-type molecules exhibit alternate modes of binding
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.270801
PMCID: PMC3256893
Accession Number: 22052913
Keywords: Animals
CHO Cells
Cricetinae
Cricetulus
Crystallography, X-Ray
Follistatin-Related Proteins/*chemistry/genetics/metabolism
Humans
*Models, Molecular
Myostatin/*chemistry/genetics/metabolism
Point Mutation
Protein Binding
Protein Structure, Quaternary
Protein Structure, Tertiary
Abstract: TGF-β family ligands are involved in a variety of critical physiological processes. For instance, the TGF-β ligand myostatin is a staunch negative regulator of muscle growth and a therapeutic target for muscle-wasting disorders. Therefore, it is important to understand the molecular mechanisms of TGF-β family regulation. One form of regulation is through inhibition by extracellular antagonists such as the follistatin (Fst)-type proteins. Myostatin is tightly controlled by Fst-like 3 (Fstl3), which is the only Fst-type molecule that has been identified in the serum bound to myostatin. Here, we present the crystal structure of myostatin in complex with Fstl3. The structure reveals that the N-terminal domain (ND) of Fstl3 interacts uniquely with myostatin as compared with activin A, because it utilizes different surfaces on the ligand. This results in conformational differences in the ND of Fstl3 that alter its position in the type I receptor-binding site of the ligand. We also show that single point mutations in the ND of Fstl3 are detrimental to ligand binding, whereas corresponding mutations in Fst have little effect. Overall, we have shown that the NDs of Fst-type molecules exhibit distinctive modes of ligand binding, which may affect overall affinity of ligand·Fst-type protein complexes.
Notes: 1083-351x
Cash, Jennifer N
Angerman, Elizabeth B
Kattamuri, Chandramohan
Nolan, Kristof
Zhao, Huaying
Sidis, Yisrael
Keutmann, Henry T
Thompson, Thomas B
R01 GM084186/GM/NIGMS NIH HHS/United States
GM084186/GM/NIGMS NIH HHS/United States
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Jan 6;287(2):1043-53. doi: 10.1074/jbc.M111.270801. Epub 2011 Nov 3.
Author Address: Department of Molecular Genetics, Biochemistry, and Microbiology, University of Cincinnati, Cincinnati, Ohio 45267, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1134
Author: Castonguay, R., Halim, D., Morin, M., Furtos, A., Lherbet, C., Bonneil, E., Thibault, P. and Keillor, J. W.
Year: 2007
Title: Kinetic characterization and identification of the acylation and glycosylation sites of recombinant human gamma-glutamyltranspeptidase
Journal: Biochemistry
Volume: 46
Issue: 43
Pages: 12253-62
Epub Date: 2007/10/11
Date: Oct 30
Short Title: Kinetic characterization and identification of the acylation and glycosylation sites of recombinant human gamma-glutamyltranspeptidase
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi700956c
Accession Number: 17924658
Keywords: Acylation
Amino Acid Sequence
Base Sequence
Chromatography, Liquid
DNA Primers
Glycosylation
Humans
Kinetics
Molecular Sequence Data
Pichia/genetics
Polymerase Chain Reaction
Recombinant Proteins/chemistry/metabolism
Tandem Mass Spectrometry
gamma-Glutamyltransferase/chemistry/isolation & purification/*metabolism
Abstract: Gamma-glutamyltranspeptidase (GGT) is a heterodimeric enzyme important for glutathione homeostasis control. It has also been implicated in many physiological disorders, including Parkinson's disease, apoptosis inhibition, and diabetes. In the first step of its ping-pong mechanism it binds glutathione, its in vivo substrate, and releases cysteinylglycine upon formation of an acyl-enzyme intermediate. This intermediate can then react with water to release glutamate as a hydrolysis product or with an amino acid or dipeptide to form a transpeptidation product. Further detailed study of the mechanism underlying these reactions is hindered at least for some GGTs by the low quantities of protein available after a multistep purification from tissue. In the present work the gene for human GGT was cloned into the pPICZalphaA vector and transformed into Pichia pastoris to express as a 68 kDa His-tagged protein. The optimized expression and secretion of this enzyme in 1 L of culture and subsequent purification by immobilized metal affinity chromatography yielded 1.6 mg of purified enzyme having a specific activity of 237 U/mg. Kinetic parameters for the transpeptidation reaction between glutathione and glycylglycine were determined by mass spectrometry, giving a kcat of 13.4 x 10(3) min-1 and apparent KM values of 1.11 mM for glutathione and 8.1 mM for glycylglycine. The GGT-mediated hydrolysis of glutathione was also studied, providing a kcat of 53 min-1 and a KM value of 7.3 microM for glutathione. Incubation of the enzyme with a mechanism-based inhibitor, enzymatic digest, and mass spectrometric analysis provided the first unambiguous identification of Thr381 as the active site nucleophile of human gamma-glutamyltranspeptidase, and confirmed four of the seven N-linked glycosylation sites. These structural and kinetic data are discussed with respect to a homology model generated to facilitate visualization.
Notes: Castonguay, Roselyne
Halim, Dany
Morin, Mylène
Furtos, Alexandra
Lherbet, Christian
Bonneil, Eric
Thibault, Pierre
Keillor, Jeffrey W
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2007 Oct 30;46(43):12253-62. doi: 10.1021/bi700956c. Epub 2007 Oct 9.
Author Address: Département de chimie, Université de Montréal, Montréal, Québec, Canada H3C 3J7.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 310
Author: Caughey, G. H., Zerweck, E. H. and Vanderslice, P.
Year: 1991
Title: Structure, chromosomal assignment, and deduced amino acid sequence of a human gene for mast cell chymase
Journal: J Biol Chem
Volume: 266
Issue: 20
Pages: 12956-63
Epub Date: 1991/07/15
Date: Jul 15
Short Title: Structure, chromosomal assignment, and deduced amino acid sequence of a human gene for mast cell chymase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2071582
Keywords: Amino Acid Sequence
Animals
Base Sequence
Chromosome Mapping
*Chromosomes, Human, Pair 14
Chymases
Cloning, Molecular
Dogs
Female
*Genes
Genomic Library
Humans
Mast Cells/*enzymology
Molecular Sequence Data
Placenta/enzymology
Pregnancy
Restriction Mapping
Sequence Homology, Nucleic Acid
Serine Endopeptidases/*genetics
Abstract: A gene encoding human chymase was cloned and sequenced. The protein-coding exons reveal a preproenzyme with a 19-amino acid signal peptide, an acidic 2-amino acid propeptide, and a 226-amino acid catalytic domain. The mature enzyme is predicted to be cationic (net charge of +13) and to be modified by N-glycosylation at two sites. The amino acid sequence is identical to the 35 residues of NH2-terminal amino acid sequence reported for human skin chymase and is identical to 29 of 31 residues of NH2-terminal and internal amino acid sequence reported for human heart chymase. The full predicted sequence of the catalytic domain reveals a high level of sequence identity to dog mast cell chymase (83%) and a lower level of identity to the sequences of rodent chymases (58-62%). In the phase and placement of introns, the organization of this human chymase gene is similar to that of several other granule-associated leukocyte serine proteases, including rat chymase II, lymphocyte granzymes, and neutrophil cathespin G and elastase. However, the gene organization differs from that of mast cell tryptase, providing additional evidence that the major mast cell serine proteases are separated by substantial evolutionary distance. Amplification of chymase gene-specific fragments from hamster/human hybrid cell line DNA suggests localization of the chymase gene to human chromosome 14. High stringency hybridization of chymase DNA to a human genomic DNA blot suggests the possibility of more than one human chymase gene. Evidence that the chymase gene is expressed in human tissues was obtained by the amplification of chymase-specific DNA from skin and placental cDNA libraries.
Notes: Caughey, G H
Zerweck, E H
Vanderslice, P
HL-01736/HL/NHLBI NIH HHS/United States
HL-24136/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1991 Jul 15;266(20):12956-63.
Author Address: Cardiovascular Research Institute, University of California, San Francisco 94143-0911.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1729
Author: Cha, M. Y., Cho, H. J., Kim, C., Jung, Y. O., Kang, M. J., Murray, M. E., Hong, H. S., Choi, Y. J., Choi, H., Kim, D. K., Choi, H., Kim, J., Dickson, D. W., Song, H. K., Cho, J. W., Yi, E. C., Kim, J., Jin, S. M. and Mook-Jung, I.
Year: 2015
Title: Mitochondrial ATP synthase activity is impaired by suppressed O-GlcNAcylation in Alzheimer's disease
Journal: Hum Mol Genet
Volume: 24
Issue: 22
Pages: 6492-504
Epub Date: 2015/09/12
Date: Nov 15
Short Title: Mitochondrial ATP synthase activity is impaired by suppressed O-GlcNAcylation in Alzheimer's disease
Alternate Journal: Human molecular genetics
ISSN: 0964-6906 (Print)
0964-6906
DOI: 10.1093/hmg/ddv358
PMCID: PMC5007609
Accession Number: 26358770
Keywords: Acetylglucosamine/metabolism
Adenosine Triphosphate/metabolism
Alzheimer Disease/enzymology/genetics/*metabolism
Animals
CHO Cells
Cricetulus
Disease Models, Animal
Glycosylation
HeLa Cells
Humans
Mice
Mice, Transgenic
Mitochondria/enzymology/metabolism
Mitochondrial Proton-Translocating ATPases/genetics/*metabolism
N-Acetylglucosaminyltransferases/*metabolism
Oxidative Phosphorylation Coupling Factors/genetics/*metabolism
Protein Processing, Post-Translational
beta-N-Acetylhexosaminidases/metabolism
Abstract: Glycosylation with O-linked β-N-acetylglucosamine (O-GlcNAc) is one of the protein glycosylations affecting various intracellular events. However, the role of O-GlcNAcylation in neurodegenerative diseases such as Alzheimer's disease (AD) is poorly understood. Mitochondrial adenosine 5'-triphosphate (ATP) synthase is a multiprotein complex that synthesizes ATP from ADP and Pi. Here, we found that ATP synthase subunit α (ATP5A) was O-GlcNAcylated at Thr432 and ATP5A O-GlcNAcylation was decreased in the brains of AD patients and transgenic mouse model, as well as Aβ-treated cells. Indeed, Aβ bound to ATP synthase directly and reduced the O-GlcNAcylation of ATP5A by inhibition of direct interaction between ATP5A and mitochondrial O-GlcNAc transferase, resulting in decreased ATP production and ATPase activity. Furthermore, treatment of O-GlcNAcase inhibitor rescued the Aβ-induced impairment in ATP production and ATPase activity. These results indicate that Aβ-mediated reduction of ATP synthase activity in AD pathology results from direct binding between Aβ and ATP synthase and inhibition of O-GlcNAcylation of Thr432 residue on ATP5A.
Notes: 1460-2083
Cha, Moon-Yong
Cho, Hyun Jin
Kim, Chaeyoung
Jung, Yang Ouk
Kang, Min Jueng
Murray, Melissa E
Hong, Hyun Seok
Choi, Young-Joo
Choi, Heesun
Kim, Dong Kyu
Choi, Hyunjung
Kim, Jisoo
Dickson, Dennis W
Song, Hyun Kyu
Cho, Jin Won
Yi, Eugene C
Kim, Jungsu
Jin, Seok Min
Mook-Jung, Inhee
P01 AG003949/AG/NIA NIH HHS/United States
AG016574/AG/NIA NIH HHS/United States
P01-AG03949/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Hum Mol Genet. 2015 Nov 15;24(22):6492-504. doi: 10.1093/hmg/ddv358. Epub 2015 Sep 10.
Author Address: Department of Biochemistry and Biomedical Sciences, Seoul National University, College of Medicine, Seoul, Korea.
Department of Neuroscience, Mayo Clinic College of Medicine, Jacksonville, FL 32224, USA.
Department of Life Sciences, Korea University, Seoul, Korea.
Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science and Technology, School of Medicine and School of Pharmacy, Seoul National University, Seoul, Korea.
Medifron-DBT, Inc., Gyeonggi-do, Korea.
Department of Integrated OMICS for Biomedical Science, Graduate School, Yonsei University, Seodaemun-gu, Seoul, Korea and.
Department of Biochemistry and Biomedical Sciences, Seoul National University, College of Medicine, Seoul, Korea, inhee@snu.ac.kr neurocom@snu.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 743
Author: Chabot, D. J., Chen, H., Dimitrov, D. S. and Broder, C. C.
Year: 2000
Title: N-linked glycosylation of CXCR4 masks coreceptor function for CCR5-dependent human immunodeficiency virus type 1 isolates
Journal: J Virol
Volume: 74
Issue: 9
Pages: 4404-13
Epub Date: 2001/02/07
Date: May
Short Title: N-linked glycosylation of CXCR4 masks coreceptor function for CCR5-dependent human immunodeficiency virus type 1 isolates
Alternate Journal: Journal of virology
ISSN: 0022-538X (Print)
0022-538x
DOI: 10.1128/jvi.74.9.4404-4413.2000
PMCID: PMC111957
Accession Number: 10756055
Keywords: Amino Acid Sequence
Animals
Cell Line
Chlorocebus aethiops
Glycosylation
HIV-1/isolation & purification/*metabolism
HeLa Cells
Humans
Molecular Sequence Data
Receptors, CCR5/*metabolism
Receptors, CXCR4/genetics/*metabolism
Tumor Cells, Cultured
Abstract: The chemokine receptors CXCR4 and CCR5 are the principal coreceptors for infection of X4 and R5 human immunodeficiency virus type 1 (HIV-1) isolates, respectively. Here we report on the unexpected observation that the removal of the N-linked glycosylation sites in CXCR4 potentially allows the protein to serve as a universal coreceptor for both X4 and R5 laboratory-adapted and primary HIV-1 strains. We hypothesize that this alteration unmasks existing common extracellular structures reflecting a conserved three-dimensional similarity of important elements of CXCR4 and CCR5 that are involved in HIV envelope glycoprotein (Env) interaction. These results may have far-reaching implications for the differential recognition of cell type-dependent glycosylated CXCR4 by HIV-1 isolates and their evolution in vivo. They also suggest a possible explanation for the various observations of restricted virus entry in some cell types and further our understanding of the framework of elements that represent the Env-coreceptor contact sites.
Notes: 1098-5514
Chabot, D J
Chen, H
Dimitrov, D S
Broder, C C
R01 AI043885/AI/NIAID NIH HHS/United States
R01AI43885/AI/NIAID NIH HHS/United States
R29AI414110/AI/NIAID NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
J Virol. 2000 May;74(9):4404-13. doi: 10.1128/jvi.74.9.4404-4413.2000.
Author Address: Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, Bethesda, Maryland 20814-4799, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1931
Author: Chai, Y., Du, Y., Zhang, S., Xiao, J., Luo, Z., He, F. and Huang, K.
Year: 2018
Title: MicroRNA-485-5p reduces O-GlcNAcylation of Bmi-1 and inhibits colorectal cancer proliferation
Journal: Exp Cell Res
Volume: 368
Issue: 1
Pages: 111-118
Epub Date: 2018/04/24
Date: Jul 1
Short Title: MicroRNA-485-5p reduces O-GlcNAcylation of Bmi-1 and inhibits colorectal cancer proliferation
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2018.04.020
Accession Number: 29680296
Keywords: Cell Line, Tumor
Cell Movement/genetics
Cell Proliferation/*genetics
Colorectal Neoplasms/*genetics
Down-Regulation
Gene Expression Regulation, Neoplastic/*genetics
Humans
MicroRNAs/*genetics
Polycomb Repressive Complex 1/*genetics
*Bmi-1
*Colorectal cancer
*Ogt
*Proliferation
*miR-485-5p
Abstract: Emerging evidences showed that miRNAs are involved in the oncogenesis of many cancers. Here, miRNA microarray analysis was performed to screen the significant miRNAs involved in the progression of colorectal cancer (CRC), miR-485-5p was chosen for further study. We found that the expression of miR-485-5p was significantly lower in CRC specimens and cell lines. In addition, low expression level of miR-485-5p is correlated with tumor progression and poor survival in CRC patients. Based on in vitro and in vivo assays, we found that miR-485-5p significantly inhibits CRC proliferation. Moreover, our results showed that miR-485-5p inhibits cell proliferation by reducing Bmi-1 protein expression, which has been reported to control the proliferation of many cancers. Mechanistically, OGT is a direct target of miR-485-5p, and miR-485-5p could inhibit the O-GlcNAcylation level of Bmi-1 by OGT. Overall, these results suggested that as a tumor suppressor, miR-485-5p may regulate CRC cells proliferation, which could regulate the O-GlcNAcylation and the stability of Bmi-1 through targeting OGT. This may give insight into a novel mechanism and therapy of CRC growth.
Notes: 1090-2422
Chai, Yong
Du, Yunyan
Zhang, Shouhua
Xiao, Juhua
Luo, Zhipeng
He, Fei
Huang, Kai
Journal Article
Research Support, Non-U.S. Gov't
United States
Exp Cell Res. 2018 Jul 1;368(1):111-118. doi: 10.1016/j.yexcr.2018.04.020. Epub 2018 Apr 20.
Author Address: Department of Ophthalmology, Jiangxi Children's Hospital, Nanchang, Jiangxi Province 330006, China.
Department of Otorhinolaryngology, Jiangxi Provincial People's Hospital, Nanchang, Jiangxi Province 330006, China.
Department of Ultrasound, Jiangxi Provincial Maternal and Child Health Hospital, Nanchang, Jiangxi Province 330006, China.
Department of Gastrointestinal Surgery, Jiangxi Provincial Cancer Hospital, Nanchang 330029, China.
Department of Gastrointestinal Surgery, Jiangxi Provincial Cancer Hospital, Nanchang 330029, China. Electronic address: huangkai186@sina.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1445
Author: Chaikuad, A., Froese, D. S., Berridge, G., von Delft, F., Oppermann, U. and Yue, W. W.
Year: 2011
Title: Conformational plasticity of glycogenin and its maltosaccharide substrate during glycogen biogenesis
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 52
Pages: 21028-33
Epub Date: 2011/12/14
Date: Dec 27
Short Title: Conformational plasticity of glycogenin and its maltosaccharide substrate during glycogen biogenesis
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1113921108
PMCID: PMC3248551
Accession Number: 22160680
Keywords: Crystallography
Glucosyltransferases/*chemistry/genetics
Glycogen/*biosynthesis
Glycoproteins/*chemistry/genetics
Humans
*Models, Molecular
Mutation, Missense/genetics
*Protein Conformation
X-Ray Diffraction
Abstract: Glycogenin initiates the synthesis of a maltosaccharide chain covalently attached to itself on Tyr195 via a stepwise glucosylation reaction, priming glycogen synthesis. We have captured crystallographic snapshots of human glycogenin during its reaction cycle, revealing a dynamic conformational switch between ground and active states mediated by the sugar donor UDP-glucose. This switch includes the ordering of a polypeptide stretch containing Tyr195, and major movement of an approximately 30-residue "lid" segment covering the active site. The rearranged lid guides the nascent maltosaccharide chain into the active site in either an intra- or intersubunit mode dependent upon chain length and steric factors and positions the donor and acceptor sugar groups for catalysis. The Thr83Met mutation, which causes glycogen storage disease XV, is conformationally locked in the ground state and catalytically inactive. Our data highlight the conformational plasticity of glycogenin and coexistence of two modes of glucosylation as integral to its catalytic mechanism.
Notes: 1091-6490
Chaikuad, Apirat
Froese, D Sean
Berridge, Georgina
von Delft, Frank
Oppermann, Udo
Yue, Wyatt W
092809/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Dec 27;108(52):21028-33. doi: 10.1073/pnas.1113921108. Epub 2011 Dec 12.
Author Address: Structural Genomics Consortium, Old Road Research Campus Building, Oxford, United Kingdom OX3 7DQ.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1720
Author: Chaiyawat, P., Chokchaichamnankit, D., Lirdprapamongkol, K., Srisomsap, C., Svasti, J. and Champattanachai, V.
Year: 2015
Title: Alteration of O-GlcNAcylation affects serine phosphorylation and regulates gene expression and activity of pyruvate kinase M2 in colorectal cancer cells
Journal: Oncol Rep
Volume: 34
Issue: 4
Pages: 1933-42
Epub Date: 2015/08/08
Date: Oct
Short Title: Alteration of O-GlcNAcylation affects serine phosphorylation and regulates gene expression and activity of pyruvate kinase M2 in colorectal cancer cells
Alternate Journal: Oncology reports
ISSN: 1021-335x
DOI: 10.3892/or.2015.4178
Accession Number: 26252736
Keywords: Acylation/genetics
Colorectal Neoplasms/*genetics/metabolism/pathology
Gene Expression Regulation, Neoplastic
HT29 Cells
Humans
N-Acetylglucosaminyltransferases/biosynthesis/*genetics
Neoplasm Proteins/biosynthesis
Phosphorylation
Protein Processing, Post-Translational/*genetics
Pyruvate Kinase/*biosynthesis/genetics
RNA Interference
Serine/metabolism
Abstract: O-GlcNAcylation is a dynamic post-translational modification that has extensive crosstalk with phosphorylation either at the same or adjacent sites of various proteins. We have previously reported that O-GlcNAcylation level was increased in primary breast and colorectal cancer, but the interplay of the two modifications remains unclear. Therefore, we explored crosstalk of the modifications by RNA interference against O-GlcNAc transferase (OGT) in colorectal cancer cells. Two-dimensional immunoblotting and mass spectrometric analysis showed that the levels of O-GlcNAc and serine phosphorylation of many proteins including serine hydroxymethyltransferase, cytokeratin-8, pyruvate kinase M2 (PKM2), heterogeneous nuclear ribonucleoprotein L, and lamin-B1, were reduced in siOGT cells compared to siScramble cells. In HT29 cells, immunoprecipitated PKM2 revealed decreased O-GlcNAc and serine phosphorylation levels after siOGT knockdown, but increased levels after treatment with Thiamet-G, an inhibitor of O-GlcNAcase (OGA). In addition, when global O-GlcNAcylation was enhanced by treating cells with Thiamet-G, PKM2 expression level was upregulated, but PKM2-specific activity was decreased. On the other hand, in OGT knockdown cells, PKM2 expression level was downregulated, but PKM2-specific activity was increased. Moreover, the metastatic colorectal cancer cells, SW620, had more O-GlcNAc-PKM2 and showed lower PKM2-specific activity compared to the non-metastatic colorectal cancer SW480 cells. These results suggested roles of O-GlcNAcylation in modulating serine phosphorylation, as well as in regulating PKM2 activity and expression. Interfering levels of O-GlcNAcylation of PKM2 may be a novel target in controlling cancer metabolism and tumorigenesis of colorectal cancer.
Notes: 1791-2431
Chaiyawat, Parunya
Chokchaichamnankit, Daranee
Lirdprapamongkol, Kriengsak
Srisomsap, Chantragan
Svasti, Jisnuson
Champattanachai, Voraratt
Journal Article
Research Support, Non-U.S. Gov't
Greece
Oncol Rep. 2015 Oct;34(4):1933-42. doi: 10.3892/or.2015.4178. Epub 2015 Aug 5.
Author Address: Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok, Thailand.
Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok, Thailand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1807
Author: Chaiyawat, P., Weeraphan, C., Netsirisawan, P., Chokchaichamnankit, D., Srisomsap, C., Svasti, J. and Champattanachai, V.
Year: 2016
Title: Elevated O-GlcNAcylation of Extracellular Vesicle Proteins Derived from Metastatic Colorectal Cancer Cells
Journal: Cancer Genomics Proteomics
Volume: 13
Issue: 5
Pages: 387-98
Epub Date: 2016/08/28
Date: 09-10
Short Title: Elevated O-GlcNAcylation of Extracellular Vesicle Proteins Derived from Metastatic Colorectal Cancer Cells
Alternate Journal: Cancer genomics & proteomics
ISSN: 1109-6535 (Print)
1109-6535
PMCID: PMC5070628
Accession Number: 27566657
Keywords: Cell Line, Tumor
Chromatography, Liquid
Colorectal Neoplasms/*metabolism/*pathology
Extracellular Vesicles/*metabolism
Glycoproteins/*metabolism
Glycosylation
Humans
Neoplasm Metastasis
Proteomics/methods
Reproducibility of Results
Tandem Mass Spectrometry
Colorectal cancer
O-GlcNAcylation
RuVB-like1
extracellular vesicles
metastasis
transitional endoplasmic reticulum ATPase
Abstract: BACKGROUND: O-GlcNAcylation is a single sugar attachment of serine and/or threonine residues on intracellular proteins. Recent reports reveal that it can modify several secretory proteins; however, the underlying mechanisms are largely unexplored. MATERIALS AND METHODS: To investigate whether extracellular vesicles (EVs) carry secretory O-GlcNAc-modified proteins that were isolated from colorectal cancer (CRC) cells, two-dimensional gel electrophoresis followed with O-GlcNAc immunoblotting and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were applied. RESULTS: It was revealed that the O-GlcNAc modification of many EV proteins was increased in metastatic cells. Among these, transitional endoplasmic reticulum ATPase (TER ATPase) and RuVB-like1 were successfully confirmed for the O-GlcNAc modification in which the levels were significantly higher in EVs of metastatic CRC cell line. CONCLUSION: These data, demonstrate that proteins carried by EVs are O-GlcNAc-modified. Importantly, elevated aberrant O-GlcNAcylation of EV proteins might serve as a potential biomarker of metastatic CRC.
Notes: 1790-6245
Chaiyawat, Parunya
Weeraphan, Churat
Netsirisawan, Pukkavadee
Chokchaichamnankit, Daranee
Srisomsap, Chantragan
Svasti, Jisnuson
Champattanachai, Voraratt
Journal Article
Cancer Genomics Proteomics. 2016 09-10;13(5):387-98.
Author Address: Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok, Thailand.
Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok, Thailand.
Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok, Thailand Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok, Thailand.
Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok, Thailand Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok, Thailand voraratt@cri.or.th.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 877
Author: Chalkley, R. J. and Burlingame, A. L.
Year: 2003
Title: Identification of novel sites of O-N-acetylglucosamine modification of serum response factor using quadrupole time-of-flight mass spectrometry
Journal: Mol Cell Proteomics
Volume: 2
Issue: 3
Pages: 182-90
Epub Date: 2003/04/10
Date: Mar
Short Title: Identification of novel sites of O-N-acetylglucosamine modification of serum response factor using quadrupole time-of-flight mass spectrometry
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M300027-MCP200
Accession Number: 12684542
Keywords: Acetylglucosamine/*chemistry
Amino Acid Sequence
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Humans
Molecular Sequence Data
Phosphorylation
Serum Response Factor/*chemistry
Spectrometry, Mass, Electrospray Ionization
Abstract: The addition of a single N-acetylglucosamine moiety O-linked to serine and threonine residues of nuclear and cytoplasmic proteins is a widespread post-translational modification. The conventional method for detecting and locating sites of modification is through a multistep radioactivity-based approach. We have recently shown that sites of O-GlcNAc modification can be determined using quadrupole time-of-flight tandem mass spectrometry (Chalkley, R. J., and Burlingame, A. L. (2001) Identification of GlcNAcylation sites of peptides and alpha-crystallin using Q-TOF mass spectrometry. J. Am. Soc. Mass Spectrom. 12, 1106-1113). In this work utilization of this new approach has revealed previously undetected sites of O-GlcNAc modification of the transcription factor serum response factor.
Notes: Chalkley, Robert J
Burlingame, A L
RR01614/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Mol Cell Proteomics. 2003 Mar;2(3):182-90. doi: 10.1074/mcp.M300027-MCP200. Epub 2003 Apr 8.
Author Address: Department of Biochemistry and Molecular Biology, University College, London, Gower Street, London WC1E 6BT, United Kingdom. robertc@itsa.ucsf.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1551
Author: Champattanachai, V., Netsirisawan, P., Chaiyawat, P., Phueaouan, T., Charoenwattanasatien, R., Chokchaichamnankit, D., Punyarit, P., Srisomsap, C. and Svasti, J.
Year: 2013
Title: Proteomic analysis and abrogated expression of O-GlcNAcylated proteins associated with primary breast cancer
Journal: Proteomics
Volume: 13
Issue: 14
Pages: 2088-99
Epub Date: 2013/04/12
Date: Jul
Short Title: Proteomic analysis and abrogated expression of O-GlcNAcylated proteins associated with primary breast cancer
Alternate Journal: Proteomics
ISSN: 1615-9853
DOI: 10.1002/pmic.201200126
Accession Number: 23576270
Keywords: Acetylglucosamine/chemistry
Biomarkers, Tumor
Breast Neoplasms/*chemistry/metabolism
Electrophoresis, Gel, Two-Dimensional
Female
Glycoproteins/*analysis/chemistry/metabolism
Heterogeneous-Nuclear Ribonucleoproteins
Humans
Immunoblotting
Lactic Acid
Models, Biological
N-Acetylglucosaminyltransferases
Proteome/*analysis/chemistry/metabolism
Reproducibility of Results
Breast cancer
Glycoproteomics
Hexosamine biosynthesis pathway
O-GlcNAc transferase
O-GlcNAcyla-tion
Warburg effect
Abstract: O-GlcNAcylation is a dynamic PTM of nuclear and cytoplasmic proteins, regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase, which catalyze the addition and removal of O-GlcNAc, respectively. This modification is associated with glucose metabolism, which plays important roles in many diseases including cancer. Although emerging evidence reveals that some tumor-associated proteins are O-GlcNAc modified, the total O-GlcNAcylation in cancer is still largely unexplored. Here, we demonstrate that O-GlcNAcylation was increased in primary breast malignant tumors, not in benign tumors and that this augmentation was associated with increased expression of OGT level. Using 2D O-GlcNAc immnoblotting and LC-MS/MS analysis, we successfully identified 29 proteins, with seven being uniquely O-GlcNAcylated or associated with O-GlcNAcylation in cancer. Of these identified proteins, some were related to the Warburg effect, including metabolic enzymes, proteins involved in stress responses and biosynthesis. In addition, proteins associated with RNA metabolism, gene expression, and cytoskeleton were highly O-GlcNAcylated or associated with O-GlcNAcylation. Moreover, OGT knockdown showed that decreasing O-GlcNAcylation was related to inhibition of the anchorage-independent growth in vitro. These data indicate that aberrant protein O-GlcNAcylation is associated with breast cancer. Abnormal modification of these O-GlcNAc-modified proteins might be one of the vital malignant characteristics of cancer.
Notes: 1615-9861
Champattanachai, Voraratt
Netsirisawan, Pukkavadee
Chaiyawat, Parunya
Phueaouan, Thanong
Charoenwattanasatien, Ratana
Chokchaichamnankit, Daranee
Punyarit, Phaibul
Srisomsap, Chantragan
Svasti, Jisnuson
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2013 Jul;13(14):2088-99. doi: 10.1002/pmic.201200126. Epub 2013 Jun 6.
Author Address: Laboratory of Biochemistry, Chulabhorn Research Institute, Bangkok, Thailand. voraratt@cri.or.th
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 283
Author: Chan, A. L., Morris, H. R., Panico, M., Etienne, A. T., Rogers, M. E., Gaffney, P., Creighton-Kempsford, L. and Dell, A.
Year: 1991
Title: A novel sialylated N-acetylgalactosamine-containing oligosaccharide is the major complex-type structure present in Bowes melanoma tissue plasminogen activator
Journal: Glycobiology
Volume: 1
Issue: 2
Pages: 173-85
Epub Date: 1991/03/01
Date: Mar
Short Title: A novel sialylated N-acetylgalactosamine-containing oligosaccharide is the major complex-type structure present in Bowes melanoma tissue plasminogen activator
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/1.2.173
Accession Number: 1823160
Keywords: Acetylgalactosamine/*analysis
Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Glycoproteins/*chemistry/genetics
Humans
Melanoma/*enzymology
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Recombinant Proteins/chemistry
Sialic Acids/*analysis
Spectrometry, Mass, Fast Atom Bombardment
Tissue Plasminogen Activator/*chemistry/genetics
Transfection
Abstract: We have employed fast atom bombardment mass spectrometry (FAB-MS) to screen the N-linked oligosaccharides of Bowes melanoma tissue plasminogen activator (mt-PA), and recombinant t-PAs produced by Chinese hamster ovary cells (rt-PA) and by a gene-enriched melanoma cell line (rmt-PA). These studies have confirmed the published structures for rt-PA, but are not in agreement with some of the structures reported for mt-PA. In the latter glycoprotein we have identified a novel structure as the major oligosaccharide attached to Asn-184 and Asn-448. This is a biantennary oligosaccharide consisting of a fucosylated trimannosyl core to which are attached two GalNAc(1----4)GlcNAc antennae, one of which carries a sialic acid linked at the 6-position of the GalNAc. Minor constituents are sialylated on both or neither antennae. The sialylated GalNAc moiety is unique in N-linked glycoproteins. The majority of complex structures in rmt-PA contain N-acetyllactosamine moieties at both the Asn-184 and Asn-448 sites with the novel oligosaccharide occurring as a minor component at the Asn-184 site. This study demonstrates the power of mass spectrometric strategies based on high-field two-sector FAB-MS for structure elucidations of natural and recombinant glycoproteins.
Notes: Chan, A L
Morris, H R
Panico, M
Etienne, A T
Rogers, M E
Gaffney, P
Creighton-Kempsford, L
Dell, A
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1991 Mar;1(2):173-85. doi: 10.1093/glycob/1.2.173.
Author Address: Department of Biochemistry, Imperial College of Science Technology and Medicine, London, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1303
Author: Chan, C. P., Mak, T. Y., Chin, K. T., Ng, I. O. and Jin, D. Y.
Year: 2010
Title: N-linked glycosylation is required for optimal proteolytic activation of membrane-bound transcription factor CREB-H
Journal: J Cell Sci
Volume: 123
Issue: Pt 9
Pages: 1438-48
Epub Date: 2010/04/02
Date: May 1
Short Title: N-linked glycosylation is required for optimal proteolytic activation of membrane-bound transcription factor CREB-H
Alternate Journal: Journal of cell science
ISSN: 0021-9533
DOI: 10.1242/jcs.067819
Accession Number: 20356926
Keywords: Animals
Brefeldin A/pharmacology
COS Cells
Cell Line, Tumor
Cell Membrane/drug effects/*metabolism
Cell Nucleus/drug effects/metabolism
Chlorocebus aethiops
Cyclic AMP Response Element-Binding Protein/chemistry/genetics/*metabolism
Endoplasmic Reticulum/drug effects/metabolism
Glycosylation/drug effects
Humans
Mice
Mutant Proteins/metabolism
Proprotein Convertases/metabolism
Protein Binding/drug effects
Protein Folding/drug effects
*Protein Processing, Post-Translational/drug effects
Protein Stability/drug effects
Protein Structure, Quaternary
Protein Transport/drug effects
Serine Endopeptidases/metabolism
Transcription, Genetic/drug effects
Transcriptional Activation/drug effects
Abstract: CREB-H is a liver-enriched bZIP transcription factor of the CREB3 subfamily. CREB-H is activated by intramembrane proteolysis that removes a C-terminal transmembrane domain. Aberrant expression of CREB-H is implicated in liver cancer. In this study we characterized N-linked glycosylation of CREB-H in the luminal domain at the C-terminus. We found that CREB-H is modified at three N-linked glycosylation sites in this region. Disruption of all three sites by site-directed mutagenesis completely abrogated N-linked glycosylation of CREB-H. The unglycosylated mutant of CREB-H was not unstable, unfolded or aggregated. Upon stimulation with an activator of intramembrane proteolysis such as brefeldin A and KDEL-tailed site 1 protease, unglycosylated or deglycosylated CREB-H was largely uncleaved, retained in an inactive form in the endoplasmic reticulum, and less capable of activating transcription driven by unfolded protein response element or C-reactive protein promoter. Taken together, our findings suggest that N-linked glycosylation is required for full activation of CREB-H through intramembrane proteolysis. Our work also reveals a novel mechanism for the regulation of CREB-H-dependent transcription.
Notes: 1477-9137
Chan, Chi-Ping
Mak, To-Yuen
Chin, King-Tung
Ng, Irene Oi-Lin
Jin, Dong-Yan
Journal Article
Research Support, Non-U.S. Gov't
England
J Cell Sci. 2010 May 1;123(Pt 9):1438-48. doi: 10.1242/jcs.067819. Epub 2010 Mar 31.
Author Address: Department of Biochemistry, The University of Hong Kong, Faculty of Medicine Building, 21 Sassoon Road, Pokfulam, Hong Kong.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1235
Author: Chan, L. F., Webb, T. R., Chung, T. T., Meimaridou, E., Cooray, S. N., Guasti, L., Chapple, J. P., Egertová, M., Elphick, M. R., Cheetham, M. E., Metherell, L. A. and Clark, A. J.
Year: 2009
Title: MRAP and MRAP2 are bidirectional regulators of the melanocortin receptor family
Journal: Proc Natl Acad Sci U S A
Volume: 106
Issue: 15
Pages: 6146-51
Epub Date: 2009/03/31
Date: Apr 14
Short Title: MRAP and MRAP2 are bidirectional regulators of the melanocortin receptor family
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0809918106
PMCID: PMC2661846
Accession Number: 19329486
Keywords: Adrenal Glands/metabolism
Amino Acid Sequence
Animals
Brain/metabolism
CHO Cells
Cricetinae
Cricetulus
Gene Expression Regulation
Glycosylation
Humans
Membrane Proteins/chemistry/genetics/*metabolism
Molecular Sequence Data
Nuclear Pore Complex Proteins/metabolism
Organ Specificity
Protein Binding
Protein Multimerization
Receptors, Melanocortin/*metabolism
Sequence Alignment
Signal Transduction
Abstract: The melanocortin receptor (MCR) family consists of 5 G protein-coupled receptors (MC1R-MC5R) with diverse physiologic roles. MC2R is a critical component of the hypothalamic-pituitary-adrenal axis, whereas MC3R and MC4R have an essential role in energy homeostasis. Mutations in MC4R are the single most common cause of monogenic obesity. Investigating the way in which these receptors signal and traffic to the cell membrane is vital in understanding disease processes related to MCR dysfunction. MRAP is an MC2R accessory protein, responsible for adrenal MC2R trafficking and function. Here we identify MRAP2 as a unique homologue of MRAP, expressed in brain and the adrenal gland. We report that MRAP and MRAP2 can interact with all 5 MCRs. This interaction results in MC2R surface expression and signaling. In contrast, MRAP and MRAP2 can reduce MC1R, MC3R, MC4R, and MC5R responsiveness to [Nle4,D-Phe7]alpha-melanocyte-stimulating hormone (NDP-MSH). Collectively, our data identify MRAP and MRAP2 as unique bidirectional regulators of the MCR family.
Notes: 1091-6490
Chan, Li F
Webb, Tom R
Chung, Teng-Teng
Meimaridou, Eirini
Cooray, Sadani N
Guasti, Leonardo
Chapple, J Paul
Egertová, Michaela
Elphick, Maurice R
Cheetham, Michael E
Metherell, Louise A
Clark, Adrian J L
G0600408/Medical Research Council/United Kingdom
G0601140/Medical Research Council/United Kingdom
G0700581/Medical Research Council/United Kingdom
G0801265/Medical Research Council/United Kingdom
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2009 Apr 14;106(15):6146-51. doi: 10.1073/pnas.0809918106. Epub 2009 Mar 27.
Author Address: Queen Mary University of London, Centre for Endocrinology, St. Bartholomew's and Royal London School of Medicine and Dentistry, London EC1M 6BQ, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 400
Author: Chan, S. J., San Segundo, B., McCormick, M. B. and Steiner, D. F.
Year: 1986
Title: Nucleotide and predicted amino acid sequences of cloned human and mouse preprocathepsin B cDNAs
Journal: Proc Natl Acad Sci U S A
Volume: 83
Issue: 20
Pages: 7721-5
Epub Date: 1986/10/01
Date: Oct
Short Title: Nucleotide and predicted amino acid sequences of cloned human and mouse preprocathepsin B cDNAs
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.83.20.7721
PMCID: PMC386793
Accession Number: 3463996
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cathepsin B/analysis/biosynthesis/*genetics
Chromosome Mapping
Cloning, Molecular
Cysteine
DNA/*analysis
Enzyme Precursors/analysis/*genetics
Glycosylation
Humans
Mice
Sequence Homology, Nucleic Acid
Abstract: Cathepsin B is a lysosomal thiol proteinase that may have additional extralysosomal functions. To further our investigations on the structure, mode of biosynthesis, and intracellular sorting of this enzyme, we have determined the complete coding sequences for human and mouse preprocathepsin B by using cDNA clones isolated from human hepatoma and kidney phage libraries. The nucleotide sequences predict that the primary structure of preprocathepsin B contains 339 amino acids organized as follows: a 17-residue NH2-terminal prepeptide sequence followed by a 62-residue propeptide region, 254 residues in mature (single chain) cathepsin B, and a 6-residue extension at the COOH terminus. A comparison of procathepsin B sequences from three species (human, mouse, and rat) reveals that the homology between the propeptides is relatively conserved with a minimum of 68% sequence identity. In particular, two conserved sequences in the propeptide that may be functionally significant include a potential glycosylation site and the presence of a single cysteine at position 59. Comparative analysis of the three sequences also suggests that processing of procathepsin B is a multistep process, during which enzymatically active intermediate forms may be generated. The availability of the cDNA clones will facilitate the identification of possible active or inactive intermediate processive forms as well as studies on the transcriptional regulation of the cathepsin B gene.
Notes: 1091-6490
Chan, S J
San Segundo, B
McCormick, M B
Steiner, D F
AM 13914/AM/NIADDK NIH HHS/United States
AM 20595/AM/NIADDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1986 Oct;83(20):7721-5. doi: 10.1073/pnas.83.20.7721.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1497
Author: Chan, W. K., Chen, V. P., Luk, W. K., Choi, R. C. and Tsim, K. W.
Year: 2012
Title: N-linked glycosylation of proline-rich membrane anchor (PRiMA) is not required for assembly and trafficking of globular tetrameric acetylcholinesterase
Journal: Neurosci Lett
Volume: 523
Issue: 1
Pages: 71-5
Epub Date: 2012/07/04
Date: Aug 8
Short Title: N-linked glycosylation of proline-rich membrane anchor (PRiMA) is not required for assembly and trafficking of globular tetrameric acetylcholinesterase
Alternate Journal: Neuroscience letters
ISSN: 0304-3940
DOI: 10.1016/j.neulet.2012.06.045
Accession Number: 22750213
Keywords: Acetylcholinesterase/*chemistry/*metabolism
Animals
Dimerization
Glycosylation
HEK293 Cells
Humans
Membrane Proteins/*chemistry/*metabolism
Mice
Multiprotein Complexes/chemistry/metabolism
Nerve Tissue Proteins/*chemistry/*metabolism
Protein Binding/physiology
Protein Transport
Abstract: Acetylcholinesterase (AChE) is organized into globular tetramers (G(4)) by a structural protein called proline-rich membrane anchor (PRiMA), anchoring it into the cell membrane of neurons in the brain. The assembly of AChE tetramers with PRiMA requires the presence of a C-terminal "t-peptide" in the AChE catalytic subunit (AChE(T)). The glycosylation of AChE(T) is known to be required for its proper assembly and trafficking; however, the role of PRiMA glycosylation in the oligomer assembly has not been revealed. PRiMA is a glycoprotein containing two putative N-linked glycosylation sites. By using site-directed mutagenesis, the asparagine-43 was identified to be the N-linked glycosylation site of PRiMA. Abolishing glycosylation on mouse PRiMA appeared not to affect its assembly with AChE(T), the enzymatic properties of AChE, and the membrane trafficking of PRiMA-linked AChE tetramers. This result is contrary to the reports that glycosylation is essential for conformation and trafficking of membrane glycoproteins.
Notes: 1872-7972
Chan, Wallace K B
Chen, Vicky P
Luk, Wilson K W
Choi, Roy C Y
Tsim, Karl W K
Journal Article
Research Support, Non-U.S. Gov't
Ireland
Neurosci Lett. 2012 Aug 8;523(1):71-5. doi: 10.1016/j.neulet.2012.06.045. Epub 2012 Jun 26.
Author Address: Division of Life Science, Center for Chinese Medicine and State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay Road, Hong Kong Special Administrative Region.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1647
Author: Chandler, K. B., Brnakova, Z., Sanda, M., Wang, S., Stalnaker, S. H., Bridger, R., Zhao, P., Wells, L., Edwards, N. J. and Goldman, R.
Year: 2014
Title: Site-specific glycan microheterogeneity of inter-alpha-trypsin inhibitor heavy chain H4
Journal: J Proteome Res
Volume: 13
Issue: 7
Pages: 3314-29
Epub Date: 2014/06/03
Date: Jul 3
Short Title: Site-specific glycan microheterogeneity of inter-alpha-trypsin inhibitor heavy chain H4
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr500394z
PMCID: PMC4084840
Accession Number: 24884609
Keywords: Amino Acid Sequence
Blood Proteins/chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Glycoproteins/*blood/chemistry
Glycosylation
HEK293 Cells
Humans
Molecular Sequence Data
Protein Isoforms
*Protein Processing, Post-Translational
Proteinase Inhibitory Proteins, Secretory/*blood/chemistry
Abstract: Inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4) is a 120 kDa acute-phase glycoprotein produced primarily in the liver, secreted into the blood, and identified in serum. ITIH4 is involved in liver development and stabilization of the extracellular matrix (ECM), and its expression is altered in liver disease. In this study, we aimed to characterize glycosylation of recombinant and serum-derived ITIH4 using analytical mass spectrometry. Recombinant ITIH4 was analyzed to optimize glycopeptide analyses, followed by serum-derived ITIH4. First, we confirmed that the four ITIH4 N-X-S/T sequons (N81, N207, N517, and N577) were glycosylated by treating ITIH4 tryptic/GluC glycopeptides with PNGaseF in the presence of (18)O water. Next, we performed glycosidase-assisted LC-MS/MS analysis of ITIH4 trypsin-GluC glycopeptides enriched via hydrophilic interaction liquid chromatography to characterize ITIH4 N-glycoforms. While microheterogeneity of N-glycoforms differed between ITIH4 protein expressed in HEK293 cells and protein isolated from serum, occupancy of N-glycosylation sites did not differ. A fifth N-glycosylation site was discovered at N274 with the rare nonconsensus NVV motif. Site N274 contained high-mannose N-linked glycans in both serum and recombinant ITIH4. We also identified isoform-specific ITIH4 O-glycoforms and documented that utilization of O-glycosylation sites on ITIH4 differed between the cell line and serum.
Notes: 1535-3907
Chandler, Kevin Brown
Brnakova, Zuzana
Sanda, Miloslav
Wang, Shuo
Stalnaker, Stephanie H
Bridger, Robert
Zhao, Peng
Wells, Lance
Edwards, Nathan J
Goldman, Radoslav
P30CA51008/CA/NCI NIH HHS/United States
R01 CA126189/CA/NCI NIH HHS/United States
CA126189/CA/NCI NIH HHS/United States
P41 GM103490/GM/NIGMS NIH HHS/United States
R01CA135069/CA/NCI NIH HHS/United States
P30 CA051008/CA/NCI NIH HHS/United States
U01 CA168926/CA/NCI NIH HHS/United States
R01 CA135069/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Proteome Res. 2014 Jul 3;13(7):3314-29. doi: 10.1021/pr500394z. Epub 2014 Jun 12.
Author Address: Department of Biochemistry and Molecular & Cellular Biology, Georgetown University , Washington, D.C. 20057, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1821
Author: Chandler, K. B., Leon, D. R., Meyer, R. D., Rahimi, N. and Costello, C. E.
Year: 2017
Title: Site-Specific N-Glycosylation of Endothelial Cell Receptor Tyrosine Kinase VEGFR-2
Journal: J Proteome Res
Volume: 16
Issue: 2
Pages: 677-688
Epub Date: 2016/12/15
Date: Feb 3
Short Title: Site-Specific N-Glycosylation of Endothelial Cell Receptor Tyrosine Kinase VEGFR-2
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/acs.jproteome.6b00738
PMCID: PMC5291786
NIHMSID: NIHMS842607
Accession Number: 27966990
Keywords: Amino Acid Sequence/genetics
Animals
Aorta/metabolism
Endothelial Cells/*metabolism
Glycopeptides/*genetics/metabolism
Glycosylation
Humans
Peptides
Polysaccharides/genetics/*metabolism
Protein Binding
Swine
Tandem Mass Spectrometry
Vascular Endothelial Growth Factor Receptor-2/*genetics/metabolism
*N-glycosylation
*Rtk
*Vegfr-2
*receptor tyrosine kinase
*vascular endothelial growth factor receptor-2
Abstract: Vascular endothelial growth factor receptor-2 (VEGFR-2) is an important receptor tyrosine kinase (RTK) that plays critical roles in both physiologic and pathologic angiogenesis. The extracellular domain of VEGFR-2 is composed of seven immunoglobulin-like domains, each with multiple potential N-glycosylation sites (sequons). N-glycosylation plays a central role in RTK ligand binding, trafficking, and stability. However, despite its importance, the functional role of N-glycosylation of VEGFR-2 remains poorly understood. The objectives of the present study were to characterize N-glycosylation sites in VEGFR-2 via enzymatic release of the glycans and concomitant incorporation of (18)O into formerly N-glycosylated sites followed by tandem mass spectrometry (MS/MS) analysis to determine N-glycosylation site occupancy and the site-specific N-glycan heterogeneity of VEGFR-2 glycopeptides. The data demonstrated that all seven VEGFR-2 immunoglobulin-like domains have at least one occupied N-glycosylation site. MS/MS analyses of glycopeptides and deamidated, deglycosylated (PNGase F-treated) peptides from ectopically expressed VEGFR-2 in porcine aortic endothelial (PAE) cells identified N-glycans at the majority of the 17 potential N-glycosylation sites on VEGFR-2 in a site-specific manner. The data presented here provide direct evidence for site-specific, heterogeneous N-glycosylation and N-glycosylation site occupancy on VEGFR-2. The study has important implications for the therapeutic targeting of VEGFR-2, ligand binding, trafficking, and signaling.
Notes: 1535-3907
Chandler, Kevin Brown
Orcid: 0000-0001-5514-9652
Leon, Deborah R
Meyer, Rosana D
Rahimi, Nader
Costello, Catherine E
Orcid: 0000-0003-1594-5122
F32 CA196157/CA/NCI NIH HHS/United States
P41 GM104603/GM/NIGMS NIH HHS/United States
R21 CA191970/CA/NCI NIH HHS/United States
S10 OD010724/OD/NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Proteome Res. 2017 Feb 3;16(2):677-688. doi: 10.1021/acs.jproteome.6b00738. Epub 2016 Dec 29.
Author Address: Center for Biomedical Mass Spectrometry, Department of Biochemistry and ‡Department of Pathology and Laboratory Medicine, Boston University School of Medicine , Boston, Massachusetts 02118, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1998
Author: Chandler, K. B., Mehta, N., Leon, D. R., Suscovich, T. J., Alter, G. and Costello, C. E.
Year: 2019
Title: Multi-isotype Glycoproteomic Characterization of Serum Antibody Heavy Chains Reveals Isotype- and Subclass-Specific N-Glycosylation Profiles
Journal: Mol Cell Proteomics
Volume: 18
Issue: 4
Pages: 686-703
Epub Date: 2019/01/20
Date: Apr
Short Title: Multi-isotype Glycoproteomic Characterization of Serum Antibody Heavy Chains Reveals Isotype- and Subclass-Specific N-Glycosylation Profiles
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.RA118.001185
PMCID: PMC6442369
Accession Number: 30659065
Keywords: Amino Acid Sequence
Glycoproteins/*blood/chemistry/metabolism
Glycosylation
Humans
Immunoglobulin Heavy Chains/*blood/chemistry
Immunoglobulin Isotypes/*blood
*Proteomics
*Antibodies*
*Glycoproteomics
*Immunoglobulin G1 (IgG1)
*Immunoglobulin G4 (IgG4)
*Immunoglobulin M (IgM)
*Immunoglobulins A1 and A2 (IgA1 and IgA2)
*Immunology*
*N-Glycosylation
*Serum/Plasma*
*sequence variant
Abstract: Antibodies are critical glycoproteins that bridge the innate and adaptive immune systems to provide protection against infection. The isotype/subclass of the antibody, the co-translational N-glycosylation on the CH2 domain, and the remodeling of the N-linked glycans during passage through the ER and Golgi are the known variables within the Fc domain that program antibody effector function. Through investigations of monoclonal therapeutics, it has been observed that addition or removal of specific monosaccharide residues from antibody N-glycans can influence the potency of antibodies, highlighting the importance of thoroughly characterizing antibody N-glycosylation. Although IgGs usually have a single N-glycosylation site and are well studied, other antibody isotypes, e.g. IgA and IgM, that are the first responders in certain diseases, have two to five sites/monomer of antibody, and little is known about their N-glycosylation. Here we employ a nLC-MS/MS method using stepped-energy higher energy collisional dissociation to characterize the N-glycan repertoire and site occupancy of circulating serum antibodies. We simultaneously determined the site-specific N-linked glycan repertoire for IgG1, IgG4, IgA1, IgA2, and IgM in individual healthy donors. Compared with IgG1, IgG4 displayed a higher relative abundance of G1S1F and a lower relative abundance of G1FB. IgA1 and IgA2 displayed mostly biantennary N-glycans. IgA2 variants with the either serine (S93) or proline (P93) were detected. In digests of the sera from a subset of donors, we detected an unmodified peptide containing a proline residue at position 93; this substitution would strongly disfavor N-glycosylation at N92. IgM sites N46, N209, and N272 displayed mostly complex glycans, whereas sites N279 and N439 displayed higher relative abundances of high-mannose glycoforms. This multi-isotype approach is a crucial step toward developing a platform to define disease-specific N-glycan signatures for different isotypes to help tune antibodies to induce protection. Data are available via ProteomeXchange with identifier PXD010911.
Notes: 1535-9484
Chandler, Kevin Brown
Mehta, Nickita
Leon, Deborah R
Suscovich, Todd J
Alter, Galit
Costello, Catherine E
S10 OD021651/OD/NIH HHS/United States
F32 CA196157/CA/NCI NIH HHS/United States
R37 AI080289/AI/NIAID NIH HHS/United States
R24 GM134210/GM/NIGMS NIH HHS/United States
S10 OD010724/OD/NIH HHS/United States
P41 GM104603/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Mol Cell Proteomics. 2019 Apr;18(4):686-703. doi: 10.1074/mcp.RA118.001185. Epub 2019 Jan 18.
Author Address: From the ‡Center for Biomedical Mass Spectrometry, Boston University School of Medicine, Boston, Massachusetts.
Ragon Institute of MGH, MIT and Harvard, Cambridge, Massachusetts.
From the ‡Center for Biomedical Mass Spectrometry, Boston University School of Medicine, Boston, Massachusetts;. Electronic address: cecmsms@bu.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1569
Author: Chandler, K. B., Pompach, P., Goldman, R. and Edwards, N.
Year: 2013
Title: Exploring site-specific N-glycosylation microheterogeneity of haptoglobin using glycopeptide CID tandem mass spectra and glycan database search
Journal: J Proteome Res
Volume: 12
Issue: 8
Pages: 3652-66
Epub Date: 2013/07/09
Date: Aug 2
Short Title: Exploring site-specific N-glycosylation microheterogeneity of haptoglobin using glycopeptide CID tandem mass spectra and glycan database search
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr400196s
PMCID: PMC3777712
NIHMSID: NIHMS506032
Accession Number: 23829323
Keywords: *Algorithms
Amino Acid Sequence
*Artifacts
Chromatography, Liquid
Databases, Factual
Glycosylation
Haptoglobins/*chemistry
Humans
Molecular Sequence Data
Peptide Fragments/*analysis
Polysaccharides/*analysis
Proteolysis
*Software
Tandem Mass Spectrometry
Trypsin/chemistry
Abstract: Glycosylation is a common protein modification with a significant role in many vital cellular processes and human diseases, making the characterization of protein-attached glycan structures important for understanding cell biology and disease processes. Direct analysis of protein N-glycosylation by tandem mass spectrometry of glycopeptides promises site-specific elucidation of N-glycan microheterogeneity, something that detached N-glycan and deglycosylated peptide analyses cannot provide. However, successful implementation of direct N-glycopeptide analysis by tandem mass spectrometry remains a challenge. In this work, we consider algorithmic techniques for the analysis of LC-MS/MS data acquired from glycopeptide-enriched fractions of enzymatic digests of purified proteins. We implement a computational strategy that takes advantage of the properties of CID fragmentation spectra of N-glycopeptides, matching the MS/MS spectra to peptide-glycan pairs from protein sequences and glycan structure databases. Significantly, we also propose a novel false discovery rate estimation technique to estimate and manage the number of false identifications. We use a human glycoprotein standard, haptoglobin, digested with trypsin and GluC, enriched for glycopeptides using HILIC chromatography, and analyzed by LC-MS/MS to demonstrate our algorithmic strategy and evaluate its performance. Our software, GlycoPeptideSearch (GPS), assigned glycopeptide identifications to 246 of the spectra at a false discovery rate of 5.58%, identifying 42 distinct haptoglobin peptide-glycan pairs at each of the four haptoglobin N-linked glycosylation sites. We further demonstrate the effectiveness of this approach by analyzing plasma-derived haptoglobin, identifying 136 N-linked glycopeptide spectra at a false discovery rate of 0.4%, representing 15 distinct glycopeptides on at least three of the four N-linked glycosylation sites. The software, GlycoPeptideSearch, is available for download from http://edwardslab.bmcb.georgetown.edu/GPS .
Notes: 1535-3907
Chandler, Kevin Brown
Pompach, Petr
Goldman, Radoslav
Edwards, Nathan
P30 CA051008/CA/NCI NIH HHS/United States
R01 CA135069/CA/NCI NIH HHS/United States
U01 CA168926/CA/NCI NIH HHS/United States
P30 CA51008/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Proteome Res. 2013 Aug 2;12(8):3652-66. doi: 10.1021/pr400196s. Epub 2013 Jul 22.
Author Address: Department of Biochemistry and Molecular & Cellular Biology, Georgetown University Medical Center, Washington, DC 20007, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1402
Author: Chandrasekhar, K. D., Lvov, A., Terrenoire, C., Gao, G. Y., Kass, R. S. and Kobertz, W. R.
Year: 2011
Title: O-glycosylation of the cardiac I(Ks) complex
Journal: J Physiol
Volume: 589
Issue: Pt 15
Pages: 3721-30
Epub Date: 2011/06/15
Date: Aug 1
Short Title: O-glycosylation of the cardiac I(Ks) complex
Alternate Journal: The Journal of physiology
ISSN: 0022-3751 (Print)
0022-3751
DOI: 10.1113/jphysiol.2011.211284
PMCID: PMC3171881
Accession Number: 21669976
Keywords: Action Potentials/physiology
Amino Acid Sequence
Animals
Arrhythmias, Cardiac/metabolism
Asparagine/metabolism
CHO Cells
Cells, Cultured
Cricetinae
Cricetulus
Glycosylation
Humans
KCNQ1 Potassium Channel/genetics/*metabolism
Mice
Mice, Transgenic/metabolism
Molecular Sequence Data
Mutation
Myocardium/*metabolism
Myocytes, Cardiac/metabolism
Polysaccharides/metabolism
Potassium Channels, Voltage-Gated/genetics/metabolism
Protein Isoforms/metabolism
*Protein Processing, Post-Translational
Protein Transport/physiology
Recombinant Fusion Proteins/genetics/*metabolism
Threonine/genetics/metabolism
Abstract: Post-translational modifications of the KCNQ1–KCNE1 (Kv7) K+ channel complex are vital for regulation of the cardiac IKs current and action potential duration. Here, we show the KCNE1 regulatory subunit is O-glycosylated with mucin-type glycans in vivo. As O-linked glycosylation sites are not recognizable by sequence gazing, we designed a novel set of glycosylation mutants and KCNE chimeras and analysed their glycan content using deglycosylation enzymes. Our results show that KCNE1 is exclusively O-glycosylated at Thr-7, which is also required for N-glycosylation at Asn-5. For wild type KCNE1, the overlapping N- and O-glycosylation sites are innocuous for subunit biogenesis; however, mutation of Thr-7 to a non-hydroxylated residue yielded mostly unglycosylated protein and a small fraction of mono-N-glycosylated protein. The compounded hypoglycosylation was equally deleterious for KCNQ1–KCNE1 cell surface expression, demonstrating that KCNE1 O-glycosylation is a post-translational modification that is integral for the proper biogenesis and anterograde trafficking of the cardiac IKs complex. The enzymatic assays and panel of glycosylation mutants used here will be valuable for identifying the different KCNE1 glycoforms in native cells and determining the roles N- and O-glycosylation play in KCNQ1–KCNE1 function and localization in cardiomyocytes
Notes: 1469-7793
Chandrasekhar, Kshama D
Lvov, Anatoli
Terrenoire, Cecile
Gao, Grace Y
Kass, Robert S
Kobertz, William R
R01 DC007669/DC/NIDCD NIH HHS/United States
R01 DC007669-05/DC/NIDCD NIH HHS/United States
DC-007669/DC/NIDCD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Physiol. 2011 Aug 1;589(Pt 15):3721-30. doi: 10.1113/jphysiol.2011.211284. Epub 2011 Jun 13.
Author Address: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-2324, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1788
Author: Chang, C. F., Hsu, L. S., Weng, C. Y., Chen, C. K., Wang, S. Y., Chou, Y. H., Liu, Y. Y., Yuan, Z. X., Huang, W. Y., Lin, H., Chen, Y. H. and Tsai, J. N.
Year: 2016
Title: N-Glycosylation of Human R-Spondin 1 Is Required for Efficient Secretion and Stability but Not for Its Heparin Binding Ability
Journal: Int J Mol Sci
Volume: 17
Issue: 6
Epub Date: 2016/06/18
Date: Jun 14
Short Title: N-Glycosylation of Human R-Spondin 1 Is Required for Efficient Secretion and Stability but Not for Its Heparin Binding Ability
Alternate Journal: International journal of molecular sciences
ISSN: 1422-0067
DOI: 10.3390/ijms17060937
PMCID: PMC4926470
Accession Number: 27314333
Keywords: Binding Sites
Endoplasmic Reticulum/metabolism
Glycosylation
HEK293 Cells
Heparin/*metabolism
Humans
Protein Binding
Protein Folding
*Protein Processing, Post-Translational
Protein Stability
*Secretory Pathway
Thrombospondins/chemistry/*metabolism
N-glycosylation
R-spondin 1
Wnt signaling
secretion
stability
Abstract: R-spondin 1 (Rspo1) plays an essential role in stem cell biology by potentiating Wnt signaling activity. Despite the fact that Rspo1 holds therapeutic potential for a number of diseases, its biogenesis is not fully elucidated. All Rspo proteins feature two amino-terminal furin-like repeats, which are responsible for Wnt signal potentiation, and a thrombospondin type 1 (TSR1) domain that can provide affinity towards heparan sulfate proteoglycans. Using chemical inhibitors, deglycosylase and site-directed mutagenesis, we found that human Rspo1 and Rspo3 are both N-glycosylated at N137, a site near the C-terminus of the furin repeat 2 domain, and Rspo2 is N-glycosylated at N160, a position near the N-terminus of TSR1 domain. Elimination of N-glycosylation at these sites affects their accumulation in media but have no effect on the ability towards heparin. Introduction of the N-glycosylation site to Rspo2 mutant at the position homologous to N137 in Rspo1 restored full glycosylation and rescued the accumulation defect of nonglycosylated Rspo2 mutant in media. Similar effect can be observed in the N137 Rspo1 or Rspo3 mutant engineered with Rspo2 N-glycosylation site. The results highlight the importance of N-glycosylation at these two positions in efficient folding and secretion of Rspo family. Finally, we further showed that human Rspo1 is subjected to endoplasmic reticulum (ER) quality control in N-glycan-dependent manner. While N-glycan of Rspo1 plays a role in its intracellular stability, it had little effect on secreted Rspo1. Our findings provide evidence for the critical role of N-glycosylation in the biogenesis of Rspo1.
Notes: 1422-0067
Chang, Chiung-Fang
Hsu, Li-Sung
Weng, Chieh-Yu
Chen, Chih-Kai
Wang, Shu-Ying
Chou, Yi-Hwa
Liu, Yan-Yu
Yuan, Zi-Xiu
Huang, Wen-Ying
Lin, Ho
Chen, Yau-Hung
Tsai, Jen-Ning
Journal Article
Int J Mol Sci. 2016 Jun 14;17(6):937. doi: 10.3390/ijms17060937.
Author Address: Department of Life Sciences, National Chung Hsing University, Taichung 40227, Taiwan. d099052009@mail.nchu.edu.tw.
Institute of Biochemistry and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. lsh316@csmu.edu.tw.
Clinical Laboratory, Chung Shan Medical University Hospital, Taichung 40201, Taiwan. lsh316@csmu.edu.tw.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. elsie1009@hotmail.com.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. okai1025@hotmail.com.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. lucywang0515@gmail.com.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. a26001234@gmail.com.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. xprfish1210@gmail.com.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. a12456910@gmail.com.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. f920719@yahoo.com.tw.
Department of Life Sciences, National Chung Hsing University, Taichung 40227, Taiwan. hlin@dragon.nchu.edu.tw.
Department of Chemistry, Tamkang University, Tamsui, New Taipei City 25137, Taiwan. yauhung@mail.tku.edu.tw.
Clinical Laboratory, Chung Shan Medical University Hospital, Taichung 40201, Taiwan. jeningts@csmu.edu.tw.
Department of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung 40201, Taiwan. jeningts@csmu.edu.tw.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 507
Author: Chang, X. B., Hou, Y. X., Jensen, T. J. and Riordan, J. R.
Year: 1994
Title: Mapping of cystic fibrosis transmembrane conductance regulator membrane topology by glycosylation site insertion
Journal: J Biol Chem
Volume: 269
Issue: 28
Pages: 18572-5
Epub Date: 1994/07/15
Date: Jul 15
Short Title: Mapping of cystic fibrosis transmembrane conductance regulator membrane topology by glycosylation site insertion
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7518437
Keywords: 1-Methyl-3-isobutylxanthine/pharmacology
Algorithms
Amino Acid Sequence
Animals
Base Sequence
Bucladesine/pharmacology
CHO Cells
Cell Membrane/metabolism/ultrastructure
Chloride Channels/biosynthesis/chemistry
Colforsin/pharmacology
Consensus Sequence
Cricetinae
Cystic Fibrosis Transmembrane Conductance Regulator
Glycosylation
Humans
Lipid Bilayers
Mammals
Membrane Proteins/biosynthesis/*chemistry/*physiology
Molecular Sequence Data
Mutagenesis, Site-Directed
*Protein Structure, Secondary
Recombinant Proteins/biosynthesis/chemistry/drug effects
Transfection
Abstract: Technical difficulties in obtaining three-dimensional structures of intrinsic membrane proteins continues to limit understanding of their function. However, considerable insight can be gained from their two-dimensional topological arrangement in the lipid bilayer. Efficient molecular genetic approaches are available to discern the topology of prokaryotic but not of eukaryotic membrane proteins. The absolute asymmetry of the sidedness of their N-glycosylation was employed here to develop such a method using the cystic fibrosis transmembrane conductance regulator (CFTR). Insertion by in vitro mutagenesis of N-glycosylation consensus sequences (NXS/T) in predicted cytoplasmic and extracytoplasmic loops between hydrophobic sequences capable of traversing the membrane established the membrane topology of CFTR. This provides the first experimental evaluation of the original topological model of CFTR based solely on hydropathy algorithms and a method which may be generally applicable for the in vivo evaluation of the topology of other mammalian membrane proteins.
Notes: Chang, X B
Hou, Y X
Jensen, T J
Riordan, J R
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1994 Jul 15;269(28):18572-5.
Author Address: Research Institute, Hospital for Sick Children, Toronto, Ontario, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1105
Author: Chao, K. L., Muthukumar, L. and Herzberg, O.
Year: 2007
Title: Structure of human hyaluronidase-1, a hyaluronan hydrolyzing enzyme involved in tumor growth and angiogenesis
Journal: Biochemistry
Volume: 46
Issue: 23
Pages: 6911-20
Epub Date: 2007/05/17
Date: Jun 12
Short Title: Structure of human hyaluronidase-1, a hyaluronan hydrolyzing enzyme involved in tumor growth and angiogenesis
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi700382g
Accession Number: 17503783
Keywords: Amino Acid Sequence
Binding Sites
Conserved Sequence
Crystallography, X-Ray
Genetic Variation
Humans
Hyaluronoglucosaminidase/*chemistry/genetics/*metabolism
Models, Molecular
Molecular Sequence Data
Neoplasms/blood supply/enzymology/*pathology
Neovascularization, Pathologic/*enzymology
Protein Conformation
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: Mammalian hyaluronidases hydrolyze hyaluronan, a polysaccharide of diverse physiological roles found in all tissues and body fluids. In addition to its function in normal cellular hyaluronan turnover, human hyaluronidase-1 is implicated in cancer proliferation, angiogenesis, and inflammatory diseases; its expression is up-regulated in advanced stages of bladder cancer, whereas the expression of the alternative splice-variants is down-regulated. The crystal structure reveals a molecule composed of two closely associated domains: a catalytic domain that adopts a distorted (beta/alpha)8 barrel resembling that of bee venom hyaluronidase, and a novel, EGF-like domain, characteristic of involvement in protein-protein interactions and regulatory processes. The structure shows that the fold of this unique EGF-like domain is intact in four alternative splice-variants, whereas the catalytic domain is likely to be unfolded. Thus, these variants may function by competing with the full-length enzyme for the putative protein partner and regulating enzymatic activity in healthy cells.
Notes: Chao, Kinlin L
Muthukumar, Lavanya
Herzberg, Osnat
P01-GM057890/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
United States
Biochemistry. 2007 Jun 12;46(23):6911-20. doi: 10.1021/bi700382g. Epub 2007 May 16.
Author Address: Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute, Rockville, Maryland 20850, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1503
Author: Chao, K. L., Tsai, I. W., Chen, C. and Herzberg, O.
Year: 2012
Title: Crystal structure of the Sema-PSI extracellular domain of human RON receptor tyrosine kinase
Journal: PLoS One
Volume: 7
Issue: 7
Pages: e41912
Epub Date: 2012/08/01
Short Title: Crystal structure of the Sema-PSI extracellular domain of human RON receptor tyrosine kinase
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0041912
PMCID: PMC3405059
Accession Number: 22848655
Keywords: Amino Acid Sequence
Crystallography, X-Ray
Extracellular Space/*enzymology
Humans
Ligands
Models, Molecular
Molecular Sequence Data
Protein Multimerization
Protein Structure, Tertiary
Proteolysis
Proto-Oncogene Proteins c-met/chemistry
Receptor Protein-Tyrosine Kinases/*chemistry/metabolism
Abstract: Human RON (Recepteur d'Origine Nantais) receptor tyrosine kinase is a cell surface receptor for Macrophage Stimulating Protein (MSP). RON mediates signal transduction pathways that regulate cell adhesion, invasion, motility and apoptosis processes. Elevated levels of RON and its alternatively spliced variants are implicated in the progression and metastasis of tumor cells. The binding of MSP α/β heterodimer to the extracellular region of RON receptor induces receptor dimerization and activation by autophosphorylation of the intracellular kinase domains. The ectodomain of RON, containing the ligand recognition and dimerization domains, is composed of a semaphorin (Sema), Plexins-Semaphorins-Integrins domain (PSI), and four Immunoglobulins-Plexins-Transcription factor (IPT) domains. High affinity association between MSP and RON is mediated by the interaction between MSP β-chain and RON Sema, although RON activation requires intact RON and MSP proteins. Here, we report the structure of RON Sema-PSI domains at 1.85 Å resolution. RON Sema domain adopts a seven-bladed β-propeller fold, followed by disulfide bond rich, cysteine-knot PSI motif. Comparison with the homologous Met receptor tyrosine kinase reveals that RON Sema-PSI contains distinguishing secondary structural features. These define the receptors' exclusive selectivity towards their respective ligands, RON for MSP and Met for HGF. The RON Sema-PSI crystal packing generates a homodimer with interface formed by the Sema domain. Mapping of the dimer interface using the RON homology to Met, MSP homology to Hepatocyte Growth Factor (HGF), and the structure of the Met/HGF complex shows the dimer interface overlapping with the putative MSPβ binding site. The crystallographically determined RON Sema-PSI homodimer may represent the dimer assembly that occurs during ligand-independent receptor activation and/or the inhibition of the constitutive activity of RONΔ160 splice variant by the soluble RON splice variant, RONΔ85.
Notes: 1932-6203
Chao, Kinlin L
Tsai, I-Wei
Chen, Chen
Herzberg, Osnat
R01 GM087922/GM/NIGMS NIH HHS/United States
R21 DA027024/DA/NIDA NIH HHS/United States
R01-GM087922/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
PLoS One. 2012;7(7):e41912. doi: 10.1371/journal.pone.0041912. Epub 2012 Jul 25.
Author Address: Institute for Bioscience and Biotechnology Research, University of Maryland, Rockville, Maryland, United States of America.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 606
Author: Chapman, B. S., Eckart, M. R., Kaufman, S. E. and Lapointe, G. R.
Year: 1996
Title: O-linked oligosaccharide on the 75-kDa neurotrophin receptor
Journal: J Neurochem
Volume: 66
Issue: 4
Pages: 1707-16
Epub Date: 1996/04/01
Date: Apr
Short Title: O-linked oligosaccharide on the 75-kDa neurotrophin receptor
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042 (Print)
0022-3042
DOI: 10.1046/j.1471-4159.1996.66041707.x
Accession Number: 8627329
Keywords: Animals
Biotin
CHO Cells/chemistry
Carbohydrates/chemistry
Cricetinae
Electrophoresis, Polyacrylamide Gel
Humans
Molecular Weight
Oligosaccharides/*analysis
Polysaccharides/chemistry/metabolism
Receptors, Nerve Growth Factor/biosynthesis/*chemistry/metabolism
Signal Transduction/physiology
Abstract: Four neurotrophic factors, important for survival and function of neurons, bind a common receptor, the 75-kDa neurotrophin receptor (NTR). An O-glycosylated peptide connects the ligand-binding domain of NTR to its transmembrane helix. This peptide, the transmembrane helix, and intracellular sequences are highly conserved in vertebrate evolution. To investigate the structure and function of O-glycosylation on NTR, we produced the extracellular domains by expression in mammalian cells. Addition during biosynthesis of O-linked glycans was evaluated, and structures were characterized by lectin blotting and glycosidase digestion. Effects of disialylation, deglycosylation, and lectin attachment on the equilibrium binding constant were measured. Addition of O-linked glycans during biosynthesis was found to have a large effect on NTR structure assessed by mobility in polyacrylamide gels. NTR O-linked glycans synthesized by cultured cells had the structure (NeuNAc)(1-2-) Gal beta 1-3GalNAc. Modification of the O-linked oligosaccharide produced small, possibly significant effects on the binding constant of NTR for nerve growth factor. The results are discussed in reference to a potential role for the stalk region in ligand binding and signaling.
Notes: Chapman, B S
Eckart, M R
Kaufman, S E
Lapointe, G R
Journal Article
England
J Neurochem. 1996 Apr;66(4):1707-16. doi: 10.1046/j.1471-4159.1996.66041707.x.
Author Address: Department of Pharmaceutical Chemistry, University of California School of Pharmacy, San Francisco, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1711
Author: Chappell, P. E., Garner, L. I., Yan, J., Metcalfe, C., Hatherley, D., Johnson, S., Robinson, C. V., Lea, S. M. and Brown, M. H.
Year: 2015
Title: Structures of CD6 and Its Ligand CD166 Give Insight into Their Interaction
Journal: Structure
Volume: 23
Issue: 8
Pages: 1426-1436
Epub Date: 2015/07/07
Date: Aug 4
Short Title: Structures of CD6 and Its Ligand CD166 Give Insight into Their Interaction
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2015.05.019
PMCID: PMC4533223
Accession Number: 26146185
Keywords: Amino Acid Motifs
Animals
Antigens, CD/*chemistry/genetics
Antigens, Differentiation, T-Lymphocyte/*chemistry/genetics
Binding Sites
CHO Cells
Cell Adhesion Molecules, Neuronal/*chemistry/genetics
Cloning, Molecular
Cricetulus
Crystallography, X-Ray
Escherichia coli/genetics/metabolism
Fetal Proteins/*chemistry/genetics
Gene Expression
Glycosylation
Humans
*Models, Molecular
Molecular Sequence Data
Mutation
*Polymorphism, Single Nucleotide
Protein Binding
Protein Interaction Domains and Motifs
Protein Multimerization
Protein Structure, Secondary
Recombinant Fusion Proteins/chemistry/genetics
Static Electricity
Abstract: CD6 is a transmembrane protein with an extracellular region containing three scavenger receptor cysteine rich (SRCR) domains. The membrane proximal domain of CD6 binds the N-terminal immunoglobulin superfamily (IgSF) domain of another cell surface receptor, CD166, which also engages in homophilic interactions. CD6 expression is mainly restricted to T cells, and the interaction between CD6 and CD166 regulates T-cell activation. We have solved the X-ray crystal structures of the three SRCR domains of CD6 and two N-terminal domains of CD166. This first structure of consecutive SRCR domains reveals a nonlinear organization. We characterized the binding sites on CD6 and CD166 and showed that a SNP in CD6 causes glycosylation that hinders the CD6/CD166 interaction. Native mass spectrometry analysis showed that there is competition between the heterophilic and homophilic interactions. These data give insight into how interactions of consecutive SRCR domains are perturbed by SNPs and potential therapeutic reagents.
Notes: 1878-4186
Chappell, Paul E
Garner, Lee I
Yan, Jun
Metcalfe, Clive
Hatherley, Deborah
Johnson, Steven
Robinson, Carol V
Lea, Susan M
Brown, Marion H
100298/Wellcome Trust/United Kingdom
100298/Z/12/Z/Wellcome Trust/United Kingdom
BB/F016085/1/Biotechnology and Biological Sciences Research Council/United Kingdom
MR/M011984/1/Medical Research Council/United Kingdom
G0400808/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Structure. 2015 Aug 4;23(8):1426-1436. doi: 10.1016/j.str.2015.05.019. Epub 2015 Jul 2.
Author Address: Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
Department of Chemistry, Physical and Theoretical Chemistry Laboratory, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK. Electronic address: marion.brown@path.ox.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 731
Author: Charlwood, J., Birrell, H. and Camilleri, P.
Year: 1999
Title: Carbohydrate release from picomole quantities of glycoprotein and characterisation of glycans by high-performance liquid chromatography and mass spectrometry
Journal: J Chromatogr B Biomed Sci Appl
Volume: 734
Issue: 1
Pages: 169-74
Epub Date: 1999/11/26
Date: Oct 29
Short Title: Carbohydrate release from picomole quantities of glycoprotein and characterisation of glycans by high-performance liquid chromatography and mass spectrometry
Alternate Journal: Journal of chromatography. B, Biomedical sciences and applications
ISSN: 1387-2273 (Print)
1387-2273
DOI: 10.1016/s0378-4347(99)00330-8
Accession Number: 10574203
Keywords: Amidohydrolases/metabolism
Aminoacridines
Chromatography, High Pressure Liquid/*methods
Disulfides/metabolism
Dithiothreitol/pharmacology
Glycoproteins/*metabolism
Humans
Immunoglobulin G/chemistry/*metabolism
Microchemistry
Oligosaccharides/analysis/isolation & purification
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Polysaccharides/chemistry/*isolation & purification/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/*methods
Trypsin/metabolism
Abstract: Samples of 5 to 20 microg of human IgG were subjected to dithiothreitol treatment to reduce disulphide bridges, followed by tryptic digestion. Glycans released from the tryptic peptide mixture by PNGase F digestion were then derivatised with 2-aminoacridone. Labelled oligosaccharides were separated by normal-phase high-performance liquid chromatography and individual components were collected for matrix-assisted laser desorption ionization time-of-flight and electrospray mass spectrometric analysis.
Notes: Charlwood, J
Birrell, H
Camilleri, P
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Chromatogr B Biomed Sci Appl. 1999 Oct 29;734(1):169-74. doi: 10.1016/s0378-4347(99)00330-8.
Author Address: SmithKline Beecham Pharmaceuticals, Harlow, Essex, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 678
Author: Charlwood, J., Birrell, H., Tolson, D. and Camilleri, P.
Year: 1998
Title: Two-dimensional chromatography in the analysis of complex glycans from transferrin
Journal: Anal Chem
Volume: 70
Issue: 13
Pages: 2530-5
Epub Date: 1998/07/17
Date: Jul 1
Short Title: Two-dimensional chromatography in the analysis of complex glycans from transferrin
Alternate Journal: Analytical chemistry
ISSN: 0003-2700 (Print)
0003-2700
DOI: 10.1021/ac9801059
Accession Number: 9666724
Keywords: Aminoacridines/chemistry
Chromatography, High Pressure Liquid
Fluorescent Dyes/chemistry
Fucose/chemistry
Humans
N-Acetylneuraminic Acid/chemistry
Polysaccharides/*analysis
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
Transferrin/*chemistry
Abstract: Oligosaccharides released from human transferrin have been derivatized with 2-aminoacridone (2-AMAC) prior to analysis by either reversed- or normal-phase high-performance liquid chromatography. These separation methods are complementary and allow distinction between isomeric mono- and disialylated oligosaccharides, which terminate with Sia alpha 2-6Gal or Sia alpha 2-3Gal at the nonreducing end. Collected fractions of 2-AMAC-derivatized glycans were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, before and after desialylation.
Notes: Charlwood, J
Birrell, H
Tolson, D
Camilleri, P
Journal Article
United States
Anal Chem. 1998 Jul 1;70(13):2530-5. doi: 10.1021/ac9801059.
Author Address: SmithKline Beecham Pharmaceuticals, New Frontiers Science Park, Harlow, Essex, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 790
Author: Charlwood, J., Dingwall, C., Matico, R., Hussain, I., Johanson, K., Moore, S., Powell, D. J., Skehel, J. M., Ratcliffe, S., Clarke, B., Trill, J., Sweitzer, S. and Camilleri, P.
Year: 2001
Title: Characterization of the glycosylation profiles of Alzheimer's beta -secretase protein Asp-2 expressed in a variety of cell lines
Journal: J Biol Chem
Volume: 276
Issue: 20
Pages: 16739-48
Epub Date: 2001/03/30
Date: May 18
Short Title: Characterization of the glycosylation profiles of Alzheimer's beta -secretase protein Asp-2 expressed in a variety of cell lines
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M009361200
Accession Number: 11278492
Keywords: Alzheimer Disease/*enzymology
Amyloid Precursor Protein Secretases
Animals
Aspartic Acid Endopeptidases/*chemistry/genetics/*metabolism
Brain/enzymology
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Endopeptidases
Glycoproteins/chemistry/genetics/*metabolism
Glycosylation
Humans
Molecular Sequence Data
Oligosaccharides/*chemistry
Polysaccharides/chemistry/*metabolism
Recombinant Proteins/chemistry/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Spodoptera
Transfection
Abstract: Amyloid 39-42 beta -peptides are the main components of amyloid plaques found in the brain of Alzheimer's disease patients. Amyloid 39-42 beta-peptide is formed from amyloid precursor protein by the sequential action of beta- and gamma-secretases. Asp-2 is a transmembrane aspartic protease expressed in the brain, shown to have beta-secretase activity. Mature Asp-2 has four N-glycosylation sites. In this report we have characterized the carbohydrate structures in this glycoprotein expressed in three different cell lines, namely Chinese hamster ovary, CV-1 origin of SV40, and baculovirus-infected SF9 cells. Biantennary and triantennary oligosaccharides of the "complex" type were released from glycoprotein expressed in the mammalian cells, whereas mannose-rich glycans were identified from glycoprotein synthesized in the baculovirus-infected cells. Site-directed mutagenesis of the asparagine residues at amino acid positions 153, 172, 223, and 354 demonstrate that the protease activity of Asp-2 is dependent on its glycosylation.
Notes: Charlwood, J
Dingwall, C
Matico, R
Hussain, I
Johanson, K
Moore, S
Powell, D J
Skehel, J M
Ratcliffe, S
Clarke, B
Trill, J
Sweitzer, S
Camilleri, P
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2001 May 18;276(20):16739-48. doi: 10.1074/jbc.M009361200. Epub 2001 Jan 30.
Author Address: Department of Analytical Sciences, SmithKline Beecham Pharmaceuticals, Harlow, Essex CM19 5AW, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2066
Author: Chattopadhyay, T., Maniyadath, B., Bagul, H. P., Chakraborty, A., Shukla, N., Budnar, S., Rajendran, A., Shukla, A., Kamat, S. S. and Kolthur-Seetharam, U.
Year: 2020
Title: Spatiotemporal gating of SIRT1 functions by O-GlcNAcylation is essential for liver metabolic switching and prevents hyperglycemia
Journal: Proc Natl Acad Sci U S A
Volume: 117
Issue: 12
Pages: 6890-6900
Epub Date: 2020/03/11
Date: Mar 24
Short Title: Spatiotemporal gating of SIRT1 functions by O-GlcNAcylation is essential for liver metabolic switching and prevents hyperglycemia
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1909943117
PMCID: PMC7104039
Accession Number: 32152092
Keywords: Acetylglucosamine/metabolism
Aging/physiology
Animals
Fasting
*Gluconeogenesis
Glycosylation
HEK293 Cells
*Homeostasis
Humans
Hyperglycemia/metabolism/pathology/*prevention & control
Insulin Resistance
Liver/immunology/*metabolism/pathology
Male
Mice
Mice, Inbred C57BL
Obesity/metabolism/pathology/prevention & control
Phosphorylation
*Protein Processing, Post-Translational
Sirtuin 1/chemistry/*metabolism
Spatio-Temporal Analysis
*Pgc1α
*fed–fast cycle
*insulin signaling
*ubiquitinylation
Abstract: Inefficient physiological transitions are known to cause metabolic disorders. Therefore, investigating mechanisms that constitute molecular switches in a central metabolic organ like the liver becomes crucial. Specifically, upstream mechanisms that control temporal engagement of transcription factors, which are essential to mediate physiological fed-fast-refed transitions are less understood. SIRT1, a NAD(+)-dependent deacetylase, is pivotal in regulating hepatic gene expression and has emerged as a key therapeutic target. Despite this, if/how nutrient inputs regulate SIRT1 interactions, stability, and therefore downstream functions are still unknown. Here, we establish nutrient-dependent O-GlcNAcylation of SIRT1, within its N-terminal domain, as a crucial determinant of hepatic functions. Our findings demonstrate that during a fasted-to-refed transition, glycosylation of SIRT1 modulates its interactions with various transcription factors and a nodal cytosolic kinase involved in insulin signaling. Moreover, sustained glycosylation in the fed state causes nuclear exclusion and cytosolic ubiquitin-mediated degradation of SIRT1. This mechanism exerts spatiotemporal control over SIRT1 functions by constituting a previously unknown molecular relay. Of note, loss of SIRT1 glycosylation discomposed these interactions resulting in aberrant gene expression, mitochondrial dysfunctions, and enhanced hepatic gluconeogenesis. Expression of nonglycosylatable SIRT1 in the liver abrogated metabolic flexibility, resulting in systemic insulin resistance, hyperglycemia, and hepatic inflammation, highlighting the physiological costs associated with its overactivation. Conversely, our study also reveals that hyperglycosylation of SIRT1 is associated with aging and high-fat-induced obesity. Thus, we establish that nutrient-dependent glycosylation of SIRT1 is essential to gate its functions and maintain physiological fitness.
Notes: 1091-6490
Chattopadhyay, Tandrika
Maniyadath, Babukrishna
Bagul, Hema P
Chakraborty, Arindam
Shukla, Namrata
Budnar, Srikanth
Rajendran, Abinaya
Shukla, Arushi
Kamat, Siddhesh S
Orcid: 0000-0001-6132-7574
Kolthur-Seetharam, Ullas
IA/I/15/2/502058/WTDBT_/DBT-Wellcome Trust India Alliance/India
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2020 Mar 24;117(12):6890-6900. doi: 10.1073/pnas.1909943117. Epub 2020 Mar 9.
Author Address: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, 400005 Maharashtra, India.
Department of Biology, Indian Institute of Science Education and Research, Pune, 411008 Maharashtra, India.
Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, 400005 Maharashtra, India; ullas@tifr.res.in.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1572
Author: Chen, C., Ke, J., Zhou, X. E., Yi, W., Brunzelle, J. S., Li, J., Yong, E. L., Xu, H. E. and Melcher, K.
Year: 2013
Title: Structural basis for molecular recognition of folic acid by folate receptors
Journal: Nature
Volume: 500
Issue: 7463
Pages: 486-9
Epub Date: 2013/07/16
Date: Aug 22
Short Title: Structural basis for molecular recognition of folic acid by folate receptors
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature12327
PMCID: PMC5797940
NIHMSID: NIHMS937267
Accession Number: 23851396
Keywords: Binding Sites/genetics
Crystallography, X-Ray
Folate Receptor 1/*chemistry/genetics/*metabolism
Folic Acid/chemistry/*metabolism
Humans
Ligands
Models, Molecular
Mutation
Protein Binding
Structure-Activity Relationship
Abstract: Folate receptors (FRα, FRβ and FRγ) are cysteine-rich cell-surface glycoproteins that bind folate with high affinity to mediate cellular uptake of folate. Although expressed at very low levels in most tissues, folate receptors, especially FRα, are expressed at high levels in numerous cancers to meet the folate demand of rapidly dividing cells under low folate conditions. The folate dependency of many tumours has been therapeutically and diagnostically exploited by administration of anti-FRα antibodies, high-affinity antifolates, folate-based imaging agents and folate-conjugated drugs and toxins. To understand how folate binds its receptors, we determined the crystal structure of human FRα in complex with folic acid at 2.8 Å resolution. FRα has a globular structure stabilized by eight disulphide bonds and contains a deep open folate-binding pocket comprised of residues that are conserved in all receptor subtypes. The folate pteroate moiety is buried inside the receptor, whereas its glutamate moiety is solvent-exposed and sticks out of the pocket entrance, allowing it to be conjugated to drugs without adversely affecting FRα binding. The extensive interactions between the receptor and ligand readily explain the high folate-binding affinity of folate receptors and provide a template for designing more specific drugs targeting the folate receptor system.
Notes: 1476-4687
Chen, Chen
Ke, Jiyuan
Zhou, X Edward
Yi, Wei
Brunzelle, Joseph S
Li, Jun
Yong, Eu-Leong
Xu, H Eric
Melcher, Karsten
R01 DK071662/DK/NIDDK NIH HHS/United States
R01 GM102545/GM/NIGMS NIH HHS/United States
R01 GM104212/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Nature. 2013 Aug 22;500(7463):486-9. doi: 10.1038/nature12327. Epub 2013 Jul 14.
Author Address: Program for Structural Biology and Drug Discovery, Van Andel Research Institute, 333 Bostwick Avenue North East, Grand Rapids, Michigan 49503, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2017
Author: Chen, C. H., Lin, K. D., Ke, L. Y., Liang, C. J., Kuo, W. C., Lee, M. Y., Lee, Y. L., Hsiao, P. J., Hsu, C. C. and Shin, S. J.
Year: 2019
Title: O-GlcNAcylation disrupts STRA6-retinol signals in kidneys of diabetes
Journal: Biochim Biophys Acta Gen Subj
Volume: 1863
Issue: 6
Pages: 1059-1069
Epub Date: 2019/03/25
Date: Jun
Short Title: O-GlcNAcylation disrupts STRA6-retinol signals in kidneys of diabetes
Alternate Journal: Biochimica et biophysica acta. General subjects
ISSN: 0304-4165
DOI: 10.1016/j.bbagen.2019.03.014
Accession Number: 30905621
Keywords: Acylation
Animals
Diabetes Mellitus, Experimental/*metabolism/pathology
Diabetic Nephropathies/*metabolism/pathology
Male
Membrane Proteins/*metabolism
Mice
*Signal Transduction
Vitamin A/*metabolism
*Diabetes
*Diabetic nephropathy
*O-GlcNAc
*Raldh
*Rbp4
*Retinol
*Stra6
Abstract: BACKGROUND: O-GlcNAcylation is an important mechanism of diabetic complication. Retinoid homeostasis regulates cell-physiological functions through STRA6-retinol signaling. Therefore, we investigated whether O-GlcNAcylation disrupted STRA6-retinol signals in diabetes. METHODS: Immunoprecipitation and proximity ligation assay were used to investigate O-GlcNAcylation of STRA6-retinol signals in kidneys of db/db and ob/ob mice. Western blot and immunohistochemistry were done for STRA6/CRBP1/LRAT/RALDH1/RARs pathway, GFAT, OGT, TGFβ(1) and collagen 1 level. HPLC and ELISA for retinol, retinal, and retinoic acid concentrations were performed in vivo and vitro. RBP4 binding with STRA6 was measured in vitro. To verify whether O-GlcNAcylation disrupted STRA6-retinol signals, treatment of TMG and OSMI-1, transfection of OGA and OGT, and OGT siRNA were performed in HK-2 cells. RESULTS: STRA6 and RALDH1 were highly O-GlcNAc-modified in glomeruli and tubules of db/db and ob/ob mice. RBP4, p-Try, p-JAK2, and p-STAT5 on STRA6 immunoprecipitate were reduced. Cellular retinol signals (CRBP1, LRAT, ADH, retinol, retinal, RA, RARα, RARγ and RXRα) remarkably decreased in kidneys of db/db, ob/ob mice and HG-cultured cells. TMG and OGT transfection induced O-GlcNAcylation of STRA6 and RALDH1, repressed RBP4-bound STRA6, and retinol signals in NG-cultured cells. OSMI-1, OGA transfection, and OGT silence reversed O-GlcNAc-modification of STRA6 and RALDH1, and rescued the decrease of retinol signals, and reversed the increase of TGFβ(1) and collagen 1 in HG-treated cells. CONCLUSIONS: O-GlcNAcylation significantly modified STRA6 and RALDH1, suppressed RBP4 binding activity, and disrupted retinol signals in the kidney of diabetes. GENERAL SIGNIFICANCE: This study first indicates that STRA6-retinol signals were directly disrupted by O-GlcNAcylation in diabetic kidney.
Notes: 1872-8006
Chen, Chao-Hung
Lin, Kun-Der
Ke, Liang-Yin
Liang, Chan-Jung
Kuo, Wen-Chen
Lee, Mei-Yueh
Lee, Yu-Li
Hsiao, Pi-Jung
Hsu, Chih-Cheng
Shin, Shyi-Jang
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta Gen Subj. 2019 Jun;1863(6):1059-1069. doi: 10.1016/j.bbagen.2019.03.014. Epub 2019 Mar 21.
Author Address: Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; Division of Endocrinology and Metabolism, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.
Division of Endocrinology and Metabolism, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan; School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
Lipid Science and Aging Research Center, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Medical Laboratory Science and Biotechnology, College of Health Sciences, Kaohsiung Medical University, Kaohsiung, Taiwan.
Lipid Science and Aging Research Center, Kaohsiung Medical University, Kaohsiung, Taiwan.
Institute of Population Health Sciences, National Health Research Institutes, Zhunan, Taiwan.
Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; Division of Endocrinology and Metabolism, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan; School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; Lipid Science and Aging Research Center, Kaohsiung Medical University, Kaohsiung, Taiwan; Institute of Medical Science and Technology, National Sun Yat-Sen University, Kaohsiung, Taiwan. Electronic address: sjshin@kmu.edu.tw.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1481
Author: Chen, C. I., Keusch, J. J., Klein, D., Hess, D., Hofsteenge, J. and Gut, H.
Year: 2012
Title: Structure of human POFUT2: insights into thrombospondin type 1 repeat fold and O-fucosylation
Journal: Embo j
Volume: 31
Issue: 14
Pages: 3183-97
Epub Date: 2012/05/17
Date: Jul 18
Short Title: Structure of human POFUT2: insights into thrombospondin type 1 repeat fold and O-fucosylation
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2012.143
PMCID: PMC3400009
Accession Number: 22588082
Keywords: Crystallography, X-Ray
Fucose/chemistry/genetics/metabolism
Fucosyltransferases/*chemistry/genetics/metabolism
Glycosylation
Humans
*Protein Folding
Protein Structure, Tertiary
Repetitive Sequences, Amino Acid
Structure-Activity Relationship
Abstract: Protein O-fucosylation is a post-translational modification found on serine/threonine residues of thrombospondin type 1 repeats (TSR). The fucose transfer is catalysed by the protein O-fucosyltransferase 2 (POFUT2) and >40 human proteins contain the TSR consensus sequence for POFUT2-dependent fucosylation. To better understand O-fucosylation on TSR, we carried out a structural and functional analysis of human POFUT2 and its TSR substrate. Crystal structures of POFUT2 reveal a variation of the classical GT-B fold and identify sugar donor and TSR acceptor binding sites. Structural findings are correlated with steady-state kinetic measurements of wild-type and mutant POFUT2 and TSR and give insight into the catalytic mechanism and substrate specificity. By using an artificial mini-TSR substrate, we show that specificity is not primarily encoded in the TSR protein sequence but rather in the unusual 3D structure of a small part of the TSR. Our findings uncover that recognition of distinct conserved 3D fold motifs can be used as a mechanism to achieve substrate specificity by enzymes modifying completely folded proteins of very wide sequence diversity and biological function.
Notes: 1460-2075
Chen, Chun-I
Keusch, Jeremy J
Klein, Dominique
Hess, Daniel
Hofsteenge, Jan
Gut, Heinz
Journal Article
EMBO J. 2012 Jul 18;31(14):3183-97. doi: 10.1038/emboj.2012.143.
Author Address: Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1657
Author: Chen, L., Dürr, K. L. and Gouaux, E.
Year: 2014
Title: X-ray structures of AMPA receptor-cone snail toxin complexes illuminate activation mechanism
Journal: Science
Volume: 345
Issue: 6200
Pages: 1021-6
Epub Date: 2014/08/12
Date: Aug 29
Short Title: X-ray structures of AMPA receptor-cone snail toxin complexes illuminate activation mechanism
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1258409
PMCID: PMC4263349
NIHMSID: NIHMS634310
Accession Number: 25103405
Keywords: Animals
Conotoxins/*chemistry
Conus Snail
Crystallography, X-Ray
*Ion Channel Gating
Ligands
Mutation
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Rats
Receptors, AMPA/*agonists/*chemistry/genetics
Abstract: AMPA-sensitive glutamate receptors are crucial to the structural and dynamic properties of the brain, to the development and function of the central nervous system, and to the treatment of neurological conditions from depression to cognitive impairment. However, the molecular principles underlying AMPA receptor activation have remained elusive. We determined multiple x-ray crystal structures of the GluA2 AMPA receptor in complex with a Conus striatus cone snail toxin, a positive allosteric modulator, and orthosteric agonists, at 3.8 to 4.1 angstrom resolution. We show how the toxin acts like a straightjacket on the ligand-binding domain (LBD) "gating ring," restraining the domains via both intra- and interdimer cross-links such that agonist-induced closure of the LBD "clamshells" is transduced into an irislike expansion of the gating ring. By structural analysis of activation-enhancing mutants, we show how the expansion of the LBD gating ring results in pulling forces on the M3 helices that, in turn, are coupled to ion channel gating.
Notes: 1095-9203
Chen, Lei
Dürr, Katharina L
Gouaux, Eric
R01 NS038631/NS/NINDS NIH HHS/United States
F32 MH100331/MH/NIMH NIH HHS/United States
F32MH100331/MH/NIMH NIH HHS/United States
Howard Hughes Medical Institute/United States
R37 NS038631/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2014 Aug 29;345(6200):1021-6. doi: 10.1126/science.1258409. Epub 2014 Aug 7.
Author Address: Vollum Institute, Oregon Health and Science University, 3181 SW Sam Jackson Park Road, Portland, OR 97239, USA.
Vollum Institute, Oregon Health and Science University, 3181 SW Sam Jackson Park Road, Portland, OR 97239, USA. Howard Hughes Medical Institute, Oregon Health and Science University, 3181 SW Sam Jackson Park Road, Portland, OR 97239, USA. gouauxe@ohsu.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1144
Author: Chen, L. M., Kelly, M. L., Rojas, J. D., Parker, M. D., Gill, H. S., Davis, B. A. and Boron, W. F.
Year: 2008
Title: Use of a new polyclonal antibody to study the distribution and glycosylation of the sodium-coupled bicarbonate transporter NCBE in rodent brain
Journal: Neuroscience
Volume: 151
Issue: 2
Pages: 374-85
Epub Date: 2007/12/07
Date: Jan 24
Short Title: Use of a new polyclonal antibody to study the distribution and glycosylation of the sodium-coupled bicarbonate transporter NCBE in rodent brain
Alternate Journal: Neuroscience
ISSN: 0306-4522 (Print)
0306-4522
DOI: 10.1016/j.neuroscience.2007.10.015
PMCID: PMC2905792
NIHMSID: NIHMS39187
Accession Number: 18061361
Keywords: Animals
Antibodies/*chemistry
Blotting, Western
Brain Chemistry/genetics/*physiology
Cells, Cultured
Chloride-Bicarbonate Antiporters/chemistry/*metabolism
Genetic Vectors
Glycosylation
Green Fluorescent Proteins/genetics
Humans
Immunohistochemistry
Mice
Molecular Sequence Data
Molecular Weight
Mutagenesis, Site-Directed
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/chemistry
Rats
Recombinant Fusion Proteins/pharmacology
Reproducibility of Results
Sodium-Bicarbonate Symporters/chemistry/*metabolism
Species Specificity
Tissue Distribution
Xenopus laevis
Abstract: NCBE (SLC4A10) is a member of the SLC4 family of bicarbonate transporters, several of which play important roles in intracellular-pH regulation and transepithelial HCO(3)(-) transport. Here we characterize a new antibody that was generated in rabbit against a fusion protein consisting of maltose-binding protein and the first 135 amino acids (aa) of the N-terminus of human NCBE. Western blotting--both of purified peptides representing the initial approximately 120 aa of the transporters and of full-length transporters expressed in Xenopus oocytes--demonstrated that the antibody is specific for NCBE versus the two most closely related proteins, NDCBE (SLC4A8) and NBCn1 (SLC4A7). Western blotting of tissue in four regions of adult mouse brain indicates that NCBE is expressed most abundantly in cerebral cortex (CX), cerebellum (CB) and hippocampus (HC), and less so in subcortex (SCX). NCBE protein was present in CX, CB, and HC microdissected to avoid choroid plexus. Immunocytochemistry shows that NCBE is present at the basolateral membrane of embryonic day 18 (E18) fetal and adult choroid plexus. NCBE protein is present by Western blot and immunocytochemistry in cultured and freshly dissociated HC neurons but not astrocytes. By Western blot, nearly all NCBE in mouse and rat brain is highly N-glycosylated (approximately 150 kDa). PNGase F reduces the molecular weight (MW) of natural NCBE in mouse brain or human NCBE expressed in oocytes to approximately the predicted MW of the unglycosylated protein. In oocytes, mutating any one of the three consensus N-glycosylation sites reduces glycosylation of the other two, and the triple mutant exhibits negligible functional expression.
Notes: 1873-7544
Chen, L-M
Kelly, M L
Rojas, J D
Parker, M D
Gill, H S
Davis, B A
Boron, W F
R01 NS018400/NS/NINDS NIH HHS/United States
F32 DK075258/DK/NIDDK NIH HHS/United States
P01 HD032573/HD/NICHD NIH HHS/United States
R37 DK030344-28/DK/NIDDK NIH HHS/United States
R01 NS018400-23/NS/NINDS NIH HHS/United States
NS18400/NS/NINDS NIH HHS/United States
R37 DK030344/DK/NIDDK NIH HHS/United States
K01 DK082646/DK/NIDDK NIH HHS/United States
P01 HD032573-130002/HD/NICHD NIH HHS/United States
HD32573/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Neuroscience. 2008 Jan 24;151(2):374-85. doi: 10.1016/j.neuroscience.2007.10.015. Epub 2007 Oct 25.
Author Address: Department of Cellular and Molecular Physiology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA. liming.chen@yale.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1938
Author: Chen, P., Tao, L., Wang, T., Zhang, J., He, A., Lam, K. H., Liu, Z., He, X., Perry, K., Dong, M. and Jin, R.
Year: 2018
Title: Structural basis for recognition of frizzled proteins by Clostridium difficile toxin B
Journal: Science
Volume: 360
Issue: 6389
Pages: 664-669
Epub Date: 2018/05/12
Date: May 11
Short Title: Structural basis for recognition of frizzled proteins by Clostridium difficile toxin B
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.aar1999
PMCID: PMC6231499
NIHMSID: NIHMS995333
Accession Number: 29748286
Keywords: Bacterial Proteins/*chemistry/metabolism
Bacterial Toxins/*chemistry/metabolism
Binding Sites
Clostridioides difficile/*pathogenicity
Clostridium Infections/*metabolism
Crystallography, X-Ray
Fatty Acids/metabolism
Frizzled Receptors/*chemistry
Humans
Protein Domains
Virulence Factors/*chemistry/*metabolism
Wnt Signaling Pathway
Abstract: Clostridium difficile infection is the most common cause of antibiotic-associated diarrhea in developed countries. The major virulence factor, C. difficile toxin B (TcdB), targets colonic epithelia by binding to the frizzled (FZD) family of Wnt receptors, but how TcdB recognizes FZDs is unclear. Here, we present the crystal structure of a TcdB fragment in complex with the cysteine-rich domain of human FZD2 at 2.5-angstrom resolution, which reveals an endogenous FZD-bound fatty acid acting as a co-receptor for TcdB binding. This lipid occupies the binding site for Wnt-adducted palmitoleic acid in FZDs. TcdB binding locks the lipid in place, preventing Wnt from engaging FZDs and signaling. Our findings establish a central role of fatty acids in FZD-mediated TcdB pathogenesis and suggest strategies to modulate Wnt signaling.
Notes: 1095-9203
Chen, Peng
Orcid: 0000-0002-6356-7683
Tao, Liang
Orcid: 0000-0003-3441-698x
Wang, Tianyu
Zhang, Jie
He, Aina
Lam, Kwok-Ho
Orcid: 0000-0002-7173-6600
Liu, Zheng
He, Xi
Perry, Kay
Orcid: 0000-0002-4046-1704
Dong, Min
Orcid: 0000-0002-1744-7293
Jin, Rongsheng
Orcid: 0000-0003-0348-7363
S10 RR029205/RR/NCRR NIH HHS/United States
P41 GM103403/GM/NIGMS NIH HHS/United States
R01 NS080833/NS/NINDS NIH HHS/United States
R01 GM057603/GM/NIGMS NIH HHS/United States
R01 AI132387/AI/NIAID NIH HHS/United States
P30 HD018655/HD/NICHD NIH HHS/United States
P30 DK034854/DK/NIDDK NIH HHS/United States
R21 AI123920/AI/NIAID NIH HHS/United States
R01 AI091823/AI/NIAID NIH HHS/United States
R01 AI125704/AI/NIAID NIH HHS/United States
R01 GM126120/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Science. 2018 May 11;360(6389):664-669. doi: 10.1126/science.aar1999.
Author Address: Department of Physiology and Biophysics, University of California, Irvine, Irvine, CA, USA.
Department of Urology, Boston Children's Hospital, Department of Microbiology and Immunobiology and Department of Surgery, Harvard Medical School, Boston, MA, USA.
Department of Oncology, Affiliated Sixth People's Hospital, Shanghai Jiaotong University, No. 600, Yishan Road, 200233 Shanghai, PRC.
F. M. Kirby Neurobiology Center, Boston Children's Hospital, Department of Neurology, Harvard Medical School, Boston, MA, USA.
NE-CAT and Department of Chemistry and Chemical Biology, Cornell University, Argonne National Laboratory, Argonne, IL, USA.
Department of Urology, Boston Children's Hospital, Department of Microbiology and Immunobiology and Department of Surgery, Harvard Medical School, Boston, MA, USA. r.jin@uci.edu min.dong@childrens.harvard.edu.
Department of Physiology and Biophysics, University of California, Irvine, Irvine, CA, USA. r.jin@uci.edu min.dong@childrens.harvard.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1563
Author: Chen, P. H., Chen, X., Lin, Z., Fang, D. and He, X.
Year: 2013
Title: The structural basis of R-spondin recognition by LGR5 and RNF43
Journal: Genes Dev
Volume: 27
Issue: 12
Pages: 1345-50
Epub Date: 2013/06/13
Date: Jun 15
Short Title: The structural basis of R-spondin recognition by LGR5 and RNF43
Alternate Journal: Genes & development
ISSN: 0890-9369 (Print)
0890-9369
DOI: 10.1101/gad.219915.113
PMCID: PMC3701190
Accession Number: 23756651
Keywords: DNA-Binding Proteins/*chemistry/genetics/metabolism
Humans
Models, Molecular
Mutation
Oncogene Proteins/*chemistry/genetics/metabolism
Protein Structure, Tertiary
Receptors, G-Protein-Coupled/*chemistry/genetics
Signal Transduction
Thrombospondins/metabolism
Ubiquitin-Protein Ligases/metabolism
Wnt Signaling Pathway
E3 ubiquitin ligase
Lgr5
R-spondin
Rnf43
Wnt signaling
furin-like repeat
Abstract: R-spondins (RSPOs) enhance Wnt signaling, affect stem cell behavior, bind to leucine-rich repeat-containing G-protein-coupled receptors 4-6, (LGR4-6) and the transmembrane E3 ubiquitin ligases RING finger 43/zinc and RING finger 3 (RNF43/ZNRF3). The structure of RSPO1 bound to both LGR5 and RNF43 ectodomains confirms their physical linkage. RSPO1 is sandwiched by LGR5 and RNF43, with its rod module of the cysteine-rich domain (CRD) contacting LGR5 and a hairpin inserted into RNF43. LGR5 does not contact RNF43 but increases the affinity of RSPO1 to RNF43, supporting LGR5 as an engagement receptor and RNF43 as an effector receptor. Disease mutations map to the RSPO1-RNF43 interface, which promises therapeutic targeting.
Notes: 1549-5477
Chen, Po-Han
Chen, Xiaoyan
Lin, Zhenghong
Fang, Deyu
He, Xiaolin
P30 CA060553/CA/NCI NIH HHS/United States
R01 GM098259/GM/NIGMS NIH HHS/United States
T32 GM008382/GM/NIGMS NIH HHS/United States
GM098259/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Genes Dev. 2013 Jun 15;27(12):1345-50. doi: 10.1101/gad.219915.113. Epub 2013 Jun 11.
Author Address: Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2055
Author: Chen, P. H., Hu, J., Wu, J., Huynh, D. T., Smith, T. J., Pan, S., Bisnett, B. J., Smith, A. B., Lu, A., Condon, B. M., Chi, J. T. and Boyce, M.
Year: 2020
Title: Gigaxonin glycosylation regulates intermediate filament turnover and may impact giant axonal neuropathy etiology or treatment
Journal: JCI Insight
Volume: 5
Issue: 1
Epub Date: 2020/01/17
Date: Jan 16
Short Title: Gigaxonin glycosylation regulates intermediate filament turnover and may impact giant axonal neuropathy etiology or treatment
Alternate Journal: JCI insight
ISSN: 2379-3708
DOI: 10.1172/jci.insight.127751
PMCID: PMC7030874
Accession Number: 31944090
Keywords: Acetylglucosamine/*metabolism
Antigens, Neoplasm/metabolism
Binding Sites
Cell Line
Cytoskeletal Proteins/genetics/*metabolism
Epigenesis, Genetic
Genetic Therapy
Giant Axonal Neuropathy/etiology/genetics/*metabolism/therapy
Glycosylation
Histone Acetyltransferases/metabolism
Humans
Hyaluronoglucosaminidase/metabolism
Intercellular Signaling Peptides and Proteins/metabolism
Intermediate Filament Proteins/*metabolism
Models, Biological
Nutritional Status
Proteasome Endopeptidase Complex/metabolism
Protein Binding
Proteostasis
Serine/metabolism
Threonine/metabolism
Ubiquitin/metabolism
Ubiquitin-Protein Ligases/metabolism
Ubiquitination
Abstract: Gigaxonin (also known as KLHL16) is an E3 ligase adaptor protein that promotes the ubiquitination and degradation of intermediate filament (IF) proteins. Mutations in human gigaxonin cause the fatal neurodegenerative disease giant axonal neuropathy (GAN), in which IF proteins accumulate and aggregate in axons throughout the nervous system, impairing neuronal function and viability. Despite this pathophysiological significance, the upstream regulation and downstream effects of normal and aberrant gigaxonin function remain incompletely understood. Here, we report that gigaxonin is modified by <italic>O</italic>-linked β-<italic>N</italic>-acetylglucosamine (O-GlcNAc), a prevalent form of intracellular glycosylation, in a nutrient- and growth factor–dependent manner. MS analyses of human gigaxonin revealed 9 candidate sites of O-GlcNAcylation, 2 of which — serine 272 and threonine 277 — are required for its ability to mediate IF turnover in gigaxonin-deficient human cell models that we created. Taken together, the results suggest that nutrient-responsive gigaxonin O-GlcNAcylation forms a regulatory link between metabolism and IF proteostasis. Our work may have significant implications for understanding the nongenetic modifiers of GAN phenotypes and for the optimization of gene therapy for this disease.
Notes: 2379-3708
Chen, Po-Han
Hu, Jimin
Wu, Jianli
Huynh, Duc T
Smith, Timothy J
Pan, Samuel
Bisnett, Brittany J
Smith, Alexander B
Lu, Annie
Condon, Brett M
Chi, Jen-Tsan
Boyce, Michael
R01 GM118847/GM/NIGMS NIH HHS/United States
R01 NS111588/NS/NINDS NIH HHS/United States
R01 GM124062/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
JCI Insight. 2019 Nov 26;5(1):e127751. doi: 10.1172/jci.insight.127751. Print 2020 Jan 16.
Author Address: Department of Biochemistry.
Department of Molecular Genetics and Microbiology, and.
Center for Genomic and Computational Biology, Duke University School of Medicine, Durham, North Carolina, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1867
Author: Chen, P. H., Smith, T. J., Wu, J., Siesser, P. F., Bisnett, B. J., Khan, F., Hogue, M., Soderblom, E., Tang, F., Marks, J. R., Major, M. B., Swarts, B. M., Boyce, M. and Chi, J. T.
Year: 2017
Title: Glycosylation of KEAP1 links nutrient sensing to redox stress signaling
Journal: Embo j
Volume: 36
Issue: 15
Pages: 2233-2250
Epub Date: 2017/07/01
Date: Aug 1
Short Title: Glycosylation of KEAP1 links nutrient sensing to redox stress signaling
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.15252/embj.201696113
PMCID: PMC5538768
Accession Number: 28663241
Keywords: Cell Line
Food
Gene Expression Profiling
*Gene Expression Regulation
*Glycosylation
Humans
Kelch-Like ECH-Associated Protein 1/*metabolism
N-Acetylglucosaminyltransferases/*metabolism
NF-E2-Related Factor 2/*metabolism
Oxidation-Reduction
*Signal Transduction
*Stress, Physiological
* Klhl
* Ogt
*Keap1
*Nrf2
*O‐GlcNAcylation
Abstract: O-GlcNAcylation is an essential, nutrient-sensitive post-translational modification, but its biochemical and phenotypic effects remain incompletely understood. To address this question, we investigated the global transcriptional response to perturbations in O-GlcNAcylation. Unexpectedly, many transcriptional effects of O-GlcNAc transferase (OGT) inhibition were due to the activation of NRF2, the master regulator of redox stress tolerance. Moreover, we found that a signature of low OGT activity strongly correlates with NRF2 activation in multiple tumor expression datasets. Guided by this information, we identified KEAP1 (also known as KLHL19), the primary negative regulator of NRF2, as a direct substrate of OGT We show that O-GlcNAcylation of KEAP1 at serine 104 is required for the efficient ubiquitination and degradation of NRF2. Interestingly, O-GlcNAc levels and NRF2 activation co-vary in response to glucose fluctuations, indicating that KEAP1 O-GlcNAcylation links nutrient sensing to downstream stress resistance. Our results reveal a novel regulatory connection between nutrient-sensitive glycosylation and NRF2 signaling and provide a blueprint for future approaches to discover functionally important O-GlcNAcylation events on other KLHL family proteins in various experimental and disease contexts.
Notes: 1460-2075
Chen, Po-Han
Smith, Timothy J
Wu, Jianli
Siesser, Priscila F
Bisnett, Brittany J
Khan, Farhan
Hogue, Maxwell
Soderblom, Erik
Tang, Flora
Marks, Jeffrey R
Major, Michael B
Swarts, Benjamin M
Boyce, Michael
Orcid: 0000-0002-2729-4876
Chi, Jen-Tsan
Orcid: 0000-0003-3433-903x
U54 CA156733/CA/NCI NIH HHS/United States
R01 CA125618/CA/NCI NIH HHS/United States
P30 DK096493/DK/NIDDK NIH HHS/United States
R01 GM118847/GM/NIGMS NIH HHS/United States
UL1 TR001117/TR/NCATS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
EMBO J. 2017 Aug 1;36(15):2233-2250. doi: 10.15252/embj.201696113. Epub 2017 Jun 29.
Author Address: Department of Molecular Genetics and Microbiology, Duke University School of Medicine, Durham, NC, USA.
Department of Biochemistry, Duke University School of Medicine, Durham, NC, USA.
Department of Cell Biology and Physiology, Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
Department of Chemistry and Biochemistry, Central Michigan University, Mount Pleasant, MI, USA.
Duke Proteomics and Metabolomics Core Facility, Center for Genomic and Computational Biology, Duke University, Durham, NC, USA.
Division of Surgical Sciences, Department of Surgery, Duke University, Durham, NC, USA.
Department of Biochemistry, Duke University School of Medicine, Durham, NC, USA michael.boyce@duke.edu jentsan.chi@duke.edu.
Department of Molecular Genetics and Microbiology, Duke University School of Medicine, Durham, NC, USA michael.boyce@duke.edu jentsan.chi@duke.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1886
Author: Chen, Q., She, J., Zeng, W., Guo, J., Xu, H., Bai, X. C. and Jiang, Y.
Year: 2017
Title: Structure of mammalian endolysosomal TRPML1 channel in nanodiscs
Journal: Nature
Volume: 550
Issue: 7676
Pages: 415-418
Epub Date: 2017/10/12
Date: Oct 19
Short Title: Structure of mammalian endolysosomal TRPML1 channel in nanodiscs
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature24035
PMCID: PMC5901962
NIHMSID: NIHMS902009
Accession Number: 29019981
Keywords: Animals
Binding Sites
Calcium
*Cryoelectron Microscopy
Endosomes/*chemistry
Hydrogen-Ion Concentration
Ion Transport
Ligands
Lysosomes/*chemistry
Mice
Models, Molecular
Mutation
Nanostructures/*chemistry
Phosphatidylinositol Phosphates/metabolism
Protein Conformation
Static Electricity
Transient Receptor Potential Channels/*chemistry/genetics/*ultrastructure
Abstract: Transient receptor potential mucolipin 1 (TRPML1) is a cation channel located within endosomal and lysosomal membranes. Ubiquitously expressed in mammalian cells, its loss-of-function mutations are the direct cause of type IV mucolipidosis, an autosomal recessive lysosomal storage disease. Here we present the single-particle electron cryo-microscopy structure of the mouse TRPML1 channel embedded in nanodiscs. Combined with mutagenesis analysis, the TRPML1 structure reveals that phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)P(2)) binds to the N terminus of the channel-distal from the pore-and the helix-turn-helix extension between segments S2 and S3 probably couples ligand binding to pore opening. The tightly packed selectivity filter contains multiple ion-binding sites, and the conserved acidic residues form the luminal Ca(2+)-blocking site that confers luminal pH and Ca(2+) modulation on channel conductance. A luminal linker domain forms a fenestrated canopy atop the channel, providing several luminal ion passages to the pore and creating a negative electrostatic trap, with a preference for divalent cations, at the luminal entrance. The structure also reveals two equally distributed S4-S5 linker conformations in the closed channel, suggesting an S4-S5 linker-mediated PtdInsP(2) gating mechanism among TRPML channels.
Notes: 1476-4687
Chen, Qingfeng
She, Ji
Zeng, Weizhong
Guo, Jiangtao
Xu, Haoxing
Bai, Xiao-Chen
Jiang, Youxing
AR060837/NH/NIH HHS/United States
R01 AR060837/AR/NIAMS NIH HHS/United States
R01 NS062792/NS/NINDS NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 GM079179/GM/NIGMS NIH HHS/United States
GM079179/NH/NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2017 Oct 19;550(7676):415-418. doi: 10.1038/nature24035. Epub 2017 Oct 11.
Author Address: Department of Physiology, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9040, USA.
Department of Biophysics, University of Texas Southwestern Medical Center, Dallas, Texas 75390-8816, USA.
Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9040, USA.
Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109-1048, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1799
Author: Chen, Q. and Yu, X.
Year: 2016
Title: OGT restrains the expansion of DNA damage signaling
Journal: Nucleic Acids Res
Volume: 44
Issue: 19
Pages: 9266-9278
Epub Date: 2016/11/02
Date: Nov 2
Short Title: OGT restrains the expansion of DNA damage signaling
Alternate Journal: Nucleic acids research
ISSN: 0305-1048 (Print)
0305-1048
DOI: 10.1093/nar/gkw663
PMCID: PMC5100584
Accession Number: 27458206
Keywords: Adaptor Proteins, Signal Transducing
Cell Cycle Checkpoints/genetics
Cell Cycle Proteins
Cell Survival/genetics
DNA Breaks, Double-Stranded
*DNA Damage
Glycosylation
HeLa Cells
Histones/metabolism
Humans
N-Acetylglucosaminyltransferases/*metabolism
Nuclear Proteins/metabolism
Phosphorylation
*Signal Transduction
Trans-Activators/metabolism
Abstract: O-linked N-acetylglucosamine linkage (O-GlcNAcylation) to serine or threonine residues regulates numerous biological processes; however, its role in DNA damage response remains elusive. Here, we found that O-GlcNAcylation is induced by DNA damage response. O-GlcNAc transferase (OGT), the solo enzyme for O-GlcNAcylation, relocates to the sites of DNA damage and induces the O-GlcNAcylation of histone H2AX and mediator of DNA damage checkpoint 1 (MDC1). The O-GlcNAcylation negatively regulates DNA double-strand break-induced phosphorylation of H2AX and MDC1 by restraining the expansion of these phosphorylation events from the sites of DNA damage. Therefore, our study reveals the molecular mechanism and biological function of OGT-dependent O-GlcNAcylation in response to DNA damage.
Notes: 1362-4962
Chen, Qiang
Yu, Xiaochun
R01 CA130899/CA/NCI NIH HHS/United States
R01 CA132755/CA/NCI NIH HHS/United States
R01 CA187209/CA/NCI NIH HHS/United States
R01 GM108647/GM/NIGMS NIH HHS/United States
Journal Article
Nucleic Acids Res. 2016 Nov 2;44(19):9266-9278. doi: 10.1093/nar/gkw663. Epub 2016 Jul 25.
Author Address: Department of Cancer Genetics and Epigenetics, Beckman Research Institute, City of Hope, 1500 E. Duarte Road, Duarte, CA 91010, USA.
Department of Cancer Genetics and Epigenetics, Beckman Research Institute, City of Hope, 1500 E. Duarte Road, Duarte, CA 91010, USA xyu@coh.org.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1221
Author: Chen, R., Jiang, X., Sun, D., Han, G., Wang, F., Ye, M., Wang, L. and Zou, H.
Year: 2009
Title: Glycoproteomics analysis of human liver tissue by combination of multiple enzyme digestion and hydrazide chemistry
Journal: J Proteome Res
Volume: 8
Issue: 2
Pages: 651-61
Epub Date: 2009/01/23
Date: Feb
Short Title: Glycoproteomics analysis of human liver tissue by combination of multiple enzyme digestion and hydrazide chemistry
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr8008012
Accession Number: 19159218
Keywords: Amino Acid Sequence
Animals
Glycoproteins/*chemistry/genetics/*metabolism
Glycosylation
Humans
*Liver/chemistry/metabolism
Molecular Sequence Data
Pepsin A/metabolism
Peptide Hydrolases/*metabolism
Proteomics/*methods
Solid Phase Extraction/*methods
Substrate Specificity
Thermolysin/metabolism
Trypsin/metabolism
Abstract: The study of protein glycosylation has lagged far behind the progress of current proteomics because of the enormous complexity, wide dynamic range distribution and low stoichiometric modification of glycoprotein. Solid phase extraction of tryptic N-glycopeptides by hydrazide chemistry is becoming a popular protocol for the analysis of N-glycoproteome. However, in silico digestion of proteins in human proteome database by trypsin indicates that a significant percentage of tryptic N-glycopeptides is not in the preferred detection mass range of shotgun proteomics approach, that is, from 800 to 3500 Da. And the quite big size of glycan groups may block trypsin to access the K, R residues near N-glycosites for digestion, which will result in generation of big glycopeptides. Thus many N-glycosites could not be localized if only trypsin was used to digest proteins. Herein, we describe a comprehensive way to analyze the N-glycoproteome of human liver tissue by combination of hydrazide chemistry method and multiple enzyme digestion. The lysate of human liver tissue was digested with three proteases, that is, trypsin, pepsin and thermolysin, with different specificities, separately. Use of trypsin alone resulted in identification of 622 N-glycosites, while using pepsin and thermolysin resulted in identification of 317 additional N-glycosites. Among the 317 additional N-glycosites, 98 (30.9%) could not be identified by trypsin in theory because the corresponding in silico tryptic peptides are either too small or too big to detect in mass spectrometer. This study clearly demonstrated that the coverage of N-glycosites could be significantly increased due to the adoption of multiple enzyme digestion. A total number of 939 N-glycosites were identified confidently, covering 523 noredundant glycoproteins from human liver tissue, which leads to the establishment of the largest data set of glycoproteome from human liver up to now.
Notes: Chen, Rui
Jiang, Xinning
Sun, Deguang
Han, Guanghui
Wang, Fangjun
Ye, Mingliang
Wang, Liming
Zou, Hanfa
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2009 Feb;8(2):651-61. doi: 10.1021/pr8008012.
Author Address: Dalian Institute of Chemical Physics, The Chinese Academy of Sciences, Dalian 116023, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1636
Author: Chen, R., Seebun, D., Ye, M., Zou, H. and Figeys, D.
Year: 2014
Title: Site-specific characterization of cell membrane N-glycosylation with integrated hydrophilic interaction chromatography solid phase extraction and LC-MS/MS
Journal: J Proteomics
Volume: 103
Pages: 194-203
Epub Date: 2014/04/12
Date: May 30
Short Title: Site-specific characterization of cell membrane N-glycosylation with integrated hydrophilic interaction chromatography solid phase extraction and LC-MS/MS
Alternate Journal: Journal of proteomics
ISSN: 1874-3919
DOI: 10.1016/j.jprot.2014.03.040
Accession Number: 24721674
Keywords: Chromatography, Liquid/methods
Glycopeptides/*isolation & purification
Glycosylation
HEK293 Cells
Humans
Hydrophobic and Hydrophilic Interactions
Membrane Proteins/*isolation & purification
Polysaccharides/chemistry
Protein Processing, Post-Translational
Solid Phase Extraction/methods
Tandem Mass Spectrometry/methods
Glycan composition
Hek 293t
Mass spectrometry
N-glycoproteome
Abstract: Glycosylation of membrane proteins plays an important role in cellular behaviors such as cell-cell interaction, immunologic recognition and cell signaling. However, the effective extraction of membrane proteins, the selective isolation of glycopeptides and the mass spectrometric characterization of glycosylation are challenging with current analytical techniques. In this study, a systematic approach was developed which combined: an integrated hydrophilic interaction chromatography solid phase interaction (HILIC SPE) for simultaneous detergent removal and glycopeptide enrichment, and mass spectrometric identification of both protein N-glycosylation sites and site-specific glycan composition. The HILIC SPE conditions were optimized to enable the use of a high concentration of strong detergents, such as SDS and Triton X-100 and to dissolve highly hydrophobic membrane proteins, thus increasing the yield of membrane protein extraction. We illustrated the performance of this approach for the study of membrane protein glycosylation from human embryonic kidney cell lines (HEK 293T). 200μg total protein digest was processed using this approach, leading to the identification of 811 N-glycosylation sites from 567 proteins within two experimental replicates. Furthermore, 177 glycopeptides representing 82 N-glycosites with both glycan composition and peptide sequence were identified by high energy collision dissociation. BIOLOGICAL SIGNIFICANCE: A method for systematic characterizing of cell membrane glycosylation has been developed in this manuscript. It is comprised of an integrated hydrophilic interaction chromatography solid phase extraction for the simultaneous detergent removal and intact glycopeptide enrichment. This HILIC SPE significantly increased the efficiency and sensitivity for glycosylation analysis and was combined with high energy collision dissociation to characterize site-specific N-glycosylation from HEK293 cell membrane. Totally 811 N-glycosylation sites from 567 proteins were identified and 177 intact glycopeptides with both glycan composition and peptides sequence were characterized, which provided a solution for site-specific N-glycosylation characterization of membrane.
Notes: 1876-7737
Chen, Rui
Seebun, Deeptee
Ye, Mingliang
Zou, Hanfa
Figeys, Daniel
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Proteomics. 2014 May 30;103:194-203. doi: 10.1016/j.jprot.2014.03.040. Epub 2014 Apr 8.
Author Address: Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa K1H 8M5, Canada; Key Lab of Separation Science for Analytical Chemistry, National Chromatography R&A Center, Dalian Institute of Chemical Physics, The Chinese Academy of Science, Dalian 116023, China.
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa K1H 8M5, Canada.
Key Lab of Separation Science for Analytical Chemistry, National Chromatography R&A Center, Dalian Institute of Chemical Physics, The Chinese Academy of Science, Dalian 116023, China.
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa K1H 8M5, Canada; Department of Chemistry, Faculty of Science, University of Ottawa, Ottawa K1N 6N5, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1210
Author: Chen, X., Liu, H., Focia, P. J., Shim, A. H. and He, X.
Year: 2008
Title: Structure of macrophage colony stimulating factor bound to FMS: diverse signaling assemblies of class III receptor tyrosine kinases
Journal: Proc Natl Acad Sci U S A
Volume: 105
Issue: 47
Pages: 18267-72
Epub Date: 2008/11/20
Date: Nov 25
Short Title: Structure of macrophage colony stimulating factor bound to FMS: diverse signaling assemblies of class III receptor tyrosine kinases
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0807762105
PMCID: PMC2587541
Accession Number: 19017797
Keywords: Calorimetry
Crystallography
Humans
Macrophage Colony-Stimulating Factor/*chemistry/metabolism
Models, Molecular
Protein Conformation
Receptor, Macrophage Colony-Stimulating Factor/*metabolism
*Signal Transduction
Abstract: Macrophage colony stimulating factor (M-CSF), through binding to its receptor FMS, a class III receptor tyrosine kinase (RTK), regulates the development and function of mononuclear phagocytes, and plays important roles in innate immunity, cancer and inflammation. We report a 2.4 A crystal structure of M-CSF bound to the first 3 domains (D1-D3) of FMS. The ligand binding mode of FMS is surprisingly different from KIT, another class III RTK, in which the major ligand-binding domain of FMS, D2, uses the CD and EF loops, but not the beta-sheet on the opposite side of the Ig domain as in KIT, to bind ligand. Calorimetric data indicate that M-CSF cannot dimerize FMS without receptor-receptor interactions mediated by FMS domains D4 and D5. Consistently, the structure contains only 1 FMS-D1-D3 molecule bound to a M-CSF dimer, due to a weak, hydrophilic M-CSF:FMS interface, and probably a conformational change of the M-CSF dimer in which binding to the second site is rendered unfavorable by FMS binding at the first site. The partial, intermediate complex suggests that FMS may be activated in two steps, with the initial engagement step distinct from the subsequent dimerization/activation step. Hence, the formation of signaling class III RTK complexes can be diverse, engaging various modes of ligand recognition and various mechanistic steps for dimerizing and activating receptors.
Notes: 1091-6490
Chen, Xiaoyan
Liu, Heli
Focia, Pamela J
Shim, Ann Hye-Ryong
He, Xiaolin
R01 GM078055/GM/NIGMS NIH HHS/United States
1R01GM078055/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2008 Nov 25;105(47):18267-72. doi: 10.1073/pnas.0807762105. Epub 2008 Nov 18.
Author Address: Department of Molecular Pharmacology and Biological Chemistry, Feinberg School of Medicine, Searle 8-417, Northwestern University, 303 East Chicago Avenue, Chicago, IL 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1148
Author: Chen, X., Liu, H., Shim, A. H., Focia, P. J. and He, X.
Year: 2008
Title: Structural basis for synaptic adhesion mediated by neuroligin-neurexin interactions
Journal: Nat Struct Mol Biol
Volume: 15
Issue: 1
Pages: 50-6
Epub Date: 2007/12/18
Date: Jan
Short Title: Structural basis for synaptic adhesion mediated by neuroligin-neurexin interactions
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb1350
PMCID: PMC2922956
NIHMSID: NIHMS226802
Accession Number: 18084303
Keywords: Alternative Splicing
Autistic Disorder/*genetics
Binding Sites
Calcium/metabolism
Calcium-Binding Proteins
Cell Adhesion/*physiology
Cell Adhesion Molecules/*chemistry/physiology
Cell Adhesion Molecules, Neuronal
Humans
Membrane Proteins/*chemistry/genetics/physiology
Models, Molecular
Nerve Tissue Proteins/*chemistry/genetics/physiology
Neural Cell Adhesion Molecules/*chemistry/physiology
Protein Conformation
Synapses/*physiology/ultrastructure
Abstract: The heterophilic synaptic adhesion molecules neuroligins and neurexins are essential for establishing and maintaining neuronal circuits by modulating the formation and maturation of synapses. The neuroligin-neurexin adhesion is Ca2+-dependent and regulated by alternative splicing. We report a structure of the complex at a resolution of 2.4 A between the mouse neuroligin-1 (NL1) cholinesterase-like domain and the mouse neurexin-1beta (NX1beta) LNS (laminin, neurexin and sex hormone-binding globulin-like) domain. The structure revealed a delicate neuroligin-neurexin assembly mediated by a hydrophilic, Ca2+-mediated and solvent-supplemented interface, rendering it capable of being modulated by alternative splicing and other regulatory factors. Thermodynamic data supported a mechanism wherein splicing site B of NL1 acts by modulating a salt bridge at the edge of the NL1-NX1beta interface. Mapping neuroligin mutations implicated in autism indicated that most such mutations are structurally destabilizing, supporting deficient neuroligin biosynthesis and processing as a common cause for this brain disorder.
Notes: 1545-9985
Chen, Xiaoyan
Liu, Heli
Shim, Ann H R
Focia, Pamela J
He, Xiaolin
R01 GM078055/GM/NIGMS NIH HHS/United States
R01 GM078055-02/GM/NIGMS NIH HHS/United States
1R01GM078055/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Struct Mol Biol. 2008 Jan;15(1):50-6. doi: 10.1038/nsmb1350. Epub 2007 Dec 16.
Author Address: Northwestern University Feinberg School of Medicine, Department of Molecular Pharmacology & Biological Chemistry, Searle 8-417, 303 East Chicago Avenue, Chicago, Illinois 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1848
Author: Chen, Y., Jin, L., Xue, B., Jin, D., Sun, F. and Wen, C.
Year: 2017
Title: NRAGE induces β-catenin/Arm O-GlcNAcylation and negatively regulates Wnt signaling
Journal: Biochem Biophys Res Commun
Volume: 487
Issue: 2
Pages: 433-437
Epub Date: 2017/04/22
Date: May 27
Short Title: NRAGE induces β-catenin/Arm O-GlcNAcylation and negatively regulates Wnt signaling
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2017.04.080
Accession Number: 28427939
Keywords: Acetylglucosamine/*metabolism
Acylation/physiology
Antigens, Neoplasm/*metabolism
Armadillo Domain Proteins/metabolism
Cell Nucleus/metabolism
DNA/*metabolism
Down-Regulation/physiology
HEK293 Cells
Humans
N-Acetylglucosaminyltransferases/*metabolism
Neoplasm Proteins/*metabolism
Wnt Signaling Pathway/*physiology
beta Catenin/*metabolism
*Nrage
*O-GlcNAc
*Wnt
*β-catenin/Arm
Abstract: The Wnt pathway is crucial for animal development, as well as tumor formation. Understanding the regulation of Wnt signaling will help to elucidate the mechanism of the cell cycle, cell differentiation and tumorigenesis. It is generally accepted that in response to Wnt signals, β-catenin accumulates in the cytoplasm and is imported into the nucleus where it recruits LEF/TCF transcription factors to activate the expression of target genes. In this study, we report that human NRAGE, a neurotrophin receptor p75 (p75NTR) binding protein, markedly suppresses the expression of genes activated by the Wnt pathway. Consistent with this finding, loss of function of NRAGE by RNA interference (RNAi) activates the Wnt pathway. Moreover, NRAGE suppresses the induction of axis duplication by microinjected β-catenin in Xenopus embryos. To our surprise, NRAGE induces nuclear localization of β-catenin and increases its DNA binding ability. Further studies reveal that NRAGE leads to the modification of β-catenin/Arm with O-linked beta-N-acetylglucosamine (O-GlcNAc), and failure of the association between β-catenin/Arm and pygopus(pygo) protein, which is required for transcriptional activation of Wnt target genes. Therefore, our findings suggest a novel mechanism for regulating Wnt signaling.
Notes: 1090-2104
Chen, Yuxin
Jin, Lei
Xue, Bin
Jin, Dong
Sun, Fenyong
Wen, Chuanjun
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2017 May 27;487(2):433-437. doi: 10.1016/j.bbrc.2017.04.080. Epub 2017 Apr 17.
Author Address: Department of Clinical Laboratory, Shanghai Tenth People's Hospital, Tongji University, Shanghai 200072, China.
College of Allied Health Professions, Shanghai University of Medicine and Health Sciences, Shanghai 201318, China.
Jiangsu Key Laboratory for Molecular and Medical Biotechnology, Nanjing Normal University, Nanjing 210097, China.
Department of Clinical Laboratory, Shanghai Tenth People's Hospital, Tongji University, Shanghai 200072, China. Electronic address: sunfenyong@126.com.
Jiangsu Key Laboratory for Molecular and Medical Biotechnology, Nanjing Normal University, Nanjing 210097, China. Electronic address: wenchuanjun123@163.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1956
Author: Chen, Y., Liu, R., Chu, Z., Le, B., Zeng, H., Zhang, X., Wu, Q., Zhu, G., Chen, Y., Liu, Y., Sun, F., Lu, Z., Qiao, Y. and Wang, J.
Year: 2018
Title: High glucose stimulates proliferative capacity of liver cancer cells possibly via O-GlcNAcylation-dependent transcriptional regulation of GJC1
Journal: J Cell Physiol
Volume: 234
Issue: 1
Pages: 606-618
Epub Date: 2018/08/06
Date: Jan
Short Title: High glucose stimulates proliferative capacity of liver cancer cells possibly via O-GlcNAcylation-dependent transcriptional regulation of GJC1
Alternate Journal: Journal of cellular physiology
ISSN: 0021-9541
DOI: 10.1002/jcp.26803
Accession Number: 30078215
Keywords: Cell Line, Tumor
Cell Proliferation/drug effects/genetics
Connexins/*genetics
Diabetes Complications/genetics/pathology
Diabetes Mellitus/*genetics/metabolism/pathology
Gene Expression Regulation, Neoplastic/drug effects
Glucose/pharmacology
Glycosylation/drug effects
Humans
Liver Neoplasms/*genetics/metabolism/pathology
N-Acetylglucosaminyltransferases/genetics
Promoter Regions, Genetic
Transcription Factors/*genetics
*Apa1
*diabetes
*liver cancer
*promoter
*transformative phenotypes
Abstract: Although it is generally accepted that diabetes is one of the most important risk factors for liver cancer, the underlying mechanism is still not well understood. The purpose of the current study is to further investigate how high concentrations of glucose (HG), a major symptom of diabetes, stimulate the development of liver malignancy. Using data mining, gap junction protein gamma 1 (GJC1) was identified as a critical proto-oncoprotein that is essential for the HG stimulation of proliferative capacity in liver cancer cells. Furthermore, enhanced transcriptional expression of GJC1 might occur after stimulation by HG. A transcription factor zinc finger protein 410 (APA1)-binding motif was found to be located at the -82 to -77 nt region within the GJC1 promoter. Without APA1, HG was unable to increase GJC1 expression. Interestingly, APA1, but not GJC1, can be O-GlcNAcylated in liver cancer cells. Moreover, O-GlcNAcylation is essential for HG-induced APA1 binding to the GJC1 promoter. Notably, global O-GlcNAcylation and expression of APA1 and GJC1 were highly elevated in liver cancer patients with diabetes compared to those in patients without diabetes. The HG-stimulated proliferative capacity was abolished upon decreasing O-GlcNAcylation, which could be reversed gradually by the simultaneous overexpression of APA1 and GJC1. Therefore, GJC1 could be a potential target for preventing liver cancer in patients with diabetes.
Notes: 1097-4652
Chen, Yan
Liu, Rui
Chu, Zhexuan
Le, Bu
Zeng, Hong
Zhang, Xiao
Wu, Qi
Zhu, Guoqing
Chen, Yuxin
Liu, Ya
Sun, Fenyong
Lu, Zhicheng
Qiao, Yongxia
Wang, Jiayi
Orcid: 0000-0003-1688-2864
Journal Article
Research Support, Non-U.S. Gov't
United States
J Cell Physiol. 2018 Jan;234(1):606-618. doi: 10.1002/jcp.26803. Epub 2018 Aug 5.
Author Address: Department of Clinical Laboratory, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, China.
Department of Scientific Research, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, China.
College of Laboratory Medicine, Chongqing Medical University, Chongqing, China.
Department of Endocrinology, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, China.
Department of Pathology, Sun Yet-Sen Memorial Hospital, Sun Yet-Sen University, Guangzhou, China.
Department of Medical Affairs Office, Shanghai seventh People's Hospital, Shanghai, China.
School of Public Health, Shanghai Jiaotong University School of Medicine, Shanghai, China.
Advanced Institute of Translational Medicine, Tongji University, Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2033
Author: Chen, Y., Zhu, G., Liu, Y., Wu, Q., Zhang, X., Bian, Z., Zhang, Y., Pan, Q. and Sun, F.
Year: 2019
Title: O-GlcNAcylated c-Jun antagonizes ferroptosis via inhibiting GSH synthesis in liver cancer
Journal: Cell Signal
Volume: 63
Pages: 109384
Epub Date: 2019/08/09
Date: Nov
Short Title: O-GlcNAcylated c-Jun antagonizes ferroptosis via inhibiting GSH synthesis in liver cancer
Alternate Journal: Cellular signalling
ISSN: 0898-6568
DOI: 10.1016/j.cellsig.2019.109384
Accession Number: 31394193
Keywords: Animals
Cell Line, Tumor
*Ferroptosis
Glutathione/*metabolism
Glycosylation
Humans
Liver Neoplasms/*metabolism
Liver Neoplasms, Experimental/*metabolism
Mice, Nude
Piperazines/*pharmacology
Proto-Oncogene Proteins c-jun/*metabolism
*Erastin
*Glutathione
*O-GlcNAcylation
*Phosphorylation
*Promoter
*Transcription
Abstract: Ferroptosis is a metabolism-related cell death. Stimulating ferroptosis in liver cancer cells is a strategy to treat liver cancer. However, how to eradicate liver cancer cells through ferroptosis and the obstacles to inducing ferroptosis in liver cancer remain unclear. Here, we observed that erastin suppressed the malignant phenotypes of liver cancer cells by inhibiting O-GlcNAcylation of c-Jun and further inhibited protein expression, transcription activity and nuclear accumulation of c-Jun. Overexpression of c-Jun-WT with simultaneous PuGNAc treatment conversely inhibited erastin-induced ferroptosis, whereas overexpression of c-Jun-WT alone or overexpression of c-Jun-S73A (a non-O-GlcNAcylated form of c-Jun) with PuGNAc treatment did not exert a similar effect. GSH downregulation induced by erastin was restored by overexpression of c-Jun-WT with simultaneous PuGNAc treatment. In addition, overexpression of c-Jun-WT, but not its S73A mutant, induced PSAT1 and CBS transcription via directly binding to their promoter regions, suggesting that GSH synthesis is regulated by O-GlcNAcylated c-Jun. A positive correlation between c-Jun O-GlcNAcylation and GSH was observed in clinical samples. Collectively, O-GlcNAcylated c-Jun represents an obstructive factor to ferroptosis, and targeting O-GlcNAcylated c-Jun might be helpful for treating liver cancer.
Notes: 1873-3913
Chen, Yan
Zhu, Guoqing
Liu, Ya
Wu, Qi
Zhang, Xiao
Bian, Zhixuan
Zhang, Yue
Pan, Qiuhui
Sun, Fenyong
Journal Article
Research Support, Non-U.S. Gov't
England
Cell Signal. 2019 Nov;63:109384. doi: 10.1016/j.cellsig.2019.109384. Epub 2019 Aug 5.
Author Address: Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
Shanghai Institute of Thoracic Tumors, Shanghai Chest Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200030, China.
Department of Laboratory Medicine, Shanghai Children's Medical Center, Shanghai Jiaotong University School of Medicine, Shanghai 200127, China.
Department of Central Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
Department of Laboratory Medicine, Shanghai Children's Medical Center, Shanghai Jiaotong University School of Medicine, Shanghai 200127, China. Electronic address: panqiuhui@263.net.
Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China. Electronic address: sunfenyongtongji@126.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1879
Author: Cheng, S., Mao, Q., Dong, Y., Ren, J., Su, L., Liu, J., Liu, Q., Zhou, J., Ye, X., Zheng, S. and Zhu, N.
Year: 2017
Title: GNB2L1 and its O-GlcNAcylation regulates metastasis via modulating epithelial-mesenchymal transition in the chemoresistance of gastric cancer
Journal: PLoS One
Volume: 12
Issue: 8
Pages: e0182696
Epub Date: 2017/08/11
Short Title: GNB2L1 and its O-GlcNAcylation regulates metastasis via modulating epithelial-mesenchymal transition in the chemoresistance of gastric cancer
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0182696
PMCID: PMC5552253
Accession Number: 28797110
Keywords: Acetylglucosamine/metabolism
Cell Line, Tumor
Cell Movement
*Drug Resistance, Neoplasm
*Epithelial-Mesenchymal Transition
GTP-Binding Proteins/*metabolism
Glycosylation
Humans
N-Acetylglucosaminyltransferases/metabolism
Neoplasm Proteins/*metabolism
*Protein Processing, Post-Translational
Receptors for Activated C Kinase
Receptors, Cell Surface/*metabolism
Stomach Neoplasms/drug therapy/*metabolism/pathology
Abstract: GNB2L1 and its O-GlcNAcylation has been reported to play roles in gastric cancer metastasis. However, the roles of GNB2L1 in chemoresistance of gastric cancer has never been determined. In the present study, we found that GNB2L1 was downregulated in chemoresistant patients of gastric cancer, and observed the decrease of GNB2L1 in protein levels instead of mRNA levels in different chemoresistant gastric cancer cell lines. Further we proved that this downregulation of GNB2L1 was resulted from its elevated O-GlcNAcylation catalyzed by OGT in both cell lines and patients. Next, we investigate the function of GNB2L1 and its O-GlcNAcylation on gastric cancer metastasis during chemoresistance, and confirmed Ser124 as the major O-GlcNAcylation site on GNB2L1 that regulated its function on metastasis. Furthermore, our data demonstrated that GNB2L1 modulated EMT via regulating the translation of EMT-related proteins in the process of chemoresistance. In summary, this study indicated that GNB2L1 and its O-GlcNAcylation regulated metastasis via modulating the translation of EMT-related proteins in the chemoresistance of gastric cancer.
Notes: 1932-6203
Cheng, Shimeng
Mao, Qiqi
Dong, Yabing
Ren, Jie
Su, Lina
Liu, Jianlan
Liu, Qingmei
Zhou, Jing
Ye, Xiaolu
Zheng, Shudan
Zhu, Ningwen
Orcid: 0000-0003-4224-4188
R01 AA020705/AA/NIAAA NIH HHS/United States
Journal Article
PLoS One. 2017 Aug 10;12(8):e0182696. doi: 10.1371/journal.pone.0182696. eCollection 2017.
Author Address: Department of Dermatology, Huashan Hospital, Fudan University, Shanghai, China.
Department of Physiology, Hainan Medical College, Haikou, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1805
Author: Cheng, S., Ren, J., Su, L., Liu, J., Liu, Q., Zhou, J., Ye, X. and Zhu, N.
Year: 2016
Title: O-GlcNAcylation of the Signaling Scaffold Protein, GNB2L1 Promotes its Degradation and Increases Metastasis of Gastric Tumours
Journal: Biochem Biophys Res Commun
Volume: 478
Issue: 4
Pages: 1497-502
Epub Date: 2016/08/18
Date: Sep 30
Short Title: O-GlcNAcylation of the Signaling Scaffold Protein, GNB2L1 Promotes its Degradation and Increases Metastasis of Gastric Tumours
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2016.08.074
Accession Number: 27530921
Keywords: Acetylglucosamine/*metabolism
Cell Line, Tumor
GTP-Binding Proteins/genetics/*metabolism
Gene Expression Regulation, Neoplastic
Glycosylation
Humans
N-Acetylglucosaminyltransferases/metabolism
Neoplasm Metastasis
Neoplasm Proteins/genetics/*metabolism
Protein Binding
Proteolysis
RNA, Messenger/genetics/metabolism
Receptors for Activated C Kinase
Receptors, Cell Surface/genetics/*metabolism
Stomach Neoplasms/genetics/*metabolism/*pathology
*Degradation
*Gnb2l1
*Gastric cancer
*Metastasis
*O-GlcNAcylation
Abstract: GNB2L1 is an intercellular scaffold protein of the Trp-Asp (WD) repeat protein family, and has been reported to play suppressive roles in the progression of gastric cancer. However, the regulatory mechanisms of GNB2L1 in gastric cancer still remain largely elusive. In the present study, we found that OGT was capable to interact with GNB2L1 directly and modify GNB2L1 with O-GlcNAcylation in gastric cancer, and this O-GlcNAcylation hindered the inhibition of GNB2L1 on migration of gastric cancer cells. Moreover, O-GlcNAcylation regulated the degradation instead of the synthesis of GNB2L1 in gastric cancer, and our data suggested the O-GlcNAcylation on GNB2L1 influenced its stability directly. In addition, the clinical data revealed the negative correlation of the protein level instead of the mRNA level of GNB2L1 with OGT expression, and showed that OGT reversed the inhibition of GNB2L1 on metastasis, and worsened the prognosis of GNB2L1(High) patients. In summary, this study indicated the O-GlcNAcylation on GNB2L1 reversed its inhibition on gastric tumour metastasis via promoting its degradation.
Notes: 1090-2104
Cheng, Shimeng
Ren, Jie
Su, Lina
Liu, Jianlan
Liu, Qingmei
Zhou, Jing
Ye, Xiaolu
Zhu, Ningwen
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2016 Sep 30;478(4):1497-502. doi: 10.1016/j.bbrc.2016.08.074. Epub 2016 Aug 13.
Author Address: Department of Dermatology, Huashan Hospital, Fudan University, Shanghai, 200040, China.
Department of Dermatology, Huashan Hospital, Fudan University, Shanghai, 200040, China. Electronic address: zhuningwen@fudan.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 772
Author: Cheng, X., Cole, R. N., Zaia, J. and Hart, G. W.
Year: 2000
Title: Alternative O-glycosylation/O-phosphorylation of the murine estrogen receptor beta
Journal: Biochemistry
Volume: 39
Issue: 38
Pages: 11609-20
Epub Date: 2000/09/20
Date: Sep 26
Short Title: Alternative O-glycosylation/O-phosphorylation of the murine estrogen receptor beta
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi000755i
Accession Number: 10995228
Keywords: Acetylglucosamine/isolation & purification/metabolism
Acylation
Amino Acid Sequence
Animals
COS Cells
Carbohydrate Conformation
Estrogen Receptor beta
Glycosylation
Mice
Molecular Sequence Data
Peptide Fragments/chemistry/genetics/metabolism
Phosphorylation
Protein Structure, Tertiary/genetics
Receptors, Estrogen/biosynthesis/*chemistry/genetics/*metabolism
Serine/genetics/metabolism
Spodoptera/genetics
Transcriptional Activation
Abstract: Estrogen receptor beta, a homologue to estrogen receptor alpha, is a new member of the steroid hormone receptor family. Recently, we documented that estrogen receptor alpha, like other transcription factors, is modified by O-linked N-acetylglucosamine (O-GlcNAc), a ubiquitous transitory posttranslational modification on nuclear and cytoplasmic proteins. Here, we report that estrogen receptor beta is alternatively modified by either O-GlcNAc or O-phosphate. Lectin chromatography of in vitro translated protein first suggested that murine estrogen receptor beta (mER-beta) is O-GlcNAcylated. Structural characterization of the carbohydrate moieties on mER-beta, overexpressed in insect Sf9 cells, confirmed the presence of O-GlcNAc. mER-beta, overexpressed in mammalian cells, is also O-GlcNAcylated. The major site of O-GlcNAc on mER-beta from Sf9 cells is Ser(16) near the N-terminus. Concomitant analyses also documented the O-phosphorylation of mER-beta at Ser(16). MALDI-TOF mass spectrometry showed alternative occupancy of this locus by these two abundant and dynamic posttranslational modifications. The localization of a major O-GlcNAc/O-phosphate site in proximity of the transactivation domain and as part of a PEST region (target sequences for rapid protein degradation) on mER-beta suggests that these modifications may play a role in regulating estrogen receptor beta transactivation and turnover.
Notes: Cheng, X
Cole, R N
Zaia, J
Hart, G W
CA42486/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2000 Sep 26;39(38):11609-20. doi: 10.1021/bi000755i.
Author Address: Department of Biological Chemistry, School of Medicine, The Johns Hopkins University, 725 North Wolfe Street, Baltimore, Maryland 21205-2185, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 787
Author: Cheng, X. and Hart, G. W.
Year: 2000
Title: Glycosylation of the murine estrogen receptor-alpha
Journal: J Steroid Biochem Mol Biol
Volume: 75
Issue: 2-3
Pages: 147-58
Epub Date: 2001/02/28
Date: Dec 15
Short Title: Glycosylation of the murine estrogen receptor-alpha
Alternate Journal: The Journal of steroid biochemistry and molecular biology
ISSN: 0960-0760 (Print)
0960-0760
DOI: 10.1016/s0960-0760(00)00167-9
Accession Number: 11226831
Keywords: Acetylglucosamine/*chemistry/metabolism
Amino Acid Sequence
Animals
Chromatography, Affinity
Electrophoresis, Polyacrylamide Gel
Estrogen Receptor alpha
Galactosyltransferases/metabolism
Glycosylation
Mice
Molecular Sequence Data
Mutagenesis, Site-Directed
Peptide Mapping
Peptides/chemistry
*Protein Processing, Post-Translational
Receptors, Estrogen/*chemistry/genetics/metabolism
Recombinant Proteins/genetics/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Transfection
Trypsin/metabolism
Abstract: O-linked N-acetylglucosamine (O-GlcNAc) is a highly dynamic and abundant modification found on nuclear and cytoplasmic proteins of nearly all eukaryotes. O-GlcNAc addition is required for life at the single cell level and is analogous to protein phosphorylation in most respects. In a previous study (M.S. Jiang, G.W. Hart, A subpopulation of estrogen receptors are modified by O-linked N-acetylglucosamine. J. Biol. Chem. 270 (1997) 2421-2428), we demonstrated that a subpopulation of the murine estrogen receptor-alpha (mER-alpha) is modified by O-GlcNAc at Thr(575). Here we mutated mER-alpha to convert Thr(575) and Ser(576) to Val and Ala, respectively. Surprisingly, this glycosylation-site mutant is still extensively modified by O-GlcNAc. Analyses of glycopeptides identified two additional sites of modification on mER-alpha, at Ser(10) and Thr(50) near the N-terminus. The major glycosylation sites are within or near PEST regions, suggesting that O-GlcNAc may regulate mER-alpha turnover.
Notes: Cheng, X
Hart, G W
CA42486/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
J Steroid Biochem Mol Biol. 2000 Dec 15;75(2-3):147-58. doi: 10.1016/s0960-0760(00)00167-9.
Author Address: Department of Biological Chemistry, School of Medicine, Johns Hopkins University, 725 N Wolfe St., Baltimore, MD 21205-2185, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1521
Author: Cheung, J., Rudolph, M. J., Burshteyn, F., Cassidy, M. S., Gary, E. N., Love, J., Franklin, M. C. and Height, J. J.
Year: 2012
Title: Structures of human acetylcholinesterase in complex with pharmacologically important ligands
Journal: J Med Chem
Volume: 55
Issue: 22
Pages: 10282-6
Epub Date: 2012/10/06
Date: Nov 26
Short Title: Structures of human acetylcholinesterase in complex with pharmacologically important ligands
Alternate Journal: Journal of medicinal chemistry
ISSN: 0022-2623
DOI: 10.1021/jm300871x
Accession Number: 23035744
Keywords: Acetylcholinesterase/*chemistry/metabolism
Acetylthiocholine/metabolism
Animals
Cholinesterase Inhibitors/*metabolism
Crystallography, X-Ray
Donepezil
Humans
Indans/*metabolism
Models, Molecular
Piperidines/*metabolism
Protein Conformation
Torpedo/*metabolism
Abstract: Human acetylcholinesterase (AChE) is a significant target for therapeutic drugs. Here we present high resolution crystal structures of human AChE, alone and in complexes with drug ligands; donepezil, an Alzheimer's disease drug, binds differently to human AChE than it does to Torpedo AChE. These crystals of human AChE provide a more accurate platform for further drug development than previously available.
Notes: 1520-4804
Cheung, Jonah
Rudolph, Michael J
Burshteyn, Fiana
Cassidy, Michael S
Gary, Ebony N
Love, James
Franklin, Matthew C
Height, Jude J
Comparative Study
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Med Chem. 2012 Nov 26;55(22):10282-6. doi: 10.1021/jm300871x. Epub 2012 Nov 12.
Author Address: New York Structural Biology Center, New York, New York 10027, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1527
Author: Chhikara, N., Saraswat, M., Tomar, A. K., Dey, S., Singh, S. and Yadav, S.
Year: 2012
Title: Human epididymis protein-4 (HE-4): a novel cross-class protease inhibitor
Journal: PLoS One
Volume: 7
Issue: 11
Pages: e47672
Epub Date: 2012/11/10
Short Title: Human epididymis protein-4 (HE-4): a novel cross-class protease inhibitor
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0047672
PMCID: PMC3489902
Accession Number: 23139753
Keywords: Chromatography, Affinity
Chromatography, Gel
Chromatography, Ion Exchange
Chromatography, Liquid
Glycosylation
Humans
Male
Mass Spectrometry
Native Polyacrylamide Gel Electrophoresis
Peptides/chemistry/metabolism
Protease Inhibitors/chemistry/isolation & purification/*metabolism
Protein Binding
Protein Multimerization
Proteins/chemistry/isolation & purification/*metabolism
Semen/metabolism
Sequence Analysis, Protein
Surface Plasmon Resonance
WAP Four-Disulfide Core Domain Protein 2
Abstract: Epididymal proteins represent the factors necessary for maturation of sperm and play a crucial role in sperm maturation. HE-4, an epididymal protein, is a member of whey acidic protein four-disulfide core (WFDC) family with no known function. A WFDC protein has a conserved WFDC domain of 50 amino acids with eight conserved cystine residue. HE-4 is a 124 amino acid long polypeptide with two WFDC domains. Here, we show that HE-4 is secreted in the human seminal fluid as a disulfide-bonded homo-trimer and is a cross-class protease inhibitor inhibits some of the serine, aspartyl and cysteine proteases tested using hemoglobin as a substrate. Using SPR we have also observed that HE-4 shows a significant binding with all these proteases. Disulfide linkages are essential for this activity. Moreover, HE-4 is N-glycosylated and highly stable on a wide range of pH and temperature. Taken together this suggests that HE-4 is a cross-class protease inhibitor which might confer protection against microbial virulence factors of proteolytic nature.
Notes: 1932-6203
Chhikara, Nirmal
Saraswat, Mayank
Tomar, Anil Kumar
Dey, Sharmistha
Singh, Sarman
Yadav, Savita
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2012;7(11):e47672. doi: 10.1371/journal.pone.0047672. Epub 2012 Nov 5.
Author Address: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1947
Author: Cho, H. J. and Mook-Jung, I.
Year: 2018
Title: O-GlcNAcylation regulates endoplasmic reticulum exit sites through Sec31A modification in conventional secretory pathway
Journal: Faseb j
Volume: 32
Issue: 9
Pages: 4641-4657
Epub Date: 2018/06/20
Date: Sep
Short Title: O-GlcNAcylation regulates endoplasmic reticulum exit sites through Sec31A modification in conventional secretory pathway
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638
DOI: 10.1096/fj.201701523R
Accession Number: 29913562
Keywords: Apoptosis Regulatory Proteins/genetics
Calcium-Binding Proteins/metabolism
Endoplasmic Reticulum/*metabolism
Golgi Apparatus/*metabolism
Humans
Protein Binding/physiology
Protein Transport/*physiology
Secretory Pathway/*physiology
Vesicular Transport Proteins/metabolism
*Alg-2
*COPII vesicles
*Eres
*ER–Golgi
*transport
Abstract: The conventional secretory pathway is indispensable for eukaryotic cells. Newly synthesized membrane and secretory proteins are released from the endoplasmic reticulum (ER) through ER-derived vesicles to their appropriate destination. Vesicle formation is important for steady protein trafficking. O-GlcNAcylation ( O-GlcNAc) is a unique protein glycosylation signature, whose dynamic regulation by O-GlcNAc transferase and O-GlcNAcase occurs exclusively for nuclear and cytoplasmic proteins. Because of this locally limited property, the role of O-GlcNAc in the conventional protein secretory pathway is unknown. We report that Sec31A on COPII vesicles, a specific coat-protein complex for anterograde trafficking in the ER-Golgi network, is O-GlcNAcylated on S964, which accelerates COPII vesicle formation through control of its binding affinity to apoptosis-linked gene 2, a calcium-binding protein. Together, O-GlcNAc on Sec31A regulates conventional secretory vesicle trafficking in the ER-Golgi network. These modifications accelerate COPII vesicle formation and accelerated anterograde transport of vesicles within the ER-Golgi networks.-Cho, H. J., Mook-Jung, I. O-GlcNAcylation regulates endoplasmic reticulum exit sites through Sec31A modification in conventional secretory pathway.
Notes: 1530-6860
Cho, Hyun Jin
Mook-Jung, Inhee
Journal Article
Research Support, Non-U.S. Gov't
United States
FASEB J. 2018 Sep;32(9):4641-4657. doi: 10.1096/fj.201701523R. Epub 2018 Apr 17.
Author Address: Department of Biochemistry and Biomedical Sciences, College of Medicine, Seoul National University, Seoul, South Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 846
Author: Cho, H. S. and Leahy, D. J.
Year: 2002
Title: Structure of the extracellular region of HER3 reveals an interdomain tether
Journal: Science
Volume: 297
Issue: 5585
Pages: 1330-3
Epub Date: 2002/08/03
Date: Aug 23
Short Title: Structure of the extracellular region of HER3 reveals an interdomain tether
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1074611
Accession Number: 12154198
Keywords: Amino Acid Sequence
Amino Acid Substitution
Animals
CHO Cells
Cricetinae
Crystallography, X-Ray
Dimerization
Epidermal Growth Factor/chemistry/metabolism
ErbB Receptors/chemistry/metabolism
Humans
Hydrogen Bonding
Ligands
Molecular Sequence Data
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Receptor, ErbB-3/*chemistry/metabolism
Recombinant Proteins/chemistry
Signal Transduction
Abstract: We have determined the 2.6 angstrom crystal structure of the entire extracellular region of human HER3 (ErbB3), a member of the epidermal growth factor receptor (EGFR) family. The structure consists of four domains with structural homology to domains found in the type I insulin-like growth factor receptor. The HER3 structure reveals a contact between domains II and IV that constrains the relative orientations of ligand-binding domains and provides a structural basis for understanding both multiple-affinity forms of EGFRs and conformational changes induced in the receptor by ligand binding during signaling. These results also suggest new therapeutic approaches to modulating the behavior of members of the EGFR family.
Notes: 1095-9203
Cho, Hyun-Soo
Leahy, Daniel J
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Science. 2002 Aug 23;297(5585):1330-3. doi: 10.1126/science.1074611. Epub 2002 Aug 1.
Author Address: Department of Biophysics and Biophysical Chemistry, Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 871
Author: Cho, H. S., Mason, K., Ramyar, K. X., Stanley, A. M., Gabelli, S. B., Denney, D. W., Jr. and Leahy, D. J.
Year: 2003
Title: Structure of the extracellular region of HER2 alone and in complex with the Herceptin Fab
Journal: Nature
Volume: 421
Issue: 6924
Pages: 756-60
Epub Date: 2003/03/01
Date: Feb 13
Short Title: Structure of the extracellular region of HER2 alone and in complex with the Herceptin Fab
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature01392
Accession Number: 12610629
Keywords: Animals
Antibodies, Monoclonal/*chemistry/*immunology
Antibodies, Monoclonal, Humanized
Binding Sites
Binding Sites, Antibody
Crystallography, X-Ray
Humans
Immunoglobulin Fab Fragments/*chemistry/*immunology
Ligands
Models, Molecular
Protein Structure, Tertiary
Rats
Receptor, ErbB-2/*chemistry/*immunology
Trastuzumab
Abstract: HER2 (also known as Neu, ErbB2) is a member of the epidermal growth factor receptor (EGFR; also known as ErbB) family of receptor tyrosine kinases, which in humans includes HER1 (EGFR, ERBB1), HER2, HER3 (ERBB3) and HER4 (ERBB4). ErbB receptors are essential mediators of cell proliferation and differentiation in the developing embryo and in adult tissues, and their inappropriate activation is associated with the development and severity of many cancers. Overexpression of HER2 is found in 20-30% of human breast cancers, and correlates with more aggressive tumours and a poorer prognosis. Anticancer therapies targeting ErbB receptors have shown promise, and a monoclonal antibody against HER2, Herceptin (also known as trastuzumab), is currently in use as a treatment for breast cancer. Here we report crystal structures of the entire extracellular regions of rat HER2 at 2.4 A and human HER2 complexed with the Herceptin antigen-binding fragment (Fab) at 2.5 A. These structures reveal a fixed conformation for HER2 that resembles a ligand-activated state, and show HER2 poised to interact with other ErbB receptors in the absence of direct ligand binding. Herceptin binds to the juxtamembrane region of HER2, identifying this site as a target for anticancer therapies.
Notes: Cho, Hyun-Soo
Mason, Karen
Ramyar, Kasra X
Stanley, Ann Marie
Gabelli, Sandra B
Denney, Dan W Jr
Leahy, Daniel J
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Nature. 2003 Feb 13;421(6924):756-60. doi: 10.1038/nature01392.
Author Address: Department of Biophysics and Biophysical Chemistry, The Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 991
Author: Choe, J., Kelker, M. S. and Wilson, I. A.
Year: 2005
Title: Crystal structure of human toll-like receptor 3 (TLR3) ectodomain
Journal: Science
Volume: 309
Issue: 5734
Pages: 581-5
Epub Date: 2005/06/18
Date: Jul 22
Short Title: Crystal structure of human toll-like receptor 3 (TLR3) ectodomain
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1115253
Accession Number: 15961631
Keywords: Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Dimerization
Glycosylation
Humans
Hydrogen Bonding
Leucine/chemistry
Ligands
Membrane Glycoproteins/*chemistry/metabolism
Models, Molecular
Molecular Sequence Data
Protein Conformation
Protein Structure, Tertiary
RNA, Double-Stranded/metabolism
Receptors, Cell Surface/*chemistry/metabolism
Repetitive Sequences, Amino Acid
Signal Transduction
Static Electricity
Surface Properties
Toll-Like Receptor 3
Toll-Like Receptors
Abstract: Toll-like receptors (TLRs) play key roles in activating immune responses during infection. The human TLR3 ectodomain structure at 2.1 angstroms reveals a large horseshoe-shaped solenoid assembled from 23 leucine-rich repeats (LRRs). Asparagines conserved in the 24-residue LRR motif contribute extensive hydrogen-bonding networks for solenoid stabilization. TLR3 is largely masked by carbohydrate, but one face is glycosylation-free, which suggests its potential role in ligand binding and oligomerization. Highly conserved surface residues and a TLR3-specific LRR insertion form a homodimer interface in the crystal, whereas two patches of positively charged residues and a second insertion would provide an appropriate binding site for double-stranded RNA.
Notes: 1095-9203
Choe, Jungwoo
Kelker, Matthew S
Wilson, Ian A
R01 AI042266/AI/NIAID NIH HHS/United States
AI-42266/AI/NIAID NIH HHS/United States
CA-58896/CA/NCI NIH HHS/United States
T32 AI077606/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Science. 2005 Jul 22;309(5734):581-5. doi: 10.1126/science.1115253. Epub 2005 Jun 16.
Author Address: Department of Molecular Biology and Skaggs Institute for Chemical Biology, Scripps Research Institute (TSRI), 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1987
Author: Choi, H., Kim, C., Song, H., Cha, M. Y., Cho, H. J., Son, S. M., Kim, H. J. and Mook-Jung, I.
Year: 2019
Title: Amyloid β-induced elevation of O-GlcNAcylated c-Fos promotes neuronal cell death
Journal: Aging Cell
Volume: 18
Issue: 1
Pages: e12872
Epub Date: 2018/12/06
Date: Feb
Short Title: Amyloid β-induced elevation of O-GlcNAcylated c-Fos promotes neuronal cell death
Alternate Journal: Aging cell
ISSN: 1474-9718 (Print)
1474-9718
DOI: 10.1111/acel.12872
PMCID: PMC6351842
Accession Number: 30515991
Keywords: Alzheimer Disease/pathology
Amyloid beta-Peptides/*metabolism/toxicity
Animals
Bcl-2-Like Protein 11/genetics/metabolism
Cell Death/drug effects
Cell Line
Gene Expression Regulation/drug effects
Glycosylation/drug effects
Humans
Mice, Transgenic
Neurons/drug effects/*pathology
Protein Stability/drug effects
Proto-Oncogene Proteins c-fos/genetics/*metabolism
Rats, Sprague-Dawley
Transcription, Genetic/drug effects
beta-N-Acetylhexosaminidases/metabolism
*Alzheimer’s disease
*O-linked β-N-acetyl glucosamine (O-GlcNAc)
*c-Fos
*glucose metabolism
*neuronal cell death
*β-amyloid (Aβ)
Abstract: Alzheimer's disease (AD) is an age-related neurodegenerative disease characterized by progressive memory loss resulting from cumulative neuronal cell death. O-linked β-N-acetyl glucosamine (O-GlcNAc) modification of the proteins reflecting glucose metabolism is altered in the brains of patients with AD. However, the link between altered O-GlcNAc modification and neuronal cell death in AD is poorly understood. Here, we examined the regulation of O-GlcNAcylation of c-Fos and the effects of O-GlcNAcylated c-Fos on neuronal cell death during AD pathogenesis. We found that amyloid beta (Aβ)-induced O-GlcNAcylation on serine-56 and 57 of c-Fos was resulted from decreased interaction between c-Fos and O-GlcNAcase and promoted neuronal cell death. O-GlcNAcylated c-Fos increased its stability and potentiated the transcriptional activity through higher interaction with c-Jun, resulting in induction of Bim expression leading to neuronal cell death. Taken together, Aβ-induced O-GlcNAcylation of c-Fos plays an important role in neuronal cell death during the pathogenesis of AD.
Notes: 1474-9726
Choi, Heesun
Kim, Chaeyoung
Song, Hyundong
Cha, Moon-Yong
Cho, Hyun Jin
Son, Sung Min
Kim, Haeng Jun
Mook-Jung, Inhee
2018R1A2A1A19019062/NRF/International
2014M3C7A1046047/NRF/International
2015M3C7A1028790/NRF/International
2012R1A5A2A44671346/MRC/International
Journal Article
Research Support, Non-U.S. Gov't
Aging Cell. 2019 Feb;18(1):e12872. doi: 10.1111/acel.12872. Epub 2018 Dec 4.
Author Address: Department of Biochemistry and Biomedical Sciences, Seoul National University, College of Medicine, Seoul, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 870
Author: Choi, I., Hu, L., Rojas, J. D., Schmitt, B. M. and Boron, W. F.
Year: 2003
Title: Role of glycosylation in the renal electrogenic Na+-HCO3- cotransporter (NBCe1)
Journal: Am J Physiol Renal Physiol
Volume: 284
Issue: 6
Pages: F1199-206
Epub Date: 2003/02/27
Date: Jun
Short Title: Role of glycosylation in the renal electrogenic Na+-HCO3- cotransporter (NBCe1)
Alternate Journal: American journal of physiology. Renal physiology
ISSN: 1931-857X (Print)
1522-1466
DOI: 10.1152/ajprenal.00131.2002
Accession Number: 12604466
Keywords: Amidohydrolases/chemistry
Amino Acid Substitution/physiology
Animals
Cell Membrane/enzymology/metabolism
Glycosylation
Histocytochemistry
Hydrogen-Ion Concentration
Immunoblotting
Kidney/*metabolism
Lectins
Microelectrodes
Molecular Weight
Mutagenesis, Site-Directed
Mutation/genetics/physiology
Oligosaccharides/metabolism
Oocytes/metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Rabbits
Rats
Rats, Sprague-Dawley
Sodium-Bicarbonate Symporters/chemistry/*metabolism/physiology
Species Specificity
Xenopus laevis
Abstract: The electrogenic Na(+)-HCO(3)(-) cotransporter NBCe1 is important for the regulation of intracellular pH (pH(i)) and for epithelial HCO(3)(-) transport in many tissues, including kidney, pancreas, and brain. In the present study, we investigate glycosylation sites in NBCe1. Treatment of rat kidney membrane extracts with peptide N-glycosidase F (PNGase F) shifted the apparent molecular weight (MW) of NBCe1 from 130 to 116, the MW predicted from the deduced amino acid sequence. Treatment with endoglycosidase F(2) or H or O-glycosidase did not affect the MW of NBCe1. Lectin-binding studies, together with the enzyme data, suggest that the N-linked carbohydrates are of tri- or tetra-antennary type. To localize glycosylation sites, we individually mutated the seven consensus N-glycosylation sites by replacing asparagine (N) with glutamine (Q) and assessing mutant transporters in Xenopus laevis oocytes. Immunoblotting of oocyte membrane extracts treated with PNGase F indicates that NBCe1 is normally glycosylated at N597 and N617 (both on the third extracellular loop). However, N592 (on the same loop) is glycosylated when the other two sites are mutated. The triple mutant (N592Q/N597Q/N617Q) is completely unglycosylated but, based on microelectrode measurements of membrane potential and pH(i) in oocytes, preserves the Na(+) and HCO(3)(-) dependence and electrogenicity of wild-type NBCe1.
Notes: Choi, Inyeong
Hu, Lihui
Rojas, José D
Schmitt, Bernhard M
Boron, Walter F
DK-30344/DK/NIDDK NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Am J Physiol Renal Physiol. 2003 Jun;284(6):F1199-206. doi: 10.1152/ajprenal.00131.2002. Epub 2003 Feb 25.
Author Address: Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06520, USA. ichoi@physio.emory.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1286
Author: Cholon, D. M., O'Neal, W. K., Randell, S. H., Riordan, J. R. and Gentzsch, M.
Year: 2010
Title: Modulation of endocytic trafficking and apical stability of CFTR in primary human airway epithelial cultures
Journal: Am J Physiol Lung Cell Mol Physiol
Volume: 298
Issue: 3
Pages: L304-14
Epub Date: 2009/12/17
Date: Mar
Short Title: Modulation of endocytic trafficking and apical stability of CFTR in primary human airway epithelial cultures
Alternate Journal: American journal of physiology. Lung cellular and molecular physiology
ISSN: 1040-0605 (Print)
1040-0605
DOI: 10.1152/ajplung.00016.2009
PMCID: PMC2838667
Accession Number: 20008117
Keywords: Cell Differentiation
Cell Membrane
*Cell Polarity
Cells, Cultured
Cystic Fibrosis Transmembrane Conductance Regulator/*metabolism
*Endocytosis
Epithelial Cells/*cytology/*metabolism
Glycosylation
Humans
Lung/*cytology
Polysaccharides/metabolism
Protein Stability
Protein Transport
Subcellular Fractions/metabolism
Surface Properties
Temperature
Abstract: CFTR is a highly regulated apical chloride channel of epithelial cells that is mutated in cystic fibrosis (CF). In this study, we characterized the apical stability and intracellular trafficking of wild-type and mutant CFTR in its native environment, i.e., highly differentiated primary human airway epithelial (HAE) cultures. We labeled the apical pool of CFTR and subsequently visualized the protein in intracellular compartments. CFTR moved from the apical surface to endosomes and then efficiently recycled back to the surface. CFTR endocytosis occurred more slowly in polarized than in nonpolarized HAE cells or in a polarized epithelial cell line. The most common mutation in CF, DeltaF508 CFTR, was rescued from endoplasmic reticulum retention by low-temperature incubation but transited from the apical membrane to endocytic compartments more rapidly and recycled less efficiently than wild-type CFTR. Incubation with small-molecule correctors resulted in DeltaF508 CFTR at the apical membrane but did not restore apical stability. To stabilize the mutant protein at the apical membrane, we found that the dynamin inhibitor Dynasore and the cholesterol-extracting agent cyclodextrin dramatically reduced internalization of DeltaF508, whereas the proteasomal inhibitor MG-132 completely blocked endocytosis of DeltaF508. On examination of intrinsic properties of CFTR that may affect its apical stability, we found that N-linked oligosaccharides were not necessary for transport to the apical membrane but were required for efficient apical recycling and, therefore, influenced the turnover of surface CFTR. Thus apical stability of CFTR in its native environment is affected by properties of the protein and modulation of endocytic trafficking.
Notes: 1522-1504
Cholon, Deborah M
O'Neal, Wanda K
Randell, Scott H
Riordan, John R
Gentzsch, Martina
R01 HL-080561/HL/NHLBI NIH HHS/United States
R01 DK-051870/DK/NIDDK NIH HHS/United States
R01 DK051619/DK/NIDDK NIH HHS/United States
P30 DK-065988/DK/NIDDK NIH HHS/United States
P50 HL-60280/HL/NHLBI NIH HHS/United States
P30 DK065988/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Am J Physiol Lung Cell Mol Physiol. 2010 Mar;298(3):L304-14. doi: 10.1152/ajplung.00016.2009. Epub 2009 Dec 11.
Author Address: Cystic Fibrosis Research Center and Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7248, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 777
Author: Chong, J. M. and Speicher, D. W.
Year: 2001
Title: Determination of disulfide bond assignments and N-glycosylation sites of the human gastrointestinal carcinoma antigen GA733-2 (CO17-1A, EGP, KS1-4, KSA, and Ep-CAM)
Journal: J Biol Chem
Volume: 276
Issue: 8
Pages: 5804-13
Epub Date: 2000/11/18
Date: Feb 23
Short Title: Determination of disulfide bond assignments and N-glycosylation sites of the human gastrointestinal carcinoma antigen GA733-2 (CO17-1A, EGP, KS1-4, KSA, and Ep-CAM)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M008839200
Accession Number: 11080501
Keywords: Amino Acid Sequence
Antigens, Neoplasm/*chemistry/genetics
Asparagine/chemistry
Carcinoma/*immunology
Cell Adhesion Molecules/*chemistry/genetics
Cysteine/chemistry
Cystine/chemistry
Disulfides/*chemistry
Epithelial Cell Adhesion Molecule
Gastrointestinal Neoplasms/*immunology
Glycoproteins/*chemistry/genetics
Humans
Models, Molecular
Molecular Sequence Data
Peptide Fragments/chemistry
Protein Structure, Tertiary
Recombinant Proteins/chemistry
Sequence Analysis, Protein
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: The GA733-2 antigen is a cell surface glycoprotein highly expressed on most human gastrointestinal carcinoma and at a lower level on most normal epithelia. It is an unusual cell-cell adhesion protein that does not exhibit any obvious relationship to the four known classes of adhesion molecules. In this study, the disulfide-bonding pattern of the GA733-2 antigen was determined using matrix-assisted laser desorption/ionization mass spectrometry and N-terminal sequencing of purified tryptic peptides treated with 2-[2'-nitrophenylsulfonyl]-3-methyl-3-bromoindolenine or partially reduced and alkylated. Numbering GA733-2 cysteines sequentially from the N terminus, the first three disulfide linkages are Cys1-Cys4, Cys2-Cys6, and Cys3-Cys5, which is a novel pattern for a cysteine-rich domain instead of the expected epidermal growth factor-like disulfide structure. The next three disulfide linkages are Cys7-Cys8, Cys9-Cys10, and Cys11-Cys12, consistent with the recently determined disulfide pattern of the thyroglobulin type 1A domain of insulin-like growth factor-binding proteins 1 and 6. Analysis of glycosylation sites showed that GA733-2 antigen contained N-linked carbohydrate but that no O-linked carbohydrate groups were detected. Of the three potential N-linked glycosylation sites, Asn175 was not glycosylated, whereas Asn88 was completely glycosylated, and Asn51 was partially glycosylated. These data show that the extracellular domain of the GA733-2 antigen consists of three distinct domains; a novel cysteine-rich N-terminal domain (GA733 type 1 motif), a cysteine-rich thyroglobulin type 1A domain (GA733 type 2 motif), and a unique nonglycosylated domain without cysteines (GA733 type 3 motif).
Notes: Chong, J M
Speicher, D W
CA10815/CA/NCI NIH HHS/United States
CA74294/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2001 Feb 23;276(8):5804-13. doi: 10.1074/jbc.M008839200. Epub 2000 Nov 15.
Author Address: Wistar Institute, Philadelphia, Pennsylvania 19104, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 829
Author: Chong, J. M., Uren, A., Rubin, J. S. and Speicher, D. W.
Year: 2002
Title: Disulfide bond assignments of secreted Frizzled-related protein-1 provide insights about Frizzled homology and netrin modules
Journal: J Biol Chem
Volume: 277
Issue: 7
Pages: 5134-44
Epub Date: 2001/12/14
Date: Feb 15
Short Title: Disulfide bond assignments of secreted Frizzled-related protein-1 provide insights about Frizzled homology and netrin modules
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M108533200
Accession Number: 11741940
Keywords: Amino Acid Sequence
Animals
Asparagine/chemistry
Binding Sites
Chromatography, High Pressure Liquid
Cysteine/chemistry
Cystine/chemistry
Disulfides
Electrophoresis, Polyacrylamide Gel
Frizzled Receptors
Glycosylation
Heparin/chemistry
Humans
Lysine/chemistry
Mass Spectrometry
Mice
Molecular Sequence Data
Mutagenesis, Site-Directed
Netrin Receptors
Phenylalanine/chemistry
Protein Binding
Protein Structure, Tertiary
Proteins/*chemistry
Receptors, Cell Surface/*chemistry
Recombinant Proteins/metabolism
Sequence Homology, Amino Acid
Serine/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Subtilisin/chemistry
Time Factors
Xenopus
Abstract: Secreted Frizzled-related protein-1 (sFRP-1), a soluble protein that binds to Wnts and modulates Wnt signaling, contains an N-terminal domain homologous to the putative Wnt-binding site of Frizzled (Fz domain) and a C-terminal heparin-binding domain with weak homology to netrin. Both domains are cysteine-rich, having 10 and 6 cysteines in the Fz and heparin-binding domains, respectively. In this study, the disulfide linkages of recombinant sFRP-1 were determined. Numbering sFRP-1 cysteines sequentially from the N terminus, the five disulfide linkages in the Fz domain are 1-5, 2-4, 3-8, 6-10, and 7-9, consistent with the disulfide pattern determined for homologous domains of several other proteins. The disulfide linkages of the heparin-binding domain are 11-14, 12-15, and 13-16. This latter set of assignments provides experimental verification of one of the disulfide patterns proposed for netrin (NTR) modules and thereby supports the prediction that the C-terminal heparin-binding domain of sFRP-1 is an NTR-type domain. Interestingly, two subsets of sFRPs appear to have alternate disulfide linkage patterns compared with sFRP-1, one of which involves the loss of a disulfide due to deletion of a single cysteine from the NTR module, whereas the remaining cysteine may pair with a new cysteine introduced in the Fz domain of the protein. Analysis of glycosylation sites showed that sFRP-1 contains a relatively large carbohydrate moiety on Asn(172) (approximately 2.8 kDa), whereas Asn(262), the second potential N-linked glycosylation site, is not modified. No O-linked carbohydrate groups were detected. There was evidence of heterogeneous proteolytic processing at both the N and C termini of the recombinant protein. The predominant N terminus was Ser(31), although minor amounts of the protein with Asp(41) and Phe(50) as the N termini were observed. The major C-terminal processing event was removal of the terminal amino acid (Lys(313)) with only a trace amount of unprocessed protein detected.
Notes: Chong, Jae Min
Uren, Aykut
Rubin, Jeffrey S
Speicher, David W
CA10815/CA/NCI NIH HHS/United States
CA74294/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2002 Feb 15;277(7):5134-44. doi: 10.1074/jbc.M108533200. Epub 2001 Dec 10.
Author Address: Wistar Institute, Philadelphia, Pennsylvania 19104, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 231
Author: Chou, C. F., Smith, A. J. and Omary, M. B.
Year: 1992
Title: Characterization and dynamics of O-linked glycosylation of human cytokeratin 8 and 18
Journal: J Biol Chem
Volume: 267
Issue: 6
Pages: 3901-6
Epub Date: 1992/02/25
Date: Feb 25
Short Title: Characterization and dynamics of O-linked glycosylation of human cytokeratin 8 and 18
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1371281
Keywords: Amino Sugars/metabolism
Carbohydrate Metabolism
Carbohydrate Sequence
Colon/cytology/metabolism
Electrophoresis, Gel, Two-Dimensional
Electrophoresis, Polyacrylamide Gel
Galactosyltransferases/metabolism
Glucosamine/metabolism
Glycosylation
Humans
Hydrogen-Ion Concentration
Hydrolysis
Keratins/*metabolism
Methionine/metabolism
Molecular Sequence Data
Peptide Mapping
Tumor Cells, Cultured
Tunicamycin/pharmacology
Uridine Diphosphate Galactose/metabolism
Abstract: The glycosylation of human cytokeratin (CK) 8 and 18 was studied after metabolic labeling of HT29 colonic cells with [3H]glucosamine. Labeling of CK8/18 was not inhibited by tunicamycin, suggesting that glycosylation was not N-linked. Acid hydrolysis of CK8 and CK18, purified from [3H]glucosamine-labeled cells, generated free glucosamine. In the presence of UDP-[3H]galactose, galactosyltransferase catalyzed the labeling of cytokeratin 8 and 18. beta-Elimination of the [3H]galactose- labeled CK8/18 generated the disaccharide N-acetyllactosaminitol, indicating that cytokeratin 8 and 18 contain single O-linked N-acetylglucosamine residues. Using chemical analysis, the stoichiometry of glycosylation was found to be 1.5 and 2 molecules of N-acetylglucosamine/protein molecule of CK8 and CK18, respectively. Peptide maps of [3H]glucosamine-labeled CK8/18 showed that multiple peptides were labeled with the amino sugar. The biosynthetic and degradation rates of the carbohydrate moiety were faster than the protein core as determined by metabolic radiolabeling or pulse-chase experiments, respectively. Our results show that CK8 and 18 are glycosylated at multiple sites with a single O-linked N-acetylglucosamine. Furthermore, CK8/18 glycosylation is a dynamic process which is likely to have functional relevance.
Notes: Chou, C F
Smith, A J
Omary, M B
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Biol Chem. 1992 Feb 25;267(6):3901-6.
Author Address: Department of Medicine, Palo Alto Veterans Administration Medical Center, California.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 533
Author: Chou, T. Y., Dang, C. V. and Hart, G. W.
Year: 1995
Title: Glycosylation of the c-Myc transactivation domain
Journal: Proc Natl Acad Sci U S A
Volume: 92
Issue: 10
Pages: 4417-21
Epub Date: 1995/05/09
Date: May 9
Short Title: Glycosylation of the c-Myc transactivation domain
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.92.10.4417
PMCID: PMC41955
Accession Number: 7753821
Keywords: Acetylglucosamine/analysis/metabolism
Amidohydrolases
Animals
Binding Sites
CHO Cells
Cattle
Cell Line
Chromatography, Affinity
Cloning, Molecular
Cricetinae
Glycoside Hydrolases
Glycosylation
Helix-Loop-Helix Motifs
Humans
Leucine Zippers
Macromolecular Substances
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Protein Biosynthesis
*Protein Processing, Post-Translational
Proto-Oncogene Proteins c-myc/biosynthesis/isolation & purification/*metabolism
Rats
Recombinant Proteins/biosynthesis/isolation & purification/metabolism
Sequence Deletion
Serine
Spodoptera
Threonine
Transcriptional Activation
Transfection
Abstract: O-linked N-acetylglucosamine (O-GlcNAc) is an abundant and dynamic posttranslational modification composed of a single monosaccharide, GlcNAc, glycosidically composed of a single monosaccharide, GlcNAc, glycosidically linked to the side-chain hydroxyl of serine or threonine residues. Although O-GlcNAc occurs on a myriad of nuclear and cytoplasmic proteins, only a few have thus far been identified. These O-GlcNAc-bearing proteins are also modified by phosphorylation and form reversible multimeric complexes. Here we present evidence for O-GlcNAc glycosylation of the oncoprotein c-Myc, a helix-loop-helix/leucine zipper phosphoprotein that heterodimerizes with Max and participates in the regulation of gene transcription in normal and neoplastic cells. O-GlcNAc modification of c-Myc is shown by three different methods: (i) demonstration of lectin binding to in vitro translated protein using a protein-protein interaction mobility-shift assay; (ii) glycosidase or glycosyltransferase treatment of in vitro translated protein analyzed by lectin affinity chromatography; and (iii) direct characterization of the sugar moieties on purified recombinant protein overexpressed in either insect cells or Chinese hamster ovary cells. Analyses of serial deletion mutants of c-Myc further suggest that the O-GlcNAc site(s) are located within or near the N-terminal transcription activation/malignant transformation domain, a region where mutations of c-Myc that are frequently found in Burkitt and AIDS-related lymphomas cluster.
Notes: 1091-6490
Chou, T Y
Dang, C V
Hart, G W
CA42486/CA/NCI NIH HHS/United States
CA57341/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1995 May 9;92(10):4417-21. doi: 10.1073/pnas.92.10.4417.
Author Address: Biochemistry, Cellular and Molecular Biology Training Program, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 520
Author: Chou, T. Y., Hart, G. W. and Dang, C. V.
Year: 1995
Title: c-Myc is glycosylated at threonine 58, a known phosphorylation site and a mutational hot spot in lymphomas
Journal: J Biol Chem
Volume: 270
Issue: 32
Pages: 18961-5
Epub Date: 1995/08/11
Date: Aug 11
Short Title: c-Myc is glycosylated at threonine 58, a known phosphorylation site and a mutational hot spot in lymphomas
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.270.32.18961
Accession Number: 7642555
Keywords: Amino Acid Sequence
Animals
Glycosylation
Lymphoma/*genetics
Molecular Sequence Data
*Mutation
Phosphorylation
Proto-Oncogene Proteins c-myc/*metabolism
Recombinant Proteins/metabolism
Spodoptera
Threonine/*metabolism
Abstract: c-Myc is a helix-loop leucine zipper phosphoprotein that heterodimerizes with Max and regulates gene transcription in cell proliferation, cell differentiation, and programmed cell death. Previously, we demonstrated that c-Myc is modified by O-linked N-acetylglucosamine (O-GlcNAc) within or nearby the N-terminal transcriptional activation domain (Chou, T.-Y., Dang, C.V., and Hart, G.W. (1995) Proc. Natl. Acad. Sci. U.S.A. 92, 4417-4421). In this paper, we identified the O-GlcNAc attachment site(s) on c-Myc. c-Myc purified from sf9 insect cells was trypsinized, and its GlcNAc moieties were enzymically labeled with [3H]galactose. The [3H]galactose-labeled glycopeptides were isolated by reverse phase high performance liquid chromatography and then subjected to gas-phase sequencing, manual Edman degradation, and laser desorption/ionization mass spectrometry. These analyses show that threonine 58, an in vivo phosphorylation site in the transactivation domain, is the major O-GlcNAc glycosylation site of c-Myc. Mutation of threonine 58, frequently found in retroviral v-Myc proteins and in human Burkitt and AIDS-related lymphomas, is associated with enhanced transforming activity and tumorigenicity. The reciprocal glycosylation and phosphorylation at this biologically significant amino acid residue may play an important role in the regulation of the functions of c-Myc.
Notes: Chou, T Y
Hart, G W
Dang, C V
CA42486/CA/NCI NIH HHS/United States
CA57341/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1995 Aug 11;270(32):18961-5. doi: 10.1074/jbc.270.32.18961.
Author Address: Biochemistry, Cellular, and Molecular Biology Training Program, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 986
Author: Christensen, B., Nielsen, M. S., Haselmann, K. F., Petersen, T. E. and Sørensen, E. S.
Year: 2005
Title: Post-translationally modified residues of native human osteopontin are located in clusters: identification of 36 phosphorylation and five O-glycosylation sites and their biological implications
Journal: Biochem J
Volume: 390
Issue: Pt 1
Pages: 285-92
Epub Date: 2005/05/05
Date: Aug 15
Short Title: Post-translationally modified residues of native human osteopontin are located in clusters: identification of 36 phosphorylation and five O-glycosylation sites and their biological implications
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20050341
PMCID: PMC1184582
Accession Number: 15869464
Keywords: Amino Acid Sequence
*Glycosylation
Humans
Molecular Sequence Data
Osteopontin
Oxidation-Reduction
Peptides/chemistry
*Phosphorylation
*Protein Processing, Post-Translational
Sequence Alignment
Sequence Homology, Amino Acid
Sialoglycoproteins/*chemistry/physiology
Abstract: OPN (osteopontin) is an integrin-binding highly phosphorylated glycoprotein, recognized as a key molecule in a multitude of biological processes such as bone mineralization, cancer metastasis, cell-mediated immune response, inflammation and cell survival. A significant regulation of OPN function is mediated through PTM (post-translational modification). Using a combination of Edman degradation and MS analyses, we have characterized the complete phosphorylation and glycosylation pattern of native human OPN. A total of 36 phosphoresidues have been localized in the sequence of OPN. There are 29 phosphorylations (Ser8, Ser10, Ser11, Ser46, Ser47, Thr50, Ser60, Ser62, Ser65, Ser83, Ser86, Ser89, Ser92, Ser104, Ser110, Ser113, Thr169, Ser179, Ser208, Ser218, Ser238, Ser247, Ser254, Ser259, Ser264, Ser275, Ser287, Ser292 and Ser294) located in the target sequence of MGCK (mammary gland casein kinase) also known as the Golgi kinase (S/T-X-E/S(P)/D). Six phosphorylations (Ser101, Ser107, Ser175, Ser199, Ser212 and Ser251) are located in the target sequence of CKII (casein kinase II) [S-X-X-E/S(P)/D] and a single phosphorylation, Ser203, is not positioned in the motif of either MGCK or CKII. The 36 phosphoresidues represent the maximal degree of modification since variability at many sites was seen. Five threonine residues are O-glycosylated (Thr118, Thr122, Thr127, Thr131 and Thr136) and two potential sites for N-glycosylation (Asn63 and Asn90) are not occupied in human milk OPN. The phosphorylations are arranged in clusters of three to five phosphoresidues and the regions containing the glycosylations and the RGD (Arg-Gly-Asp) integrin-binding sequence are devoid of phosphorylations. Knowledge about the positions and nature of PTMs in OPN will allow a rational experimental design of functional studies aimed at understanding the structural and functional interdependences in diverse biological processes in which OPN is a key molecule.
Notes: 1470-8728
Christensen, Brian
Nielsen, Mette S
Haselmann, Kim F
Petersen, Torben E
Sørensen, Esben S
Journal Article
Biochem J. 2005 Aug 15;390(Pt 1):285-92. doi: 10.1042/BJ20050341.
Author Address: Protein Chemistry Laboratory, Department of Molecular Biology, Science Park, University of Aarhus, Gustav Wieds Vej 10C, DK-8000 Aarhus C, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 898
Author: Chruszcz, M., Laidler, P., Monkiewicz, M., Ortlund, E., Lebioda, L. and Lewinski, K.
Year: 2003
Title: Crystal structure of a covalent intermediate of endogenous human arylsulfatase A
Journal: J Inorg Biochem
Volume: 96
Issue: 2-3
Pages: 386-92
Epub Date: 2003/07/31
Date: Aug 1
Short Title: Crystal structure of a covalent intermediate of endogenous human arylsulfatase A
Alternate Journal: Journal of inorganic biochemistry
ISSN: 0162-0134 (Print)
0162-0134
DOI: 10.1016/s0162-0134(03)00176-4
Accession Number: 12888274
Keywords: Binding Sites
Calcium/analysis/chemistry
Cerebroside-Sulfatase/*chemistry
Crystallization
Humans
Magnesium/analysis/chemistry
Molecular Structure
Phosphates/chemistry
Placenta/enzymology
Protein Conformation
Abstract: The structures of human arylsulfatase A crystals soaked in solutions containing 4-methylumbelliferyl phosphate and O-phospho-DL-tyrosine have been determined at 2.7- and 3.2-A resolution, respectively. The formylglycine in position 69, a residue crucial for catalytic activity, was unambiguously identified in both structures as forming a covalent bond to the phosphate moiety. A hydroxyl group is present at the Cbeta of residue 69 and the formation of one out of two possible stereomeric forms is strongly favoured. The structures confirm the importance of the gem-diol intermediate in the arylsulfatase's catalytic mechanism. The presence of an apparently stable covalent bond is consistent with the weak phosphatase activity observed for human arylsulfatase A. The structures of the complexes suggest that phosphate ions and phosphate esters inhibit arylsulfatase in non-covalent and covalent modes, respectively. The metal ion present in the active site of arylsulfatase A isolated from human placenta is Ca(2+) and not Mg(2+) as was found in the structure of the recombinant enzyme.
Notes: Chruszcz, Maksymilian
Laidler, Piotr
Monkiewicz, Maciej
Ortlund, Eric
Lebioda, Lukasz
Lewinski, Krzysztof
Journal Article
Research Support, Non-U.S. Gov't
United States
J Inorg Biochem. 2003 Aug 1;96(2-3):386-92. doi: 10.1016/s0162-0134(03)00176-4.
Author Address: Faculty of Chemistry, Jagiellonian University, ul. Ingardena 3, 30-060, Krakow, Poland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1624
Author: Chu, C. S., Lo, P. W., Yeh, Y. H., Hsu, P. H., Peng, S. H., Teng, Y. C., Kang, M. L., Wong, C. H. and Juan, L. J.
Year: 2014
Title: O-GlcNAcylation regulates EZH2 protein stability and function
Journal: Proc Natl Acad Sci U S A
Volume: 111
Issue: 4
Pages: 1355-60
Epub Date: 2014/01/30
Date: Jan 28
Short Title: O-GlcNAcylation regulates EZH2 protein stability and function
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1323226111
PMCID: PMC3910655
Accession Number: 24474760
Keywords: Acetylglucosamine/*metabolism
DNA Methylation
Down-Regulation
Enhancer of Zeste Homolog 2 Protein
Gene Knockdown Techniques
Genes, Tumor Suppressor
Humans
N-Acetylglucosaminyltransferases/genetics/metabolism
Polycomb Repressive Complex 2/*metabolism
Protein Stability
Abstract: O-linked N-acetylglucosamine (GlcNAc) transferase (OGT) is the only known enzyme that catalyzes the O-GlcNAcylation of proteins at the Ser or Thr side chain hydroxyl group. OGT participates in transcriptional and epigenetic regulation, and dysregulation of OGT has been implicated in diseases such as cancer. However, the underlying mechanism is largely unknown. Here we show that OGT is required for the trimethylation of histone 3 at K27 to form the product H3K27me3, a process catalyzed by the histone methyltransferase enhancer of zeste homolog 2 (EZH2) in the polycomb repressive complex 2 (PRC2). H3K27me3 is one of the most important histone modifications to mark the transcriptionally silenced chromatin. We found that the level of H3K27me3, but not other H3 methylation products, was greatly reduced upon OGT depletion. OGT knockdown specifically down-regulated the protein stability of EZH2, without altering the levels of H3K27 demethylases UTX and JMJD3, and disrupted the integrity of the PRC2 complex. Furthermore, the interaction of OGT and EZH2/PRC2 was detected by coimmunoprecipitation and cosedimentation experiments. Importantly, we identified that serine 75 is the site for EZH2 O-GlcNAcylation, and the EZH2 mutant S75A exhibited reduction in stability. Finally, microarray and ChIP analysis have characterized a specific subset of potential tumor suppressor genes subject to repression via the OGT-EZH2 axis. Together these results indicate that OGT-mediated O-GlcNAcylation at S75 stabilizes EZH2 and hence facilitates the formation of H3K27me3. The study not only uncovers a functional posttranslational modification of EZH2 but also reveals a unique epigenetic role of OGT in regulating histone methylation.
Notes: 1091-6490
Chu, Chi-Shuen
Lo, Pei-Wen
Yeh, Yi-Hsien
Hsu, Pang-Hung
Peng, Shih-Huan
Teng, Yu-Ching
Kang, Ming-Lun
Wong, Chi-Huey
Juan, Li-Jung
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2014 Jan 28;111(4):1355-60. doi: 10.1073/pnas.1323226111. Epub 2014 Jan 13.
Author Address: Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1862
Author: Chun, Y. S., Kwon, O. H. and Chung, S.
Year: 2017
Title: O-GlcNAcylation of amyloid-β precursor protein at threonine 576 residue regulates trafficking and processing
Journal: Biochem Biophys Res Commun
Volume: 490
Issue: 2
Pages: 486-491
Epub Date: 2017/06/19
Date: Aug 19
Short Title: O-GlcNAcylation of amyloid-β precursor protein at threonine 576 residue regulates trafficking and processing
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2017.06.067
Accession Number: 28624365
Keywords: Acetylglucosamine/*metabolism
Acylation
Alzheimer Disease/metabolism
Amyloid beta-Protein Precursor/analysis/genetics/*metabolism
Cell Membrane/metabolism
Glycosylation
HeLa Cells
Humans
Point Mutation
Protein Processing, Post-Translational
Protein Transport
Threonine/analysis/genetics/metabolism
trans-Golgi Network/metabolism
*Alzheimer's disease
*Amyloid
*Amyloid-β protein precursor
*O-GlcNAcylation
Abstract: The pathological hallmark of Alzheimer's disease (AD) is associated with the accumulation of amyloid-β (Aβ) derived from proteolytic processing of amyloid-β precursor protein (APP). APP undergoes post-translational modification including N- and O-glycosylation. O-GlcNAcylation is a novel type of O-glycosylation, mediated by O-GlcNAc transferase attaching O-β-N-acetylglucosamine (O-GlcNAc) to serine/threonine residues of the target proteins. O-GlcNAc is removed by O-GlcNAcase. We have previously reported that increasing O-GlcNAcylated APP using the O-GlcNAcase inhibitor, PUGNAc, increases its trafficking rate to the plasma membrane and decreases its endocytosis rate, resulting in decreased Aβ production. However, O-GlcNAc modification sites in APP are unknown. In this study, we mutated three predicted O-GlcNAc modification threonine residues of APP into alanines (T291A, T292A, and T576A) and expressed them in HeLa cells. These APP mutants showed reduced O-GlcNAcylation levels, indicating that these sites were endogenously O-GlcNAcylated. Thr 576 was the major O-GlcNAcylation site when cell was treated with PUGNAc. We also showed that the effects of PUGNAc on APP trafficking to the plasma membrane and Aβ production were prevented in the T576A mutant. These results implicate Thr 576 as the major O-GlcNAcylation site in APP and indicate that O-GlcNAcylation of this residue regulates its trafficking and processing. Thus, specific O-GlcNAcylation of APP at Thr 576 may be a novel and promising drug target for AD therapeutics.
Notes: 1090-2104
Chun, Yoon Sun
Kwon, Oh-Hoon
Chung, Sungkwon
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2017 Aug 19;490(2):486-491. doi: 10.1016/j.bbrc.2017.06.067. Epub 2017 Jun 15.
Author Address: Department of Physiology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746, South Korea.
Department of Physiology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746, South Korea. Electronic address: schung@skku.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 405
Author: Chung, D. W., Fujikawa, K., McMullen, B. A. and Davie, E. W.
Year: 1986
Title: Human plasma prekallikrein, a zymogen to a serine protease that contains four tandem repeats
Journal: Biochemistry
Volume: 25
Issue: 9
Pages: 2410-7
Epub Date: 1986/05/06
Date: May 6
Short Title: Human plasma prekallikrein, a zymogen to a serine protease that contains four tandem repeats
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00357a017
Accession Number: 3521732
Keywords: Amino Acid Sequence
Antibodies/isolation & purification
Base Sequence
Cloning, Molecular
DNA/analysis
Endopeptidases/*genetics
Humans
Kallikreins/*genetics
Peptide Fragments/analysis
Prekallikrein/blood/*genetics
Repetitive Sequences, Nucleic Acid
Serine Endopeptidases
Abstract: The amino acid sequence of human plasma prekallikrein was determined by a combination of automated Edman degradation and cDNA sequencing techniques. Human plasma prekallikrein was fragmented with cyanogen bromide, and 13 homogeneous peptides were isolated and sequenced. Cyanogen bromide peptides containing carbohydrate were further digested with trypsin, and the peptides containing carbohydrate were isolated and sequenced. Five asparagine-linked carbohydrate attachment sites were identified. The sequence determined by Edman degradation was aligned with the amino acid sequence predicted from cDNAs isolated from a lambda gt11 expression library. This library contained cDNA inserts prepared from human liver poly(A) RNA. Analysis of the cDNA indicated that human plasma prekallikrein is synthesized as a precursor with a signal peptide of 19 amino acids. The mature form of the protein that circulates in blood is a single-chain polypeptide of 619 amino acids. Plasma prekallikrein is converted to plasma kallikrein by factor XIIa by the cleavage of an internal Arg-Ile bond. Plasma kallikrein is composed of a heavy chain (371 amino acids) and a light chain (248 amino acids), and these 2 chains are held together by a disulfide bond. The heavy chain of plasma kallikrein originates from the amino-terminal end of the zymogen and is composed of 4 tandem repeats that are 90 or 91 amino acid residues in length. These repeat sequences are also homologous to those in human factor XI. The light chain of plasma kallikrein contains the catalytic portion of the enzyme and is homologous to the trypsin family of serine proteases.
Notes: Chung, D W
Fujikawa, K
McMullen, B A
Davie, E W
HL 16919/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1986 May 6;25(9):2410-7. doi: 10.1021/bi00357a017.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 421
Author: Chung, L. P., Bentley, D. R. and Reid, K. B.
Year: 1985
Title: Molecular cloning and characterization of the cDNA coding for C4b-binding protein, a regulatory protein of the classical pathway of the human complement system
Journal: Biochem J
Volume: 230
Issue: 1
Pages: 133-41
Epub Date: 1985/08/15
Date: Aug 15
Short Title: Molecular cloning and characterization of the cDNA coding for C4b-binding protein, a regulatory protein of the classical pathway of the human complement system
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2300133
PMCID: PMC1152596
Accession Number: 3840370
Keywords: Amino Acid Sequence
Base Sequence
Carrier Proteins/*genetics
*Cloning, Molecular
*Complement Inactivator Proteins
Complement Pathway, Classical
DNA/*genetics
Electrophoresis, Agar Gel
Genetic Code
*Glycoproteins
Humans
Oligonucleotides/chemical synthesis
Abstract: By using synthetic oligonucleotides as probes, plasmid clones containing portions of cDNA coding for human C4b-binding protein were isolated from a liver cDNA library. The entire amino acid sequence of the C4b-binding protein can be predicted from this study of the cloned cDNA when allied to a previous sequence study at the protein level [Chung, Gagnon & Reid (1985) Mol. Immunol. 22, 427-435], in which over 55% of the amino acid sequence, including the N-terminal 62 residues, was obtained. The plasmid clones isolated allowed the unambiguous determination of 1717 nucleotides of cDNA sequence between the codon for the 32nd amino acid in the sequence of C4b-binding protein and the 164th nucleotide in the 3' non-translated region. The sequence studies show that the secreted form of C4b-binding protein, found in plasma, is composed of chains of apparent Mr 70 000 that contains 549 amino acid residues. Examination of the protein and cDNA sequence results show that there are at least two polymorphic sites in the molecule. One is at position 44, which can be glutamine or threonine, and the other is at position 309, which can be tyrosine or histidine. Northern-blot analysis indicated that the mRNA for C4b-binding protein is approx. 2.5 kilobases long. The N-terminal 491 amino acids of C4b-binding protein can be divided into eight internal homologous regions, each approx. 60 amino acids long, which can be aligned by the presence in each region of four half-cystine, one tryptophan and several other conserved residues. These regions in C4b-binding protein are homologous with the three internal-homology regions that have been reported to be present within the Ba region of the complement enzyme factor B and also to the internal-homology regions found in the non-complement beta 2-glycoprotein I.
Notes: 1470-8728
Chung, L P
Bentley, D R
Reid, K B
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1985 Aug 15;230(1):133-41. doi: 10.1042/bj2300133.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 726
Author: Cihlar, T., Lin, D. C., Pritchard, J. B., Fuller, M. D., Mendel, D. B. and Sweet, D. H.
Year: 1999
Title: The antiviral nucleotide analogs cidofovir and adefovir are novel substrates for human and rat renal organic anion transporter 1
Journal: Mol Pharmacol
Volume: 56
Issue: 3
Pages: 570-80
Epub Date: 1999/08/27
Date: Sep
Short Title: The antiviral nucleotide analogs cidofovir and adefovir are novel substrates for human and rat renal organic anion transporter 1
Alternate Journal: Molecular pharmacology
ISSN: 0026-895X (Print)
0026-895x
DOI: 10.1124/mol.56.3.570
Accession Number: 10462545
Keywords: Adenine/*analogs & derivatives/metabolism/pharmacology
Amino Acid Sequence
Animals
Anion Transport Proteins
Antiviral Agents/*metabolism/pharmacology
Base Sequence
Biological Transport
Carrier Proteins/*genetics/metabolism
Cidofovir
Cloning, Molecular
Cytosine/*analogs & derivatives/metabolism/pharmacology
DNA, Complementary/analysis
Humans
Immunoblotting
Kidney/*drug effects/metabolism
Molecular Sequence Data
Nucleosides/metabolism
*Organophosphonates
Organophosphorus Compounds/*metabolism/pharmacology
Rats
Abstract: Nephrotoxicity is the dose-limiting clinical adverse effect of cidofovir and adefovir, two potent antiviral therapeutics. Because renal uptake likely plays a role in the etiology of cidofovir- and adefovir-associated nephrotoxicity, we attempted to identify a renal transporter capable of interacting with these therapeutics. A cDNA clone was isolated from a human renal library and designated human organic anion transporter 1 (hOAT1). Northern analysis detected a specific 2.5-kilobase pair hOAT1 transcript only in human kidney. However, reverse transcription-polymerase chain reaction revealed hOAT1 expression in human brain and skeletal muscle, as well. Immunoblot analysis of human kidney cortex demonstrated that hOAT1 is an 80- to 90-kilodalton heterogeneous protein modified by abundant N-glycosylation. Xenopus laevis oocytes expressing hOAT1 supported probenecid-sensitive uptake of [(3)H]p-aminohippurate (K(m) = 4 microM), which was trans-stimulated in oocytes preloaded with glutarate. Importantly, both hOAT1 and rat renal organic anion transporter 1 (rROAT1) mediated saturable, probenecid-sensitive uptake of cidofovir, adefovir, and other nucleoside phosphonate antivirals. The affinity of hOAT1 toward cidofovir and adefovir (K(m) = 46 and 30 microM, respectively) was 5- to 9-fold higher compared with rROAT1 (K(m) = 238 and 270 microM, respectively). These data indicate that hOAT1 may significantly contribute to the accumulation of cidofovir and adefovir in renal proximal tubules and, thus, play an active role in the mechanism of nephrotoxicity associated with these antiviral therapeutics.
Notes: Cihlar, T
Lin, D C
Pritchard, J B
Fuller, M D
Mendel, D B
Sweet, D H
Journal Article
United States
Mol Pharmacol. 1999 Sep;56(3):570-80. doi: 10.1124/mol.56.3.570.
Author Address: Gilead Sciences, Foster City, California, USA. tomas_cihlar@gilead.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1263
Author: Clark, N. E. and Garman, S. C.
Year: 2009
Title: The 1.9 a structure of human alpha-N-acetylgalactosaminidase: The molecular basis of Schindler and Kanzaki diseases
Journal: J Mol Biol
Volume: 393
Issue: 2
Pages: 435-47
Epub Date: 2009/08/18
Date: Oct 23
Short Title: The 1.9 a structure of human alpha-N-acetylgalactosaminidase: The molecular basis of Schindler and Kanzaki diseases
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2009.08.021
PMCID: PMC2771859
NIHMSID: NIHMS139243
Accession Number: 19683538
Keywords: Amino Acid Sequence
Animals
Cell Line
Crystallography, X-Ray
Humans
Insecta
Lysosomal Storage Diseases/*enzymology
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Tertiary
Sequence Homology, Amino Acid
alpha-N-Acetylgalactosaminidase/*chemistry/genetics/metabolism
Abstract: alpha-N-acetylgalactosaminidase (alpha-NAGAL; E.C. 3.2.1.49) is a lysosomal exoglycosidase that cleaves terminal alpha-N-acetylgalactosamine residues from glycopeptides and glycolipids. In humans, a deficiency of alpha-NAGAL activity results in the lysosomal storage disorders Schindler disease and Kanzaki disease. To better understand the molecular defects in the diseases, we determined the crystal structure of human alpha-NAGAL after expressing wild-type and glycosylation-deficient glycoproteins in recombinant insect cell expression systems. We measured the enzymatic parameters of our purified wild-type and mutant enzymes, establishing their enzymatic equivalence. To investigate the binding specificity and catalytic mechanism of the human alpha-NAGAL enzyme, we determined three crystallographic complexes with different catalytic products bound in the active site of the enzyme. To better understand how individual defects in the alpha-NAGAL glycoprotein lead to Schindler disease, we analyzed the effect of disease-causing mutations on the three-dimensional structure.
Notes: 1089-8638
Clark, Nathaniel E
Garman, Scott C
R01 DK076877/DK/NIDDK NIH HHS/United States
R01 DK076877-03/DK/NIDDK NIH HHS/United States
Y01 GM000080/GM/NIGMS NIH HHS/United States
R01 DK76877/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Mol Biol. 2009 Oct 23;393(2):435-47. doi: 10.1016/j.jmb.2009.08.021. Epub 2009 Aug 14.
Author Address: Department of Biochemistry and Molecular Biology, University of Massachusetts, Amherst, 01003, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1202
Author: Clarke, A. J., Hurtado-Guerrero, R., Pathak, S., Schüttelkopf, A. W., Borodkin, V., Shepherd, S. M., Ibrahim, A. F. and van Aalten, D. M.
Year: 2008
Title: Structural insights into mechanism and specificity of O-GlcNAc transferase
Journal: Embo j
Volume: 27
Issue: 20
Pages: 2780-8
Epub Date: 2008/09/27
Date: Oct 22
Short Title: Structural insights into mechanism and specificity of O-GlcNAc transferase
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2008.186
PMCID: PMC2556091
Accession Number: 18818698
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Humans
Models, Biological
Molecular Conformation
Molecular Sequence Data
N-Acetylglucosaminyltransferases/*chemistry/metabolism
Phosphorylation
Protein Binding
Protein Processing, Post-Translational
Sequence Homology, Amino Acid
Signal Transduction
Substrate Specificity
Xenopus
Abstract: Post-translational modification of protein serines/threonines with N-acetylglucosamine (O-GlcNAc) is dynamic, inducible and abundant, regulating many cellular processes by interfering with protein phosphorylation. O-GlcNAcylation is regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase, both encoded by single, essential, genes in metazoan genomes. It is not understood how OGT recognises its sugar nucleotide donor and performs O-GlcNAc transfer onto proteins/peptides, and how the enzyme recognises specific cellular protein substrates. Here, we show, by X-ray crystallography and mutagenesis, that OGT adopts the (metal-independent) GT-B fold and binds a UDP-GlcNAc analogue at the bottom of a highly conserved putative peptide-binding groove, covered by a mobile loop. Strikingly, the tetratricopeptide repeats (TPRs) tightly interact with the active site to form a continuous 120 A putative interaction surface, whereas the previously predicted phosphatidylinositide-binding site locates to the opposite end of the catalytic domain. On the basis of the structure, we identify truncation/point mutants of the TPRs that have differential effects on activity towards proteins/peptides, giving first insights into how OGT may recognise its substrates.
Notes: 1460-2075
Clarke, Andrew J
Hurtado-Guerrero, Ramon
Pathak, Shalini
Schüttelkopf, Alexander W
Borodkin, Vladimir
Shepherd, Sharon M
Ibrahim, Adel F M
van Aalten, Daan M F
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2008 Oct 22;27(20):2780-8. doi: 10.1038/emboj.2008.186. Epub 2008 Sep 25.
Author Address: Division of Biological Chemistry & Drug Discovery, College of Life Sciences, University of Dundee, Dundee, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1256
Author: Clayton, A., Siebold, C., Gilbert, R. J., Sutton, G. C., Harlos, K., McIlhinney, R. A., Jones, E. Y. and Aricescu, A. R.
Year: 2009
Title: Crystal structure of the GluR2 amino-terminal domain provides insights into the architecture and assembly of ionotropic glutamate receptors
Journal: J Mol Biol
Volume: 392
Issue: 5
Pages: 1125-32
Epub Date: 2009/08/05
Date: Oct 9
Short Title: Crystal structure of the GluR2 amino-terminal domain provides insights into the architecture and assembly of ionotropic glutamate receptors
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2009.07.082
Accession Number: 19651138
Keywords: Crystallography, X-Ray
Humans
Models, Molecular
Protein Multimerization
Protein Structure, Quaternary
Protein Structure, Tertiary
Receptors, AMPA/*chemistry
Abstract: Ionotropic glutamate receptors are functionally diverse but have a common architecture, including the 400-residue amino-terminal domain (ATD). We report a 1.8-A resolution crystal structure of human GluR2-ATD. This dimeric structure provides a mechanism for how the ATDs can drive receptor assembly and subtype-restricted composition. Lattice contacts in a 4.1-A resolution crystal form reveal a tetrameric (dimer-dimer) arrangement consistent with previous cellular and cryo-electron microscopic data for full-length AMPA receptors.
Notes: 1089-8638
Clayton, Amber
Siebold, Christian
Gilbert, Robert J C
Sutton, Geoffrey C
Harlos, Karl
McIlhinney, R A Jeffrey
Jones, E Yvonne
Aricescu, A Radu
G0900084/Medical Research Council/United Kingdom
G0500367/Medical Research Council/United Kingdom
G0700232/Medical Research Council/United Kingdom
MC_U138197108/Medical Research Council/United Kingdom
G0500365/Medical Research Council/United Kingdom
Wellcome Trust/United Kingdom
G9900061/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2009 Oct 9;392(5):1125-32. doi: 10.1016/j.jmb.2009.07.082. Epub 2009 Aug 3.
Author Address: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 763
Author: Coddeville, B., Regoeczi, E., Strecker, G., Plancke, Y. and Spik, G.
Year: 2000
Title: Structural analysis of trisialylated biantennary glycans isolated from mouse serum transferrin. Characterization of the sequence Neu5Gc(alpha 2-3)Gal(beta 1-3)[Neu5Gc(alpha 2-6)]GlcNAc(beta 1-2)Man
Journal: Biochim Biophys Acta
Volume: 1475
Issue: 3
Pages: 321-8
Epub Date: 2000/09/27
Date: Jul 26
Short Title: Structural analysis of trisialylated biantennary glycans isolated from mouse serum transferrin. Characterization of the sequence Neu5Gc(alpha 2-3)Gal(beta 1-3)[Neu5Gc(alpha 2-6)]GlcNAc(beta 1-2)Man
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/s0304-4165(00)00083-0
Accession Number: 10913832
Keywords: Animals
Carbohydrate Sequence
Chromatography, Ion Exchange
Disaccharides/chemistry
Female
Glycopeptides/chemistry/isolation & purification
Magnetic Resonance Spectroscopy
Male
Mice
Molecular Sequence Data
Molecular Structure
N-Acetylneuraminic Acid
Neuraminic Acids/analysis
Polysaccharides/*chemistry/isolation & purification
Transferrin/*chemistry/isolation & purification
Abstract: Five variants of mouse serum transferrin (mTf, designated mTf-I to mTf-V) with respect to carbohydrate composition have been isolated by DEAE-cellulose chromatography in the following relative percentages: mTf-I: 0.55; mTf-II: 0.79; mTf-III: 71.80; mTf-VI: 21. 90 and mTf-V: 4.96. The primary structures of the major glycans from mTf-III and mTf-IV were determined by methylation analysis and 1H-nuclear magnetic resonance (NMR) spectroscopy. All glycans possessed a common trimannosyl-N,N'-diacetylchitobiose core. From the glycovariant mTf-III two isomers of a conventional biantennary N-acetyllactosamine type were isolated, in which two N-glycolylneuraminic acid (Neu5Gc) residues are linked to galactose either by a (alpha 2-6) or (alpha 2-3) linkage. A subpopulation of this glycovariant contains a fucose residue (alpha 1-6)-linked to GlcNAc-1. The structure of the major glycan found in variant mTf-IV contained an additional Neu5Gc and possessed the following new type of linkage: Neu5Gc(alpha 2-3)Gal(beta 1-3)[Neu5Gc(alpha 2-6)]GlcNAc(beta 1-2 )Man(alpha 1-3). In addition to this glycan, a minor compound contained the same antennae linked to Man(alpha 1-6). In fraction mTf-V, which was found to be very heterogeneous by (1)H NMR analysis, carbohydrate composition and methylation analysis suggested the presence of tri'-antennary glycans sialylated by Neu5Gc alpha-2,6- and alpha-2, 3-linked to the terminal galactose residues. In summary, mTf glycans differed from those of other analyzed mammalian transferrins by the presence of Neu5Gc and by a Neu5Gc(alpha 2-6)GlcNAc linkage in trisialylated biantennary structures, reflecting in mouse liver, a high activity of CMP-Neu5Ac hydroxylase and (alpha 2-6)GlcNAc sialyltransferase.
Notes: Coddeville, B
Regoeczi, E
Strecker, G
Plancke, Y
Spik, G
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2000 Jul 26;1475(3):321-8. doi: 10.1016/s0304-4165(00)00083-0.
Author Address: Laboratoire de Chimie Biologique de l'Université des Sciences et Technologies de Lille, UMR CNRS No. 8576, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 745
Author: Cointe, D., Béliard, R., Jorieux, S., Leroy, Y., Glacet, A., Verbert, A., Bourel, D. and Chirat, F.
Year: 2000
Title: Unusual N-glycosylation of a recombinant human erythropoietin expressed in a human lymphoblastoid cell line does not alter its biological properties
Journal: Glycobiology
Volume: 10
Issue: 5
Pages: 511-9
Epub Date: 2000/04/15
Date: May
Short Title: Unusual N-glycosylation of a recombinant human erythropoietin expressed in a human lymphoblastoid cell line does not alter its biological properties
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/10.5.511
Accession Number: 10764840
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cell Line
Chemical Fractionation
Cricetinae
Erythropoietin/*chemistry/genetics/*metabolism
Glycopeptides/analysis/isolation & purification
Glycosylation
Humans
Lymphocytes/*metabolism
Methylation
Molecular Sequence Data
Monosaccharides/analysis
N-Acetylneuraminic Acid/metabolism
Oligosaccharides/analysis
Pharmacokinetics
Recombinant Proteins
Sialyl Lewis X Antigen
Structure-Activity Relationship
Abstract: Erythropoietin (Epo) is a 166 amino acids protein containing three N-glycosylation sites (Asn-24, Asn-38, and Asn-83) and 1 O- glycosylation site (Ser-126) and involved in the regulation of the level of red blood cells. Today, only one recombinant human Epo (rHuEpo), produced in CHO cell line, is extensively used in therapy to cure severe anemia. The structure of the glycan chains of this rHuEpo slightly differ of those of the urinary human Epo (uHuEpo), considered as the natural Epo molecule. In an attempt to produce a rHuEpo as close as possible to the uHuEpo, Epo gene was expressed in a human lymphoblastoid cell line, named RPMI 1788. In order to fully characterize the Epo-RPMI, structural characterizations of the protein skeleton as well as glycan chains were undergone. As expected, the amino acid sequence of the Epo-RPMI conformed to that of uHuEpo. Surprisingly, the structure of some N-glycan chains, as mainly determined by ESI-MS, revealed some unusual characteristics. Thus, 80% of N-glycans possess a bisecting GlcNAc residue, 25% bear a second fucose residue which is present, in a large part, in a sialyl Le(x)motif, and 13% contain more than three LacNAc repeats (up to five per molecule). Despite these unusual structural characteristics, the data concerning the in vitro and in vivo biological activities were not impaired when compared to Epo-CHO and uHuEpo.
Notes: Cointe, D
Béliard, R
Jorieux, S
Leroy, Y
Glacet, A
Verbert, A
Bourel, D
Chirat, F
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2000 May;10(5):511-9. doi: 10.1093/glycob/10.5.511.
Author Address: Laboratoire de Glycobiologie Structurale et Fonctionnelle, Unité Mixte de Recherche du CNRS No 8576, Université des Sciences et Technologies de Lille, F-59655 Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 705
Author: Cole, A. R., Hall, N. E., Treutlein, H. R., Eddes, J. S., Reid, G. E., Moritz, R. L. and Simpson, R. J.
Year: 1999
Title: Disulfide bond structure and N-glycosylation sites of the extracellular domain of the human interleukin-6 receptor
Journal: J Biol Chem
Volume: 274
Issue: 11
Pages: 7207-15
Epub Date: 1999/03/06
Date: Mar 12
Short Title: Disulfide bond structure and N-glycosylation sites of the extracellular domain of the human interleukin-6 receptor
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.274.11.7207
Accession Number: 10066782
Keywords: Amino Acid Sequence
Animals
Antigens, CD/metabolism
CHO Cells
Cricetinae
Cytokine Receptor gp130
Disulfides/chemistry/*metabolism
Glycosylation
Humans
Membrane Glycoproteins/metabolism
Models, Molecular
Molecular Sequence Data
Molecular Structure
Peptide Mapping
Receptors, Interleukin-6/chemistry/*metabolism
Recombinant Proteins/chemistry/metabolism
Sequence Homology, Amino Acid
Signal Transduction
Trypsin/metabolism
Abstract: The high affinity interleukin-6 (IL-6) receptor is a hexameric complex consisting of two molecules each of IL-6, IL-6 receptor (IL-6R), and the high affinity converter and signaling molecule, gp130. The extracellular "soluble" part of the IL-6R (sIL-6R) consists of three domains: an amino-terminal Ig-like domain and two fibronectin-type III (FN III) domains. The two FN III domains comprise the cytokine-binding domain defined by a set of 4 conserved cysteine residues and a WSXWS sequence motif. Here, we have determined the disulfide structure of the human sIL-6R by peptide mapping in the absence and presence of reducing agent. Mass spectrometric analysis of these peptides revealed four disulfide bonds and two free cysteines. The disulfides Cys102-Cys113 and Cys146-Cys157 are consistent with known cytokine-binding domain motifs, and Cys28-Cys77 with known Ig superfamily domains. An unusual cysteine connectivity between Cys6-Cys174, which links the Ig-like and NH2-terminal FN III domains causing them to fold back onto each other, has not previously been observed among cytokine receptors. The two free cysteines (Cys192 and Cys258) were detected as cysteinyl-cysteines, although a small proportion of Cys258 was reactive with the alkylating agent 4-vinylpyridine. Of the four potential N-glycosylation sites, carbohydrate moieties were identified on Asn36, Asn74, and Asn202, but not on Asn226.
Notes: Cole, A R
Hall, N E
Treutlein, H R
Eddes, J S
Reid, G E
Moritz, R L
Simpson, R J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1999 Mar 12;274(11):7207-15. doi: 10.1074/jbc.274.11.7207.
Author Address: Joint Protein Structure Laboratory, Ludwig Institute for Cancer Research (Melbourne Tumour Biology Branch) and The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3050, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 719
Author: Cole, R. N. and Hart, G. W.
Year: 1999
Title: Glycosylation sites flank phosphorylation sites on synapsin I: O-linked N-acetylglucosamine residues are localized within domains mediating synapsin I interactions
Journal: J Neurochem
Volume: 73
Issue: 1
Pages: 418-28
Epub Date: 1999/07/01
Date: Jul
Short Title: Glycosylation sites flank phosphorylation sites on synapsin I: O-linked N-acetylglucosamine residues are localized within domains mediating synapsin I interactions
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042 (Print)
0022-3042
DOI: 10.1046/j.1471-4159.1999.0730418.x
Accession Number: 10386995
Keywords: Acetylglucosamine/*analysis
Amino Acid Sequence
Animals
Binding Sites
Calcium-Calmodulin-Dependent Protein Kinase Type 2
Calcium-Calmodulin-Dependent Protein Kinases/metabolism
Carbohydrate Conformation
Chromatography, High Pressure Liquid
Glycosylation
Mass Spectrometry
Molecular Sequence Data
Peptide Fragments/chemistry/isolation & purification
Phosphorylation
Rats
Rats, Sprague-Dawley
Synapsins/*chemistry/*metabolism
Abstract: Synapsin I is concentrated in nerve terminals, where it appears to anchor synaptic vesicles to the cytoskeleton and thereby ensures a steady supply of fusion-competent synaptic vesicles. Although phosphorylation-dependent binding of synapsin I to cytoskeletal elements and synaptic vesicles is well characterized, little is known about synapsin I's O-linked N-acetylglucosamine (O-GlcNAc) modifications. Here, we identified seven in vivo O-GlcNAcylation sites on synapsin I by analysis of HPLC-purified digests of rat brain synapsin I. The seven O-GlcNAcylation sites (Ser55, Thr56, Thr87, Ser516, Thr524, Thr562, and Ser576) in synapsin I are clustered around its five phosphorylation sites in domains B and D. The proximity of phosphorylation sites to O-GlcNAcylation sites in the regulatory domains of synapsin I suggests that O-GlcNAcylation may modulate phosphorylation and indirectly affect synapsin I interactions. With use of synthetic peptides, however, the presence of an O-GlcNAc at sites Thr562 and Ser576 resulted in only a 66% increase in the Km of calcium/calmodulin-dependent protein kinase II phosphorylation of site Ser566 with no effect on its Vmax. We conclude that O-GlcNAcylation likely plays a more direct role in synapsin I interactions than simply modulating the protein's phosphorylation.
Notes: Cole, R N
Hart, G W
NSO9415/NS/NINDS NIH HHS/United States
R01 HD13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
J Neurochem. 1999 Jul;73(1):418-28. doi: 10.1046/j.1471-4159.1999.0730418.x.
Author Address: Department of Biological Chemistry, School of Medicine, Johns Hopkins University, Baltimore, Maryland 21205-2185, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1770
Author: Coleman, J. A., Green, E. M. and Gouaux, E.
Year: 2016
Title: X-ray structures and mechanism of the human serotonin transporter
Journal: Nature
Volume: 532
Issue: 7599
Pages: 334-9
Epub Date: 2016/04/07
Date: Apr 21
Short Title: X-ray structures and mechanism of the human serotonin transporter
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature17629
PMCID: PMC4898786
NIHMSID: NIHMS764795 on the online version of the paper.
Accession Number: 27049939
Keywords: Allosteric Regulation/drug effects
Allosteric Site/drug effects
Antidepressive Agents/chemistry/metabolism/pharmacology
Citalopram/chemistry/metabolism/pharmacology
Crystallography, X-Ray
Dopamine Plasma Membrane Transport Proteins/chemistry
Drug Design
Extracellular Space/metabolism
Humans
Immunoglobulin Fab Fragments/immunology
Intracellular Space/metabolism
Ions/chemistry/metabolism
Ligands
Models, Molecular
Paroxetine/chemistry/metabolism/pharmacology
Protein Binding/drug effects
Protein Conformation/drug effects
Protein Stability
Serotonin/metabolism
Serotonin Plasma Membrane Transport Proteins/*chemistry/immunology/*metabolism
Structure-Activity Relationship
Abstract: The serotonin transporter (SERT) terminates serotonergic signalling through the sodium- and chloride-dependent reuptake of neurotransmitter into presynaptic neurons. SERT is a target for antidepressant and psychostimulant drugs, which block reuptake and prolong neurotransmitter signalling. Here we report X-ray crystallographic structures of human SERT at 3.15 Å resolution bound to the antidepressants (S)-citalopram or paroxetine. Antidepressants lock SERT in an outward-open conformation by lodging in the central binding site, located between transmembrane helices 1, 3, 6, 8 and 10, directly blocking serotonin binding. We further identify the location of an allosteric site in the complex as residing at the periphery of the extracellular vestibule, interposed between extracellular loops 4 and 6 and transmembrane helices 1, 6, 10 and 11. Occupancy of the allosteric site sterically hinders ligand unbinding from the central site, providing an explanation for the action of (S)-citalopram as an allosteric ligand. These structures define the mechanism of antidepressant action in SERT, and provide blueprints for future drug design.
Notes: 1476-4687
Coleman, Jonathan A
Green, Evan M
Gouaux, Eric
R37 MH070039/MH/NIMH NIH HHS/United States
5R37MH070039/MH/NIMH NIH HHS/United States
Canadian Institutes of Health Research/Canada
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2016 Apr 21;532(7599):334-9. doi: 10.1038/nature17629. Epub 2016 Apr 6.
Author Address: Vollum Institute, Oregon Health &Science University, Portland, Oregon 97239, USA.
Howard Hughes Medical Institute, Oregon Health &Science University, Portland, Oregon 97239, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1670
Author: Coles, C. H., Mitakidis, N., Zhang, P., Elegheert, J., Lu, W., Stoker, A. W., Nakagawa, T., Craig, A. M., Jones, E. Y. and Aricescu, A. R.
Year: 2014
Title: Structural basis for extracellular cis and trans RPTPσ signal competition in synaptogenesis
Journal: Nat Commun
Volume: 5
Pages: 5209
Epub Date: 2014/11/12
Date: Nov 11
Short Title: Structural basis for extracellular cis and trans RPTPσ signal competition in synaptogenesis
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms6209
PMCID: PMC4239663
NIHMSID: NIHMS634923
Accession Number: 25385546
Keywords: Animals
Cell Differentiation/physiology
Chick Embryo
Coculture Techniques
Crystallization
Extracellular Matrix Proteins/chemistry/*physiology
Humans
Ligands
Mice
Neurogenesis/*physiology
Neurons/cytology/physiology
Protein Binding
Protein Structure, Tertiary
Proteoglycans/chemistry/physiology
Receptor, trkC/chemistry/physiology
Receptor-Like Protein Tyrosine Phosphatases, Class 2/*chemistry/*physiology
Signal Transduction/physiology
Synapses/*physiology
Abstract: Receptor protein tyrosine phosphatase sigma (RPTPσ) regulates neuronal extension and acts as a presynaptic nexus for multiple protein and proteoglycan interactions during synaptogenesis. Unknown mechanisms govern the shift in RPTPσ function, from outgrowth promotion to synaptic organization. Here, we report crystallographic, electron microscopic and small-angle X-ray scattering analyses, which reveal sufficient inter-domain flexibility in the RPTPσ extracellular region for interaction with both cis (same cell) and trans (opposite cell) ligands. Crystal structures of RPTPσ bound to its postsynaptic ligand TrkC detail an interaction surface partially overlapping the glycosaminoglycan-binding site. Accordingly, heparan sulphate and heparin oligomers compete with TrkC for RPTPσ binding in vitro and disrupt TrkC-dependent synaptic differentiation in neuronal co-culture assays. We propose that transient RPTPσ ectodomain emergence from the presynaptic proteoglycan layer allows capture by TrkC to form a trans-synaptic complex, the consequent reduction in RPTPσ flexibility potentiating interactions with additional ligands to orchestrate excitatory synapse formation.
Notes: 2041-1723
Coles, Charlotte H
Mitakidis, Nikolaos
Zhang, Peng
Elegheert, Jonathan
Lu, Weixian
Stoker, Andrew W
Nakagawa, Terunaga
Craig, Ann Marie
Jones, E Yvonne
Aricescu, A Radu
090532/Z/09/Z/Wellcome Trust/United Kingdom
R01HD061543/HD/NICHD NIH HHS/United States
L009609/Medical Research Council/United Kingdom
R01 HD061543/HD/NICHD NIH HHS/United States
MH070860/MH/NIMH NIH HHS/United States
G0900084/Medical Research Council/United Kingdom
R01 MH070860/MH/NIMH NIH HHS/United States
090532/Wellcome Trust/United Kingdom
10976/Cancer Research UK/United Kingdom
G0700232/Medical Research Council/United Kingdom
G9900061/Medical Research Council/United Kingdom
A10976/Cancer Research UK/United Kingdom
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Commun. 2014 Nov 11;5:5209. doi: 10.1038/ncomms6209.
Author Address: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK.
Brain Research Centre and Department of Psychiatry, University of British Columbia, Vancouver, British Columbia, Canada V6T 2B5.
Cancer Section, Institute of Child Health, University College London, 30 Guilford Street, London WC1N 1EH, UK.
Department of Molecular Physiology and Biophysics, Vanderbilt University, School of Medicine, 702 Light Hall (0615), Nashville, Tennessee 37232-0615, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 415
Author: Collard, M. W. and Griswold, M. D.
Year: 1987
Title: Biosynthesis and molecular cloning of sulfated glycoprotein 2 secreted by rat Sertoli cells
Journal: Biochemistry
Volume: 26
Issue: 12
Pages: 3297-303
Epub Date: 1987/06/16
Date: Jun 16
Short Title: Biosynthesis and molecular cloning of sulfated glycoprotein 2 secreted by rat Sertoli cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00386a008
Accession Number: 3651384
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cells, Cultured
Cloning, Molecular
Clusterin
DNA/metabolism
Glycoproteins/biosynthesis/*genetics/metabolism
Male
*Molecular Chaperones
Plasmids
Polyribosomes
RNA, Messenger/genetics/isolation & purification
Rats
Sertoli Cells/*metabolism
Transcription, Genetic
Abstract: Sulfated glycoprotein 2 (SGP-2) is the major protein secreted by rat Sertoli cells. Pulse-chase labeling shows that SGP-2 is synthesized as a cotranslationally glycosylated 64-kDa precursor that is modified to a negatively charged 73-kDa form before intracellular cleavage to the mature 47- and 34-kDa subunits. A plasmid cDNA library was constructed from immunopurified mRNA, and a recombinant clone containing the entire protein coding sequence of SGP-2 was isolated. The 1857-nucleotide cDNA consists of a 297-nucleotide 5' noncoding segment, a 1341-nucleotide coding segment, and a 219-nucleotide 3' noncoding sequence. The 5' noncoding region contains five ATG codons followed by four short open reading frames. The derived SGP-2 sequence has a molecular weight of 51,379 and contains six potential N-glycosylation sites. Proteolytic processing sites for the preproprotein were determined by amino-terminal sequencing of the isolated SGP-2 subunits. Northern blots show a wide tissue distribution for the 2.0-kb SGP-2 message, and computer sequence analysis indicates a significant relationship between SGP-2 and human apolipoprotein A-I.
Notes: Collard, M W
Griswold, M D
HD-10808/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1987 Jun 16;26(12):3297-303. doi: 10.1021/bi00386a008.
Author Address: Biochemistry/Biophysics Program, Washington State University, Pullman 99164-4660.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1628
Author: Cominelli, A., Halbout, M., N'Kuli, F., Lemoine, P., Courtoy, P. J., Marbaix, E., Tyteca, D. and Henriet, P.
Year: 2014
Title: A unique C-terminal domain allows retention of matrix metalloproteinase-27 in the endoplasmic reticulum
Journal: Traffic
Volume: 15
Issue: 4
Pages: 401-17
Epub Date: 2014/02/20
Date: Apr
Short Title: A unique C-terminal domain allows retention of matrix metalloproteinase-27 in the endoplasmic reticulum
Alternate Journal: Traffic (Copenhagen, Denmark)
ISSN: 1398-9219
DOI: 10.1111/tra.12149
Accession Number: 24548619
Keywords: Amino Acid Sequence
Endoplasmic Reticulum/*enzymology
Humans
Matrix Metalloproteinases/chemistry/*metabolism
Molecular Sequence Data
Sequence Homology, Amino Acid
Subcellular Fractions/enzymology
Ergic
Triton X-114
brefeldin A
intermediate compartment
intracellular MMP
membrane
proteinase K
surface biotinylation
Abstract: Matrix metalloproteinase-27 (MMP-27) is poorly characterized. Sequence comparison suggests that a C-terminal extension (CTE) includes a potential transmembrane domain as in some membrane-type (MT)-MMPs. Having noticed that MMP-27 was barely secreted, we investigated its subcellular localization and addressed CTE contribution for MMP-27 retention. Intracellular MMP-27 was sensitive to endoglycosidase H. Subcellular fractionation and confocal microscopy evidenced retention of endogenous MMP-27 or recombinant rMMP-27 in the endoplasmic reticulum (ER) with locked exit across the intermediate compartment (ERGIC). Conversely, truncated rMMP-27 without CTE accessed downstream secretory compartments (ERGIC and Golgi) and was constitutively secreted. CTE addition to rMMP-10 (a secreted MMP) caused ER retention and blocked secretion. Addition of a PKA target sequence to the cytosolic C-terminus of transmembrane MT1-MMP/MMP-14 led to effective phosphorylation upon forskolin stimulation, but not for MMP-27, excluding transmembrane anchorage. Moreover, MMP-27 was protected from digestion by proteinase K. Finally, MT1-MMP/MMP-14 but neither endogenous nor recombinant MMP-27 partitioned in the detergent phase after Triton X-114 extraction, indicating that MMP-27 is not an integral membrane protein. In conclusion, MMP-27 is efficiently retained within the ER due to its unique CTE, which does not lead to stable membrane insertion. This could represent a novel ER retention system.
Notes: 1600-0854
Cominelli, Antoine
Halbout, Mathias
N'Kuli, Francisca
Lemoine, Pascale
Courtoy, Pierre J
Marbaix, Etienne
Tyteca, Donatienne
Henriet, Patrick
Journal Article
Research Support, Non-U.S. Gov't
England
Traffic. 2014 Apr;15(4):401-17. doi: 10.1111/tra.12149. Epub 2014 Feb 6.
Author Address: Cell Biology Unit, de Duve Institute, Université catholique de Louvain, Avenue Hippocrate 75, Box B1.75.05, B-1200, Brussels, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1009
Author: Compaan, D. M., Gonzalez, L. C., Tom, I., Loyet, K. M., Eaton, D. and Hymowitz, S. G.
Year: 2005
Title: Attenuating lymphocyte activity: the crystal structure of the BTLA-HVEM complex
Journal: J Biol Chem
Volume: 280
Issue: 47
Pages: 39553-61
Epub Date: 2005/09/20
Date: Nov 25
Short Title: Attenuating lymphocyte activity: the crystal structure of the BTLA-HVEM complex
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M507629200
Accession Number: 16169851
Keywords: Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Humans
In Vitro Techniques
Light
Lymphocytes/immunology
Models, Molecular
Molecular Mimicry
Molecular Sequence Data
Multiprotein Complexes/chemistry
Mutagenesis, Insertional
Protein Structure, Tertiary
Receptors, Immunologic/*chemistry/genetics/physiology
Receptors, Tumor Necrosis Factor/*chemistry/genetics/physiology
Receptors, Tumor Necrosis Factor, Member 14
Receptors, Virus/*chemistry/genetics/physiology
Recombinant Proteins/chemistry/genetics
Scattering, Radiation
Sequence Homology, Amino Acid
Viral Envelope Proteins/chemistry
Abstract: Five CD28-like proteins exert positive or negative effects on immune cells. Only four of these five receptors interact with members of the B7 family. The exception is BTLA (B and T lymphocyte attenuator), which instead interacts with the tumor necrosis factor receptor superfamily member HVEM (herpes virus entry mediator). To better understand this interaction, we determined the 2.8-A crystal structure of the BTLA-HVEM complex. This structure shows that BTLA binds the N-terminal cysteine-rich domain of HVEM and employs a unique binding surface compared with other CD28-like receptors. Moreover, the structure shows that BTLA recognizes the same surface on HVEM as gD (herpes virus glycoprotein D) and utilizes a similar binding motif. Light scattering analysis demonstrates that the extracellular domain of BTLA is monomeric and that BTLA and HVEM form a 1:1 complex. Alanine-scanning mutagenesis of HVEM was used to further define critical binding residues. Finally, BTLA adopts an immunoglobulin I-set fold. Despite structural similarities to other CD28-like members, BTLA represents a unique co-receptor.
Notes: Compaan, Deanne M
Gonzalez, Lino C
Tom, Irene
Loyet, Kelly M
Eaton, Dan
Hymowitz, Sarah G
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Biol Chem. 2005 Nov 25;280(47):39553-61. doi: 10.1074/jbc.M507629200. Epub 2005 Sep 16.
Author Address: Department of Protein Engineering, Genentech, Inc., S. San Francisco, California 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1067
Author: Compaan, D. M. and Hymowitz, S. G.
Year: 2006
Title: The crystal structure of the costimulatory OX40-OX40L complex
Journal: Structure
Volume: 14
Issue: 8
Pages: 1321-30
Epub Date: 2006/08/15
Date: Aug
Short Title: The crystal structure of the costimulatory OX40-OX40L complex
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2006.06.015
Accession Number: 16905106
Keywords: Amino Acid Sequence
Animals
Antigens, Differentiation/*chemistry/genetics
Crystallography, X-Ray
Enzyme-Linked Immunosorbent Assay
Humans
Membrane Glycoproteins/*chemistry/genetics
Mice
*Models, Molecular
Molecular Sequence Data
Multiprotein Complexes/*chemistry
Mutagenesis
OX40 Ligand
Protein Binding
Sequence Alignment
Tumor Necrosis Factors/*chemistry/genetics
Abstract: OX40 is a T cell costimulator activated by OX40L. Blockade of the OX40L-OX40 interaction has ameliorative effects in animal models of T cell pathologies. In order to better understand the interaction between OX40 and OX40L, we have determined the crystal structure of murine OX40L and of the human OX40-OX40L complex at 1.45 and 2.4 A, respectively. These structures show that OX40L is an unusually small member of the tumor necrosis factor superfamily (TNFSF). The arrangement of the OX40L protomers forming the functional trimer is atypical and differs from that of other members by a 15 degrees rotation of each protomer with respect to the trimer axis, resulting in an open assembly. Site-directed changes of the interfacial residues of OX40L suggest this interface lacks a single "hot spot" and that instead, binding energy is dispersed over at least two distinct areas. These structures demonstrate the structural plasticity of TNFSF members and their interactions with receptors.
Notes: Compaan, Deanne M
Hymowitz, Sarah G
RR-01646/RR/NCRR NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
United States
Structure. 2006 Aug;14(8):1321-30. doi: 10.1016/j.str.2006.06.015.
Author Address: Department of Protein Engineering, Genentech, Incorporated, 1 DNA Way, South San Francisco, California 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 848
Author: Compton, S. J., Sandhu, S., Wijesuriya, S. J. and Hollenberg, M. D.
Year: 2002
Title: Glycosylation of human proteinase-activated receptor-2 (hPAR2): role in cell surface expression and signalling
Journal: Biochem J
Volume: 368
Issue: Pt 2
Pages: 495-505
Epub Date: 2002/08/13
Date: Dec 1
Short Title: Glycosylation of human proteinase-activated receptor-2 (hPAR2): role in cell surface expression and signalling
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20020706
PMCID: PMC1222997
Accession Number: 12171601
Keywords: Alanine
Amino Acid Substitution
Animals
CHO Cells
Calcium/metabolism
Cell Membrane/genetics/*metabolism
Cricetinae
Glycosylation
Humans
Molecular Weight
Mutation
N-Acetylneuraminic Acid/chemistry/metabolism
Oligopeptides/metabolism/pharmacology
Receptor, PAR-2
Receptors, Thrombin/chemistry/drug effects/genetics/*metabolism
Recombinant Proteins/genetics/metabolism
Serine Endopeptidases/metabolism/pharmacology
*Signal Transduction
Trypsin/metabolism/pharmacology
Tryptases
Abstract: We have analysed the role of N-linked glycosylation in regulating human proteinase-activated receptor-2 (hPAR(2)) expression and function. Epitope-tagged wild-type hPAR(2) (wt-hPAR(2)) or hPAR(2) that lacked glycosylation sequons (following site-directed mutagenesis) in either the N-terminus [hPAR(2)N30A (Asn(30)-->Ala)], extracellular loop 2 [ECL2; hPAR(2)N222Q (Asn(222)-->Gln) or hPAR(2)N222A (Asn(222)-->Ala)] or both (hPAR(2)N30A,N222A or hPAR(2)N30A,N222Q) were expressed in the Chinese-hamster ovary (CHO) fibroblast cell line, Pro5. Western blot analysis of wt-hPAR(2) showed mature wt-hPAR(2) to have a molecular mass of 55-100 kDa, and 33-48 kDa following N -glycosidase F deglycosylation. FACS analysis and immunocytochemistry of the wt-hPAR(2) and PAR(2) mutant cell lines revealed that removal of both glycosylation sequons decreases (50% of wt-hPAR(2)) cell surface expression. Western blot analysis indicated that both N-linked sites are glycosylated. In functional studies, hPAR(2)N30A displayed a selective and significant increase in sensitivity towards tryptase. Interestingly, hPAR(2)N222A displayed a loss in sensitivity towards all PAR(2) agonists tested. However, further analysis revealed receptor sensitivity to alanine mutations in this domain, as the more conservative substitution hPAR(2)N222Q displayed no change in response to PAR(2) agonists. hPAR(2)N30A,N222Q displayed increased sensitivity towards tryptase, but a loss in sensitivity towards trypsin and the synthetic peptide SLIGRL-NH(2), although this loss in sensitivity towards trypsin and SLIGRL-NH(2) was secondary to changes in cell-surface expression. Finally, expression of sialic-acid-deficient wt-hPAR(2) in the CHO Lec2 glycosylation-deficient mutant cell line, showed a 40 kDa loss in molecular mass, in addition to a marked and selective increase in sensitivity towards tryptase. We conclude that hPAR(2) N-linked glycosylation and sialylation regulates receptor expression and/or signalling.
Notes: 1470-8728
Compton, Steven J
Sandhu, Sabrina
Wijesuriya, Suranga J
Hollenberg, Morley D
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2002 Dec 1;368(Pt 2):495-505. doi: 10.1042/BJ20020706.
Author Address: Mucosal Inflammation Research Group, Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1. S.J.Compton@hull.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 734
Author: Conklin, D., Gilbertson, D., Taft, D. W., Maurer, M. F., Whitmore, T. E., Smith, D. L., Walker, K. M., Chen, L. H., Wattler, S., Nehls, M. and Lewis, K. B.
Year: 1999
Title: Identification of a mammalian angiopoietin-related protein expressed specifically in liver
Journal: Genomics
Volume: 62
Issue: 3
Pages: 477-82
Epub Date: 2000/01/25
Date: Dec 15
Short Title: Identification of a mammalian angiopoietin-related protein expressed specifically in liver
Alternate Journal: Genomics
ISSN: 0888-7543 (Print)
0888-7543
DOI: 10.1006/geno.1999.6041
Accession Number: 10644446
Keywords: Amino Acid Motifs/genetics
Amino Acid Sequence
Angiopoietin-1
Angiopoietin-2
Angiopoietin-like Proteins
*Angiopoietins
Animals
Base Sequence
Binding Sites/genetics
Blotting, Northern
Chromosomes, Human, Pair 1/genetics
Glycosylation
Growth Substances/biosynthesis/*genetics
Humans
*Intercellular Signaling Peptides and Proteins
Liver/*metabolism
Membrane Glycoproteins/*genetics
Mice
Molecular Sequence Data
Organ Specificity
Physical Chromosome Mapping
Proteins/*genetics
RNA, Messenger/biosynthesis
Sequence Analysis, DNA
Sequence Homology, Amino Acid
Abstract: Based on searches of EST databases for signal sequences and amphipathic helices, we have identified and cloned an angiopoietin-like gene, ANGPTL3. Multiple tissue Northern blots show that ANGPTL3 is expressed principally in the liver. ANGPTL3 is expressed early during liver development, and expression is maintained in adult liver. Human ANGPTL3 is a 460-amino-acid polypeptide with the characteristic structure of angiopoietins: a signal peptide, an extended helical domain predicted to form dimeric or trimeric coiled-coils, a short linker peptide, and a globular fibrinogen homology domain (FHD). Murine ANGPTL3 is a 455-acid polypeptide encoded by seven exons on mouse chromosome 4, spanning about 11 kb of DNA. ANGPTL3 contains the four conserved cysteines implicated in the intramolecular disulfide bonds within the FHD, but it does not contain two other cysteines that are found within the FHD of angiopoietins 1, 2, and 4. ANGPTL3 also does not contain the characteristic calcium binding motif found in the other angiopoietins. By radiation hybrid mapping and the use of surrounding genes, human ANGPTL3 maps to the 1p31 region.
Notes: Conklin, D
Gilbertson, D
Taft, D W
Maurer, M F
Whitmore, T E
Smith, D L
Walker, K M
Chen, L H
Wattler, S
Nehls, M
Lewis, K B
Journal Article
United States
Genomics. 1999 Dec 15;62(3):477-82. doi: 10.1006/geno.1999.6041.
Author Address: ZymoGenetics, Inc., 1201 Eastlake Avenue East, Seattle, Washington 98012, USA. conklin@zgi.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 417
Author: Conradt, H. S., Egge, H., Peter-Katalinic, J., Reiser, W., Siklosi, T. and Schaper, K.
Year: 1987
Title: Structure of the carbohydrate moiety of human interferon-beta secreted by a recombinant Chinese hamster ovary cell line
Journal: J Biol Chem
Volume: 262
Issue: 30
Pages: 14600-5
Epub Date: 1987/10/25
Date: Oct 25
Short Title: Structure of the carbohydrate moiety of human interferon-beta secreted by a recombinant Chinese hamster ovary cell line
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3667593
Keywords: Animals
Carbohydrate Sequence
Cell Line
Cricetinae
Glycosylation
Interferon Type I/*analysis/isolation & purification/metabolism
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/*analysis
Recombinant Proteins/analysis
Sialyltransferases/pharmacology
Abstract: The carbohydrate structure of the major oligosaccharide of human interferon-beta (IFN-beta) synthesized by a genetically engineered Chinese hamster ovary cell line has been determined. Analysis of the glycopeptidase F-released carbohydrates by sequential exoglycosidase treatment, methylation analysis, and fast atom bombardment-mass spectrometry revealed that 95% of the IFN-beta oligosaccharides had the following structure: (Formula: see text). The remaining 5% of the carbohydrates are probably tri- or higher antennary oligosaccharide chains. The major oligosaccharide of the recombinant IFN-beta is remarkably homogeneous with respect to terminal galactose sialylation. NeuAc, which is alpha 2-3-linked to galactose in the human IFN-beta secreted by Chinese hamster ovary cells, can be re-incorporated with an alpha 2-6 linkage in vitro, into enzymatically desialylated IFN-beta using rat liver Gal beta 1-4GlcNAc alpha 2-6 sialyltransferase. The sugar chain is important for maintaining protein solubility as shown by the fact that IFN-beta protein precipitates after deglycosylation with glycopeptidase F.
Notes: Conradt, H S
Egge, H
Peter-Katalinic, J
Reiser, W
Siklosi, T
Schaper, K
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1987 Oct 25;262(30):14600-5.
Author Address: Department of Genetics, Gesellschaft für Biotechnologische Forschung mbH, Braunschweig, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 427
Author: Conradt, H. S., Geyer, R., Hoppe, J., Grotjahn, L., Plessing, A. and Mohr, H.
Year: 1985
Title: Structures of the major carbohydrates of natural human interleukin-2
Journal: Eur J Biochem
Volume: 153
Issue: 2
Pages: 255-61
Epub Date: 1985/12/02
Date: Dec 2
Short Title: Structures of the major carbohydrates of natural human interleukin-2
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1985.tb09295.x
Accession Number: 3935432
Keywords: Amino Acid Sequence
Amino Acids/analysis
Binding Sites
Calcimycin/pharmacology
Carbohydrates/blood
Chromatography, Gel
Chromatography, High Pressure Liquid
DNA/analysis
Electrophoresis, Polyacrylamide Gel
Humans
Interleukin-2/*analysis
Lymphocytes/metabolism
Mass Spectrometry
Methylation
Tetradecanoylphorbol Acetate/pharmacology
Abstract: Purified human interleukin-2 secreted by peripheral blood lymphocytes from healthy donors was found to exist in several forms. These forms were (partially) resolved by reversed-phase high-performance liquid chromatography and sodium dodecyl sulfate/polyacrylamide gel electrophoresis. Two major polypeptide species (interleukin-2 N1 and N2, 16.5 kDa) were shown to be glycosylated on the basis of [3H]galactose/[3H]glucosamine incorporation and determination of amino sugars after acid hydrolysis. A third component (interleukin-2 M, 14.5 kDa) represents a nonglycosylated form. The amino acid composition and the NH2-terminal sequence of both forms are consistent with the data deduced from the cDNA coding for interleukin-2 after removal of a leader peptide of 20 amino acids. Carbohydrates are O-linked to the IL-2 protein via threonine-3 of the polypeptide chain. The oligosaccharides were released by reductive beta-elimination and were purified by gel filtration and high-performance liquid chromatography. Applying methylation analysis, exoglycosidase digestion and fast atom bombardment mass spectrometry the following major carbohydrate structures were identified: N1, NeuAc(alpha 2-3)Gal(beta 1-3)GalNAc-ol; and N2, NeuAc(alpha 2-3)Gal(beta 1-3)[NeuAc(alpha 2-6)]GalNAc-ol.
Notes: Conradt, H S
Geyer, R
Hoppe, J
Grotjahn, L
Plessing, A
Mohr, H
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1985 Dec 2;153(2):255-61. doi: 10.1111/j.1432-1033.1985.tb09295.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 362
Author: Conradt, H. S., Nimtz, M., Dittmar, K. E., Lindenmaier, W., Hoppe, J. and Hauser, H.
Year: 1989
Title: Expression of human interleukin-2 in recombinant baby hamster kidney, Ltk-, and Chinese hamster ovary cells. Structure of O-linked carbohydrate chains and their location within the polypeptide
Journal: J Biol Chem
Volume: 264
Issue: 29
Pages: 17368-73
Epub Date: 1989/10/15
Date: Oct 15
Short Title: Expression of human interleukin-2 in recombinant baby hamster kidney, Ltk-, and Chinese hamster ovary cells. Structure of O-linked carbohydrate chains and their location within the polypeptide
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2793860
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/*analysis
Cell Line
Cloning, Molecular
Cricetinae
*Gene Expression
Glycosylation
Humans
Interleukin-2/analysis/*genetics
Plasmids
Protein Processing, Post-Translational
Recombinant Proteins/analysis/*genetics
Transfection
Abstract: The similarity or identity of O-glycosylation in glycoproteins from natural sources or produced in heterologous cell lines, a central problem for the development of many biotechnologically relevant production processes, was examined using interleukin-2 (IL-2) as a model. Human interleukin-2 was constitutively expressed in several mammalian cell lines in high amounts. The recombinant proteins were purified to homogeneity and their carbohydrate structures were analyzed. Only the NeuAc alpha 2-3Gal beta 1-3[NeuAc alpha 2-6]GalNAc oligosaccharide structure or the NeuAc alpha 2-3Gal beta 1-3GalNAc were found in all IL-2 preparations secreted from recombinant Ltk-, Chinese hamster ovary, and baby hamster kidney cell lines. The O-linked chains were exclusively linked to Thr in position 3 of the polypeptide chain which is the carbohydrate attachment site in natural human IL-2. The proportions of O-glycosylated versus nonglycosylated forms of the protein secreted by each recombinant cell line were independent of productivity or of cell culture conditions. Our results show that O-glycosylated human IL-2 can be produced by applying recombinant DNA technology in heterologous cell lines with the same type of post-translational modification that is observed for the protein secreted from natural T lymphocytes.
Notes: Conradt, H S
Nimtz, M
Dittmar, K E
Lindenmaier, W
Hoppe, J
Hauser, H
Journal Article
United States
J Biol Chem. 1989 Oct 15;264(29):17368-73.
Author Address: Department of Cell Biology and Genetics, Gesellschaft für Biotechnologische Forschung mbH, Braunschweig, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1043
Author: Conte, M., Arcaro, A., D'Angelo, D., Gnata, A., Mamone, G., Ferranti, P., Formisano, S. and Gentile, F.
Year: 2006
Title: A single chondroitin 6-sulfate oligosaccharide unit at Ser-2730 of human thyroglobulin enhances hormone formation and limits proteolytic accessibility at the carboxyl terminus. Potential insights into thyroid homeostasis and autoimmunity
Journal: J Biol Chem
Volume: 281
Issue: 31
Pages: 22200-22211
Epub Date: 2006/05/09
Date: Aug 4
Short Title: A single chondroitin 6-sulfate oligosaccharide unit at Ser-2730 of human thyroglobulin enhances hormone formation and limits proteolytic accessibility at the carboxyl terminus. Potential insights into thyroid homeostasis and autoimmunity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M513382200
Accession Number: 16679516
Keywords: Amino Acid Sequence
Autoimmunity
Chondroitin Sulfates/*physiology
Homeostasis
Hormones/*biosynthesis
Humans
Oligosaccharides
Peptide Hydrolases/metabolism
Serine
Thyroglobulin/chemistry/*metabolism/*physiology
Thyroid Gland/*immunology/physiology
Thyroiditis, Autoimmune/etiology
Thyroxine/biosynthesis
Triiodothyronine/biosynthesis
Abstract: We localized the site of type D (chondroitin 6-sulfate) oligosaccharide unit addition to human thyroglobulin (hTg). hTg was chromatographically separated into chondroitin 6-sulfate-containing (hTg-CS) and chondroitin 6-sulfate-devoid (hTg-CS0) molecules on the basis of their D-glucuronic acid content. In an ample number of hTg preparations, the fraction of hTg-CS in total hTg ranged from 32.0 to 71.6%. By exploiting the electrophoretic mobility shift and metachromasia conferred by chondroitin 6-sulfate upon the products of limited proteolysis of hTg, chondroitin 6-sulfate was first restricted to a carboxyl-terminal region, starting at residue 2514. A single chondroitin 6-sulfate-containing nonapeptide was isolated in pure form from the products of digestion of hTg with endoproteinase Glu-C, and its sequence was determined as LTAGXGLRE (residues 2726-2734, X being Ser2730 linked to the oligosaccharide chain). In an in vitro assay of enzymatic iodination, hTg-CS produced higher yields of 3,5,5 '-triiodothyronine (T3) (171%) and 3,5,3',5'-tetraiodothyronine (T4) (134%) than hTg-CS0. Unfractionated hTg behaved as hTg-CS. Thus, chondroitin 6-sulfate addition to a subset of hTg molecules enhanced the overall level of T4 and, in particular, T3 formation. Furthermore, the chondroitin 6-sulfate oligosaccharide unit of hTg-CS protected peptide bond Lys2714-Gly2715 from proteolysis, during the limited digestion of hTg-CS with trypsin. These findings provide insights into the molecular mechanism of regulation of the hormonogenic efficiency and of the T4/T3 ratio in hTg. The potential implications in the ability of hTg to function as an autoantigen and into the pathogenesis of thyroidal and extra-thyroidal manifestations of autoimmune thyroid disease are discussed.
Notes: Conte, Marisa
Arcaro, Alessia
D'Angelo, Daniela
Gnata, Ariele
Mamone, Gianfranco
Ferranti, Pasquale
Formisano, Silvestro
Gentile, Fabrizio
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Aug 4;281(31):22200-22211. doi: 10.1074/jbc.M513382200. Epub 2006 May 5.
Author Address: Dipartimento di Biologia e Patologia Cellulare e Molecolare, Università di Napoli Federico II, Via S. Pansini 5, 80131 Napoli.
Dipartimento di Scienze per la Salute, Università del Molise, Via F. De Sanctis, 86100 Campobasso.
Istituto di Scienze dell'Alimentazione del Consiglio Nazionale delle Ricerche, Via Roma 52a/c, 83100 Avellino.
Dipartimento di Scienza degli Alimenti, Università di Napoli Federico II, Parco Gussone, 80055 Portici, Italy.
Dipartimento di Scienze per la Salute, Università del Molise, Via F. De Sanctis, 86100 Campobasso. Electronic address: gentilefabrizio@unimol.it.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 423
Author: Cool, D. E., Edgell, C. J., Louie, G. V., Zoller, M. J., Brayer, G. D. and MacGillivray, R. T.
Year: 1985
Title: Characterization of human blood coagulation factor XII cDNA. Prediction of the primary structure of factor XII and the tertiary structure of beta-factor XIIa
Journal: J Biol Chem
Volume: 260
Issue: 25
Pages: 13666-76
Epub Date: 1985/11/05
Date: Nov 5
Short Title: Characterization of human blood coagulation factor XII cDNA. Prediction of the primary structure of factor XII and the tertiary structure of beta-factor XIIa
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3877053
Keywords: Amino Acid Sequence
Base Sequence
DNA/*analysis
*Factor XII/analysis/*genetics
Factor XIIa
Humans
*Peptide Fragments
Protein Conformation
Sequence Homology, Nucleic Acid
Abstract: A human liver cDNA library was screened by colony hybridization with two mixtures of synthetic oligodeoxyribonucleotides as probes. These oligonucleotides encoded regions of beta-factor XIIa as predicted from the amino acid sequence. Four positive clones were isolated that contained DNA coding for most of factor XII mRNA. DNA sequence analysis of these overlapping clones showed that they contained DNA coding for part of an amino-terminal extension, the complete amino acid sequence of plasma factor XII, a TGA stop codon, a 3' untranslated region of 150 nucleotides, and a poly(A)+ tail. The cDNA sequence predicts that plasma factor XII consists of 596 amino acid residues. Within the predicted amino acid sequence of factor XII, we have identified three peptide bonds that are cleaved by kallikrein during the formation of beta-factor XIIa. Comparison of the structure of factor XII with other proteins revealed extensive sequence identity with regions of tissue-type plasminogen activator (the epidermal growth factor-like region and the kringle region) and fibronectin (type I and type II homologies). As the type II region of fibronectin contains a collagen-binding site, the homologous region in factor XII may be responsible for the binding of factor XII to collagen. The carboxyl-terminal region of factor XII shares considerable amino acid sequence homology with other serine proteases including trypsin and many clotting factors. A preliminary structural model of beta-factor XIIa is proposed based on the known high resolution x-ray diffraction structures of trypsin, chymotrypsin, and elastase.
Notes: Cool, D E
Edgell, C J
Louie, G V
Zoller, M J
Brayer, G D
MacGillivray, R T
HL06350/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1985 Nov 5;260(25):13666-76.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 810
Author: Cornelis, S., Uttenweiler-Joseph, S., Panneels, V., Vassart, G. and Costagliola, S.
Year: 2001
Title: Purification and characterization of a soluble bioactive amino-terminal extracellular domain of the human thyrotropin receptor
Journal: Biochemistry
Volume: 40
Issue: 33
Pages: 9860-9
Epub Date: 2001/08/15
Date: Aug 21
Short Title: Purification and characterization of a soluble bioactive amino-terminal extracellular domain of the human thyrotropin receptor
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi0107389
Accession Number: 11502179
Keywords: Amino Acid Sequence
Animals
Antibodies/chemistry
Antibodies, Monoclonal/chemistry
Binding Sites
CHO Cells
Chromatography, Agarose
Cricetinae
Dose-Response Relationship, Drug
Flow Cytometry
Glycosylation
Graves Disease/metabolism
Humans
Imidazoles/pharmacology
Immunoglobulins/chemistry
Kinetics
Mass Spectrometry
Molecular Sequence Data
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Receptors, Thyrotropin/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: The amino-terminal ectodomain of the human TSH receptor has been expressed at the surface of CHO cells as a glycosylphosphatidylinositol-anchored molecule containing a 10-residue histidine tag close to its C terminus. The soluble ectodomain could be released from the cells by treatment with a glycosylphosphatidylinositol-phospholipase C and purified to apparent homogeneity by cobalt-Sepharose chromatography. Two nanomoles of material was obtained, which was suitable for analysis by mass spectrometry. This allowed the identification of four out of the six potential N-glycosylation sites as being effectively glycosylated. A proportion of the purified soluble ectodomain displayed specific binding of (125)I-labeled TSH, allowing for the first time performance of classical saturation binding experiments. Two classes of high-affinity binding sites were identified: site A, K(d) 0.014 nM; site B, K(d) 0.83 nM. The significance of site A, whose affinity is much higher than for the holoreceptor at the surface of intact cells, remains to be clarified. The purified ectodomain was capable of inhibiting efficiently the thyroid stimulating activity of immunoglobulins from patients with Graves' disease. It allowed computation of the amounts of these immunoglobulins in patient's serum, giving values up to 10 microg/mL. Contrary to all currently available assays, the soluble ectodomain of the TSH receptor purified in a functionally competent conformation allows direct studies of its interactions with TSH and autoantibodies and opens the way to structural studies.
Notes: Cornelis, S
Uttenweiler-Joseph, S
Panneels, V
Vassart, G
Costagliola, S
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2001 Aug 21;40(33):9860-9. doi: 10.1021/bi0107389.
Author Address: IRIBHN and Department of Medical Genetics, ULB, Brussels, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1033
Author: Corradi, H. R., Schwager, S. L., Nchinda, A. T., Sturrock, E. D. and Acharya, K. R.
Year: 2006
Title: Crystal structure of the N domain of human somatic angiotensin I-converting enzyme provides a structural basis for domain-specific inhibitor design
Journal: J Mol Biol
Volume: 357
Issue: 3
Pages: 964-74
Epub Date: 2006/02/16
Date: Mar 31
Short Title: Crystal structure of the N domain of human somatic angiotensin I-converting enzyme provides a structural basis for domain-specific inhibitor design
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2006.01.048
Accession Number: 16476442
Keywords: Amino Acid Sequence
Angiotensin-Converting Enzyme Inhibitors/*chemical synthesis/*metabolism
Binding Sites
Catalytic Domain
Crystallization
Crystallography, X-Ray
Drug Design
Humans
Molecular Sequence Data
Peptidyl-Dipeptidase A/*chemistry/genetics/*metabolism
Protein Structure, Tertiary
Abstract: Human somatic angiotensin I-converting enzyme (sACE) is a key regulator of blood pressure and an important drug target for combating cardiovascular and renal disease. sACE comprises two homologous metallopeptidase domains, N and C, joined by an inter-domain linker. Both domains are capable of cleaving the two hemoregulatory peptides angiotensin I and bradykinin, but differ in their affinities for a range of other substrates and inhibitors. Previously we determined the structure of testis ACE (C domain); here we present the crystal structure of the N domain of sACE (both in the presence and absence of the antihypertensive drug lisinopril) in order to aid the understanding of how these two domains differ in specificity and function. In addition, the structure of most of the inter-domain linker allows us to propose relative domain positions for sACE that may contribute to the domain cooperativity. The structure now provides a platform for the design of "domain-specific" second-generation ACE inhibitors.
Notes: Corradi, Hazel R
Schwager, Sylva L U
Nchinda, Aloysius T
Sturrock, Edward D
Acharya, K Ravi
070060/Wellcome Trust/United Kingdom
071047/Wellcome Trust/United Kingdom
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2006 Mar 31;357(3):964-74. doi: 10.1016/j.jmb.2006.01.048. Epub 2006 Jan 31.
Author Address: Department of Biology and Biochemistry, University of Bath, Claverton Down, Bath BA2 7AY, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2058
Author: Coscia, F., Taler-Verčič, A., Chang, V. T., Sinn, L., O'Reilly, F. J., Izoré, T., Renko, M., Berger, I., Rappsilber, J., Turk, D. and Löwe, J.
Year: 2020
Title: The structure of human thyroglobulin
Journal: Nature
Volume: 578
Issue: 7796
Pages: 627-630
Epub Date: 2020/02/07
Date: Feb
Short Title: The structure of human thyroglobulin
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/s41586-020-1995-4
PMCID: PMC7170718
NIHMSID: EMS85501
Accession Number: 32025030
Keywords: Bacterial Proteins/chemistry
*Cryoelectron Microscopy
HEK293 Cells
Humans
Maltose-Binding Proteins/chemistry
Models, Molecular
Mutation
Reproducibility of Results
Solvents/chemistry
Thyroglobulin/*chemistry/genetics/*ultrastructure
Thyroid Hormones/biosynthesis/metabolism
Tyrosine/chemistry/genetics/metabolism
Abstract: Thyroglobulin (TG) is the protein precursor of thyroid hormones, which are essential for growth, development and the control of metabolism in vertebrates(1,2). Hormone synthesis from TG occurs in the thyroid gland via the iodination and coupling of pairs of tyrosines, and is completed by TG proteolysis(3). Tyrosine proximity within TG is thought to enable the coupling reaction but hormonogenic tyrosines have not been clearly identified, and the lack of a three-dimensional structure of TG has prevented mechanistic understanding(4). Here we present the structure of full-length human thyroglobulin at a resolution of approximately 3.5 Å, determined by cryo-electron microscopy. We identified all of the hormonogenic tyrosine pairs in the structure, and verified them using site-directed mutagenesis and in vitro hormone-production assays using human TG expressed in HEK293T cells. Our analysis revealed that the proximity, flexibility and solvent exposure of the tyrosines are the key characteristics of hormonogenic sites. We transferred the reaction sites from TG to an engineered tyrosine donor-acceptor pair in the unrelated bacterial maltose-binding protein (MBP), which yielded hormone production with an efficiency comparable to that of TG. Our study provides a framework to further understand the production and regulation of thyroid hormones.
Notes: 1476-4687
Coscia, Francesca
Taler-Verčič, Ajda
Chang, Veronica T
Sinn, Ludwig
O'Reilly, Francis J
Izoré, Thierry
Renko, Miha
Berger, Imre
Rappsilber, Juri
Turk, Dušan
Löwe, Jan
203149/WT_/Wellcome Trust/United Kingdom
202754/WT_/Wellcome Trust/United Kingdom
103139/Z/13/Z/WT_/Wellcome Trust/United Kingdom
WT_/Wellcome Trust/United Kingdom
103139/WT_/Wellcome Trust/United Kingdom
MC_U105184326/MRC_/Medical Research Council/United Kingdom
Journal Article
Nature. 2020 Feb;578(7796):627-630. doi: 10.1038/s41586-020-1995-4. Epub 2020 Feb 5.
Author Address: MRC Laboratory of Molecular Biology, Cambridge, UK.
Jožef Stefan Institute, Ljubljana, Slovenia.
Centre of Excellence for Integrated Approaches in Chemistry and Biology of Proteins, Ljubljana, Slovenia.
Institute of Biotechnology, Technische Universität Berlin, Berlin, Germany.
Max Planck Bristol Centre for Minimal Biology, University of Bristol, Bristol, UK.
Wellcome Centre for Cell Biology, University of Edinburgh, Edinburgh, UK.
Jožef Stefan Institute, Ljubljana, Slovenia. dusan.turk@ijs.si.
Centre of Excellence for Integrated Approaches in Chemistry and Biology of Proteins, Ljubljana, Slovenia. dusan.turk@ijs.si.
MRC Laboratory of Molecular Biology, Cambridge, UK. jyl@mrc-lmb.cam.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1467
Author: Cotella, D., Radicke, S., Cipriani, V., Cavaletto, M., Merlin, S., Follenzi, A., Ravens, U., Wettwer, E., Santoro, C. and Sblattero, D.
Year: 2012
Title: N-glycosylation of the mammalian dipeptidyl aminopeptidase-like protein 10 (DPP10) regulates trafficking and interaction with Kv4 channels
Journal: Int J Biochem Cell Biol
Volume: 44
Issue: 6
Pages: 876-85
Epub Date: 2012/03/06
Date: Jun
Short Title: N-glycosylation of the mammalian dipeptidyl aminopeptidase-like protein 10 (DPP10) regulates trafficking and interaction with Kv4 channels
Alternate Journal: The international journal of biochemistry & cell biology
ISSN: 1357-2725
DOI: 10.1016/j.biocel.2012.02.011
Accession Number: 22387313
Keywords: Amino Acid Sequence
Animals
Asparagine/genetics/metabolism
CHO Cells
Cell Line
Cricetinae
Cricetulus
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases/chemistry/genetics/*metabolism
Electrophoresis, Polyacrylamide Gel
Flow Cytometry
Glycosylation
Humans
Immunoprecipitation
Mass Spectrometry
Molecular Sequence Data
Mutagenesis
Protein Binding
Protein Transport
Shal Potassium Channels/*metabolism
Abstract: The dipeptidyl aminopeptidase-like protein 10 (DPP10) is a type II transmembrane protein homologue to the serine protease DPPIV/CD26 but enzymatically inactive. In the mammalian brain, DPP10 forms a complex with voltage-gated potassium channels of the Kv4 family, regulating their cell surface expression and biophysical properties. DPP10 is a glycoprotein containing eight predicted N-glycosylation sites in the extracellular domain. In this study we investigated the role of N-glycosylation on DPP10 trafficking and functional activity. Using site-directed mutagenesis (N to Q) we showed that N-glycosylation occured at six positions. Glycosylation at these specific residues was necessary for DPP10 trafficking to the plasma membrane as observed by flow cytometry. The surface expression levels of the substitutions N90Q, N119Q, N257Q and N342Q were reduced by more than 60%. Hence the interaction with the Kv4.3/KChIP2a channel complex was disrupted preventing the hastening effect of wild type DPP10 on current kinetics. Interestingly, N257 was crucial for this function and its substitution to glutamine completely blocked DPP10 sorting to the cell surface and prevented DPP10 dimerization. In summary, we demonstrated that glycosylation was necessary for both DPP10 trafficking to the cell surface and functional interaction with Kv4 channels.
Notes: 1878-5875
Cotella, Diego
Radicke, Susanne
Cipriani, Valentina
Cavaletto, Maria
Merlin, Simone
Follenzi, Antonia
Ravens, Ursula
Wettwer, Erich
Santoro, Claudio
Sblattero, Daniele
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Int J Biochem Cell Biol. 2012 Jun;44(6):876-85. doi: 10.1016/j.biocel.2012.02.011. Epub 2012 Feb 22.
Author Address: Department of Health Sciences and Interdisciplinary Research Centre on Autoimmune Diseases (IRCAD), Università del Piemonte Orientale A. Avogadro, Via Solaroli 17, 28100 Novara, Italy. diego.cotella@med.unipmn.it
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 556
Author: Counillon, L., Pouysségur, J. and Reithmeier, R. A.
Year: 1994
Title: The Na+/H+ exchanger NHE-1 possesses N- and O-linked glycosylation restricted to the first N-terminal extracellular domain
Journal: Biochemistry
Volume: 33
Issue: 34
Pages: 10463-9
Epub Date: 1994/08/30
Date: Aug 30
Short Title: The Na+/H+ exchanger NHE-1 possesses N- and O-linked glycosylation restricted to the first N-terminal extracellular domain
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00200a030
Accession Number: 8068684
Keywords: Amino Acid Sequence
Animals
Base Sequence
Binding Sites
Cell Line
Cell Membrane/chemistry
Consensus Sequence
Conserved Sequence
Cricetinae
DNA, Complementary/genetics
Glycosylation
Humans
Models, Molecular
Molecular Sequence Data
Molecular Structure
Mutagenesis, Site-Directed
Rats
Sodium-Hydrogen Exchangers/*chemistry/genetics
Trout
Abstract: The ubiquitously-expressed human Na+H+ exchanger (NHE-1) contains three consensus sites (Asn-X-Ser/Thr) for N-linked glycosylation at asparagines 75, 370, and 410. The first extracellular loop is rich in serine and threonine residues which may contain O-linked carbohydrate. In order to determine unambiguously the sites of glycosylation and their role in biosynthesis and cation transport, site-directed mutagenesis at the individual potential N-glycosylation sites (Asn to Asp) was performed and all possible double and triple mutants were constructed. The mutated DNAs were expressed in PS120 hamster fibroblasts lacking endogenous exchanger, and the transfected cells were selected by their ability to survive acute intracellular acidification. All constructs produced functional exchangers that had transport rates and pharmacological profiles that were similar to that of wild-type. Immunoblot analysis of the expressed proteins with and without N-glycosidase F treatment showed that only the first N-glycosylation site (Asn 75) is utilized. In addition, treatment of NHE-1 with neuraminidase and O-glycosidase demonstrated that NHE-1 also contains O-linked oligosaccharide. Two forms of NHE-1 was consistently observed, a mature form with a molecular mass of 110,000 Da which contains N-linked and O-linked oligosaccharide and is expressed at the cell surface, and a lower molecular mass form (85,000 Da) present in the endoplasmic reticulum which only contains N-linked high-mannose oligosaccharide. NHE-3, an apically-expressed epithelial isoform which does not possess the N75 N-linked putative glycosylation site and any extracellular loops enriched in serine and threonine residues, does not exhibit any detectable glycosylation.
Notes: Counillon, L
Pouysségur, J
Reithmeier, R A
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1994 Aug 30;33(34):10463-9. doi: 10.1021/bi00200a030.
Author Address: Centre de Biochimie-CNRS, Université de Nice, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1985
Author: Cox, N. J., Luo, P. M., Smith, T. J., Bisnett, B. J., Soderblom, E. J. and Boyce, M.
Year: 2018
Title: A Novel Glycoproteomics Workflow Reveals Dynamic O-GlcNAcylation of COPγ1 as a Candidate Regulator of Protein Trafficking
Journal: Front Endocrinol (Lausanne)
Volume: 9
Pages: 606
Epub Date: 2018/11/22
Short Title: A Novel Glycoproteomics Workflow Reveals Dynamic O-GlcNAcylation of COPγ1 as a Candidate Regulator of Protein Trafficking
Alternate Journal: Frontiers in endocrinology
ISSN: 1664-2392 (Print)
1664-2392
DOI: 10.3389/fendo.2018.00606
PMCID: PMC6232944
Accession Number: 30459710
Keywords: COPI vesicle trafficking
O-GlcNAc
Silac
click chemistry
glycoproteomics
protein secretion
Abstract: O-linked β-N-acetylglucosamine (O-GlcNAc) is an abundant and essential intracellular form of protein glycosylation in animals and plants. In humans, dysregulation of O-GlcNAcylation occurs in a wide range of diseases, including cancer, diabetes, and neurodegeneration. Since its discovery more than 30 years ago, great strides have been made in understanding central aspects of O-GlcNAc signaling, including identifying thousands of its substrates and characterizing the enzymes that govern it. However, while many O-GlcNAcylated proteins have been reported, only a small subset of these change their glycosylation status in response to a typical stimulus or stress. Identifying the functionally important O-GlcNAcylation changes in any given signaling context remains a significant challenge in the field. To address this need, we leveraged chemical biology and quantitative mass spectrometry methods to create a new glycoproteomics workflow for profiling stimulus-dependent changes in O-GlcNAcylated proteins. In proof-of-principle experiments, we used this new workflow to interrogate changes in O-GlcNAc substrates in mammalian protein trafficking pathways. Interestingly, our results revealed dynamic O-GlcNAcylation of COPγ1, an essential component of the coat protein I (COPI) complex that mediates Golgi protein trafficking. Moreover, we detected 11 O-GlcNAc moieties on COPγ1 and found that this modification is reduced by a model secretory stress that halts COPI trafficking. Our results suggest that O-GlcNAcylation may regulate the mammalian COPI system, analogous to its previously reported roles in other protein trafficking pathways. More broadly, our glycoproteomics workflow is applicable to myriad systems and stimuli, empowering future studies of O-GlcNAc in a host of biological contexts.
Notes: 1664-2392
Cox, Nathan J
Luo, Peter M
Smith, Timothy J
Bisnett, Brittany J
Soderblom, Erik J
Boyce, Michael
R01 GM117473/GM/NIGMS NIH HHS/United States
Journal Article
Front Endocrinol (Lausanne). 2018 Oct 15;9:606. doi: 10.3389/fendo.2018.00606. eCollection 2018.
Author Address: Department of Biochemistry, Duke University School of Medicine, Durham, NC, United States.
Proteomics and Metabolomics Core Facility, Center for Genomic and Computational Biology, Duke University, Durham, NC, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1893
Author: Cox, N. J., Unlu, G., Bisnett, B. J., Meister, T. R., Condon, B. M., Luo, P. M., Smith, T. J., Hanna, M., Chhetri, A., Soderblom, E. J., Audhya, A., Knapik, E. W. and Boyce, M.
Year: 2018
Title: Dynamic Glycosylation Governs the Vertebrate COPII Protein Trafficking Pathway
Journal: Biochemistry
Volume: 57
Issue: 1
Pages: 91-107
Epub Date: 2017/11/22
Date: Jan 9
Short Title: Dynamic Glycosylation Governs the Vertebrate COPII Protein Trafficking Pathway
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/acs.biochem.7b00870
PMCID: PMC5767944
NIHMSID: NIHMS927295
Accession Number: 29161034
Keywords: Acetylglucosamine/*metabolism
Acylation
Animals
COP-Coated Vesicles/*metabolism
Cell Line
Collagen/metabolism
Craniofacial Abnormalities/metabolism
Disease Models, Animal
Glycosylation
Humans
Organelles/metabolism
Protein Conformation
Protein Processing, Post-Translational
Protein Transport
Vertebrates
Vesicular Transport Proteins/chemistry/genetics/*metabolism
Zebrafish
Abstract: The COPII coat complex, which mediates secretory cargo trafficking from the endoplasmic reticulum, is a key control point for subcellular protein targeting. Because misdirected proteins cannot function, protein sorting by COPII is critical for establishing and maintaining normal cell and tissue homeostasis. Indeed, mutations in COPII genes cause a range of human pathologies, including cranio-lenticulo-sutural dysplasia (CLSD), which is characterized by collagen trafficking defects, craniofacial abnormalities, and skeletal dysmorphology. Detailed knowledge of the COPII pathway is required to understand its role in normal cell physiology and to devise new treatments for disorders in which it is disrupted. However, little is known about how vertebrates dynamically regulate COPII activity in response to developmental, metabolic, or pathological cues. Several COPII proteins are modified by O-linked β-N-acetylglucosamine (O-GlcNAc), a dynamic form of intracellular protein glycosylation, but the biochemical and functional effects of these modifications remain unclear. Here, we use a combination of chemical, biochemical, cellular, and genetic approaches to demonstrate that site-specific O-GlcNAcylation of COPII proteins mediates their protein-protein interactions and modulates cargo secretion. In particular, we show that individual O-GlcNAcylation sites of SEC23A, an essential COPII component, are required for its function in human cells and vertebrate development, because mutation of these sites impairs SEC23A-dependent in vivo collagen trafficking and skeletogenesis in a zebrafish model of CLSD. Our results indicate that O-GlcNAc is a conserved and critical regulatory modification in the vertebrate COPII-dependent trafficking pathway.
Notes: 1520-4995
Cox, Nathan J
Orcid: 0000-0002-8573-5596
Unlu, Gokhan
Bisnett, Brittany J
Meister, Thomas R
Condon, Brett M
Luo, Peter M
Smith, Timothy J
Hanna, Michael
Chhetri, Abhishek
Soderblom, Erik J
Audhya, Anjon
Knapik, Ela W
Boyce, Michael
Orcid: 0000-0002-2729-4876
T32 GM008688/GM/NIGMS NIH HHS/United States
R01 HL092217/HL/NHLBI NIH HHS/United States
R01 GM110567/GM/NIGMS NIH HHS/United States
T32 GM007184/GM/NIGMS NIH HHS/United States
R01 GM117473/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochemistry. 2018 Jan 9;57(1):91-107. doi: 10.1021/acs.biochem.7b00870. Epub 2017 Dec 15.
Author Address: Departments of Medicine and Cell and Developmental Biology, Vanderbilt University Medical Center , Nashville, Tennessee 37232, United States.
Department of Biomolecular Chemistry, University of Wisconsin-Madison School of Medicine and Public Health , Madison, Wisconsin 53706, United States.
Duke Proteomics and Metabolomics Core Facility, Center for Genomic and Computational Biology, Duke University , Durham, North Carolina 27710, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 755
Author: Cruz, H. J., Peixoto, C. M., Nimtz, M., Alves, P. M., Dias, E. M., Moreira, J. L. and Carrondo, M. J.
Year: 2000
Title: Metabolic shifts do not influence the glycosylation patterns of a recombinant fusion protein expressed in BHK cells
Journal: Biotechnol Bioeng
Volume: 69
Issue: 2
Pages: 129-39
Epub Date: 2000/06/22
Date: Jul 20
Short Title: Metabolic shifts do not influence the glycosylation patterns of a recombinant fusion protein expressed in BHK cells
Alternate Journal: Biotechnology and bioengineering
ISSN: 0006-3592 (Print)
0006-3592
DOI: 10.1002/(sici)1097-0290(20000720)69:2<129::aid-bit2>3.0.co;2-5
Accession Number: 10861392
Keywords: Animals
Biotechnology
Carbohydrate Sequence
Cell Line
Cricetinae
Energy Metabolism
Gene Expression
Glycosylation
Humans
Immunoglobulin G/chemistry/genetics/metabolism
Interleukin-2/chemistry/genetics/metabolism
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/chemistry
Recombinant Fusion Proteins/chemistry/genetics/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: BHK-21 cells expressing a human IgG-IL2 fusion protein, with potential application in tumor-targeted therapy, were grown under different nutrient conditions in a continuous system for a time period of 80 days. At very low-glucose (< 0.5 mM) or glutamine (< 0. 2 mM) concentrations, a shift toward an energetically more efficient metabolism was observed. Cell-specific productivity was maintained under metabolically shifted growth conditions and at the same time an almost identical intracellular ATP content, obtained by in vivo (31)P NMR experiments, was observed. No significant differences in the oligosaccharide structures were detected from the IgG-IL2 fusion protein preparations obtained by growing cells under the different metabolic states. By using oligosaccharide mapping and MALDI/TOF-MS, only neutral diantennary oligosaccharides with or without core alpha1-6-linked fucose were detected that carried no, one or two beta1-4-linked galactose. Although the O-linked oligosaccharide structures that are present in the IL2 moiety of the protein were studied with less detail, the data obtained from the hydrazinolysis procedure point to the presence of the classical NeuAcalpha2-3Galbeta1-3GalNAc structure. Here, it is shown that under different defined cellular metabolic states, the quality of a recombinant product in terms of O- and N-linked oligosaccharides is stable, even after a prolonged cultivation period. Moreover, unaffected intracellular ATP levels under the different metabolic states were observed.
Notes: Cruz, H J
Peixoto, C M
Nimtz, M
Alves, P M
Dias, E M
Moreira, J L
Carrondo, M J
Journal Article
Research Support, Non-U.S. Gov't
United States
Biotechnol Bioeng. 2000 Jul 20;69(2):129-39. doi: 10.1002/(sici)1097-0290(20000720)69:2<129::aid-bit2>3.0.co;2-5.
Author Address: IBET/ITQB- Instituto de Biologia Experimental e Tecnológica / Instituto de Tecnologia Química e Biológica, Apartado 12, P-2781-901 Oeiras, Portugal.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 458
Author: Cummings, R. D., Kornfeld, S., Schneider, W. J., Hobgood, K. K., Tolleshaug, H., Brown, M. S. and Goldstein, J. L.
Year: 1983
Title: Biosynthesis of N- and O-linked oligosaccharides of the low density lipoprotein receptor
Journal: J Biol Chem
Volume: 258
Issue: 24
Pages: 15261-73
Epub Date: 1983/12/25
Date: Dec 25
Short Title: Biosynthesis of N- and O-linked oligosaccharides of the low density lipoprotein receptor
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6317691
Keywords: Adrenal Cortex/analysis
Animals
Carcinoma, Squamous Cell/metabolism
Cattle
Cell Line
Fibroblasts/metabolism
Humans
Mice
Molecular Weight
Neuraminidase/metabolism
Oligosaccharides/*biosynthesis
Protein Precursors/*biosynthesis
Receptors, Cell Surface/*biosynthesis
Receptors, LDL
Tunicamycin/pharmacology
Abstract: In human fibroblasts, the receptor for low density lipoprotein (LDL) is synthesized as a precursor of apparent Mr = 120,000 which is converted to a mature form of apparent Mr = 160,000, as determined by migration in sodium dodecyl sulfate (SDS)-polyacrylamide gels (Tolleshaug, H., Goldstein, J. L., Schneider, W. J., and Brown, M. S. (1982) Cell 30, 715-724). The current paper describes the relationship of N- and O-glycosylation to this post-translational modification. Oligosaccharides were analyzed from precursor and mature forms of LDL receptors that had been immunoprecipitated from cells grown in media containing radioactive sugars. In human epidermoid carcinoma A-431 cells, the receptor precursor appears to contain one N-linked high mannose oligosaccharide and approximately 6-9 N-acetylgalactosamine residues linked O-glycosidically to Ser/Thr residues. In the mature receptor, the O-linked oligosaccharides are mono- and disialylated species having the core structure of galactose leads to N-acetylgalactosamine leads to Ser/Thr. The single N-linked oligosaccharide of the mature receptor can either be a tri- or tetraantennary complex-type species. Similar results were obtained with normal human fibroblast receptor except that the O-linked oligosaccharides on the precursor are neutral disaccharides, of which one component is GalNAc and the N-linked complex type unit on the mature receptor is less branched. Since the addition of GalNAc residues to Ser/Thr residues precedes the conversion of N-linked high mannose-type oligosaccharides to complex-type structures, the transfer of N-acetylgalactosamine must occur prior to the entry of glycoproteins into the region of the Golgi containing the processing enzyme alpha-mannosidase I. We also studied the receptor from tunicamycin-treated cells and after treatment with neuraminidase. In addition, we analyzed the receptor synthesized by a lectin-resistant clone of Chinese hamster ovary cells that is deficient in adding galactose residues to both N- and O-linked oligosaccharides. These studies suggest that the apparent differences in molecular weight between the precursor and mature forms of the LDL receptor are largely, if not entirely, due to the addition of sialic acid and galactose residues to the O-linked GalNAc residues.(ABSTRACT TRUNCATED AT 400 WORDS)
Notes: Cummings, R D
Kornfeld, S
Schneider, W J
Hobgood, K K
Tolleshaug, H
Brown, M S
Goldstein, J L
R01 CA08759/CA/NCI NIH HHS/United States
5 T32 HD07103/HD/NICHD NIH HHS/United States
HL 20948/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1983 Dec 25;258(24):15261-73.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 824
Author: da Silva Correia, J. and Ulevitch, R. J.
Year: 2002
Title: MD-2 and TLR4 N-linked glycosylations are important for a functional lipopolysaccharide receptor
Journal: J Biol Chem
Volume: 277
Issue: 3
Pages: 1845-54
Epub Date: 2001/11/14
Date: Jan 18
Short Title: MD-2 and TLR4 N-linked glycosylations are important for a functional lipopolysaccharide receptor
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109910200
Accession Number: 11706042
Keywords: Antigens, Surface/genetics/metabolism/*physiology
Base Sequence
Blotting, Western
Cell Line
DNA Primers
*Drosophila Proteins
Glycosylation
Humans
Lipopolysaccharide Receptors/chemistry/metabolism/*physiology
Lipopolysaccharides/metabolism
Lymphocyte Antigen 96
Membrane Glycoproteins/genetics/metabolism/*physiology
Mutagenesis, Site-Directed
Precipitin Tests
Receptors, Cell Surface/genetics/metabolism/*physiology
Signal Transduction
Toll-Like Receptor 4
Toll-Like Receptors
Abstract: The lipopolysaccharide (LPS) receptor is a multi-protein complex that consists of at least three proteins, CD14, TLR4, and MD-2. Because each of these proteins is glycosylated, we have examined the functional role of N-linked carbohydrates of both MD-2 and TLR4. We demonstrate that MD-2 contains 2 N-glycosylated sites at positions Asn(26) and Asn(114), whereas the amino-terminal ectodomain of human TLR4 contains 9 N-linked glycosylation sites. Site-directed mutagenesis studies showed that cell surface expression of MD-2 did not depend on the presence of either N-linked site, whereas in contrast, TLR4 mutants carrying substitutions in Asn(526) or Asn(575) failed to be transported to the cell surface. Using a UV-activated derivative of Re595 LPS (ASD-Re595 LPS) in cross-linking assays, we demonstrated a critical role of MD-2 and TLR4 carbohydrates in LPS cross-linking to the LPS receptor. The ability of the various glycosylation mutants to support cell activation was also evaluated in transiently transfected HeLa cells. The double mutant of MD-2 failed to support LPS-induced activation of an interleukin-8 (IL-8) promoter-driven luciferase reporter to induce IL-8 secretion or to activate amino-terminal c-Jun kinase (JNK). Similar results were observed with TLR4 mutants lacking three or more N-linked glycosylation sites. Surprisingly, the reduction in activation resulting from expression of the Asn mutants of MD-2 and TLR4 can be partially reversed by co-expression with CD14. This suggests that the functional integrity of the LPS receptor depends both on the surface expression of at least three proteins, CD14, MD-2, and TLR4, and that N-linked sites of both MD-2 and TLR4 are essential in maintaining the functional integrity of this receptor.
Notes: da Silva Correia, Jean
Ulevitch, Richard J
AI15136/AI/NIAID NIH HHS/United States
GM28485/GM/NIGMS NIH HHS/United States
GM37696/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2002 Jan 18;277(3):1845-54. doi: 10.1074/jbc.M109910200. Epub 2001 Nov 12.
Author Address: Scripps Research Institute, Department of Immunology, La Jolla, California 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 488
Author: Dahr, W., Beyreuther, K., Kordowicz, M. and Krüger, J.
Year: 1982
Title: N-terminal amino acid sequence of sialoglycoprotein D (glycophorin C) from human erythrocyte membranes
Journal: Eur J Biochem
Volume: 125
Issue: 1
Pages: 57-62
Epub Date: 1982/06/15
Date: Jun 15
Short Title: N-terminal amino acid sequence of sialoglycoprotein D (glycophorin C) from human erythrocyte membranes
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1982.tb06650.x
Accession Number: 7106126
Keywords: Amino Acid Sequence
Chemical Phenomena
Chemistry
Chromatography, Gel
*Glycophorins
Humans
*Sialoglycoproteins
Abstract: The amino acid sequence of the N-terminal tryptic glycopeptide from a minor human erythrocyte membrane sialoglycoprotein (component D or glycophorin C) was determined by manual sequencing. The glycosylation sites were identified by a new procedure for the detection of the glycosylated derivatives released by Edman degradation. The fragment, comprising 47 residues, was found to contain an average of about 12 O-glycosidically linked oligosaccharides and one asparagine-linked carbohydrate chain. An identical hexapeptide sequence occurring in two regions of the glycopeptide provides evidence that it has developed by an internal gene duplication during evolution. In addition, a part of its structure shows a striking similarity to the sequence of a certain region of the MN and Ss erythrocyte membrane sialoglycoproteins (glycophorins A and B), suggesting that the molecules might be related.
Notes: Dahr, W
Beyreuther, K
Kordowicz, M
Krüger, J
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1982 Jun 15;125(1):57-62. doi: 10.1111/j.1432-1033.1982.tb06650.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1186
Author: Dai, S., Crawford, F., Marrack, P. and Kappler, J. W.
Year: 2008
Title: The structure of HLA-DR52c: comparison to other HLA-DRB3 alleles
Journal: Proc Natl Acad Sci U S A
Volume: 105
Issue: 33
Pages: 11893-7
Epub Date: 2008/08/14
Date: Aug 19
Short Title: The structure of HLA-DR52c: comparison to other HLA-DRB3 alleles
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0805810105
PMCID: PMC2515224
Accession Number: 18697946
Keywords: *Alleles
Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
HLA-DR Antigens/*chemistry/genetics/immunology/*metabolism
Humans
Hydrogen Bonding
Models, Molecular
Molecular Sequence Data
Peptides/chemistry/genetics/metabolism
Protein Binding
Protein Structure, Tertiary
Sequence Alignment
Abstract: Class II major histocompatibility complex (MHCII) molecules present antigens to CD4(+) T cells. In addition to the most commonly studied human MHCII isotype, HLA-DR, whose beta chain is encoded by the HLA-DRB1 locus, several other isotypes that use the same alpha chain but have beta chains encoded by other genes. These other DR molecules also are expressed in antigen-presenting cells and are known to participate in peptide presentation to T cells and to be recognized as alloantigens by other T cells. Like some of the HLA-DRB1 alleles, several of these alternate DR molecules have been associated with specific autoimmune diseases and T cell hypersensitivity. Here we present the structure of an HLA-DR molecule (DR52c) containing one of these alternate beta chains (HLA-DRB3*0301) bound to a self-peptide derived from the Tu elongation factor. The molecule shares structurally conserved elements with other MHC class II molecules but has some unique features in the peptide-binding groove. Comparison of the three major HLA-DBR3 alleles (DR52a, b, and c) suggests that they were derived from one another by recombination events that scrambled the four major peptide-binding pockets at peptide positions 1, 4, 6, and 9 but left virtually no polymorphisms elsewhere in the molecules.
Notes: 1091-6490
Dai, Shaodong
Crawford, Frances
Marrack, Philippa
Kappler, John W
R01 AI018785/AI/NIAID NIH HHS/United States
AI-17134/AI/NIAID NIH HHS/United States
AI-18785/AI/NIAID NIH HHS/United States
R01 AI017134/AI/NIAID NIH HHS/United States
R37 AI018785/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
R56 AI017134/AI/NIAID NIH HHS/United States
AI-22295/AI/NIAID NIH HHS/United States
R56 AI018785/AI/NIAID NIH HHS/United States
P01 AI022295/AI/NIAID NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2008 Aug 19;105(33):11893-7. doi: 10.1073/pnas.0805810105. Epub 2008 Aug 12.
Author Address: Integrated Department of Immunology, Howard Hughes Medical Institute, National Jewish Medical and Research Center, Denver, CO 80206, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1566
Author: Dall, E. and Brandstetter, H.
Year: 2013
Title: Mechanistic and structural studies on legumain explain its zymogenicity, distinct activation pathways, and regulation
Journal: Proc Natl Acad Sci U S A
Volume: 110
Issue: 27
Pages: 10940-5
Epub Date: 2013/06/19
Date: Jul 2
Short Title: Mechanistic and structural studies on legumain explain its zymogenicity, distinct activation pathways, and regulation
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1300686110
PMCID: PMC3703970
Accession Number: 23776206
Keywords: Amino Acid Sequence
Catalytic Domain/genetics
Crystallography, X-Ray
Cysteine Endopeptidases/*chemistry/genetics/*physiology
Enzyme Activation/genetics
Enzyme Precursors/chemistry/genetics/metabolism
Enzyme Stability/genetics
Humans
Hydrogen-Ion Concentration
Kinetics
Models, Molecular
Molecular Sequence Data
Protein Conformation
Sequence Homology, Amino Acid
Signal Transduction/genetics/*physiology
allostery
context-dependent activities
death domain
electrostatic stability switch
kcat-substrate specificity
Abstract: The cysteine protease legumain plays important functions in immunity and cancer at different cellular locations, some of which appeared conflicting with its proteolytic activity and stability. Here, we report crystal structures of legumain in the zymogenic and fully activated form in complex with different substrate analogs. We show that the eponymous asparagine-specific endopeptidase activity is electrostatically generated by pH shift. Completely unexpectedly, the structure points toward a hidden carboxypeptidase activity that develops upon proteolytic activation with the release of an activation peptide. These activation routes reconcile the enigmatic pH stability of legumain, e.g., lysosomal, nuclear, and extracellular activities with relevance in immunology and cancer. Substrate access and turnover is controlled by selective protonation of the S1 pocket (KM) and the catalytic nucleophile (kcat), respectively. The multibranched and context-dependent activation process of legumain illustrates how proteases can act not only as signal transducers but as decision makers.
Notes: 1091-6490
Dall, Elfriede
Brandstetter, Hans
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2013 Jul 2;110(27):10940-5. doi: 10.1073/pnas.1300686110. Epub 2013 Jun 17.
Author Address: Department of Molecular Biology, University of Salzburg, A-5020 Salzburg, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1059
Author: Dalpathado, D. S., Irungu, J., Go, E. P., Butnev, V. Y., Norton, K., Bousfield, G. R. and Desaire, H.
Year: 2006
Title: Comparative glycomics of the glycoprotein follicle stimulating hormone: glycopeptide analysis of isolates from two mammalian species
Journal: Biochemistry
Volume: 45
Issue: 28
Pages: 8665-73
Epub Date: 2006/07/13
Date: Jul 18
Short Title: Comparative glycomics of the glycoprotein follicle stimulating hormone: glycopeptide analysis of isolates from two mammalian species
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi060435k
Accession Number: 16834341
Keywords: Amino Acid Sequence
Animals
Follicle Stimulating Hormone/*chemistry
Follicle Stimulating Hormone, beta Subunit/*chemistry
Fourier Analysis
Glycopeptides/chemistry
Glycoprotein Hormones, alpha Subunit/*chemistry
Glycoproteins/chemistry
Glycosylation
Horses
Humans
Mass Spectrometry
Molecular Sequence Data
Polysaccharides/analysis
Abstract: Follicle stimulating hormone (FSH) is one of the important hormones that regulate gonadal functions. This hormone is glycosylated, and the glycans greatly influence the biological properties. In the present study the negatively charged glycopeptides of equine and human pituitary follicle stimulating hormone (eFSH and hFSH) have been characterized in a glycosylation site-specific manner using FT-ICR-MS and Edman sequencing. The characteristic pattern of glycan distribution at each glycosylation site has been deduced and compared between horse and human FSH preparations. The data suggest that site-specific differences exist between glycoforms of human and equine FSH. For instance, except for one site in the beta subunit (Asn7) of hFSH all other sites in both species have sulfated glycoforms. Also, glycoforms at Asn52 of hFSH are all complex type, whereas in eFSH, both complex and hybrid structures exist at this site. There is also a higher percentage of sulfated glycans in the latter site compared to the former. This is the first study that characterizes the glycans from this hormone in a glycosylation site-specific manner, and these data can be used to begin correlative studies between glycosylation structure and hormone function.
Notes: Dalpathado, Dilusha S
Irungu, Janet
Go, Eden P
Butnev, Vladimir Y
Norton, Katie
Bousfield, George R
Desaire, Heather
1 P20RR17708/RR/NCRR NIH HHS/United States
R01GM077226/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
United States
Biochemistry. 2006 Jul 18;45(28):8665-73. doi: 10.1021/bi060435k.
Author Address: Department of Chemistry, University of Kansas, Lawrence, Kansas 66045, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 550
Author: D'Andrea, G., D'Alessandro, A. M., Salucci, M. L. and Oratore, A.
Year: 1994
Title: Primary structure of the major glycan from human seminal transferrin
Journal: J Protein Chem
Volume: 13
Issue: 1
Pages: 31-6
Epub Date: 1994/01/01
Date: Jan
Short Title: Primary structure of the major glycan from human seminal transferrin
Alternate Journal: Journal of protein chemistry
ISSN: 0277-8033 (Print)
0277-8033
DOI: 10.1007/bf01891990
Accession Number: 8011069
Keywords: Amidohydrolases
Carbohydrate Conformation
Carbohydrate Sequence
Detergents
Humans
Hydrogen
Magnetic Resonance Spectroscopy/methods
Male
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Polysaccharides/*chemistry/isolation & purification
Semen/*chemistry
Transferrin/*chemistry/isolation & purification
Abstract: Human seminal transferrin (HSmT) is an iron-containing glycoprotein whose structural properties have not been adequately investigated. The carbohydrate content of the purified glycoprotein amount to 6.1%, and monosaccharide analysis revealed the major oligosaccharide moiety to be of the N-glycoside type. The carbohydrate chains were released from the iron-free form by digestion with peptide N-glycosidase F (PNGase F) in the presence of detergents such as SDS and beta-octylglucoside. After ethanol precipitation and fractionation on Bio-Gel P-6 and Bio-Gel P-2, the oligosaccharide was further purified on Mono-Q and desalted on Bio-Gel P-2. By 600-MHz 1H-NMR spectroscopy, the primary structure of the major N-linked oligosaccharide component was established to be: [formula: see text]
Notes: D'Andrea, G
D'Alessandro, A M
Salucci, M L
Oratore, A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Protein Chem. 1994 Jan;13(1):31-6. doi: 10.1007/BF01891990.
Author Address: Department of Sciences and Biomedical Technologies and Biometrics, University of L'Aquila, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1374
Author: Daou, S., Mashtalir, N., Hammond-Martel, I., Pak, H., Yu, H., Sui, G., Vogel, J. L., Kristie, T. M. and Affar el, B.
Year: 2011
Title: Crosstalk between O-GlcNAcylation and proteolytic cleavage regulates the host cell factor-1 maturation pathway
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 7
Pages: 2747-52
Epub Date: 2011/02/03
Date: Feb 15
Short Title: Crosstalk between O-GlcNAcylation and proteolytic cleavage regulates the host cell factor-1 maturation pathway
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1013822108
PMCID: PMC3041071
Accession Number: 21285374
Keywords: Blotting, Western
Cell Line
Cell Nucleus/*metabolism
Chromatin Immunoprecipitation
Fluorescent Antibody Technique
Gene Expression Regulation/genetics/*physiology
Host Cell Factor C1/*metabolism
Humans
Immediate-Early Proteins/metabolism
Immunoprecipitation
Mutagenesis
N-Acetylglucosaminyltransferases/*metabolism
Reverse Transcriptase Polymerase Chain Reaction
Simplexvirus/metabolism
Abstract: Host Cell Factor 1 (HCF-1) plays critical roles in regulating gene expression in a plethora of physiological processes. HCF-1 is first synthesized as a precursor, and subsequently specifically proteolytically cleaved within a large middle region termed the proteolytic processing domain (PPD). Although the underlying mechanism remains enigmatic, proteolysis of HCF-1 regulates its transcriptional activity and is important for cell cycle progression. Here we report that HCF-1 proteolysis is a regulated process. We demonstrate that a large proportion of the signaling enzyme O-linked-N-acetylglucosaminyl transferase (OGT) is complexed with HCF-1 and this interaction is essential for HCF-1 cleavage. Moreover, HCF-1 is, in turn, required for stabilizing OGT in the nucleus. We provide evidence indicating that OGT regulates HCF-1 cleavage via interaction with and O-GlcNAcylation of the HCF-1 PPD. In contrast, although OGT also interacts with the basic domain in the HCF-1 amino-terminal subunit, neither the interaction nor the O-GlcNAcylation of this region are required for proteolysis. Moreover, we show that OGT-mediated modulation of HCF-1 impacts the expression of the herpes simplex virus immediate-early genes, targets of HCF-1 during the initiation of viral infection. Together the data indicate that O-GlcNAcylation of HCF-1 is a signal for its proteolytic processing and reveal a unique crosstalk between these posttranslational modifications. Additionally, interactions of OGT with multiple HCF-1 domains may indicate that OGT has several functions in association with HCF-1.
Notes: 1091-6490
Daou, Salima
Mashtalir, Nazar
Hammond-Martel, Ian
Pak, Helen
Yu, Helen
Sui, Guangchao
Vogel, Jodi L
Kristie, Thomas M
Affar, El Bachir
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Proc Natl Acad Sci U S A. 2011 Feb 15;108(7):2747-52. doi: 10.1073/pnas.1013822108. Epub 2011 Feb 1.
Author Address: Maisonneuve-Rosemont Hospital Research Center, Department of Medicine, University of Montréal, Montréal, QC, Canada H1T 2M4.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1941
Author: Darabedian, N., Gao, J., Chuh, K. N., Woo, C. M. and Pratt, M. R.
Year: 2018
Title: The Metabolic Chemical Reporter 6-Azido-6-deoxy-glucose Further Reveals the Substrate Promiscuity of O-GlcNAc Transferase and Catalyzes the Discovery of Intracellular Protein Modification by O-Glucose
Journal: J Am Chem Soc
Volume: 140
Issue: 23
Pages: 7092-7100
Epub Date: 2018/05/18
Date: Jun 13
Short Title: The Metabolic Chemical Reporter 6-Azido-6-deoxy-glucose Further Reveals the Substrate Promiscuity of O-GlcNAc Transferase and Catalyzes the Discovery of Intracellular Protein Modification by O-Glucose
Alternate Journal: Journal of the American Chemical Society
ISSN: 0002-7863 (Print)
0002-7863
DOI: 10.1021/jacs.7b13488
PMCID: PMC6540071
NIHMSID: NIHMS1022466
Accession Number: 29771506
Keywords: Animals
Azides/chemical synthesis/chemistry/*metabolism
Cell Line, Tumor
Chlorocebus aethiops
Deoxyglucose/*analogs & derivatives/chemical synthesis/*metabolism
Glycosylation
Humans
Mice
N-Acetylglucosaminyltransferases/*metabolism
Protein Processing, Post-Translational
Proteins/*metabolism
Substrate Specificity
Uridine Diphosphate Sugars/biosynthesis
beta-N-Acetylhexosaminidases/metabolism
Abstract: Metabolic chemical reporters of glycosylation in combination with bioorthogonal reactions have been known for two decades and have been used by many different research laboratories for the identification and visualization of glycoconjugates. More recently, however, they have begun to see utility for the investigation of cellular metabolism and the tolerance of biosynthetic enzymes and glycosyltransferases to different sugars. Here, we take this concept one step further by using the metabolic chemical reporter 6-azido-6-deoxy-glucose (6AzGlc). We show that treatment of mammalian cells with the per- O-acetylated version of 6AzGlc results in robust labeling of a variety of proteins. Notably, the pattern of this labeling was consistent with O-GlcNAc modifications, suggesting that the enzyme O-GlcNAc transferase is quite promiscuous for its donor sugar substrates. To confirm this possibility, we show that 6AzGlc-treatment results in the labeling of known O-GlcNAcylated proteins, that the UDP-6AzGlc donor sugar is indeed produced in living cells, and that recombinant OGT will accept UDP-6AzGlc as a substrate in vitro. Finally, we use proteomics to first identify several bona fide 6AzGlc-modifications in mammalian cells and then an endogenous O-glucose modification on host cell factor. These results support the conclusion that OGT can endogenously modify proteins with both N-acetyl-glucosamine and glucose, raising the possibility that intracellular O-glucose modification may be a widespread modification under certain conditions or in particular tissues.
Notes: 1520-5126
Darabedian, Narek
Gao, Jinxu
Chuh, Kelly N
Woo, Christina M
Orcid: 0000-0001-8687-9105
Pratt, Matthew R
Orcid: 0000-0003-3205-5615
P30 CA014089/CA/NCI NIH HHS/United States
R01 GM125939/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Am Chem Soc. 2018 Jun 13;140(23):7092-7100. doi: 10.1021/jacs.7b13488. Epub 2018 May 30.
Author Address: Department of Chemistry and Chemical Biology , Harvard University , Cambridge , Massachusetts 02138 , United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1965
Author: Darabedian, N., Thompson, J. W., Chuh, K. N., Hsieh-Wilson, L. C. and Pratt, M. R.
Year: 2018
Title: Optimization of Chemoenzymatic Mass Tagging by Strain-Promoted Cycloaddition (SPAAC) for the Determination of O-GlcNAc Stoichiometry by Western Blotting
Journal: Biochemistry
Volume: 57
Issue: 40
Pages: 5769-5774
Epub Date: 2018/09/01
Date: Oct 9
Short Title: Optimization of Chemoenzymatic Mass Tagging by Strain-Promoted Cycloaddition (SPAAC) for the Determination of O-GlcNAc Stoichiometry by Western Blotting
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/acs.biochem.8b00648
PMCID: PMC6211186
NIHMSID: NIHMS991462
Accession Number: 30169966
Keywords: Acetylglucosamine/*chemistry
Blotting, Western
*Cycloaddition Reaction
Glycosylation
*Protein Processing, Post-Translational
Proteins/*chemistry
Abstract: The dynamic modification of intracellular proteins by O-linked β -N-acetylglucosamine (O-GlcNAcylation) plays critical roles in many cellular processes. Although various methods have been developed for O-GlcNAc detection, there are few techniques for monitoring glycosylation stoichiometry and state (i.e., mono-, di-, etc., O-GlcNAcylated). Measuring the levels of O-GlcNAcylation on a given substrate protein is important for understanding the biology of this critical modification and for prioritizing substrates for functional studies. One powerful solution to this limitation involves the chemoenzymatic installation of polyethylene glycol polymers of defined molecular mass onto O-GlcNAcylated proteins. These "mass tags" produce shifts in protein migration during sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) that can be detected by Western blotting. Broad adoption of this method by the scientific community has been limited, however, by a lack of commercially available reagents and well-defined protein standards. Here, we develop a "click chemistry" approach to this method using entirely commercial reagents and confirm the accuracy of the approach using a semisynthetic O-GlcNAcylated protein. Our studies establish a new, expedited experimental workflow and standardized methods that can be readily utilized by non-experts to quantify the O-GlcNAc stoichiometry and state on endogenous proteins in any cell or tissue lysate.
Notes: 1520-4995
Darabedian, Narek
Thompson, John W
Chuh, Kelly N
Hsieh-Wilson, Linda C
Orcid: 0000-0001-5661-1714
Pratt, Matthew R
Orcid: 0000-0003-3205-5615
T32 GM007616/GM/NIGMS NIH HHS/United States
T32 GM008042/GM/NIGMS NIH HHS/United States
R01 GM114537/GM/NIGMS NIH HHS/United States
R01 GM084724/GM/NIGMS NIH HHS/United States
RF1 AG060540/AG/NIA NIH HHS/United States
F30 AG055314/AG/NIA NIH HHS/United States
R01 GM125939/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Biochemistry. 2018 Oct 9;57(40):5769-5774. doi: 10.1021/acs.biochem.8b00648. Epub 2018 Sep 24.
Author Address: Division of Chemistry and Chemical Engineering , California Institute of Technology , Pasadena , California 91125 , United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 996
Author: Darlington, P. J., Kirchhof, M. G., Criado, G., Sondhi, J. and Madrenas, J.
Year: 2005
Title: Hierarchical regulation of CTLA-4 dimer-based lattice formation and its biological relevance for T cell inactivation
Journal: J Immunol
Volume: 175
Issue: 2
Pages: 996-1004
Epub Date: 2005/07/09
Date: Jul 15
Short Title: Hierarchical regulation of CTLA-4 dimer-based lattice formation and its biological relevance for T cell inactivation
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
DOI: 10.4049/jimmunol.175.2.996
Accession Number: 16002699
Keywords: Alanine/genetics
Antigens, CD/chemistry/*metabolism/physiology
Antigens, Differentiation/*chemistry/genetics/metabolism/*physiology
B7-1 Antigen/chemistry/*metabolism/physiology
B7-2 Antigen
CTLA-4 Antigen
Cell Line, Transformed
Cysteine/genetics
Dimerization
Disulfides/chemistry
Glycosylation
Humans
Immunosuppressive Agents/chemistry/*metabolism
Jurkat Cells
Ligands
Lymphocyte Activation/genetics/*immunology
Membrane Glycoproteins/chemistry/*metabolism/physiology
Membrane Microdomains/chemistry/genetics/metabolism
Point Mutation
Protein Binding/genetics/immunology
T-Lymphocytes/chemistry/*immunology/*metabolism
Abstract: CTLA-4 is an activation-induced, homodimeric inhibitory receptor in T cells. Recent crystallographic reports have suggested that it may form lattice-like arrays on the cell surface upon binding B7.1/B7.2 (CD80, CD86) molecules. To test the biological relevance of these CTLA-4-B7 lattices, we introduced a C122A point mutation in human CTLA-4, because this residue was shown to be essential for dimerization in solution. Surprisingly, we found that up to 35% of C122A CTLA-4 dimerized in human T lymphocytes. Moreover, C122A CTLA-4 partitioned within lipid rafts, colocalized with the TCR in the immunological synapse, and inhibited T cell activation. C122-independent dimerization of CTLA-4 involved N-glycosylation, because further mutation of the N78 and N110 glycosylation sites abrogated dimerization. Despite being monomeric, the N78A/N110A/C122A triple mutant CTLA-4 localized in the immunological synapse and inhibited T cell activation. Such functionality correlated with B7-induced dimerization of these mutant molecules. Based on these data, we propose a model of hierarchical regulation of CTLA-4 oligomerization by which B7 binding ultimately determines the formation of dimer-dependent CTLA-4 lattices that may be necessary for triggering B7-dependent T cell inactivation.
Notes: Darlington, Peter J
Kirchhof, Mark G
Criado, Gabriel
Sondhi, Jitin
Madrenas, Joaquín
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 2005 Jul 15;175(2):996-1004. doi: 10.4049/jimmunol.175.2.996.
Author Address: FOCIS Center for Clinical Immunology and Immunotherapeutics, Robarts Research Institute, 100 Perth Drive, London, Ontario, Canada N6A 5K8.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1981
Author: Darula, Z., Pap, Á and Medzihradszky, K. F.
Year: 2019
Title: Extended Sialylated O-Glycan Repertoire of Human Urinary Glycoproteins Discovered and Characterized Using Electron-Transfer/Higher-Energy Collision Dissociation
Journal: J Proteome Res
Volume: 18
Issue: 1
Pages: 280-291
Epub Date: 2018/11/09
Date: Jan 4
Short Title: Extended Sialylated O-Glycan Repertoire of Human Urinary Glycoproteins Discovered and Characterized Using Electron-Transfer/Higher-Energy Collision Dissociation
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/acs.jproteome.8b00587
Accession Number: 30407017
Keywords: Chromatography, Liquid
Glycopeptides/analysis
Glycoproteins/*urine
Humans
N-Acetylneuraminic Acid/chemistry
Polysaccharides/*analysis/chemistry
Proteomics/*methods
Tandem Mass Spectrometry/methods
*EThcD
*O-acetyl-sialic acid
*O-glycopeptides
*affinity chromatography
*blood-type antigen
*disialic acid
*mass spectrometry
*urinary glycoproteins
*wheat germ agglutinin
Abstract: A relatively novel activation technique, electron-transfer/higher-energy collision dissociation (EThcD) was used in the LC-MS/MS analysis of tryptic glycopeptides enriched with wheat germ agglutinin from human urine samples. We focused on the characterization of mucin-type O-glycopeptides. EThcD in a single spectrum provided information on both the peptide modified and the glycan carried. Unexpectedly, glycan oxonium ions indicated the presence of O-acetyl, and even O-diacetyl-sialic acids. B and Y fragment ions revealed that (i) in core 1 structures the Gal residue featured the O-acetyl-sialic acid, when there was only one in the glycan; (ii) several glycopeptides featured core 1 glycans with disialic acids, in certain instances O-acetylated; (iii) the disialic acid was linked to the GalNAc residue whatever the degree of O-acetylation; (iv) core 2 isomers with a single O-acetyl-sialic acid were chromatographically resolved. Glycan fragmentation also helped to decipher additional core 2 oligosaccharides: a LacdiNAc-like structure, glycans carrying sialyl Lewis(X/A) at different stages of O-acetylation, and blood antigens. A sialo core 3 structure was also identified. We believe this is the first study when such structures were characterized from a very complex mixture and were linked not only to a specific protein, but also the sites of modifications have been determined.
Notes: 1535-3907
Darula, Zsuzsanna
Pap, Ádám
Orcid: 0000-0003-1521-2017
Medzihradszky, Katalin F
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2019 Jan 4;18(1):280-291. doi: 10.1021/acs.jproteome.8b00587. Epub 2018 Nov 19.
Author Address: Biological Research Centre of the Hungarian Academy of Sciences , Temesvari krt. 62. , H-6726 Szeged , Hungary.
Doctoral School in Biology, Faculty of Science and Informatics , University of Szeged , Kozep fasor 52. , H-6726 Szeged , Hungary.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1982
Author: Das, S., Bailey, S. K., Metge, B. J., Hanna, A., Hinshaw, D. C., Mota, M., Forero-Torres, A., Chatham, J. C., Samant, R. S. and Shevde, L. A.
Year: 2019
Title: O-GlcNAcylation of GLI transcription factors in hyperglycemic conditions augments Hedgehog activity
Journal: Lab Invest
Volume: 99
Issue: 2
Pages: 260-270
Epub Date: 2018/11/14
Date: Feb
Short Title: O-GlcNAcylation of GLI transcription factors in hyperglycemic conditions augments Hedgehog activity
Alternate Journal: Laboratory investigation; a journal of technical methods and pathology
ISSN: 0023-6837 (Print)
0023-6837
DOI: 10.1038/s41374-018-0122-8
PMCID: PMC6857801
NIHMSID: NIHMS1503942 that could be perceived to bias their work.
Accession Number: 30420690
Keywords: Acetylglucosamine/*metabolism
Carrier Proteins/metabolism
Cell Line, Tumor
Glucose/*metabolism
Hedgehog Proteins/*metabolism
Humans
Hyperglycemia
Membrane Proteins/metabolism
N-Acetylglucosaminyltransferases/*metabolism
Thyroid Hormones/metabolism
Transcription Factors/metabolism
Abstract: Modification of proteins by O-linked β-N-acetylglucosamine (O-GlcNAc) promotes tumor cell survival, proliferation, epigenetic changes, angiogenesis, invasion, and metastasis. Here we demonstrate that in conditions of elevated glucose, there is increased expression of key drug resistance proteins (ABCB1, ABCG2, ERCC1, and XRCC1), all of which are regulated by the Hedgehog pathway. In elevated glucose conditions, we determined that the Hedgehog pathway transcription factors, GLI1 and GLI2, are modified by O-GlcNAcylation. This modification functionally enhanced their transcriptional activity. The activity of GLI was enhanced when O-GlcNAcase was inhibited, while inhibiting O-GlcNAc transferase caused a decrease in GLI activity. The metabolic impact of hyperglycemic conditions impinges on maintaining PKM2 in the less active state that facilitates the availability of glycolytic intermediates for biosynthetic pathways. Interestingly, under elevated glucose conditions, PKM2 directly influenced GLI activity. Specifically, abrogating PKM2 expression caused a significant decline in GLI activity and expression of drug resistance proteins. Cumulatively, our results suggest that elevated glucose conditions upregulate chemoresistance through elevated transcriptional activity of the Hedgehog/GLI pathway. Interfering in O-GlcNAcylation of the GLI transcription factors may be a novel target in controlling cancer progression and drug resistance of breast cancer.
Notes: 1530-0307
Das, Shamik
Bailey, Sarah K
Metge, Brandon J
Hanna, Ann
Hinshaw, Dominique C
Mota, Mateus
Forero-Torres, Andres
Chatham, John C
Samant, Rajeev S
Shevde, Lalita A
I01 BX003374/BX/BLRD VA/United States
R01 CA169202/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Lab Invest. 2019 Feb;99(2):260-270. doi: 10.1038/s41374-018-0122-8. Epub 2018 Nov 12.
Author Address: Department of Pathology, The University of Alabama at Birmingham, Birmingham, USA.
Department of Medicine, The University of Alabama at Birmingham, Birmingham, USA.
Comprehensive Diabetes Center, The University of Alabama at Birmingham, Birmingham, USA.
Comprehensive Cancer Center, The University of Alabama at Birmingham, Birmingham, USA.
Department of Pathology, The University of Alabama at Birmingham, Birmingham, USA. lalitasamant@uabmc.edu.
Comprehensive Cancer Center, The University of Alabama at Birmingham, Birmingham, USA. lalitasamant@uabmc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1182
Author: Das, S., Hahn, Y., Walker, D. A., Nagata, S., Willingham, M. C., Peehl, D. M., Bera, T. K., Lee, B. and Pastan, I.
Year: 2008
Title: Topology of NGEP, a prostate-specific cell:cell junction protein widely expressed in many cancers of different grade level
Journal: Cancer Res
Volume: 68
Issue: 15
Pages: 6306-12
Epub Date: 2008/08/05
Date: Aug 1
Short Title: Topology of NGEP, a prostate-specific cell:cell junction protein widely expressed in many cancers of different grade level
Alternate Journal: Cancer research
ISSN: 0008-5472 (Print)
0008-5472
DOI: 10.1158/0008-5472.can-08-0870
PMCID: PMC2562772
NIHMSID: NIHMS54452
Accession Number: 18676855
Keywords: Anoctamins
Humans
Male
Membrane Proteins/chemistry/immunology/*metabolism
Microscopy, Confocal
Microscopy, Fluorescence
Prostate/*metabolism
Prostatic Neoplasms/metabolism
Protein Conformation
Abstract: New gene expressed in prostate (NGEP) is a prostate-specific polytopic membrane protein found at high concentrations at cell:cell contact regions. To determine if NGEP is a useful target for antibody-based therapy of prostate cancer, we performed an immunohistochemical analysis of 126 human prostate carcinoma samples using polyclonal anti-NGEP sera and found that 91% of the cancers express NGEP protein. To elucidate the topology of NGEP and guide the development of monoclonal antibodies (mAb) reacting with the extracellular regions of NGEP, a hemagglutinin epitope tag was inserted at several positions within the NGEP sequence. The tagged proteins were expressed in 293T cells and locations of the tags were determined by immunofluorescence in intact or permeabilized cells. The results indicate that NGEP contains eight transmembrane domains with both the NH(2) and COOH termini of NGEP located inside the cell. We produced mAb to three regions that are predicted to be intracellular based on the epitope tag data (amino acids 1-352, 441-501, and 868-933), and as predicted, the mAb only detected the protein in permeabilized cells. NGEP is a glycoprotein with predicted glycosylation sites at N809 and N824. When these residues were converted to glutamine, glycosylation was abolished, confirming that the residues are extracellular. Our findings on the expression and the orientation of the NGEP protein serve as an important framework for the development of mAb targeting the extracellular regions of NGEP that could be used for prostate cancer immunotherapy.
Notes: 1538-7445
Das, Sudipto
Hahn, Yoonsoo
Walker, Dawn A
Nagata, Satoshi
Willingham, Mark C
Peehl, Donna M
Bera, Tapan K
Lee, Byungkook
Pastan, Ira
Z01 BC008759-16/Intramural NIH HHS/United States
Z01 BC010298-10/Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Cancer Res. 2008 Aug 1;68(15):6306-12. doi: 10.1158/0008-5472.CAN-08-0870.
Author Address: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland 20892-4264, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 879
Author: Datta, R., Choudhury, P., Ghosh, A. and Datta, B.
Year: 2003
Title: A glycosylation site, 60SGTS63, of p67 is required for its ability to regulate the phosphorylation and activity of eukaryotic initiation factor 2alpha
Journal: Biochemistry
Volume: 42
Issue: 18
Pages: 5453-60
Epub Date: 2003/05/07
Date: May 13
Short Title: A glycosylation site, 60SGTS63, of p67 is required for its ability to regulate the phosphorylation and activity of eukaryotic initiation factor 2alpha
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi020699g
Accession Number: 12731887
Keywords: Acetylglucosaminidase/genetics/*metabolism
Amino Acid Substitution
*Aminopeptidases
Animals
Blotting, Western
Carcinoma, Hepatocellular/genetics/metabolism/pathology
DNA Primers/chemistry
Eukaryotic Initiation Factor-2/*metabolism
Gene Expression Regulation
Glutathione Transferase/*metabolism
Glycoproteins/*metabolism
Glycosylation
Green Fluorescent Proteins
Liver Neoplasms/genetics/metabolism/pathology
Luminescent Proteins/genetics/*metabolism
Methionine/chemistry
Mutagenesis, Site-Directed
Mutation/genetics
Phosphorylation
Polymerase Chain Reaction
Precipitin Tests
Protein Conformation
Rats
Recombinant Fusion Proteins/chemistry/genetics/metabolism
Serine/chemistry
Threonine/chemistry
Tumor Cells, Cultured
eIF-2 Kinase/chemistry/genetics/metabolism
Abstract: Eukaryotic initiation factor 2- (eIF2-) associated glycoprotein p67 blocks eIF2alpha phosphorylation by kinases, and its N-terminal 1-97 amino acid segment can induce efficient translation. To investigate whether glycosylation at the serine/threonine clusters at this region is important in protein synthesis, we selected (27)TSST(30) and (60)SGTS(63) clusters for further analysis. By site-directed mutagenesis, (27)TSST(30) and (60)SGTS(63) clusters were substituted with (27)AAGA(30) and (60)AGAA(63) amino acid residues in full-length p67, and their EGFP fusions were constitutively expressed in rat tumor hepatoma cells (KRC-7). The (60)AGAA(63) mutant blocked eIF2alpha phosphorylation less than either wild-type p67 or the (27)AAGA(30) mutant. The (60)AGAA(63) mutant also showed a low level of protein synthesis rate, a lower level of glycosylation, increased turnover rate, and weaker binding to eIF2alpha. These results suggest that glycosylation within the (60)SGTS(63) sequence of p67 plays an important role in its stability and thus its regulation of protein synthesis by modulating the phosphorylation of the alpha-subunit of eIF2.
Notes: Datta, Rekha
Choudhury, Papiya
Ghosh, Arnab
Datta, Bansidhar
GM59190/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2003 May 13;42(18):5453-60. doi: 10.1021/bi020699g.
Author Address: Department of Chemistry, Kent State University, Kent, Ohio 44242, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 321
Author: David, G., Lories, V., Decock, B., Marynen, P., Cassiman, J. J. and Van den Berghe, H.
Year: 1990
Title: Molecular cloning of a phosphatidylinositol-anchored membrane heparan sulfate proteoglycan from human lung fibroblasts
Journal: J Cell Biol
Volume: 111
Issue: 6 Pt 2
Pages: 3165-76
Epub Date: 1990/12/01
Date: Dec
Short Title: Molecular cloning of a phosphatidylinositol-anchored membrane heparan sulfate proteoglycan from human lung fibroblasts
Alternate Journal: The Journal of cell biology
ISSN: 0021-9525 (Print)
0021-9525
DOI: 10.1083/jcb.111.6.3165
PMCID: PMC2116352
Accession Number: 2148568
Keywords: Amino Acid Sequence
Base Sequence
Cell Membrane/metabolism
Chondroitin Sulfate Proteoglycans/*genetics/metabolism
Cloning, Molecular
Culture Media
Fibroblasts
Glycolipids/metabolism
Glycosylation
Glycosylphosphatidylinositols
Heparan Sulfate Proteoglycans
Heparitin Sulfate/*genetics/metabolism
Humans
Lung
Molecular Sequence Data
Phosphatidylinositols/metabolism
Abstract: Two mAbs raised against the 64-kD core protein of a membrane heparan sulfate proteoglycan from human lung fibroblasts also recognize a nonhydrophobic proteoglycan which accumulates in the culture medium of the cells. Pulse-chase studies suggest that the hydrophobic cell-associated forms act as precursors for the nonhydrophobic medium-released species. The core proteins of the medium-released proteoglycans are slightly smaller than those of the hydrophobic cell-associated species, but the NH2-terminal amino acid sequences of both forms are identical. The characterization of human lung fibroblast cDNAs that encode the message for these core proteins and the effect of bacterial phosphatidylinositol-specific phospholipase C suggest that the hydrophobic proteoglycan is membrane-anchored through a phospholipid tail. These data identify a novel membrane proteoglycan in human lung fibroblasts and imply that the shedding of this proteoglycan may be related to the presence of the phospholipid anchor.
Notes: 1540-8140
David, G
Lories, V
Decock, B
Marynen, P
Cassiman, J J
Van den Berghe, H
Journal Article
Research Support, Non-U.S. Gov't
J Cell Biol. 1990 Dec;111(6 Pt 2):3165-76. doi: 10.1083/jcb.111.6.3165.
Author Address: Center for Human Genetics, University of Leuven, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 290
Author: Davidson, D. J. and Castellino, F. J.
Year: 1991
Title: Oligosaccharide structures present on asparagine-289 of recombinant human plasminogen expressed in a Chinese hamster ovary cell line
Journal: Biochemistry
Volume: 30
Issue: 3
Pages: 625-33
Epub Date: 1991/01/22
Date: Jan 22
Short Title: Oligosaccharide structures present on asparagine-289 of recombinant human plasminogen expressed in a Chinese hamster ovary cell line
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00217a006
Accession Number: 1899031
Keywords: Animals
Asparagine/*genetics
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Cricetulus
Female
Glucan 1,4-beta-Glucosidase
Glycosylation
Humans
Hydrolysis
Molecular Sequence Data
Oligonucleotides/chemistry/*metabolism
Ovary/*metabolism
Plasminogen/biosynthesis/*genetics
beta-Glucosidase
Abstract: The oligosaccharide structures linked to Asn289 of a recombinant (r) variant (R561S) human plasminogen (HPg) expressed in Chinese hamster ovary (CHO) cells, after transfection of these cells with a plasmid containing the cDNA coding for the variant HPg, have been determined. Employing high-performance anion-exchange liquid chromatography mapping of the oligosaccharide units cleaved from the protein by glycopeptidase F, compared with elution positions of standard oligosaccharides, coupled with monosaccharide compositional determinations and analyses of sequential exoglycosidase digestions and specific lectin binding, we find that considerable microheterogeneity in oligosaccharide structure exists at this sole potential N-linked glycosylation site on HPg. A variety of high-mannose structures, as well as bi-, tri-, and tetraantennary complex-type carbohydrate, has been found, in relative amounts of 1-25% of the total oligosaccharides. The complex-type structures contain variable amounts of sialic acid (Sia), ranging from 0 to 5 mol/mol of oligosaccharide in the different glycan structures. Neither hybrid-type molecules, N-acetylglucosamine bisecting oligosaccharides, nor N-acetyllactosaminyl-repeat structures were found to be present in the complex-type carbohydrate pool in observable amounts. Of interest, a significant portion of the Sia exists an outer arm structures in an (alpha 2,6) linkage to the penultimate galactose, a novel finding in CHO cell-directed glycosylation.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Davidson, D J
Castellino, F J
HL-13423/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1991 Jan 22;30(3):625-33. doi: 10.1021/bi00217a006.
Author Address: Department of Chemistry and Biochemistry, University of Notre Dame, Indiana 46556.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 308
Author: Davidson, D. J. and Castellino, F. J.
Year: 1991
Title: Asparagine-linked oligosaccharide processing in lepidopteran insect cells. Temporal dependence of the nature of the oligosaccharides assembled on asparagine-289 of recombinant human plasminogen produced in baculovirus vector infected Spodoptera frugiperda (IPLB-SF-21AE) cells
Journal: Biochemistry
Volume: 30
Issue: 25
Pages: 6165-74
Epub Date: 1991/06/25
Date: Jun 25
Short Title: Asparagine-linked oligosaccharide processing in lepidopteran insect cells. Temporal dependence of the nature of the oligosaccharides assembled on asparagine-289 of recombinant human plasminogen produced in baculovirus vector infected Spodoptera frugiperda (IPLB-SF-21AE) cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00239a013
Accession Number: 2059624
Keywords: Amidohydrolases/chemistry/metabolism
Animals
Asparagine/*metabolism
Baculoviridae/genetics
Carbohydrate Conformation
Carbohydrate Sequence
Cells, Cultured
DNA/biosynthesis
Gene Expression
Genetic Vectors
Glycoside Hydrolases/metabolism
Glycosylation
Hexosyltransferases/genetics
Humans
Hydrolysis
Mannose/chemistry
Molecular Sequence Data
Moths/*genetics/metabolism/microbiology
Oligosaccharides/*metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Plasminogen/biosynthesis/chemistry/*genetics
Recombinant Proteins
Time Factors
Abstract: Previous studies from this laboratory have established that lepidopteran insect cells possess the glycosylation machinery needed to assemble N-linked complex-type oligosaccharides on Asn289 of recombinant human plasminogen (r-HPg). In the present paper, we show that the nature of N289-linked glycosylation of [R561E]r-HPg expressed in Spodoptera frugiperda (IPLB-SF-21AE) cells is dependent upon the length of time of infection of the cells with the recombinant baculovirus/HPg-cDNA construct. At the earliest postinfection (p.i.) time period studied, i.e., 0-20 h, virtually all (96%) of the oligosaccharides released with glycopeptidase F from N289 of the expressed r-HPg were of the high-mannose type and comprised nearly the full range of such structures, containing 3-9 mannose units. At a time window of 60-96 h, p.i., essentially all of the oligosaccharides (92% of the total) assembled on N289 of rHPg were of the biantennary, triantennary, and tetraantennary complex classes, with varying extents of outer arm completion. At an intermediate time period window, of 20-60 h, p.i., a mixture of complex-type oligosaccharides, totaling approximately 77% of the glycans, with various levels of branching and outer arm completion, and high-mannose type of oligosaccharides, totaling approximately 23% of the glycans, was assembled on N289 of the r-HPg produced. These studies demonstrate that lepidopteran insect cells contain the glycosyltransferase genes required for assembly of N-linked complex oligosaccharide and that these transferases are utilized under proper conditions. The time dependency of the assembly of complex-type oligosaccharides on r-HPg indicates that an activation of the appropriate glycosyl transferases and/or transferase genes can take place. Thus, one consequence of the infective process with the recombinant baculovirus/HPg-cDNA construct is to alter the normal glycosylation characteristics of insect cells and to allow complex-type oligosaccharide processing to occur.
Notes: Davidson, D J
Castellino, F J
HL-13423/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1991 Jun 25;30(25):6165-74. doi: 10.1021/bi00239a013.
Author Address: Department of Chemistry and Biochemistry, University of Notre Dame, Indiana 46556.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 309
Author: Davidson, D. J. and Castellino, F. J.
Year: 1991
Title: Structures of the asparagine-289-linked oligosaccharides assembled on recombinant human plasminogen expressed in a Mamestra brassicae cell line (IZD-MBO503)
Journal: Biochemistry
Volume: 30
Issue: 27
Pages: 6689-96
Epub Date: 1991/07/09
Date: Jul 9
Short Title: Structures of the asparagine-289-linked oligosaccharides assembled on recombinant human plasminogen expressed in a Mamestra brassicae cell line (IZD-MBO503)
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00241a008
Accession Number: 2065054
Keywords: Animals
Asparagine/*chemistry
Baculoviridae/genetics
Carbohydrate Sequence
Cell Line
Chromatography, High Pressure Liquid
DNA/genetics
Electrophoresis, Polyacrylamide Gel
Genes, Viral
Glycosylation
Humans
Lepidoptera/cytology
Molecular Sequence Data
Oligosaccharides/*chemistry
Plasminogen/*chemistry/genetics
Recombinant Proteins/chemistry/genetics
Abstract: In this report, we have fortified and extended a previous investigation [Davidson, D. J., Fraser, M. J., & Castellino, F. J. (1990) Biochemistry 29, 5584-5590] in which we demonstrated for the first time that lepidopteran insect (Spodoptera frugiperda) cells (IPLB-SF-21AE) were capable of assembling N-linked complex oligosaccharide on a human protein (plasminogen), the cDNA of which had been inserted into these cells via recombinant DNA technology with a baculovirus vector. In order to investigate whether a more general capability of lepidopteran insect cells to produce complex oligosaccharide existed, and to identify the chemical nature of the types of oligosaccharides that such insect cells were able to assemble, we have infected Mamestra brassicae (IZD-MBO503) cells for 48 h with a recombinant (r) baculovirus containing the [R561E]human plasminogen (HPg) cDNA and characterized the nature of the glycopeptidase F (GF) released N-linked oligosaccharides contained on Asn289 of the r-HPg expressed by these cells. We found that approximately 63% of the total N-linked oligosaccharides were of the complex type, with bisialo-biantennary (28%), asialo-biantennary (7%), fucosylated bisialo-biantennary (25%), and fucosylated asialo-biantennary (3%) oligosaccharides representing the major complex-type carbohydrate species. The remainder of the oligosaccharides were of the high-mannose type, with (mannose)9(N-acetylglucosamine)2 (22%), (mannose)5(N-acetylglucosamine)2 (13%), and (mannose)3(N-acetylglucosamine)2 (2%) representing the major oligosaccharides observed.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Davidson, D J
Castellino, F J
HL-13423/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1991 Jul 9;30(27):6689-96. doi: 10.1021/bi00241a008.
Author Address: Department of Chemistry and Biochemistry, University of Notre Dame, Indiana 46556.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 334
Author: Davidson, D. J., Fraser, M. J. and Castellino, F. J.
Year: 1990
Title: Oligosaccharide processing in the expression of human plasminogen cDNA by lepidopteran insect (Spodoptera frugiperda) cells
Journal: Biochemistry
Volume: 29
Issue: 23
Pages: 5584-90
Epub Date: 1990/06/12
Date: Jun 12
Short Title: Oligosaccharide processing in the expression of human plasminogen cDNA by lepidopteran insect (Spodoptera frugiperda) cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00475a024
Accession Number: 2386787
Keywords: Animals
Base Sequence
Carbohydrate Sequence
DNA/*genetics
Gene Expression
Glycosylation
Humans
Molecular Sequence Data
Moths/genetics
Mutation
Oligosaccharides/*metabolism
Plasminogen/genetics/*metabolism
Recombinant Proteins/genetics
Abstract: A comparison has been made between the Asn289-linked oligosaccharide structures of human plasma plasminogen and a recombinant human plasminogen, expressed in lepidopteran insect (Spodoptera frugiperda) cells, after infection of these cells with a recombinant baculovirus containing the entire human plasminogen cDNA. Using anion-exchange liquid chromatography mapping of the oligosaccharide units cleaved from the proteins by glycopeptidase F, compared with elution positions of standard oligosaccharide structures, coupled with monosaccharide compositional analysis, we find that the human plasma protein contained only bisialo-biantennary complex-type carbohydrate and asialo-biantennary complex carbohydrate, confirming earlier work published by this laboratory. The glycosylation pattern of the insect cell expressed recombinant human plasminogen showed considerable microheterogeneity, with identifiable high-mannose carbohydrate (Man9GlcNAc2) and truncated high-mannose oligosaccharide (Man5GlcNAc2, Man4GlcNAc2, and Man3GlcNAc2). Of major importance, approximately 40% of the oligosaccharide population consisted of complex carbohydrate (bisialo-biantennary), identical in structure with that of the human plasma protein. This is the first direct identification of complex carbohydrate in proteins produced in insect cells and demonstrates that trimming and processing of high-mannose carbohydrate into complex-type oligosaccharide can occur. Our data indicate that both normal and alternate pathways exist in these cells for incorporation and trimming of high-mannose oligosaccharides and that mannosidases, as well as galactosyl-, hexosaminidasyl-, and sialyltransferases are present, and/or can be induced, in these cells. From these observations, we conclude that amino acid sequences and/or protein conformational properties can control oligosaccharide processing events.
Notes: Davidson, D J
Fraser, M J
Castellino, F J
HL-13423/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1990 Jun 12;29(23):5584-90. doi: 10.1021/bi00475a024.
Author Address: Department of Chemistry and Biochemistry, University of Notre Dame, Indiana 46556.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 751
Author: Davies, J. P. and Ioannou, Y. A.
Year: 2000
Title: Topological analysis of Niemann-Pick C1 protein reveals that the membrane orientation of the putative sterol-sensing domain is identical to those of 3-hydroxy-3-methylglutaryl-CoA reductase and sterol regulatory element binding protein cleavage-activating protein
Journal: J Biol Chem
Volume: 275
Issue: 32
Pages: 24367-74
Epub Date: 2000/05/24
Date: Aug 11
Short Title: Topological analysis of Niemann-Pick C1 protein reveals that the membrane orientation of the putative sterol-sensing domain is identical to those of 3-hydroxy-3-methylglutaryl-CoA reductase and sterol regulatory element binding protein cleavage-activating protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M002184200
Accession Number: 10821832
Keywords: Amino Acid Sequence
Animals
Binding Sites
COS Cells
*Carrier Proteins
Filipin/analysis
Glycosylation
Humans
Hydroxymethylglutaryl CoA Reductases/*chemistry
Intracellular Signaling Peptides and Proteins
*Membrane Glycoproteins
Membrane Proteins/*chemistry
Models, Molecular
Molecular Sequence Data
Niemann-Pick Diseases
Protein Structure, Secondary
Proteins/analysis/*chemistry/*metabolism
Recombinant Proteins/analysis/chemistry/metabolism
Sterols/metabolism
Transfection
Abstract: The Niemann-Pick C1 (NPC1) protein is predicted to be a polytopic glycoprotein, and it contains a region with extensive homology to the sterol-sensing domains (SSD) of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-R) and sterol regulatory element binding protein cleavage-activating protein (SCAP). To aid the functional characterization of NPC1, a model of NPC1 topology was evaluated by expression of epitope-tagged NPC1 proteins and investigation of epitope accessibility in selectively permeabilized cells. These results were further confirmed by expression of NPC1 and identification of glycosylated domains that are located in the lumen of the endoplasmic reticulum. Our data indicate that this glycoprotein contains 13 transmembrane domains, 3 large and 4 small luminal loops, 6 small cytoplasmic loops, and a cytoplasmic tail. Furthermore, our data show that the putative SSD of NPC1 is oriented in the same manner as those of HMG-R and SCAP, providing strong evidence that this domain is functionally important.
Notes: Davies, J P
Ioannou, Y A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2000 Aug 11;275(32):24367-74. doi: 10.1074/jbc.M002184200.
Author Address: Department of Human Genetics, Mount Sinai School of Medicine, New York, New York 10029, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 536
Author: Davis, D., Liu, X. and Segaloff, D. L.
Year: 1995
Title: Identification of the sites of N-linked glycosylation on the follicle-stimulating hormone (FSH) receptor and assessment of their role in FSH receptor function
Journal: Mol Endocrinol
Volume: 9
Issue: 2
Pages: 159-70
Epub Date: 1995/02/01
Date: Feb
Short Title: Identification of the sites of N-linked glycosylation on the follicle-stimulating hormone (FSH) receptor and assessment of their role in FSH receptor function
Alternate Journal: Molecular endocrinology (Baltimore, Md.)
ISSN: 0888-8809 (Print)
0888-8809
DOI: 10.1210/mend.9.2.7776966
Accession Number: 7776966
Keywords: Amidohydrolases/pharmacology
Animals
Asparagine/*metabolism
Cell Line
Cell Membrane/metabolism
Glycoside Hydrolases/pharmacology
Glycosylation
Humans
Mutagenesis, Site-Directed
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Protein Folding
Rats
Receptors, FSH/*metabolism
Transfection
Tunicamycin/pharmacology
Abstract: The FSH receptor (FSHR) contains a large extracellular domain in which exist three potential sites for N-linked glycosylation. A truncated form of the FSHR representing only the extracellular domain was created and expressed in mammalian cells. We show that this truncated receptor is glycosylated, through the carbohydrates are not as fully processed as those of the full-length receptor. This truncated receptor, which remains intracellular, binds FSH with an affinity comparable to that of the full-length FSHR. Therefore, although other regions of the FSHR may contribute to hormone binding, the extracellular domain alone can confer high affinity binding. The above results suggest that N-linked FSHR carbohydrates may, in some way, be required for FSH binding. Therefore, further experiments, done in the context of the full-length receptor, were performed to determine the actual sites of glycosylation in the FSHR as well as to elucidate their role in the functions of the FSHR. Site-directed mutagenesis was done to individually or collectively disrupt the potential sites for N-linked glycosylation. Western blot analyses of the wild type vs. mutant receptors demonstrate that, of the three potential sites for N-linked glycosylation, Asn's 174 and 276 are actually glycosylated. Binding assays demonstrate that these two N-linked FSHR carbohydrates are redundant in function since carbohydrate at either Asn174 or Asn276 allows the receptor to be expressed on the cell surface and to bind FSH with normal affinity. However, FSH binding activity is not observed with nonglycosylated mutant receptors where both sites have been collectively disrupted. Similarly, when cells expressing the wild type FSHR were treated with tunicamycin to prevent N-linked glycosylation, the resulting nonglycosylated FSHR was not able to bind FSH. In contrast, normal high affinity binding of FSH was maintained when N-linked carbohydrates were enzymatically removed from wild type receptors. Our results demonstrate that while N-linked carbohydrates on the FSH receptor are not required directly for the binding of hormone, a carbohydrate at either Asn174 or Asn276 is required for the efficient folding of the nascent receptor protein into a conformation that allows high affinity binding of hormone.
Notes: Davis, D
Liu, X
Segaloff, D L
DK-25295/DK/NIDDK NIH HHS/United States
HD-00968/HD/NICHD NIH HHS/United States
HD-28970/HD/NICHD NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Mol Endocrinol. 1995 Feb;9(2):159-70. doi: 10.1210/mend.9.2.7776966.
Author Address: Department of Physiology and Biophysics, University of Iowa College of Medicine, Iowa City 52242, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 386
Author: Davis, L. I. and Blobel, G.
Year: 1987
Title: Nuclear pore complex contains a family of glycoproteins that includes p62: glycosylation through a previously unidentified cellular pathway
Journal: Proc Natl Acad Sci U S A
Volume: 84
Issue: 21
Pages: 7552-6
Epub Date: 1987/11/01
Date: Nov
Short Title: Nuclear pore complex contains a family of glycoproteins that includes p62: glycosylation through a previously unidentified cellular pathway
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.84.21.7552
PMCID: PMC299337
Accession Number: 3313397
Keywords: Animals
Antibodies, Monoclonal
Antigen-Antibody Complex
Fluorescent Antibody Technique
Glycoproteins/*biosynthesis
Glycoside Hydrolases
Kinetics
Liver/*metabolism
Molecular Weight
Nuclear Envelope/*metabolism
Peptide Mapping
Rats
Rats, Inbred BUF
Uridine Diphosphate Galactose/metabolism
Abstract: Using a monoclonal antibody (mAb 414), we previously identified a protein of 62 kDa (p62) that was localized to the nuclear pore complex by immunoelectron microscopy. We also showed that p62 binds specifically to wheat germ agglutinin. Therefore, we proposed that this nuclear pore complex protein might be a member of a recently characterized family of glycoproteins that are labeled by in vitro galactosylation of rat liver nuclei and contain O-linked monosaccharidic GlcNAc residues. In support of this, we now show that incubation with N-acetylglucosaminidase reduces the molecular mass of p62 by approximately 3 kDa because of the removal of terminal GlcNAc residues. Moreover, p62 can be galactosylated in vitro by using UDP-[3H]galactose and galactosyltransferase. We also show that most of the GlcNAc residues are added within 5 min of synthesis, when p62 is soluble and cytosolic. Thus, the addition of GlcNAc is carried out in the cytoplasm and is clearly distinct from the N- and O-linked glycosylation pathways of the endoplasmic reticulum and Golgi complex. Using another mAb with a broad specificity for nuclear GlcNAc-containing proteins, we show by immunofluorescence and protein blotting of subnuclear fractions that some of these proteins are in the interior of the nucleus, and others are most likely located in the pore complex.
Notes: 1091-6490
Davis, L I
Blobel, G
GM27155/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1987 Nov;84(21):7552-6. doi: 10.1073/pnas.84.21.7552.
Author Address: Laboratory of Cell Biology, Howard Hughes Medical Institute, Rockefeller University, New York, NY 10021.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 313
Author: Davis, S. J., Ward, H. A., Puklavec, M. J., Willis, A. C., Williams, A. F. and Barclay, A. N.
Year: 1990
Title: High level expression in Chinese hamster ovary cells of soluble forms of CD4 T lymphocyte glycoprotein including glycosylation variants
Journal: J Biol Chem
Volume: 265
Issue: 18
Pages: 10410-8
Epub Date: 1990/06/25
Date: Jun 25
Short Title: High level expression in Chinese hamster ovary cells of soluble forms of CD4 T lymphocyte glycoprotein including glycosylation variants
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2113054
Keywords: Amino Acid Sequence
Animals
CD4 Antigens/*genetics/immunology/isolation & purification
Cell Line
Cricetinae
Cricetulus
Female
*Gene Expression
Genes, MHC Class II
*Genetic Variation
Glycopeptides/isolation & purification
Glycosylation
Molecular Sequence Data
Ovary
Peptide Fragments/isolation & purification
Rats
T-Lymphocytes/*immunology
Transfection
Abstract: The CD4 cell surface antigen is of interest as a marker of T lymphocytes that recognize foreign antigens in the context of MHC Class II antigen, as a receptor for the human immunodeficiency virus (HIV) and as a member of the immunoglobulin superfamily (IgSF) with four Ig-like domains present in the extracellular domain. In order to produce large amounts of soluble CD4 for x-ray crystallography and other molecular studies, a recently developed expression system based on selection via glutamine synthetase was used. Expression was attempted for rat CD4 corresponding to the full extracellular sequence (sCD4; domains 1-4), the NH2-terminal half (domains 1 and 2) and the first domain alone. Stable transfected Chinese hamster ovary cell lines were obtained that expressed sCD4 and sCD4 (half) at typical maximal levels in spent tissue culture supernatant of greater than 80 and 25 mg/liter, respectively. Domain 1 alone was not expressed and introduction of a N-linked glycosylation site did not facilitate expression. The role of glycosylation in the expression of sCD4 was investigated by mutagenesis of the constructs to remove each of the two N-linked glycosylation sites in turn and both together. All three forms were expressed at 60-120 mg/liter. The sCD4 (half) was not expressed after deletion of its N-linked site. The disulfide bonds of sCD4 were determined to be within domains 1, 2, and 4 and isolation of glycopeptides showed that both N-linked sites were glycosylated. Analysis of the hydrodynamic properties of sCD4 suggested that the molecule adopted an extended conformation in solution rather than folding to form a compact structure like an Fab. The possibility of dimerisation of CD4 was investigated but sCD4 dimers were not seen at an affinity cut-off of about 4 x 10(5) M-1.
Notes: Davis, S J
Ward, H A
Puklavec, M J
Willis, A C
Williams, A F
Barclay, A N
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1990 Jun 25;265(18):10410-8.
Author Address: Medical Research Council, Cellular Immunology Unit, Sir William Dunn School of Pathology, University of Oxford, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 438
Author: Dawson, G. and Clamp, J. R.
Year: 1968
Title: Investigations on the oligosaccharide units of an A myeloma globulin
Journal: Biochem J
Volume: 107
Issue: 3
Pages: 341-52
Epub Date: 1968/04/01
Date: Apr
Short Title: Investigations on the oligosaccharide units of an A myeloma globulin
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj1070341
PMCID: PMC1198669
Accession Number: 4172099
Keywords: Chemical Phenomena
Chemistry, Physical
Chromatography, Gas
Chromatography, Gel
Colorimetry
Electrophoresis
Fucose/analysis
Galactose/analysis
Glucosamine/analysis
Mannose/analysis
Multiple Myeloma/*blood
Neuraminic Acids/analysis
Oligosaccharides/*analysis
Oxidation-Reduction
gamma-Globulins/*analysis
Abstract: The carbohydrate content of an A myeloma globulin was investigated. The carbohydrate content was found to be unchanged when the protein was isolated from the patient over a period of 18 months. The various polymeric forms of the protein contained similar proportions of carbohydrate. The A myeloma globulin contained approx. 2 residues of 6-deoxy-l-galactose (l-fucose), 14-15 of d-mannose, 12-13 of d-galactose, 12-13 of 2-acetamido-2-deoxy-d-glucose (N-acetyl-d-glucosamine), 6 of 2-acetamido-2-deoxy-d-galactose (N-acetyl-d-galactosamine) and 5 of N-acetylneuraminic acid (sialic acid), and these were distributed between six oligosaccharide units all of which were present on the heavy polypeptide chains. The oligosaccharide units showed two kinds of heterogeneity, which have been termed central and peripheral. Central heterogeneity was shown by the presence of three completely different core units, which had the following compositions: (1) 3 residues of d-galactose and 3 of 2-acetamido-2-deoxy-d-galactose, joined to protein by an O-glycosidic linkage between acetamidohexose and serine; (2) 3 residues of d-mannose, 2 of d-galactose and 3 of 2-acetamido-2-deoxy-d-glucose, joined to protein by an N-glycosidic linkage between acetamidohexose and aspartic acid; (3) 4 residues of d-mannose and 3 of 2-acetamido-2-deoxy-d-glucose with a linkage similar to that in (2). The core oligosaccharide units showed peripheral heterogeneity in the attachment of 6-deoxy-l-galactose, 2-acetamido-2-deoxy-d-glucose and N-acetylneuraminic acid. Tentative structures are proposed for these various types of oligosaccharide unit. Glycopeptides were isolated in which the sialic acid content exceeded that of d-galactose. Explanations are given for the electrophoretic mobility and staining characteristics of the various glycopeptides.
Notes: 1470-8728
Dawson, G
Clamp, J R
Journal Article
Biochem J. 1968 Apr;107(3):341-52. doi: 10.1042/bj1070341.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1468
Author: de Baaij, J. H., Stuiver, M., Meij, I. C., Lainez, S., Kopplin, K., Venselaar, H., Müller, D., Bindels, R. J. and Hoenderop, J. G.
Year: 2012
Title: Membrane topology and intracellular processing of cyclin M2 (CNNM2)
Journal: J Biol Chem
Volume: 287
Issue: 17
Pages: 13644-55
Epub Date: 2012/03/09
Date: Apr 20
Short Title: Membrane topology and intracellular processing of cyclin M2 (CNNM2)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.342204
PMCID: PMC3340180
Accession Number: 22399287
Keywords: Animals
COS Cells
Cation Transport Proteins/*genetics
Cell Membrane/metabolism
Chlorocebus aethiops
Cyclins/*genetics
Cystathionine beta-Synthase/metabolism
Endoplasmic Reticulum/metabolism
Humans
Immunohistochemistry/methods
Magnesium/chemistry/metabolism
Mice
Mice, Inbred C57BL
Mutagenesis
*Mutation
Protein Isoforms
Protein Sorting Signals
Protein Structure, Tertiary
Tissue Distribution
Abstract: Recently, mutations in the cyclin M2 (CNNM2) gene were identified to be causative for severe hypomagnesemia. In kidney, CNNM2 is a basolaterally expressed protein with predominant expression in the distal convoluted tubule. Transcellular magnesium (Mg(2+)) reabsorption in the distal convoluted tubule represents the final step before Mg(2+) is excreted into the urine, thus fine-tuning its final excretion via a tightly regulated mechanism. The present study aims to get insight in the structure of CNNM2 and to characterize its post-translational modifications. Here, membrane topology studies using intramolecular epitopes and immunocytochemistry showed that CNNM2 has an extracellular N terminus and an intracellular C terminus. This suggests that one of the predicted transmembrane regions might be re-entrant. By homology modeling, we demonstrated that the loss-of-function mutation as found in patients disturbs the potential ATP binding by the intracellular cystathionine β-synthase domains. In addition, the cellular processing pathway of CNNM2 was exposed in detail. In the endoplasmic reticulum, the signal peptidase complex cleaves off a large N-terminal signal peptide of about 64 amino acids. Mutagenesis screening showed that CNNM2 is glycosylated at residue Asn-112, stabilizing CNNM2 on the plasma membrane. Interestingly, co-immunoprecipitation studies evidenced that CNNM2a forms heterodimers with the smaller isoform CNNM2b. These new findings on CNNM2 structure and processing may aid to elucidate the physiological role of CNNM2 in Mg(2+) reabsorption in the kidney.
Notes: 1083-351x
de Baaij, Jeroen H F
Stuiver, Marchel
Meij, Iwan C
Lainez, Sergio
Kopplin, Kathrin
Venselaar, Hanka
Müller, Dominik
Bindels, René J M
Hoenderop, Joost G J
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Apr 20;287(17):13644-55. doi: 10.1074/jbc.M112.342204. Epub 2012 Mar 7.
Author Address: Department of Physiology, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, 6500 HB Nijmegen, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 551
Author: de Beer, T., van Zuylen, C. W., Hård, K., Boelens, R., Kaptein, R., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1994
Title: Rapid and simple approach for the NMR resonance assignment of the carbohydrate chains of an intact glycoprotein. Application of gradient-enhanced natural abundance 1H-13C HSQC and HSQC-TOCSY to the alpha-subunit of human chorionic gonadotropin
Journal: FEBS Lett
Volume: 348
Issue: 1
Pages: 1-6
Epub Date: 1994/07/04
Date: Jul 4
Short Title: Rapid and simple approach for the NMR resonance assignment of the carbohydrate chains of an intact glycoprotein. Application of gradient-enhanced natural abundance 1H-13C HSQC and HSQC-TOCSY to the alpha-subunit of human chorionic gonadotropin
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(94)00547-8
Accession Number: 8026573
Keywords: Carbohydrate Sequence
Chorionic Gonadotropin/*chemistry
Glycoproteins/*chemistry
Humans
Magnetic Resonance Spectroscopy/*methods
Molecular Sequence Data
Abstract: The structure assessment of an intact glycoprotein in solution requires an extensive assignment of the carbohydrate NMR resonances. However, assignment of homonuclear spectra is very complicated because of the severe overlap of protein and carbohydrate signals. Application of pulsed field gradients allowed high quality natural abundance 1H-13C HSQC and HSQC-TOCSY spectra to be recorded of the alpha-subunit of human chorionic gonadotropin. Most carbohydrate 1H-13C correlations appear in a distinct region between the aromatic region and the protein C alpha-H alpha region. The enormous reduction in overlap led to fast and unambiguous assignment of the anomeric 1H-13C correlations. Subsequently, correlations of the monosaccharide skeleton atoms were readily assigned in the HSQC-TOCSY spectrum.
Notes: de Beer, T
van Zuylen, C W
Hård, K
Boelens, R
Kaptein, R
Kamerling, J P
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1994 Jul 4;348(1):1-6. doi: 10.1016/0014-5793(94)00547-8.
Author Address: Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 514
Author: de Beer, T., Vliegenthart, J. F., Löffler, A. and Hofsteenge, J.
Year: 1995
Title: The hexopyranosyl residue that is C-glycosidically linked to the side chain of tryptophan-7 in human RNase Us is alpha-mannopyranose
Journal: Biochemistry
Volume: 34
Issue: 37
Pages: 11785-9
Epub Date: 1995/09/19
Date: Sep 19
Short Title: The hexopyranosyl residue that is C-glycosidically linked to the side chain of tryptophan-7 in human RNase Us is alpha-mannopyranose
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00037a016
Accession Number: 7547911
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Endoribonucleases/*chemistry
Glycosylation
Humans
Magnetic Resonance Spectroscopy
Mannose/*chemistry
Molecular Sequence Data
Molecular Structure
Trisaccharides/chemistry
Tryptophan/chemistry
Abstract: Recently, the novel C-glycosidic linkage of a hexopyranosyl residue to the indole ring of tryptophan residue 7 of human RNase U(s) was reported [Hofsteenge, J., Müller, D. R., de Beer, T., Löffler A., Richter, W. J., & Vliegenthart, J. F. G. (1994) Biochemistry 33, 13524-13530]. Identification of this monosaccharide is a prerequisite for studies of its biosynthesis and its biological relevance. Using vicinal proton-proton coupling constants and rotating-frame nuclear Overhauser enhancements, ewe demonstrate that the C-linked substituent is alpha-mannopyranose. Furthermore, the nuclear magnetic resonance (NMR) data indicate that the mannopyranose moiety in a glycopeptide derived from RNase U(s) adopts several conformations on the NMR time scale.
Notes: de Beer, T
Vliegenthart, J F
Löffler, A
Hofsteenge, J
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1995 Sep 19;34(37):11785-9. doi: 10.1021/bi00037a016.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 350
Author: de Bruijn, M. H. and Fey, G. H.
Year: 1985
Title: Human complement component C3: cDNA coding sequence and derived primary structure
Journal: Proc Natl Acad Sci U S A
Volume: 82
Issue: 3
Pages: 708-12
Epub Date: 1985/02/01
Date: Feb
Short Title: Human complement component C3: cDNA coding sequence and derived primary structure
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.82.3.708
PMCID: PMC397115
Accession Number: 2579379
Keywords: Amino Acid Sequence
Base Sequence
Complement C3/*genetics
Complement C4/analysis
DNA/*analysis
Humans
Protein Precursors/genetics
RNA, Messenger/metabolism
alpha-Macroglobulins/analysis
Abstract: The complete cDNA coding sequence and derived amino acid sequence of human complement component C3 are presented. The encoded precursor molecule contains a signal peptide of 22 amino acid residues, the beta chain (645 residues), and the alpha chain (992 residues). The two chains are joined by four arginine residues not present in the mature protein. Several functionally important sites have been localized, such as the thiolester site, the cleavage site liberating the anaphylatoxin, and two sites of cleavage by the serine protease factor I, as well as a peptide fragment with leukocyte mobilizing activity. At least two carbohydrate attachment sites, one on each chain, have been identified. Human C3 has 79% identity to mouse C3 at the nucleotide level and 77% identity at the amino acid level. The protease alpha 2-macroglobulin and complement component C4 show considerable homology to C3, suggesting that the three proteins have evolved from a common ancestor.
Notes: 1091-6490
de Bruijn, M H
Fey, G H
AI-19651/AI/NIAID NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1985 Feb;82(3):708-12. doi: 10.1073/pnas.82.3.708.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 341
Author: De Caro, A. M., Adrich, Z., Fournet, B., Capon, C., Bonicel, J. J., De Caro, J. D. and Rovery, M.
Year: 1989
Title: N-terminal sequence extension in the glycosylated forms of human pancreatic stone protein. The 5-oxoproline N-terminal chain is O-glycosylated on the 5th amino acid residue
Journal: Biochim Biophys Acta
Volume: 994
Issue: 3
Pages: 281-4
Epub Date: 1989/02/23
Date: Feb 23
Short Title: N-terminal sequence extension in the glycosylated forms of human pancreatic stone protein. The 5-oxoproline N-terminal chain is O-glycosylated on the 5th amino acid residue
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/0167-4838(89)90305-1
Accession Number: 2493268
Keywords: Acetylgalactosamine/analysis
Acetylglucosamine/analysis
Amino Acid Sequence
Amino Acids/analysis
*Calcium-Binding Proteins/analysis/metabolism
Carbohydrates/analysis
Glycosylation
Humans
Hydrogen-Ion Concentration
Lithostathine
Molecular Sequence Data
*Nerve Tissue Proteins
Pancreatic Juice/analysis
Peptide Fragments/metabolism
Phosphoproteins
*Pyrrolidinones
*Pyrrolidonecarboxylic Acid
Solubility
Threonine/*metabolism
Trypsin/metabolism
Abstract: The pancreatic stone protein isolated from human calculi (PSP) derives from the immunoreactive protein forms detected in human pancreatic juice (PSP S2-5) through the tryptic cleavage of the Arg-11-Ile-12 bond. Among the eleven amino acids of the PSP S2-5 N-terminal extension Z-E-A-Q-T-E-L-P-Q-A-R, the first residue is an oxoproline and the fifth, a threonine, bears the single carbohydrate chain of the protein molecules. Variations in the glycan chain composition account for the differences in the Mr of PSP S2-5. The PSP S2-5 forms are very soluble in aqueous solutions between the pH values 5.0-9.0, whereas the proteolysated form is scarcely soluble.
Notes: De Caro, A M
Adrich, Z
Fournet, B
Capon, C
Bonicel, J J
De Caro, J D
Rovery, M
Journal Article
Netherlands
Biochim Biophys Acta. 1989 Feb 23;994(3):281-4. doi: 10.1016/0167-4838(89)90305-1.
Author Address: Unité de Recherches de Physiologie et de Pathologie Digestive, Marseille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 854
Author: de Marco, M. C., Martín-Belmonte, F., Kremer, L., Albar, J. P., Correas, I., Vaerman, J. P., Marazuela, M., Byrne, J. A. and Alonso, M. A.
Year: 2002
Title: MAL2, a novel raft protein of the MAL family, is an essential component of the machinery for transcytosis in hepatoma HepG2 cells
Journal: J Cell Biol
Volume: 159
Issue: 1
Pages: 37-44
Epub Date: 2002/10/09
Date: Oct 14
Short Title: MAL2, a novel raft protein of the MAL family, is an essential component of the machinery for transcytosis in hepatoma HepG2 cells
Alternate Journal: The Journal of cell biology
ISSN: 0021-9525 (Print)
0021-9525
DOI: 10.1083/jcb.200206033
PMCID: PMC2173496
Accession Number: 12370246
Keywords: Actins/metabolism
Antibodies, Monoclonal/immunology/metabolism
CD59 Antigens/metabolism
Carcinoma, Hepatocellular/*metabolism
Carrier Proteins/genetics/*metabolism
Cell Polarity
Humans
Immunoglobulin A/metabolism
Liver/cytology
Liver Neoplasms/*metabolism
Membrane Microdomains/*chemistry/metabolism
Myelin and Lymphocyte-Associated Proteolipid Proteins
Protein Transport/*physiology
Proteolipids/genetics/*metabolism
Receptors, Fc/metabolism
Tumor Cells, Cultured
*Vesicular Transport Proteins
Abstract: Transcytosis is used alone (e.g., hepatoma HepG2 cells) or in combination with a direct pathway from the Golgi (e.g., epithelial MDCK cells) as an indirect route for targeting proteins to the apical surface. The raft-associated MAL protein is an essential element of the machinery for the direct route in MDCK cells. Herein, we present the functional characterization of MAL2, a member of the MAL protein family, in polarized HepG2 cells. MAL2 resided selectively in rafts and is predominantly distributed in a compartment localized beneath the subapical F-actin cytoskeleton. MAL2 greatly colocalized in subapical endosome structures with transcytosing molecules en route to the apical surface. Depletion of endogenous MAL2 drastically blocked transcytotic transport of exogenous polymeric immunoglobulin receptor and endogenous glycosylphosphatidylinositol-anchored protein CD59 to the apical membrane. MAL2 depletion did not affect the internalization of these molecules but produced their accumulation in perinuclear endosome elements that were accessible to transferrin. Normal transcytosis persisted in cells that expressed exogenous MAL2 designed to resist the depletion treatment. MAL2 is therefore essential for transcytosis in HepG2 cells.
Notes: 1540-8140
de Marco, María C
Martín-Belmonte, Fernando
Kremer, Leonor
Albar, Juan P
Correas, Isabel
Vaerman, Jean P
Marazuela, Mónica
Byrne, Jennifer A
Alonso, Miguel A
Journal Article
Research Support, Non-U.S. Gov't
J Cell Biol. 2002 Oct 14;159(1):37-44. doi: 10.1083/jcb.200206033. Epub 2002 Oct 7.
Author Address: Centro de Biología Molecular "Severo Ochoa", Centro Nacional de Biotecnología, Universidad Autónoma de Madrid and Consejo Superior de Investigaciones Científicas, Cantoblanco, 28049 Madrid, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1794
Author: de Queiroz, R. M., Madan, R., Chien, J., Dias, W. B. and Slawson, C.
Year: 2016
Title: Changes in O-Linked N-Acetylglucosamine (O-GlcNAc) Homeostasis Activate the p53 Pathway in Ovarian Cancer Cells
Journal: J Biol Chem
Volume: 291
Issue: 36
Pages: 18897-914
Epub Date: 2016/07/13
Date: Sep 2
Short Title: Changes in O-Linked N-Acetylglucosamine (O-GlcNAc) Homeostasis Activate the p53 Pathway in Ovarian Cancer Cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.734533
PMCID: PMC5009264
Accession Number: 27402830
Keywords: Acetylglucosamine/genetics/*metabolism
Active Transport, Cell Nucleus/drug effects/genetics
Cell Line, Tumor
Cell Nucleus/genetics/*metabolism/pathology
Female
Gene Silencing
*Homeostasis
Humans
Mutation
N-Acetylglucosaminyltransferases/genetics/metabolism
Ovarian Neoplasms/genetics/*metabolism/pathology
*Protein Processing, Post-Translational
Protein Stability/drug effects
Pyrans/pharmacology
RNA, Messenger/biosynthesis/genetics
RNA, Neoplasm/biosynthesis/genetics
Thiazoles/pharmacology
Tumor Suppressor Protein p53/genetics/*metabolism
beta-N-Acetylhexosaminidases/antagonists & inhibitors/genetics/metabolism
*O-GlcNAcase
*O-linked N-acetylglucosamine (O-GlcNAc)
*O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
*cell growth
*ovarian cancer
*p53
Abstract: O-GlcNAcylation is a dynamic post-translational modification consisting of the addition of a single N-acetylglucosamine sugar to serine and threonine residues in proteins by the enzyme O-linked β-N-acetylglucosamine transferase (OGT), whereas the enzyme O-GlcNAcase (OGA) removes the modification. In cancer, tumor samples present with altered O-GlcNAcylation; however, changes in O-GlcNAcylation are not consistent between tumor types. Interestingly, the tumor suppressor p53 is modified by O-GlcNAc, and most solid tumors contain mutations in p53 leading to the loss of p53 function. Because ovarian cancer has a high frequency of p53 mutation rates, we decided to investigate the relationship between O-GlcNAcylation and p53 function in ovarian cancer. We measured a significant decrease in O-GlcNAcylation of tumor tissue in an ovarian tumor microarray. Furthermore, O-GlcNAcylation was increased, and OGA protein and mRNA levels were decreased in ovarian tumor cell lines not expressing the protein p53. Treatment with the OGA inhibitor Thiamet-G (TMG), silencing of OGA, or overexpression of OGA and OGT led to p53 stabilization, increased nuclear localization, and increased protein and mRNA levels of p53 target genes. These data suggest that changes in O-GlcNAc homeostasis activate the p53 pathway. Combination treatment of the chemotherapeutic cisplatin with TMG decreased tumor cell growth and enhanced cell cycle arrest without impairing cytotoxicity. The effects of TMG on tumor cell growth were partially dependent on wild type p53 activation. In conclusion, changes in O-GlcNAc homeostasis activate the wild type p53 pathway in ovarian cancer cells, and OGA inhibition has the potential as an adjuvant treatment for ovarian carcinoma.
Notes: 1083-351x
de Queiroz, Rafaela Muniz
Madan, Rashna
Chien, Jeremy
Dias, Wagner Barbosa
Slawson, Chad
P30 CA168524/CA/NCI NIH HHS/United States
P30 GM103326/GM/NIGMS NIH HHS/United States
R01 DK100595/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2016 Sep 2;291(36):18897-914. doi: 10.1074/jbc.M116.734533. Epub 2016 Jul 11.
Author Address: From the Laboratório de Glicobiologia Estrutural e Funcional, Instituto de Biofísica Carlos Chagas Filho, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Cidade Universitária, Rio de Janeiro, 21941-902RJ, Brazil.
Division of Hematology/Oncology, Department of Pathology, University of Kansas Medical Center, Kansas City, Kansas 66160.
Cancer Biology and.
From the Laboratório de Glicobiologia Estrutural e Funcional, Instituto de Biofísica Carlos Chagas Filho, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Cidade Universitária, Rio de Janeiro, 21941-902RJ, Brazil, diaswb@biof.ufrj.br.
the Departments of Biochemistry and Molecular Biology and cslawson@kumc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 517
Author: De Reggi, M., Capon, C., Gharib, B., Wieruszeski, J. M., Michel, R. and Fournet, B.
Year: 1995
Title: The glycan moiety of human pancreatic lithostathine. Structure characterization and possible pathophysiological implications
Journal: Eur J Biochem
Volume: 230
Issue: 2
Pages: 503-10
Epub Date: 1995/06/01
Date: Jun 1
Short Title: The glycan moiety of human pancreatic lithostathine. Structure characterization and possible pathophysiological implications
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1995.tb20589.x
Accession Number: 7607222
Keywords: Calcium-Binding Proteins/*chemistry/physiology
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Humans
Lithostathine
Molecular Sequence Data
Nerve Tissue Proteins/*chemistry/physiology
Pancreas/*chemistry
Polysaccharides/*chemistry
Protein Conformation
Protein Processing, Post-Translational
Abstract: Lithostathine, also known as pancreatic stone protein, pancreatic thread protein or regenerating protein, is a glycoprotein which is normally found in the exocrine pancreas, whereas in other tissues it appears either only under pathological conditions, such as Alzheimer's disease (brain), cancer (colon) or during regeneration (endocrine pancreas). In the latter case, it has been shown recently that it acts as a growth factor which stimulates islet regeneration. Little is known about its glycan moiety, which conceivably might be involved in this tissue specificity and pathophysiological characteristics. Therefore we isolated the major oligosaccharide chains of human pancreatic lithostathine and determined their sequences by means of NMR analysis. We obtained eleven different glycoforms and we were able to determine the sequence of seven of them. They all were from the same site of glycosylation (Thr5) and displayed the same core 2 structure: GlcNAc(beta 1-6)[Gal(beta 1-3)]GalNAc alpha-. They ranged in size from 4 to 9 sugar residues. Elongation was found to proceed from a common tetrasaccharidic core: Gal(beta 1-4)GlcNAc(beta 1-6)[Gal(beta 1-3)]GalNAc-ol through N-acetyllactosamine units. The non-reducing ends of some oligosaccharides carry the antigenic determinant H, with presence of external Fuc linked only in (alpha 1-2) to Gal. All the glycans, except one, carry a sialic acid in (alpha 2-3) linkage to Gal, with one disialylated form which displays a supplementary (alpha 2-6) linkage. These findings are consistent with the polymorphism of the protein, shown by means of SDS gel electrophoresis and isoelectric focusing, either in its native form or after enzymic processing. Moreover, sialylation seems to protect to some extent the Arg11-Ile12 bond from in situ hydrolysis, thus preventing the harmful precipitation of the C-terminal polypeptide in the pancreatic ducts.
Notes: De Reggi, M
Capon, C
Gharib, B
Wieruszeski, J M
Michel, R
Fournet, B
Journal Article
England
Eur J Biochem. 1995 Jun 1;230(2):503-10. doi: 10.1111/j.1432-1033.1995.tb20589.x.
Author Address: INSERM U399, Faculté de Médecine, Marseille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1422
Author: Deane, J. E., Graham, S. C., Kim, N. N., Stein, P. E., McNair, R., Cachón-González, M. B., Cox, T. M. and Read, R. J.
Year: 2011
Title: Insights into Krabbe disease from structures of galactocerebrosidase
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 37
Pages: 15169-73
Epub Date: 2011/08/31
Date: Sep 13
Short Title: Insights into Krabbe disease from structures of galactocerebrosidase
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1105639108
PMCID: PMC3174575
Accession Number: 21876145
Keywords: Animals
Binding Sites
Crystallography, X-Ray
Galactosylceramidase/*chemistry/genetics
Galactosylceramides/chemistry/metabolism
HEK293 Cells
Humans
Leukodystrophy, Globoid Cell/*enzymology/genetics
Mice
Models, Molecular
Mutation/genetics
Protein Structure, Secondary
Substrate Specificity
Abstract: Krabbe disease is a devastating neurodegenerative disease characterized by widespread demyelination that is caused by defects in the enzyme galactocerebrosidase (GALC). Disease-causing mutations have been identified throughout the GALC gene. However, a molecular understanding of the effect of these mutations has been hampered by the lack of structural data for this enzyme. Here we present the crystal structures of GALC and the GALC-product complex, revealing a novel domain architecture with a previously uncharacterized lectin domain not observed in other hydrolases. All three domains of GALC contribute residues to the substrate-binding pocket, and disease-causing mutations are widely distributed throughout the protein. Our structures provide an essential insight into the diverse effects of pathogenic mutations on GALC function in human Krabbe variants and a compelling explanation for the severity of many mutations associated with fatal infantile disease. The localization of disease-associated mutations in the structure of GALC will facilitate identification of those patients that would be responsive to pharmacological chaperone therapies. Furthermore, our structure provides the atomic framework for the design of such drugs.
Notes: 1091-6490
Deane, Janet E
Graham, Stephen C
Kim, Nee Na
Stein, Penelope E
McNair, Rosamund
Cachón-González, M Begoña
Cox, Timothy M
Read, Randy J
082961/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Sep 13;108(37):15169-73. doi: 10.1073/pnas.1105639108. Epub 2011 Aug 29.
Author Address: Department of Haematology, Cambridge Institute for Medical Research, University of Cambridge, Addenbrooke's Hospital, Hills Road, Cambridge CB2 0XY, United Kingdom. jed55@cam.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1344
Author: Deans, M. R., Peterson, J. M. and Wong, G. W.
Year: 2010
Title: Mammalian Otolin: a multimeric glycoprotein specific to the inner ear that interacts with otoconial matrix protein Otoconin-90 and Cerebellin-1
Journal: PLoS One
Volume: 5
Issue: 9
Pages: e12765
Epub Date: 2010/09/22
Date: Sep 15
Short Title: Mammalian Otolin: a multimeric glycoprotein specific to the inner ear that interacts with otoconial matrix protein Otoconin-90 and Cerebellin-1
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0012765
PMCID: PMC2939893
Accession Number: 20856818
Keywords: Amino Acid Sequence
Animals
Calcium-Binding Proteins
Cloning, Molecular
Ear, Inner/chemistry/*metabolism
Extracellular Matrix Proteins/chemistry/genetics/*metabolism
Gene Expression Regulation
HEK293 Cells
Humans
Mammals/genetics/metabolism
Mice/genetics/*metabolism
Molecular Sequence Data
Nerve Tissue Proteins/genetics/*metabolism
Oncorhynchus keta
Protein Binding
Protein Multimerization
Protein Precursors/genetics/*metabolism
Sequence Alignment
Abstract: BACKGROUND: The mammalian otoconial membrane is a dense extracellular matrix containing bio-mineralized otoconia. This structure provides the mechanical stimulus necessary for hair cells of the vestibular maculae to respond to linear accelerations and gravity. In teleosts, Otolin is required for the proper anchoring of otolith crystals to the sensory maculae. Otoconia detachment and subsequent entrapment in the semicircular canals can result in benign paroxysmal positional vertigo (BPPV), a common form of vertigo for which the molecular basis is unknown. Several cDNAs encoding protein components of the mammalian otoconia and otoconial membrane have recently been identified, and mutations in these genes result in abnormal otoconia formation and balance deficits. PRINCIPAL FINDINGS: Here we describe the cloning and characterization of mammalian Otolin, a protein constituent of otoconia and the otoconial membrane. Otolin is a secreted glycoprotein of ∼70 kDa, with a C-terminal globular domain that is homologous to the immune complement C1q, and contains extensive posttranslational modifications including hydroxylated prolines and glycosylated lysines. Like all C1q/TNF family members, Otolin multimerizes into higher order oligomeric complexes. The expression of otolin mRNA is restricted to the inner ear, and immunohistochemical analysis identified Otolin protein in support cells of the vestibular maculae and semi-circular canal cristae. Additionally, Otolin forms protein complexes with Cerebellin-1 and Otoconin-90, two protein constituents of the otoconia, when expressed in vitro. Otolin was also found in subsets of support cells and non-sensory cells of the cochlea, suggesting that Otolin is also a component of the tectorial membrane. CONCLUSION: Given the importance of Otolin in lower organisms, the molecular cloning and biochemical characterization of the mammalian Otolin protein may lead to a better understanding of otoconial development and vestibular dysfunction.
Notes: 1932-6203
Deans, Michael R
Peterson, Jonathan M
Wong, G William
R03 DC009490/DC/NIDCD NIH HHS/United States
F32 DK084607/DK/NIDDK NIH HHS/United States
P60DK079637/DK/NIDDK NIH HHS/United States
P60 DK079637/DK/NIDDK NIH HHS/United States
F32DK084607/DK/NIDDK NIH HHS/United States
DK084171/DK/NIDDK NIH HHS/United States
R01 DK084171/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
PLoS One. 2010 Sep 15;5(9):e12765. doi: 10.1371/journal.pone.0012765.
Author Address: Department of Otolaryngology, Head and Neck Surgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2015
Author: Debarnot, C., Monneau, Y. R., Roig-Zamboni, V., Delauzun, V., Le Narvor, C., Richard, E., Hénault, J., Goulet, A., Fadel, F., Vivès, R. R., Priem, B., Bonnaffé, D., Lortat-Jacob, H. and Bourne, Y.
Year: 2019
Title: Substrate binding mode and catalytic mechanism of human heparan sulfate d-glucuronyl C5 epimerase
Journal: Proc Natl Acad Sci U S A
Volume: 116
Issue: 14
Pages: 6760-6765
Epub Date: 2019/03/16
Date: Apr 2
Short Title: Substrate binding mode and catalytic mechanism of human heparan sulfate d-glucuronyl C5 epimerase
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1818333116
PMCID: PMC6452739
Accession Number: 30872481
Keywords: Binding Sites
Carbohydrate Epimerases/*chemistry/genetics
Catalysis
Crystallography, X-Ray
Glucuronic Acid/*chemistry
HEK293 Cells
Heparin/*chemistry
Humans
Oligosaccharides/*chemistry
Structure-Activity Relationship
Substrate Specificity
*C5 epimerization
*X-ray crystallography
*catalytic mechanism
*heparan sulfate
*substrate distortion
Abstract: Heparan sulfate (HS) is a linear, complex polysaccharide that modulates the biological activities of proteins through binding sites made by a series of Golgi-localized enzymes. Of these, glucuronyl C5-epimerase (Glce) catalyzes C5-epimerization of the HS component, d-glucuronic acid (GlcA), into l-iduronic acid (IdoA), which provides internal flexibility to the polymer and forges protein-binding sites to ensure polymer function. Here we report crystal structures of human Glce in the unbound state and of an inactive mutant, as assessed by real-time NMR spectroscopy, bound with a (GlcA-GlcNS)(n) substrate or a (IdoA-GlcNS)(n) product. Deep infiltration of the oligosaccharides into the active site cleft imposes a sharp kink within the central GlcNS-GlcA/IdoA-GlcNS trisaccharide motif. An extensive network of specific interactions illustrates the absolute requirement of N-sulfate groups vicinal to the epimerization site for substrate binding. At the epimerization site, the GlcA/IdoA rings are highly constrained in two closely related boat conformations, highlighting ring-puckering signatures during catalysis. The structure-based mechanism involves the two invariant acid/base residues, Glu499 and Tyr578, poised on each side of the target uronic acid residue, thus allowing reversible abstraction and readdition of a proton at the C5 position through a neutral enol intermediate, reminiscent of mandelate racemase. These structures also shed light on a convergent mechanism of action between HS epimerases and lyases and provide molecular frameworks for the chemoenzymatic synthesis of heparin or HS analogs.
Notes: 1091-6490
Debarnot, Claire
Monneau, Yoan R
Roig-Zamboni, Véronique
Delauzun, Vincent
Le Narvor, Christine
Richard, Emeline
Hénault, Jérôme
Goulet, Adeline
Fadel, Firas
Vivès, Romain R
Priem, Bernard
Bonnaffé, David
Lortat-Jacob, Hugues
Bourne, Yves
Orcid: 0000-0003-3850-0548
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2019 Apr 2;116(14):6760-6765. doi: 10.1073/pnas.1818333116. Epub 2019 Mar 14.
Author Address: Architecture et Fonction des Macromolécules Biologiques, CNRS, Aix-Marseille University, 13288 Marseille, France.
Institut de Biologie Structurale, University Grenoble Alpes, Commissariat à l'Énergie Atomique et aux Énergies Alternatives, CNRS, 38044 Grenoble, France.
Institut de Chimie Moléculaire et des Matériaux d'Orsay, University Paris Sud, CNRS, Université Paris-Saclay, 91405 Orsay, France.
Centre de Recherches sur les Macromolécules Végétales, CNRS, University Grenoble Alpes, 38041 Grenoble, France.
Architecture et Fonction des Macromolécules Biologiques, CNRS, Aix-Marseille University, 13288 Marseille, France; Yves.Bourne@afmb.univ-mrs.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1131
Author: Debela, M., Goettig, P., Magdolen, V., Huber, R., Schechter, N. M. and Bode, W.
Year: 2007
Title: Structural basis of the zinc inhibition of human tissue kallikrein 5
Journal: J Mol Biol
Volume: 373
Issue: 4
Pages: 1017-31
Epub Date: 2007/09/21
Date: Nov 2
Short Title: Structural basis of the zinc inhibition of human tissue kallikrein 5
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2007.08.042
Accession Number: 17881000
Keywords: Amino Acid Sequence
Binding Sites
Crystallography, X-Ray/methods
Humans
Kallikreins/*chemistry/genetics/*metabolism
Kinetics
Leupeptins/metabolism
Models, Molecular
Molecular Sequence Data
Protein Structure, Secondary
Protein Structure, Tertiary
Sequence Homology, Amino Acid
Substrate Specificity
Zinc/chemistry/*metabolism
Abstract: Human kallikrein 5 (hK5) is a member of the tissue kallikrein family of serine peptidases. It has trypsin-like substrate specificity, is inhibited by metal ions, and is abundantly expressed in human skin, where it is believed to play a central role in desquamation. To further understand the interaction of hK5 with substrates and metal ions, active recombinant hK5 was crystallized in complex with the tripeptidyl aldehyde inhibitor leupeptin, and structures at 2.3 A resolution were obtained with and without Zn2+. While the overall structure and the specificity of S1 pocket for basic side-chains were similar to that of hK4, a closely related family member, both differed in their interaction with Zn2+. Unlike hK4, the 75-loop of hK5 is not structured to bind a Zn2+. Instead, Zn2+ binds adjacent to the active site, becoming coordinated by the imidazole rings of His99 and His96 not present in hK4. This zinc binding is accompanied by a large shift in the backbone conformation of the 99-loop and by large movements of both His side-chains. Modeling studies show that in the absence of bound leupeptin, Zn2+ is likely further coordinated by the imidazolyl side-chain of the catalytic His57 which can, similar to equivalent His57 imidazole groups in the related rat kallikrein proteinase tonin and in an engineered metal-binding rat trypsin, rotate out of its triad position to provide the third co-ordination site of the bound Zn2+, rendering Zn2+-bound hK5 inactive. In solution, this mode of binding likely occurs in the presence of free and substrate saturated hK5, as kinetic analyses of Zn2+ inhibition indicate a non-competitive mechanism. Supporting the His57 re-orientation, Zn2+ does not fully inhibit hK5 hydrolysis of tripeptidyl substrates containing a P2-His residue. The P2 and His57 imidazole groups would lie next to each other in the enzyme-substrate complex, indicating that incomplete inhibition is due to competition between both imidazole groups for Zn2+. The His96-99-57 triad is thus suggested to be responsible for the Zn2+-mediated inhibition of hK5 catalysis.
Notes: Debela, Mekdes
Goettig, Peter
Magdolen, Viktor
Huber, Robert
Schechter, Norman M
Bode, Wolfram
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2007 Nov 2;373(4):1017-31. doi: 10.1016/j.jmb.2007.08.042. Epub 2007 Aug 24.
Author Address: Max-Planck-Institut für Biochemie, Proteinase Research Group, Am Klopferspitz 18, 82152 Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1605
Author: Deeb, S. J., Cox, J., Schmidt-Supprian, M. and Mann, M.
Year: 2014
Title: N-linked glycosylation enrichment for in-depth cell surface proteomics of diffuse large B-cell lymphoma subtypes
Journal: Mol Cell Proteomics
Volume: 13
Issue: 1
Pages: 240-51
Epub Date: 2013/11/06
Date: Jan
Short Title: N-linked glycosylation enrichment for in-depth cell surface proteomics of diffuse large B-cell lymphoma subtypes
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M113.033977
PMCID: PMC3879617
Accession Number: 24190977
Keywords: Gene Expression Regulation, Neoplastic
Glycoproteins/*biosynthesis
Glycosylation
Humans
Lymphoma, Large B-Cell, Diffuse/*genetics/pathology
Neoplasm Proteins/*biosynthesis
*Proteomics
Abstract: Global analysis of lymphoma genome integrity and transcriptomes tremendously advanced our understanding of their biology. Technological advances in mass spectrometry-based proteomics promise to complete the picture by allowing the global quantification of proteins and their post-translational modifications. Here we use N-glyco FASP, a recently developed mass spectrometric approach using lectin-enrichment, in conjunction with a super-SILAC approach to quantify N-linked glycoproteins in lymphoma cells. From patient-derived diffuse large B-cell lymphoma cell lines, we mapped 2383 glycosites on 1321 protein groups, which were highly enriched for cell membrane proteins. This N-glyco subproteome alone allowed the segregation of the ABC from the GCB subtypes of diffuse large B-cell lymphoma, which before gene expression studies had been considered one disease entity. Encouragingly, many of the glycopeptides driving the segregation belong to proteins previously characterized as segregators in a deep proteome study of these subtypes (S. J. Deeb et al. MCP 2012 PMID 22442255). This conforms to the high correlation that we observed between the expression level of the glycosites and their corresponding proteins. Detailed examination of glycosites and glycoprotein expression levels uncovered, among other interesting findings, enrichment of transcription factor binding motifs, including known NF-kappa-B related ones. Thus, enrichment of a class of post-translationally modified peptides can classify cancer types as well as reveal cancer specific mechanistic changes.
Notes: 1535-9484
Deeb, Sally J
Cox, Juergen
Schmidt-Supprian, Marc
Mann, Matthias
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2014 Jan;13(1):240-51. doi: 10.1074/mcp.M113.033977. Epub 2013 Nov 4.
Author Address: Proteomics and Signal Transduction Department, Max-Planck Institute of Biochemistry, D-82152 Martinsried, Germany;
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1762
Author: Deen, A. J., Arasu, U. T., Pasonen-Seppänen, S., Hassinen, A., Takabe, P., Wojciechowski, S., Kärnä, R., Rilla, K., Kellokumpu, S., Tammi, R., Tammi, M. and Oikari, S.
Year: 2016
Title: UDP-sugar substrates of HAS3 regulate its O-GlcNAcylation, intracellular traffic, extracellular shedding and correlate with melanoma progression
Journal: Cell Mol Life Sci
Volume: 73
Issue: 16
Pages: 3183-204
Epub Date: 2016/02/18
Date: Aug
Short Title: UDP-sugar substrates of HAS3 regulate its O-GlcNAcylation, intracellular traffic, extracellular shedding and correlate with melanoma progression
Alternate Journal: Cellular and molecular life sciences : CMLS
ISSN: 1420-682x
DOI: 10.1007/s00018-016-2158-5
Accession Number: 26883802
Keywords: Acetylglucosamine/metabolism
Acylation
Animals
COS Cells
Cell Line
Cell Line, Tumor
Chlorocebus aethiops
Disease Progression
Endocytosis
Glucuronosyltransferase/*metabolism
Humans
Hyaluronan Synthases
Hyaluronic Acid/*metabolism
Melanoma/*metabolism/pathology
Protein Transport
Skin/*metabolism/pathology
Skin Neoplasms/*metabolism/pathology
Uridine Diphosphate N-Acetylglucosamine/metabolism
Uridine Diphosphate Sugars/*metabolism
*4mu
*Gnpda
*Glucosamine
*Mannose
*Ogt
*Ugdh
Abstract: Hyaluronan content is a powerful prognostic factor in many cancer types, but the molecular basis of its synthesis in cancer still remains unclear. Hyaluronan synthesis requires the transport of hyaluronan synthases (HAS1-3) from Golgi to plasma membrane (PM), where the enzymes are activated. For the very first time, the present study demonstrated a rapid recycling of HAS3 between PM and endosomes, controlled by the cytosolic levels of the HAS substrates UDP-GlcUA and UDP-GlcNAc. Depletion of UDP-GlcNAc or UDP-GlcUA shifted the balance towards HAS3 endocytosis, and inhibition of hyaluronan synthesis. In contrast, UDP-GlcNAc surplus suppressed endocytosis and lysosomal decay of HAS3, favoring its retention in PM, stimulating hyaluronan synthesis, and HAS3 shedding in extracellular vesicles. The concentration of UDP-GlcNAc also controlled the level of O-GlcNAc modification of HAS3. Increasing O-GlcNAcylation reproduced the effects of UDP-GlcNAc surplus on HAS3 trafficking, while its suppression showed the opposite effects, indicating that O-GlcNAc signaling is associated to UDP-GlcNAc supply. Importantly, a similar correlation existed between the expression of GFAT1 (the rate limiting enzyme in UDP-GlcNAc synthesis) and hyaluronan content in early and deep human melanomas, suggesting the association of UDP-sugar metabolism in initiation of melanomagenesis. In general, changes in glucose metabolism, realized through UDP-sugar contents and O-GlcNAc signaling, are important in HAS3 trafficking, hyaluronan synthesis, and correlates with melanoma progression.
Notes: 1420-9071
Deen, Ashik Jawahar
Arasu, Uma Thanigai
Pasonen-Seppänen, Sanna
Hassinen, Antti
Takabe, Piia
Wojciechowski, Sara
Kärnä, Riikka
Rilla, Kirsi
Kellokumpu, Sakari
Tammi, Raija
Tammi, Markku
Oikari, Sanna
Journal Article
Switzerland
Cell Mol Life Sci. 2016 Aug;73(16):3183-204. doi: 10.1007/s00018-016-2158-5. Epub 2016 Feb 16.
Author Address: Institute of Biomedicine, School of Medicine, University of Eastern Finland, 70210, Kuopio, Finland. ashik.jawahardeen@uef.fi.
Institute of Biomedicine, School of Medicine, University of Eastern Finland, 70210, Kuopio, Finland.
Faculty of Biochemistry and Molecular Medicine, University of Oulu, 90014, Oulu, Finland.
A. I. Virtanen Institute for Molecular Sciences, School of Medicine, University of Eastern Finland, 70210, Kuopio, Finland.
Institute of Biomedicine, School of Medicine, University of Eastern Finland, 70210, Kuopio, Finland. sanna.oikari@uef.fi.
Institute of Dentistry, School of Medicine, University of Eastern Finland, 70210, Kuopio, Finland. sanna.oikari@uef.fi.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 925
Author: Dela Cruz, C. S., Lee, Y., Viswanathan, S. R., El-Guindy, A. S., Gerlach, J., Nikiforow, S., Shedd, D., Gradoville, L. and Miller, G.
Year: 2004
Title: N-linked glycosylation is required for optimal function of Kaposi's sarcoma herpesvirus-encoded, but not cellular, interleukin 6
Journal: J Exp Med
Volume: 199
Issue: 4
Pages: 503-14
Epub Date: 2004/02/19
Date: Feb 16
Short Title: N-linked glycosylation is required for optimal function of Kaposi's sarcoma herpesvirus-encoded, but not cellular, interleukin 6
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.20031205
PMCID: PMC2211829
Accession Number: 14970177
Keywords: B-Lymphocytes/immunology
Cell Division/immunology
Cell Line
Cell Line, Tumor
Cloning, Molecular
Contactins
Escherichia coli/genetics
Glycosylation
Herpesvirus 8, Human/*immunology
Humans
Interleukin-6/genetics/*immunology
Lymphocyte Activation/immunology
Neural Cell Adhesion Molecules/immunology
Peptide Fragments/chemistry
Plasmids
Recombinant Proteins/immunology
Abstract: Kaposi's sarcoma-associated herpesvirus interleukin-6 (vIL-6) is a structural and functional homologue of the human cytokine IL-6 (hIL-6). hIL-6 and vIL-6 exhibit similar biological functions and both act via the gp130 receptor subunit to activate the Janus tyrosine kinase (JAK)1 and signal transducer and activator of transcription (STAT)1/3 pathway. Here we show that vIL-6 is N-linked glycosylated at N78 and N89 and demonstrate that N-linked glycosylation at site N89 of vIL-6 markedly enhances binding to gp130, signaling through the JAK1-STAT1/3 pathway and functions in a cytokine-dependent cell proliferation bioassay. Although hIL-6 is also N-glycosylated at N73 and multiply O-glycosylated, neither N-linked nor O-linked glycosylation is necessary for IL-6 receptor alpha-dependent binding to gp130 or signaling through JAK1-STAT1/3. As distinct from vIL-6, unglycosylated hIL-6 is as potent as glycosylated hIL-6 in stimulating B cell proliferation. These findings highlight distinct functional roles of N-linked glycosylation in viral and cellular IL-6.
Notes: 1540-9538
Dela Cruz, Charles S
Lee, Yoomi
Viswanathan, Srinivas R
El-Guindy, Ayman S
Gerlach, Jennifer
Nikiforow, Sarah
Shedd, Duane
Gradoville, Lyn
Miller, George
R01 CA070036/CA/NCI NIH HHS/United States
CA70036/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
J Exp Med. 2004 Feb 16;199(4):503-14. doi: 10.1084/jem.20031205.
Author Address: Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, CT 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 959
Author: Delker, S. L., West, A. P., Jr., McDermott, L., Kennedy, M. W. and Bjorkman, P. J.
Year: 2004
Title: Crystallographic studies of ligand binding by Zn-alpha2-glycoprotein
Journal: J Struct Biol
Volume: 148
Issue: 2
Pages: 205-13
Epub Date: 2004/10/13
Date: Nov
Short Title: Crystallographic studies of ligand binding by Zn-alpha2-glycoprotein
Alternate Journal: Journal of structural biology
ISSN: 1047-8477 (Print)
1047-8477
DOI: 10.1016/j.jsb.2004.04.009
Accession Number: 15477100
Keywords: Adipokines
Binding Sites
Carrier Proteins/*chemistry/*genetics
Crystallography, X-Ray/methods
Dansyl Compounds/chemistry
Fatty Acids/chemistry
Fluorescent Dyes/chemistry
Glycoproteins/*chemistry/*genetics
Glycosylation
Humans
Ligands
Models, Molecular
Peptides/chemistry
Polyethylene Glycols/chemistry
Protein Structure, Secondary
Recombinant Proteins/chemistry
Zinc/*chemistry
Abstract: Zn-alpha2-glycoprotein (ZAG) is a 41 kDa soluble protein that is present in most bodily fluids. The previously reported 2.8 A crystal structure of ZAG isolated from human serum demonstrated the structural similarity between ZAG and class I major histocompatibility complex (MHC) molecules and revealed a non-peptidic ligand in the ZAG counterpart of the MHC peptide-binding groove. Here we present crystallographic studies to explore further the nature of the non-peptidic ligand in the ZAG groove. Comparison of the structures of several forms of recombinant ZAG, including a 1.95 A structure derived from ZAG expressed in insect cells, suggests that the non-peptidic ligand in the current structures and in the structure of serum ZAG is a polyethylene glycol (PEG), which is present in the crystallization conditions used. Further support for PEG binding in the ZAG groove is provided by the finding that PEG displaces a fluorophore-tagged fatty acid from the ZAG binding site. From these results we hypothesize that our purified forms of ZAG do not contain a bound endogenous ligand, but that the ZAG groove is capable of binding hydrophobic molecules, which may relate to its function.
Notes: Delker, Silvia L
West, Anthony P Jr
McDermott, Lindsay
Kennedy, Malcolm W
Bjorkman, Pamela J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Struct Biol. 2004 Nov;148(2):205-13. doi: 10.1016/j.jsb.2004.04.009.
Author Address: Division of Biology, 114-96 Pasadena, CA 91125, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 915
Author: Dell, A., Chalabi, S., Easton, R. L., Haslam, S. M., Sutton-Smith, M., Patankar, M. S., Lattanzio, F., Panico, M., Morris, H. R. and Clark, G. F.
Year: 2003
Title: Murine and human zona pellucida 3 derived from mouse eggs express identical O-glycans
Journal: Proc Natl Acad Sci U S A
Volume: 100
Issue: 26
Pages: 15631-6
Epub Date: 2003/12/16
Date: Dec 23
Short Title: Murine and human zona pellucida 3 derived from mouse eggs express identical O-glycans
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.2635507100
PMCID: PMC307619
Accession Number: 14673092
Keywords: Animals
Carbohydrate Sequence
Egg Proteins/*chemistry/genetics/*isolation & purification
Female
Humans
Membrane Glycoproteins/*chemistry/genetics/*isolation & purification
Mice
Mice, Transgenic
Molecular Sequence Data
Oligopeptides/chemistry
Ovary/cytology/physiology
Ovum/*physiology
Polysaccharides/*chemistry/isolation & purification
*Receptors, Cell Surface
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Sperm-Ovum Interactions
Zona Pellucida Glycoproteins
Abstract: Murine sperm initiate fertilization by binding to the outer covering of the egg known as the murine zona pellucida (mZP). This binding is thought to require the interaction of O-glycans linked to a specific mZP glycoprotein (mZP3) with egg-binding proteins coating the sperm plasma membrane. The precise molecular basis of this interaction remains to be resolved. In this study, we analyzed the O-glycosylation of the individual mZP glycoproteins by using ultrasensitive MS methods. We found that the majority of the O-glycans that are linked to mZP3 are core type 2 sequences terminated with sialic acid, lacNAc (Galbeta1-4GlcNAc), lacdiNAc (Gal-NAcbeta1-4GlcNAc), Galalpha1-3Gal, and NeuAcalpha2-3[GalNAcbeta1-4]Galbeta1-4 (Sda antigen). Many of these terminal sequences have been implicated previously in murine sperm-egg binding. Core type 1 O-glycans are also present and are generally unmodified, although some are terminated with sialic acid, beta-linked N-acetylhexosamine, or NeuAcalpha2-3[GalNAcbeta1-4]Galbeta1-4. Eggs expressing human ZP (huZP) glycoprotein huZP3, derived from transgenic mice, bind murine but not human sperm, implying that huZP3 acquires the same O-glycans as native mZP3. Sequencing of huZP3-associated O-glycans confirms that this implication is correct. The data obtained in this investigation may prove to be very useful for studies to determine the precise molecular basis of initial murine sperm-egg binding.
Notes: 1091-6490
Dell, Anne
Chalabi, Sara
Easton, Richard L
Haslam, Stuart M
Sutton-Smith, Mark
Patankar, Manish S
Lattanzio, Frank
Panico, Maria
Morris, Howard R
Clark, Gary F
WT_/Wellcome Trust/United Kingdom
R01 HD035652/HD/NICHD NIH HHS/United States
HD 35652/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2003 Dec 23;100(26):15631-6. doi: 10.1073/pnas.2635507100. Epub 2003 Dec 12.
Author Address: Department of Biological Sciences, Imperial College London, London SW7 2AZ, United Kingdom. a.dell@ic.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 516
Author: Dell, A., Morris, H. R., Easton, R. L., Panico, M., Patankar, M., Oehniger, S., Koistinen, R., Koistinen, H., Seppala, M. and Clark, G. F.
Year: 1995
Title: Structural analysis of the oligosaccharides derived from glycodelin, a human glycoprotein with potent immunosuppressive and contraceptive activities
Journal: J Biol Chem
Volume: 270
Issue: 41
Pages: 24116-26
Epub Date: 1995/10/13
Date: Oct 13
Short Title: Structural analysis of the oligosaccharides derived from glycodelin, a human glycoprotein with potent immunosuppressive and contraceptive activities
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.270.41.24116
Accession Number: 7592613
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
*Contraceptive Agents
Cyanogen Bromide
Epitopes/analysis/chemistry
Gas Chromatography-Mass Spectrometry
Glycodelin
*Glycoproteins
Humans
*Immunosuppressive Agents
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/chemistry
Pregnancy Proteins/*chemistry
Spectrometry, Mass, Fast Atom Bombardment
Abstract: Glycodelin, also known as placental protein 14 (PP14) or progesterone-associated endometrial protein (PAEP), is a human glycoprotein with potent immunosuppressive and contraceptive activities. In this paper we report the first characterization of glycodelin-derived oligosaccharides. Using strategies based upon fast atom bombardment and electrospray mass spectrometry we have established that glycodelin is glycosylated at Asn-28 and Asn-63. The Asn-28 site carries high mannose, hybrid and complex-type structures, whereas the second site is exclusively occupied by complex-type glycans. The major non-reducing epitopes in the complex-type glycans are: Gal beta 1-4GlcNAc (lacNAc), GalNAc beta 1-4GlcNAc (lacdiNAc), NeuAc alpha 2-6Gal beta 1-4GlcNAc (sialylated lacNAc), NeuAc alpha 2-6Gal beta 1-4GlcNAc (sialylated lacdiNAc), Gal beta 1-4(Fuc alpha 1-3)GlcNAc (Lewisx), and GalNAc beta 1-4(Fuc alpha 1-3)GlcNAc (lacdiNAc analogue of Lewisx). It is possible that the oligosaccharides bearing sialylated lacNAc or lacdiNAc antennae may manifest immunosuppressive effects by specifically blocking adhesive and activation-related events mediated by CD22, the human B cell associated receptor. Oligosaccharides with fucosylated lacdiNAc antennae have previously been shown to potently block selectin-mediated adhesions and may perform the same function in glycodelin. The potent inhibitory effect of glycodelin on initial human sperm-zona pellucida binding is consistent with our previous suggestion that this cell adhesion event requires a selectin-like adhesion process. This result also raises the possibility that a convergence between immune and gamete recognition processes may have occurred in the types of carbohydrate ligands recognized in the human.
Notes: Dell, A
Morris, H R
Easton, R L
Panico, M
Patankar, M
Oehniger, S
Koistinen, R
Koistinen, H
Seppala, M
Clark, G F
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1995 Oct 13;270(41):24116-26. doi: 10.1074/jbc.270.41.24116.
Author Address: Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1120
Author: Dellisanti, C. D., Yao, Y., Stroud, J. C., Wang, Z. Z. and Chen, L.
Year: 2007
Title: Crystal structure of the extracellular domain of nAChR alpha1 bound to alpha-bungarotoxin at 1.94 A resolution
Journal: Nat Neurosci
Volume: 10
Issue: 8
Pages: 953-62
Epub Date: 2007/07/24
Date: Aug
Short Title: Crystal structure of the extracellular domain of nAChR alpha1 bound to alpha-bungarotoxin at 1.94 A resolution
Alternate Journal: Nature neuroscience
ISSN: 1097-6256 (Print)
1097-6256
DOI: 10.1038/nn1942
Accession Number: 17643119
Keywords: Acetylcholine/pharmacology
Animals
Binding Sites/drug effects/genetics
Bungarotoxins/chemistry/*metabolism
COS Cells
Chlorocebus aethiops
Cryoelectron Microscopy
Membrane Potentials/drug effects/genetics/radiation effects
Mice
Models, Molecular
Muscles/chemistry
Mutation
Nuclear Magnetic Resonance, Biomolecular
Patch-Clamp Techniques
Protein Structure, Tertiary
Receptors, Nicotinic/biosynthesis/*metabolism/ultrastructure
Recombinant Proteins/genetics/metabolism
Sequence Homology, Amino Acid
Transfection
Abstract: We determined the crystal structure of the extracellular domain of the mouse nicotinic acetylcholine receptor (nAChR) alpha1 subunit bound to alpha-bungarotoxin at 1.94 A resolution. This structure is the first atomic-resolution view of a nAChR subunit extracellular domain, revealing receptor-specific features such as the main immunogenic region (MIR), the signature Cys-loop and the N-linked carbohydrate chain. The toxin binds to the receptor through extensive protein-protein and protein-sugar interactions. To our surprise, the structure showed a well-ordered water molecule and two hydrophilic residues deep in the core of the alpha1 subunit. The two hydrophilic core residues are highly conserved in nAChRs, but correspond to hydrophobic residues in the nonchannel homolog acetylcholine-binding proteins. We carried out site-directed mutagenesis and electrophysiology analyses to assess the functional role of the glycosylation and the hydrophilic core residues. Our structural and functional studies show essential features of the nAChR and provide new insights into the gating mechanism.
Notes: Dellisanti, Cosma D
Yao, Yun
Stroud, James C
Wang, Zuo-Zhong
Chen, Lin
R01 GM064642/GM/NIGMS NIH HHS/United States
R01 GM077320/GM/NIGMS NIH HHS/United States
R01 HL076334/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Nat Neurosci. 2007 Aug;10(8):953-62. doi: 10.1038/nn1942. Epub 2007 Jul 22.
Author Address: Molecular and Computation Biology, University of Southern California, 1050 Childs Way, RIH201, Los Angeles, California 90089-2910, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1159
Author: Dentin, R., Hedrick, S., Xie, J., Yates, J., 3rd and Montminy, M.
Year: 2008
Title: Hepatic glucose sensing via the CREB coactivator CRTC2
Journal: Science
Volume: 319
Issue: 5868
Pages: 1402-5
Epub Date: 2008/03/08
Date: Mar 7
Short Title: Hepatic glucose sensing via the CREB coactivator CRTC2
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1151363
Accession Number: 18323454
Keywords: Amino Acid Substitution
Animals
Blood Glucose/metabolism
Cell Nucleus/metabolism
Cells, Cultured
Cyclic AMP Response Element-Binding Protein/metabolism
Cytoplasm/metabolism
Diabetes Mellitus/metabolism
*Gluconeogenesis
Glucose/*metabolism
Glycosylation
Glycosyltransferases/metabolism
Hepatocytes/metabolism
Humans
Insulin/metabolism
Liver/*metabolism
Male
Mice
Mice, Inbred C57BL
Phosphorylation
RNA Interference
Signal Transduction
Trans-Activators/genetics/*metabolism
Transcription Factors
beta-N-Acetylhexosaminidases/metabolism
Abstract: Chronic hyperglycemia contributes to the development of diabetes-associated complications. Increases in the concentration of circulating glucose activate the hexosamine biosynthetic pathway (HBP) and promote the O-glycosylation of proteins by O-glycosyl transferase (OGT). We show that OGT triggered hepatic gluconeogenesis through the O-glycosylation of the transducer of regulated cyclic adenosine monophosphate response element-binding protein (CREB) 2 (TORC2 or CRTC2). CRTC2 was O-glycosylated at sites that normally sequester CRTC2 in the cytoplasm through a phosphorylation-dependent mechanism. Decreasing amounts of O-glycosylated CRTC2 by expression of the deglycosylating enzyme O-GlcNAcase blocked effects of glucose on gluconeogenesis, demonstrating the importance of the HBP in the development of glucose intolerance.
Notes: 1095-9203
Dentin, Renaud
Hedrick, Susan
Xie, Jianxin
Yates, John 3rd
Montminy, Marc
R37 GM037828/GM/NIGMS NIH HHS/United States
R01 GM037828/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Science. 2008 Mar 7;319(5868):1402-5. doi: 10.1126/science.1151363.
Author Address: Salk Institute for Biological Studies, 10010 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 336
Author: Derisbourg, P., Wieruszeski, J. M., Montreuil, J. and Spik, G.
Year: 1990
Title: Primary structure of glycans isolated from human leucocyte lactotransferrin. Absence of fucose residues questions the proposed mechanism of hyposideraemia
Journal: Biochem J
Volume: 269
Issue: 3
Pages: 821-5
Epub Date: 1990/08/01
Date: Aug 1
Short Title: Primary structure of glycans isolated from human leucocyte lactotransferrin. Absence of fucose residues questions the proposed mechanism of hyposideraemia
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2690821
PMCID: PMC1131661
Accession Number: 2390069
Keywords: Anemia, Hypochromic/blood
Carbohydrate Sequence
Fucose/metabolism
Humans
Hydrazines/metabolism
Lactoferrin/*blood/isolation & purification/metabolism
Lactoglobulins/*blood
Leukocytes/*analysis
Magnetic Resonance Spectroscopy
Mass Spectrometry/methods
Milk, Human/analysis
Molecular Sequence Data
Oligosaccharides/metabolism
Polysaccharides/*blood/isolation & purification
Abstract: Lactotransferrin was highly purified from lysates of human neutrophilic leucocytes by immuno-affinity chromatography. A comparative analysis of the molar carbohydrate compositions of human leucocyte lactotransferrin and human milk lactotransferrin reveals that the glycans of leucocyte lactotransferrin differ essentially by the absence of fucose residues. Structural analysis combining methylation-mass spectrometry and 400 MHz 1H-n.m.r. spectrometry of oligosaccharide alditols released from human leucocyte lactotransferrin shows the presence of two disialylated and non-fucosylated biantennary glycans of the N-acetyl-lactosaminic type. These results question a previously proposed mechanism for hyposideraemia in which the leucocyte lactotransferrin was involved and in which the fucose residues played a key role.
Notes: 1470-8728
Derisbourg, P
Wieruszeski, J M
Montreuil, J
Spik, G
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1990 Aug 1;269(3):821-5. doi: 10.1042/bj2690821.
Author Address: Laboratoire de Chimie Biologique, Université des Sciences et Techniques de Lille Flandres-Artois, Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1253
Author: Desko, M. M., Gross, D. A. and Kohler, J. J.
Year: 2009
Title: Effects of N-glycosylation on the activity and localization of GlcNAc-6-sulfotransferase 1
Journal: Glycobiology
Volume: 19
Issue: 10
Pages: 1068-77
Epub Date: 2009/07/03
Date: Oct
Short Title: Effects of N-glycosylation on the activity and localization of GlcNAc-6-sulfotransferase 1
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwp092
Accession Number: 19571171
Keywords: Amino Acid Sequence
Animals
Carbohydrate Sequence
Epitopes/immunology
Glycosylation
HeLa Cells
Humans
Lewis X Antigen/analogs & derivatives
Mice
Molecular Sequence Data
Mutation/genetics
Oligosaccharides/metabolism
Polysaccharides/metabolism
Sequence Alignment
Sialyl Lewis X Antigen/analogs & derivatives
Sulfotransferases/chemistry/genetics/immunology/*metabolism
Abstract: N-Acetylglucosamine-6-sulfotransferase-1 (GlcNAc6ST-1) is a Golgi-resident glycoprotein that is responsible for sulfation of the l-selectin ligand on endothelial cells. Here, we report the sites at which GlcNAc6ST-1 is modified with N-linked glycans and the effects that each glycan has on enzyme activity, specificity, and localization. We determined that glycans are added at three of four potential N-linked glycosylation sites: N196, N410, and N428. The N428 glycan is required for the production of sulfated cell surface glycans: cells expressing a mutant enzyme lacking this glycan were unable to sulfate the sialyl Lewis X tetrasaccharide or a putative extended core 1 O-linked glycan. The N196 and N410 glycans differentially affect sulfation of two different substrates: cells that express an enzyme lacking the N410 glycan are able to sulfate the sialyl Lewis X substrate, but produce reduced levels of a sulfated peripheral lymph node addressin epitope and cells that express an enzyme lacking the N196 glycan are able to produce a sulfated peripheral lymph node addressin epitope, but are impaired in their ability to sulfate sialyl Lewis X. The glycans' effects on enzyme activity may be mediated, in part, by changes in enzyme localization. While most mutants that lacked glycans localized normally within the Golgi, the N428A mutant and a mutant lacking all glycans were also found to localize ectopically. Altered trafficking of mutants may be associated with the mechanisms by which misglycosylated enzyme is degraded.
Notes: 1460-2423
Desko, Marguerite M
Gross, David A
Kohler, Jennifer J
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2009 Oct;19(10):1068-77. doi: 10.1093/glycob/cwp092. Epub 2009 Jul 1.
Author Address: Department of Chemistry, Stanford University, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1073
Author: Deuschl, F., Kollmann, K., von Figura, K. and Lübke, T.
Year: 2006
Title: Molecular characterization of the hypothetical 66.3-kDa protein in mouse: lysosomal targeting, glycosylation, processing and tissue distribution
Journal: FEBS Lett
Volume: 580
Issue: 24
Pages: 5747-52
Epub Date: 2006/09/30
Date: Oct 16
Short Title: Molecular characterization of the hypothetical 66.3-kDa protein in mouse: lysosomal targeting, glycosylation, processing and tissue distribution
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2006.09.029
Accession Number: 17007843
Keywords: Animals
Cell Line, Tumor
Glycoproteins/genetics/isolation & purification/*metabolism
Glycosylation
Humans
Lysosomes/*metabolism
Mice
Molecular Weight
Organ Specificity
Protein Processing, Post-Translational
Substrate Specificity
Abstract: Recently, we and others identified the 66.3-kDa protein as one of several putative novel lysosomal matrix proteins by analyzing mannose 6-phosphate receptors binding proteins [Kollmann K., Mutenda K.E., Balleininger M., Eckermann E., von Figura K., Schmidt B., Lübke T. (2005) Identification of novel lysosomal matrix proteins by proteome analysis. Proteomics 5(15), 3966-3678, Sleat D.E., Lackland H., Wang Y., Sohar I., Xiao G., Li H., Lobel P. (2005) The human brain mannose 6-phosphate glycoproteome: a complex mixture composed of multiple isoforms of many soluble lysosomal proteins. Proteomics. 5(6), 1520-1532]. Here, we describe the expression of the mouse 66.3-kDa protein in HT1080 cells in which it is synthesized as a precursor of about 75kDa and subsequently processed by limited proteolysis to mature polypeptides accumulating in the lysosomal compartment. The lysosomal localisation of the endogenous 66.3-kDa protein was verified by indirect immunofluorescence in mouse embryonic fibroblasts and by subcellular fractionation of tyloxapol-filled mouse liver lysosomes. Northern blot analysis reveals high transcriptional levels in testis, liver and kidney, whereas Western blot analysis shows high protein levels in brain, heart, lung and spleen. Interestingly, in mouse the endogenous 66.3-kDa protein is processed in a highly tissue-dependent manner to mature forms.
Notes: Deuschl, Florian
Kollmann, Katrin
von Figura, Kurt
Lübke, Torben
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2006 Oct 16;580(24):5747-52. doi: 10.1016/j.febslet.2006.09.029. Epub 2006 Sep 22.
Author Address: Zentrum Biochemie und Molekulare Zellbiologie, Abteilung Biochemie II, Georg-August Universität Göttingen, Heinrich-Düker-Weg 12, D-37073Göttingen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1432
Author: Di Giovine, P., Settembre, E. C., Bhargava, A. K., Luftig, M. A., Lou, H., Cohen, G. H., Eisenberg, R. J., Krummenacher, C. and Carfi, A.
Year: 2011
Title: Structure of herpes simplex virus glycoprotein D bound to the human receptor nectin-1
Journal: PLoS Pathog
Volume: 7
Issue: 9
Pages: e1002277
Epub Date: 2011/10/08
Date: Sep
Short Title: Structure of herpes simplex virus glycoprotein D bound to the human receptor nectin-1
Alternate Journal: PLoS pathogens
ISSN: 1553-7366 (Print)
1553-7366
DOI: 10.1371/journal.ppat.1002277
PMCID: PMC3182920
Accession Number: 21980294
Keywords: Cell Adhesion Molecules/*chemistry/genetics/metabolism
Herpesvirus 1, Human/*chemistry/physiology
Humans
Nectins
Protein Binding
Protein Structure, Quaternary
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Virus/*chemistry/genetics/metabolism
Structure-Activity Relationship
Viral Envelope Proteins/*chemistry/genetics/metabolism
Virus Internalization
Abstract: Binding of herpes simplex virus (HSV) glycoprotein D (gD) to a cell surface receptor is required to trigger membrane fusion during entry into host cells. Nectin-1 is a cell adhesion molecule and the main HSV receptor in neurons and epithelial cells. We report the structure of gD bound to nectin-1 determined by x-ray crystallography to 4.0 Å resolution. The structure reveals that the nectin-1 binding site on gD differs from the binding site of the HVEM receptor. A surface on the first Ig-domain of nectin-1, which mediates homophilic interactions of Ig-like cell adhesion molecules, buries an area composed by residues from both the gD N- and C-terminal extensions. Phenylalanine 129, at the tip of the loop connecting β-strands F and G of nectin-1, protrudes into a groove on gD, which is otherwise occupied by C-terminal residues in the unliganded gD and by N-terminal residues in the gD/HVEM complex. Notably, mutation of Phe129 to alanine prevents nectin-1 binding to gD and HSV entry. Together these data are consistent with previous studies showing that gD disrupts the normal nectin-1 homophilic interactions. Furthermore, the structure of the complex supports a model in which gD-receptor binding triggers HSV entry through receptor-mediated displacement of the gD C-terminal region.
Notes: 1553-7374
Di Giovine, Paolo
Settembre, Ethan C
Bhargava, Arjun K
Luftig, Micah A
Lou, Huan
Cohen, Gary H
Eisenberg, Roselyn J
Krummenacher, Claude
Carfi, Andrea
R01 AI076231-15/AI/NIAID NIH HHS/United States
R56 AI076231/AI/NIAID NIH HHS/United States
R01 AI056045-07/AI/NIAID NIH HHS/United States
R01 AI076231/AI/NIAID NIH HHS/United States
AI-076231/AI/NIAID NIH HHS/United States
AI-073384/AI/NIAID NIH HHS/United States
R01 NS036731/NS/NINDS NIH HHS/United States
R37-AI-18289/AI/NIAID NIH HHS/United States
AI-056045/AI/NIAID NIH HHS/United States
R21 AI073384/AI/NIAID NIH HHS/United States
R01 AI056045/AI/NIAID NIH HHS/United States
R37 AI018289/AI/NIAID NIH HHS/United States
R37 AI018289-31/AI/NIAID NIH HHS/United States
R21 AI056045/AI/NIAID NIH HHS/United States
R37 AI018289-30/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
PLoS Pathog. 2011 Sep;7(9):e1002277. doi: 10.1371/journal.ppat.1002277. Epub 2011 Sep 29.
Author Address: Department of Biochemistry and Molecular Biology, IRBM P. Angeletti, Pomezia, Rome, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1478
Author: Dias, W. B., Cheung, W. D. and Hart, G. W.
Year: 2012
Title: O-GlcNAcylation of kinases
Journal: Biochem Biophys Res Commun
Volume: 422
Issue: 2
Pages: 224-8
Epub Date: 2012/05/09
Date: Jun 1
Short Title: O-GlcNAcylation of kinases
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/j.bbrc.2012.04.124
PMCID: PMC3387735
NIHMSID: NIHMS378885
Accession Number: 22564745
Keywords: Acetylglucosamine/*chemistry
Acylation
HEK293 Cells
Humans
N-Acetylglucosaminyltransferases/chemistry/genetics
Phosphorylation
Phosphotransferases (Alcohol Group Acceptor)/*chemistry/isolation & purification
Protein Array Analysis
Protein Conformation
Recombinant Proteins/chemistry/genetics
Substrate Specificity
Abstract: Recent evidence indicates that site-specific crosstalk between O-GlcNAcylation and phosphorylation and the O-GlcNAcylation of kinases play an important role in regulating cell signaling. However, relatively few kinases have been analyzed for O-GlcNAcylation. Here, we identify additional kinases that are substrates for O-GlcNAcylation using an in vitro OGT assay on a functional kinase array. Forty-two kinases were O-GlcNAcylated in vitro, representing 39% of the kinases on the array. In addition, we confirmed the in vivo O-GlcNAcylation of three identified kinases. Our results suggest that O-GlcNAcylation may directly regulate a substantial number of kinases and illustrates the increasingly complex relationship between O-GlcNAcylation and phosphorylation in cellular signaling.
Notes: 1090-2104
Dias, Wagner B
Cheung, Win D
Hart, Gerald W
R01 DK61671/DK/NIDDK NIH HHS/United States
N01-HV-28180/HV/NHLBI NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
N01 HV028180/HV/NHLBI NIH HHS/United States
N01HV28180/HL/NHLBI NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
R01 CA42486/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Biochem Biophys Res Commun. 2012 Jun 1;422(2):224-8. doi: 10.1016/j.bbrc.2012.04.124. Epub 2012 Apr 30.
Author Address: Department of Biological Chemistry, Johns Hopkins University, School of Medicine, Baltimore, MD 21205, USA. diaswb@biof.ufrj.br
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1246
Author: Dias, W. B., Cheung, W. D., Wang, Z. and Hart, G. W.
Year: 2009
Title: Regulation of calcium/calmodulin-dependent kinase IV by O-GlcNAc modification
Journal: J Biol Chem
Volume: 284
Issue: 32
Pages: 21327-37
Epub Date: 2009/06/10
Date: Aug 7
Short Title: Regulation of calcium/calmodulin-dependent kinase IV by O-GlcNAc modification
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.007310
PMCID: PMC2755857
Accession Number: 19506079
Keywords: Acetylglucosamine/*chemistry
Animals
Calcium-Calmodulin-Dependent Protein Kinase Type 4/*metabolism
Calcium-Calmodulin-Dependent Protein Kinases/*metabolism
Cell Line
Cerebellum/metabolism
*Gene Expression Regulation, Enzymologic
Humans
Models, Biological
Mutagenesis, Site-Directed
Phosphorylation
Protein Structure, Tertiary
Rats
Threonine/chemistry
Abstract: Similar to phosphorylation, GlcNAcylation (the addition of O-GlcNAc to Ser(Thr) residues on polypeptides) is an abundant, dynamic, and inducible post-translational modification. GlcNAcylated proteins are crucial in regulating virtually all cellular processes, including signaling, cell cycle, and transcription. Here we show that calcium/calmodulin-dependent kinase IV (CaMKIV) is highly GlcNAcylated in vivo. In addition, we show that upon activation of HEK293 cells, hemagglutinin-tagged CaMKIV GlcNAcylation rapidly decreases, in a manner directly opposing its phosphorylation at Thr-200. Correspondingly, there is an increase in CaMKIV interaction with O-GlcNAcase during CaMKIV activation. Furthermore, we identify at least five sites of GlcNAcylation on CaMKIV. Using site-directed mutagenesis, we determine that the GlcNAcylation sites located in the active site of CaMKIV can modulate its phosphorylation at Thr-200 and its activity toward cAMP-response element-binding transcription factor. Our results strongly indicate that the O-GlcNAc modification participates in the regulation of CaMKIV activation and function, possibly coordinating nutritional signals with the immune and nervous systems. This is the first example of an O-GlcNAc/phosphate cycle involving O-GlcNAc transferase/kinase cross-talk.
Notes: 1083-351x
Dias, Wagner B
Cheung, Win D
Wang, Zihao
Hart, Gerald W
DK61671/DK/NIDDK NIH HHS/United States
N01-HV-28180/HV/NHLBI NIH HHS/United States
CA42486/CA/NCI NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
N01HV28180/HL/NHLBI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2009 Aug 7;284(32):21327-37. doi: 10.1074/jbc.M109.007310. Epub 2009 Jun 8.
Author Address: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 974
Author: Dickmanns, A., Schmidt, B., Rudolph, M. G., Mariappan, M., Dierks, T., von Figura, K. and Ficner, R.
Year: 2005
Title: Crystal structure of human pFGE, the paralog of the Calpha-formylglycine-generating enzyme
Journal: J Biol Chem
Volume: 280
Issue: 15
Pages: 15180-7
Epub Date: 2005/02/03
Date: Apr 15
Short Title: Crystal structure of human pFGE, the paralog of the Calpha-formylglycine-generating enzyme
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M414317200
Accession Number: 15687489
Keywords: Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Calcium/chemistry
Cations
Cloning, Molecular
Crystallography, X-Ray
Dimerization
Disulfides/chemistry
Humans
Models, Molecular
Molecular Sequence Data
Oxidoreductases Acting on Sulfur Group Donors
Peptides/chemistry
Protein Binding
Protein Conformation
Protein Folding
Protein Processing, Post-Translational
Protein Structure, Secondary
Protein Structure, Tertiary
Sequence Homology, Amino Acid
Sulfatases/*chemistry/physiology
X-Ray Diffraction
Abstract: In eukaryotes, sulfate esters are degraded by sulfatases, which possess a unique Calpha-formylglycine residue in their active site. The defect in post-translational formation of the Calpha-formylglycine residue causes a severe lysosomal storage disorder in humans. Recently, FGE (formylglycine-generating enzyme) has been identified as the protein required for this specific modification. Using sequence comparisons, a protein homologous to FGE was found and denoted pFGE (paralog of FGE). pFGE binds a sulfatase-derived peptide bearing the FGE recognition motif, but it lacks formylglycine-generating activity. Both proteins belong to a large family of pro- and eukaryotic proteins containing the DUF323 domain, a formylglycine-generating enzyme domain of unknown three-dimensional structure. We have crystallized the glycosylated human pFGE and determined its crystal structure at a resolution of 1.86 A. The structure reveals a novel fold, which we denote the FGE fold and which therefore serves as a paradigm for the DUF323 domain. It is characterized by an asymmetric partitioning of secondary structure elements and is stabilized by two calcium cations. A deep cleft on the surface of pFGE most likely represents the sulfatase polypeptide binding site. The asymmetric unit of the pFGE crystal contains a homodimer. The putative peptide binding site is buried between the monomers, indicating a biological significance of the dimer. The structure suggests the capability of pFGE to form a heterodimer with FGE.
Notes: Dickmanns, Achim
Schmidt, Bernhard
Rudolph, Markus G
Mariappan, Malaiyalam
Dierks, Thomas
von Figura, Kurt
Ficner, Ralf
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2005 Apr 15;280(15):15180-7. doi: 10.1074/jbc.M414317200. Epub 2005 Feb 1.
Author Address: Abteilung Molekulare Strukturbiologie, Institut für Mikrobiologie und Genetik, Georg-August-Universität, Justus-von-Liebig Weg 9, D-37077 Göttingen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 988
Author: Dierks, T., Dickmanns, A., Preusser-Kunze, A., Schmidt, B., Mariappan, M., von Figura, K., Ficner, R. and Rudolph, M. G.
Year: 2005
Title: Molecular basis for multiple sulfatase deficiency and mechanism for formylglycine generation of the human formylglycine-generating enzyme
Journal: Cell
Volume: 121
Issue: 4
Pages: 541-552
Epub Date: 2005/05/24
Date: May 20
Short Title: Molecular basis for multiple sulfatase deficiency and mechanism for formylglycine generation of the human formylglycine-generating enzyme
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2005.03.001
Accession Number: 15907468
Keywords: Alanine/analogs & derivatives/biosynthesis/metabolism
Amino Acid Sequence
Binding Sites/physiology
Calcium/metabolism
Catalytic Domain
Crystallography, X-Ray
Cysteine/*analogs & derivatives/metabolism
Glycine/*analogs & derivatives/*biosynthesis/metabolism
Humans
Models, Molecular
Molecular Conformation
Molecular Sequence Data
Mutation, Missense/physiology
Oxidation-Reduction
Oxidoreductases Acting on Sulfur Group Donors
Oxygen/metabolism
Protein Structure, Secondary/physiology
Sequence Homology, Amino Acid
Sphingolipidoses/genetics/*metabolism
Sulfatases/*chemistry/genetics/*metabolism
Tumor Cells, Cultured
Abstract: Sulfatases are enzymes essential for degradation and remodeling of sulfate esters. Formylglycine (FGly), the key catalytic residue in the active site, is unique to sulfatases. In higher eukaryotes, FGly is generated from a cysteine precursor by the FGly-generating enzyme (FGE). Inactivity of FGE results in multiple sulfatase deficiency (MSD), a fatal autosomal recessive syndrome. Based on the crystal structure, we report that FGE is a single-domain monomer with a surprising paucity of secondary structure and adopts a unique fold. The effect of all 18 missense mutations found in MSD patients is explained by the FGE structure, providing a molecular basis of MSD. The catalytic mechanism of FGly generation was elucidated by six high-resolution structures of FGE in different redox environments. The structures allow formulation of a novel oxygenase mechanism whereby FGE utilizes molecular oxygen to generate FGly via a cysteine sulfenic acid intermediate.
Notes: Dierks, Thomas
Dickmanns, Achim
Preusser-Kunze, Andrea
Schmidt, Bernhard
Mariappan, Malaiyalam
von Figura, Kurt
Ficner, Ralf
Rudolph, Markus Georg
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell. 2005 May 20;121(4):541-552. doi: 10.1016/j.cell.2005.03.001.
Author Address: Department of Biochemistry II, University of Göttingen, D-37073 Göttingen, Germany.
Department of Molecular Structural Biology, University of Göttingen, D-37077 Göttingen, Germany.
Department of Biochemistry II, University of Göttingen, D-37073 Göttingen, Germany. Electronic address: kfigura@gwdg.de.
Department of Molecular Structural Biology, University of Göttingen, D-37077 Göttingen, Germany. Electronic address: markus.rudolph@bio.uni-goettingen.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 908
Author: Dietrich, A., Mederos y Schnitzler, M., Emmel, J., Kalwa, H., Hofmann, T. and Gudermann, T.
Year: 2003
Title: N-linked protein glycosylation is a major determinant for basal TRPC3 and TRPC6 channel activity
Journal: J Biol Chem
Volume: 278
Issue: 48
Pages: 47842-52
Epub Date: 2003/09/13
Date: Nov 28
Short Title: N-linked protein glycosylation is a major determinant for basal TRPC3 and TRPC6 channel activity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M302983200
Accession Number: 12970363
Keywords: Amino Acid Motifs
Calcium Channels/*chemistry
Cations
Cell Line
Diglycerides/chemistry
Electrophoresis, Polyacrylamide Gel
Electrophysiology
Glycosylation
Humans
Immunoblotting
Ion Channels/*chemistry
Manganese/metabolism
*Membrane Proteins
Microscopy, Fluorescence
Mutagenesis, Site-Directed
Mutation
Protein Binding
Protein Processing, Post-Translational
Recombinant Fusion Proteins/metabolism
TRPC Cation Channels
TRPC6 Cation Channel
TRPM Cation Channels
Time Factors
Transfection
Tunicamycin/pharmacology
Type C Phospholipases/chemistry
Abstract: The TRPC family of receptor-activated cation channels (TRPC channels) can be subdivided into four subfamilies based on sequence homology as well as functional similarities. Members of the TRPC3/6/7 subfamily share common biophysical characteristics and are activated by diacylglycerol in a membrane-delimited manner. At present, it is only poorly understood whether members of the TRPC3/6/7 subfamily are functionally redundant or whether they serve distinct cellular roles. By electrophysiological and fluorescence imaging strategies we show that TRPC3 displays considerable constitutive activity, while TRPC6 is a tightly regulated channel. To identify potential molecular correlates accounting for the functional difference, we analyzed the glycosylation pattern of TRPC6 compared with TRPC3. Two NX(S/T) motifs in TRPC6 were mutated (Asn to Gln) by in vitro mutagenesis to delete one or both extracellular N-linked glycosylation sites. Immunoblotting analysis of HEK 293 cell lysates expressing TRPC6 wild type and mutants favors a model of TRPC6 that is dually glycosylated within the first (e1) and second extracellular loop (e2) as opposed to the monoglycosylated TRPC3 channel (Vannier, B., Zhu, X., Brown, D., and Birnbaumer, L. (1998) J. Biol. Chem. 273, 8675-8679). Elimination of the e2 glycosylation site, missing in the monoglycosylated TRPC3, was sufficient to convert the tightly receptor-regulated TRPC6 into a constitutively active channel, displaying functional characteristics of TRPC3. Reciprocally, engineering of an additional second glycosylated site in TRPC3 to mimic the glycosylation status in TRPC6 markedly reduced TRPC3 basal activity. We conclude that the glycosylation pattern plays a pivotal role for the tight regulation of TRPC6 through phospholipase C-activating receptors.
Notes: Dietrich, Alexander
Mederos y Schnitzler, Michael
Emmel, Jens
Kalwa, Hermann
Hofmann, Thomas
Gudermann, Thomas
Journal Article
United States
J Biol Chem. 2003 Nov 28;278(48):47842-52. doi: 10.1074/jbc.M302983200. Epub 2003 Sep 11.
Author Address: Institut für Pharmakologie und Toxikologie, Philipps-Universität-Marburg, 35033 Marburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 608
Author: Dietrich, J., Neisig, A., Hou, X., Wegener, A. M., Gajhede, M. and Geisler, C.
Year: 1996
Title: Role of CD3 gamma in T cell receptor assembly
Journal: J Cell Biol
Volume: 132
Issue: 3
Pages: 299-310
Epub Date: 1996/02/01
Date: Feb
Short Title: Role of CD3 gamma in T cell receptor assembly
Alternate Journal: The Journal of cell biology
ISSN: 0021-9525 (Print)
0021-9525
DOI: 10.1083/jcb.132.3.299
PMCID: PMC2120718
Accession Number: 8636209
Keywords: Amino Acid Sequence
Animals
Birds
CD3 Complex/*chemistry/metabolism
Cell Membrane/immunology
Computer Simulation
Cytosol/immunology
Humans
Macromolecular Substances
Mice
Models, Molecular
Models, Structural
Molecular Sequence Data
Protein Folding
*Protein Structure, Secondary
Rats
Receptor-CD3 Complex, Antigen, T-Cell/*biosynthesis/*chemistry
Receptors, Antigen, T-Cell, alpha-beta/chemistry/metabolism
Receptors, Antigen, T-Cell, gamma-delta/*chemistry/metabolism
Sequence Homology, Amino Acid
Sheep
T-Lymphocytes/*immunology
Abstract: The T cell receptor (TCR) consists of the Ti alpha beta heterodimer and the associated CD3 gamma delta epsilon and zeta 2 chains. The structural relationships between the subunits of the TCR complex are still not fully known. In this study we examined the role of the extracellular (EC), transmembrane (TM), and cytoplasmic (CY) domain of CD3 gamma in assembly and cell surface expression of the complete TCR in human T cells. A computer model indicated that the EC domain of CD3 gamma folds as an Ig domain. Based on this model and on alignment studies, two potential interaction sites were predicted in the EC domain of CD3 gamma. Site-directed mutagenesis demonstrated that these sites play a crucial role in TCR assembly probably by binding to CD3 epsilon. Mutagenesis of N-linked glycosylation sites showed that glycosylation of CD3 gamma is not required for TCR assembly and expression. In contrast, treatment of T cells with tunicamycin suggested that N-linked glycosylation of CD3 delta is required for TCR assembly. Site-directed mutagenesis of the acidic amino acid in the TM domain of CD3 gamma demonstrated that this residue is involved in TCR assembly probably by binding to Ti beta. Deletion of the entire CY domain of CD3 gamma did not prevent assembly and expression of the TCR. In conclusion, this study demonstrated that specific TCR interaction sites exist in both the EC and TM domain of CD3 gamma. Furthermore, the study indicated that, in contrast to CD3 gamma, glycosylation of CD3 delta is required for TCR assembly and expression.
Notes: 1540-8140
Dietrich, J
Neisig, A
Hou, X
Wegener, A M
Gajhede, M
Geisler, C
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
J Cell Biol. 1996 Feb;132(3):299-310. doi: 10.1083/jcb.132.3.299.
Author Address: Institute of Medical Microbiology and Immunology, University of Copenhagen, Panum Institute, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 459
Author: Dijkema, R., Pouwels, P., de Reus, A. and Schellekens, H.
Year: 1984
Title: Structure and expression in Escherichia coli of a cloned rat interferon-alpha gene
Journal: Nucleic Acids Res
Volume: 12
Issue: 2
Pages: 1227-42
Epub Date: 1984/01/25
Date: Jan 25
Short Title: Structure and expression in Escherichia coli of a cloned rat interferon-alpha gene
Alternate Journal: Nucleic acids research
ISSN: 0305-1048 (Print)
0305-1048
DOI: 10.1093/nar/12.2.1227
PMCID: PMC318569
Accession Number: 6320120
Keywords: Animals
Base Sequence
*Cloning, Molecular
DNA/metabolism
DNA Replication
DNA Restriction Enzymes
DNA, Recombinant/metabolism
Escherichia coli/*genetics
Female
Gene Amplification
*Genes
Interferon Type I/*genetics
Oocytes/metabolism
RNA, Messenger/genetics
Rats
Transcription, Genetic
Xenopus
Abstract: DNA synthesized by in vitro transcription on rat interferon (IFN) mRNA has been cloned and amplified as recombinant DNA. The nucleotide sequence of these rat IFN cDNA clones revealed i. the partial presence of the coding region of the gene and ii all cDNA clones were derived from the same subtype of rat IFN-alpha mRNA. Purified inserted fragments were used as a hybridisation probe against chromosomal "Southern blots" to show that at least twelve rat IFN-alpha-related sequences are present in the genome. A lambda-linked rat gene library was screened with the cDNA probes, resulting in an equivalent number of rat IFN-alpha-related hybrid phages. By use of a 3'-noncoding region as a probe, the chromosomal counterpart of the cDNA clones could be detected and the nucleotide sequence of its coding region has been determined. Expression of the coding region in E. coli yielded biologically active IFN, when tested for in vitro or in vivo antiviral activity.
Notes: 1362-4962
Dijkema, R
Pouwels, P
de Reus, A
Schellekens, H
Journal Article
Nucleic Acids Res. 1984 Jan 25;12(2):1227-42. doi: 10.1093/nar/12.2.1227.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1747
Author: Ding, X., Jiang, W., Zhou, P., Liu, L., Wan, X., Yuan, X., Wang, X., Chen, M., Chen, J., Yang, J., Kong, C., Li, B., Peng, C., Wong, C. C., Hou, F. and Zhang, Y.
Year: 2015
Title: Mixed Lineage Leukemia 5 (MLL5) Protein Stability Is Cooperatively Regulated by O-GlcNac Transferase (OGT) and Ubiquitin Specific Protease 7 (USP7)
Journal: PLoS One
Volume: 10
Issue: 12
Pages: e0145023
Epub Date: 2015/12/19
Short Title: Mixed Lineage Leukemia 5 (MLL5) Protein Stability Is Cooperatively Regulated by O-GlcNac Transferase (OGT) and Ubiquitin Specific Protease 7 (USP7)
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0145023
PMCID: PMC4683056
Accession Number: 26678539
Keywords: Adenocarcinoma/*metabolism
Amino Acid Sequence
DNA-Binding Proteins/chemistry/genetics/*metabolism
Female
HEK293 Cells
HeLa Cells
Humans
Molecular Sequence Data
N-Acetylglucosaminyltransferases/genetics/*metabolism
Protein Binding
Protein Stability
Ubiquitin Thiolesterase/genetics/*metabolism
Ubiquitin-Specific Peptidase 7
Ubiquitination
Up-Regulation
Uterine Cervical Neoplasms/*metabolism
Abstract: Mixed lineage leukemia 5 (MLL5) protein is a trithorax family histone 3 lysine 4 (H3K4) methyltransferase that regulates diverse biological processes, including cell cycle progression, hematopoiesis and cancer. The mechanisms by which MLL5 protein stability is regulated have remained unclear to date. Here, we showed that MLL5 protein stability is cooperatively regulated by O-GlcNAc transferase (OGT) and ubiquitin-specific protease 7 (USP7). Depletion of OGT in cells led to a decrease in the MLL5 protein level through ubiquitin/proteasome-dependent proteolytic degradation, whereas ectopic expression of OGT protein suppressed MLL5 ubiquitylation. We further identified deubiquitinase USP7 as a novel MLL5-associated protein using mass spectrometry. USP7 stabilized the MLL5 protein through direct binding and deubiquitylation. Loss of USP7 induced degradation of MLL5 protein. Conversely, overexpression of USP7, but not a catalytically inactive USP7 mutant, led to decreased ubiquitylation and increased MLL5 stability. Co-immunoprecipitation and co-immunostaining assays revealed that MLL5, OGT and USP7 interact with each other to form a stable ternary complex that is predominantly located in the nucleus. In addition, upregulation of MLL5 expression was correlated with increased expression of OGT and USP7 in human primary cervical adenocarcinomas. Our results collectively reveal a novel molecular mechanism underlying regulation of MLL5 protein stability and provide new insights into the functional interplay among O-GlcNAc transferase, deubiquitinase and histone methyltransferase.
Notes: 1932-6203
Ding, Xiaodan
Jiang, Wei
Zhou, Peipei
Liu, Lulu
Wan, Xiaoling
Yuan, Xiujie
Wang, Xizi
Chen, Miao
Chen, Jun
Yang, Jing
Kong, Chao
Li, Bin
Peng, Chao
Wong, Catherine C L
Hou, Fajian
Zhang, Yan
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2015 Dec 17;10(12):e0145023. doi: 10.1371/journal.pone.0145023. eCollection 2015.
Author Address: Department of Immunology, Nanjing Medical University, Jiangsu, China.
Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
Shanghai Red House Obstetrics and Gynecology Hospital, Fudan University, Shanghai, China.
Institute of Biology and Medical Sciences, Soochow University, Jiangsu, China.
College of life science, Sun Yet-Sen University, Guangzhou, China.
College of Life Science, Shanghai Normal University, Shanghai, China.
National Center for Protein Science Shanghai, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 981
Author: Diop, N. K. and Hrycyna, C. A.
Year: 2005
Title: N-Linked glycosylation of the human ABC transporter ABCG2 on asparagine 596 is not essential for expression, transport activity, or trafficking to the plasma membrane
Journal: Biochemistry
Volume: 44
Issue: 14
Pages: 5420-9
Epub Date: 2005/04/06
Date: Apr 12
Short Title: N-Linked glycosylation of the human ABC transporter ABCG2 on asparagine 596 is not essential for expression, transport activity, or trafficking to the plasma membrane
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi0479858
Accession Number: 15807535
Keywords: ATP Binding Cassette Transporter, Subfamily G, Member 2
ATP-Binding Cassette Transporters/chemistry/*metabolism
Amino Acid Sequence
Asparagine/chemistry/*metabolism
Glycosylation
HeLa Cells
Humans
Immunoprecipitation
Microscopy, Confocal
Microscopy, Fluorescence
Molecular Sequence Data
Neoplasm Proteins/chemistry/*metabolism
Protein Transport
Tunicamycin/pharmacology
Abstract: The human ATP-binding cassette half-transporter ABCG2 is a 72 kDa plasma membrane protein that can confer multidrug resistance to cells in culture when overexpressed. Both transiently and stably expressed ABCG2 are glycosylated, and treatment with peptide N-glycosidase F reduces the apparent molecular mass on SDS-PAGE gels to approximately 60 kDa. Sequence analysis revealed three potential N-linked glycosylation sites in human ABCG2 at amino acids 418, 557, and 596. Site-directed mutagenesis experiments, in which each Asn was changed to Gln independently, revealed that only asparagine 596 is N-linked glycosylated. These data provide the first direct identification of the modified residue in ABCG2 and evidence for the localization of loop 5 to the extracellular space, previously only predicted from hydropathy analysis. Immunoblot and pulse-chase analyses revealed that the glycosylation-deficient ABCG2 (N596Q) variant and the glycosylated parent transporter are expressed equivalently at steady state and have similar half-lives. Cell surface analysis of ABCG2 expression showed comparable amounts of the N596Q variant present at the plasma membrane compared to the glycosylated ABCG2 protein. The ABCG2 (N596Q) variant is also functional, demonstrating rhodamine 123 transport in intact cells comparable to that in cells expressing glycosylated ABCG2. Furthermore, in crude membrane preparations, neither the basal nor the prazosin-stimulated ( approximately 2-fold) ATPase activities of ABCG2 (N596Q) were affected compared to glycosylated ABCG2. Although subtle defects in transporter trafficking and function may exist, these data taken together suggest that N-glycosylation at arginine 596 is not essential for the expression, trafficking to the plasma membrane, or the overall function of ABCG2.
Notes: Diop, Ndeye K
Hrycyna, Christine A
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2005 Apr 12;44(14):5420-9. doi: 10.1021/bi0479858.
Author Address: Department of Chemistry and Purdue Cancer Center, Purdue University, 560 Oval Drive, West Lafayette, Indiana 47907-2084, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 434
Author: DiScipio, R. G. and Hugli, T. E.
Year: 1985
Title: The architecture of complement component C9 and poly(C9)
Journal: J Biol Chem
Volume: 260
Issue: 27
Pages: 14802-9
Epub Date: 1985/11/25
Date: Nov 25
Short Title: The architecture of complement component C9 and poly(C9)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4055801
Keywords: Amino Acid Sequence
Antibodies
Antigen-Antibody Complex
Carbohydrates/analysis
*Complement C9/isolation & purification
Humans
Macromolecular Substances
Microscopy, Electron
Molecular Weight
Oligosaccharides/analysis
Protein Conformation
Abstract: The architecture of human complement component C9 and poly(C9) was investigated by transmission electron microscopy. Monomeric native C9 (Mr = 66,000) exhibits an ellipsoid appearance (70 X 50 A) with a crevice visible on one face. C9 polymerizes spontaneously to form hollow tubular structures consisting of 12-16 monomeric subunits. Poly(C9) is a cylinder (150 A-outer diameters and 90 A-inner diameter) rimmed by a torus (46-A thick) on one end. Electron micrographs of poly(C9) indicate that the torus is formed by radial strands of polypeptide. Each subunit of poly(C9) is apparently tilted relative to the central axis of the cylindrical structure. C9 can be cleaved by alpha-thrombin into two single-chain polypeptide fragments: C9a (Mr = 28,000) and C9b (Mr = 38,000), which are the amino- and carboxyl-terminal segments of the protein, respectively. The cleaved form of the protein, C9a,b, can be induced to polymerize under suitable conditions to form sodium dodecyl sulfate-resistant poly(C9), indicating that the resistance of poly(C9) to denaturation is a collective feature of both C9a and C9b. The C9a and C9b polypeptide regions have been mapped on poly(C9) by immunoelectron microscopy. Determinants for the C9a region were observed about the torus, base, and on the midsection of the poly(C9) cylinder. C9b epitopes are concentrated predominantly about the torus and base, but were rarely observed on the midsection of poly(C9). Thus, the C9a and C9b segments of the C9 polypeptide are not clearly segregated in poly(C9). The locations of oligosaccharide units on poly(C9) were visualized by electron microscopy after labeling of the complex with concanavalin A bound to colloidal gold. The oligosaccharide positions were found on the periphery of the torus and base. In summary, C9 appears to be a single-domain protein. Polymerization involves a major rearrangement. To form a subunit of poly(C9) the polypeptide chain must form at least one major fold parallel to the central axis of the tubule.
Notes: DiScipio, R G
Hugli, T E
AI17354/AI/NIAID NIH HHS/United States
AI22415/AI/NIAID NIH HHS/United States
HL16411/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1985 Nov 25;260(27):14802-9.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 238
Author: Do, S. I. and Cummings, R. D.
Year: 1992
Title: Presence of O-linked oligosaccharide on a threonine residue in the human transferrin receptor
Journal: Glycobiology
Volume: 2
Issue: 4
Pages: 345-53
Epub Date: 1992/08/01
Date: Aug
Short Title: Presence of O-linked oligosaccharide on a threonine residue in the human transferrin receptor
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/2.4.345
Accession Number: 1421756
Keywords: Amino Acid Sequence
Amino Acids/analysis
Binding Sites
Carbohydrate Conformation
Carbohydrate Sequence
Glycosylation
Humans
Hydrogen-Ion Concentration
Immunosorbent Techniques
Leukemia, Erythroblastic, Acute
Molecular Sequence Data
Oligosaccharides/*analysis
Oxidation-Reduction
Peptide Fragments/analysis/metabolism
Receptors, Transferrin/*analysis/chemistry/metabolism
Threonine/*analysis
Trypsin
Tumor Cells, Cultured
Abstract: We have previously demonstrated that the human transferrin receptor (TfR) of approximately 90 kDa contains Ser/Thr-linked (O-linked) oligosaccharides. In the present study, we report our identification of the site of attachment of the O-linked oligosaccharides in the receptor. A 70 kDa fragment from the external domain of the TfR was generated by trypsin treatment of the [3H]glucosamine-labelled receptor purified from human K562 cells. The beta-elimination of the intact TfR, but not the 70 kDa fragment, released Gal-[3H]Gal-NAcitol, indicating that the 70 kDa fragment lacks O-linked oligosaccharides. In the remaining 20 kDa fragment there are three potential sites (Thr96, Thr104 and Ser106) for O-glycosylation in the extracellular domain. To identify which of these residues are O-glycosylated, both the [3H]Thr- and [3H]Ser-labelled TfR were directly treated with mild base to effect beta-elimination, and the radiolabelled amino acids and their derivatives were analysed. Approximately 2% of the total radiolabelled Thr, but no radiolabelled Ser, was converted to expected beta-elimination products by this treatment. These and other results demonstrate that only one O-linked oligosaccharide is present in the TfR and that it occurs on either Thr96 or Thr104. From human serum we purified the cleaved, soluble form of the TfR (s-TfR), which contains Thr104, but lacks Thr96. The s-TfR was sensitive to O-glycanase and bound to Jacalin lectin, indicating that the s-TfR contains an O-linked oligosaccharide.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Do, S I
Cummings, R D
CA37626/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1992 Aug;2(4):345-53. doi: 10.1093/glycob/2.4.345.
Author Address: Department of Biochemistry, University of Georgia, Athens 30602.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 339
Author: Do, S. I., Enns, C. and Cummings, R. D.
Year: 1990
Title: Human transferrin receptor contains O-linked oligosaccharides
Journal: J Biol Chem
Volume: 265
Issue: 1
Pages: 114-25
Epub Date: 1990/01/05
Date: Jan 5
Short Title: Human transferrin receptor contains O-linked oligosaccharides
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2403553
Keywords: Acetylgalactosamine/analysis
Borohydrides
Carbohydrate Conformation
Chromatography, Affinity
Galactose/analysis
Glucosamine/metabolism
Humans
Hydrogen-Ion Concentration
Immunosorbent Techniques
Mannose/analysis/metabolism
N-Acetylneuraminic Acid
Oligosaccharides/*isolation & purification
Receptors, Transferrin/*analysis
Sialic Acids/analysis
Tumor Cells, Cultured
Abstract: We have investigated the oligosaccharides in the human transferrin receptor from three different cell lines. During our studies on the structures of the N-linked oligosaccharides of the receptor, we discovered that the receptor contains O-linked oligosaccharides. This report describes the isolation and characterization of these O-linked oligosaccharides. Three different human cell lines--K562, A431, and BeWo--were grown in media containing either [2-3H] mannose or [6-3H]glucosamine. The newly synthesized and radiolabeled transferrin receptors were purified by immunoprecipitation from cell extracts and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The receptor was proteolytically digested or treated directly with mild base/borohydride. The released radiolabeled glycopeptides and oligosaccharides were separated by a variety of chromatographic techniques, and their structures were analyzed. The transferrin receptor from all three cell types contains O-linked oligosaccharides that are released from peptide by mild base/borohydride treatment. The receptor from K562 cells contains at least one O-linked oligosaccharide having two sialic acid residues and a core structure of the disaccharide galactose-N-acetyl-galactosamine. In contrast, the O-linked oligosaccharides in the transferring receptors from both A431 and BeWo cell lines are not as highly sialylated and were identified as both the neutral disaccharide galactose-N-acetylgalactosamine and the neutral monosaccharide N-acetylgalactosamine. In addition, the receptors from all three cell lines contain both complex-type and high mannose-type N-linked oligosaccharides. The complex-type chains in the receptor from A431 cells have properties of blood group A antigens, whereas oligosaccharides in receptors from both BeWo and K562 cells lack these properties. These results are interesting since both A431 and BeWo cells, but not K562 cells, are positive for blood group A antigens. Thus, our results demonstrate that the human transferrin receptor contains O-linked oligosaccharides and that there are differences in the structures of both the O-linked and complex-type N-linked oligosaccharides on the receptors synthesized by different cell types.
Notes: Do, S I
Enns, C
Cummings, R D
CA37626-04/CA/NCI NIH HHS/United States
DK40608/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1990 Jan 5;265(1):114-25.
Author Address: Department of Biochemistry, University of Georgia, Athens 30602.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 217
Author: Dobrowolska, A., Rewkiewicz-Dziarska, A. and Szarska, I.
Year: 1976
Title: The effect of exogenic thyroxine on activity of the thyroid gland, blood serum proteins and leukocytes in the common vole Microtus arvalis, Pallas
Journal: Comp Biochem Physiol A Comp Physiol
Volume: 53
Issue: 4
Pages: 323-6
Epub Date: 1976/01/01
Short Title: The effect of exogenic thyroxine on activity of the thyroid gland, blood serum proteins and leukocytes in the common vole Microtus arvalis, Pallas
Alternate Journal: Comparative biochemistry and physiology. A, Comparative physiology
ISSN: 0300-9629 (Print)
0300-9629
DOI: 10.1016/s0300-9629(76)80148-x
Accession Number: 3312
Keywords: Animals
Arvicolinae/*physiology
Blood Proteins/*metabolism
Leukocytes/drug effects/*metabolism
Male
Rodentia/*physiology
Serum Albumin/metabolism
Serum Globulins/metabolism
Thyroid Gland/cytology/drug effects/*physiology
Thyroxine/*pharmacology
Notes: Dobrowolska, A
Rewkiewicz-Dziarska, A
Szarska, I
Journal Article
England
Comp Biochem Physiol A Comp Physiol. 1976;53(4):323-6. doi: 10.1016/s0300-9629(76)80148-x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1920
Author: Dodson, M., Liu, P., Jiang, T., Ambrose, A. J., Luo, G., Rojo de la Vega, M., Cholanians, A. B., Wong, P. K., Chapman, E. and Zhang, D. D.
Year: 2018
Title: Increased O-GlcNAcylation of SNAP29 Drives Arsenic-Induced Autophagic Dysfunction
Journal: Mol Cell Biol
Volume: 38
Issue: 11
Epub Date: 2018/03/07
Date: Jun 1
Short Title: Increased O-GlcNAcylation of SNAP29 Drives Arsenic-Induced Autophagic Dysfunction
Alternate Journal: Molecular and cellular biology
ISSN: 0270-7306 (Print)
0270-7306
DOI: 10.1128/mcb.00595-17
PMCID: PMC5954189
Accession Number: 29507186
Keywords: Animals
Arsenic/*pharmacology
Autophagy/*drug effects
HeLa Cells
Humans
Lysosomes/*metabolism
Membrane Fusion/physiology
Mice
NIH 3T3 Cells
Phagosomes/metabolism
Protein Binding
Qb-SNARE Proteins/genetics/*metabolism
Qc-SNARE Proteins/genetics/*metabolism
*O-GlcNAc
*Snap29
*SNARE complex
*Stx17
*Vamp8
*arsenic
*autophagy
Abstract: Environmental exposure to arsenic is linked to adverse health effects, including cancer and diabetes. Pleiotropic cellular effects are observed with arsenic exposure. Previously, we demonstrated that arsenic dysregulated the autophagy pathway at low, environmentally relevant concentrations. Here we show that arsenic blocks autophagy by preventing autophagosome-lysosome fusion. Specifically, arsenic disrupts formation of the STX17-SNAP29-VAMP8 SNARE complex, where SNAP29 mediates vesicle fusion through bridging STX17-containing autophagosomes to VAMP8-bearing lysosomes. Mechanistically, arsenic inhibits SNARE complex formation, at least in part, by enhancing O-GlcNAcylation of SNAP29. Transfection of O-GlcNAcylation-defective, but not wild-type, SNAP29 into clustered regularly interspaced short palindromic repeat (CRISPR)-mediated SNAP29 knockout cells abolishes arsenic-mediated autophagy inhibition. These findings reveal a mechanism by which low levels of arsenic perturb proteostasis through inhibition of SNARE complex formation, providing a possible therapeutic target for disease intervention in the more than 200 million people exposed to unsafe levels of arsenic.
Notes: 1098-5549
Dodson, Matthew
Liu, Pengfei
Jiang, Tao
Ambrose, Andrew J
Luo, Gang
Rojo de la Vega, Montserrat
Cholanians, Aram B
Wong, Pak Kin
Chapman, Eli
Zhang, Donna D
R01 ES023758/ES/NIEHS NIH HHS/United States
P30 ES006694/ES/NIEHS NIH HHS/United States
T32 GM008804/GM/NIGMS NIH HHS/United States
R01 DK109555/DK/NIDDK NIH HHS/United States
R01 ES026845/ES/NIEHS NIH HHS/United States
R01 ES015010/ES/NIEHS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Mol Cell Biol. 2018 May 15;38(11):e00595-17. doi: 10.1128/MCB.00595-17. Print 2018 Jun 1.
Author Address: Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona, USA.
Department of Biomedical Engineering, The Pennsylvania State University, University Park, Pennsylvania, USA.
Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona, USA dzhang@pharmacy.arizona.edu.
Arizona Cancer Center, University of Arizona, Tucson, Arizona, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1118
Author: Donadio, A. C., Lobo, C., Tosina, M., de la Rosa, V., Martín-Rufián, M., Campos-Sandoval, J. A., Matés, J. M., Márquez, J., Alonso, F. J. and Segura, J. A.
Year: 2008
Title: Antisense glutaminase inhibition modifies the O-GlcNAc pattern and flux through the hexosamine pathway in breast cancer cells
Journal: J Cell Biochem
Volume: 103
Issue: 3
Pages: 800-11
Epub Date: 2007/07/07
Date: Feb 15
Short Title: Antisense glutaminase inhibition modifies the O-GlcNAc pattern and flux through the hexosamine pathway in breast cancer cells
Alternate Journal: Journal of cellular biochemistry
ISSN: 0730-2312
DOI: 10.1002/jcb.21449
Accession Number: 17614351
Keywords: Animals
Breast Neoplasms/*enzymology/*genetics/pathology
Gene Expression/genetics
Glutaminase/antagonists & inhibitors/genetics/*metabolism
Glutamine/metabolism
Hexosamines/biosynthesis/*metabolism
Humans
Mice
N-Acetylglucosaminyltransferases/metabolism
Protein Processing, Post-Translational/genetics
Proteomics
RNA Interference
Sp1 Transcription Factor/*biosynthesis/metabolism
Tumor Cells, Cultured
Uridine Diphosphate N-Acetylglucosamine/metabolism
Abstract: Glutamine behaves as a key nutrient for tumors and rapidly dividing cells. Glutaminase is the main glutamine-utilizing enzyme in these cells, and its activity correlates with glutamine consumption and growth rate. We have carried out the antisense L-type glutaminase inhibition in human MCF7 breast cancer cells, in order to study its effect on the hexosamine pathway and the pattern of protein O-glycosylation. The antisense mRNA glutaminase expressing cells, named ORF19, presented a 50% lower proliferation rate than parental cells, showing a more differentiated phenotype. ORF19 cells had an 80% reduction in glutamine:fructose-6-P amidotransferase activity, which is the rate-limiting step of the hexosamine pathway. Although the overall cellular protein O-glycosylation did not change, the O-glycosylation status of several key proteins was altered. O-glycosylation of O-GlcNAc transferase (OGT), the enzyme that links N-acetylglucosamine to proteins, was fivefold lower in ORF19 than in wild type cells. Inhibition of glutaminase also provoked a 10-fold increase in Sp1 expression, and a significant decrease in the ratio of O-glycosylated to total protein for both Sp1 and the Rpt2 proteasome component. These changes were accompanied by a higher Sp1 transcriptional activity. Proteome analysis of O-glycosylated proteins permitted the detection of two new OGT target proteins: the chaperonin TCP-1 theta and the oncogene Ets-related protein isoform 7. Taken together, our results support the hexosamine pathway and the O-glycosylation of proteins being a sensor mechanism of the nutritional and energetic states of the cell.
Notes: 1097-4644
Donadio, Ana C
Lobo, Carolina
Tosina, Marta
de la Rosa, Vanessa
Martín-Rufián, Mercedes
Campos-Sandoval, José A
Matés, José M
Márquez, Javier
Alonso, Francisco J
Segura, Juan A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Cell Biochem. 2008 Feb 15;103(3):800-11. doi: 10.1002/jcb.21449.
Author Address: Departamento de Bioquímica Clínica, Centro de Investigaciones en Bioquímica Clínica e Inmunología, CIBICI-CONICET, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Haya de Torre y Medina Allende, Ciudad Universitaria, Córdoba, Argentina.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 580
Author: Dong, D. L., Xu, Z. S., Chevrier, M. R., Cotter, R. J., Cleveland, D. W. and Hart, G. W.
Year: 1993
Title: Glycosylation of mammalian neurofilaments. Localization of multiple O-linked N-acetylglucosamine moieties on neurofilament polypeptides L and M
Journal: J Biol Chem
Volume: 268
Issue: 22
Pages: 16679-87
Epub Date: 1993/08/05
Date: Aug 5
Short Title: Glycosylation of mammalian neurofilaments. Localization of multiple O-linked N-acetylglucosamine moieties on neurofilament polypeptides L and M
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8344946
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Cattle
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Neurofilament Proteins/*metabolism
Peptide Mapping
Rats
Sequence Homology, Amino Acid
Abstract: Neurofilaments are neuronal intermediate filaments that play an important role in the growth and maintenance of large myelinated axons. Mammalian neurofilaments are composed of three polypeptide subunits, designed as NF-L, NF-M, and NF-H, all of which are phosphorylated. Here, we demonstrate by several criteria that neurofilament polypeptides are also modified by an abundant type of intracellular protein glycosylation in which single N-acetylglucosamine monosaccharides are O-glycosidically (O-GlcNAc) linked to serine or threonine residues. In purified neurofilament proteins, the O-GlcNAc modifications occur at a stoichiometry of approximately 0.1 and 0.15 mol of GlcNAc/mol of NF-L and NF-M, respectively. The predominant sites of O-GlcNAc attachment on NF-L and NF-M are identified using proteolysis, purification of the glycopeptides, and subsequent analysis by automated gas-phase sequencing, manual Edman degradation, and laser desorption mass spectrometry. For NF-L, both major sites of glycosylation (Thr21 and Ser27) are located at the NH2-terminal head domain. For NF-M, one major site (Thr48) lies within the NH2-terminal head domain, whereas the other (Thr431) is located at the tail domain. Deletions encompassing these sites have been shown previously to have a dominant detrimental effect upon neurofilament assembly, raising questions about the specific function(s) of the saccharide moieties at these sites. Specific identification of these O-GlcNAc attachment sites has set the stage for more detailed mutagenic analysis of O-GlcNAc functions on neurofilaments.
Notes: Dong, D L
Xu, Z S
Chevrier, M R
Cotter, R J
Cleveland, D W
Hart, G W
1P30A128748/PHS HHS/United States
HD13563/HD/NICHD NIH HHS/United States
NS27036/NS/NINDS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1993 Aug 5;268(22):16679-87.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 616
Author: Dong, D. L., Xu, Z. S., Hart, G. W. and Cleveland, D. W.
Year: 1996
Title: Cytoplasmic O-GlcNAc modification of the head domain and the KSP repeat motif of the neurofilament protein neurofilament-H
Journal: J Biol Chem
Volume: 271
Issue: 34
Pages: 20845-52
Epub Date: 1996/08/23
Date: Aug 23
Short Title: Cytoplasmic O-GlcNAc modification of the head domain and the KSP repeat motif of the neurofilament protein neurofilament-H
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.34.20845
Accession Number: 8702840
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Glycopeptides/chemistry
Glycosylation
Male
Molecular Sequence Data
Nerve Tissue Proteins/chemistry
Neurofilament Proteins/chemistry/*metabolism
Peptide Mapping
Protein Processing, Post-Translational
Rats
Repetitive Sequences, Nucleic Acid
Serine/chemistry
Spinal Cord/chemistry
Abstract: Neurofilaments, the major intermediate filaments in large myelinated neurons, are essential for specifying proper axonal caliber. Mammalian neurofilaments are obligate heteropolymers assembled from three polypeptides, neurofilament (NF)-H, NF-M, and NF-L, each of which undergoes phosphorylation at multiple sites. NF-M and NF-L are known to be modified by O-linked N-acetylglucosamine (O-GlcNAc) (Dong, D. L.-Y., Xu, Z.-S., Chevrier, M. R., Cotter, R. J., Cleveland, D. W., and Hart, G. W. (1993) J. Biol. Chem. 268, 16679-16687). Here we further report that NF-H is extensively modified by O-GlcNAc at Thr53, Ser54, and Ser56 in the head domain and, somewhat surprisingly, at multiple sites within the Lys-Ser-Pro repeat motif in the tail domain, a region in assembled neurofilaments known to be nearly stoichiometrically phosphorylated on each of the approximately 50 KSP repeats. Beyond the earlier identified sites on NF-M and NF-L, O-GlcNAc sites on Thr19 and Ser34 of NF-M and Ser34 and Ser48 of NF-L are also determined here, all of which are localized in head domain sequences critical for filament assembly. The proximity of O-GlcNAc and phosphorylation sites in both head and tail domains of each subunit indicates that these modifications may influence one another and play a role in filament assembly and network formation.
Notes: Dong, D L
Xu, Z S
Hart, G W
Cleveland, D W
HD13563/HD/NICHD NIH HHS/United States
NS27036/NS/NINDS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1996 Aug 23;271(34):20845-52. doi: 10.1074/jbc.271.34.20845.
Author Address: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1215
Author: Dong, G., Wearsch, P. A., Peaper, D. R., Cresswell, P. and Reinisch, K. M.
Year: 2009
Title: Insights into MHC class I peptide loading from the structure of the tapasin-ERp57 thiol oxidoreductase heterodimer
Journal: Immunity
Volume: 30
Issue: 1
Pages: 21-32
Epub Date: 2009/01/03
Date: Jan 16
Short Title: Insights into MHC class I peptide loading from the structure of the tapasin-ERp57 thiol oxidoreductase heterodimer
Alternate Journal: Immunity
ISSN: 1074-7613 (Print)
1074-7613
DOI: 10.1016/j.immuni.2008.10.018
PMCID: PMC2650231
NIHMSID: NIHMS92170
Accession Number: 19119025
Keywords: Animals
Cell Line
Crystallography, X-Ray
Dimerization
Histocompatibility Antigens Class I/*immunology
Humans
Membrane Transport Proteins/*chemistry/metabolism
*Models, Molecular
Peptides/*immunology
Protein Disulfide Reductase (Glutathione)/*chemistry/metabolism
Protein Disulfide-Isomerases/*chemistry/metabolism
Protein Structure, Quaternary
Abstract: Tapasin is a glycoprotein critical for loading major histocompatibility complex (MHC) class I molecules with high-affinity peptides. It functions within the multimeric peptide-loading complex (PLC) as a disulfide-linked, stable heterodimer with the thiol oxidoreductase ERp57, and this covalent interaction is required to support optimal PLC activity. Here, we present the 2.6 A resolution structure of the tapasin-ERp57 core of the PLC. The structure revealed that tapasin interacts with both ERp57 catalytic domains, accounting for the stability of the heterodimer, and provided an example of a protein disulfide isomerase family member interacting with substrate. Mutational analysis identified a conserved surface on tapasin that interacted with MHC class I molecules and was critical for peptide loading and editing functions of the tapasin-ERp57 heterodimer. By combining the tapasin-ERp57 structure with those of other defined PLC components, we present a molecular model that illuminates the processes involved in MHC class I peptide loading.
Notes: 1097-4180
Dong, Gang
Wearsch, Pamela A
Peaper, David R
Cresswell, Peter
Reinisch, Karin M
R01 GM070521/GM/NIGMS NIH HHS/United States
R01 GM070521-03/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
GM070521/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Immunity. 2009 Jan 16;30(1):21-32. doi: 10.1016/j.immuni.2008.10.018.
Author Address: Department of Cell Biology, Yale University School of Medicine, New Haven, CT 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1386
Author: Dong, L., Vecchio, A. J., Sharma, N. P., Jurban, B. J., Malkowski, M. G. and Smith, W. L.
Year: 2011
Title: Human cyclooxygenase-2 is a sequence homodimer that functions as a conformational heterodimer
Journal: J Biol Chem
Volume: 286
Issue: 21
Pages: 19035-46
Epub Date: 2011/04/07
Date: May 27
Short Title: Human cyclooxygenase-2 is a sequence homodimer that functions as a conformational heterodimer
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.231969
PMCID: PMC3099718
Accession Number: 21467029
Keywords: Animals
Catalytic Domain
Cell Line
Crystallography, X-Ray
Cyclooxygenase 2/*chemistry/genetics/metabolism
Cyclooxygenase 2 Inhibitors/chemistry
Heme/chemistry/genetics/metabolism
Humans
Mice
Protein Binding
Protein Multimerization/*physiology
Protein Structure, Quaternary
Recombinant Proteins/chemistry/genetics/metabolism
Abstract: Prostaglandin endoperoxide H synthases 1 and 2, also known as cyclooxygenases (COXs) 1 and 2, convert arachidonic acid (AA) to prostaglandin endoperoxide H(2). Prostaglandin endoperoxide H synthases are targets of nonspecific nonsteroidal anti-inflammatory drugs and COX-2-specific inhibitors called coxibs. PGHS-2 is a sequence homodimer. Each monomer has a peroxidase and a COX active site. We find that human PGHS-2 functions as a conformational heterodimer having a catalytic monomer (E(cat)) and an allosteric monomer (E(allo)). Heme binds tightly only to the peroxidase site of E(cat), whereas substrates, as well as certain inhibitors (e.g. celecoxib), bind the COX site of E(cat). E(cat) is regulated by E(allo) in a manner dependent on what ligand is bound to E(allo). Substrate and nonsubstrate fatty acids (FAs) and some COX inhibitors (e.g. naproxen) preferentially bind to the COX site of E(allo). AA can bind to E(cat) and E(allo), but the affinity of AA for E(allo) is 25 times that for E(cat). Palmitic acid, an efficacious stimulator of human PGHS-2, binds only E(allo) in palmitic acid/murine PGHS-2 co-crystals. Nonsubstrate FAs can potentiate or attenuate actions of COX inhibitors depending on the FA and whether the inhibitor binds E(cat) or E(allo). Our studies suggest that the concentration and composition of the free FA pool in the environment in which PGHS-2 functions in cells, the FA tone, is a key factor regulating PGHS-2 activity and its responses to COX inhibitors. We suggest that differences in FA tone occurring with different diets will likely affect both base-line prostanoid synthesis and responses to COX inhibitors.
Notes: 1083-351x
Dong, Liang
Vecchio, Alex J
Sharma, Narayan P
Jurban, Brice J
Malkowski, Michael G
Smith, William L
R01 GM077176/GM/NIGMS NIH HHS/United States
GM 068848/GM/NIGMS NIH HHS/United States
GM 077176/GM/NIGMS NIH HHS/United States
P41 RR001646/RR/NCRR NIH HHS/United States
RR-01646/RR/NCRR NIH HHS/United States
DMR0225180/GM/NIGMS NIH HHS/United States
R01 GM068848/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2011 May 27;286(21):19035-46. doi: 10.1074/jbc.M111.231969. Epub 2011 Apr 5.
Author Address: Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, Michigan 48109, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1201
Author: Dong, M., Liu, H., Tepp, W. H., Johnson, E. A., Janz, R. and Chapman, E. R.
Year: 2008
Title: Glycosylated SV2A and SV2B mediate the entry of botulinum neurotoxin E into neurons
Journal: Mol Biol Cell
Volume: 19
Issue: 12
Pages: 5226-37
Epub Date: 2008/09/26
Date: Dec
Short Title: Glycosylated SV2A and SV2B mediate the entry of botulinum neurotoxin E into neurons
Alternate Journal: Molecular biology of the cell
ISSN: 1059-1524 (Print)
1059-1524
DOI: 10.1091/mbc.e08-07-0765
PMCID: PMC2592654
Accession Number: 18815274
Keywords: Amino Acid Sequence
Animals
Botulinum Toxins/*metabolism
Cells, Cultured
Gangliosides/metabolism
Glycosylation
Hippocampus/cytology
Humans
Membrane Glycoproteins/genetics/*metabolism
Membrane Potentials/physiology
Mice
Mice, Knockout
Molecular Sequence Data
Nerve Tissue Proteins/genetics/*metabolism
Neurons/cytology/*metabolism
Protein Isoforms/genetics/*metabolism
Rats
Recombinant Fusion Proteins/genetics/metabolism
Sequence Alignment
Synaptic Vesicles/metabolism
*Virus Internalization
Abstract: Botulinum neurotoxin E (BoNT/E) can cause paralysis in humans and animals by blocking neurotransmitter release from presynaptic nerve terminals. How this toxin targets and enters neurons is not known. Here we identified two isoforms of the synaptic vesicle protein SV2, SV2A and SV2B, as the protein receptors for BoNT/E. BoNT/E failed to enter neurons cultured from SV2A/B knockout mice; entry was restored by expressing SV2A or SV2B, but not SV2C. Mice lacking SV2B displayed reduced sensitivity to BoNT/E. The fourth luminal domain of SV2A or SV2B alone, expressed in chimeric receptors by replacing the extracellular domain of the low-density lipoprotein receptor, can restore the binding and entry of BoNT/E into neurons lacking SV2A/B. Furthermore, we found disruption of a N-glycosylation site (N573Q) within the fourth luminal domain of SV2A rendered the mutant unable to mediate the entry of BoNT/E and also reduced the entry of BoNT/A. Finally, we demonstrate that BoNT/E failed to bind and enter ganglioside-deficient neurons; entry was rescued by loading exogenous gangliosides into neuronal membranes. Together, the data reported here demonstrate that glycosylated SV2A and SV2B act in conjunction with gangliosides to mediate the entry of BoNT/E into neurons.
Notes: 1939-4586
Dong, Min
Liu, Huisheng
Tepp, William H
Johnson, Eric A
Janz, Roger
Chapman, Edwin R
R01 EY016452-03/EY/NEI NIH HHS/United States
R01 EY016452/EY/NEI NIH HHS/United States
1-U54-AI-057153/AI/NIAID NIH HHS/United States
EY016452/EY/NEI NIH HHS/United States
R01 AI057744/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
GM62116/GM/NIGMS NIH HHS/United States
U54 AI057153/AI/NIAID NIH HHS/United States
U54 GM062116/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Biol Cell. 2008 Dec;19(12):5226-37. doi: 10.1091/mbc.e08-07-0765. Epub 2008 Sep 24.
Author Address: Howard Hughes Medical Institute and Department of Physiology, University of Wisconsin, Madison, WI 53706, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1833
Author: Dong, X., Zhao, B., Iacob, R. E., Zhu, J., Koksal, A. C., Lu, C., Engen, J. R. and Springer, T. A.
Year: 2017
Title: Force interacts with macromolecular structure in activation of TGF-β
Journal: Nature
Volume: 542
Issue: 7639
Pages: 55-59
Epub Date: 2017/01/25
Date: Feb 2
Short Title: Force interacts with macromolecular structure in activation of TGF-β
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature21035
PMCID: PMC5586147
NIHMSID: NIHMS875443
Accession Number: 28117447
Keywords: Amino Acid Sequence
Antigens, Neoplasm/*chemistry/*metabolism
Binding Sites
Crystallography, X-Ray
Evolution, Molecular
Humans
Integrins/*chemistry/*metabolism
Models, Molecular
Protein Binding
Protein Conformation
Transforming Growth Factor beta1/*agonists/*chemistry/metabolism
Abstract: Integrins are adhesion receptors that transmit force across the plasma membrane between extracellular ligands and the actin cytoskeleton. In activation of the transforming growth factor-β1 precursor (pro-TGF-β1), integrins bind to the prodomain, apply force, and release the TGF-β growth factor. However, we know little about how integrins bind macromolecular ligands in the extracellular matrix or transmit force to them. Here we show how integrin α(V)β(6) binds pro-TGF-β1 in an orientation biologically relevant for force-dependent release of TGF-β from latency. The conformation of the prodomain integrin-binding motif differs in the presence and absence of integrin binding; differences extend well outside the interface and illustrate how integrins can remodel extracellular matrix. Remodelled residues outside the interface stabilize the integrin-bound conformation, adopt a conformation similar to earlier-evolving family members, and show how macromolecular components outside the binding motif contribute to integrin recognition. Regions in and outside the highly interdigitated interface stabilize a specific integrin/pro-TGF-β orientation that defines the pathway through these macromolecules which actin-cytoskeleton-generated tensile force takes when applied through the integrin β-subunit. Simulations of force-dependent activation of TGF-β demonstrate evolutionary specializations for force application through the TGF-β prodomain and through the β- and not α-subunit of the integrin.
Notes: 1476-4687
Dong, Xianchi
Zhao, Bo
Iacob, Roxana E
Zhu, Jianghai
Koksal, Adem C
Lu, Chafen
Engen, John R
Springer, Timothy A
R01 AR067288/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2017 Feb 2;542(7639):55-59. doi: 10.1038/nature21035. Epub 2017 Jan 25.
Author Address: Children's Hospital Boston and Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Department of Chemistry and Chemical Biology, Northeastern University, Boston, Massachusetts 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 381
Author: D'Onofrio, M., Starr, C. M., Park, M. K., Holt, G. D., Haltiwanger, R. S., Hart, G. W. and Hanover, J. A.
Year: 1988
Title: Partial cDNA sequence encoding a nuclear pore protein modified by O-linked N-acetylglucosamine
Journal: Proc Natl Acad Sci U S A
Volume: 85
Issue: 24
Pages: 9595-9
Epub Date: 1988/12/01
Date: Dec
Short Title: Partial cDNA sequence encoding a nuclear pore protein modified by O-linked N-acetylglucosamine
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.85.24.9595
PMCID: PMC282808
Accession Number: 3200844
Keywords: Acetylglucosamine/*pharmacology
Amino Acid Sequence
Animals
Base Sequence
Cloning, Molecular
DNA/*analysis
Glucosamine/*analogs & derivatives
Liver/analysis
Membrane Glycoproteins/genetics
Membrane Proteins/*genetics/isolation & purification
Molecular Sequence Data
Molecular Weight
Nuclear Envelope/*analysis
Nuclear Pore Complex Proteins
Peptide Mapping
Rats
Abstract: The nuclear pore complex contains a family of proteins ranging in molecular mass from 35 to 220 kDa that are glycosylated with O-linked N-acetylglucosamine (GlcNAc) residues. We sought to determine the primary sequence of a nuclear pore protein modified by O-linked GlcNAc. The major (62 kDa) nuclear pore glycoprotein (np62) was purified from rat liver nuclear envelopes by immunoaffinity chromatography and preparative gel electrophoresis. After CNBr fragmentation, a glycopeptide was isolated and microsequenced. An oligonucleotide probe based on this sequence information was used to screen a lambda gt11 cDNA library constructed from poly(A) mRNA of the rat thyroid cell line FRTL-5. A clone (B5) was isolated and shown to hybridize to a single 2.5-kilobase species in poly(A) mRNA from rat liver and FRTL-5. This insert was sequenced and found to contain a 691-base-pair cDNA encoding a 155-amino acid open reading frame. This open reading frame contained a CNBr fragment identical to the original glycopeptide sequence and a second CNBr fragment corresponding to a nonglycosylated peptide that was also isolated from the purified pore glycoprotein. The B5 cDNA produced a beta-galactosidase fusion protein of the size predicted by the open reading frame. Analysis of the residues making up a presumptive glycosylation site suggests that the sequence is unlike any known sites for enzymatic N- or O-linked glycosylation. The partial sequence of the 62-kDa nuclear pore glycoprotein shows little similarity to other characterized proteins and elucidates structural features of a member of the family of nuclear pore glycoproteins.
Notes: 1091-6490
D'Onofrio, M
Starr, C M
Park, M K
Holt, G D
Haltiwanger, R S
Hart, G W
Hanover, J A
Journal Article
Proc Natl Acad Sci U S A. 1988 Dec;85(24):9595-9. doi: 10.1073/pnas.85.24.9595.
Author Address: Laboratory of Biochemistry and Metabolism, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, MD 20892.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1667
Author: Donovan, K., Alekseev, O., Qi, X., Cho, W. and Azizkhan-Clifford, J.
Year: 2014
Title: O-GlcNAc modification of transcription factor Sp1 mediates hyperglycemia-induced VEGF-A upregulation in retinal cells
Journal: Invest Ophthalmol Vis Sci
Volume: 55
Issue: 12
Pages: 7862-73
Epub Date: 2014/10/30
Date: Oct 28
Short Title: O-GlcNAc modification of transcription factor Sp1 mediates hyperglycemia-induced VEGF-A upregulation in retinal cells
Alternate Journal: Investigative ophthalmology & visual science
ISSN: 0146-0404 (Print)
0146-0404
DOI: 10.1167/iovs.14-14048
PMCID: PMC4541483
Accession Number: 25352121
Keywords: Animals
Cell Line
Disease Models, Animal
Endothelial Cells/metabolism
Epithelial Cells/metabolism
Humans
Hyperglycemia/*metabolism
N-Acetylglucosaminyltransferases/antagonists & inhibitors/*physiology
Rats
Retina/*metabolism
Retinal Pigment Epithelium/metabolism
Sp1 Transcription Factor/*physiology
Transcriptional Activation
Up-Regulation/drug effects
Vascular Endothelial Growth Factor A/*metabolism
O-GlcNAc
Ogt
Sp1
Vegf-a
diabetic retinopathy
Abstract: PURPOSE: Proangiogenic protein VEGF-A contributes significantly to retinal lesions and neovascularization in diabetic retinopathy (DR). In preclinical DR, hyperglycemia can upregulate VEGF-A in retinal cells. The VEGF-A promoter is responsive to the transcription factor specificity protein 1 (Sp1). The O-GlcNAc modification is driven by glucose concentration and has a profound effect on Sp1 activity. This study investigated the effects of hyperglycemia on Sp1-mediated expression of VEGF-A in the retinal endothelium and pigment epithelium. METHODS: Hyperglycemia-exposed ARPE-19 (human retinal pigment epithelial cells) and TR-iBRB (rat retinal microendothelial cells) were assayed for levels of VEGF-A by qRT-PCR, Western blot, and ELISA. Small molecule inhibitors of O-GlcNAc transferase (OGT) or O-GlcNAcase (OGA) were used to manipulate O-GlcNAc levels. Vascular endothelial growth factor-A protein and transcript were measured in cells depleted of OGT or Sp1 by shRNA. The proximal VEGF-A promoter was analyzed for glucose sensitivity by luciferase assay. Chromatin immunoprecipitation (ChIP) was used to assess Sp1 occupancy on the VEGF-A promoter. RESULTS: Hyperglycemia increased VEGF-A promoter activity and upregulated VEGF-A transcript and protein. Elevation of O-GlcNAc by OGA inhibitors was sufficient to increase VEGF-A. O-GlcNAc transferase inhibition abrogated glucose-driven VEGF-A. Cellular depletion of OGT or Sp1 by shRNA significantly abrogated glucose-induced changes in VEGF-A. ChIP analysis showed that hyperglycemia significantly increased binding of Sp1 to the VEGF-A promoter. CONCLUSIONS: Hyperglycemia-driven VEGF-A production is mediated by elevated O-GlcNAc modification of the Sp1 transcription factor. This mechanism may be significant in the pathogenesis of preclinical DR through VEGF-A upregulation.
Notes: 1552-5783
Donovan, Kelly
Alekseev, Oleg
Qi, Xin
Cho, William
Azizkhan-Clifford, Jane
F30 DK094612/DK/NIDDK NIH HHS/United States
1F30DK094612-O1A1/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Invest Ophthalmol Vis Sci. 2014 Oct 28;55(12):7862-73. doi: 10.1167/iovs.14-14048.
Author Address: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania, United States.
Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania, United States Department of Ophthalmology, The Second Xiangya Hospital of Central South University, Changsha, Hunan, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1766
Author: Dörr, K., Kilch, T., Kappel, S., Alansary, D., Schwär, G., Niemeyer, B. A. and Peinelt, C.
Year: 2016
Title: Cell type-specific glycosylation of Orai1 modulates store-operated Ca2+ entry
Journal: Sci Signal
Volume: 9
Issue: 418
Pages: ra25
Epub Date: 2016/03/10
Date: Mar 8
Short Title: Cell type-specific glycosylation of Orai1 modulates store-operated Ca2+ entry
Alternate Journal: Science signaling
ISSN: 1945-0877
DOI: 10.1126/scisignal.aaa9913
Accession Number: 26956484
Keywords: Antigens, CD/genetics/metabolism
Calcium/*metabolism
Calcium Signaling/*physiology
Endoplasmic Reticulum/genetics/metabolism
Gene Knockdown Techniques
Glycosylation
Humans
Jurkat Cells
ORAI1 Protein/genetics/*metabolism
Sialyltransferases/genetics/metabolism
Abstract: N-glycosylation of cell surface proteins affects protein function, stability, and interaction with other proteins. Orai channels, which mediate store-operated Ca(2+) entry (SOCE), are composed of N-glycosylated subunits. Upon activation by Ca(2+) sensor proteins (stromal interaction molecules STIM1 or STIM2) in the endoplasmic reticulum, Orai Ca(2+) channels in the plasma membrane mediate Ca(2+) influx. Lectins are carbohydrate-binding proteins, and Siglecs are a family of sialic acid-binding lectins with immunoglobulin-like repeats. Using Western blot analysis and lectin-binding assays from various primary human cells and cancer cell lines, we found that glycosylation of Orai1 is cell type-specific. Ca(2+) imaging experiments and patch-clamp experiments revealed that mutation of the only glycosylation site of Orai1 (Orai1N223A) enhanced SOCE in Jurkat T cells. Knockdown of the sialyltransferase ST6GAL1 reduced α-2,6-linked sialic acids in the glycan structure of Orai1 and was associated with increased Ca(2+) entry in Jurkat T cells. In human mast cells, inhibition of sialyl sulfation altered the N-glycan of Orai1 (and other proteins) and increased SOCE. These data suggest that cell type-specific glycosylation influences the interaction of Orai1 with specific lectins, such as Siglecs, which then attenuates SOCE. In summary, the glycosylation state of Orai1 influences SOCE-mediated Ca(2+) signaling and, thus, may contribute to pathophysiological Ca(2+) signaling observed in immune disease and cancer.
Notes: 1937-9145
Dörr, Kathrin
Kilch, Tatiana
Kappel, Sven
Alansary, Dalia
Schwär, Gertrud
Niemeyer, Barbara A
Peinelt, Christine
Journal Article
Research Support, Non-U.S. Gov't
United States
Sci Signal. 2016 Mar 8;9(418):ra25. doi: 10.1126/scisignal.aaa9913.
Author Address: Biophysics, Center for Integrative Physiology and Molecular Medicine, School of Medicine, Saarland University, Homburg 66421, Germany. Molecular Biophysics, Center for Integrative Physiology and Molecular Medicine, School of Medicine, Saarland University, Homburg 66421, Germany. Center of Human and Molecular Biology, Saarland University, Homburg 66421, Germany.
Biophysics, Center for Integrative Physiology and Molecular Medicine, School of Medicine, Saarland University, Homburg 66421, Germany. Center of Human and Molecular Biology, Saarland University, Homburg 66421, Germany.
Molecular Biophysics, Center for Integrative Physiology and Molecular Medicine, School of Medicine, Saarland University, Homburg 66421, Germany.
Biophysics, Center for Integrative Physiology and Molecular Medicine, School of Medicine, Saarland University, Homburg 66421, Germany. Molecular Biophysics, Center for Integrative Physiology and Molecular Medicine, School of Medicine, Saarland University, Homburg 66421, Germany.
Biophysics, Center for Integrative Physiology and Molecular Medicine, School of Medicine, Saarland University, Homburg 66421, Germany. Center of Human and Molecular Biology, Saarland University, Homburg 66421, Germany. bpcpei@uks.eu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1372
Author: Doubravska, L., Krausova, M., Gradl, D., Vojtechova, M., Tumova, L., Lukas, J., Valenta, T., Pospichalova, V., Fafilek, B., Plachy, J., Sebesta, O. and Korinek, V.
Year: 2011
Title: Fatty acid modification of Wnt1 and Wnt3a at serine is prerequisite for lipidation at cysteine and is essential for Wnt signalling
Journal: Cell Signal
Volume: 23
Issue: 5
Pages: 837-48
Epub Date: 2011/01/20
Date: May
Short Title: Fatty acid modification of Wnt1 and Wnt3a at serine is prerequisite for lipidation at cysteine and is essential for Wnt signalling
Alternate Journal: Cellular signalling
ISSN: 0898-6568
DOI: 10.1016/j.cellsig.2011.01.007
Accession Number: 21244856
Keywords: Amino Acid Sequence
Amino Acid Substitution
Animals
Cell Line
Cysteine/*metabolism
Embryonic Development
Humans
Lipoylation
Mice
Molecular Sequence Data
Mutation
Rats
Serine/*metabolism
Wnt Proteins/genetics/*metabolism
Wnt1 Protein/genetics/*metabolism
Wnt3 Protein
Wnt3A Protein
Xenopus/embryology/metabolism
Xenopus Proteins
Abstract: The Wnt family of proteins is a group of extracellular signalling molecules that regulate cell-fate decisions in developing and adult tissues. It is presumed that all 19 mammalian Wnt family members contain two types of post-translational modification: the covalent attachment of fatty acids at two distinct positions, and the N-glycosylation of multiple asparagines. We examined how these modifications contribute to the secretion, extracellular movement and signalling activity of mouse Wnt1 and Wnt3a ligands. We revealed that O-linked acylation of serine is required for the subsequent S-palmitoylation of cysteine. As such, mutant proteins that lack the crucial serine residue are not lipidated. Interestingly, although double-acylation of Wnt1 was indispensable for signalling in mammalian cells, in Xenopus embryos the S-palmitoyl-deficient form retained the signalling activity. In the case of Wnt3a, the functional duality of the attached acyls was less prominent, since the ligand lacking S-linked palmitate was still capable of signalling in various cellular contexts. Finally, we show that the signalling competency of both Wnt1 and Wnt3a is related to their ability to associate with the extracellular matrix.
Notes: 1873-3913
Doubravska, Lenka
Krausova, Michaela
Gradl, Dietmar
Vojtechova, Martina
Tumova, Lucie
Lukas, Jan
Valenta, Tomas
Pospichalova, Vendula
Fafilek, Bohumil
Plachy, Jiri
Sebesta, Ondrej
Korinek, Vladimir
Journal Article
Research Support, Non-U.S. Gov't
England
Cell Signal. 2011 May;23(5):837-48. doi: 10.1016/j.cellsig.2011.01.007. Epub 2011 Jan 16.
Author Address: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Videnska 1083, 142 20 Prague 4, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 713
Author: Doucey, M. A., Hess, D., Blommers, M. J. and Hofsteenge, J.
Year: 1999
Title: Recombinant human interleukin-12 is the second example of a C-mannosylated protein
Journal: Glycobiology
Volume: 9
Issue: 5
Pages: 435-41
Epub Date: 1999/04/20
Date: May
Short Title: Recombinant human interleukin-12 is the second example of a C-mannosylated protein
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/9.5.435
Accession Number: 10207176
Keywords: Amino Acid Sequence
Animals
B-Lymphocytes/metabolism
CHO Cells
Cricetinae
Glycosylation
Humans
In Vitro Techniques
Interleukin-12/*chemistry/metabolism
Magnetic Resonance Spectroscopy
Mannose/*chemistry/metabolism
Mannosyltransferases/metabolism
Peptide Fragments/chemistry/metabolism
Recombinant Proteins/chemistry/metabolism
Substrate Specificity
Abstract: The beta-chain of human interleukin 12 (IL-12) contains at position 319-322, the sequence Trp-x-x-Trp. In human RNase 2 this is the recognition motif for a new, recently discovered posttranslational modification, i.e., the C-glycosidic attachment of a mannosyl residue to the side chain of tryptophan. Analysis of C-terminal peptides of recombinant IL-12 (rHuIL-12) by mass spectrometry and NMR spectroscopy revealed that Trp-319beta is (partially) C-mannosylated. This finding was extended by in vitro mannosylation experiments, using a synthetic peptide derived from the same region of the protein as an acceptor. Furthermore, human B-lymphoblastoid cells, which secrete IL-12, were found to contain an enzyme that carries out the C-mannosylation reaction. This shows that nonrecombinant IL-12 is potentially C-mannosylated as well. This is only the second report on a C-mannosylated protein. However, the occurrence of the C-mannosyltransferase activity in a variety of cells and tissues, and the presence of the recognition motif in many proteins indicate that more C-mannosylated proteins may be found.
Notes: Doucey, M A
Hess, D
Blommers, M J
Hofsteenge, J
Journal Article
England
Glycobiology. 1999 May;9(5):435-41. doi: 10.1093/glycob/9.5.435.
Author Address: Friedrich Miescher-Institut, P.O. Box 2543, CH-4002 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1074
Author: Dragoni, I., Guida, E. and McIntyre, P.
Year: 2006
Title: The cold and menthol receptor TRPM8 contains a functionally important double cysteine motif
Journal: J Biol Chem
Volume: 281
Issue: 49
Pages: 37353-60
Epub Date: 2006/10/04
Date: Dec 8
Short Title: The cold and menthol receptor TRPM8 contains a functionally important double cysteine motif
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M607227200
Accession Number: 17015441
Keywords: Amino Acid Motifs
Amino Acid Sequence
Amino Acid Substitution
Animals
Base Sequence
Binding Sites/genetics
CHO Cells
Cell Line
Cold Temperature
Cricetinae
Cysteine/chemistry
DNA Primers/genetics
Dimerization
Glycosylation
In Vitro Techniques
Menthol/metabolism
Mice
Models, Molecular
Molecular Sequence Data
Molecular Weight
Mutagenesis, Site-Directed
Protein Structure, Quaternary
Recombinant Fusion Proteins/chemistry/genetics/metabolism
Sequence Homology, Amino Acid
TRPM Cation Channels/*chemistry/genetics/*metabolism
Abstract: We have investigated the glycosylation, disulfide bonding, and subunit structure of mouse TRPM8. To do this, amino-terminal c-myc or hemagglutinin epitope-tagged proteins were incorporated and expressed in Chinese hamster ovary cells. These modifications had no obvious effects on channel function in intracellular calcium imaging assays upon application of agonists, icilin or menthol, and cold temperatures. Unmodified TRPM8 migrates with an apparent mass of 129 kDa and can be glycosylated in Chinese hamster ovary cells to give glycoproteins with apparent masses of 136 and 147 kDa. We identified two potential N-linked glycosylation sites in TRPM8 (Asn-821 and Asn-934) and mutated them to show that only the site in the putative pore region at position 934 is modified and that glycosylation of this site is not absolutely necessary for cell surface expression or responsiveness to icilin, menthol, and cool temperatures. Enzymatic cleavage of the carbohydrate chains indicated that they are complex carbohydrate. The glycosylation site is flanked in the pore by two cysteine residues that we mutated, to prove that they are involved in a conserved double cysteine motif, which is essential for channel function. Mutation of either of these cysteines abolishes function and forces the formation of a non-functional complex of the size of a homodimer. The double cysteine mutant is also non-functional. Finally, we showed in Perfluoro-octanoic acid-polyacrylamide gels that TRPM8 can form a tetramer (in addition to dimer and trimer forms), consistent with current thinking that functional TRP ion channels are tetrameric.
Notes: Dragoni, Ilaria
Guida, Elizabeth
McIntyre, Peter
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Dec 8;281(49):37353-60. doi: 10.1074/jbc.M607227200. Epub 2006 Oct 2.
Author Address: Department of Pharmacology, University of Melbourne, Grattan Street Carlton, Victoria 3010, Australia and Novartis Institutes for Biomedical Research, 5 Gower Place, London WC1E 6BS, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1917
Author: Draime, A., Bridoux, L., Belpaire, M., Pringels, T., Degand, H., Morsomme, P. and Rezsohazy, R.
Year: 2018
Title: The O-GlcNAc transferase OGT interacts with and post-translationally modifies the transcription factor HOXA1
Journal: FEBS Lett
Volume: 592
Issue: 7
Pages: 1185-1201
Epub Date: 2018/02/22
Date: Apr
Short Title: The O-GlcNAc transferase OGT interacts with and post-translationally modifies the transcription factor HOXA1
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1002/1873-3468.13015
Accession Number: 29465778
Keywords: Animals
COS Cells
Chlorocebus aethiops
HEK293 Cells
Homeodomain Proteins/genetics/*metabolism
Humans
Mice
N-Acetylglucosaminyltransferases/genetics/*metabolism
NIH 3T3 Cells
Protein Domains
Protein Processing, Post-Translational/*physiology
Transcription Factors/genetics/*metabolism
* Ogt
*HOX proteins
*Hoxa1
*Post-translational modifications
*protein-protein interaction
Abstract: HOXA1 belongs to the HOX family of transcription factors which are key regulators of animal development. Little is known about the molecular pathways controlling HOXA1. Recent data from our group revealed distinct partner proteins interacting with HOXA1. Among them, OGT is an O-linked N-acetylglucosamine (O-GlcNAc) transferase modifying a variety of proteins involved in different cellular processes including transcription. Here, we confirm OGT as a HOXA1 interactor, we characterise which domains of HOXA1 and OGT are required for the interaction, and we provide evidence that OGT post-translationally modifies HOXA1. Mass spectrometry experiments indeed reveal that HOXA1 can be phosphorylated on the AGGTVGSPQYIHHSY peptide and that upon OGT expression, the phosphate adduct is replaced by an O-GlcNAc group.
Notes: 1873-3468
Draime, Amandine
Bridoux, Laure
Belpaire, Magali
Pringels, Tamara
Degand, Hervé
Morsomme, Pierre
Rezsohazy, René
Letter
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2018 Apr;592(7):1185-1201. doi: 10.1002/1873-3468.13015. Epub 2018 Mar 13.
Author Address: Animal Molecular and Cellular Biology, Institut des Sciences de la Vie (ISV), Université catholique de Louvain, Louvain-la-Neuve, Belgium.
Molecular Physiology, Institut des Sciences de la Vie (ISV), Université catholique de Louvain, Louvain-la-Neuve, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1929
Author: Drake, W. R., Hou, C. W., Zachara, N. E. and Grimes, C. L.
Year: 2018
Title: New use for CETSA: monitoring innate immune receptor stability via post-translational modification by OGT
Journal: J Bioenerg Biomembr
Volume: 50
Issue: 3
Pages: 231-240
Epub Date: 2018/04/20
Date: Jun
Short Title: New use for CETSA: monitoring innate immune receptor stability via post-translational modification by OGT
Alternate Journal: Journal of bioenergetics and biomembranes
ISSN: 0145-479X (Print)
0145-479x
DOI: 10.1007/s10863-018-9754-z
PMCID: PMC6126917
NIHMSID: NIHMS961325
Accession Number: 29671171
Keywords: Crohn Disease/genetics
Glycosylation
Humans
Ligands
Mutation
N-Acetylglucosaminyltransferases/*metabolism
Nod2 Signaling Adaptor Protein/genetics/metabolism
Protein Binding
*Protein Processing, Post-Translational
*Protein Stability
*Cetsa
*Crohn’s disease
*NLRs
*O-GlcNAcylation
*Ogt
*Peptidoglycan
Abstract: O-GlcNAcylation is a dynamic and functionally diverse post-translational modification shown to affect thousands of proteins, including the innate immune receptor nucleotide-binding oligomerization domain-containing protein 2 (Nod2). Mutations of Nod2 (R702W, G908R and 1007 fs) are associated with Crohn's disease and have lower stabilities compared to wild type. Cycloheximide (CHX)-chase half-life assays have been used to show that O-GlcNAcylation increases the stability and response of both wild type and Crohn's variant Nod2, R702W. A more rapid method to assess stability afforded by post-translational modifications is necessary to fully comprehend the correlation between NLR stability and O-GlcNAcylation. Here, a recently developed cellular thermal shift assay (CETSA) that is typically used to demonstrate protein-ligand binding was adapted to detect shifts in protein stabilization upon increasing O-GlcNAcylation levels in Nod2. This assay was used as a method to predict if other Crohn's associated Nod2 variants were O-GlcNAcylated, and also identified the modification on another NLR, Nod1. Classical immunoprecipitations and NF-κB transcriptional assays were used to confirm the presence and effect of this modification on these proteins. The results presented here demonstrate that CETSA is a convenient method that can be used to detect the stability effect of O-GlcNAcylation on O-GlcNAc-transferase (OGT) client proteins and will be a powerful tool in studying post-translational modification.
Notes: 1573-6881
Drake, Walter R
Hou, Ching-Wen
Zachara, Natasha E
Grimes, Catherine Leimkuhler
P01 HL107153/HL/NHLBI NIH HHS/United States
P20 GM104316/GM/NIGMS NIH HHS/United States
T32 GM008550/GM/NIGMS NIH HHS/United States
T32 GM133395/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Bioenerg Biomembr. 2018 Jun;50(3):231-240. doi: 10.1007/s10863-018-9754-z. Epub 2018 Apr 18.
Author Address: Department of Chemistry and Biochemistry, University of Delaware, Newark, DE, 19716, USA.
Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, MD, 21205, USA.
Department of Chemistry and Biochemistry, University of Delaware, Newark, DE, 19716, USA. cgrimes@udel.edu.
Department of Biological Sciences, University of Delaware, Newark, DE, 19716, USA. cgrimes@udel.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1176
Author: Dreses-Werringloer, U., Lambert, J. C., Vingtdeux, V., Zhao, H., Vais, H., Siebert, A., Jain, A., Koppel, J., Rovelet-Lecrux, A., Hannequin, D., Pasquier, F., Galimberti, D., Scarpini, E., Mann, D., Lendon, C., Campion, D., Amouyel, P., Davies, P., Foskett, J. K., Campagne, F. and Marambaud, P.
Year: 2008
Title: A polymorphism in CALHM1 influences Ca2+ homeostasis, Abeta levels, and Alzheimer's disease risk
Journal: Cell
Volume: 133
Issue: 7
Pages: 1149-61
Epub Date: 2008/07/01
Date: Jun 27
Short Title: A polymorphism in CALHM1 influences Ca2+ homeostasis, Abeta levels, and Alzheimer's disease risk
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2008.05.048
PMCID: PMC2577842
NIHMSID: NIHMS56596
Accession Number: 18585350
Keywords: Aged
Aged, 80 and over
Alzheimer Disease/*genetics
Amino Acid Sequence
Amyloid beta-Peptides/*metabolism
Calcium/*metabolism
Calcium Channels
Cell Membrane/metabolism
Chromosomes, Human, Pair 10
Cytosol/metabolism
Female
*Genetic Predisposition to Disease
Genome, Human
Humans
Male
Membrane Glycoproteins/chemistry/*genetics/*metabolism
Middle Aged
Molecular Sequence Data
Phylogeny
*Polymorphism, Genetic
Sequence Alignment
Abstract: Alzheimer's disease (AD) is a genetically heterogeneous disorder characterized by early hippocampal atrophy and cerebral amyloid-beta (Abeta) peptide deposition. Using TissueInfo to screen for genes preferentially expressed in the hippocampus and located in AD linkage regions, we identified a gene on 10q24.33 that we call CALHM1. We show that CALHM1 encodes a multipass transmembrane glycoprotein that controls cytosolic Ca(2+) concentrations and Abeta levels. CALHM1 homomultimerizes, shares strong sequence similarities with the selectivity filter of the NMDA receptor, and generates a large Ca(2+) conductance across the plasma membrane. Importantly, we determined that the CALHM1 P86L polymorphism (rs2986017) is significantly associated with AD in independent case-control studies of 3404 participants (allele-specific OR = 1.44, p = 2 x 10(-10)). We further found that the P86L polymorphism increases Abeta levels by interfering with CALHM1-mediated Ca(2+) permeability. We propose that CALHM1 encodes an essential component of a previously uncharacterized cerebral Ca(2+) channel that controls Abeta levels and susceptibility to late-onset AD.
Notes: 1097-4172
Dreses-Werringloer, Ute
Lambert, Jean-Charles
Vingtdeux, Valérie
Zhao, Haitian
Vais, Horia
Siebert, Adam
Jain, Ankit
Koppel, Jeremy
Rovelet-Lecrux, Anne
Hannequin, Didier
Pasquier, Florence
Galimberti, Daniela
Scarpini, Elio
Mann, David
Lendon, Corinne
Campion, Dominique
Amouyel, Philippe
Davies, Peter
Foskett, J Kevin
Campagne, Fabien
Marambaud, Philippe
R01 MH059937/MH/NIMH NIH HHS/United States
R01 MH059937-09/MH/NIMH NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Cell. 2008 Jun 27;133(7):1149-61. doi: 10.1016/j.cell.2008.05.048.
Author Address: Litwin-Zucker Research Center for the Study of Alzheimer's Disease, The Feinstein Institute for Medical Research, North Shore-LIJ, Manhasset, NY 11030, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1513
Author: Drougat, L., Olivier-Van Stichelen, S., Mortuaire, M., Foulquier, F., Lacoste, A. S., Michalski, J. C., Lefebvre, T. and Vercoutter-Edouart, A. S.
Year: 2012
Title: Characterization of O-GlcNAc cycling and proteomic identification of differentially O-GlcNAcylated proteins during G1/S transition
Journal: Biochim Biophys Acta
Volume: 1820
Issue: 12
Pages: 1839-48
Epub Date: 2012/09/13
Date: Dec
Short Title: Characterization of O-GlcNAc cycling and proteomic identification of differentially O-GlcNAcylated proteins during G1/S transition
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbagen.2012.08.024
Accession Number: 22967762
Keywords: Acetylglucosamine/*metabolism
Blotting, Western
Electrophoresis, Gel, Two-Dimensional
Fluorescent Antibody Technique
G1 Phase/*physiology
Humans
Immunoprecipitation
MCF-7 Cells
N-Acetylglucosaminyltransferases/*metabolism
Phosphorylation
*Protein Processing, Post-Translational
*Proteomics
S Phase/*physiology
Signal Transduction
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: BACKGROUND: DNA replication represents a critical step of the cell cycle which requires highly controlled and ordered regulatory mechanisms to ensure the integrity of genome duplication. Among a plethora of elements, post-translational modifications (PTMs) ensure the spatiotemporal regulation of pivotal proteins orchestrating cell division. Despite increasing evidences showing that O-GlcNAcylation regulates mitotic events, the impact of this PTM in the early steps of the cell cycle remains poorly understood. METHODS AND RESULTS: Quiescent MCF7 cells were stimulated by serum mitogens and cell cycle progression was determined by flow cytometry. The levels of O-GlcNAc modified proteins, O-GlcNAc Transferase (OGT) and O-GlcNAcase (OGA) were examined by Western blotting and OGA activity was measured during the progression of cells towards S phase. A global decrease in O-GlcNAcylation was observed at S phase entry, concomitantly to an increase in the activity of OGA. A combination of two-dimensional electrophoresis, Western blotting and mass spectrometry was then used to detect and identify cell cycle-dependent putative O-GlcNAcylated proteins. 58 cytoplasmic and nuclear proteins differentially O-GlcNAcylated through G1/S transition were identified and the O-GlcNAc variations of Cytokeratin 8, hnRNP K, Caprin-1, Minichromosome Maintenance proteins MCM3, MCM6 and MCM7 were validated by immunoprecipitation. CONCLUSIONS: The dynamics of O-GlcNAc is regulated during G1/S transition and observed on key proteins involved in the cytoskeleton networks, mRNA processing, translation, protein folding and DNA replication. GENERAL SIGNIFICANCE: Our results led us to propose that O-GlcNAcylation joins the PTMs that take part in the regulation of DNA replication initiation.
Notes: Drougat, Ludivine
Olivier-Van Stichelen, Stéphanie
Mortuaire, Marlène
Foulquier, François
Lacoste, Anne-Sophie
Michalski, Jean-Claude
Lefebvre, Tony
Vercoutter-Edouart, Anne-Sophie
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2012 Dec;1820(12):1839-48. doi: 10.1016/j.bbagen.2012.08.024. Epub 2012 Sep 3.
Author Address: Université des Sciences et Technologies de Lille, Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1916
Author: Duan, F., Wu, H., Jia, D., Wu, W., Ren, S., Wang, L., Song, S., Guo, X., Liu, F., Ruan, Y. and Gu, J.
Year: 2018
Title: O-GlcNAcylation of RACK1 promotes hepatocellular carcinogenesis
Journal: J Hepatol
Volume: 68
Issue: 6
Pages: 1191-1202
Epub Date: 2018/02/18
Date: Jun
Short Title: O-GlcNAcylation of RACK1 promotes hepatocellular carcinogenesis
Alternate Journal: Journal of hepatology
ISSN: 0168-8278
DOI: 10.1016/j.jhep.2018.02.003
Accession Number: 29454068
Keywords: Amino Acid Substitution
Animals
Carcinogens/chemistry/metabolism
Carcinoma, Hepatocellular/*etiology/metabolism
Cell Line, Tumor
Disease Progression
Glycosylation
Humans
Liver Neoplasms/*etiology/metabolism
Mice
Mice, Inbred BALB C
Mice, Inbred C57BL
Mice, Mutant Strains
Mice, Nude
Mice, Transgenic
Mutagenesis, Site-Directed
Neoplasm Proteins/chemistry/genetics/*metabolism
Neoplasm Transplantation
Protein Kinase C beta/metabolism
Protein Stability
Receptors for Activated C Kinase/chemistry/genetics/*metabolism
Serine/chemistry
*Hepatocellular carcinoma
*O-GlcNAcylation
*Pkcβii
*Rack1
*Ribosome
*Translation
Abstract: BACKGROUND & AIMS: Aberrant oncogenic mRNA translation and protein O-linked β-N-acetylglucosaminylation (O-GlcNAcylation) are general features during tumorigenesis. Nevertheless, whether and how these two pathways are interlinked remain unknown. Our previous study indicated that ribosomal receptor for activated C-kinase 1 (RACK1) promoted chemoresistance and growth in hepatocellular carcinoma (HCC). The aim of this study is to examine the role of RACK1 O-GlcNAcylation in oncogene translation and HCC carcinogenesis. METHODS: The site(s) of RACK1 for O-GlcNAcylation was mapped by mass spectrometry analysis. HCC cell lines were employed to examine the effects of RACK1 O-GlcNAcylation on the translation of oncogenic factors and behaviors of tumor cells in vitro. Transgenic knock-in mice were used to detect the role of RACK1 O-GlcNAcylation in modulating HCC tumorigenesis in vivo. The correlation of RACK1 O-GlcNAcylation with tumor progression and relapse were analyzed in clinical HCC samples. RESULTS: We found that ribosomal RACK1 was highly modified by O-GlcNAc at Ser122. O-GlcNAcylation of RACK1 enhanced its protein stability, ribosome binding and interaction with PKCβII (PRKCB), leading to increased eukaryotic translation initiation factor 4E phosphorylation and translation of potent oncogenes in HCC cells. Genetic ablation of RACK1 O-GlcNAcylation at Ser122 dramatically suppressed tumorigenesis, angiogenesis, and metastasis in vitro and in diethylnitrosamine (DEN)-induced HCC mouse model. Increased RACK1 O-GlcNAcylation was also observed in HCC patient samples and correlated with tumor development and recurrence after chemotherapy. CONCLUSIONS: These findings demonstrate that RACK1 acts as key mediator linking O-GlcNAc metabolism to cap-dependent translation during HCC tumorigenesis. Targeting RACK1 O-GlcNAcylation provides promising options for HCC treatment. LAY SUMMARY: O-GlcNAcylation of ribosomal receptor for activated C-kinase 1 at the amino acid serine122 promotes its stability, ribosome localization and interaction with the protein kinase, PKCβII, thus driving the translation of oncogenes and tumorigenesis of hepatocellular carcinoma. Increased O-GlcNAcylation of ribosomal receptor for activated C-kinase 1 is positively correlated with tumor growth, metastasis and recurrence in patients with hepatocellular carcinoma.
Notes: 1600-0641
Duan, Fangfang
Wu, Hao
Jia, Dongwei
Wu, Weicheng
Ren, Shifang
Wang, Lan
Song, Shushu
Guo, Xinying
Liu, Fenglin
Ruan, Yuanyuan
Gu, Jianxin
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Hepatol. 2018 Jun;68(6):1191-1202. doi: 10.1016/j.jhep.2018.02.003. Epub 2018 Feb 15.
Author Address: Key Laboratory of Glycoconjugate Research Ministry of Health, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China; Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China.
Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai 200032, China. Electronic address: liu.fenglin@zs-hospital.sh.cn.
Key Laboratory of Glycoconjugate Research Ministry of Health, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China; Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China. Electronic address: yuanyuanruan@fudan.edu.cn.
Key Laboratory of Glycoconjugate Research Ministry of Health, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China; Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China; Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 803
Author: Duan, J., Dahlbäck, B. and Villoutreix, B. O.
Year: 2001
Title: Proposed lipocalin fold for apolipoprotein M based on bioinformatics and site-directed mutagenesis
Journal: FEBS Lett
Volume: 499
Issue: 1-2
Pages: 127-32
Epub Date: 2001/06/22
Date: Jun 15
Short Title: Proposed lipocalin fold for apolipoprotein M based on bioinformatics and site-directed mutagenesis
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/s0014-5793(01)02544-3
Accession Number: 11418126
Keywords: Amino Acid Sequence
Apolipoproteins/*chemistry/genetics/*metabolism
Apolipoproteins M
Bacterial Outer Membrane Proteins/*chemistry/metabolism
Binding Sites
*Computational Biology
Computer Simulation
Databases as Topic
*Escherichia coli Proteins
Glycosylation
Humans
Lipocalins
Lipoproteins/*chemistry/metabolism
Models, Molecular
Molecular Sequence Data
*Mutagenesis, Site-Directed
Protein Conformation
*Protein Folding
Solvents
Static Electricity
Structure-Activity Relationship
Abstract: Apolipoprotein M (apoM) is a novel apolipoprotein that is predominantly present in high-density lipoprotein. Sensitive sequence searches, threading and comparative model building experiments revealed apoM to be structurally related to the lipocalin protein family. In a 3D model, characterized by an eight-stranded anti-parallel beta-barrel, a segment including Asn135 could adopt a closed or open conformation. Using site-directed mutagenesis, we demonstrated Asn135 in wild-type apoM to be glycosylated, suggesting that the segment is solvent exposed. ApoM displays two strong acidic patches of potential functional importance, one around the N-terminus and the other next to the opening of the beta-barrel.
Notes: Duan, J
Dahlbäck, B
Villoutreix, B O
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2001 Jun 15;499(1-2):127-32. doi: 10.1016/s0014-5793(01)02544-3.
Author Address: Center for Structural Biochemistry, Department of Biosciences at Novum, Karolinska Institute, Huddinge, Sweden. jianxin.duan@csb.ki.se
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1339
Author: Duggan, K. C., Walters, M. J., Musee, J., Harp, J. M., Kiefer, J. R., Oates, J. A. and Marnett, L. J.
Year: 2010
Title: Molecular basis for cyclooxygenase inhibition by the non-steroidal anti-inflammatory drug naproxen
Journal: J Biol Chem
Volume: 285
Issue: 45
Pages: 34950-9
Epub Date: 2010/09/03
Date: Nov 5
Short Title: Molecular basis for cyclooxygenase inhibition by the non-steroidal anti-inflammatory drug naproxen
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.162982
PMCID: PMC2966109
Accession Number: 20810665
Keywords: Amino Acid Substitution
Animals
Anti-Inflammatory Agents, Non-Steroidal/*chemistry
Crystallography, X-Ray
Cyclooxygenase 2/*chemistry
Humans
Mice
Mutagenesis, Site-Directed
Mutation, Missense
Naproxen/*chemistry
Protein Binding
Abstract: Naproxen ((S)-6-methoxy-α-methyl-2-naphthaleneacetic acid) is a powerful non-selective non-steroidal anti-inflammatory drug that is extensively used as a prescription and over-the-counter medication. Naproxen exhibits gastrointestinal toxicity, but its cardiovascular toxicity may be reduced compared with other drugs in its class. Despite the fact that naproxen has been marketed for many years, the molecular basis of its interaction with cyclooxygenase (COX) enzymes is unknown. We performed a detailed study of naproxen-COX-2 interactions using site-directed mutagenesis, structure-activity analysis, and x-ray crystallography. The results indicate that each of the pendant groups of the naphthyl scaffold are essential for COX inhibition, and only minimal substitutions are tolerated. Mutation of Trp-387 to Phe significantly reduced inhibition by naproxen, a result that appears unique to this inhibitor. Substitution of S or CH(2) for the O atom of the p-methoxy group yielded analogs that were not affected by the W387F substitution and that exhibited increased COX-2 selectivity relative to naproxen. Crystallization and x-ray analysis yielded structures of COX-2 complexed to naproxen and its methylthio analog at 1.7 and 2.3 Å resolution, respectively. The combination of mutagenesis, structure analysis, and x-ray crystallography provided comprehensive information on the unique interactions responsible for naproxen binding to COX-2.
Notes: 1083-351x
Duggan, Kelsey C
Walters, Matthew J
Musee, Joel
Harp, Joel M
Kiefer, James R
Oates, John A
Marnett, Lawrence J
CA89450/CA/NCI NIH HHS/United States
GM15431/GM/NIGMS NIH HHS/United States
R01 CA089450/CA/NCI NIH HHS/United States
P01 GM015431/GM/NIGMS NIH HHS/United States
P50 GM015431/GM/NIGMS NIH HHS/United States
T90 DA022873/DA/NIDA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2010 Nov 5;285(45):34950-9. doi: 10.1074/jbc.M110.162982. Epub 2010 Sep 1.
Author Address: AB Hancock Jr Memorial Laboratory for Cancer Research, Department of Biochemistry, Vanderbilt Institute for Chemical Biology, the Center in Molecular Toxicology, Nashville, Tennessee 37232-0146.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 886
Author: Dvir, H., Harel, M., McCarthy, A. A., Toker, L., Silman, I., Futerman, A. H. and Sussman, J. L.
Year: 2003
Title: X-ray structure of human acid-beta-glucosidase, the defective enzyme in Gaucher disease
Journal: EMBO Rep
Volume: 4
Issue: 7
Pages: 704-9
Epub Date: 2003/06/07
Date: Jul
Short Title: X-ray structure of human acid-beta-glucosidase, the defective enzyme in Gaucher disease
Alternate Journal: EMBO reports
ISSN: 1469-221X (Print)
1469-221x
DOI: 10.1038/sj.embor.embor873
PMCID: PMC1326319
Accession Number: 12792654
Keywords: Amino Acid Sequence
Binding Sites
Catalytic Domain
Crystallography, X-Ray
Gaucher Disease/*enzymology/genetics
Glucosylceramidase/*chemistry/genetics/isolation & purification/metabolism
Humans
Models, Molecular
Molecular Sequence Data
Mutation/genetics
Protein Conformation
Abstract: Gaucher disease, the most common lysosomal storage disease, is caused by mutations in the gene that encodes acid-beta-glucosidase (GlcCerase). Type 1 is characterized by hepatosplenomegaly, and types 2 and 3 by early or chronic onset of severe neurological symptoms. No clear correlation exists between the approximately 200 GlcCerase mutations and disease severity, although homozygosity for the common mutations N370S and L444P is associated with non- neuronopathic and neuronopathic disease, respectively. We report the X-ray structure of GlcCerase at 2.0 A resolution. The catalytic domain consists of a (beta/alpha)(8) TIM barrel, as expected for a member of the glucosidase hydrolase A clan. The distance between the catalytic residues E235 and E340 is consistent with a catalytic mechanism of retention. N370 is located on the longest alpha-helix (helix 7), which has several other mutations of residues that point into the TIM barrel. Helix 7 is at the interface between the TIM barrel and a separate immunoglobulin-like domain on which L444 is located, suggesting an important regulatory or structural role for this non-catalytic domain. The structure provides the possibility of engineering improved GlcCerase for enzyme-replacement therapy, and for designing structure-based drugs aimed at restoring the activity of defective GlcCerase.
Notes: 1469-3178
Dvir, Hay
Harel, Michal
McCarthy, Andrew A
Toker, Lilly
Silman, Israel
Futerman, Anthony H
Sussman, Joel L
Journal Article
Research Support, Non-U.S. Gov't
EMBO Rep. 2003 Jul;4(7):704-9. doi: 10.1038/sj.embor.embor873.
Author Address: [1] Departments of Structural Biology and Neurobiology, Weizmann Institute of Science, Rehovot 76100, Israel
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 741
Author: Easton, R. L., Patankar, M. S., Lattanzio, F. A., Leaven, T. H., Morris, H. R., Clark, G. F. and Dell, A.
Year: 2000
Title: Structural analysis of murine zona pellucida glycans. Evidence for the expression of core 2-type O-glycans and the Sd(a) antigen
Journal: J Biol Chem
Volume: 275
Issue: 11
Pages: 7731-42
Epub Date: 2000/03/14
Date: Mar 17
Short Title: Structural analysis of murine zona pellucida glycans. Evidence for the expression of core 2-type O-glycans and the Sd(a) antigen
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.275.11.7731
Accession Number: 10713085
Keywords: Animals
Carbohydrate Sequence
Female
Gas Chromatography-Mass Spectrometry
Glycopeptides/chemistry
*Glycoside Hydrolases
Lectins/metabolism
Mannosidases/metabolism
Methylation
Mice
Models, Chemical
Models, Molecular
Molecular Sequence Data
Neuraminidase/metabolism
Oligosaccharides/*chemistry
Polysaccharides/*chemistry
Protein Binding
Sequence Analysis
Spectrometry, Mass, Fast Atom Bombardment
Zona Pellucida/*chemistry
alpha-Mannosidase
beta-Galactosidase/metabolism
beta-N-Acetylhexosaminidases/metabolism
Abstract: Murine sperm initiate fertilization by binding to specific oligosaccharides linked to the zona pellucida, the specialized matrix coating the egg. Biophysical analyses have revealed the presence of both high mannose and complex-type N-glycans in murine zona pellucida. The predominant high mannose-type glycan had the composition Man(5)GlcNAc(2), but larger oligosaccharides of this type were also detected. Biantennary, triantennary, and tetraantennary complex-type N-glycans were found to be terminated with the following antennae: Galbeta1-4GlcNAc, NeuAcalpha2-3Galbeta1-4GlcNAc, NeuGcalpha2-3Galbeta1-4GlcNAc, the Sd(a) antigen (NeuAcalpha2-3[GalNAcbeta1-4]Galbeta1-4GlcNAc, NeuGcalpha2-3[GalNAcbeta1-4]Galbeta1-4GlcNAc), and terminal GlcNAc. Polylactosamine-type sequence was also detected on a subset of the antennae. Analysis of the O-glycans indicated that the majority were core 2-type (Galbeta1-4GlcNAcbeta1-6[Galbeta1-3]GalNAc). The beta1-6-linked branches attached to these O-glycans were terminated with the same sequences as the N-glycans, except for terminal GlcNAc. Glycans bearing Galbeta1-4GlcNAcbeta1-6 branches have previously been suggested to mediate initial murine gamete binding. Oligosaccharides terminated with GalNAcbeta1-4Gal have been implicated in the secondary binding interaction that occurs following the acrosome reaction. The significant implications of these observations are discussed.
Notes: Easton, R L
Patankar, M S
Lattanzio, F A
Leaven, T H
Morris, H R
Clark, G F
Dell, A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2000 Mar 17;275(11):7731-42. doi: 10.1074/jbc.275.11.7731.
Author Address: Department of Biochemistry, Imperial College of Science, Technology and Medicine, London SW7 2AY, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1285
Author: Egenberger, B., Polleichtner, G., Wischmeyer, E. and Döring, F.
Year: 2010
Title: N-linked glycosylation determines cell surface expression of two-pore-domain K+ channel TRESK
Journal: Biochem Biophys Res Commun
Volume: 391
Issue: 2
Pages: 1262-7
Epub Date: 2009/12/17
Date: Jan 8
Short Title: N-linked glycosylation determines cell surface expression of two-pore-domain K+ channel TRESK
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2009.12.056
Accession Number: 20006580
Keywords: Amino Acid Sequence
Animals
Cell Membrane/*metabolism
Glycosylation
Humans
Mice
Molecular Sequence Data
Oocytes
Patch-Clamp Techniques
Potassium Channels/genetics/*metabolism
Xenopus
Abstract: Within the first external loop of mouse and human TRESK subunits one or two N-glycosylation consensus sites were identified, respectively. Using site directed mutagenesis and Western immunoblotting a single residue of both orthologues was found to be glycosylated upon heterologous expression. Two-electrode voltage-clamp recordings from Xenopus oocytes revealed that current amplitudes of N-glycosylation mutants were reduced by 80% as compared to wildtype TRESK. To investigate membrane targeting, GFP-tagged TRESK subunits were expressed in Xenopus oocytes and fluorescence intensity at the cell surface was measured by confocal microscopy. Signals of the N-glycosylation mutants were reduced by >50%, indicating that their lower current amplitudes substantially result from inadequate surface expression of the channel.
Notes: 1090-2104
Egenberger, Brigitte
Polleichtner, Georg
Wischmeyer, Erhard
Döring, Frank
Journal Article
United States
Biochem Biophys Res Commun. 2010 Jan 8;391(2):1262-7. doi: 10.1016/j.bbrc.2009.12.056. Epub 2009 Dec 16.
Author Address: Institute of Physiology, Department of Neurophysiology, University of Würzburg, 97070 Würzburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 466
Author: Eiffert, H., Quentin, E., Decker, J., Hillemeir, S., Hufschmidt, M., Klingmüller, D., Weber, M. H. and Hilschmann, N.
Year: 1984
Title: [The primary structure of human free secretory component and the arrangement of disulfide bonds]
Journal: Hoppe Seylers Z Physiol Chem
Volume: 365
Issue: 12
Pages: 1489-95
Epub Date: 1984/12/01
Date: Dec
Short Title: [The primary structure of human free secretory component and the arrangement of disulfide bonds]
Alternate Journal: Hoppe-Seyler's Zeitschrift fur physiologische Chemie
ISSN: 0018-4888 (Print)
0018-4888
Original Publication: Die Primärstruktur der menschlichen freien Sekretkomponente und die Anordnung der Disulfidbrücken.
Accession Number: 6526384
Keywords: Amino Acid Sequence
Disulfides/*analysis
Glycoproteins/*analysis
Humans
Hydrolysis
Immunoglobulin A/analysis
Protein Conformation
Abstract: The amino-acid sequence and the arrangement of the disulfide bonds of the human free secretory component were completely elucidated by the methods of protein chemistry. The free secretory component is a monomeric glycoprotein (Mr approximately 86000), consisting of 558 amino acids with 7 carbohydrate chains bound to asparagine. The protein contains 20 cysteine residues but, as a special feature, no methionine. The polypeptide chain is divided into five regions of internal homology, 104 to 114 amino acids in length. The 20 cysteine residues form 10 disulfide bonds, 9 of which confirm the internal homology by their characteristic arrangement. The free secretory component also shows homology to immunoglobulins in some sections. A computer-supported tertiary structure is proposed for the free secretory component.
Notes: Eiffert, H
Quentin, E
Decker, J
Hillemeir, S
Hufschmidt, M
Klingmüller, D
Weber, M H
Hilschmann, N
English Abstract
Journal Article
Germany
Hoppe Seylers Z Physiol Chem. 1984 Dec;365(12):1489-95.
Database Provider: NLM
Language: ger

Reference Type:  Journal Article
Record Number: 1330
Author: Eigenbrot, C., Ultsch, M., Dubnovitsky, A., Abrahmsén, L. and Härd, T.
Year: 2010
Title: Structural basis for high-affinity HER2 receptor binding by an engineered protein
Journal: Proc Natl Acad Sci U S A
Volume: 107
Issue: 34
Pages: 15039-44
Epub Date: 2010/08/11
Date: Aug 24
Short Title: Structural basis for high-affinity HER2 receptor binding by an engineered protein
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1005025107
PMCID: PMC2930565
Accession Number: 20696930
Keywords: Amino Acid Sequence
Binding Sites
Biophysical Phenomena
Crystallography, X-Ray
Epitopes/chemistry/metabolism
Humans
In Vitro Techniques
Models, Molecular
Molecular Sequence Data
Nuclear Magnetic Resonance, Biomolecular
Protein Engineering
Protein Stability
Protein Structure, Secondary
Protein Structure, Tertiary
Receptor, ErbB-2/chemistry/immunology/*metabolism
Recombinant Fusion Proteins/*chemistry/genetics/immunology/*metabolism
Thermodynamics
Abstract: The human epidermal growth factor receptor 2 (HER2) is specifically overexpressed in tumors of several cancers, including an aggressive form of breast cancer. It is therefore a target for both cancer diagnostics and therapy. The 58 amino acid residue Zher2 affibody molecule was previously engineered as a high-affinity binder of HER2. Here we determined the structure of Zher2 in solution and the crystal structure of Zher2 in complex with the HER2 extracellular domain. Zher2 binds to a conformational epitope on HER2 that is distant from those recognized by the therapeutic antibodies trastuzumab and pertuzumab. Its small size and lack of interference may provide Zher2 with advantages for diagnostic use or even for delivery of therapeutic agents to HER2-expressing tumors when trastuzumab or pertuzumab are already employed. Biophysical characterization shows that Zher2 is thermodynamically stable in the folded state yet undergoing conformational interconversion on a submillisecond time scale. The data suggest that it is the HER2-binding conformation that is formed transiently prior to binding. Still, binding is very strong with a dissociation constant K(D) = 22 pM, and perfect conformational homogeneity is therefore not necessarily required in engineered binding proteins. A comparison of the original Z domain scaffold to free and bound Zher2 structures reveals how high-affinity binding has evolved during selection and affinity maturation and suggests how a compromise between binding surface optimization and stability and dynamics of the unbound state has been reached.
Notes: 1091-6490
Eigenbrot, Charles
Ultsch, Mark
Dubnovitsky, Anatoly
Abrahmsén, Lars
Härd, Torleif
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2010 Aug 24;107(34):15039-44. doi: 10.1073/pnas.1005025107. Epub 2010 Aug 9.
Author Address: Department of Structural Biology, Genentech Inc, 1 DNA Way, South San Francisco, CA 94080, USA. eigenbrot.c@gene.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1796
Author: Elegheert, J., Kakegawa, W., Clay, J. E., Shanks, N. F., Behiels, E., Matsuda, K., Kohda, K., Miura, E., Rossmann, M., Mitakidis, N., Motohashi, J., Chang, V. T., Siebold, C., Greger, I. H., Nakagawa, T., Yuzaki, M. and Aricescu, A. R.
Year: 2016
Title: Structural basis for integration of GluD receptors within synaptic organizer complexes
Journal: Science
Volume: 353
Issue: 6296
Pages: 295-9
Epub Date: 2016/07/16
Date: Jul 15
Short Title: Structural basis for integration of GluD receptors within synaptic organizer complexes
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.aae0104
PMCID: PMC5291321
NIHMSID: EMS71210
Accession Number: 27418511
Keywords: Animals
Ligands
*Long-Term Synaptic Depression
Mice
Nerve Tissue Proteins/*chemistry/metabolism
*Neurogenesis
Protein Multimerization
Protein Precursors/*chemistry/metabolism
Protein Structure, Tertiary
Purkinje Cells/metabolism/physiology
Receptors, Glutamate/*chemistry/metabolism
Signal Transduction
Synapses/metabolism/*physiology
Abstract: Ionotropic glutamate receptor (iGluR) family members are integrated into supramolecular complexes that modulate their location and function at excitatory synapses. However, a lack of structural information beyond isolated receptors or fragments thereof currently limits the mechanistic understanding of physiological iGluR signaling. Here, we report structural and functional analyses of the prototypical molecular bridge linking postsynaptic iGluR δ2 (GluD2) and presynaptic β-neurexin 1 (β-NRX1) via Cbln1, a C1q-like synaptic organizer. We show how Cbln1 hexamers "anchor" GluD2 amino-terminal domain dimers to monomeric β-NRX1. This arrangement promotes synaptogenesis and is essential for D: -serine-dependent GluD2 signaling in vivo, which underlies long-term depression of cerebellar parallel fiber-Purkinje cell (PF-PC) synapses and motor coordination in developing mice. These results lead to a model where protein and small-molecule ligands synergistically control synaptic iGluR function.
Notes: 1095-9203
Elegheert, Jonathan
Kakegawa, Wataru
Clay, Jordan E
Shanks, Natalie F
Behiels, Ester
Matsuda, Keiko
Kohda, Kazuhisa
Miura, Eriko
Rossmann, Maxim
Mitakidis, Nikolaos
Motohashi, Junko
Chang, Veronica T
Siebold, Christian
Greger, Ingo H
Nakagawa, Terunaga
Yuzaki, Michisuke
Aricescu, A Radu
090532/Z/09/Z/Wellcome Trust/United Kingdom
R01HD061543/HD/NICHD NIH HHS/United States
L009609/Medical Research Council/United Kingdom
R01 HD061543/HD/NICHD NIH HHS/United States
MC_U105174197/Medical Research Council/United Kingdom
MR/L009609/1/Medical Research Council/United Kingdom
MR/L017776/1/Medical Research Council/United Kingdom
G0700232/Medical Research Council/United Kingdom
C20724/A14414/Cancer Research UK/United Kingdom
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2016 Jul 15;353(6296):295-9. doi: 10.1126/science.aae0104.
Author Address: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK.
Department of Physiology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan.
Department of Molecular Physiology and Biophysics, School of Medicine, Vanderbilt University, Nashville, TN 37232-0615, USA.
Neurobiology Division, MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH, UK.
Department of Physiology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. radu@strubi.ox.ac.uk myuzaki@a5.keio.jp.
Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK. radu@strubi.ox.ac.uk myuzaki@a5.keio.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 658
Author: Elliott, M. M., Kardana, A., Lustbader, J. W. and Cole, L. A.
Year: 1997
Title: Carbohydrate and peptide structure of the alpha- and beta-subunits of human chorionic gonadotropin from normal and aberrant pregnancy and choriocarcinoma
Journal: Endocrine
Volume: 7
Issue: 1
Pages: 15-32
Epub Date: 1997/08/01
Date: Aug
Short Title: Carbohydrate and peptide structure of the alpha- and beta-subunits of human chorionic gonadotropin from normal and aberrant pregnancy and choriocarcinoma
Alternate Journal: Endocrine
ISSN: 1355-008X (Print)
1355-008x
DOI: 10.1007/bf02778058
Accession Number: 9449027
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Carbohydrates/*chemistry
Choriocarcinoma/chemistry/*urine
Chorionic Gonadotropin, beta Subunit, Human/isolation & purification/*urine
Female
Glycoprotein Hormones, alpha Subunit/isolation & purification/*urine
Humans
Hydatidiform Mole/chemistry/urine
Hydrogen Bonding
Hydrolysis
Molecular Sequence Data
Oligosaccharides/chemistry
Peptides/*chemistry
Pregnancy/*urine
Pregnancy Complications, Neoplastic/*urine
Pregnancy in Diabetics/urine
Uterine Neoplasms/chemistry/*urine
Abstract: Human chorionic gonadotropin (hCG), purified from the urine of 14 individuals with normal pregnancy, diabetic pregnancy, hydatidiform mole, or choriocarcinoma, plus two hCG standard preparations, was examined for concurrent peptide-sequence and asparagine (N)- and serine (O)-linked carbohydrate heterogeneity. Protein-sequence analysis was used to measure amino-terminal heterogeneity and the "nicking" of internal peptide bonds. The use of high-pH anion-exchange chromatography coupled with the increased sensitivity of pulsed amperometric detection (HPAE/PAD) revealed that distinct proportions of both hCG alpha- and beta-subunits from normal and aberrant pregnancy are hyperglycosylated, and that it is the extent of the specific subunit hyperglycosylation that significantly increases in malignant disease. Peptide-bond nicking was restricted to a single linkage (beta 47-48) in normal and diabetic pregnancy, but occurred at two sites in standard preparations, at three sites in hydatidiform mole, and at three sites in choriocarcinoma beta-subunit. In the carbohydrate moiety, alpha-subunit from normal pregnancy hCG contained nonfucosylated, mono- and biantennary N-linked structures (49.3 and 36.7%, means); fucosylated biantennary and triantennary oligosaccharides were also identified (7.3 and 6.9%). In choriocarcinoma alpha-subunit, the level of fucosylated biantennary increased, offset by a parallel decrease in the predominant biantennary structure of normal pregnancy (P < 0.0001). The beta-subunit from normal pregnancy hCG contained fucosylated and nonfucosylated biantennary N-linked structures; however, mono- and triantennary oligosaccharides were also identified (4.6 and 13.7%). For O-linked glycans, in beta-subunit from normal pregnancy, disaccharide-core structure predominated, whereas tetrasaccharide-core structure was also detected (15.6%). A trend was demonstrated in beta-subunit: the proportions of the nonpredominating N- and O-linked oligosaccharides increased stepwise from normal pregnancy to hydatidiform mole to choriocarcinoma. The increases were: for monoantennary oligosaccharide, 4.6 to 6.8 to 11.2%; for triantennary, 13.7 to 26.7 to 51.5% and, for O-linked tetrasaccharide-core structure, 15.6 to 23.0 to 74.8%. For hCG from individual diabetic pregnancy, the principal N-linked structure (34.7%) was consistent with a biantennary oligosaccharide previously reported only in carcinoma; and sialylation of both N- and O-linked antennae was significantly decreased compared to that of normal pregnancy. Taken collectively, the distinctive patterns of subunit-specific, predominant oligosaccharides appear to reflect the steric effect of local protein structure during glycosylation processes. The evidence of alternative or "hyperbranched" glycoforms on both alpha- and beta-subunits, seen at low levels in normal pregnancy and at increased or even predominant levels in malignant disease, suggests alternative substrate accessibility for Golgi processing enzymes, alpha 1,6 fucosyltransferase and N-acetylglucosaminyltransferase IV, in distinct proportions of subunit molecules.
Notes: Elliott, M M
Kardana, A
Lustbader, J W
Cole, L A
CA-44131/CA/NCI NIH HHS/United States
HD-15454/HD/NICHD NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Endocrine. 1997 Aug;7(1):15-32. doi: 10.1007/BF02778058.
Author Address: Department of Obstetrics and Gynecology, Yale University School of Medicine, New Haven, CT 06510, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1272
Author: Ely, L. K., Fischer, S. and Garcia, K. C.
Year: 2009
Title: Structural basis of receptor sharing by interleukin 17 cytokines
Journal: Nat Immunol
Volume: 10
Issue: 12
Pages: 1245-51
Epub Date: 2009/10/20
Date: Dec
Short Title: Structural basis of receptor sharing by interleukin 17 cytokines
Alternate Journal: Nature immunology
ISSN: 1529-2908 (Print)
1529-2908
DOI: 10.1038/ni.1813
PMCID: PMC2783927
NIHMSID: NIHMS146774 file a patent to use information gained from this crystal structure to design IL-17 antagonists.
Accession Number: 19838198
Keywords: Amino Acid Sequence
Cell Line
Conserved Sequence
Crystallography, X-Ray
Humans
Interleukin-17/*chemistry/immunology/*metabolism
Models, Molecular
Molecular Sequence Data
*Protein Multimerization
Protein Structure, Quaternary
Protein Structure, Tertiary
Receptors, Interleukin-17/*chemistry/immunology/*metabolism
Sequence Alignment
Abstract: Interleukin 17 (IL-17)-producing helper T cells (T(H)-17 cells), together with their effector cytokines, including members of the IL-17 family, are emerging as key mediators of chronic inflammatory and autoimmune disorders. Here we present the crystal structure of a complex of IL-17 receptor A (IL-17RA) bound to IL-17F in a 1:2 stoichiometry. The mechanism of complex formation was unique for cytokines and involved the engagement of IL-17 by two fibronectin-type domains of IL-17RA in a groove between the IL-17 homodimer interface. Binding of the first receptor to the IL-17 cytokines modulated the affinity and specificity of the second receptor-binding event, thereby promoting heterodimeric versus homodimeric complex formation. IL-17RA used a common recognition strategy to bind to several members of the IL-17 family, which allows it to potentially act as a shared receptor in multiple different signaling complexes.
Notes: 1529-2916
Ely, Lauren K
Fischer, Suzanne
Garcia, K Christopher
R01 AI051321-05/AI/NIAID NIH HHS/United States
R37 AI051321/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
AI51321/AI/NIAID NIH HHS/United States
R01 AI051321/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Immunol. 2009 Dec;10(12):1245-51. doi: 10.1038/ni.1813. Epub 2009 Oct 18.
Author Address: Howard Hughes Medical Institute, Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1364
Author: Encarnação, M., Kollmann, K., Trusch, M., Braulke, T. and Pohl, S.
Year: 2011
Title: Post-translational modifications of the gamma-subunit affect intracellular trafficking and complex assembly of GlcNAc-1-phosphotransferase
Journal: J Biol Chem
Volume: 286
Issue: 7
Pages: 5311-8
Epub Date: 2010/12/22
Date: Feb 18
Short Title: Post-translational modifications of the gamma-subunit affect intracellular trafficking and complex assembly of GlcNAc-1-phosphotransferase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.202382
PMCID: PMC3037643
Accession Number: 21173149
Keywords: Animals
COS Cells
Chlorocebus aethiops
Cricetinae
Golgi Apparatus/*enzymology
Humans
Mice
Protein Multimerization/*physiology
Protein Processing, Post-Translational/*physiology
Protein Subunits/genetics/*metabolism
Protein Transport/physiology
Transferases (Other Substituted Phosphate Groups)/genetics/*metabolism
Abstract: GlcNAc-1-phosphotransferase plays a key role in the generation of mannose 6-phosphate, a recognition marker essential for efficient transport of lysosomal hydrolases to lysosomes. The enzyme complex is composed of six subunits (α(2)β(2)γ(2)). The α- and β-subunits are catalytically active, whereas the function of the γ-subunit is still unclear. We have investigated structural properties, localization, and intracellular transport of the human and mouse γ-subunits and the molecular requirements for the assembly of the phosphotransferase complex. The results showed that endogenous and overexpressed γ-subunits were localized in the cis-Golgi apparatus. Secreted forms of γ-subunits were detectable in media of cultured cells as well as in human serum. The γ-subunit contains two in vivo used N-glycosylation sites at positions 88 and 115, equipped with high mannose-type oligosaccharides. (35)S pulse-chase experiments and size exclusion chromatography revealed that the majority of non-glycosylated γ-subunit mutants were integrated in high molecular mass complexes, failed to exit the endoplasmic reticulum (ER), and were rapidly degraded. The substitution of cysteine 245 involved in dimerization of γ-subunits impaired neither ER exit nor trafficking through the secretory pathway. Monomeric γ-subunits failed, however, to associate with other GlcNAc-1-phosphotransferase subunits. The data provide evidence that assembly of the GlcNAc-1-phosphotransferase complex takes place in the ER and requires dimerization of the γ-subunits.
Notes: 1083-351x
Encarnação, Marisa
Kollmann, Katrin
Trusch, Maria
Braulke, Thomas
Pohl, Sandra
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 Feb 18;286(7):5311-8. doi: 10.1074/jbc.M110.202382. Epub 2010 Dec 20.
Author Address: Department of Biochemistry, Children's Hospital, University Medical Center Hamburg-Eppendorf, 20246 Hamburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 487
Author: Endo, M., Suzuki, K., Schmid, K., Fournet, B., Karamanos, Y., Montreuil, J., Dorland, L., van Halbeek, H. and Vliegenthart, J. F.
Year: 1982
Title: The structures and microheterogeneity of the carbohydrate chains of human plasma ceruloplasmin. A study employing 500-MHz 1H-NMR spectroscopy
Journal: J Biol Chem
Volume: 257
Issue: 15
Pages: 8755-60
Epub Date: 1982/08/10
Date: Aug 10
Short Title: The structures and microheterogeneity of the carbohydrate chains of human plasma ceruloplasmin. A study employing 500-MHz 1H-NMR spectroscopy
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7096333
Keywords: Carbohydrate Sequence
*Ceruloplasmin
Humans
Magnetic Resonance Spectroscopy
Notes: Endo, M
Suzuki, K
Schmid, K
Fournet, B
Karamanos, Y
Montreuil, J
Dorland, L
van Halbeek, H
Vliegenthart, J F
GM-10374/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1982 Aug 10;257(15):8755-60.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 602
Author: Endo, T., Fujiwara, T., Ikehara, Y. and Kobata, A.
Year: 1996
Title: Comparative study of the sugar chains of alkaline phosphatases purified from rat liver and rat AH-130 hepatoma cells. Occurrence of fucosylated high-mannose-type and hybrid-type sugar chains
Journal: Eur J Biochem
Volume: 236
Issue: 2
Pages: 579-90
Epub Date: 1996/03/01
Date: Mar 1
Short Title: Comparative study of the sugar chains of alkaline phosphatases purified from rat liver and rat AH-130 hepatoma cells. Occurrence of fucosylated high-mannose-type and hybrid-type sugar chains
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1996.t01-1-00579.x
Accession Number: 8612632
Keywords: Alkaline Phosphatase/*chemistry
Animals
Carbohydrate Sequence
Glycoproteins/*chemistry
Liver/*enzymology
Liver Neoplasms, Experimental/*enzymology
Molecular Sequence Data
Oligosaccharides/chemistry
Oxidation-Reduction
Periodic Acid/chemistry
Rats
Abstract: The N-linked sugar chains of alkaline phosphatases, purified from rat AH-130 hepatoma and from normal rat liver, were released quantitatively as oligosaccharides by hydrazinolysis and were labeled by reduction with NaB3H4. A comparative study of their structures revealed that following structural differences are induced by hepatocyte carcinogenesis: complex-type tetraantennary sugar chains and hybrid-type sugar chains appear; outer-chain moieties of the sugar chains of the hepatoma enzyme contain exclusively the Gal(Beta 1-4)GlcNAc groups (type 2 chains) but those of the normal enzyme contain other Gal(Beta 1-)GlcNAc groups and type 2 chains; and novel fucosylated high-mannose-type sugar chains are found in the oligosaccharides of the hepatoma enzyme.
Notes: Endo, T
Fujiwara, T
Ikehara, Y
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1996 Mar 1;236(2):579-90. doi: 10.1111/j.1432-1033.1996.t01-1-00579.x.
Author Address: Department of Biochemistry, Institute of Medical Science, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 331
Author: Endo, T., Higashino, K., Hada, T., Imanishi, H., Muratani, K., Kochibe, N. and Kobata, A.
Year: 1990
Title: Structures of the asparagine-linked oligosaccharides of an alkaline phosphatase, kasahara isozyme, purified from FL amnion cells
Journal: Cancer Res
Volume: 50
Issue: 4
Pages: 1079-84
Epub Date: 1990/02/15
Date: Feb 15
Short Title: Structures of the asparagine-linked oligosaccharides of an alkaline phosphatase, kasahara isozyme, purified from FL amnion cells
Alternate Journal: Cancer research
ISSN: 0008-5472 (Print)
0008-5472
Accession Number: 2297756
Keywords: Alkaline Phosphatase/*analysis
Amnion/cytology/enzymology
Carbohydrate Sequence
Cell Line
Chemical Phenomena
Chemistry
Chromatography, Gel
Chromatography, Paper
Humans
Isoenzymes/*analysis
Methylation
Molecular Sequence Data
Oligosaccharides/*analysis
Abstract: Asparagine-linked oligosaccharides were quantitatively released by hydrazinolysis from an alkaline phosphatase, Kasahara isozyme, which was purified from FL amnion cells. Almost all of the oligosaccharides (98%) were acidic components, all of which can be converted to neutral oligosaccharides upon sialidase digestion. Structural analysis of the oligosaccharides by sequential exoglycosidase digestion in combination with methylation analysis revealed that the alkaline phosphatase of FL cells contains sialylated mono-, bi-, tri-, and tetraantennary complex type sugar chains with the Gal beta 1----4GlcNAc beta 1---- outer chains. Some of the tetraantennary sugar chains contain a single Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4GlcNAc beta 1---- outer chain on their Man alpha 1----6 arm. Both fucosylated and nonfucosylated trimannosyl cores were found in the sugar chains. However, it is of interest that the core portion of monoantennary oligosaccharide was not fucosylated and that of the tetraantennary oligosaccharide with a tetrasaccharide outer chain was completely fucosylated.
Notes: Endo, T
Higashino, K
Hada, T
Imanishi, H
Muratani, K
Kochibe, N
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
Cancer Res. 1990 Feb 15;50(4):1079-84.
Author Address: Department of Biochemistry, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 328
Author: Endo, T., Kasahara, M. and Kobata, A.
Year: 1990
Title: Structures of the asparagine-linked sugar chain of glucose transporter from human erythrocytes
Journal: Biochemistry
Volume: 29
Issue: 39
Pages: 9126-34
Epub Date: 1990/10/02
Date: Oct 2
Short Title: Structures of the asparagine-linked sugar chain of glucose transporter from human erythrocytes
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00491a005
Accession Number: 2271582
Keywords: Amino Sugars/*chemistry
*Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Ion Exchange
Erythrocytes/*chemistry
Glycophorins/*chemistry
Glycoside Hydrolases
Humans
Molecular Sequence Data
Monosaccharide Transport Proteins/*chemistry
Oligosaccharides
Abstract: The asparagine-linked sugar chain of glucose transporter from human erythrocytes was quantitatively released as oligosaccharides from the polypeptide backbone by hydrazinolysis. They were converted to radioactive oligosaccharides by NaB3H4 reduction after N-acetylation and fractionated by anion-exchange column chromatography and Bio-Gel P-4 column chromatography after sialidase treatment. Structural study of each oligosaccharide by exo- and endoglycosidase digestion and methylation analysis indicated that the glycoprotein contains a high-mannose-type oligosaccharide, Man9.GlcNAc.GlcNAc, and biantennary complex-type oligosaccharides with Man alpha 1----6(+/- GlcNAc beta 1----4)(Man alpha 1----3) Man beta beta 1----4GlcNAc beta 1----4(+/- Fuc alpha 1----6)GlcNAc as their cores and the poly-N-acetyllactosamine composed of about 16 N-acetyllactosaminyl units as their outer chains. These structural features of the sugar moiety of glucose transporter are quite different from those of two major intrinsic glycoproteins of human erythrocytes, glycophorin A and band 3.
Notes: Endo, T
Kasahara, M
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1990 Oct 2;29(39):9126-34. doi: 10.1021/bi00491a005.
Author Address: Department of Biochemistry, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 397
Author: Engel, J., Taylor, W., Paulsson, M., Sage, H. and Hogan, B.
Year: 1987
Title: Calcium binding domains and calcium-induced conformational transition of SPARC/BM-40/osteonectin, an extracellular glycoprotein expressed in mineralized and nonmineralized tissues
Journal: Biochemistry
Volume: 26
Issue: 22
Pages: 6958-65
Epub Date: 1987/11/03
Date: Nov 3
Short Title: Calcium binding domains and calcium-induced conformational transition of SPARC/BM-40/osteonectin, an extracellular glycoprotein expressed in mineralized and nonmineralized tissues
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00396a015
Accession Number: 3427055
Keywords: Amino Acid Sequence
Animals
Calcium/*pharmacology
Carrier Proteins/isolation & purification/*metabolism
Cell Line
Circular Dichroism
Glycoproteins/*metabolism
Mice
Models, Molecular
Molecular Sequence Data
Molecular Weight
Osteonectin
Protein Conformation
Abstract: SPARC, BM-40, and osteonectin are identical or very closely related extracellular proteins of apparent Mr 43,000 (Mr 33,000 predicted from sequence). They were originally isolated from parietal endoderm cells, basement membrane producing tumors, and bone, respectively, but are rather widely distributed in various tissues. In view of the calcium binding activity reported for osteonectin, we analyzed the SPARC sequence and found two putative calcium binding domains. One is an N-terminal acidic region with clusters of glutamic acid residues. This region, although neither gamma-carboxylated nor homologous, resembles the gamma-carboxyglutamic acid (Gla) domain of vitamin K dependent proteins of the blood clotting system in charge density, size of negatively charged clusters, and linkage to the rest of the molecule by a cysteine-rich domain. The other region is an EF-hand calcium binding domain located near the C-terminus. A disulfide bond between the E and F helix is predicted from modeling the EF-hand structure with the known coordinates of intestinal calcium binding protein. The disulfide bridge apparently serves to stabilize the isolated calcium loop in the extracellular protein. As observed for cytoplasmic EF-hand-containing proteins and for Gla domain containing proteins, a major conformational transition is induced in BM-40 upon binding of several Ca2+ ions. This is accompanied by a 35% increase in alpha-helicity. A pronounced sigmoidicity of the dependence of the circular dichroism signal at 220 nm on calcium concentration indicates that the process is cooperative. In view of its properties, abundance, and wide distribution, it is proposed that SPARC/BM-40/osteonectin has a rather general regulatory function in calcium-dependent processes of the extracellular matrix.
Notes: Engel, J
Taylor, W
Paulsson, M
Sage, H
Hogan, B
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1987 Nov 3;26(22):6958-65. doi: 10.1021/bi00396a015.
Author Address: Department of Biophysical Chemistry, Biozentrum, Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 289
Author: Enghild, J. J., Salvesen, G., Hefta, S. A., Thøgersen, I. B., Rutherfurd, S. and Pizzo, S. V.
Year: 1991
Title: Chondroitin 4-sulfate covalently cross-links the chains of the human blood protein pre-alpha-inhibitor
Journal: J Biol Chem
Volume: 266
Issue: 2
Pages: 747-51
Epub Date: 1991/01/15
Date: Jan 15
Short Title: Chondroitin 4-sulfate covalently cross-links the chains of the human blood protein pre-alpha-inhibitor
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1898736
Keywords: Amino Acids/chemistry
Blood Proteins/*metabolism
Blotting, Western
Carbohydrates/chemistry
Chondroitin Sulfates/*metabolism
Cross-Linking Reagents
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Mass Spectrometry
Protein Precursors/*metabolism
Trypsin Inhibitors/*metabolism
Abstract: The human blood protein pre-alpha-inhibitor is composed of one heavy and one light protein chain. The chains are covalently linked to each other by a structure that has not previously been described, which we designate a protein-glycosaminoglycan-protein (PGP) cross-link. A combination of protein and carbohydrate analytical techniques indicates that the interchain linkage is mediated by a chondroitin 4-sulfate glycosaminoglycan that originates from a typical O-glycosidic link to Ser-10 of the light chain. The heavy chain is esterified, via the alpha-carbon of its C-terminal Asp, to C-6 of an internal N-acetylgalactosamine of the glycosaminoglycan chain. This PGP cross-link may be present in other proteins, but could have been overlooked due to the heterogeneous behavior of proteins containing glycosaminoglycan.
Notes: Enghild, J J
Salvesen, G
Hefta, S A
Thøgersen, I B
Rutherfurd, S
Pizzo, S V
CA-33572/CA/NCI NIH HHS/United States
HL-24066/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1991 Jan 15;266(2):747-51.
Author Address: Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 525
Author: Enghild, J. J., Salvesen, G., Thøgersen, I. B., Valnickova, Z., Pizzo, S. V. and Hefta, S. A.
Year: 1993
Title: Presence of the protein-glycosaminoglycan-protein covalent cross-link in the inter-alpha-inhibitor-related proteinase inhibitor heavy chain 2/bikunin
Journal: J Biol Chem
Volume: 268
Issue: 12
Pages: 8711-6
Epub Date: 1993/04/25
Date: Apr 25
Short Title: Presence of the protein-glycosaminoglycan-protein covalent cross-link in the inter-alpha-inhibitor-related proteinase inhibitor heavy chain 2/bikunin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7682553
Keywords: Alpha-Globulins/*metabolism
Amino Acid Sequence
Blood Proteins/metabolism
Carbohydrate Sequence
Electrophoresis, Polyacrylamide Gel
Glycoproteins/blood/chemistry/*metabolism
Glycosaminoglycans/*metabolism
Humans
Mass Spectrometry
*Membrane Glycoproteins
Molecular Sequence Data
Protease Inhibitors/blood/chemistry/*metabolism
*Trypsin Inhibitor, Kunitz Soybean
Abstract: HC2/bikunin is a human plasma proteinase inhibitor composed of two polypeptide chains that resist dissociation under reducing conditions in SDS-polyacrylamide gel electrophoresis. This observation suggests that a nondisulfide cross-link is responsible for the association of these two polypeptide chains. In this study, we have utilized a variety of techniques to investigate the structural basis for this observation. We show that the cross-link between the two protein chains is sensitive to chondroitin sulfate-degrading enzymes and to 50 mM NaOH, properties shared by the protein-glycosaminoglycan-protein cross-link found in the related pre-alpha-inhibitor (Enghild, J. J., Salvesen, G., Hefta, S., Thøgersen, I. B., Rutherfurd, S., and Pizzo, S. V. (1991) J. Biol. Chem. 266, 747-751). Biochemical and mass spectrometric analysis of the peptides containing the cross-link indicate that it is mediated by a chondroitin-4-sulfate chain that originates from a typical O-glycosidic link to Ser10 of bikunin. The COOH-terminal Asp648 residue of heavy chain 2 is esterified via the alpha-carbon to C-6 of an internal N-acetylgalactosamine of the chondroitin-4-sulfate chain. This suggests that the protein-glycosaminoglycan-protein cross-link that assembles the chains of pre-alpha-inhibitor is identical to that which assembles HC2/bikunin, and is probably a characteristic of the bikunin proteins.
Notes: Enghild, J J
Salvesen, G
Thøgersen, I B
Valnickova, Z
Pizzo, S V
Hefta, S A
CA33572/CA/NCI NIH HHS/United States
HL-24066/HL/NHLBI NIH HHS/United States
HL-49542/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1993 Apr 25;268(12):8711-6.
Author Address: Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1588
Author: Erickson, J. R., Pereira, L., Wang, L., Han, G., Ferguson, A., Dao, K., Copeland, R. J., Despa, F., Hart, G. W., Ripplinger, C. M. and Bers, D. M.
Year: 2013
Title: Diabetic hyperglycaemia activates CaMKII and arrhythmias by O-linked glycosylation
Journal: Nature
Volume: 502
Issue: 7471
Pages: 372-6
Epub Date: 2013/10/01
Date: Oct 17
Short Title: Diabetic hyperglycaemia activates CaMKII and arrhythmias by O-linked glycosylation
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature12537
PMCID: PMC3801227
NIHMSID: NIHMS515163
Accession Number: 24077098
Keywords: Acetylglucosamine/metabolism
Animals
Arrhythmias, Cardiac/complications/enzymology/*metabolism
Benzylamines/pharmacology
Brain/drug effects/enzymology
Calcium/metabolism
Calcium-Calmodulin-Dependent Protein Kinase Type 2/*metabolism
Diabetes Complications/enzymology/*metabolism
Diazooxonorleucine/pharmacology
Enzyme Activation/drug effects
Glucose/metabolism/pharmacology
Glycosylation/drug effects
Humans
Hyperglycemia/complications/enzymology/*metabolism
Mice
Myocardium/cytology/enzymology
Myocytes, Cardiac/enzymology/metabolism
Rats
Sarcoplasmic Reticulum/metabolism
Sulfonamides/pharmacology
Abstract: Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is an enzyme with important regulatory functions in the heart and brain, and its chronic activation can be pathological. CaMKII activation is seen in heart failure, and can directly induce pathological changes in ion channels, Ca(2+) handling and gene transcription. Here, in human, rat and mouse, we identify a novel mechanism linking CaMKII and hyperglycaemic signalling in diabetes mellitus, which is a key risk factor for heart and neurodegenerative diseases. Acute hyperglycaemia causes covalent modification of CaMKII by O-linked N-acetylglucosamine (O-GlcNAc). O-GlcNAc modification of CaMKII at Ser 279 activates CaMKII autonomously, creating molecular memory even after Ca(2+) concentration declines. O-GlcNAc-modified CaMKII is increased in the heart and brain of diabetic humans and rats. In cardiomyocytes, increased glucose concentration significantly enhances CaMKII-dependent activation of spontaneous sarcoplasmic reticulum Ca(2+) release events that can contribute to cardiac mechanical dysfunction and arrhythmias. These effects were prevented by pharmacological inhibition of O-GlcNAc signalling or genetic ablation of CaMKIIδ. In intact perfused hearts, arrhythmias were aggravated by increased glucose concentration through O-GlcNAc- and CaMKII-dependent pathways. In diabetic animals, acute blockade of O-GlcNAc inhibited arrhythmogenesis. Thus, O-GlcNAc modification of CaMKII is a novel signalling event in pathways that may contribute critically to cardiac and neuronal pathophysiology in diabetes and other diseases.
Notes: 1476-4687
Erickson, Jeffrey R
Pereira, Laetitia
Wang, Lianguo
Han, Guanghui
Ferguson, Amanda
Dao, Khanha
Copeland, Ronald J
Despa, Florin
Hart, Gerald W
Ripplinger, Crystal M
Bers, Donald M
P30AG010129/AG/NIA NIH HHS/United States
R01 HL111600/HL/NHLBI NIH HHS/United States
T32 HL086350/HL/NHLBI NIH HHS/United States
P01 HL080101/HL/NHLBI NIH HHS/United States
1R01HL118474-01A1/HL/NHLBI NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
R37HL30077/HL/NHLBI NIH HHS/United States
R01 HL030077/HL/NHLBI NIH HHS/United States
R01DK61671/DK/NIDDK NIH HHS/United States
R01HL111600/HL/NHLBI NIH HHS/United States
P30 AG010129/AG/NIA NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
R01 HL105242/HL/NHLBI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
T32HL86350/HL/NHLBI NIH HHS/United States
R01 HL118474/HL/NHLBI NIH HHS/United States
P01HL080101/HL/NHLBI NIH HHS/United States
R37 HL030077/HL/NHLBI NIH HHS/United States
P01HL107153/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Nature. 2013 Oct 17;502(7471):372-6. doi: 10.1038/nature12537. Epub 2013 Sep 29.
Author Address: Department of Pharmacology, University of California, Davis, Davis, California 95616, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1079
Author: Erler, I., Al-Ansary, D. M., Wissenbach, U., Wagner, T. F., Flockerzi, V. and Niemeyer, B. A.
Year: 2006
Title: Trafficking and assembly of the cold-sensitive TRPM8 channel
Journal: J Biol Chem
Volume: 281
Issue: 50
Pages: 38396-404
Epub Date: 2006/10/27
Date: Dec 15
Short Title: Trafficking and assembly of the cold-sensitive TRPM8 channel
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M607756200
Accession Number: 17065148
Keywords: Animals
Cell Line
*Cold Temperature
Humans
Immunoprecipitation
Mutagenesis
Mutagenesis, Site-Directed
Protein Transport
Recombinant Proteins/genetics/metabolism
TRPM Cation Channels/genetics/*metabolism
Two-Hybrid System Techniques
Abstract: TRPM (transient receptor potential melastatin-like) channels are distinct from many other members of the transient receptor potential family in regard to their overall size (>1000 amino acids), the lack of N-terminal ankyrin-like repeats, and hydrophobicity predictions that may allow for more than six transmembrane regions. Common to each TRPM member is a prominent C-terminal coiled coil region. Here we have shown that TRPM8 channels assemble as multimers using the putative coiled coil region within the intracellular C terminus and that this assembly can be disturbed by a single point mutation within the coiled coil region. This mutant neither gives rise to functional channels nor do its subunits interact or form protein complexes that correspond to a multimer. However, they are still transported to the plasma membrane. Furthermore, wild-type currents can be suppressed by expressing the membrane-attached C-terminal region of TRPM8. To separate assembly from trafficking, we investigated the maturation of TRPM8 protein by identifying and mutating the relevant N-linked glycosylation site and showing that glycosylation is neither essential for multimerization nor for transport to the plasma membrane per se but appears to facilitate efficient multimerization and transport.
Notes: Erler, Isabell
Al-Ansary, Dalia M M
Wissenbach, Ulrich
Wagner, Thomas F J
Flockerzi, Veit
Niemeyer, Barbara A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Dec 15;281(50):38396-404. doi: 10.1074/jbc.M607756200. Epub 2006 Oct 25.
Author Address: Department of Pharmacology and Toxicology, University of Saarland, Medical Campus, 66421 Homburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 273
Author: Escribano, J., Lopex-Otin, C., Hjerpe, A., Grubb, A. and Mendez, E.
Year: 1990
Title: Location and characterization of the three carbohydrate prosthetic groups of human protein HC
Journal: FEBS Lett
Volume: 266
Issue: 1-2
Pages: 167-70
Epub Date: 1990/06/18
Date: Jun 18
Short Title: Location and characterization of the three carbohydrate prosthetic groups of human protein HC
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(90)81531-r
Accession Number: 1694784
Keywords: Alpha-Globulins/*ultrastructure
Amino Acid Sequence
Amino Acids/analysis
Carbohydrates/analysis
Chromatography, High Pressure Liquid
Glycoproteins/*ultrastructure
Humans
Molecular Sequence Data
Peptide Fragments/analysis
Abstract: Three different carbohydrate prosthetic groups associated to three chymotryptic peptides, Q1, Q2 and Q3, were isolated from the reduced and carboxymethylated human protein HC. The first oligosaccharide forms an O-glycosidic linkage with a threonine residue at position 5 in the polypeptide chain of protein HC. The second and third carbohydrate prosthetic groups form N-linkages with asparagine residues at positions 17 and 96. Oligosaccharides present in Q1 contain 1 residue of NANA, 2 of GalNAc and 1 of Gal corresponding to the following structure: -O-GalNAc-GalNAc-Gal-NANA. Q2 contains 3 NANA, 9 GlcNAc, 2 Gal and 3 Man, and Q3 contains 2 NANA, 5 GlcNAc, 1 Gal and 2 Man. The sugar compositions of Q2 and Q3 oligosaccharides are compatible with that of the complex kind. The amount of oligosaccharides present in Q1, Q2 and Q3 corresponded respectively to 3.0%, 12.2% and 7.3% of the weight of protein HC. No difference was found between the carbohydrate composition of urinary and plasma protein HC.
Notes: Escribano, J
Lopex-Otin, C
Hjerpe, A
Grubb, A
Mendez, E
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1990 Jun 18;266(1-2):167-70. doi: 10.1016/0014-5793(90)81531-r.
Author Address: Servicio de Endocrinologia, Hospital Ramon y Cajal, Madrid, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 623
Author: Esposito, C., Pucci, P., Amoresano, A., Marino, G., Cozzolino, A. and Porta, R.
Year: 1996
Title: Transglutaminase from rat coagulating gland secretion. Post-translational modifications and activation by phosphatidic acids
Journal: J Biol Chem
Volume: 271
Issue: 44
Pages: 27416-23
Epub Date: 1996/11/01
Date: Nov 1
Short Title: Transglutaminase from rat coagulating gland secretion. Post-translational modifications and activation by phosphatidic acids
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.44.27416
Accession Number: 8910321
Keywords: Amino Acid Sequence
Animals
Binding Sites
Calcium/analysis
Carbohydrate Sequence
Cations, Divalent/pharmacology
Cysteine
Enzyme Activation
Glycoproteins/chemistry/metabolism
Guinea Pigs
Kinetics
Male
Molecular Sequence Data
Oligosaccharides/chemistry/isolation & purification
Peptide Fragments/chemistry/isolation & purification
Phosphatidic Acids/*pharmacology
Prostate/*enzymology
*Protein Processing, Post-Translational
Rats
Rats, Wistar
Sequence Homology, Amino Acid
Sexual Maturation
Spectrometry, Mass, Fast Atom Bombardment
Transglutaminases/chemistry/*metabolism
Abstract: Structural and biochemical characteristics of transglutaminase purified by a rapid chromatographic procedure from the rat coagulating gland (anterior prostate) secretion are reported. Fast atom bombardment mapping and automated Edman degradation experiments allowed us to verify that at least 85% of the entire transglutaminase amino acid sequence is identical to that derived from the cDNA of the major androgen-dependent rat prostate protein called DP1. The enzyme was found NH2 terminally blocked and largely post-translationally modified, since the presence of N-linked oligosaccharides, as well as of complex lipidic structures, was observed. Mass spectral analysis showed that Asn-408 and -488 are the glycosylated sites, the N-linked structures identified belonging to both high-mannose and complex type glycans. The presence of myo-inositol, of glycerol bound fatty acids, and the high content of mannose residues, are in agreement with previous observations suggesting that a lipid anchor is bound to coagulating gland secretion transglutaminase. Furthermore, two tightly bound calcium ions per molecule of enzyme were detected. Finally, a strong stimulation of the enzyme activity in vitro by both SDS and a variety of phosphatidic acids was observed. The reported structural and functional peculiarities should definitively lead to consider the prostate enzyme as a new member (type IV) of the transglutaminase family.
Notes: Esposito, C
Pucci, P
Amoresano, A
Marino, G
Cozzolino, A
Porta, R
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1996 Nov 1;271(44):27416-23. doi: 10.1074/jbc.271.44.27416.
Author Address: Department of Biochemistry and Biophysics, 2nd University of Naples, 80138 Naples, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 976
Author: Ethier, M., Krokhin, O., Ens, W., Standing, K. G., Wilkins, J. A. and Perreault, H.
Year: 2005
Title: Global and site-specific detection of human integrin alpha 5 beta 1 glycosylation using tandem mass spectrometry and the StrOligo algorithm
Journal: Rapid Commun Mass Spectrom
Volume: 19
Issue: 5
Pages: 721-7
Epub Date: 2005/02/11
Short Title: Global and site-specific detection of human integrin alpha 5 beta 1 glycosylation using tandem mass spectrometry and the StrOligo algorithm
Alternate Journal: Rapid communications in mass spectrometry : RCM
ISSN: 0951-4198 (Print)
0951-4198
DOI: 10.1002/rcm.1844
Accession Number: 15702487
Keywords: *Algorithms
Binding Sites
Glycosylation
Humans
Integrin alpha5beta1/*analysis/*chemistry
Polysaccharides/*analysis/*chemistry
Protein Binding
Spectrometry, Mass, Electrospray Ionization/*methods
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/*methods
Abstract: Glycans are oligosaccharides associated with proteins, and are known to confer specific functions and conformations on glycoproteins. As protein tridimensional structures are related to function, the study of glycans and their impact on protein folding can provide important information to the field of proteomics. The subdiscipline of glycomics (or glycoproteomics) is rapidly growing in importance as glycans in proteins have shown to be involved in protein-protein or protein-(drug, virus, antibody) interactions. Glycomics studies most often aim at identifying glycosylation sites, and thus are performed on deglycosylated proteins resulting in loss of site-specific details concerning the glycosylation. In order to obtain such details by mass spectrometry (MS), either whole glycoproteins must be digested and analyzed as mixtures of peptides and glycopeptides, or glycans must be isolated from glycopeptide fractions and analyzed as pools. This article describes parallel experiments involving both approaches, designed to take advantage of the StrOligo algorithm functionalities with the aim of characterizing glycosylation microheterogeneity on a specific site. A hybrid quadrupole-quadrupole-time-of-flight (QqTOF) instrument equipped with a matrix-assisted laser desorption/ionization (MALDI) source was used. Glycosylation of alpha 5 beta 1 subunits of human integrin was studied to test the methodology. The sample was divided in two aliquots, and glycans from the first aliquot were released enzymatically, labelled with 2-aminobenzamide, and identified using tandem mass spectrometry (MS/MS) and the StrOligo program. The other aliquot was digested with trypsin and the resulting peptides separated by reversed-phase high-performance liquid chromatography (HPLC). A specific collected fraction was then analyzed by MS before and after glycan release. These spectra allowed, by comparison, detection of a glycopeptide (several glycoforms) and elucidation of peptide sequence. Compositions of glycans present were proposed, and identification of possible glycan structures was conducted using MS/MS and StrOligo.
Notes: Ethier, Martin
Krokhin, Oleg
Ens, Werner
Standing, Kenneth G
Wilkins, John A
Perreault, Hélène
59240/PHS HHS/United States
Comparative Study
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Rapid Commun Mass Spectrom. 2005;19(5):721-7. doi: 10.1002/rcm.1844.
Author Address: Department of Chemistry, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1056
Author: Evans, E. J., Castro, M. A., O'Brien, R., Kearney, A., Walsh, H., Sparks, L. M., Tucknott, M. G., Davies, E. A., Carmo, A. M., van der Merwe, P. A., Stuart, D. I., Jones, E. Y., Ladbury, J. E., Ikemizu, S. and Davis, S. J.
Year: 2006
Title: Crystal structure and binding properties of the CD2 and CD244 (2B4)-binding protein, CD48
Journal: J Biol Chem
Volume: 281
Issue: 39
Pages: 29309-20
Epub Date: 2006/06/29
Date: Sep 29
Short Title: Crystal structure and binding properties of the CD2 and CD244 (2B4)-binding protein, CD48
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M601314200
Accession Number: 16803907
Keywords: Animals
Antigens, CD/*chemistry/metabolism
CD2 Antigens/*chemistry/metabolism
CD48 Antigen
CD58 Antigens/chemistry
Evolution, Molecular
Humans
Ligands
Membrane Glycoproteins/*chemistry/metabolism
Models, Molecular
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Rats
Receptors, Immunologic/*chemistry/metabolism
Signaling Lymphocytic Activation Molecule Family
Thermodynamics
Abstract: The structural analysis of surface proteins belonging to the CD2 subset of the immunoglobulin superfamily has yielded important insights into transient cellular interactions. In mice and rats, CD2 and CD244 (2B4), which are expressed predominantly on T cells and natural killer cells, respectively, bind the same, broadly expressed ligand, CD48. Structures of CD2 and CD244 have been solved previously, and we now present the structure of the receptor-binding domain of rat CD48. The receptor-binding surface of CD48 is unusually flat, as in the case of rat CD2, and shares a high degree of electrostatic complementarity with the equivalent surface of CD2. The relatively simple arrangement of charged residues and this flat topology explain why CD48 cross-reacts with CD2 and CD244 and, in rats, with the CD244-related protein, 2B4R. Comparisons of modeled complexes of CD2 and CD48 with the complex of human CD2 and CD58 are suggestive of there being substantial plasticity in the topology of ligand binding by CD2. Thermodynamic analysis of the native CD48-CD2 interaction indicates that binding is driven by equivalent, weak enthalpic and entropic effects, in contrast to the human CD2-CD58 interaction, for which there is a large entropic barrier. Overall, the structural and biophysical comparisons of the CD2 homologues suggest that the evolutionary diversification of interacting cell surface proteins is rapid and constrained only by the requirement that binding remains weak and specific.
Notes: Evans, Edward J
Castro, Mónica A A
O'Brien, Ronan
Kearney, Alice
Walsh, Heather
Sparks, Lisa M
Tucknott, Michael G
Davies, Elizabeth A
Carmo, Alexandre M
van der Merwe, P Anton
Stuart, David I
Jones, E Yvonne
Ladbury, John E
Ikemizu, Shinji
Davis, Simon J
G0500365/Medical Research Council/United Kingdom
G9722488/Medical Research Council/United Kingdom
G9900061/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Sep 29;281(39):29309-20. doi: 10.1074/jbc.M601314200. Epub 2006 Jun 27.
Author Address: Nuffield Department of Clinical Medicine, The University of Oxford and MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Headington, Oxford OX3 9DS, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 975
Author: Evans, E. J., Esnouf, R. M., Manso-Sancho, R., Gilbert, R. J., James, J. R., Yu, C., Fennelly, J. A., Vowles, C., Hanke, T., Walse, B., Hünig, T., Sørensen, P., Stuart, D. I. and Davis, S. J.
Year: 2005
Title: Crystal structure of a soluble CD28-Fab complex
Journal: Nat Immunol
Volume: 6
Issue: 3
Pages: 271-9
Epub Date: 2005/02/08
Date: Mar
Short Title: Crystal structure of a soluble CD28-Fab complex
Alternate Journal: Nature immunology
ISSN: 1529-2908 (Print)
1529-2908
DOI: 10.1038/ni1170
Accession Number: 15696168
Keywords: Abatacept
Amino Acid Sequence
Animals
CD28 Antigens/*chemistry/genetics/immunology
Crystallography
Immunoconjugates/metabolism
Immunoglobulin Fab Fragments/*chemistry/genetics/metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Sequence Alignment
Abstract: Naive T cell activation requires signaling by the T cell receptor and by nonclonotypic cell surface receptors. The most important costimulatory protein is the monovalent homodimer CD28, which interacts with CD80 and CD86 expressed on antigen-presenting cells. Here we present the crystal structure of a soluble form of CD28 in complex with the Fab fragment of a mitogenic antibody. Structural comparisons redefine the evolutionary relationships of CD28-related proteins, antigen receptors and adhesion molecules and account for the distinct ligand-binding and stoichiometric properties of CD28 and the related, inhibitory homodimer CTLA-4. Cryo-electron microscopy-based comparisons of complexes of CD28 with mitogenic and nonmitogenic antibodies place new constraints on models of antibody-induced receptor triggering. This work completes the initial structural characterization of the CD28-CTLA-4-CD80-CD86 signaling system.
Notes: Evans, Edward J
Esnouf, Robert M
Manso-Sancho, Raquel
Gilbert, Robert J C
James, John R
Yu, Chao
Fennelly, Janet A
Vowles, Cheryl
Hanke, Thomas
Walse, Björn
Hünig, Thomas
Sørensen, Poul
Stuart, David I
Davis, Simon J
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Immunol. 2005 Mar;6(3):271-9. doi: 10.1038/ni1170. Epub 2005 Feb 6.
Author Address: Nuffield Department of Clinical Medicine, The University of Oxford, John Radcliffe Hospital, Headington, Oxford, OX3 9DU, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 706
Author: Everse, S. J., Spraggon, G., Veerapandian, L. and Doolittle, R. F.
Year: 1999
Title: Conformational changes in fragments D and double-D from human fibrin(ogen) upon binding the peptide ligand Gly-His-Arg-Pro-amide
Journal: Biochemistry
Volume: 38
Issue: 10
Pages: 2941-6
Epub Date: 1999/03/13
Date: Mar 9
Short Title: Conformational changes in fragments D and double-D from human fibrin(ogen) upon binding the peptide ligand Gly-His-Arg-Pro-amide
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi982626w
Accession Number: 10074346
Keywords: Binding Sites
Calcium/physiology
Calcium-Binding Proteins/metabolism
Computer Simulation
Crystallization
Crystallography, X-Ray
Fibrin Fibrinogen Degradation Products/*chemistry/*metabolism
Humans
Ligands
Models, Molecular
Oligopeptides/*metabolism
Peptide Fragments/chemistry
Protein Binding
Protein Conformation
Abstract: The structure of fragment double-D from human fibrin has been solved in the presence and absence of the peptide ligands that simulate the two knobs exposed by the removal of fibrinopeptides A and B, respectively. All told, six crystal structures have been determined, three of which are reported here for the first time: namely, fragments D and double-D with the peptide GHRPam alone and double-D in the absence of any peptide ligand. Comparison of the structures has revealed a series of conformational changes that are brought about by the various knob-hole interactions. Of greatest interest is a moveable "flap" of two negatively charged amino acids (Glubeta397 and Aspbeta398) whose side chains are pinned back to the coiled coil with a calcium atom bridge until GHRPam occupies the beta-chain pocket. Additionally, in the absence of the peptide ligand GPRPam, GHRPam binds to the gamma-chain pocket, a new calcium-binding site being formed concomitantly.
Notes: Everse, S J
Spraggon, G
Veerapandian, L
Doolittle, R F
HL-26873/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1999 Mar 9;38(10):2941-6. doi: 10.1021/bi982626w.
Author Address: Center for Molecular Genetics, University of California, San Diego, La Jolla 92093-0634, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 675
Author: Everse, S. J., Spraggon, G., Veerapandian, L., Riley, M. and Doolittle, R. F.
Year: 1998
Title: Crystal structure of fragment double-D from human fibrin with two different bound ligands
Journal: Biochemistry
Volume: 37
Issue: 24
Pages: 8637-42
Epub Date: 1998/06/24
Date: Jun 16
Short Title: Crystal structure of fragment double-D from human fibrin with two different bound ligands
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi9804129
Accession Number: 9628725
Keywords: Crystallography, X-Ray
Fibrin Fibrinogen Degradation Products/*chemistry
Humans
Ligands
Molecular Sequence Data
Peptides
*Protein Conformation
Abstract: Factor XIII-cross-linked fragment D (double-D) from human fibrin was crystallized in the presence of two different peptide ligands and the X-ray structure determined at 2.3 A. The peptide Gly-Pro-Arg-Pro-amide, which is an analogue of the knob exposed by the thrombin-catalyzed removal of fibrinopeptide A, was found to reside in the gamma-chain holes, and the peptide Gly-His-Arg-Pro-amide, which corresponds to the beta-chain knob, was found in the homologous beta-chain holes. The structure shows for the first time that the beta-chain knob does indeed bind to a homologous hole on the beta-chain. The gamma- and beta-chain holes are structurally very similar, and it is remarkable they are able to distinguish between these two peptides that differ by a single amino acid. Additionally, we have found that the beta-chain domain, like its gamma-chain counterpart, binds calcium.
Notes: Everse, S J
Spraggon, G
Veerapandian, L
Riley, M
Doolittle, R F
HL-26873/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1998 Jun 16;37(24):8637-42. doi: 10.1021/bi9804129.
Author Address: Center for Molecular Genetics, University of California, San Diego, La Jolla, California 92093-0634, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1187
Author: Eydoux, C., Spinelli, S., Davis, T. L., Walker, J. R., Seitova, A., Dhe-Paganon, S., De Caro, A., Cambillau, C. and Carrière, F.
Year: 2008
Title: Structure of human pancreatic lipase-related protein 2 with the lid in an open conformation
Journal: Biochemistry
Volume: 47
Issue: 36
Pages: 9553-64
Epub Date: 2008/08/16
Date: Sep 9
Short Title: Structure of human pancreatic lipase-related protein 2 with the lid in an open conformation
Alternate Journal: Biochemistry
ISSN: 0006-2960
DOI: 10.1021/bi8005576
Accession Number: 18702514
Keywords: Animals
Cholinesterase Inhibitors/chemistry
Crystallography, X-Ray
Glycosylation
Humans
Kinetics
Lipase/*chemistry/genetics/metabolism
Lipids/*chemistry/genetics
Paraoxon/chemistry
Protein Structure, Tertiary/physiology
Rats
Abstract: Access to the active site of pancreatic lipase (PL) is controlled by a surface loop, the lid, which normally undergoes conformational changes only upon addition of lipids or amphiphiles. Structures of PL with their lids in the open and functional conformation have required cocrystallization with amphiphiles. Here we report two crystal structures of wild-type and unglycosylated human pancreatic lipase-related protein 2 (HPLRP2) with the lid in an open conformation in the absence of amphiphiles. These structures solved independently are strikingly similar, with some residues of the lid being poorly defined in the electron-density map. The open conformation of the lid is however different from that previously observed in classical liganded PL, suggesting different kinetic properties for HPLRP2. Here we show that the HPLRP2 is directly inhibited by E600, does not present interfacial activation, and acts preferentially on substrates forming monomers or small aggregates (micelles) dispersed in solution like monoglycerides, phospholipids and galactolipids, whereas classical PL displays reverse properties and a high specificity for unsoluble substrates like triglycerides and diglycerides forming oil-in-water interfaces. These biochemical properties imply that the lid of HPLRP2 is likely to spontaneously adopt in solution the open conformation observed in the crystal structure. This open conformation generates a large cavity capable of accommodating the digalactose polar head of galactolipids, similar to that previously observed in the active site of the guinea pig PLRP2, but absent from the classical PL. Most of the structural and kinetic properties of HPLRP2 were found to be different from those of rat PLRP2, the structure of which was previously obtained with the lid in a closed conformation. Our findings illustrate the essential role of the lid in determining the substrate specificity and the mechanism of action of lipases.
Notes: 1520-4995
Eydoux, Cécilia
Spinelli, Silvia
Davis, Tara L
Walker, John R
Seitova, Alma
Dhe-Paganon, Sirano
De Caro, Alain
Cambillau, Christian
Carrière, Frédéric
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
Biochemistry. 2008 Sep 9;47(36):9553-64. doi: 10.1021/bi8005576. Epub 2008 Aug 15.
Author Address: CNRS UPR9025 Laboratoire d'Enzymologie Interfaciale et de Physiologie de la Lipolyse, Marseille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1149
Author: Fabrichny, I. P., Leone, P., Sulzenbacher, G., Comoletti, D., Miller, M. T., Taylor, P., Bourne, Y. and Marchot, P.
Year: 2007
Title: Structural analysis of the synaptic protein neuroligin and its beta-neurexin complex: determinants for folding and cell adhesion
Journal: Neuron
Volume: 56
Issue: 6
Pages: 979-91
Epub Date: 2007/12/21
Date: Dec 20
Short Title: Structural analysis of the synaptic protein neuroligin and its beta-neurexin complex: determinants for folding and cell adhesion
Alternate Journal: Neuron
ISSN: 0896-6273 (Print)
0896-6273
DOI: 10.1016/j.neuron.2007.11.013
PMCID: PMC2703725
NIHMSID: NIHMS109548
Accession Number: 18093521
Keywords: Animals
Calcium/metabolism
Carotenoids/*chemistry/*physiology
Cell Adhesion/*physiology
Cell Line, Transformed
Humans
Membrane Proteins/*chemistry/*physiology
Models, Molecular
Molecular Sequence Data
Mutation
Protein Binding/physiology
*Protein Folding
Protein Structure, Tertiary
Rats
Recombinant Proteins/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Transfection/methods
Abstract: The neuroligins are postsynaptic cell adhesion proteins whose associations with presynaptic neurexins participate in synaptogenesis. Mutations in the neuroligin and neurexin genes appear to be associated with autism and mental retardation. The crystal structure of a neuroligin reveals features not found in its catalytically active relatives, such as the fully hydrophobic interface forming the functional neuroligin dimer; the conformations of surface loops surrounding the vestigial active center; the location of determinants that are critical for folding and processing; and the absence of a macromolecular dipole and presence of an electronegative, hydrophilic surface for neurexin binding. The structure of a beta-neurexin-neuroligin complex reveals the precise orientation of the bound neurexin and, despite a limited resolution, provides substantial information on the Ca2+-dependent interactions network involved in trans-synaptic neurexin-neuroligin association. These structures exemplify how an alpha/beta-hydrolase fold varies in surface topography to confer adhesion properties and provide templates for analyzing abnormal processing or recognition events associated with autism.
Notes: 1097-4199
Fabrichny, Igor P
Leone, Philippe
Sulzenbacher, Gerlind
Comoletti, Davide
Miller, Meghan T
Taylor, Palmer
Bourne, Yves
Marchot, Pascale
P42 ES010337-099005/ES/NIEHS NIH HHS/United States
GM07752/GM/NIGMS NIH HHS/United States
P42 ES010337/ES/NIEHS NIH HHS/United States
R01 GM018360/GM/NIGMS NIH HHS/United States
T32 GM007752/GM/NIGMS NIH HHS/United States
UO-1 ES 10337/ES/NIEHS NIH HHS/United States
R37 GM018360/GM/NIGMS NIH HHS/United States
R37-GM 18360/GM/NIGMS NIH HHS/United States
Comment
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Neuron. 2007 Dec 20;56(6):979-91. doi: 10.1016/j.neuron.2007.11.013.
Author Address: Biochimie des Interactions Moléculaires et Cellulaires, CNRS FRE-2738, Institut Fédératif de Recherche Jean Roche, Université de la Méditerranée, Faculté de Médecine Secteur Nord, F-13916 Marseille Cedex 20, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1656
Author: Faid, V., Denguir, N., Chapuis, V., Bihoreau, N. and Chevreux, G.
Year: 2014
Title: Site-specific N-glycosylation analysis of human factor XI: Identification of a noncanonical NXC glycosite
Journal: Proteomics
Volume: 14
Issue: 21-22
Pages: 2460-70
Epub Date: 2014/08/06
Date: Nov
Short Title: Site-specific N-glycosylation analysis of human factor XI: Identification of a noncanonical NXC glycosite
Alternate Journal: Proteomics
ISSN: 1615-9853
DOI: 10.1002/pmic.201400038
Accession Number: 25092234
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Chromatography, Liquid
Factor XI/*chemistry
Glycosylation
Humans
Molecular Sequence Data
Polysaccharides/*analysis
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tandem Mass Spectrometry
Coagulation factor XI
Glycoproteomics
Mass spectrometry
NXC glycosite
Abstract: Human factor XI (hFXI) is a 160-kDa disulphide-linked homodimer zymogen involved in the coagulation cascade. Its deficiency results in bleeding diathesis referred to as hemophilia C. hFXI bears five N-glycosylation consensus sites per monomer, N72 , N108 , N335 on the heavy chain and N432 , N473 on the light chain. This study reports the first in-depth glycosylation analysis of hFXI based on advanced MS approaches. Hydrophilic interaction LC and MS characterization and quantification of the N-glycans showed that the two major forms are complex biantennary mono-α2,6-sialylated (A2 S1 , 20%) and bis-α2,6-sialylated structures (A2 S2 , 66%). Minor triantennary structures (A3 S3 F, ∼1.5%; A3 S3 , ∼2%) were also identified. MS analyses of intact hFXI revealed full occupation of two of the three heavy-chain glycosites and almost full-site occupancy of the light chain. Analysis of hFXI glycopeptides by LC-MS/MS enabled site-specific glycan profiling and occupancy. It was evidenced that N335 was not glycosylated and that N72 and N108 were fully occupied, whereas N432 and N473 were occupied at about 92 and 95%, respectively. We also identified a new glycosite of the noncanonical format NXC at N145 , occupied at around 5%. These data provide valuable structural information useful to understand the potential roles of N-glycosylation on hFXI function and could serve as a structural reference.
Notes: 1615-9861
Faid, Valegh
Denguir, Nouha
Chapuis, Violaine
Bihoreau, Nicolas
Chevreux, Guillaume
Journal Article
Germany
Proteomics. 2014 Nov;14(21-22):2460-70. doi: 10.1002/pmic.201400038.
Author Address: Analytical Department, LFB Biotechnologies, Courtaboeuf, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1925
Author: Fan, Q., Moen, A., Anonsen, J. H., Bindesbøll, C., Sæther, T., Carlson, C. R. and Grønning-Wang, L. M.
Year: 2018
Title: O-GlcNAc site-mapping of liver X receptor-α and O-GlcNAc transferase
Journal: Biochem Biophys Res Commun
Volume: 499
Issue: 2
Pages: 354-360
Epub Date: 2018/03/27
Date: May 5
Short Title: O-GlcNAc site-mapping of liver X receptor-α and O-GlcNAc transferase
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2018.03.164
Accession Number: 29577901
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Cell Line, Tumor
Humans
Liver X Receptors/chemistry/*metabolism
Mutation/genetics
N-Acetylglucosaminyltransferases/chemistry/*metabolism
Protein Binding
Protein Domains
Transcription, Genetic
*Etd ms
*Hcd ms
*Lxrα
*O-GlcNAc
*Ogt
Abstract: The Liver X Receptor α (LXRα) belongs to the nuclear receptor superfamily and plays an essential role in regulating cholesterol, lipid and glucose metabolism and inflammatory responses. We have previously shown that LXRα is post-translationally modified by O-linked β-N-acetyl-glucosamine (O-GlcNAc) with increased transcriptional activity. Moreover, we showed that LXRα associates with O-GlcNAc transferase (OGT) in vitro and in vivo in mouse liver. In this study, we report that human LXRα is O-GlcNAc modified in its N-terminal domain (NTD) by identifying a specific O-GlcNAc site S49 and a novel O-GlcNAc modified peptide (20)LWKPGAQDASSQAQGGSSCILRE(42). However, O-GlcNAc site-mutations did not modulate LXRα transactivation of selected target gene promoters in vitro. Peptide array and co-immunoprecipitation assays demonstrate that LXRα interacts with OGT in its NTD and ligand-binding domain (LBD) in a ligand-independent fashion. Moreover, we map two new O-GlcNAc sites in the longest OGT isoform (ncOGT): S437 in the tetratricopeptide repeat (TPR) 13 domain and T1043 in the far C-terminus, and a new O-GlcNAc modified peptide (amino acids 826-832) in the intervening region (Int-D) within the catalytic domain. We also map four new O-GlcNAc sites in the short isoform sOGT: S391, T393, S399 and S437 in the TPRs 11-13 domain. Future studies will reveal the biological role of identified O-GlcNAc sites in LXRα and OGT.
Notes: 1090-2104
Fan, Qiong
Moen, Anders
Anonsen, Jan Haug
Bindesbøll, Christian
Sæther, Thomas
Carlson, Cathrine Rein
Grønning-Wang, Line M
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2018 May 5;499(2):354-360. doi: 10.1016/j.bbrc.2018.03.164. Epub 2018 Mar 26.
Author Address: Department of Nutrition, Institute of Basic Medical Sciences, University of Oslo, 1046 Blindern, 0317 Oslo, Norway.
Department of Biosciences, Faculty of Mathematics and Natural Sciences, University of Oslo, 1041 Blindern, 0317 Oslo, Norway.
Department of Biosciences, Faculty of Mathematics and Natural Sciences, University of Oslo, 1041 Blindern, 0317 Oslo, Norway; CIME, Center for Integrative Microbial Evolution, University of Oslo, 1041 Blindern, 0317 Oslo, Norway.
Department of Molecular Medicine, Institute of Basic Medical Sciences, University of Oslo, 1046 Blindern, 0317 Oslo, Norway.
Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway.
Department of Nutrition, Institute of Basic Medical Sciences, University of Oslo, 1046 Blindern, 0317 Oslo, Norway. Electronic address: lmgronningwang@gmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 971
Author: Fan, Q. R. and Hendrickson, W. A.
Year: 2005
Title: Structure of human follicle-stimulating hormone in complex with its receptor
Journal: Nature
Volume: 433
Issue: 7023
Pages: 269-77
Epub Date: 2005/01/22
Date: Jan 20
Short Title: Structure of human follicle-stimulating hormone in complex with its receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature03206
PMCID: PMC5514322
NIHMSID: NIHMS874812 financial interests.
Accession Number: 15662415
Keywords: Amino Acid Sequence
Binding Sites
Crystallization
Dimerization
Follicle Stimulating Hormone, Human/*chemistry/*metabolism
Glycosylation
Humans
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Conformation
Protein Subunits/chemistry/metabolism
Receptors, FSH/*chemistry/*metabolism
Scattering, Radiation
Static Electricity
Structure-Activity Relationship
Substrate Specificity
Abstract: Follicle-stimulating hormone (FSH) is central to reproduction in mammals. It acts through a G-protein-coupled receptor on the surface of target cells to stimulate testicular and ovarian functions. We present here the 2.9-A-resolution structure of a partially deglycosylated complex of human FSH bound to the extracellular hormone-binding domain of its receptor (FSHR(HB)). The hormone is bound in a hand-clasp fashion to an elongated, curved receptor. The buried interface of the complex is large (2,600 A2) and has a high charge density. Our analysis suggests that all glycoprotein hormones bind to their receptors in this mode and that binding specificity is mediated by key interaction sites involving both the common alpha- and hormone-specific beta-subunits. On binding, FSH undergoes a concerted conformational change that affects protruding loops implicated in receptor activation. The FSH-FSHR(HB) complexes form dimers in the crystal and at high concentrations in solution. Such dimers may participate in transmembrane signal transduction.
Notes: 1476-4687
Fan, Qing R
Hendrickson, Wayne A
R01 GM068671/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Nature. 2005 Jan 20;433(7023):269-77. doi: 10.1038/nature03206.
Author Address: Department of Biochemistry and Molecular Biophysics, Columbia University, New York, New York 10032, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 683
Author: Fang, B., Hanover, J. A. and Miller, M. W.
Year: 1998
Title: An evaluation of sialation of the nucleoporin p62
Journal: Arch Biochem Biophys
Volume: 357
Issue: 1
Pages: 95-100
Epub Date: 1998/08/29
Date: Sep 1
Short Title: An evaluation of sialation of the nucleoporin p62
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1998.0767
Accession Number: 9721187
Keywords: Animals
Cell Cycle
Glycosylation
HeLa Cells
Humans
Lectins/metabolism
Membrane Glycoproteins/*metabolism
Mice
N-Acetylneuraminic Acid/*metabolism
Neuroblastoma
Nuclear Pore Complex Proteins
Nuclear Proteins/*metabolism
Rats
Tumor Cells, Cultured
Xenopus laevis
Abstract: Many nuclear and cytosolic proteins are modified by single residues of O-linked N-acetyl-D-glucosamine. These include many proteins found in nuclear pore complexes required for transport of macromolecules between the nucleus and the cytoplasm. The best characterized pore glycoprotein, p62, mediates its function as one component of a protein complex essential for nuclear transport. Although p62 sugar residues are not essential for nuclear transport, they appear to oppose protein phosphorylation occurring at sites predicted to destabilize protein-protein interactions of the p62 complex. Recently, a p62-like protein isolated from mouse neuroblastoma cells was reported to be modified by both GlcNAc and sialic acid. As there is little precedent for nucleoplasmic sialation, the finding that a characterized nuclear pore protein is sialated is significant because it may regulate pore function. To assess the biological importance of p62 sialation, GlcNAc and sialic acid-specific lectins were used to examine the state of p62 glycosylation in cells commonly used to study nuclear transport: frog eggs and normal rat kidney and HeLa fibroblasts. In addition, four mouse neuroblastoma cell lines derived from the same tumor were examined. The glycosylation of p62 in these cells appears to involve only single O-linked GlcNAc moieties; no significant sialation was detected.
Notes: Fang, B
Hanover, J A
Miller, M W
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Arch Biochem Biophys. 1998 Sep 1;357(1):95-100. doi: 10.1006/abbi.1998.0767.
Author Address: Department of Biological Sciences, Wright State University, Dayton, Ohio, 45435-0001, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 839
Author: Farzan, M., Babcock, G. J., Vasilieva, N., Wright, P. L., Kiprilov, E., Mirzabekov, T. and Choe, H.
Year: 2002
Title: The role of post-translational modifications of the CXCR4 amino terminus in stromal-derived factor 1 alpha association and HIV-1 entry
Journal: J Biol Chem
Volume: 277
Issue: 33
Pages: 29484-9
Epub Date: 2002/05/30
Date: Aug 16
Short Title: The role of post-translational modifications of the CXCR4 amino terminus in stromal-derived factor 1 alpha association and HIV-1 entry
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M203361200
Accession Number: 12034737
Keywords: Amino Acid Sequence
Animals
Cell Line
Chemokine CXCL12
Chemokines, CXC/chemistry/*metabolism/physiology
Chondroitin Sulfates/metabolism
Dogs
HIV-1/*physiology
Humans
Membrane Fusion/*physiology
Molecular Sequence Data
*Protein Processing, Post-Translational
Receptors, CXCR4/chemistry/*metabolism/physiology
Sequence Homology, Amino Acid
Serine/metabolism
Tyrosine/analogs & derivatives/metabolism
Abstract: The chemokine receptor CXCR4 plays critical roles in development, immune function, and human immunodeficiency virus type 1 (HIV-1) entry. Here we demonstrate that, like the CC-chemokine receptors CCR5 and CCR2b, CXCR4 is posttranslationally modified by sulfation of its amino-terminal tyrosines. The sulfate group at tyrosine 21 contributes substantially to the ability of CXCR4 to bind its ligand, stromal derived factor 1 alpha. Tyrosine sulfation plays a less significant role in CXCR4-dependent HIV-1 entry than in CCR5-dependent HIV-1 entry. In some cell lines, CXCR4 is efficiently modified by a chondroitin sulfate chain at serine 18, but neither HIV-1 entry nor stromal derived factor 1 alpha binding was affected by loss of this glycosaminoglycan. These data demonstrate a functional role for tyrosine sulfate in the CXC-chemokine receptor family and underscore a general difference in HIV-1 utilization of CCR5 and CXCR4.
Notes: Farzan, Michael
Babcock, Gregory J
Vasilieva, Natalya
Wright, Paulette L
Kiprilov, Enko
Mirzabekov, Tajib
Choe, Hyeryun
R01 AI043891/AI/NIAID NIH HHS/United States
AI43891/AI/NIAID NIH HHS/United States
AI48425/AI/NIAID NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2002 Aug 16;277(33):29484-9. doi: 10.1074/jbc.M203361200. Epub 2002 May 28.
Author Address: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 707
Author: Farzan, M., Mirzabekov, T., Kolchinsky, P., Wyatt, R., Cayabyab, M., Gerard, N. P., Gerard, C., Sodroski, J. and Choe, H.
Year: 1999
Title: Tyrosine sulfation of the amino terminus of CCR5 facilitates HIV-1 entry
Journal: Cell
Volume: 96
Issue: 5
Pages: 667-76
Epub Date: 1999/03/25
Date: Mar 5
Short Title: Tyrosine sulfation of the amino terminus of CCR5 facilitates HIV-1 entry
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/s0092-8674(00)80577-2
Accession Number: 10089882
Keywords: Amino Acid Sequence
Animals
CD4 Antigens/metabolism
Carbohydrate Conformation
Cells, Cultured
Chemokine CCL4
Chlorates/pharmacology
Dogs
Glycosylation
HIV Envelope Protein gp120/metabolism
HIV-1/*physiology
HeLa Cells
Humans
Macrophage Inflammatory Proteins/metabolism
Molecular Sequence Data
*Protein Processing, Post-Translational/drug effects
Receptors, CCR5/*chemistry/physiology
Receptors, CXCR4/chemistry/physiology
Recombinant Fusion Proteins/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Sulfotransferases/metabolism
Tumor Cells, Cultured
Tyrosine/*analogs & derivatives/biosynthesis/physiology
Abstract: Chemokine receptors and related seven-transmembrane-segment (7TMS) receptors serve as coreceptors for entry of human and simian immunodeficiency viruses (HIV-1, HIV-2, and SIV) into target cells. Each of these otherwise diverse coreceptors contains an N-terminal region that is acidic and tyrosine rich. Here, we show that the chemokine receptor CCR5, a principal HIV-1 coreceptor, is posttranslationally modified by O-linked glycosylation and by sulfation of its N-terminal tyrosines. Sulfated tyrosines contribute to the binding of CCR5 to MIP-1 alpha, MIP-1 beta, and HIV-1 gp120/CD4 complexes and to the ability of HIV-1 to enter cells expressing CCR5 and CD4. CXCR4, another important HIV-1 coreceptor, is also sulfated. Tyrosine sulfation may contribute to the natural function of many 7TMS receptors and may be a modification common to primate immunodeficiency virus coreceptors.
Notes: Farzan, M
Mirzabekov, T
Kolchinsky, P
Wyatt, R
Cayabyab, M
Gerard, N P
Gerard, C
Sodroski, J
Choe, H
AI 24755/AI/NIAID NIH HHS/United States
AI 28691/AI/NIAID NIH HHS/United States
AI 41851/AI/NIAID NIH HHS/United States
etc.
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Cell. 1999 Mar 5;96(5):667-76. doi: 10.1016/s0092-8674(00)80577-2.
Author Address: Dana-Farber Cancer Institute, Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA. farzan@mbcrr.harvard.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 786
Author: Faull, K. F., Johnson, J., Kim, M. J., To, T., Whitelegge, J. P., Stevens, R. L., Fluharty, C. B. and Fluharty, A. L.
Year: 2000
Title: Structure of the asparagine-linked sugar chains of porcine kidney and human urine cerebroside sulfate activator protein
Journal: J Mass Spectrom
Volume: 35
Issue: 12
Pages: 1416-24
Epub Date: 2001/02/17
Date: Dec
Short Title: Structure of the asparagine-linked sugar chains of porcine kidney and human urine cerebroside sulfate activator protein
Alternate Journal: Journal of mass spectrometry : JMS
ISSN: 1076-5174 (Print)
1076-5174
DOI: 10.1002/1096-9888(200012)35:12<1416::aid-jms75>3.0.co;2-k
Accession Number: 11180632
Keywords: Animals
Asparagine/*chemistry
*Carbohydrate Conformation
Chromatography, High Pressure Liquid
Glycoproteins/*chemistry/urine
Humans
Kidney/*chemistry
Molecular Structure
Saposins
Spectrometry, Mass, Electrospray Ionization
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Sphingolipid Activator Proteins
Swine
Abstract: The specific sugar residues and their linkages in the oligosaccharides from pig kidney and human urine cerebroside sulfate activator proteins (saposin B), although previously hypothesized, have been unambiguously characterized. Exhaustive sequential exoglycosidase digestion of the trimethyl-p-aminophenyl derivatives, followed by either matrix-assisted laser desorption/ionization and/or mass spectrometry, was used to define the residues and their linkages. The oligosaccharides were enzymatically released from the proteins by treatment with peptidyl-N-glycosidase F and separated from the proteins by reversed-phase high-performance liquid chromatography (HPLC). Reducing termini were converted to the trimethyl-p-aminophenyl derivative and the samples were further purified by normal-phase HPLC. The derivatized carbohydrates were then treated sequentially with a series of exoglycosidases of defined specificity, and the products of each digestion were examined by mass spectrometry. The pentasaccharides from pig kidney and human urine protein were shown to be of the asparagine-linked complex type composed of mannose-alpha 1-6-mannose-beta 1-4-N-acetylglucosamine-N-acetylglucosamine(alpha 1-6-fucose). This highly degraded structure probably represents the final product of intra-lysosomal exoglycosidase digestion. Oligosaccharide sequencing by specific exoglycosidase degradation coupled with mass spectrometry is more rapid than conventional oligosaccharide sequencing. The procedures developed will be useful for sequencing other oligosaccharides including those from other members of the lipid-binding protein class to which cerebroside sulfate activator belongs. (c) 2000 John Wiley & Sons, Ltd.
Notes: Faull, K F
Johnson, J
Kim, M J
To, T
Whitelegge, J P
Stevens, R L
Fluharty, C B
Fluharty, A L
NS31271/NS/NINDS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
J Mass Spectrom. 2000 Dec;35(12):1416-24. doi: 10.1002/1096-9888(200012)35:12<1416::AID-JMS75>3.0.CO;2-K.
Author Address: Department of Chemistry and Biochemistry, UCLA, Los Angeles, California, 90095, USA. faull@chem.ucla.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 881
Author: Feinberg, H., Uitdehaag, J. C., Davies, J. M., Wallis, R., Drickamer, K. and Weis, W. I.
Year: 2003
Title: Crystal structure of the CUB1-EGF-CUB2 region of mannose-binding protein associated serine protease-2
Journal: Embo j
Volume: 22
Issue: 10
Pages: 2348-59
Epub Date: 2003/05/14
Date: May 15
Short Title: Crystal structure of the CUB1-EGF-CUB2 region of mannose-binding protein associated serine protease-2
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/cdg236
PMCID: PMC155994
Accession Number: 12743029
Keywords: Amino Acid Sequence
Animals
CHO Cells
Calcium/metabolism
Calorimetry
Cricetinae
Crystallography, X-Ray
Dimerization
Humans
Mannose-Binding Lectin/chemistry/metabolism
Mannose-Binding Protein-Associated Serine Proteases
Models, Molecular
Molecular Sequence Data
Protein Binding
*Protein Conformation
Rats
Sequence Alignment
Serine Endopeptidases/*chemistry/genetics/metabolism
Abstract: Serum mannose-binding proteins (MBPs) are C-type lectins that recognize cell surface carbohydrate structures on pathogens, and trigger killing of these targets by activating the complement pathway. MBPs circulate as a complex with MBP-associated serine proteases (MASPs), which become activated upon engagement of a target cell surface. The minimal functional unit for complement activation is a MASP homodimer bound to two MBP trimeric subunits. MASPs have a modular structure consisting of an N-terminal CUB domain, a Ca(2+)-binding EGF-like domain, a second CUB domain, two complement control protein modules and a C-terminal serine protease domain. The CUB1-EGF-CUB2 region mediates homodimerization and binding to MBP. The crystal structure of the MASP-2 CUB1-EGF-CUB2 dimer reveals an elongated structure with a prominent concave surface that is proposed to be the MBP-binding site. A model of the full six-domain structure and its interaction with MBPs suggests mechanisms by which binding to a target cell transmits conformational changes from MBP to MASP that allow activation of its protease activity.
Notes: 1460-2075
Feinberg, Hadar
Uitdehaag, Joost C M
Davies, Jason M
Wallis, Russell
Drickamer, Kurt
Weis, William I
R01 GM050565/GM/NIGMS NIH HHS/United States
GM50565/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
EMBO J. 2003 May 15;22(10):2348-59. doi: 10.1093/emboj/cdg236.
Author Address: Departments of Structural Biology and of Molecular and Cellular Physiology, Stanford University School of Medicine, 299 Campus Drive West, CA 94305-5126, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1179
Author: Fenaille, F., Groseil, C., Ramon, C., Riandé, S., Siret, L., Chtourou, S. and Bihoreau, N.
Year: 2008
Title: Mass spectrometric characterization of N- and O-glycans of plasma-derived coagulation factor VII
Journal: Glycoconj J
Volume: 25
Issue: 9
Pages: 827-42
Epub Date: 2008/07/22
Date: Dec
Short Title: Mass spectrometric characterization of N- and O-glycans of plasma-derived coagulation factor VII
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080
DOI: 10.1007/s10719-008-9143-7
Accession Number: 18642129
Keywords: Alkylation
Amino Acid Sequence
Chromatography, Liquid
Electrophoresis, Capillary
Factor VII/analysis/*chemistry
Fluorescence
Glycopeptides/analysis/chemistry
Glycosylation
Humans
*Mass Spectrometry
Molecular Sequence Data
Molecular Weight
Polysaccharides/*chemistry
Protein Subunits/analysis/chemistry
Spectrometry, Mass, Electrospray Ionization
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tandem Mass Spectrometry
Trypsin/metabolism
Abstract: Factor VII (FVII) is a vitamin K-dependent glycoprotein which, in its activated form (FVIIa), participates in the coagulation process by activating factor X and factor IX. FVII is secreted as single peptide chain of 406 residues. Plasma-derived FVII undergoes many post-translational modifications such as gamma-carboxylation, N- and O-glycosylation, beta-hydroxylation. Despite glycosylation of recombinant FVIIa has been fully characterized, nothing is reported on the N- and O-glycans of plasma-derived FVII (pd-FVII) and on their structural heterogeneity at each glycosylation site. N- and O-glycosylation sites and site specific heterogeneity of pd-FVII were studied by various complementary qualitative and quantitative techniques. A MALDI-MS analysis of the native protein indicated that FVII is a 50.1 kDa glycoprotein modified on two sites by diantennary, disialylated non-fucosylated (A2S2) glycans. LC-ESIMS/MS analysis revealed that both light chain and heavy chain were N-glycosylated mainly by A2S2 but also by triantennary sialylated glycans. Nevertheless, lower amounts of triantennary structures were found on Asn(322) compared to Asn(145). Moreover, the triantennary glycans were shown to be fucosylated. In parallel, quantitative analysis of the isolated glycans by capillary electrophoresis indicated that the diantennary structures represented about 50% of the total glycan content. Glycan sequencing using different glycanases led to the identification of triantennary difucosylated structures. Last, MS and MS/MS analysis revealed that FVII is O-glycosylated on the light chain at position Ser(60) and Ser(52) which are modified by oligosaccharide structures such as fucose and Glc(Xyl)(0-1-2), respectively. These latter three O-glycans coexist in equal amounts in plasma-derived FVII.
Notes: 1573-4986
Fenaille, François
Groseil, Catherine
Ramon, Christine
Riandé, Sandrine
Siret, Laurent
Chtourou, Sami
Bihoreau, Nicolas
Journal Article
United States
Glycoconj J. 2008 Dec;25(9):827-42. doi: 10.1007/s10719-008-9143-7. Epub 2008 Jul 20.
Author Address: Laboratoire Français du Fractionnement et des Biotechnologies, 3 avenue des Tropiques, BP 305, Les Ulis, 91958, Courtaboeuf cedex, France. francois.fenaille@cea.fr
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1117
Author: Fenaille, F., Le Mignon, M., Groseil, C., Ramon, C., Riandé, S., Siret, L. and Bihoreau, N.
Year: 2007
Title: Site-specific N-glycan characterization of human complement factor H
Journal: Glycobiology
Volume: 17
Issue: 9
Pages: 932-44
Epub Date: 2007/06/27
Date: Sep
Short Title: Site-specific N-glycan characterization of human complement factor H
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwm060
Accession Number: 17591618
Keywords: Amino Acid Sequence
Chromatography, Liquid
Complement Factor H/*chemistry
Electrophoresis, Capillary
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/chemistry
Peptide Mapping
Polysaccharides/*chemistry
Protein Structure, Tertiary
Spectrometry, Mass, Electrospray Ionization
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Time Factors
Abstract: Human complement factor H (CFH) is a plasma glycoprotein involved in the regulation of the alternative pathway of the complement system. A deficiency in CFH is a cause of severe pathologies like atypical haemolytic uraemic syndrome (aHUS). CFH is a 155-kDa glycoprotein containing nine potential N-glycosylation sites. In the current study, we present a quantitative glycosylation analysis of CFH using capillary electrophoresis and a complete site-specific N-glycan characterization using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) and liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESIMS/MS). A 17.9-kDa mass decrease, observed after glycosidase treatment, indicated that N-glycosylation is the major post-translational modification of CFH. This mass difference is consistent with CFH glycosylation by diantennary disialylated glycans of 2204 Da on eight sites. CFH was not sensitive to endoglycosidase H (Endo H) deglycosylation, indicating the absence of hybrid and oligomannose structures. Quantitative analysis showed that CFH is mainly glycosylated by complex, diantennary disialylated, non-fucosylated glycans. Disialylated fucosylated and monosialylated non-fucosylated oligosaccharides were also identified. MS analysis allowed complete characterization of the protein backbone, verification of the glycosylation sites and site-specific N-glycan identification. The absence of glycosylation at Asn199 of the NGSP sequence of CFH is shown. Asn511, Asn700, Asn784, Asn804, Asn864, Asn893, Asn1011 and Asn1077 are glycosylated essentially by diantennary disialylated structures with a relative distribution varying between 45% for Asn804 and 75% for Asn864. Diantennary monosialylated glycans and triantennary trisialylated fucosylated and non-fucosylated structures have also been identified. Interestingly, the sialylation level along with the amount of triantennary structures decreases from the N- to the C-terminal side of the protein.
Notes: Fenaille, François
Le Mignon, Maxime
Groseil, Catherine
Ramon, Christine
Riandé, Sandrine
Siret, Laurent
Bihoreau, Nicolas
Journal Article
England
Glycobiology. 2007 Sep;17(9):932-44. doi: 10.1093/glycob/cwm060. Epub 2007 Jun 25.
Author Address: Laboratoire français du Fractionnement et des Biotechnologies, Développement Biopharmaceutique, 3 avenue des Tropiques, BP305 Les Ulis, 91958 Courtaboeuf cedex, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1288
Author: Feng, D., Kim, T., Ozkan, E., Light, M., Torkin, R., Teng, K. K., Hempstead, B. L. and Garcia, K. C.
Year: 2010
Title: Molecular and structural insight into proNGF engagement of p75NTR and sortilin
Journal: J Mol Biol
Volume: 396
Issue: 4
Pages: 967-84
Epub Date: 2009/12/29
Date: Mar 5
Short Title: Molecular and structural insight into proNGF engagement of p75NTR and sortilin
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2009.12.030
PMCID: PMC2847487
NIHMSID: HHMIMS184600
Accession Number: 20036257
Keywords: Adaptor Proteins, Vesicular Transport/*chemistry/genetics/*metabolism
Amino Acid Substitution
Animals
Binding Sites
Calcium/metabolism
Cell Line
Crystallography, X-Ray
Humans
In Vitro Techniques
Ligands
Mice
Models, Molecular
Multiprotein Complexes/chemistry
Mutagenesis, Site-Directed
Nerve Growth Factor/*chemistry/genetics/*metabolism
Nerve Tissue Proteins
Protein Interaction Domains and Motifs
Protein Multimerization
Protein Precursors/*chemistry/genetics/*metabolism
Protein Processing, Post-Translational
Protein Stability
Rats
Receptors, Growth Factor
Receptors, Nerve Growth Factor/*chemistry/genetics/*metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Signal Transduction
Surface Plasmon Resonance
Abstract: Nerve growth factor (NGF) is initially synthesized as a precursor, proNGF, that is cleaved to release its C-terminal mature form. Recent studies suggested that proNGF is not an inactive precursor but acts as a signaling ligand distinct from its mature counterpart. proNGF and mature NGF initiate opposing biological responses by utilizing both distinct and shared receptor components. In this study, we carried out structural and biochemical characterization of proNGF interactions with p75NTR and sortilin. We crystallized proNGF complexed to p75NTR and present the structure at 3.75-A resolution. The structure reveals a 2:2 symmetric binding mode, as compared with the asymmetric structure of a previously reported crystal structure of mature NGF complexed to p75NTR and the 2:2 symmetric complex of neurotrophin-3 (NT-3) and p75NTR. Here, we discuss the possible origins and implications of the different stoichiometries. In the proNGF-p75NTR complex, the pro regions of proNGF are mostly disordered and two hairpin loops (loop 2) at the top of the NGF dimer have undergone conformational changes in comparison with mature NT structures, suggesting possible interactions with the propeptide. We further explored the binding characteristics of proNGF to sortilin using surface plasmon resonance and cell-based assays and determined that calcium ions promote the formation of a stable ternary complex of proNGF-sortilin-p75NTR. These results, together with those of previous structural and mechanistic studies of NT-receptor interactions, suggest the potential for distinct signaling activities through p75NTR mediated by different NT-induced conformational changes.
Notes: 1089-8638
Feng, Dan
Kim, Taeho
Ozkan, Engin
Light, Matthew
Torkin, Risa
Teng, Kenneth K
Hempstead, Barbara L
Garcia, K Christopher
R01 NS057627/NS/NINDS NIH HHS/United States
R0I NS057627/NS/NINDS NIH HHS/United States
R0I NS30658/NS/NINDS NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 AI051321/AI/NIAID NIH HHS/United States
R01-AI51321/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Mol Biol. 2010 Mar 5;396(4):967-84. doi: 10.1016/j.jmb.2009.12.030. Epub 2009 Dec 28.
Author Address: Department of Cellular and Molecular Physiology, Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2039
Author: Feng, D., Sheng-Dong, L., Tong, W. and Zhen-Xian, D.
Year: 2020
Title: O-GlcNAcylation of RAF1 increases its stabilization and induces the renal fibrosis
Journal: Biochim Biophys Acta Mol Basis Dis
Volume: 1866
Issue: 3
Pages: 165556
Epub Date: 2019/09/16
Date: Mar 1
Short Title: O-GlcNAcylation of RAF1 increases its stabilization and induces the renal fibrosis
Alternate Journal: Biochimica et biophysica acta. Molecular basis of disease
ISSN: 0925-4439
DOI: 10.1016/j.bbadis.2019.165556
Accession Number: 31521821
Keywords: Acetylglucosamine/metabolism
Acylation/*physiology
Animals
Cell Line
Disease Models, Animal
Epithelial-Mesenchymal Transition/physiology
Fibrosis/*metabolism
Humans
Kidney Diseases/*metabolism
Male
Protein Processing, Post-Translational/physiology
Proto-Oncogene Proteins c-raf/*metabolism
Rats
Rats, Sprague-Dawley
Signal Transduction/physiology
Ubiquitination/physiology
Ureteral Obstruction/metabolism
*Emt
*Ogt
*Raf1
*Renal epithelium
Abstract: Epithelial-mesenchymal transition (EMT) is considered to be one of the most important mechanisms for the progression of renal interstitial fibrosis (RIF). Recently the relationship between post-translational modifications and EMT has been reported. O-GlcNAcylation, one of the key post-translational modifications, was rarely mentioned about its role in EMT, especially in EMT during the process of RIF. The current study aimed to determine whether O-GlcNAcylation participates in the regulation of EMT during RIF. We proved that O-GlcNAcylation prompted the EMT of HK2 cells. Mass spectral analysis identified RAF1 to be one of the O-GlcNAcylated proteins. Moreover, O-GlcNAcylation of RAF1 stabilized RAF1 protein and prompted EMT of HK2 cells. In terms of mechanism, we verified that O-GlcNAcylation of RAF1 inhibited its ubiquitination and thus stabilized RAF1. The upregulation of RAF1 and O-GlcNAcylation products (O-GlcNAc) in vivo were also observed in unilateral ureteral obstruction (UUO) animal models. Collectively, our study indicated that O-GlcNAcylation suppressed the ubiquitination of RAF1, stabilized RAF1 and then modulated the EMT in HK2 cells. These results may give us several new targets for the treatment of RIF.
Notes: 1879-260x
Feng, Du
Sheng-Dong, Li
Tong, Wu
Zhen-Xian, Du
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta Mol Basis Dis. 2020 Mar 1;1866(3):165556. doi: 10.1016/j.bbadis.2019.165556. Epub 2019 Sep 12.
Author Address: Department of Nephrology, Shengjing Hospital of China Medical University, Shenyang 110004, China. Electronic address: dolphine_doctor@hotmail.com.
Department of Endocrinology & Metabolism, the 1st Affiliated Hospital, China Medical University, Shenyang 110001, China.
Department of Endocrinology & Metabolism, the 1st Affiliated Hospital, China Medical University, Shenyang 110001, China. Electronic address: dzx_doctor@qq.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1575
Author: Feng, Z., Hui, Y., Ling, L., Xiaoyan, L., Yuqiu, W., Peng, W. and Lianwen, Z.
Year: 2013
Title: FBXW10 is negatively regulated in transcription and expression level by protein O-GlcNAcylation
Journal: Biochem Biophys Res Commun
Volume: 438
Issue: 2
Pages: 427-32
Epub Date: 2013/08/01
Date: Aug 23
Short Title: FBXW10 is negatively regulated in transcription and expression level by protein O-GlcNAcylation
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2013.07.091
Accession Number: 23899520
Keywords: F-Box Proteins/genetics/*metabolism
Gene Expression Profiling
*Gene Expression Regulation
HEK293 Cells
Humans
N-Acetylglucosaminyltransferases/*metabolism
Plasmids
Protein Processing, Post-Translational
RNA, Messenger/metabolism
*Transcription, Genetic
Ubiquitin/chemistry
Ubiquitination
Cross-talk
F-box protein
Fbxo4
Fbxw10
O-GlcNAcylation
Abstract: Intricate cross-talks exist among multiple post-translational modifications that play critical roles in various cellular events, such as the control of gene expression and regulation of protein function. Here, the cross-talk between O-GlcNAcylation and ubiquitination was investigated in HEK293T cells. By PCR array, 84 ubiquitination-related genes were explored in transcription level in response to the elevation of total protein O-GlcNAcylation due to over-expression of OGT, inhibition of OGA or GlcN treatment. Varied genes were transcriptionally regulated by using different method. But FBXW10, an F-box protein targeting specific proteins for ubiquitination, could be negatively regulated in all ways, suggesting its regulation by protein O-GlcNAcylation. By RT-PCR and Western blot analysis, it was found that FBXW10 could be sharply down-regulated in mRNA and protein level in GlcN-treated cells in a time-dependent way, in line with the enhancement of protein O-GlcNAcylation. It was also found that endogenous FBXW10 was modified by O-GlcNAc in HEK293T cells, implying O-GlcNAcylation might regulate FBXW10 in multiple levels. These findings indicate that O-GlcNAcylation is involved in the regulation of ubiquitination-related genes, and help us understand the cross-talk between O-GlcNAcylation and ubiquitination.
Notes: 1090-2104
Feng, Zhou
Hui, Yan
Ling, Li
Xiaoyan, Liu
Yuqiu, Wang
Peng, Wang
Lianwen, Zhang
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2013 Aug 23;438(2):427-32. doi: 10.1016/j.bbrc.2013.07.091. Epub 2013 Jul 27.
Author Address: State Key Laboratory of Medicinal Chemical Biology, College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300071, PR China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 872
Author: Ferguson, K. M., Berger, M. B., Mendrola, J. M., Cho, H. S., Leahy, D. J. and Lemmon, M. A.
Year: 2003
Title: EGF activates its receptor by removing interactions that autoinhibit ectodomain dimerization
Journal: Mol Cell
Volume: 11
Issue: 2
Pages: 507-17
Epub Date: 2003/03/07
Date: Feb
Short Title: EGF activates its receptor by removing interactions that autoinhibit ectodomain dimerization
Alternate Journal: Molecular cell
ISSN: 1097-2765 (Print)
1097-2765
DOI: 10.1016/s1097-2765(03)00047-9
Accession Number: 12620237
Keywords: Crystallography, X-Ray
Dimerization
Epidermal Growth Factor/*metabolism
ErbB Receptors/antagonists & inhibitors/*chemistry/genetics/*metabolism
Humans
Hydrogen-Ion Concentration
In Vitro Techniques
Models, Molecular
Mutation
Protein Conformation
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Abstract: Epidermal growth factor (EGF) receptor is the prototype of the ErbB (HER) family receptor tyrosine kinases (RTKs), which regulate cell growth and differentiation and are implicated in many human cancers. EGF activates its receptor by inducing dimerization of the 621 amino acid EGF receptor extracellular region. We describe the 2.8 A resolution crystal structure of this entire extracellular region (sEGFR) in an unactivated state. The structure reveals an autoinhibited configuration, where the dimerization interface recently identified in activated sEGFR structures is completely occluded by intramolecular interactions. To activate the receptor, EGF binding must promote a large domain rearrangement that exposes this dimerization interface. This contrasts starkly with other RTK activation mechanisms and suggests new approaches for designing ErbB receptor antagonists.
Notes: Ferguson, Kathryn M
Berger, Mitchell B
Mendrola, Jeannine M
Cho, Hyun Soo
Leahy, Daniel J
Lemmon, Mark A
R01-CA090466/CA/NCI NIH HHS/United States
K01-CA092246/CA/NCI NIH HHS/United States
R01-CA79992/CA/NCI NIH HHS/United States
R21-CA87182/CA/NCI NIH HHS/United States
K01 CA092246/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Mol Cell. 2003 Feb;11(2):507-17. doi: 10.1016/s1097-2765(03)00047-9.
Author Address: Department of Biochemistry and Biophysics, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA. fertuso2@mail.med.upenn.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 637
Author: Ferlinz, K., Hurwitz, R., Moczall, H., Lansmann, S., Schuchman, E. H. and Sandhoff, K.
Year: 1997
Title: Functional characterization of the N-glycosylation sites of human acid sphingomyelinase by site-directed mutagenesis
Journal: Eur J Biochem
Volume: 243
Issue: 1-2
Pages: 511-7
Epub Date: 1997/01/15
Date: Jan 15
Short Title: Functional characterization of the N-glycosylation sites of human acid sphingomyelinase by site-directed mutagenesis
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1997.511_1a.x
Accession Number: 9030779
Keywords: Animals
Asparagine/metabolism
COS Cells
Glycoproteins/metabolism
Glycosylation
Humans
Membrane Glycoproteins/metabolism
Molecular Weight
Mutagenesis, Site-Directed
Phosphorylation
Protein Processing, Post-Translational
Recombinant Proteins
Sphingomyelin Phosphodiesterase/chemistry/*metabolism
Structure-Activity Relationship
Abstract: Most soluble lysosomal enzymes require a mannose-6-phosphate recognition marker present on asparagine-linked oligosaccharides for proper targeting to lysosomes. We have determined the influence of the six potential N-linked oligosaccharide chains of human acid sphingomyelinase (ASM) on catalytic activity, targeting, and processing of the enzyme. Each N-glycosylation site was modified by site-directed mutagenesis and subsequently expressed in COS-1 cells. Evidence is presented that five of these sites are used. Elimination of the four N-terminal glycosylation sites does not disturb lysosomal targeting, processing, or enzymatic activity. However, removal of the two C-terminal N-glycosylation sites inhibits the formation of mature enzyme. Absence of glycosylation site five resulted in rapid cleavage of the primary translation product to an enzymatically inactive protein which accumulated inside the endoplasmic reticulum/Golgi, whereas deletion of glycosylation site six led to the formation of an inactive ASM precursor, also retained inside the endoplasmic reticulum/Golgi. Our results also provide evidence that the site of early proteolytic cleavage of newly synthesized ASM must be located between the second and third glycosylation sites.
Notes: Ferlinz, K
Hurwitz, R
Moczall, H
Lansmann, S
Schuchman, E H
Sandhoff, K
HD 28607/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Eur J Biochem. 1997 Jan 15;243(1-2):511-7. doi: 10.1111/j.1432-1033.1997.511_1a.x.
Author Address: Institut für Organische Chemie und Biochemie, Bonn, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1556
Author: Ferluga, S., Hantgan, R., Goldgur, Y., Himanen, J. P., Nikolov, D. B. and Debinski, W.
Year: 2013
Title: Biological and structural characterization of glycosylation on ephrin-A1, a preferred ligand for EphA2 receptor tyrosine kinase
Journal: J Biol Chem
Volume: 288
Issue: 25
Pages: 18448-57
Epub Date: 2013/05/11
Date: Jun 21
Short Title: Biological and structural characterization of glycosylation on ephrin-A1, a preferred ligand for EphA2 receptor tyrosine kinase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.464008
PMCID: PMC3689987
Accession Number: 23661698
Keywords: Amino Acid Sequence
Binding Sites/genetics
Binding, Competitive
Blotting, Western
Cell Line, Tumor
Cell Movement
Cell Proliferation
Crystallography, X-Ray
Ephrin-A1/chemistry/genetics/*metabolism
Ephrin-A2/genetics/metabolism
Glycosylation
Humans
Ligands
Models, Molecular
Molecular Sequence Data
Mutation
Protein Binding
Protein Multimerization
Protein Structure, Tertiary
Receptor, EphA2/chemistry/genetics/*metabolism
Sequence Homology, Amino Acid
*Signal Transduction
Surface Plasmon Resonance
Crystal Structure
EphA2
Ligand-binding Protein
Receptor
Receptors
Signaling
ephrin-A1
Abstract: The EphA2 receptor tyrosine kinase is overexpressed in a number of malignancies and is activated by ephrin ligands, most commonly by ephrin-A1. The crystal structure of the ligand-receptor complex revealed a glycosylation on the Asn-26 of ephrin-A1. Here we report for the first time the significance of the glycosylation in the biology of EphA2 and ephrin-A1. Ephrin-A1 was enzymatically deglycosylated, and its activity was evaluated in several assays using glioblastoma (GBM) cells and recombinant EphA2. We found that deglycosylated ephrin-A1 does not efficiently induce EphA2 receptor internalization and degradation, and does not activate the downstream signaling pathways involved in cell migration and proliferation. Data obtained by surface plasmon resonance confirms that deglycosylated ephrin-A1 does not bind EphA2 with high affinity. Mutations in the glycosylation site on ephrin-A1 result in protein aggregation and mislocalization. Analysis of Eph/ephrin crystal structures reveals an interaction between the ligand's carbohydrates and two residues of EphA2: Asp-78 and Lys-136. These findings suggest that the glycosylation on ephrin-A1 plays a critical role in the binding and activation of the EphA2 receptor.
Notes: 1083-351x
Ferluga, Sara
Hantgan, Roy
Goldgur, Yehuda
Himanen, Juha P
Nikolov, Dimitar B
Debinski, Waldemar
CA74145/CA/NCI NIH HHS/United States
P30 CA012197/CA/NCI NIH HHS/United States
R01 CA074145/CA/NCI NIH HHS/United States
R01 CA139099/CA/NCI NIH HHS/United States
R01 NS038486/NS/NINDS NIH HHS/United States
NS38486/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2013 Jun 21;288(25):18448-57. doi: 10.1074/jbc.M113.464008. Epub 2013 May 9.
Author Address: Department of Neurosurgery, Comprehensive Cancer Center, Wake Forest School of Medicine, Wake Forest University, Winston-Salem, North Carolina 27157, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1414
Author: Ferrara, C., Grau, S., Jäger, C., Sondermann, P., Brünker, P., Waldhauer, I., Hennig, M., Ruf, A., Rufer, A. C., Stihle, M., Umaña, P. and Benz, J.
Year: 2011
Title: Unique carbohydrate-carbohydrate interactions are required for high affinity binding between FcgammaRIII and antibodies lacking core fucose
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 31
Pages: 12669-74
Epub Date: 2011/07/20
Date: Aug 2
Short Title: Unique carbohydrate-carbohydrate interactions are required for high affinity binding between FcgammaRIII and antibodies lacking core fucose
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1108455108
PMCID: PMC3150898 and Roche Glycart AG.
Accession Number: 21768335
Keywords: Animals
Antibodies/chemistry/*immunology/metabolism
Antibody-Dependent Cell Cytotoxicity/immunology
Antigen-Antibody Complex/chemistry/immunology/metabolism
Binding, Competitive/immunology
CHO Cells
Carbohydrates/chemistry/*immunology
Cells, Cultured
Cricetinae
Cricetulus
Crystallography, X-Ray
Fucose/chemistry/*immunology/metabolism
Glycosylation
Humans
Immunoglobulin Fc Fragments/chemistry/immunology/metabolism
Immunoglobulin G/chemistry/immunology/metabolism
Kinetics
Leukocytes, Mononuclear/immunology
Models, Molecular
Molecular Structure
Protein Binding/immunology
Protein Structure, Tertiary
Receptors, IgG/chemistry/*immunology/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Surface Plasmon Resonance
Abstract: Antibody-mediated cellular cytotoxicity (ADCC), a key immune effector mechanism, relies on the binding of antigen-antibody complexes to Fcγ receptors expressed on immune cells. Antibodies lacking core fucosylation show a large increase in affinity for FcγRIIIa leading to an improved receptor-mediated effector function. Although afucosylated IgGs exist naturally, a next generation of recombinant therapeutic, glycoenginereed antibodies is currently being developed to exploit this finding. In this study, the crystal structures of a glycosylated Fcγ receptor complexed with either afucosylated or fucosylated Fc were determined allowing a detailed, molecular understanding of the regulatory role of Fc-oligosaccharide core fucosylation in improving ADCC. The structures reveal a unique type of interface consisting of carbohydrate-carbohydrate interactions between glycans of the receptor and the afucosylated Fc. In contrast, in the complex structure with fucosylated Fc, these contacts are weakened or nonexistent, explaining the decreased affinity for the receptor. These findings allow us to understand the higher efficacy of therapeutic antibodies lacking the core fucose and also suggest a unique mechanism by which the immune system can regulate antibody-mediated effector functions.
Notes: 1091-6490
Ferrara, Claudia
Grau, Sandra
Jäger, Christiane
Sondermann, Peter
Brünker, Peter
Waldhauer, Inja
Hennig, Michael
Ruf, Armin
Rufer, Arne Christian
Stihle, Martine
Umaña, Pablo
Benz, Jörg
Journal Article
Proc Natl Acad Sci U S A. 2011 Aug 2;108(31):12669-74. doi: 10.1073/pnas.1108455108. Epub 2011 Jul 18.
Author Address: Pharma Research and Early Development, Roche Glycart AG, CH-8952 Schlieren, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1790
Author: Ferrer, C. M., Lu, T. Y., Bacigalupa, Z. A., Katsetos, C. D., Sinclair, D. A. and Reginato, M. J.
Year: 2017
Title: O-GlcNAcylation regulates breast cancer metastasis via SIRT1 modulation of FOXM1 pathway
Journal: Oncogene
Volume: 36
Issue: 4
Pages: 559-569
Epub Date: 2016/06/28
Date: Jan 26
Short Title: O-GlcNAcylation regulates breast cancer metastasis via SIRT1 modulation of FOXM1 pathway
Alternate Journal: Oncogene
ISSN: 0950-9232 (Print)
0950-9232
DOI: 10.1038/onc.2016.228
PMCID: PMC5192006
NIHMSID: NIHMS779391 The authors declare no conflict of interest.
Accession Number: 27345396
Keywords: Animals
Breast Neoplasms/genetics/*metabolism/pathology
Cell Line, Tumor
Cell Proliferation/physiology
Female
Forkhead Box Protein M1/genetics/*metabolism
Glycosylation
Heterografts
Humans
MCF-7 Cells
Mice
Mice, Inbred NOD
N-Acetylglucosaminyltransferases/*metabolism
Neoplasm Metastasis
Sirtuin 1/genetics/*metabolism
Abstract: Tumors utilize aerobic glycolysis to support growth and invasion. However, the molecular mechanisms that link metabolism with invasion are not well understood. The nutrient sensor O-linked-β-N-acetylglucosamine (O-GlcNAc) transferase (OGT) modifies intracellular proteins with N-acetylglucosamine. Cancers display elevated O-GlcNAcylation and suppression of O-GlcNAcylation inhibits cancer invasion and metastasis. Here, we show that the regulation of cancer invasion by OGT is dependent on the NAD(+)-dependent deacetylase SIRT1. Reducing O-GlcNAcylation elevates SIRT1 levels and activity in an AMPK (AMP-activated protein kinase α)-dependent manner. Reduced O-GlcNAcylation in cancer cells leads to SIRT1-mediated proteasomal degradation of oncogenic transcription factor FOXM1 in an MEK/ERK-dependent manner. SIRT1 is critical for OGT-mediated regulation of FOXM1 ubiquitination and reducing SIRT1 activity reverses OGT-mediated regulation of FOXM1. Moreover, we show that SIRT1 levels are required for OGT-mediated regulation of invasion and metastasis in breast cancer cells. Thus, O-GlcNAcylation is a central component linking metabolism to invasion and metastasis via an SIRT1/ERK/FOXM1 axis.
Notes: 1476-5594
Ferrer, C M
Lu, T Y
Bacigalupa, Z A
Katsetos, C D
Sinclair, D A
Reginato, M J
F31 CA183574/CA/NCI NIH HHS/United States
R01 AG028730/AG/NIA NIH HHS/United States
R01 CA155413/CA/NCI NIH HHS/United States
R37 AG028730/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Oncogene. 2017 Jan 26;36(4):559-569. doi: 10.1038/onc.2016.228. Epub 2016 Jun 27.
Author Address: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, USA.
Department of Pathology and Laboratory Medicine, Drexel University College of Medicine, Philadelphia, PA, USA.
Department of Pediatrics, Drexel University College of Medicine, St. Christopher's Hospital for Children, Philadelphia, PA, USA.
Paul F. Glenn Labs for the Biological Mechanisms of Aging, Department of Genetics, Harvard Medical School, Boston, MA, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 500
Author: Fiat, A. M., Jollès, J., Aubert, J. P., Loucheux-Lefebvre, M. H. and Jollès, P.
Year: 1980
Title: Localisation and importance of the sugar part of human casein
Journal: Eur J Biochem
Volume: 111
Issue: 2
Pages: 333-9
Epub Date: 1980/10/01
Date: Oct
Short Title: Localisation and importance of the sugar part of human casein
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1980.tb04946.x
Accession Number: 7460900
Keywords: Amino Acid Sequence
Carbohydrates/analysis
*Caseins
Glycopeptides/analysis
Humans
Peptide Fragments/analysis
Pronase
Protein Conformation
Trypsin
Abstract: The study of glycopeptides obtained after enzymic digestions of human caseinoglycopeptide demonstrated the presence of several (up to 10) prosthetic sugar groups distributed throughout the peptide chain instead of only one in the case of cow and sheep kappa-caseinoglycopeptides. One of the glycopeptides allowed us to complete the C-terminal sequence of human caseinoglycopeptide previously published in part by others. The predicted secondary structure was in accordance with the high number of sugar groups situated all but one in beta-turns. A significant microheterogeneity seems to occur at the sugar level of human caseinoglycopeptide, which might thus present differences from one milk to another.
Notes: Fiat, A M
Jollès, J
Aubert, J P
Loucheux-Lefebvre, M H
Jollès, P
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1980 Oct;111(2):333-9. doi: 10.1111/j.1432-1033.1980.tb04946.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 746
Author: Fiedler, T. J., Davey, C. A. and Fenna, R. E.
Year: 2000
Title: X-ray crystal structure and characterization of halide-binding sites of human myeloperoxidase at 1.8 A resolution
Journal: J Biol Chem
Volume: 275
Issue: 16
Pages: 11964-71
Epub Date: 2000/04/15
Date: Apr 21
Short Title: X-ray crystal structure and characterization of halide-binding sites of human myeloperoxidase at 1.8 A resolution
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.275.16.11964
Accession Number: 10766826
Keywords: Binding Sites
Bromides/*metabolism
Crystallography, X-Ray
Dimerization
Heme/analysis
Humans
Hydrogen Bonding
Models, Chemical
Models, Molecular
Molecular Sequence Data
Peroxidase/*chemistry
Protein Conformation
Abstract: The x-ray crystal structure of human myeloperoxidase has been extended to 1.8 A resolution, using x-ray data recorded at -180 degrees C (r = 0.197, free r = 0.239). Results confirm that the heme is covalently attached to the protein via two ester linkages between the carboxyl groups of Glu(242) and Asp(94) and modified methyl groups on pyrrole rings A and C of the heme as well as a sulfonium ion linkage between the sulfur atom of Met(243) and the beta-carbon of the vinyl group on pyrrole ring A. In the native enzyme a bound chloride ion has been identified at the amino terminus of the helix containing the proximal His(336). Determination of the x-ray crystal structure of a myeloperoxidase-bromide complex (r = 0.243, free r = 0.296) has shown that this chloride ion can be replaced by bromide. Bromide is also seen to bind, at partial occupancy, in the distal heme cavity, in close proximity to the distal His(95), where it replaces the water molecule hydrogen bonded to Gln(91). The bromide-binding site in the distal cavity appears to be the halide-binding site responsible for shifts in the Soret band of the absorption spectrum of myeloperoxidase. It is proposed that halide binding to this site inhibits the enzyme by effectively competing with H(2)O(2) for access to the distal histidine, whereas in compound I, the same site may be the halide substrate-binding site.
Notes: Fiedler, T J
Davey, C A
Fenna, R E
GM 49149/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2000 Apr 21;275(16):11964-71. doi: 10.1074/jbc.275.16.11964.
Author Address: Department of Biochemistry and Molecular Biology, University of Miami Medical School, Miami, Florida 33136, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 354
Author: Field, M. C., Dwek, R. A., Edge, C. J. and Rademacher, T. W.
Year: 1989
Title: O-linked oligosaccharides from human serum immunoglobulin A1
Journal: Biochem Soc Trans
Volume: 17
Issue: 6
Pages: 1034-5
Epub Date: 1989/12/01
Date: Dec
Short Title: O-linked oligosaccharides from human serum immunoglobulin A1
Alternate Journal: Biochemical Society transactions
ISSN: 0300-5127 (Print)
0300-5127
DOI: 10.1042/bst0171034
Accession Number: 2628063
Keywords: Carbohydrate Sequence
Humans
Immunoglobulin A/*analysis
Molecular Conformation
Molecular Sequence Data
Molecular Structure
Oligosaccharides/*blood
Notes: Field, M C
Dwek, R A
Edge, C J
Rademacher, T W
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem Soc Trans. 1989 Dec;17(6):1034-5. doi: 10.1042/bst0171034.
Author Address: Department of Biochemistry, Oxford University, U.K.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2026
Author: Filhoulaud, G., Benhamed, F., Pagesy, P., Bonner, C., Fardini, Y., Ilias, A., Movassat, J., Burnol, A. F., Guilmeau, S., Kerr-Conte, J., Pattou, F., Issad, T. and Postic, C.
Year: 2019
Title: O-GlcNacylation Links TxNIP to Inflammasome Activation in Pancreatic β Cells
Journal: Front Endocrinol (Lausanne)
Volume: 10
Pages: 291
Epub Date: 2019/06/06
Short Title: O-GlcNacylation Links TxNIP to Inflammasome Activation in Pancreatic β Cells
Alternate Journal: Frontiers in endocrinology
ISSN: 1664-2392 (Print)
1664-2392
DOI: 10.3389/fendo.2019.00291
PMCID: PMC6536593
Accession Number: 31164864
Keywords: O-GlcNAcylation
TXNIP (thioredoxin-interacting protein)
hyperglycemia
inflammasome
pancreatic beta cells
Abstract: Thioredoxin interacting protein (TxNIP), which strongly responds to glucose, has emerged as a central mediator of glucotoxicity in pancreatic β cells. TxNIP is a scaffold protein interacting with target proteins to inhibit or stimulate their activity. Recent studies reported that high glucose stimulates the interaction of TxNIP with the inflammasome protein NLRP3 (NLR family, pyrin domain containing 3) to increase interleukin-1 β (IL1β) secretion by pancreatic β cells. To better understand the regulation of TxNIP by glucose in pancreatic β cells, we investigated the implication of O-linked β-N-acetylglucosamine (O-GlcNAcylation) in regulating TxNIP at the posttranslational level. O-GlcNAcylation of proteins is controlled by two enzymes: the O-GlcNAc transferase (OGT), which transfers a monosaccharide to serine/threonine residues on target proteins, and the O-GlcNAcase (OGA), which removes it. Our study shows that TxNIP is subjected to O-GlcNAcylation in response to high glucose concentrations in β cell lines. Modification of the O-GlcNAcylation pathway through manipulation of OGT or OGA expression or activity significantly modulates TxNIP O-GlcNAcylation in INS1 832/13 cells. Interestingly, expression and O-GlcNAcylation of TxNIP appeared to be increased in islets of diabetic rodents. At the mechanistic level, the induction of the O-GlcNAcylation pathway in human and rat islets promotes inflammasome activation as evidenced by enhanced cleaved IL1β. Overexpression of OGT in HEK293 or INS1 832/13 cells stimulates TxNIP and NLRP3 interaction, while reducing TxNIP O-GlcNAcylation through OGA overexpression destabilizes this interaction. Altogether, our study reveals that O-GlcNAcylation represents an important regulatory mechanism for TxNIP activity in β cells.
Notes: 1664-2392
Filhoulaud, Gaelle
Benhamed, Fadila
Pagesy, Patrick
Bonner, Caroline
Fardini, Yann
Ilias, Anissa
Movassat, Jamileh
Burnol, Anne-Françoise
Guilmeau, Sandra
Kerr-Conte, Julie
Pattou, François
Issad, Tarik
Postic, Catherine
Journal Article
Front Endocrinol (Lausanne). 2019 May 21;10:291. doi: 10.3389/fendo.2019.00291. eCollection 2019.
Author Address: INSERM U1016, Institut Cochin, Paris, France.
CNRS UMR 8104, Paris, France.
Université Paris Descartes, Sorbonne Paris Cité, Paris, France.
Pasteur Institute de Lille, Lille, France.
INSERM U1190 - EGID, Lille, France.
UMR8251-CNRS, Paris, France.
Université Paris Diderot, Sorbonne Paris Cité, Paris, France.
CHU, Lille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 338
Author: Fisher, K. J., Tollersrud, O. K. and Aronson, N. N., Jr.
Year: 1990
Title: Cloning and sequence analysis of a cDNA for human glycosylasparaginase. A single gene encodes the subunits of this lysosomal amidase
Journal: FEBS Lett
Volume: 269
Issue: 2
Pages: 440-4
Epub Date: 1990/09/03
Date: Sep 3
Short Title: Cloning and sequence analysis of a cDNA for human glycosylasparaginase. A single gene encodes the subunits of this lysosomal amidase
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(90)81211-6
Accession Number: 2401370
Keywords: Amidohydrolases/*genetics
Amino Acid Sequence
Animals
Aspartylglucosylaminase/*genetics/metabolism
Base Sequence
Blotting, Northern
Cloning, Molecular/methods
DNA/*genetics/isolation & purification
Female
*Genes
Humans
Liver/enzymology
Lysosomes/*enzymology
Macromolecular Substances
Molecular Sequence Data
Placenta/*enzymology
Polymerase Chain Reaction
Pregnancy
Protein Conformation
Rats
Abstract: We have isolated a full-length cDNA (HPAsn.6) for human placenta glycosylasparaginase using a 221-bp PCR amplified fragment containing rat liver asparaginase gene sequences. The deduced amino acid sequence from the human clone showed sequence identity to both the alpha and beta subunits of the rat enzyme. The human enzyme is encoded as a 34.6 kDa polypeptide that is post-translationally processed to generate two subunits of approx. 19.5 (alpha) and 15 (beta) kDa. A charge enriched region is present at the predicted site where cleavage occurs. Using polyclonal antibodies against the alpha and beta subunits of rat liver asparaginase, we have shown that the human enzyme is similar in structure to the rat enzyme.
Notes: Fisher, K J
Tollersrud, O K
Aronson, N N Jr
DK-33314/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
FEBS Lett. 1990 Sep 3;269(2):440-4. doi: 10.1016/0014-5793(90)81211-6.
Author Address: Department of Molecular and Cell Biology, Althouse Laboratory, Pennsylvania State University, University Park 16802.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 679
Author: Flahaut, C., Capon, C., Balduyck, M., Ricart, G., Sautiere, P. and Mizon, J.
Year: 1998
Title: Glycosylation pattern of human inter-alpha-inhibitor heavy chains
Journal: Biochem J
Volume: 333 ( Pt 3)
Issue: Pt 3
Pages: 749-56
Epub Date: 1998/07/25
Date: Aug 1
Short Title: Glycosylation pattern of human inter-alpha-inhibitor heavy chains
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj3330749
PMCID: PMC1219641
Accession Number: 9677337
Keywords: Alpha-Globulins/chemistry/*metabolism
Amino Acid Sequence
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Glycopeptides/chemistry/metabolism
Glycosylation
Humans
Macromolecular Substances
Molecular Sequence Data
Serine Proteinase Inhibitors/chemistry/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Human inter-alpha-inhibitor (IalphaI) is a plasma serine-proteinase inhibitor. It consists of three polypeptide chains covalently linked by a glycosaminoglycan chain: a light chain named bikunin carrying the anti-proteinase activity and two heavy chains, H1 and H2, which exhibit specific properties, e.g. they interact with hyaluronan thus stabilizing the extracellular matrix. In this study, using matrix-assisted laser desorption ionization-time-of-flight MS and amino acid sequencing of tryptic peptides, we provide a detailed analysis of the glycosylation pattern of both heavy chains. H1 carries two complex-type N-glycans of predominantly biantennary structure linked to asparagine residues at positions 256 and 559 respectively. In contrast, the oligosaccharides attached to H2 are a complex-type N-glycan in the N-terminal region of the protein (Asn64) and three to four type-1 core-structure O-glycans mono- or di-sialylated, clustered in the C-terminal region. We propose that these O-glycans might function as a recognition signal for the H2 heavy chain. The biological implications of this hypothesis, notably for the biosynthetic pathway of IalphaI, are discussed.
Notes: 1470-8728
Flahaut, C
Capon, C
Balduyck, M
Ricart, G
Sautiere, P
Mizon, J
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1998 Aug 1;333 ( Pt 3)(Pt 3):749-56. doi: 10.1042/bj3330749.
Author Address: Laboratoire de Biochimie, Faculté de Pharmacie, Université de Lille II, Avenue du Professeur Laguesse, B.P. 83, F-59006 Lille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 455
Author: Flanagan, J. G. and Rabbitts, T. H.
Year: 1982
Title: The sequence of a human immunoglobulin epsilon heavy chain constant region gene, and evidence for three non-allelic genes
Journal: Embo j
Volume: 1
Issue: 5
Pages: 655-60
Epub Date: 1982/01/01
Short Title: The sequence of a human immunoglobulin epsilon heavy chain constant region gene, and evidence for three non-allelic genes
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
PMCID: PMC553102
Accession Number: 6234164
Keywords: Alleles
Amino Acid Sequence
Bacteriophage lambda/genetics
Base Sequence
Cell Line
Cloning, Molecular
*Genes
Humans
Immunoglobulin Constant Regions/*genetics
Immunoglobulin Heavy Chains/*genetics
Immunoglobulin epsilon-Chains/*genetics
Immunoglobulins/*genetics
Nucleic Acid Hybridization
Plasmacytoma/immunology
Polymorphism, Genetic
Abstract: An immunoglobulin epsilon heavy chain gene was isolated from a DNA library of the human epsilon chain-producing myeloma 266B1 , using a JH gene region probe. The gene was shown to be the one expressed in the myeloma by Southern hybridisation analysis and by comparison of nucleotide sequences with the known amino acid sequence of the epsilon chain made by the myeloma. The gene consists of a variable region segment separated from a constant region segment by a 3.5-kb intervening sequence. The complete sequence of the constant region gene segment shows that this segment is split by intervening sequences into four coding segments corresponding to the four constant region domains of the protein. Using the cloned epsilon constant region gene segment as a probe we obtained evidence, from Southern hybridisation analysis, for three non-allelic epsilon constant region genes. An order on the chromosome for these three genes can be predicted from their pattern of retention in myeloma 266B1 DNA.
Notes: 1460-2075
Flanagan, J G
Rabbitts, T H
Journal Article
Embo j. 1982;1(5):655-60.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1335
Author: Fogel, A. I., Li, Y., Giza, J., Wang, Q., Lam, T. T., Modis, Y. and Biederer, T.
Year: 2010
Title: N-glycosylation at the SynCAM (synaptic cell adhesion molecule) immunoglobulin interface modulates synaptic adhesion
Journal: J Biol Chem
Volume: 285
Issue: 45
Pages: 34864-74
Epub Date: 2010/08/27
Date: Nov 5
Short Title: N-glycosylation at the SynCAM (synaptic cell adhesion molecule) immunoglobulin interface modulates synaptic adhesion
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.120865
PMCID: PMC2966101
Accession Number: 20739279
Keywords: Animals
COS Cells
Cell Adhesion/physiology
Cell Adhesion Molecules, Neuronal/*chemistry/genetics/metabolism
Chlorocebus aethiops
Crystallography, X-Ray
Glycosylation
Humans
Mass Spectrometry
Mutation
N-Acetylneuraminic Acid/*chemistry/genetics/metabolism
Presynaptic Terminals/*chemistry/metabolism
Protein Processing, Post-Translational/physiology
Protein Structure, Tertiary
Abstract: Select adhesion molecules connect pre- and postsynaptic membranes and organize developing synapses. The regulation of these trans-synaptic interactions is an important neurobiological question. We have previously shown that the synaptic cell adhesion molecules (SynCAMs) 1 and 2 engage in homo- and heterophilic interactions and bridge the synaptic cleft to induce presynaptic terminals. Here, we demonstrate that site-specific N-glycosylation impacts the structure and function of adhesive SynCAM interactions. Through crystallographic analysis of SynCAM 2, we identified within the adhesive interface of its Ig1 domain an N-glycan on residue Asn(60). Structural modeling of the corresponding SynCAM 1 Ig1 domain indicates that its glycosylation sites Asn(70)/Asn(104) flank the binding interface of this domain. Mass spectrometric and mutational studies confirm and characterize the modification of these three sites. These site-specific N-glycans affect SynCAM adhesion yet act in a differential manner. Although glycosylation of SynCAM 2 at Asn(60) reduces adhesion, N-glycans at Asn(70)/Asn(104) of SynCAM 1 increase its interactions. The modification of SynCAM 1 with sialic acids contributes to the glycan-dependent strengthening of its binding. Functionally, N-glycosylation promotes the trans-synaptic interactions of SynCAM 1 and is required for synapse induction. These results demonstrate that N-glycosylation of SynCAM proteins differentially affects their binding interface and implicate post-translational modification as a mechanism to regulate trans-synaptic adhesion.
Notes: 1083-351x
Fogel, Adam I
Li, Yue
Giza, Joanna
Wang, Qing
Lam, Tukiet T
Modis, Yorgo
Biederer, Thomas
T32 GM007223/GM/NIGMS NIH HHS/United States
R01 DA018928/DA/NIDA NIH HHS/United States
T32 GM007367/GM/NIGMS NIH HHS/United States
P30 DA018343/DA/NIDA NIH HHS/United States
P30 EB009998/EB/NIBIB NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2010 Nov 5;285(45):34864-74. doi: 10.1074/jbc.M110.120865. Epub 2010 Aug 25.
Author Address: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1464
Author: Fong, J. J., Nguyen, B. L., Bridger, R., Medrano, E. E., Wells, L., Pan, S. and Sifers, R. N.
Year: 2012
Title: β-N-Acetylglucosamine (O-GlcNAc) is a novel regulator of mitosis-specific phosphorylations on histone H3
Journal: J Biol Chem
Volume: 287
Issue: 15
Pages: 12195-203
Epub Date: 2012/03/01
Date: Apr 6
Short Title: β-N-Acetylglucosamine (O-GlcNAc) is a novel regulator of mitosis-specific phosphorylations on histone H3
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.315804
PMCID: PMC3320971
Accession Number: 22371497
Keywords: Acetylglucosamine/metabolism/*physiology
Amino Acid Sequence
G2 Phase
Glycosylation
HeLa Cells
Histones/chemistry/*metabolism
Humans
Immunoprecipitation
*Mitosis
Molecular Sequence Data
Peptide Fragments/chemistry
Phosphorylation
*Protein Processing, Post-Translational
Threonine/metabolism
Abstract: O-Linked β-N-acetylglucosamine, or O-GlcNAc, is a dynamic post-translational modification that cycles on and off serine and threonine residues of nucleocytoplasmic proteins. The O-GlcNAc modification shares a complex relationship with phosphorylation, as both modifications are capable of mutually inhibiting the occupation of each other on the same or nearby amino acid residue. In addition to diabetes, cancer, and neurodegenerative diseases, O-GlcNAc appears to play a significant role in cell growth and cell cycle progression, although the precise mechanisms are still not well understood. A recent study also found that all four core nucleosomal histones (H2A, H2B, H3, and H4) are modified with O-GlcNAc, although no specific sites on H3 were reported. Here, we describe that histone H3, a protein highly phosphorylated during mitosis, is modified with O-GlcNAc. Several biochemical assays were used to validate that H3 is modified with O-GlcNAc. Mass spectrometry analysis identified threonine 32 as a novel O-GlcNAc site. O-GlcNAc was detected at higher levels on H3 during interphase than mitosis, which inversely correlated with phosphorylation. Furthermore, increased O-GlcNAcylation was observed to reduce mitosis-specific phosphorylation at serine 10, serine 28, and threonine 32. Finally, inhibiting OGA, the enzyme responsible for removing O-GlcNAc, hindered the transition from G2 to M phase of the cell cycle, displaying a phenotype similar to preventing mitosis-specific phosphorylation on H3. Taken together, these data indicate that O-GlcNAcylation regulates mitosis-specific phosphorylations on H3, providing a mechanistic switch that orchestrates the G2-M transition of the cell cycle.
Notes: 1083-351x
Fong, Jerry J
Nguyen, Brenda L
Bridger, Robert
Medrano, Estela E
Wells, Lance
Pan, Shujuan
Sifers, Richard N
P41 GM103490/GM/NIGMS NIH HHS/United States
R01 CA084282/CA/NCI NIH HHS/United States
R01CA84282/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2012 Apr 6;287(15):12195-203. doi: 10.1074/jbc.M111.315804. Epub 2012 Feb 27.
Author Address: Department of Pathology and Immunology, Baylor College of Medicine, Houston, Texas 77030-3498, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 370
Author: Foster, D. C., Yoshitake, S. and Davie, E. W.
Year: 1985
Title: The nucleotide sequence of the gene for human protein C
Journal: Proc Natl Acad Sci U S A
Volume: 82
Issue: 14
Pages: 4673-7
Epub Date: 1985/07/01
Date: Jul
Short Title: The nucleotide sequence of the gene for human protein C
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.82.14.4673
PMCID: PMC390448
Accession Number: 2991887
Keywords: Amino Acid Sequence
Base Sequence
Blood Coagulation Factors/*genetics
DNA Restriction Enzymes
Factor IX/genetics
*Genes
Glycoproteins/*genetics
Humans
Nucleic Acid Hybridization
Protein C
Protein Conformation
Abstract: A human genomic DNA library was screened for the gene for protein C by using a cDNA probe coding for the human protein. Three different overlapping lambda Charon 4A phage were isolated that contain inserts for the gene for protein C. The complete sequence of the gene was determined by the dideoxy method and shown to span about 11 kilobases of DNA. The coding and 3' noncoding portion of the gene consists of eight exons and seven introns. The eight exons code for a preproleader sequence of 42 amino acids, a light chain of 155 amino acids, a connecting dipeptide of Lys-Arg, and a heavy chain of 262 amino acids. The preproleader sequence and the connecting dipeptide are removed during processing, resulting in the mature protein composed of a heavy and a light chain held together by a disulfide bond. The heavy chain also contains the catalytic region for the serine protease. Two Alu sequences and two homologous repeats of about 160 nucleotides were found in intron E. The seven introns in the gene for protein C are located in essentially the same positions in the amino acid sequence as the seven introns in the gene for human factor IX, while the first three introns in protein C are located in the same positions as the first three in the gene for human prothrombin.
Notes: 1091-6490
Foster, D C
Yoshitake, S
Davie, E W
GM 07270/GM/NIGMS NIH HHS/United States
HL 16919/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1985 Jul;82(14):4673-7. doi: 10.1073/pnas.82.14.4673.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 224
Author: Fournet, B., Montreuil, J., Strecker, G., Dorland, L., Haverkamp, J., Vliegenthart, F. G., Binette, J. P. and Schmid, K.
Year: 1978
Title: Determination of the primary structures of 16 asialo-carbohydrate units derived from human plasma alpha 1-acid glycoprotein by 360-MHZ 1H NMR spectroscopy and permethylation analysis
Journal: Biochemistry
Volume: 17
Issue: 24
Pages: 5206-14
Epub Date: 1978/11/28
Date: Nov 28
Short Title: Determination of the primary structures of 16 asialo-carbohydrate units derived from human plasma alpha 1-acid glycoprotein by 360-MHZ 1H NMR spectroscopy and permethylation analysis
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00617a021
Accession Number: 728395
Keywords: Carbohydrates/*analysis
Glycopeptides/analysis
Humans
Magnetic Resonance Spectroscopy
Methylation
Molecular Conformation
Monosaccharides/analysis
Oligosaccharides/analysis
*Orosomucoid
Sialic Acids
Notes: Fournet, B
Montreuil, J
Strecker, G
Dorland, L
Haverkamp, J
Vliegenthart, F G
Binette, J P
Schmid, K
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1978 Nov 28;17(24):5206-14. doi: 10.1021/bi00617a021.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 934
Author: Franklin, M. C., Carey, K. D., Vajdos, F. F., Leahy, D. J., de Vos, A. M. and Sliwkowski, M. X.
Year: 2004
Title: Insights into ErbB signaling from the structure of the ErbB2-pertuzumab complex
Journal: Cancer Cell
Volume: 5
Issue: 4
Pages: 317-28
Epub Date: 2004/04/20
Date: Apr
Short Title: Insights into ErbB signaling from the structure of the ErbB2-pertuzumab complex
Alternate Journal: Cancer cell
ISSN: 1535-6108 (Print)
1535-6108
DOI: 10.1016/s1535-6108(04)00083-2
Accession Number: 15093539
Keywords: Animals
Antibodies, Monoclonal/*chemistry/genetics/metabolism
Antibodies, Monoclonal, Humanized
Binding Sites
Binding Sites, Antibody
CHO Cells
Cricetinae
Crystallography, X-Ray
Dimerization
Humans
Ligands
Models, Molecular
Mutagenesis, Site-Directed
Protein Structure, Tertiary
Receptor, ErbB-2/*chemistry/genetics/metabolism
*Signal Transduction
Abstract: We have determined the 3.2 A X-ray crystal structure of the extracellular domain of the human epidermal growth factor receptor 2 (ErbB2 or HER2) in a complex with the antigen binding fragment of pertuzumab, an anti-ErbB2 monoclonal antibody also known as 2C4 or Omnitarg. Pertuzumab binds to ErbB2 near the center of domain II, sterically blocking a binding pocket necessary for receptor dimerization and signaling. The ErbB2-pertuzumab structure, combined with earlier mutagenesis data, defines the pertuzumab residues essential for ErbB2 interaction. To analyze the ErbB2 side of the interface, we have mutated a number of residues contacting pertuzumab and examined the effects of these mutations on pertuzumab binding and ErbB2-ErbB3 heterodimerization. We have also shown that conserved residues previously shown to be necessary for EGF receptor homodimerization may be dispensible for ErbB2-ErbB3 heterodimerization.
Notes: Franklin, Matthew C
Carey, Kendall D
Vajdos, Felix F
Leahy, Daniel J
de Vos, Abraham M
Sliwkowski, Mark X
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
Cancer Cell. 2004 Apr;5(4):317-28. doi: 10.1016/s1535-6108(04)00083-2.
Author Address: Department of Protein Engineering, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94114 USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1419
Author: Franklin, M. C., Navarro, E. C., Wang, Y., Patel, S., Singh, P., Zhang, Y., Persaud, K., Bari, A., Griffith, H., Shen, L., Balderes, P. and Kussie, P.
Year: 2011
Title: The structural basis for the function of two anti-VEGF receptor 2 antibodies
Journal: Structure
Volume: 19
Issue: 8
Pages: 1097-107
Epub Date: 2011/08/11
Date: Aug 10
Short Title: The structural basis for the function of two anti-VEGF receptor 2 antibodies
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126
DOI: 10.1016/j.str.2011.01.019
Accession Number: 21827946
Keywords: Amino Acid Motifs
Amino Acid Sequence
Angiogenesis Inhibitors/*chemistry
Antibodies, Monoclonal/*chemistry
Antibodies, Monoclonal, Humanized
Antibody Specificity
Binding Sites, Antibody
Crystallography, X-Ray
Enzyme-Linked Immunosorbent Assay
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Binding
Protein Structure, Quaternary
Protein Structure, Tertiary
Vascular Endothelial Growth Factor Receptor-2/*chemistry/genetics
Abstract: The anti-VEGF receptor 2 antibody IMC-1121B is a promising antiangiogenic drug being tested for treatment of breast and gastric cancer. We have determined the structure of the 1121B Fab fragment in complex with domain 3 of VEGFR2, as well as the structure of a different neutralizing anti-VEGFR2 antibody, 6.64, also in complex with VEGFR2 domain 3. The two Fab fragments bind at opposite ends of VEGFR2 domain 3; 1121B directly blocks VEGF binding, whereas 6.64 may prevent receptor dimerization by perturbing the domain 3:domain 4 interface. Mutagenesis reveals that residues essential for VEGF, 1121B, and 6.64 binding are nonoverlapping among the three contact patches.
Notes: 1878-4186
Franklin, Matthew C
Navarro, Elizabeth C
Wang, Yujie
Patel, Sheetal
Singh, Pinki
Zhang, Yi
Persaud, Kris
Bari, Amtul
Griffith, Heather
Shen, Leyi
Balderes, Paul
Kussie, Paul
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
United States
Structure. 2011 Aug 10;19(8):1097-107. doi: 10.1016/j.str.2011.01.019.
Author Address: Department of Immunology, ImClone Systems, New York, NY 10014, USA. mfranklin@nysbc.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 465
Author: Frantíková, V., Borvák, J., Kluh, I. and Morávek, L.
Year: 1984
Title: Amino acid sequence of the N-terminal region of human hemopexin
Journal: FEBS Lett
Volume: 178
Issue: 2
Pages: 213-6
Epub Date: 1984/12/10
Date: Dec 10
Short Title: Amino acid sequence of the N-terminal region of human hemopexin
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(84)80603-1
Accession Number: 6510521
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Chromatography, Gel
Chromatography, High Pressure Liquid
Cyanogen Bromide
Disulfides
*Hemopexin
Humans
Oligosaccharides
*Peptide Fragments
Abstract: Cyanogen bromide digestion of hemopexin at its 6 methionine residues results in 7 fragments (CB1-CB7) partially connected by disulfide bridges. By sequence studies of fragments CB1-CB4 and peptides prepared by their enzyme cleavage, a continuous amino acid sequence of the N-terminal region of human hemopexin, comprising 220 amino acid residues, was determined. The presence of intramolecular disulfide bonds, connecting half-cystine residues 126/130 and 165/170, was proved in fragments CB2 and CB3. Fragments CB1-CB4 include 5 sites, where hexosamine oligosaccharides are attached (positions 1,41,164,217 and probably 223). In the sequenced region two sites sensitive to acid hydrolysis--bonds ... Asp--Pro ... in positions 20/21 and 187/188 were found. In spite of the fact that pooled material of many donors was studied, no sequence heterogeneity was discovered.
Notes: Frantíková, V
Borvák, J
Kluh, I
Morávek, L
Journal Article
England
FEBS Lett. 1984 Dec 10;178(2):213-6. doi: 10.1016/0014-5793(84)80603-1.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1097
Author: Frasco, M. F., Colletier, J. P., Weik, M., Carvalho, F., Guilhermino, L., Stojan, J. and Fournier, D.
Year: 2007
Title: Mechanisms of cholinesterase inhibition by inorganic mercury
Journal: Febs j
Volume: 274
Issue: 7
Pages: 1849-61
Epub Date: 2007/03/16
Date: Apr
Short Title: Mechanisms of cholinesterase inhibition by inorganic mercury
Alternate Journal: The FEBS journal
ISSN: 1742-464X (Print)
1742-464x
DOI: 10.1111/j.1742-4658.2007.05732.x
Accession Number: 17355286
Keywords: Acetylcholinesterase/chemistry/genetics
Animals
Binding Sites
Butyrylcholinesterase/chemistry/genetics
Cholinesterase Inhibitors/*chemistry
Cholinesterases/*chemistry/genetics
Crystallography, X-Ray
Cysteine/chemistry
Drosophila melanogaster/enzymology/genetics
Electrophorus/genetics/metabolism
Humans
Kinetics
Light
Mercuric Chloride/*chemistry
Models, Chemical
Models, Molecular
Nitrobenzenes/chemistry
Phenylacetates/chemistry
Protein Conformation
Recombinant Proteins/chemistry
Scattering, Radiation
Torpedo/genetics/metabolism
Abstract: The poorly known mechanism of inhibition of cholinesterases by inorganic mercury (HgCl2) has been studied with a view to using these enzymes as biomarkers or as biological components of biosensors to survey polluted areas. The inhibition of a variety of cholinesterases by HgCl2 was investigated by kinetic studies, X-ray crystallography, and dynamic light scattering. Our results show that when a free sensitive sulfhydryl group is present in the enzyme, as in Torpedo californica acetylcholinesterase, inhibition is irreversible and follows pseudo-first-order kinetics that are completed within 1 h in the micromolar range. When the free sulfhydryl group is not sensitive to mercury (Drosophila melanogaster acetylcholinesterase and human butyrylcholinesterase) or is otherwise absent (Electrophorus electricus acetylcholinesterase), then inhibition occurs in the millimolar range. Inhibition follows a slow binding model, with successive binding of two mercury ions to the enzyme surface. Binding of mercury ions has several consequences: reversible inhibition, enzyme denaturation, and protein aggregation, protecting the enzyme from denaturation. Mercury-induced inactivation of cholinesterases is thus a rather complex process. Our results indicate that among the various cholinesterases that we have studied, only Torpedo californica acetylcholinesterase is suitable for mercury detection using biosensors, and that a careful study of cholinesterase inhibition in a species is a prerequisite before using it as a biomarker to survey mercury in the environment.
Notes: Frasco, Manuela F
Colletier, Jacques-Philippe
Weik, Martin
Carvalho, Félix
Guilhermino, Lúcia
Stojan, Jure
Fournier, Didier
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2007 Apr;274(7):1849-61. doi: 10.1111/j.1742-4658.2007.05732.x. Epub 2007 Mar 12.
Author Address: ICBAS, Instituto de Ciências Biomédicas de Abel Salazar, Universidade do Porto, Portugal.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1185
Author: Frederick, J. P., Tafari, A. T., Wu, S. M., Megosh, L. C., Chiou, S. T., Irving, R. P. and York, J. D.
Year: 2008
Title: A role for a lithium-inhibited Golgi nucleotidase in skeletal development and sulfation
Journal: Proc Natl Acad Sci U S A
Volume: 105
Issue: 33
Pages: 11605-12
Epub Date: 2008/08/13
Date: Aug 19
Short Title: A role for a lithium-inhibited Golgi nucleotidase in skeletal development and sulfation
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0801182105
PMCID: PMC2575314
Accession Number: 18695242
Keywords: Animals
Animals, Newborn
Body Patterning
Bone and Bones/*embryology/*enzymology
Cartilage/embryology/enzymology
Cells, Cultured
Chondrodysplasia Punctata/embryology/enzymology/genetics
Chondroitin/metabolism
Embryo, Nonmammalian/embryology/enzymology
Enzyme Inhibitors/*pharmacology
Gene Expression Regulation, Developmental
Gene Expression Regulation, Enzymologic
Golgi Apparatus/*drug effects/*enzymology
Growth Plate/abnormalities/enzymology
Heparitin Sulfate/metabolism
Lithium/*pharmacology
Male
Mice
Mice, Transgenic
Nucleotidases/*antagonists & inhibitors/genetics/metabolism
Phylogeny
Sulfur/*metabolism
Abstract: Sulfation is an important biological process that modulates the function of numerous molecules. It is directly mediated by cytosolic and Golgi sulfotransferases, which use 3'-phosphoadenosine 5'-phosphosulfate to produce sulfated acceptors and 3'-phosphoadenosine 5'-phosphate (PAP). Here, we identify a Golgi-resident PAP 3'-phosphatase (gPAPP) and demonstrate that its activity is potently inhibited by lithium in vitro. The inactivation of gPAPP in mice led to neonatal lethality, lung abnormalities resembling atelectasis, and dwarfism characterized by aberrant cartilage morphology. The phenotypic similarities of gPAPP mutant mice to chondrodysplastic models harboring mutations within components of the sulfation pathway lead to the discovery of undersulfated chondroitin in the absence of functional enzyme. Additionally, we observed loss of gPAPP leads to perturbations in the levels of heparan sulfate species in lung tissue and whole embryos. Our data are consistent with a model that clearance of the nucleotide product of sulfotransferases within the Golgi plays an important role in glycosaminoglycan sulfation, provide a unique genetic basis for chondrodysplasia, and define a function for gPAPP in the formation of skeletal elements derived through endochondral ossification.
Notes: 1091-6490
Frederick, Joshua P
Tafari, A Tsahai
Wu, Sheue-Mei
Megosh, Louis C
Chiou, Shean-Tai
Irving, Ryan P
York, John D
R01 HL055672/HL/NHLBI NIH HHS/United States
R01 HL-55672/HL/NHLBI NIH HHS/United States
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2008 Aug 19;105(33):11605-12. doi: 10.1073/pnas.0801182105. Epub 2008 Aug 11.
Author Address: Department of Pharmacology and Cancer Biology, Howard Hughes Medical Institute, Duke University Medical Center, Durham, NC 27710, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1173
Author: Fredslund, F., Laursen, N. S., Roversi, P., Jenner, L., Oliveira, C. L., Pedersen, J. S., Nunn, M. A., Lea, S. M., Discipio, R., Sottrup-Jensen, L. and Andersen, G. R.
Year: 2008
Title: Structure of and influence of a tick complement inhibitor on human complement component 5
Journal: Nat Immunol
Volume: 9
Issue: 7
Pages: 753-60
Epub Date: 2008/06/10
Date: Jul
Short Title: Structure of and influence of a tick complement inhibitor on human complement component 5
Alternate Journal: Nature immunology
ISSN: 1529-2908
DOI: 10.1038/ni.1625
Accession Number: 18536718
Keywords: Animals
Arthropod Proteins
Binding Sites
Carrier Proteins
Complement C3
Complement C5/*chemistry/*metabolism
Crystallography, X-Ray
Humans
Insect Proteins/chemistry/*metabolism
*Protein Structure, Quaternary
Surface Plasmon Resonance
Abstract: To provide insight into the structural and functional properties of human complement component 5 (C5), we determined its crystal structure at a resolution of 3.1 A. The core of C5 adopted a structure resembling that of C3, with the domain arrangement at the position corresponding to the C3 thioester being very well conserved. However, in contrast to C3, the convertase cleavage site in C5 was ordered and the C345C domain flexibly attached to the core of C5. Binding of the tick C5 inhibitor OmCI to C5 resulted in stabilization of the global conformation of C5 but did not block the convertase cleavage site. The structure of C5 may render possible a structure-based approach for the design of new selective complement inhibitors.
Notes: 1529-2916
Fredslund, Folmer
Laursen, Nick S
Roversi, Pietro
Jenner, Lasse
Oliveira, Cristiano L P
Pedersen, Jan S
Nunn, Miles A
Lea, Susan M
Discipio, Richard
Sottrup-Jensen, Lars
Andersen, Gregers R
BB/D003628/1/Biotechnology and Biological Sciences Research Council/United Kingdom
G0400775/Medical Research Council/United Kingdom
Journal Article
United States
Nat Immunol. 2008 Jul;9(7):753-60. doi: 10.1038/ni.1625. Epub 2008 Jun 8.
Author Address: Department of Molecular Biology, University of Aarhus, DK-8000 Aarhus, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 788
Author: Freeman, S., Birrell, H. C., D'Alessio, K., Erickson-Miller, C., Kikly, K. and Camilleri, P.
Year: 2001
Title: A comparative study of the asparagine-linked oligosaccharides on siglec-5, siglec-7 and siglec-8, expressed in a CHO cell line, and their contribution to ligand recognition
Journal: Eur J Biochem
Volume: 268
Issue: 5
Pages: 1228-37
Epub Date: 2001/03/07
Date: Mar
Short Title: A comparative study of the asparagine-linked oligosaccharides on siglec-5, siglec-7 and siglec-8, expressed in a CHO cell line, and their contribution to ligand recognition
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.2001.01985.x
Accession Number: 11231274
Keywords: Amino Acid Motifs
Amino Acid Sequence
Animals
Antigens, CD/chemistry/genetics/metabolism
Antigens, Differentiation, B-Lymphocyte/chemistry/genetics/metabolism
Antigens, Differentiation, Myelomonocytic/*chemistry/genetics/*metabolism
Asparagine/*metabolism
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Erythrocytes/metabolism
Glycosylation
Humans
*Lectins
Ligands
Mass Spectrometry
Membrane Glycoproteins/*chemistry/genetics/*metabolism
Molecular Sequence Data
Mutagenesis, Site-Directed/genetics
N-Acetylneuraminic Acid/metabolism
Neuraminidase/metabolism
Oligosaccharides/*analysis/chemistry/metabolism
Protein Binding
Sequence Alignment
alpha-L-Fucosidase/metabolism
Abstract: The siglecs (sialic acid-binding immunoglobulin-like lectins) mediate sialic acid-dependent cellular interactions and may in some cases signal through SH2-binding domains. In addition to the previously characterized siglecs, sialoadhesin, CD22, CD33 and myelin-associated glycoprotein, several new ones, siglec-5, siglec-7 and siglec-8, have recently been cloned. Although these novel receptors have generated considerable interest as therapeutic targets because of their expression pattern on immune cells, very little is known about how their lectin activity is regulated. Previous studies with sialoadhesin, CD22 and CD33 have shown that siglec glycosylation has significant effects on binding. To determine any differences in the glycan composition of siglec-5, siglec-7 and siglec-8 that may modify their function, we released and characterized the N-linked oligosaccharide distribution in these three glycoproteins. The glycan pools from siglec-5 and siglec-7 contained a larger proportion of sialylated and core-fucosylated biantennary, triantennary and tetra-antennary oligosaccharides, whereas the carbohydrate mixture released from siglec-8 is noticeably less sialylated and is more abundant in 'high-mannose'-type glycans. In addition, we show that, in contrast with CD22 and CD33, mutating the conserved potentially N-linked glycosylation site in the first domain has no effect on binding mediated by siglec-5 or siglec-7.
Notes: Freeman, S
Birrell, H C
D'Alessio, K
Erickson-Miller, C
Kikly, K
Camilleri, P
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 2001 Mar;268(5):1228-37. doi: 10.1046/j.1432-1327.2001.01985.x.
Author Address: University of Oxford, Institute of Molecular Medicine, John Radcliffe Hospital, Headington, Oxford, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 691
Author: Fremont, D. H., Crawford, F., Marrack, P., Hendrickson, W. A. and Kappler, J.
Year: 1998
Title: Crystal structure of mouse H2-M
Journal: Immunity
Volume: 9
Issue: 3
Pages: 385-93
Epub Date: 1998/10/13
Date: Sep
Short Title: Crystal structure of mouse H2-M
Alternate Journal: Immunity
ISSN: 1074-7613 (Print)
1074-7613
DOI: 10.1016/s1074-7613(00)80621-4
Accession Number: 9768758
Keywords: Amino Acid Sequence
Animals
Conserved Sequence
Crystallography, X-Ray
HLA-D Antigens/*chemistry/isolation & purification/physiology
Histocompatibility Antigens Class I/chemistry
Histocompatibility Antigens Class II/chemistry
Mice
Mice, Inbred C57BL
Molecular Sequence Data
Protein Conformation
Protein Structure, Tertiary
Solubility
Abstract: H2-M (HLA-DM in humans) resides in an acidic endosomal compartment, where it facilitates the loading of antigenic peptides into the peptide-binding groove of class II MHC. The crystal structure of a soluble form of H2-M has been solved to 3.1 A resolution, revealing a heterodimer with structural similarities to the MHC family of proteins. In contrast to its antigen-presenting cousins, the membrane distal alpha helices of H2-M pack closely together, occluding most of the binding groove except for a single large pocket near the center. The structure of H2-M has several unique features that may play a role in its function as a molecular chaperone and peptide exchange factor.
Notes: Fremont, D H
Crawford, F
Marrack, P
Hendrickson, W A
Kappler, J
AI-17134/AI/NIAID NIH HHS/United States
AI-18785/AI/NIAID NIH HHS/United States
AI-22295/AI/NIAID NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Immunity. 1998 Sep;9(3):385-93. doi: 10.1016/s1074-7613(00)80621-4.
Author Address: Center for Immunology, Department of Pathology, Washington University School of Medicine, St. Louis, Missouri 63110, USA. fremont@immunology.wustl.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1829
Author: Freund, P., Kerenyi, M. A., Hager, M., Wagner, T., Wingelhofer, B., Pham, H. T. T., Elabd, M., Han, X., Valent, P., Gouilleux, F., Sexl, V., Krämer, O. H., Groner, B. and Moriggl, R.
Year: 2017
Title: O-GlcNAcylation of STAT5 controls tyrosine phosphorylation and oncogenic transcription in STAT5-dependent malignancies
Journal: Leukemia
Volume: 31
Issue: 10
Pages: 2132-2142
Epub Date: 2017/01/12
Date: Oct
Short Title: O-GlcNAcylation of STAT5 controls tyrosine phosphorylation and oncogenic transcription in STAT5-dependent malignancies
Alternate Journal: Leukemia
ISSN: 0887-6924 (Print)
0887-6924
DOI: 10.1038/leu.2017.4
PMCID: PMC5629373
Accession Number: 28074064
Keywords: Acetylglucosamine/*metabolism
Animals
Cell Line
*Cell Transformation, Neoplastic
Female
Gene Expression Regulation, Neoplastic
Genes, Reporter
Glycosylation
Humans
Interleukin-3/pharmacology
Lymphoid Tissue/cytology
Male
Mice
Mutagenesis, Site-Directed
Myeloproliferative Disorders/*etiology/genetics
Phosphorylation
Phosphotyrosine/metabolism
*Protein Processing, Post-Translational
Radiation Chimera
Recombinant Fusion Proteins/metabolism
STAT5 Transcription Factor/genetics/*metabolism
Signal Transduction
T-Lymphocytes/cytology/drug effects/metabolism
Threonine/metabolism
*Transcriptional Activation
Tumor Suppressor Proteins/genetics/*metabolism
Abstract: The signal transducer and activator of transcription 5 (STAT5) regulates differentiation, survival, proliferation and transformation of hematopoietic cells. Upon cytokine stimulation, STAT5 tyrosine phosphorylation (pYSTAT5) is transient, while in diverse neoplastic cells persistent overexpression and enhanced pYSTAT5 are frequently found. Post-translational modifications might contribute to enhanced STAT5 activation in the context of transformation, but the strength and duration of pYSTAT5 are incompletely understood. We found that O-GlcNAcylation and tyrosine phosphorylation act together to trigger pYSTAT5 levels and oncogenic transcription in neoplastic cells. The expression of a mutated hyperactive gain-of-function (GOF) STAT5 without O-GlcNAcylation resulted in decreased tyrosine phosphorylation, oligomerization and transactivation potential and complete loss of oncogenic transformation capacity. The lack of O-GlcNAcylation diminished phospho-ERK and phospho-AKT levels. Our data show that O-GlcNAcylation of STAT5 is an important process that contributes to oncogenic transcription through enhanced STAT5 tyrosine phosphorylation and oligomerization driving myeloid transformation. O-GlcNAcylation of STAT5 could be required for nutrient sensing and metabolism of cancer cells.
Notes: 1476-5551
Freund, P
Kerenyi, M A
Hager, M
Wagner, T
Wingelhofer, B
Pham, H T T
Elabd, M
Han, X
Valent, P
Gouilleux, F
Sexl, V
Krämer, O H
Groner, B
Moriggl, R
Orcid: 0000-0003-0918-9463
R21 AI103388/AI/NIAID NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Leukemia. 2017 Oct;31(10):2132-2142. doi: 10.1038/leu.2017.4. Epub 2017 Jan 11.
Author Address: Ludwig Boltzmann Institute for Cancer Research, Vienna, Austria.
Department of Biomedical Science, Institute of Animal Breeding and Genetics, University of Veterinary Medicine, Vienna, Austria.
Department of Pharmacology, Boehringer Ingelheim RCV GmbH &Co KG, Vienna, Austria.
Department of Biochemistry, Center for Molecular Biomedicine, Friedrich Schiller University Jena, Jena, Germany.
Division of Gastroenterology, Hepatology and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA.
Key Laboratory of Human Disease Comparative Medicine, Ministry of Health; Institute of Laboratory Animal Science, Chinese Academy of Medical Sciences (CAMS) and Peking Union Medical College (PUMC), Beijing, China.
Department of Internal Medicine I, Division Hematology and Hemostaseology and Ludwig Boltzmann Cluster Oncology, Medical University of Vienna, Vienna, Austria.
CNRS UMR 7292, Université François Rabelais, Tours, France.
Department of Biomedical Science, Institute of Pharmacology and Toxicology, University of Veterinary Medicine, Vienna, Austria.
Department of Toxicology, University Medical Center, Mainz, Germany.
Georg Speyer Haus, Institute for Tumor Biology and Experimental Therapy, Frankfurt am Main, Germany.
Medical University of Vienna, Vienna, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 955
Author: Friedmann, E., Lemberg, M. K., Weihofen, A., Dev, K. K., Dengler, U., Rovelli, G. and Martoglio, B.
Year: 2004
Title: Consensus analysis of signal peptide peptidase and homologous human aspartic proteases reveals opposite topology of catalytic domains compared with presenilins
Journal: J Biol Chem
Volume: 279
Issue: 49
Pages: 50790-8
Epub Date: 2004/09/24
Date: Dec 3
Short Title: Consensus analysis of signal peptide peptidase and homologous human aspartic proteases reveals opposite topology of catalytic domains compared with presenilins
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M407898200
Accession Number: 15385547
Keywords: Aspartic Acid Endopeptidases/*chemistry
Binding Sites
Blotting, Western
Catalysis
Catalytic Domain
Cell Line
Cell-Free System
Cloning, Molecular
Cytosol/metabolism
DNA, Complementary/metabolism
Electrophoresis, Polyacrylamide Gel
Fluorescent Antibody Technique, Indirect
Gene Library
Glycosylation
HeLa Cells
Humans
Membrane Proteins/*chemistry
Microscopy, Fluorescence
Oligonucleotide Array Sequence Analysis
Phylogeny
Plasmids/metabolism
Polysaccharides/chemistry
Presenilin-1
Protein Biosynthesis
Protein Structure, Tertiary
Protein Transport
RNA, Messenger/metabolism
Tissue Distribution
Transcription, Genetic
Abstract: The human genome encodes seven intramembrane-cleaving GXGD aspartic proteases. These are the two presenilins that activate signaling molecules and are implicated in Alzheimer's disease, signal peptide peptidase (SPP), required for immune surveillance, and four SPP-like candidate proteases (SPPLs), of unknown function. Here we describe a comparative analysis of the topologies of SPP and its human homologues, SPPL2a, -2b, -2c, and -3. We demonstrate that their N-terminal extensions are located in the extracellular space and, except for SPPL3, are modified with N-glycans. Whereas SPPL2a, -2b, and -2c contain a signal sequence, SPP and SPPL3 contain a type I signal anchor sequence for initiation of protein translocation and membrane insertion. The hydrophilic loops joining the transmembrane regions, which contain the catalytic residues, are facing the exoplasm. The C termini of all these proteins are exposed toward the cytosol. Taken together, our study demonstrates that SPP and its homologues are all of the same principal structure with a catalytic domain embedded in the membrane in opposite orientation to that of presenilins. Other than presenilins, SPPL2a, -2b, -2c, and -3 are therefore predicted to cleave type II-oriented substrate peptides like the prototypic protease SPP.
Notes: Friedmann, Elena
Lemberg, Marius K
Weihofen, Andreas
Dev, Kumlesh K
Dengler, Uwe
Rovelli, Giorgio
Martoglio, Bruno
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2004 Dec 3;279(49):50790-8. doi: 10.1074/jbc.M407898200. Epub 2004 Sep 21.
Author Address: Institute of Biochemistry, Swiss Federal Institute of Technology (ETH), ETH-Hoenggerberg, 8093 Zurich, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 960
Author: Fritz, T. A., Hurley, J. H., Trinh, L. B., Shiloach, J. and Tabak, L. A.
Year: 2004
Title: The beginnings of mucin biosynthesis: the crystal structure of UDP-GalNAc:polypeptide alpha-N-acetylgalactosaminyltransferase-T1
Journal: Proc Natl Acad Sci U S A
Volume: 101
Issue: 43
Pages: 15307-12
Epub Date: 2004/10/16
Date: Oct 26
Short Title: The beginnings of mucin biosynthesis: the crystal structure of UDP-GalNAc:polypeptide alpha-N-acetylgalactosaminyltransferase-T1
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0405657101
PMCID: PMC524453
Accession Number: 15486088
Keywords: Animals
Binding Sites
Catalytic Domain
Crystallography, X-Ray
Lectins/metabolism
Mice
Models, Molecular
Mucins/*biosynthesis
N-Acetylgalactosaminyltransferases/*chemistry/genetics/metabolism
Protein Conformation
Abstract: UDP-GalNAc:polypeptide alpha-N-acetylgalactosaminyltransferases (ppGaNTases) initiate the formation of mucin-type, O-linked glycans by catalyzing the transfer of alpha-N-acetylgalactosamine from UDP-GalNAc to Ser or Thr residues of core proteins to form the Tn antigen (GalNAc-alpha-1-O-Ser/Thr). ppGaNTases are unique among glycosyltransferases in containing a C-terminal lectin domain. We present the x-ray crystal structure of a ppGaNTase, murine ppGaNTase-T1, and show that it folds to form distinct catalytic and lectin domains. The association of the two domains forms a large cleft in the surface of the enzyme that contains a Mn2+ ion complexed by invariant D209 and H211 of the "DXH" motif and by invariant H344. Each of the three potential lectin domain carbohydrate-binding sites (alpha, beta, and gamma) is located on the active-site face of the enzyme, suggesting a mechanism by which the transferase may accommodate multiple conformations of glycosylated acceptor substrates. A model of a mucin 1 glycopeptide substrate bound to the enzyme shows that the spatial separation between the lectin alpha site and a modeled active site UDP-GalNAc is consistent with the in vitro pattern of glycosylation observed for this peptide catalyzed by ppGaNTase-T1. The structure also provides a template for the larger ppGaNTase family, and homology models of several ppGaNTase isoforms predict dramatically different surface chemistries consistent with isoform-selective acceptor substrate recognition.
Notes: 1091-6490
Fritz, Timothy A
Hurley, James H
Trinh, Loc-Ba
Shiloach, Joseph
Tabak, Lawrence A
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2004 Oct 26;101(43):15307-12. doi: 10.1073/pnas.0405657101. Epub 2004 Oct 14.
Author Address: Section on Biological Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Department of Health and Human Services, Bethesda, MD 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 244
Author: Fu, D. and van Halbeek, H.
Year: 1992
Title: N-glycosylation site mapping of human serotransferrin by serial lectin affinity chromatography, fast atom bombardment-mass spectrometry, and 1H nuclear magnetic resonance spectroscopy
Journal: Anal Biochem
Volume: 206
Issue: 1
Pages: 53-63
Epub Date: 1992/10/01
Date: Oct
Short Title: N-glycosylation site mapping of human serotransferrin by serial lectin affinity chromatography, fast atom bombardment-mass spectrometry, and 1H nuclear magnetic resonance spectroscopy
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/s0003-2697(05)80010-7
Accession Number: 1456441
Keywords: Amino Acid Sequence
Binding Sites
Carbohydrate Sequence
Chromatography, Affinity/*methods
Glycopeptides/chemistry/isolation & purification
Glycosylation
Humans
*Lectins
Magnetic Resonance Spectroscopy/methods
Molecular Sequence Data
Oligosaccharides/chemistry/isolation & purification
Peptide Mapping
Spectrometry, Mass, Fast Atom Bombardment
Transferrin/*chemistry/isolation & purification/metabolism
Trypsin/metabolism
Abstract: This report describes the N-glycosylation site mapping of human serotransferrin (h-STF). Reduced and S-carboxymethylated h-STF was digested with trypsin or chymotrypsin. Glycopeptides in the proteolytic digests were isolated by serial concanavalin A (Con A), Sambucus nigra agglutinin (SNA), and Phaseolus vulgaris leukoagglutinin (LPHA) affinity chromatography and subjected to preliminary analysis by 1H NMR spectroscopy. The glycopeptide fractions were then individually digested with N-glycanase. One part of the digest of each fraction was analyzed by fast atom bombardment-mass spectrometry (FAB-MS) to identify the peptide sequences of the glycosylation sites. The other part was used to isolate the oligosaccharide by the corresponding lectin affinity chromatography and to characterize the structures of the isolated oligosaccharides by 1H NMR spectroscopy and FAB-MS. The oligosaccharides in the Con A-bound fraction were shown to have bi-alpha(2-->6)-sialyl, diantennary structures. The SNA-bound fraction was shown to contain trisialyl, triantennary structures. Di- and triantennary oligosaccharides were found to occur on each of the two N-glycosylation sites of h-STF (Asn413 and Asn611) in the ratio of approximately 85:15. The SNA-bound glycopeptides were further fractionated by LPHA affinity chromatography. Two different oligosaccharides were characterized, namely, a trisialyl 2,4-triantennary and a trisialyl 2,6-triantennary glycan. The ratio of 2,4-triantennary vs 2,6-triantennary oligosaccharides attached to glycosylation site Asn413 was found to be approximately 5:1, whereas the two isomeric triantennary oligosaccharides were found to be attached to glycosylation site Asn611 in the ratio approximately 1:1.
Notes: Fu, D
van Halbeek, H
P01-AI-27135/AI/NIAID NIH HHS/United States
P41-RR-05351/RR/NCRR NIH HHS/United States
S10-RR-04720/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Anal Biochem. 1992 Oct;206(1):53-63. doi: 10.1016/s0003-2697(05)80010-7.
Author Address: Complex Carbohydrate Research Center, University of Georgia, Athens 30602-4712.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1626
Author: Fu, H. L., Valiathan, R. R., Payne, L., Kumarasiri, M., Mahasenan, K. V., Mobashery, S., Huang, P. and Fridman, R.
Year: 2014
Title: Glycosylation at Asn211 regulates the activation state of the discoidin domain receptor 1 (DDR1)
Journal: J Biol Chem
Volume: 289
Issue: 13
Pages: 9275-87
Epub Date: 2014/02/11
Date: Mar 28
Short Title: Glycosylation at Asn211 regulates the activation state of the discoidin domain receptor 1 (DDR1)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.541102
PMCID: PMC3979393
Accession Number: 24509848
Keywords: Amino Acid Motifs
Amino Acid Sequence
Animals
*Asparagine
Cell Line
Collagen Type I/pharmacology
Conserved Sequence
Discoidin Domain Receptor 1
Endocytosis/drug effects
Glycosylation
Humans
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutation
Protein Multimerization
Protein Structure, Quaternary
Receptor Protein-Tyrosine Kinases/*chemistry/genetics/*metabolism
Collagen
Mutagenesis Site-specific
Receptor Regulation
Receptor Tyrosine Kinase
Abstract: Discoidin domain receptor 1 (DDR1) belongs to a unique family of receptor tyrosine kinases that signal in response to collagens. DDR1 undergoes autophosphorylation in response to collagen binding with a slow and sustained kinetics that is unique among members of the receptor tyrosine kinase family. DDR1 dimerization precedes receptor activation suggesting a structural inhibitory mechanism to prevent unwarranted phosphorylation. However, the mechanism(s) that maintains the autoinhibitory state of the DDR1 dimers is unknown. Here, we report that N-glycosylation at the Asn(211) residue plays a unique role in the control of DDR1 dimerization and autophosphorylation. Using site-directed mutagenesis, we found that mutations that disrupt the conserved (211)NDS N-glycosylation motif, but not other N-glycosylation sites (Asn(260), Asn(371), and Asn(394)), result in collagen I-independent constitutive phosphorylation. Mass spectrometry revealed that the N211Q mutant undergoes phosphorylation at Tyr(484), Tyr(520), Tyr(792), and Tyr(797). The N211Q traffics to the cell surface, and its ectodomain displays collagen I binding with an affinity similar to that of the wild-type DDR1 ectodomain. However, unlike the wild-type receptor, the N211Q mutant exhibits enhanced receptor dimerization and sustained activation upon ligand withdrawal. Taken together, these data suggest that N-glycosylation at the highly conserved (211)NDS motif evolved to act as a negative repressor of DDR1 phosphorylation in the absence of ligand. The presence of glycan moieties at that site may help to lock the collagen-binding domain in the inactive state and prevent unwarranted signaling by receptor dimers. These studies provide a novel insight into the structural mechanisms that regulate DDR activation.
Notes: 1083-351x
Fu, Hsueh-Liang
Valiathan, Rajeshwari R
Payne, Leo
Kumarasiri, Malika
Mahasenan, Kiran V
Mobashery, Shahriar
Huang, Paul
Fridman, Rafael
089028/Wellcome Trust/United Kingdom
R01 CA061986/CA/NCI NIH HHS/United States
BB/I014276/1/Biotechnology and Biological Sciences Research Council/United Kingdom
CA-61986/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2014 Mar 28;289(13):9275-87. doi: 10.1074/jbc.M113.541102. Epub 2014 Feb 7.
Author Address: From the Department of Pathology, Wayne State University, Detroit, Michigan 48201.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 319
Author: Fujii, S., Nishiura, T., Nishikawa, A., Miura, R. and Taniguchi, N.
Year: 1990
Title: Structural heterogeneity of sugar chains in immunoglobulin G. Conformation of immunoglobulin G molecule and substrate specificities of glycosyltransferases
Journal: J Biol Chem
Volume: 265
Issue: 11
Pages: 6009-18
Epub Date: 1990/04/15
Date: Apr 15
Short Title: Structural heterogeneity of sugar chains in immunoglobulin G. Conformation of immunoglobulin G molecule and substrate specificities of glycosyltransferases
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2138613
Keywords: Animals
Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Cattle
Chromatography, High Pressure Liquid
Galactosyltransferases/*metabolism
Glucosyltransferases/*metabolism
Glycosylation
Humans
*Immunoglobulin G/*genetics/isolation & purification/metabolism
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Multiple Myeloma/immunology
*N-Acetylglucosaminyltransferases
Oligosaccharides/*biosynthesis/isolation & purification
Protein Processing, Post-Translational
Reference Values
Substrate Specificity
Abstract: The heterogeneous asparagine-linked sugar chains of bovine and human immunoglobulins G were separated into 12 components by reversed-phase high performance liquid chromatography, and their structures were determined by 1H NMR spectroscopy. Both immunoglobulin (Ig) G sources contained eight non-bisected biantennary complexes and four bisected biantennary complexes. In the non-bisected sugar chains of bovine IgG, galactosylation of the Man alpha 1-3 branch predominated over that of the Man alpha 1-6, whereas in the bisected complexes galactosylation of the Man alpha 1-6 branch predominated. This difference can be explained by the substrate specificities of the galactosyl-transferases and of the N-acetylglucosaminyltransferase III involved in their synthesis. The sugar chains of human IgG1 differs in the distribution of its galactose residues from bovine IgG and human IgG2. The Man alpha 1-6 branch of all IgG1s was more highly galactosylated than the Man alpha 1-3 branch even in the non-bisected complexes. Such findings are in conflict with the substrate specificities of galactosyltransferases. Whereas these enzymes derivatized more of the Man alpha 1-6 branch of native human IgG1, in denatured protein more of the Man alpha 1-3 branch was galactosylated. Thus, protein conformation may influence the structure of its sugar chains.
Notes: Fujii, S
Nishiura, T
Nishikawa, A
Miura, R
Taniguchi, N
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1990 Apr 15;265(11):6009-18.
Author Address: Department of Biochemistry, Osaka University Medical School, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1442
Author: Fujiki, R., Hashiba, W., Sekine, H., Yokoyama, A., Chikanishi, T., Ito, S., Imai, Y., Kim, J., He, H. H., Igarashi, K., Kanno, J., Ohtake, F., Kitagawa, H., Roeder, R. G., Brown, M. and Kato, S.
Year: 2011
Title: GlcNAcylation of histone H2B facilitates its monoubiquitination
Journal: Nature
Volume: 480
Issue: 7378
Pages: 557-60
Epub Date: 2011/11/29
Date: Nov 27
Short Title: GlcNAcylation of histone H2B facilitates its monoubiquitination
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature10656
PMCID: PMC7289526
NIHMSID: NIHMS1591273
Accession Number: 22121020
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Cell Line
HeLa Cells
Histones/chemistry/genetics/*metabolism
Humans
Models, Molecular
Mutation
Protein Structure, Tertiary
Recombinant Proteins/chemistry/metabolism
Ubiquitination
Abstract: Chromatin reorganization is governed by multiple post-translational modifications of chromosomal proteins and DNA. These histone modifications are reversible, dynamic events that can regulate DNA-driven cellular processes. However, the molecular mechanisms that coordinate histone modification patterns remain largely unknown. In metazoans, reversible protein modification by O-linked N-acetylglucosamine (GlcNAc) is catalysed by two enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). However, the significance of GlcNAcylation in chromatin reorganization remains elusive. Here we report that histone H2B is GlcNAcylated at residue S112 by OGT in vitro and in living cells. Histone GlcNAcylation fluctuated in response to extracellular glucose through the hexosamine biosynthesis pathway (HBP). H2B S112 GlcNAcylation promotes K120 monoubiquitination, in which the GlcNAc moiety can serve as an anchor for a histone H2B ubiquitin ligase. H2B S112 GlcNAc was localized to euchromatic areas on fly polytene chromosomes. In a genome-wide analysis, H2B S112 GlcNAcylation sites were observed widely distributed over chromosomes including transcribed gene loci, with some sites co-localizing with H2B K120 monoubiquitination. These findings suggest that H2B S112 GlcNAcylation is a histone modification that facilitates H2BK120 monoubiquitination, presumably for transcriptional activation.
Notes: 1476-4687
Fujiki, Ryoji
Hashiba, Waka
Sekine, Hiroki
Yokoyama, Atsushi
Chikanishi, Toshihiro
Ito, Saya
Imai, Yuuki
Kim, Jaehoon
He, Housheng Hansen
Igarashi, Katsuhide
Kanno, Jun
Ohtake, Fumiaki
Kitagawa, Hirochika
Roeder, Robert G
Brown, Myles
Kato, Shigeaki
R01 CA129325/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Nature. 2011 Nov 27;480(7378):557-60. doi: 10.1038/nature10656.
Author Address: Institute of Molecular and Cellular Biosciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 617
Author: Fujinaga, M., Chernaia, M. M., Halenbeck, R., Koths, K. and James, M. N.
Year: 1996
Title: The crystal structure of PR3, a neutrophil serine proteinase antigen of Wegener's granulomatosis antibodies
Journal: J Mol Biol
Volume: 261
Issue: 2
Pages: 267-78
Epub Date: 1996/08/16
Date: Aug 16
Short Title: The crystal structure of PR3, a neutrophil serine proteinase antigen of Wegener's granulomatosis antibodies
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1006/jmbi.1996.0458
Accession Number: 8757293
Keywords: Amino Acid Sequence
Autoantigens/*chemistry
Binding Sites
Carbohydrate Sequence
Crystallography, X-Ray
Glycosylation
Granulomatosis with Polyangiitis/*immunology
Humans
Leukocyte Elastase
Models, Molecular
Molecular Sequence Data
Myeloblastin
Neutrophils/*immunology
Pancreatic Elastase/chemistry
Protein Conformation
Recombinant Proteins/chemistry
Sequence Alignment
Serine Endopeptidases/*chemistry
Substrate Specificity
Abstract: The crystal structure of PR3, a serine proteinase from the azurophilic granules of human polymorphonuclear neutrophils, has been solved by molecular replacement using the human leukocyte elastase structure. The PR3 structure has been refined to an R-factor (= sigma parallel Fo magnitude of-Fc parallel/sigma magnitude of Fo) of 0.201 for all data in the range of 10.0 to 2.2 A resolution. The enzyme was crystallized in space group P21 with four molecules in the asymmetric unit (Vm approximately equal to 2.6 A/Da). The overall fold consists of two domains of beta-barrel structures typical of the chymotrypsin family of serine proteinases. In general, the substrate binding sites, S4 to S3', are more polar than comparable sites in the related proteinase, human leukocyte elastase. The experimentally observed preference of PR3 for small aliphatic residues at the P1 position of a substrate is explained by the Val to Ile substitution at position 190 when compared to the elastase structure. The substitution of Ala by Asp at position 213 at the back of S1 should not affect its specificity greatly, as the Asp side-chain points back into the interior of the protein. The PR3 structure includes a disaccharide unit (N-linked 2-acetamido-2-deoxy-beta-D-glucopyranose and 1,6-linked alpha-L-fucopyranose) covalently attached to Asn 159. The linear antigenic sites of PR3 reported to react with Wegener's granulomatosis autoantibodies occur in regions of the three-dimensional structure that may implicate the inactive pro-form of the enzyme in the pathogenesis of the disease.
Notes: Fujinaga, M
Chernaia, M M
Halenbeck, R
Koths, K
James, M N
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 1996 Aug 16;261(2):267-78. doi: 10.1006/jmbi.1996.0458.
Author Address: Department of Biochemistry, University of Alberta, Edmonton, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 542
Author: Fujita, S., Endo, T., Ju, J., Kean, E. L. and Kobata, A.
Year: 1994
Title: Structural studies of the N-linked sugar chains of human rhodopsin
Journal: Glycobiology
Volume: 4
Issue: 5
Pages: 633-40
Epub Date: 1994/10/01
Date: Oct
Short Title: Structural studies of the N-linked sugar chains of human rhodopsin
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/4.5.633
Accession Number: 7881178
Keywords: Amino Acids/analysis
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Electrophoresis, Polyacrylamide Gel
Humans
Molecular Sequence Data
Oligosaccharides/chemistry/isolation & purification
Rhodopsin/*chemistry
Abstract: Human rhodopsin is a glycoprotein containing two N-linked sugar chains. After the isolation and purification of rhodopsins from human retinas, structural studies of their N-linked sugar chains were performed. The sugar moieties, quantitatively released as oligosaccharides from the polypeptide backbone by hydrazinolysis, were converted to radioactive oligosaccharides by reduction with NaB3H4 after N-acetylation. As indicated by high-voltage paper electrophoresis, > 96% of the sugar chains were free of sialic acid and the remaining were sialylated derivatives. Structural studies of each oligosaccharide by lectin affinity column chromatography, and sequential exoglycosidase digestion in combination with methylation analysis, revealed that almost all of the oligosaccharides were hybrid-type sugar chains. While the major oligosaccharide species of bovine and human rhodopsin are identical, in contrast to the sugar chains of bovine rhodopsin, human rhodopsin also contains sialylated isomers and a high concentration of a galactosylated isomer. These results suggest that species-specific processing of the sugar chains of rhodopsin occurs.
Notes: Fujita, S
Endo, T
Ju, J
Kean, E L
Kobata, A
EY 00393/EY/NEI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1994 Oct;4(5):633-40. doi: 10.1093/glycob/4.5.633.
Author Address: Department of Biochemistry, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1055
Author: Fujita, T., Utsunomiya, I., Ren, J., Matsushita, Y., Kawai, M., Sasaki, S., Hoshi, K., Miyatake, T. and Taguchi, K.
Year: 2006
Title: Glycosylation and cell surface expression of Kv1.2 potassium channel are regulated by determinants in the pore region
Journal: Neurochem Res
Volume: 31
Issue: 5
Pages: 589-96
Epub Date: 2006/06/14
Date: May
Short Title: Glycosylation and cell surface expression of Kv1.2 potassium channel are regulated by determinants in the pore region
Alternate Journal: Neurochemical research
ISSN: 0364-3190 (Print)
0364-3190
DOI: 10.1007/s11064-006-9056-4
Accession Number: 16770729
Keywords: Amino Acid Sequence
Animals
Antineoplastic Combined Chemotherapy Protocols
Cells
Cricetinae
Cricetulus
Cyclophosphamide
Doxorubicin
Glycosylation
Kv1.2 Potassium Channel/*chemistry/genetics/*metabolism
Molecular Sequence Data
Mutagenesis, Site-Directed
Patch-Clamp Techniques
Potassium/metabolism
*Protein Structure, Tertiary
Protein Subunits/chemistry/genetics/metabolism
Rats
Vincristine
Abstract: Voltage-gated K(+) channels contain six membrane spanning segments and a pore-forming domain. We used site-directed mutation to examine the role of specific amino acids in the extracellular region of the pore in Kv1.2. When expressed in CHO cells, a K(+) current was not observed for mutants S356A, S360A, T383A and T384A. However, coexpression of the Kvbeta2 subunit and the S360A mutant resulted in a robust peak current. Immunocytochemistry for Kv1.2 showed staining throughout the cytoplasm in cells coexpressing the beta2 and S360A, whereas only the perinuclear region was stained in cells expressing the S360A mutant. Western blotting revealed that the major immunoreactive protein in wild-type- and mutant-expressing cells is 60-kDa, but 87-kDa bands were also detected in cells expressing wild-type Kv1.2 and cells coexpressing beta2and S360A. These results suggest that amino acids in the pore region help regulate ion permeability or cellular trafficking by affecting glycosylation of Kv1.2.
Notes: Fujita, Tetsuhiro
Utsunomiya, Iku
Ren, Jin
Matsushita, Yousuke
Kawai, Miwa
Sasaki, Sachie
Hoshi, Keiko
Miyatake, Tadashi
Taguchi, Kyoji
Journal Article
Research Support, Non-U.S. Gov't
United States
Neurochem Res. 2006 May;31(5):589-96. doi: 10.1007/s11064-006-9056-4. Epub 2006 May 23.
Author Address: Department of Pharmacotherapeutics, Showa Pharmaceutical University, Tokyo, Machida, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 574
Author: Fujiwara, S., Shinkai, H., Mann, K. and Timpl, R.
Year: 1993
Title: Structure and localization of O- and N-linked oligosaccharide chains on basement membrane protein nidogen
Journal: Matrix
Volume: 13
Issue: 3
Pages: 215-22
Epub Date: 1993/05/01
Date: May
Short Title: Structure and localization of O- and N-linked oligosaccharide chains on basement membrane protein nidogen
Alternate Journal: Matrix (Stuttgart, Germany)
ISSN: 0934-8832 (Print)
0934-8832
DOI: 10.1016/s0934-8832(11)80005-3
Accession Number: 8326911
Keywords: Amino Acid Sequence
Animals
Basement Membrane/*chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Membrane Glycoproteins/*chemistry
Mice
Molecular Sequence Data
Neoplasm Proteins/chemistry
Neoplasms, Experimental/chemistry
Oligosaccharides/*chemistry
Abstract: The carbohydrate content of mouse nidogen predicts the occupation of two N- and about seven O-linked acceptor sites. The corresponding oligosaccharides were examined by sequential exoglycosidase digestions. The data indicate N-linked substitutions by several bi-, tri- and tetraantennary complex types of oligosaccharides which are further modified by additional lactosamines and terminal alpha-galactose and/or sialic acid. Mannose-rich oligosaccharides were of low abundance. O-linked structures included a di- and tetrasaccharide core structure that were in addition sialylated and may be similar to structures found in fetuin. Evidence is provided that the two sequence-predicted asparagine acceptors are almost fully substituted. Sequence analysis of tryptic peptides identified Thr-271, Ser-303, Thr-309, Thr-317, Thr-320, Thr-892 and Thr-905 as the most likely sites for galactosamine substitutions. These residues are located in the flexible link connecting the N-terminal globular domains G1 and G2 of nidogen and at the border between the rod and the C-terminal globe G3. Four of them showed Pro in the -1 or +3 position. All these Ser, Thr and Pro residues but not the N-linked attachment sites are identical in human nidogen.
Notes: Fujiwara, S
Shinkai, H
Mann, K
Timpl, R
Journal Article
Research Support, Non-U.S. Gov't
Germany
Matrix. 1993 May;13(3):215-22. doi: 10.1016/s0934-8832(11)80005-3.
Author Address: Department of Dermatology, Oita, Medical University, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 414
Author: Fukuda, M., Lauffenburger, M., Sasaki, H., Rogers, M. E. and Dell, A.
Year: 1987
Title: Structures of novel sialylated O-linked oligosaccharides isolated from human erythrocyte glycophorins
Journal: J Biol Chem
Volume: 262
Issue: 25
Pages: 11952-7
Epub Date: 1987/09/05
Date: Sep 5
Short Title: Structures of novel sialylated O-linked oligosaccharides isolated from human erythrocyte glycophorins
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3624241
Keywords: Carbohydrate Conformation
Chromatography, Gel
Chromatography, High Pressure Liquid
Glycophorins/*analysis
Humans
Mass Spectrometry
Methylation
Oligosaccharides/*analysis
Sialic Acids/analysis
Sialoglycoproteins/*analysis
Abstract: The O-linked oligosaccharides attached to human erythrocyte glycophorins were extensively characterized. In addition to the previously described disialylated tetrasaccharide, NeuNAc alpha 2----3Gal beta 1----3 (Neu-NAc alpha 2----6)GalNAcOH and monosialylated trisaccharide, NeuNAc alpha 2----3Gal beta 1----3GalNAcOH, novel trisialylated oligosaccharides were isolated. Methylation analysis, fast atom bombardment-mass spectrometry, and enzymatic degradation were used to elucidate the following novel structures: formula; see text: These results suggest that O-linked oligosaccharides with a disialosyl group, NeuNAc alpha 2----8NeuNAc alpha 2----, may be present in various tissues.
Notes: Fukuda, M
Lauffenburger, M
Sasaki, H
Rogers, M E
Dell, A
R01 CA33000/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1987 Sep 5;262(25):11952-7.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 572
Author: Fukushima, K., Watanabe, H., Takeo, K., Nomura, M., Asahi, T. and Yamashita, K.
Year: 1993
Title: N-linked sugar chain structure of recombinant human lymphotoxin produced by CHO cells: the functional role of carbohydrate as to its lectin-like character and clearance velocity
Journal: Arch Biochem Biophys
Volume: 304
Issue: 1
Pages: 144-53
Epub Date: 1993/07/01
Date: Jul
Short Title: N-linked sugar chain structure of recombinant human lymphotoxin produced by CHO cells: the functional role of carbohydrate as to its lectin-like character and clearance velocity
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1993.1332
Accession Number: 8323280
Keywords: Amino Acid Sequence
Animals
CHO Cells
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Cricetinae
Glycoproteins/*chemistry
Humans
Lymphotoxin-alpha/*chemistry/pharmacokinetics
Metabolic Clearance Rate
Methylation
Molecular Sequence Data
Molecular Structure
Mucoproteins/metabolism
Oligosaccharides/chemistry
Protein Binding
Rats
Recombinant Proteins/chemistry/pharmacokinetics
Structure-Activity Relationship
Uromodulin
Abstract: Recombinant human lymphotoxin (rhLT) produced by CHO cells transfected with human LT genomic DNA was purified to homogeneity, but approximately 5% of the molecules were devoid of the last two amino terminal residues. A peptide N-glycosylated at Asn62 (Tr-45) and one partially O-glycosylated at Thr7 (Tr-14) on cleavage with trypsin were separated by reverse phase HPLC. The N-linked sugar chains of Tr-45 were released quantitatively as oligosaccharides on hydrazinolysis (100 degrees C, 8 h), followed by N-acetylation. After being reduced with either NaB3H4 or NaB2H4, their structures were determined by a combination of serial lectin affinity chromatography, exoglycosidase digestion, and methylation analysis: 82.7% of the sugar chains occur as biantennary complex-type sugar chains, the remainder being C-2 and C-2,4/C-2,6 branched triantennary, and C-2,4 and C-2,6 branched tetraantennary complex-type sugar chains with a fucosylated mannose core. Their sialic acid residues occur only as the Neu5Ac alpha 2-->3Gal group. The clearance velocity from the bloodstream dramatically increased with desialylation, and rhLT tends to have accumulated in the kidney, indicating that there may exist other mechanisms for clearance from the circulation besides the galactose-binding protein in hepatocytes and the filtration system of the kidney. Desialylated rhLT showed a lectin-like binding character to uromodulin similar to that of tumor necrosis factor, although intact rhLT did not. The interaction between desialylated rhLT and uromodulin was inhibited by N,N'-diacetylchitobiose and [Man alpha 1-->6(Man alpha 1-->3)Man alpha 1-->6](Man alpha 1-->2Man alpha 1-->3)Man beta 1-->4GlcNAc beta 1-->4GlcNAc-->Asn. These results indicate that the lectin-like domain of rhLT is exposed on its desialylation.
Notes: Fukushima, K
Watanabe, H
Takeo, K
Nomura, M
Asahi, T
Yamashita, K
Journal Article
Research Support, Non-U.S. Gov't
United States
Arch Biochem Biophys. 1993 Jul;304(1):144-53. doi: 10.1006/abbi.1993.1332.
Author Address: Department of Biochemistry, Sasaki Institute, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 744
Author: Fukuta, K., Abe, R., Yokomatsu, T., Kono, N., Asanagi, M., Omae, F., Minowa, M. T., Takeuchi, M. and Makino, T.
Year: 2000
Title: Remodeling of sugar chain structures of human interferon-gamma
Journal: Glycobiology
Volume: 10
Issue: 4
Pages: 421-30
Epub Date: 2000/04/15
Date: Apr
Short Title: Remodeling of sugar chain structures of human interferon-gamma
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/10.4.421
Accession Number: 10764830
Keywords: Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/*chemistry
Chromatography, High Pressure Liquid
Cricetinae
Flow Cytometry
Humans
Interferon-gamma/*chemistry/genetics
Molecular Sequence Data
N-Acetylglucosaminyltransferases/genetics/metabolism
N-Acetylneuraminic Acid/metabolism
Recombinant Proteins/metabolism
Transfection
Abstract: Natural human interferon (IFN)-gamma has mainly biantennary complex-type sugar chains and scarcely has multiantennary structures. We attempted to remodel the sugar chain structures using IFN-gamma as a model glycoprotein. To obtain the branching glycoforms of IFN-gamma, we introduced the genes for GnT-IV (UDP-N-acetylglucosamine:alpha-1,3-D-mannoside beta-1, 4-N-acetylglucosaminyltransferase) and/or GnT-V (UDP-N-acetylglucosamine:alpha-1,6-D-mannoside beta-1, 6-N-acetylglucosaminyltransferase) into Chinese hamster ovary (CHO) cells producing human IFN-gamma. The parental CHO cells produced IFN-gamma with biantennary sugar chains mainly. When the GnT-IV activity was increased, triantennary sugar chains with a branch produced by GnT-IV increased up to 66.9% of the total sugar chains. When the GnT-V activity was increased, triantennary sugar chains with a corresponding branch increased up to 55.7% of the total sugar chains. When the GnT-IV and -V activities were increased at a time, tetraantennary sugar chains increased up to 56.2% of the total sugar chains. The proportion of these multiantennary sugar chains corresponded to the intracellular activities of GnT-IV and -V. What is more, lectin blot and flow cytometric analysis indicated that the multi-branch structure of the sugar chains was increased not only on IFN-gamma, one of the secretory glycoproteins, but also on almost CHO cellular proteins by introducing either or both of the GnT genes. The results suggest that the branching structure of sugar chains of glycoproteins could be controlled by cellular GnT-IV and GnT-V activities. This technology can produce glycoforms out of natural occurrence, which should enlarge the potency of glycoprotein therapeutics.
Notes: Fukuta, K
Abe, R
Yokomatsu, T
Kono, N
Asanagi, M
Omae, F
Minowa, M T
Takeuchi, M
Makino, T
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2000 Apr;10(4):421-30. doi: 10.1093/glycob/10.4.421.
Author Address: Life Science Laboratory, Mitsui Chemicals Inc., 1144 Togo, Mobara, Chiba 297-0017, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 756
Author: Fukuta, K., Abe, R., Yokomatsu, T., Kono, N., Nagatomi, Y., Asanagi, M., Shimazaki, Y. and Makino, T.
Year: 2000
Title: Comparative study of the N-glycans of human monoclonal immunoglobulins M produced by hybridoma and parental cells
Journal: Arch Biochem Biophys
Volume: 378
Issue: 1
Pages: 142-50
Epub Date: 2000/06/28
Date: Jun 1
Short Title: Comparative study of the N-glycans of human monoclonal immunoglobulins M produced by hybridoma and parental cells
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.2000.1806
Accession Number: 10871054
Keywords: Amino Acid Sequence
Antibodies, Monoclonal/biosynthesis/*chemistry/genetics
Base Sequence
Carbohydrate Sequence
Carbohydrates/chemistry
Cell Line, Transformed
Cloning, Molecular
DNA Primers/genetics
Galactosyltransferases/metabolism
Humans
Hybridomas/enzymology/*immunology
Immunoglobulin M/biosynthesis/*chemistry/genetics
Immunoglobulin Variable Region/biosynthesis/chemistry/genetics
Immunoglobulin mu-Chains/biosynthesis/chemistry/genetics
Molecular Sequence Data
N-Acetylglucosaminyltransferases/metabolism
Polysaccharides/*chemistry
Sialic Acids/analysis
Tumor Cells, Cultured
Abstract: Cell-cell hybridization is one method of establishing cell lines capable of producing an abundance of antibodies. In order to clearly characterize antibodies produced by hybridomas, the influence of cell-cell hybridization on the glycosylation of produced antibodies should be studied. In this report, we describe structural changes of the N-glycans in immunoglobulin M (IgM) produced by a hybridoma cell line termed 3-4, which was established through hybridization of an IgM-producing Epstein-Barr virus transformed human B-cell line termed No. 12, and a human myeloma cell line termed P109. We analyzed the structures of sugar chains on the constant region of the mu-chain of IgMs produced by parental No. 12 cells and hybridoma 3-4 cells. In both parental cells and hybridoma cells, the predominant structures at Asn171, Asn332, and N395 were fully galactosylated biantennary complex types, with or without core fucose and/or bisecting GlcNAc. However, the amount of bisecting GlcNAc was markedly decreased in the hybridoma cells. Therefore, the activity of UDP-N-acetylglucosamine:beta-D-mannoside beta-1,4-N-acetylglucosaminyltransferase (GnT-III) responsible for the formation of bisecting GlcNAc was measured in parental cells and hybridoma cells. No. 12 cells showed some GnT-III activity, whereas P109 cells showed no such activity. The corresponding level of activity observed in hybridoma 3-4 cells was much lower than that in No. 12 cells. The above results demonstrated a reduction in the intracellular activity of GnT-III in the hybridoma cells, which was largely due to the influence of P109 cells. Moreover, the sugar chain structures of IgMs produced by the cells reflected the level of GnT-III activity.
Notes: Fukuta, K
Abe, R
Yokomatsu, T
Kono, N
Nagatomi, Y
Asanagi, M
Shimazaki, Y
Makino, T
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Arch Biochem Biophys. 2000 Jun 1;378(1):142-50. doi: 10.1006/abbi.2000.1806.
Author Address: Life Science Laboratory, Mitsui Chemicals, Inc., Mobara, Chiba, Japan. Kazuhiro.Fukuta@mitsui.chem.co.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 807
Author: Fukuta, K., Abe, R., Yokomatsu, T., Minowa, M. T., Takeuchi, M., Asanagi, M. and Makino, T.
Year: 2001
Title: The widespread effect of beta 1,4-galactosyltransferase on N-glycan processing
Journal: Arch Biochem Biophys
Volume: 392
Issue: 1
Pages: 79-86
Epub Date: 2001/07/27
Date: Aug 1
Short Title: The widespread effect of beta 1,4-galactosyltransferase on N-glycan processing
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.2001.2421
Accession Number: 11469797
Keywords: Animals
CHO Cells
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Cricetinae
Humans
Interferon-gamma/biosynthesis/chemistry/genetics
Mannose/chemistry
Mannosidases/metabolism
Molecular Sequence Data
N-Acetylglucosaminyltransferases/metabolism
N-Acetyllactosamine Synthase/genetics/*metabolism
Polysaccharides/chemistry/*metabolism
Recombinant Proteins/genetics/metabolism
alpha-Mannosidase
Abstract: We investigated beta 1,4-GalT (UDP-galactose: beta-d-N-acetylglucosaminide beta 1,4-galactosyltransferase) in terms of intracellular competition with GnT-IV (UDP-N-acetylglucosamine: alpha1,3-d-mannoside beta1,4-N-acetylglucosaminyltransferase) and GnT-V (UDP-N-acetylglucosamine: alpha1,6-d-mannoside beta 1,6-N-acetylglucosaminyltransferase). The beta 1,4-GalT-I gene was introduced into Chinese hamster ovary (CHO) cells producing human interferon (hIFN)-gamma (IM4/V/IV cells) and five clones expressing various levels of beta 1,4-GalT were isolated. As we previously reported, parental IM4/V/IV cells express high levels of GnT-IVa and -V and produce hIFN-gamma having primarily tetraantennary sugar chains. The branching of sugar chains on hIFN-gamma was suppressed in the beta 1,4-GalT-enhanced clones to a level corresponding to the intracellular activity of beta 1,4-GalT relative to GnTs. Moreover, the contents of hybrid-type and high-mannose-type sugar chains increased in these clones. The results showed that beta 1,4-GalT widely affects N-glycan processing by competing with GnT-IV, GnT-V, and alpha-mannosidase II in cells and also by some other mechanisms that suppress the conversion of high-mannose-type sugar chains to the hybrid type.
Notes: Fukuta, K
Abe, R
Yokomatsu, T
Minowa, M T
Takeuchi, M
Asanagi, M
Makino, T
Journal Article
Research Support, Non-U.S. Gov't
United States
Arch Biochem Biophys. 2001 Aug 1;392(1):79-86. doi: 10.1006/abbi.2001.2421.
Author Address: Life Science Laboratory, Mitsui Chemicals, Incorporated, 1144 Togo, Mobara, Chiba 297-0017, Japan. kazuhiro.fukuta@mitsui-chem.co.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 750
Author: Fukuta, K., Abe, R., Yokomatsu, T., Omae, F., Asanagi, M. and Makino, T.
Year: 2000
Title: Control of bisecting GlcNAc addition to N-linked sugar chains
Journal: J Biol Chem
Volume: 275
Issue: 31
Pages: 23456-61
Epub Date: 2000/05/19
Date: Aug 4
Short Title: Control of bisecting GlcNAc addition to N-linked sugar chains
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M002693200
Accession Number: 10816579
Keywords: Acetylglucosamine/*metabolism
Antibodies, Monoclonal/biosynthesis
B-Lymphocytes/*metabolism
Carbohydrate Sequence
Clone Cells
Glycosylation
Humans
Immunoglobulin M/*biosynthesis/*chemistry
Molecular Sequence Data
Mutation
N-Acetylglucosaminyltransferases/genetics/*metabolism
N-Acetyllactosamine Synthase/genetics/*metabolism
Protein Processing, Post-Translational
Transfection
Abstract: In the present study, experimental control of the formation of bisecting GlcNAc was investigated, and the competition between beta-1,4-GalT (UDP-galactose:N-acetylglucosamine beta-1, 4-galactosyltransferase) and GnT-III (UDP-N-acetylglucosamine:beta-d-mannoside beta-1, 4-N-acetylglucosaminyltransferase) was examined. We isolated a beta-1,4-GalT-I single knockout human B cell clone producing monoclonal IgM and several transfectant clones that overexpressed beta-1,4-GalT-I or GnT-III. In the beta-1,4-GalT-I-single knockout cells, the extent of bisecting GlcNAc addition to the sugar chains of IgM was increased, where beta-1,4-GalT activity was reduced to about half that in the parental cells, and GnT-III activity was unaltered. In the beta-1,4-GalT-I transfectants, the extent of bisecting GlcNAc addition was reduced although GnT-III activity was not altered significantly. In the GnT-III transfectants, the extent of bisecting GlcNAc addition increased along with the increase in levels of GnT-III activity. The extent of bisecting GlcNAc addition to the sugar chains of IgM was significantly correlated with the level of intracellular beta-1,4-GalT activity relative to that of GnT-III. These results were interpreted as indicating that beta-1, 4-GalT competes with GnT-III for substrate in the cells.
Notes: Fukuta, K
Abe, R
Yokomatsu, T
Omae, F
Asanagi, M
Makino, T
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2000 Aug 4;275(31):23456-61. doi: 10.1074/jbc.M002693200.
Author Address: Life Science Laboratory, Mitsui Chemicals, Inc., 1144 Togo, Mobara, Chiba 297-0017, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 785
Author: Funatsu, O., Sato, T., Kotovuori, P., Gahmberg, C. G., Ikekita, M. and Furukawa, K.
Year: 2001
Title: Structural study of N-linked oligosaccharides of human intercellular adhesion molecule-3 (CD50)
Journal: Eur J Biochem
Volume: 268
Issue: 4
Pages: 1020-9
Epub Date: 2001/02/17
Date: Feb
Short Title: Structural study of N-linked oligosaccharides of human intercellular adhesion molecule-3 (CD50)
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.2001.01960.x
Accession Number: 11179968
Keywords: *Antigens, CD
*Antigens, Differentiation
Carbohydrate Sequence
Cell Adhesion Molecules/*chemistry
Chromatography, Liquid
Concanavalin A/chemistry
Electrophoresis, Paper
Glycoside Hydrolases/chemistry
Humans
Lectins/chemistry
Leukocytes/chemistry
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Abstract: The N-linked oligosaccharides were released from purified human intercellular adhesion molecule (ICAM)-3 by hydrazinolysis. Approximately 6 mol of oligosaccharides were released from 1 mol of ICAM-3. The oligosaccharides reduced with NaB[3H]4 were separated into neutral and acidic fractions by paper electrophoresis. Most of the acidic oligosaccharides were converted to neutral ones by digestion with sialidase, indicating that they are sialyl derivatives. The neutral and sialidase-treated acidic oligosaccharides were fractionated by serial lectin column chromatography followed by Bio-Gel P-4 column chromatography. Structural studies of each oligosaccharide by sequential exo- and endo-glycosidase digestion and by methylation analysis revealed that N-linked oligosaccharides of ICAM-3 are mainly of tri- and tetra-antennary complex-type, about 60% of which contain two to three poly N-acetyllactosamine chains terminated with the type 1 structure and those without the type 1 structure per oligosaccharide. In addition, a small amount of the high mannose-type oligosaccharide with six alpha-mannose residues, which could act as a ligand for the dendritic cell-specific ICAM-3 grabbing nonintegrin, was detected.
Notes: Funatsu, O
Sato, T
Kotovuori, P
Gahmberg, C G
Ikekita, M
Furukawa, K
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 2001 Feb;268(4):1020-9. doi: 10.1046/j.1432-1327.2001.01960.x.
Author Address: Department of Biosignal Research, Tokyo Metropolitan Institute of Gerontology, Itabashi-ku, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1571
Author: Furger, E., Frei, D. C., Schibli, R., Fischer, E. and Prota, A. E.
Year: 2013
Title: Structural basis for universal corrinoid recognition by the cobalamin transport protein haptocorrin
Journal: J Biol Chem
Volume: 288
Issue: 35
Pages: 25466-25476
Epub Date: 2013/07/13
Date: Aug 30
Short Title: Structural basis for universal corrinoid recognition by the cobalamin transport protein haptocorrin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.483271
PMCID: PMC3757208
Accession Number: 23846701
Keywords: Amino Acid Motifs
Animals
Cobamides/*chemistry/genetics/metabolism
Crystallography, X-Ray
HEK293 Cells
Humans
Mice
Protein Structure, Tertiary
Structure-Activity Relationship
Transcobalamins/*chemistry/genetics/metabolism
Biosynthesis
Cobalamin
Cobinamide
Cofactors
Corrinoids
Crystal Structure
Haptocorrin
Metabolism
Nutrition
Vitamins
Abstract: Cobalamin (Cbl; vitamin B12) is an essential micronutrient synthesized only by bacteria. Mammals have developed a sophisticated uptake system to capture the vitamin from the diet. Cbl transport is mediated by three transport proteins: transcobalamin, intrinsic factor, and haptocorrin (HC). All three proteins have a similar overall structure but a different selectivity for corrinoids. Here, we present the crystal structures of human HC in complex with cyanocobalamin and cobinamide at 2.35 and 3.0 Å resolution, respectively. The structures reveal that many of the interactions with the corrin ring are conserved among the human Cbl transporters. However, the non-conserved residues Asn-120, Arg-357, and Asn-373 form distinct interactions allowing for stabilization of corrinoids other than Cbl. A central binding motif forms interactions with the e- and f-side chains of the corrin ring and is conserved in corrinoid-binding proteins of other species. In addition, the α- and β-domains of HC form several unique interdomain contacts and have a higher shape complementarity than those of intrinsic factor and transcobalamin. The stabilization of ligands by all of these interactions is reflected in higher melting temperatures of the protein-ligand complexes. Our structural analysis offers fundamental insights into the unique binding behavior of HC and completes the picture of Cbl interaction with its three transport proteins.
Notes: 1083-351x
Furger, Evelyne
Frei, Dominik C
Schibli, Roger
Fischer, Eliane
Prota, Andrea E
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2013 Aug 30;288(35):25466-25476. doi: 10.1074/jbc.M113.483271. Epub 2013 Jul 11.
Author Address: From the Center for Radiopharmaceutical Sciences and.
Laboratory of Biomolecular Research, Department of Biology and Chemistry, Paul Scherrer Institut, CH-5232 Villigen PSI and.
From the Center for Radiopharmaceutical Sciences and; the Institute of Pharmaceutical Sciences, Eidgenössische Technische Hochschule (ETH) Zürich, CH-8093 Zürich, Switzerland.
From the Center for Radiopharmaceutical Sciences and. Electronic address: eliane.fischer@psi.ch.
Laboratory of Biomolecular Research, Department of Biology and Chemistry, Paul Scherrer Institut, CH-5232 Villigen PSI and. Electronic address: andrea.prota@psi.ch.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 894
Author: Furmanek, A., Hess, D., Rogniaux, H. and Hofsteenge, J.
Year: 2003
Title: The WSAWS motif is C-hexosylated in a soluble form of the erythropoietin receptor
Journal: Biochemistry
Volume: 42
Issue: 28
Pages: 8452-8
Epub Date: 2003/07/16
Date: Jul 22
Short Title: The WSAWS motif is C-hexosylated in a soluble form of the erythropoietin receptor
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi034112p
Accession Number: 12859190
Keywords: Amino Acid Substitution
Animals
Cloning, Molecular
Conserved Sequence
Genetic Vectors
Hexoses/chemistry/*metabolism
Mass Spectrometry
Mice
Models, Molecular
Mutagenesis, Site-Directed
Oligopeptides/chemistry/metabolism
Peptide Fragments/*chemistry/metabolism
Protein Conformation
Protein Transport
Receptors, Erythropoietin/*chemistry
Recombinant Proteins/chemistry
Solubility
Abstract: The WSXWS motif is a highly conserved structural feature of the type I cytokine receptor family. It has previously been demonstrated that mutations in the (232)WSAWS(236) motif in the erythropoietin receptor (EPOR) can result in strongly inhibited surface expression, due to defective intracellular transport [Hilton, D. J., et al. (1996) J. Biol. Chem. 271, 4699-4708]. Here we report that the first tryptophan in the motif of the recombinant extracellular domain of EPOR (sEPOR) expressed in HEK-EBNA cells carries a C-linked hexosyl residue. The S233A mutation completely abolished secretion of sEPOR, whereas the A234E mutation resulted in enhanced secretion. Comparison of the level of C-hexosylation in the wild-type protein and in the mutant proteins isolated from the conditioned medium and/or the cells suggested that C-hexosylation of the motif did not play a role in the correct intracellular transport of sEPOR.
Notes: Furmanek, Aleksandra
Hess, Daniel
Rogniaux, Hélène
Hofsteenge, Jan
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2003 Jul 22;42(28):8452-8. doi: 10.1021/bi034112p.
Author Address: Friedrich Miescher Institute, Novartis Research Foundation, Maulbeerstrasse 66, 4058 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 661
Author: Furukawa, K., Funakoshi, Y., Autero, M., Horejsi, V., Kobata, A. and Gahmberg, C. G.
Year: 1998
Title: Structural study of the O-linked sugar chains of human leukocyte tyrosine phosphatase CD45
Journal: Eur J Biochem
Volume: 251
Issue: 1-2
Pages: 288-94
Epub Date: 1998/03/10
Date: Jan 15
Short Title: Structural study of the O-linked sugar chains of human leukocyte tyrosine phosphatase CD45
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.1998.2510288.x
Accession Number: 9492296
Keywords: Carbohydrate Sequence
Chromatography, Liquid/methods
Humans
Leukocyte Common Antigens/*chemistry
Leukocytes/chemistry
Molecular Sequence Data
Oligosaccharides/*chemistry
Abstract: The O-linked sugar chains of the human leukocyte cell surface glycoprotein CD45 were released as tritium-labeled oligosaccharides by beta-elimination in the presence of NaB3H4. Mono Q column chromatography revealed that they comprise neutral (64%) and acidic (36%) oligosaccharides, the latter of which were converted to neutral ones by Arthrobacter ureafaciens sialidase treatment. Structural studies of each oligosaccharide fractionated on a Bio-Gel P-4 column by sequential exoglycosidase digestion and by methylation analysis revealed that human leukocyte CD45 contains mainly core 1 and core 2 oligosaccharides, 15% of which are modified with poly (N-acetyllactosamine) chains in different extensions. CD45 consists of several isoforms which were isolated after cell surface sialic acid residues were labeled by periodate/NaB3H4 treatment. Bio-Gel P-6 column chromatography of a mixture of the tritium-labeled glycopeptide/oligosaccharides obtained by pronase-digestion followed by mild alkaline borohydride treatment showed that distribution of the sialylated core 2 oligosaccharides is different among CD45 isoforms.
Notes: Furukawa, K
Funakoshi, Y
Autero, M
Horejsi, V
Kobata, A
Gahmberg, C G
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1998 Jan 15;251(1-2):288-94. doi: 10.1046/j.1432-1327.1998.2510288.x.
Author Address: Department of Biosignal Research, Tokyo Metropolitan Institute of Gerontology, Japan. furukawa@tmig.or.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 694
Author: Gadroy, P., Stridsberg, M., Capon, C., Michalski, J. C., Strub, J. M., Van Dorsselaer, A., Aunis, D. and Metz-Boutigue, M. H.
Year: 1998
Title: Phosphorylation and O-glycosylation sites of human chromogranin A (CGA79-439) from urine of patients with carcinoid tumors
Journal: J Biol Chem
Volume: 273
Issue: 51
Pages: 34087-97
Epub Date: 1998/12/16
Date: Dec 18
Short Title: Phosphorylation and O-glycosylation sites of human chromogranin A (CGA79-439) from urine of patients with carcinoid tumors
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.51.34087
Accession Number: 9852066
Keywords: Amino Acid Sequence
Animals
Biomarkers, Tumor/urine
Carcinoid Tumor/secondary/*urine
Cattle
Chromatography, High Pressure Liquid
Chromogranin A
Chromogranins/*chemistry/*metabolism/urine
Glycopeptides/chemistry/isolation & purification
Glycosylation
Humans
Liver Neoplasms/secondary/urine
Molecular Sequence Data
Peptide Fragments/chemistry/ultrastructure
Phosphopeptides/chemistry/isolation & purification
Phosphorylation
*Protein Processing, Post-Translational
Sequence Alignment
Sequence Homology, Amino Acid
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Because of their water-soluble properties, chromogranins (CGs) and chromogranin-derived fragments are released together with catecholamines from adrenal chromaffin cells during stress situations and can be detected in the blood by radiochemical and enzyme assays. It is well known that chromogranins can serve as immunocytochemical markers for neuroendocrine tissues and as a diagnostic tool for neuroendocrine tumors. In 1993, large CGA-derived fragments have been shown to be excreted into the urine in patients with carcinoid tumors and the present study deals with the characterization of the post-translational modifications (phosphorylation and O-glycosylation) located along the largest natural CGA-derived fragment CGA79-439. Using mild proteolysis of peptidic material, high performance liquid chromatography, sequencing, and mass spectrometry analysis, six post-translational modifications were detected along the C-terminal CGA-derived fragment CGA79-439. Three O-linked glycosylation sites were located in the core of the protein on Thr163, Thr165, and Thr233, consisting in di-, tri-, and tetrasaccharides. Three phosphorylation sites were located in the middle and C-terminal domain, on serine residues Ser200, Ser252, and Ser315. These modified sites were compared with sequences of others species and discussed in relation with the post-translational modifications that we have reported previously for bovine CGA.
Notes: Gadroy, P
Stridsberg, M
Capon, C
Michalski, J C
Strub, J M
Van Dorsselaer, A
Aunis, D
Metz-Boutigue, M H
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1998 Dec 18;273(51):34087-97. doi: 10.1074/jbc.273.51.34087.
Author Address: INSERM, Unité 338, Biologie de la Communication Cellulaire, 67084 Strasbourg, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1704
Author: Gagnon, J., Daou, S., Zamorano, N., Iannantuono, N. V., Hammond-Martel, I., Mashtalir, N., Bonneil, E., Wurtele, H., Thibault, P. and Affar el, B.
Year: 2015
Title: Undetectable histone O-GlcNAcylation in mammalian cells
Journal: Epigenetics
Volume: 10
Issue: 8
Pages: 677-91
Epub Date: 2015/06/16
Short Title: Undetectable histone O-GlcNAcylation in mammalian cells
Alternate Journal: Epigenetics
ISSN: 1559-2294 (Print)
1559-2294
DOI: 10.1080/15592294.2015.1060387
PMCID: PMC4622518
Accession Number: 26075789
Keywords: Acylation
Animals
Chromatin/genetics
DNA-Binding Proteins/*genetics
*Epigenesis, Genetic
Glycosylation
HEK293 Cells
Histones/*genetics/metabolism
Host Cell Factor C1/*genetics
Humans
N-Acetylglucosaminyltransferases/genetics/metabolism
Protein Processing, Post-Translational/genetics
Proto-Oncogene Proteins/*genetics
beta-N-Acetylhexosaminidases/*genetics/metabolism
Chromatin
Epigenetics
H2B K120ub, Histone H2B lysine 120 monoubiquitination
H2B S112 O-GlcNAc, Histone H2B serine 112 O-GlcNAc
Hcf-1
HCF-1, Host Cell Factor-1
Histone
O-GlcNAc
O-GlcNAc, O-Linked N-acetylglucosamine
O-GlcNAcylation
OGA, O-GlcNAcase
Ogt
OGT, O-Linked N-acetylglucosamine transferase
PUGNAc, O-(2-acetamido-2-deoxyglucopyranosylidene) amino N-phenylcarbamate
Polycomb
Tet2
TET2, Ten-Eleven Translocation protein 2
UDP-GlcNAc, Uridine Diphosphate N-Acetylglucosamine
WGA, Wheat Germ Agglutinin.
posttranslational modification
Abstract: O-GlcNAcylation is a posttranslational modification catalyzed by the O-Linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) and reversed by O-GlcNAcase (OGA). Numerous transcriptional regulators, including chromatin modifying enzymes, transcription factors, and co-factors, are targeted by O-GlcNAcylation, indicating that this modification is central for chromatin-associated processes. Recently, OGT-mediated O-GlcNAcylation was reported to be a novel histone modification, suggesting a potential role in directly coordinating chromatin structure and function. In contrast, using multiple biochemical approaches, we report here that histone O-GlcNAcylation is undetectable in mammalian cells. Conversely, O-GlcNAcylation of the transcription regulators Host Cell Factor-1 (HCF-1) and Ten-Eleven Translocation protein 2 (TET2) could be readily observed. Our study raises questions on the occurrence and abundance of O-GlcNAcylation as a histone modification in mammalian cells and reveals technical complications regarding the detection of genuine protein O-GlcNAcylation. Therefore, the identification of the specific contexts in which histone O-GlcNAcylation might occur is still to be established.
Notes: 1559-2308
Gagnon, Jessica
Daou, Salima
Zamorano, Natalia
Iannantuono, Nicholas V G
Hammond-Martel, Ian
Mashtalir, Nazar
Bonneil, Eric
Wurtele, Hugo
Thibault, Pierre
Affar, El Bachir
MOP-115132/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
Epigenetics. 2015;10(8):677-91. doi: 10.1080/15592294.2015.1060387.
Author Address: a Maisonneuve-Rosemont Hospital Research Center and Department of Medicine; University of Montréal ; Montréal, Québec , Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1044
Author: Gandy, J. C., Rountree, A. E. and Bijur, G. N.
Year: 2006
Title: Akt1 is dynamically modified with O-GlcNAc following treatments with PUGNAc and insulin-like growth factor-1
Journal: FEBS Lett
Volume: 580
Issue: 13
Pages: 3051-8
Epub Date: 2006/05/11
Date: May 29
Short Title: Akt1 is dynamically modified with O-GlcNAc following treatments with PUGNAc and insulin-like growth factor-1
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2006.04.051
PMCID: PMC2493066
NIHMSID: NIHMS56278
Accession Number: 16684529
Keywords: Acetylglucosamine/analogs & derivatives/pharmacology
Cell Line, Tumor
Cell Nucleus/*enzymology
Cytosol/enzymology
Glycoproteins/*metabolism
Humans
Insulin-Like Growth Factor I/pharmacology
Oximes/pharmacology
Phenylcarbamates/pharmacology
Phosphorylation
*Protein Processing, Post-Translational
Proto-Oncogene Proteins c-akt/*metabolism
Abstract: The Ser/Thr kinase Akt1 is activated by growth factors subsequent to its phosphorylation on Thr308 and Ser473. In the present study, Akt1 was found to be constitutively modified with O-GlcNAc. Treatment of SH-SY5Y cells with O(2-acetamido-2-deoxy-D-glucopyranosylidene)amino-N-phenylcarbamate (PUGNAc), which inhibits the enzymatic removal of O-GlcNAc from proteins, increased cytosolic O-GlcNAc-Akt1 levels. Treatment of cells with insulin-like growth factor-1 (IGF-1) also increased O-GlcNAc-Akt1 levels and increased Akt1 phosphorylation. PUGNAc treatment did not attenuate IGF-1 induced Akt1 phosphorylation. These results indicate that Akt1 can be simultaneously modified with O-GlcNAc and phosphorylated. However, PUGNAc induced the nuclear accumulation of Akt1 suggesting that the O-GlcNAc-modification on Akt1 may play a role in Akt1 nuclear localization.
Notes: Gandy, Johanna C
Rountree, Abigail E
Bijur, Gautam N
R01 NS044853/NS/NINDS NIH HHS/United States
R01 NS044853-04/NS/NINDS NIH HHS/United States
NS044853/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
FEBS Lett. 2006 May 29;580(13):3051-8. doi: 10.1016/j.febslet.2006.04.051. Epub 2006 Apr 27.
Author Address: Department of Psychiatry and Behavioral Neurobiology, University of Alabama at Birmingham, Sparks Center, Room 1009, Birmingham, AL 35294-0017, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1162
Author: Gao, J., Chen, T., Hu, G., Gong, Y., Qiang, B., Yuan, J. and Peng, X.
Year: 2008
Title: Nectin-like molecule 1 is a glycoprotein with a single N-glycosylation site at N290KS which influences its adhesion activity
Journal: Biochim Biophys Acta
Volume: 1778
Issue: 6
Pages: 1429-35
Epub Date: 2008/04/19
Date: Jun
Short Title: Nectin-like molecule 1 is a glycoprotein with a single N-glycosylation site at N290KS which influences its adhesion activity
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbamem.2008.03.013
Accession Number: 18420026
Keywords: Adult
Amino Acid Substitution
Animals
Axons/*metabolism
Brain/embryology/*metabolism
Cell Adhesion/physiology
Cell Adhesion Molecules
Cell Line
Glycosylation
Humans
Immunoglobulins/genetics/*metabolism
Membrane Glycoproteins/genetics/*metabolism
Membrane Proteins/genetics/*metabolism
Mice
Mutation, Missense
Nerve Tissue Proteins/genetics/*metabolism
Species Specificity
Synapses/*metabolism
Abstract: Nectin-like molecule 1 (NECL1)/CADM3/IGSF4B/TSLL1/SynCAM3, from now on referred to as NECL1, is a neural tissue-specific immunoglobulin-like cell-cell adhesion molecule which has Ca(2+)-independent homo- or heterophilic cell-cell adhesion activity and plays an important role in the formation of synapses, axon bundles and myelinated axons. Here we first detected the expression of NECL1 in human fetal and adult brains, and mouse brains at different developmental stages. The results indicated that two bands with molecular weights of about 62 kDa and 48 kDa were found in human fetal brain, while only one band with a molecular weight of about 48 kDa was found in human adult brain; two bands with molecular weights of about 62 kDa and 48 kDa whose expression level gradually increased were also found from mouse E16 to P14, while only one band with a molecular weight of about 48 kDa was found from P14. Bioinformatics analysis showed there were two putative N-glycosylation sites within human NECL1 at positions N25LS and N290KS and within mouse Necl1 at positions N23LS and N288KS, respectively. There was no O-glycosylation site in either human NECL1 or mouse Necl1. Based on the results of N-Glycosidase F treatment with human fetal brain tissue and lysates from transient transfection with human wild-type or glycosylation site mutant NECL1 in 293ET cells, we demonstrated that human NECL1 is an N-linked glycoprotein with a single glycosylation site at position N290KS. Cell aggregation assay further showed there was an increased adhesion activity after the glycosylation site mutation of NECL1 molecule.
Notes: Gao, Jing
Chen, Tao
Hu, Guangyu
Gong, Yanhua
Qiang, Boqin
Yuan, Jiangang
Peng, Xiaozhong
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2008 Jun;1778(6):1429-35. doi: 10.1016/j.bbamem.2008.03.013. Epub 2008 Mar 30.
Author Address: National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, National Human Genome Center, Beijing, China. gaojing_pumc@yahoo.com.cn
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1974
Author: Gao, L., Zhao, R., Wang, Y., Lu, M., Yang, D., Fa, M. and Yao, X.
Year: 2018
Title: Surface plasmon resonance biosensor for the accurate and sensitive quantification of O-GlcNAc based on cleavage by β-D-N-acetylglucosaminidase
Journal: Anal Chim Acta
Volume: 1040
Pages: 90-98
Epub Date: 2018/10/18
Date: Dec 21
Short Title: Surface plasmon resonance biosensor for the accurate and sensitive quantification of O-GlcNAc based on cleavage by β-D-N-acetylglucosaminidase
Alternate Journal: Analytica chimica acta
ISSN: 0003-2670
DOI: 10.1016/j.aca.2018.07.058
Accession Number: 30327117
Keywords: Acetylglucosamine/*blood/metabolism
*Biosensing Techniques
Cell Line
Gold/chemistry
Humans
Metal Nanoparticles/chemistry
N-Acetylglucosaminyltransferases/*metabolism
*Surface Plasmon Resonance
Accurate detection
Clinical diagnosis
O-GlcNAc
Spr
β-D-N-Acetylglucosaminidase
Abstract: Abnormal O-linked-N-acetylglucosamine (O-GlcNAc) concentrations have been associated with a variety of diseases (e.g., cancer, Alzheimer's disease, cardiovascular disease, etc.). However, O-GlcNAc detection is complicated, time-consuming and has poor specificity, therefore, the accurate detection of O-GlcNAc is difficult. In this study, an accurate and sensitive surface plasmon resonance (SPR) biosensor for O-GlcNAc detection that is based on β-D-N-acetylglucosaminidase (OGA) and Au nanoparticles (AuNPs) was developed. In this strategy, AuNPs were used to amplify the SPR signal and improve the biosensor's sensitivity; OGA was used to cleave O-GlcNAc from O-GlcNAcylated biomolecules. The interaction between AuNPs labeled wheat germ agglutinin (AuNPs/WGA) and O-GlcNAcylated biomolecules on a modified Au film treated with and without OGA was recorded by SPR. The change of the SPR signal moves linearly with the amount of O-GlcNAc on the Au film and thus could be used for the detection of O-GlcNAc. By recording the difference of the SPR signals, this method can avoid disturbances from other sugars and nonspecific adsorption of AuNPs and thus enable the accurate detection of O-GlcNAc. The accurate detection range of O-GlcNAc was 4.65 × 10(-12) to 4.65 × 10(-7) M which was obtained by quantifying the amount of a standard O-GlcNAcylated peptide (O-GlcNAc-CREB), and the detection limit is 4.65 × 10(-13) M. More importantly, the strategy was successfully used to detect O-GlcNAc in a real α-crystallin protein, cancer cell lysates and blood samples with satisfactory results. The study's results imply that this accurate and sensitive method has the potential to be applied in the early clinical diagnosis of O-GlcNAc-related diseases.
Notes: 1873-4324
Gao, Li
Zhao, Ruihuan
Wang, Yiwen
Lu, Mei
Yang, Dingding
Fa, Mengmei
Yao, Xin
Journal Article
Netherlands
Anal Chim Acta. 2018 Dec 21;1040:90-98. doi: 10.1016/j.aca.2018.07.058. Epub 2018 Jul 30.
Author Address: School of Chemical Science, University of Chinese Academy of Sciences, Beijing, 100049, PR China; Nanchang Institute of Technology, Nanchang 330044, PR China.
School of Chemical Science, University of Chinese Academy of Sciences, Beijing, 100049, PR China.
School of Chemical Science, University of Chinese Academy of Sciences, Beijing, 100049, PR China; State Key Laboratory of Natural and Biomimetic Drugs, Peking University, Beijing, 100191, PR China. Electronic address: yaox@ucas.ac.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2006
Author: Gao, S., Miao, Y., Liu, Y., Liu, X., Fan, X., Lin, Y., Qian, P., Zhou, J., Dai, Y., Xia, L., Zhu, P. and Zhu, J.
Year: 2019
Title: Reciprocal Regulation Between O-GlcNAcylation and β-Catenin Facilitates Cell Viability and Inhibits Apoptosis in Liver Cancer
Journal: DNA Cell Biol
Volume: 38
Issue: 4
Pages: 286-296
Epub Date: 2019/02/15
Date: Apr
Short Title: Reciprocal Regulation Between O-GlcNAcylation and β-Catenin Facilitates Cell Viability and Inhibits Apoptosis in Liver Cancer
Alternate Journal: DNA and cell biology
ISSN: 1044-5498
DOI: 10.1089/dna.2018.4447
Accession Number: 30762425
Keywords: Acetylglucosamine/*metabolism
*Apoptosis
Carcinogenesis
Cell Survival
Disease Progression
Hep G2 Cells
Humans
Liver Neoplasms/enzymology/*metabolism/*pathology
Nucleotidyltransferases/metabolism
Up-Regulation
beta Catenin/*metabolism
O-GlcNAcylation
Uap1
liver cancer
β-catenin
Abstract: Abnormal expression of O-Linked β-N-acetylglucosamine (O-GlcNAc) and β-catenin is a general feature of cancer and contributes to transformed phenotypes. In this study, we identified the interaction between O-GlcNAc and β-catenin, and explored their effects on the progression of liver cancer. Our results demonstrated that upregulation of O-GlcNAc was induced by high glucose, whereas the application of PuGNAc and GlcNAc increased β-catenin protein expression levels, as well as the protein's stability and nuclear accumulation in the liver cancer cell lines HEP-G2 and HuH-7. In addition, overexpression of β-catenin could increase O-GlcNAc expression levels through upregulation of uridine 5'-diphosphate (UDP)-N-acetylglucosamine pyrophosphorylase 1 (UAP1) protein expression, protein stability, and inhibition of its ubiquitination. Moreover, the O-GlcNAcylation of β-catenin promoted the proliferation, colony formation, and repressed the induction of apoptosis in HEP-G2 and HuH-7 cells. Knockdown of β-catenin reduced cell proliferation, colony formation, and tumorigenesis, and promoted cell apoptosis through the downregulation of UAP1 expression. In conclusion, this study revealed that the reciprocal regulation between O-GlcNAcylation and β-catenin facilitated the proliferation of liver cancer.
Notes: 1557-7430
Gao, Sicheng
Miao, Yun
Liu, Yijie
Liu, Xing
Fan, Xingliang
Lin, Yan
Qian, Pingan
Zhou, Jun
Dai, Yaoyao
Xia, Li
Zhu, Po
Zhu, Junfeng
Journal Article
United States
DNA Cell Biol. 2019 Apr;38(4):286-296. doi: 10.1089/dna.2018.4447. Epub 2019 Feb 14.
Author Address: 1 Department of Hepatology, Shanghai Municipal Hospital of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, P.R. China.
2 Department of Rehabilitation, Shanghai General Hospital, Shanghai, P.R. China.
3 Department of Rehabilitation, Shanghai University of Traditional Chinese Medicine, Shanghai, P.R. China.
4 Department of Central Laboratory Medicine, Shanghai Municipal Hospital of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, P.R. China.
5 Department of Liver Disease, Chongqing Traditional Chinese Medicine Hospital, Chongqing, P.R. China.
6 Department of Neurology, Minhang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai, P.R. China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2002
Author: Gao, Y., Liu, J., Bai, Z., Sink, S., Zhao, C., Lorenzo, F. R. and McClain, D. A.
Year: 2019
Title: Iron down-regulates leptin by suppressing protein O-GlcNAc modification in adipocytes, resulting in decreased levels of O-glycosylated CREB
Journal: J Biol Chem
Volume: 294
Issue: 14
Pages: 5487-5495
Epub Date: 2019/02/03
Date: Apr 5
Short Title: Iron down-regulates leptin by suppressing protein O-GlcNAc modification in adipocytes, resulting in decreased levels of O-glycosylated CREB
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA118.005183
PMCID: PMC6462527
Accession Number: 30709903
Keywords: 3T3-L1 Cells
Adipocytes/*metabolism
Animals
Cyclic AMP Response Element-Binding Protein/*metabolism
Down-Regulation/*drug effects
Glucosamine/metabolism
Glycosylation/drug effects
Iron/metabolism/*pharmacology
Leptin/*biosynthesis
Mice
*Models, Biological
Promoter Regions, Genetic
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc)
*adipocyte
*diabetes
*iron
*leptin
this article
Abstract: We previously reported that iron down-regulates transcription of the leptin gene by increasing occupancy of phosphorylated cAMP response element-binding protein (pCREB) at two sites in the leptin gene promoter. Several nutrient-sensing pathways including O-GlcNAcylation also regulate leptin. We therefore investigated whether O-glycosylation plays a role in iron- and CREB-mediated regulation of leptin. We found that high iron decreases protein O-GlcNAcylation both in cultured 3T3-L1 adipocytes and in mice fed high-iron diets and down-regulates leptin mRNA and protein levels. Glucosamine treatment, which bypasses the rate-limiting step in the synthesis of substrate for glycosylation, increased both O-GlcNAc and leptin, whereas inhibition of O-glycosyltransferase (OGT) decreased O-GlcNAc and leptin. The increased leptin levels induced by glucosamine were susceptible to the inhibition by iron, but in the case of OGT inhibition, iron did not further decrease leptin. Mice with deletion of the O-GlcNAcase gene, either via whole-body heterozygous deletion or through adipocyte-targeted homozygous deletion, exhibited increased O-GlcNAc levels in adipose tissue and increased leptin levels that were inhibited by iron. Of note, iron increased the occupancy of pCREB and decreased the occupancy of O-GlcNAcylated CREB on the leptin promoter. These patterns observed in our experimental models suggest that iron exerts its effects on leptin by decreasing O-glycosylation and not by increasing protein deglycosylation and that neither O-GlcNAcase nor OGT mRNA and protein levels are affected by iron. We conclude that iron down-regulates leptin by decreasing CREB glycosylation, resulting in increased CREB phosphorylation and leptin promoter occupancy by pCREB.
Notes: 1083-351x
Gao, Yan
Liu, Jingfang
Bai, Zhenzhong
Sink, Sandy
Zhao, Chengyu
Lorenzo, Felipe Ramos
McClain, Donald A
Orcid: 0000-0002-3310-2359
I01 BX001140/BX/BLRD VA/United States
R01 DK081842/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2019 Apr 5;294(14):5487-5495. doi: 10.1074/jbc.RA118.005183. Epub 2019 Feb 1.
Author Address: From the Department of Internal Medicine, Wake Forest School of Medicine, Winston Salem, North Carolina 27157 and.
From the Department of Internal Medicine, Wake Forest School of Medicine, Winston Salem, North Carolina 27157 and dmcclain@wakehealth.edu.
the W. G. Hefner Veterans Affairs Medical Center, Salisbury, North Carolina 28144.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1063
Author: Garcia-Alles, L. F., Versluis, K., Maveyraud, L., Vallina, A. T., Sansano, S., Bello, N. F., Gober, H. J., Guillet, V., de la Salle, H., Puzo, G., Mori, L., Heck, A. J., De Libero, G. and Mourey, L.
Year: 2006
Title: Endogenous phosphatidylcholine and a long spacer ligand stabilize the lipid-binding groove of CD1b
Journal: Embo j
Volume: 25
Issue: 15
Pages: 3684-92
Epub Date: 2006/07/29
Date: Aug 9
Short Title: Endogenous phosphatidylcholine and a long spacer ligand stabilize the lipid-binding groove of CD1b
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7601244
PMCID: PMC1538567
Accession Number: 16874306
Keywords: Antigens, CD1/*chemistry/metabolism
Binding Sites
Crystallography, X-Ray
Humans
Hydrophobic and Hydrophilic Interactions
Isoelectric Focusing
Ligands
Mass Spectrometry
Models, Molecular
Phosphatidylcholines/*chemistry/metabolism
Protein Binding
Protein Conformation
Protein Folding
Abstract: CD1 proteins present lipid antigens to T cells. The antigens are acquired in the endosomal compartments. This raises the question of how the large hydrophobic CD1 pockets are preserved between the moment of biosynthesis in the endoplasmic reticulum and arrival to the endosomes. To address this issue, the natural ligands associated with a soluble form of human CD1b have been investigated. Using isoelectric focusing, native mass spectrometry and resolving the crystal structure at 1.8 A resolution, we found that human CD1b is simultaneously associated with endogenous phosphatidylcholine (PC) and a 41-44 carbon atoms-long spacer molecule. The two lipids appear to work in concert to stabilize the CD1b groove, their combined size slightly exceeding the maximal groove capacity. We propose that the spacer serves to prevent binding of ligands with long lipid tails, whereas short-chain lipids might still displace the PC, which is exposed at the groove entrance. The data presented herein explain how the CD1b groove is preserved, and provide a rationale for the in vivo antigen-binding properties of CD1b.
Notes: 1460-2075
Garcia-Alles, Luis F
Versluis, Kees
Maveyraud, Laurent
Vallina, Ana Tesouro
Sansano, Sebastiano
Bello, Nana Fatimath
Gober, Hans-Jürgen
Guillet, Valérie
de la Salle, Henri
Puzo, Germain
Mori, Lucia
Heck, Albert J R
De Libero, Gennaro
Mourey, Lionel
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2006 Aug 9;25(15):3684-92. doi: 10.1038/sj.emboj.7601244. Epub 2006 Jul 27.
Author Address: CNRS, UMR5089, Département Mécanismes Moléculaires des Infections Mycobactériennes, Institut de Pharmacologie et de Biologie Structurale, Toulouse, France. Luis-Fernando.Garcia-Alles@ipbs.fr
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1006
Author: García-Castellanos, R., Bonet-Figueredo, R., Pallarés, I., Ventura, S., Avilés, F. X., Vendrell, J. and Gomis-Rütha, F. X.
Year: 2005
Title: Detailed molecular comparison between the inhibition mode of A/B-type carboxypeptidases in the zymogen state and by the endogenous inhibitor latexin
Journal: Cell Mol Life Sci
Volume: 62
Issue: 17
Pages: 1996-2014
Epub Date: 2005/08/11
Date: Sep
Short Title: Detailed molecular comparison between the inhibition mode of A/B-type carboxypeptidases in the zymogen state and by the endogenous inhibitor latexin
Alternate Journal: Cellular and molecular life sciences : CMLS
ISSN: 1420-682X (Print)
1420-682x
DOI: 10.1007/s00018-005-5174-4
Accession Number: 16091843
Keywords: Amino Acid Sequence
Animals
Antigens/*chemistry/*pharmacology
Carboxypeptidase B/antagonists & inhibitors/chemistry
Carboxypeptidases A/*antagonists & inhibitors/*chemistry
Crystallography, X-Ray
Enzyme Precursors/*antagonists & inhibitors/*chemistry
Humans
Male
Mice
Molecular Sequence Data
Prostatic Neoplasms/*enzymology
Protein Conformation
Abstract: Treatment of advanced stages of prostate carcinoma with histone-deacetylase inhibitors entails expression of human procarboxypeptidase-A4 (hPCPA4). The three-dimensional structure of hPCPA4 has been solved and shows the features of related metallocarboxypeptidase zymogens, with a preformed alpha/beta/-hydrolase active-enzyme moiety (hCPA4) and an inhibiting pro-domain (PD). The protease moiety recalls a sphere, out of which a spherical cone has been cut. This results in a funnel-like structure, at the bottom of which the active-site cleft resides. The border of this funnel is shaped by loops, which are responsible for the interaction with the PD, characterised by a large interface area and relatively few contacts. Such an inhibitory mode is evocative of the recently reported structure of the human inhibitor latexin in its complex with hCPA4. The main contacting structure of latexin is similar to the one employed for PD inhibition. In both cases, active-site blocking relies mainly on a loop provided by the central part of a beta sheet.
Notes: García-Castellanos, R
Bonet-Figueredo, R
Pallarés, I
Ventura, S
Avilés, F X
Vendrell, J
Gomis-Rütha, F X
Journal Article
Research Support, Non-U.S. Gov't
Switzerland
Cell Mol Life Sci. 2005 Sep;62(17):1996-2014. doi: 10.1007/s00018-005-5174-4.
Author Address: Institut de Biologia Molecular de Barcelona, C.I.D. - C.S.I.C., C/Jordi Girona 18-26, 08034, Barcelona, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1088
Author: Garlatti, V., Belloy, N., Martin, L., Lacroix, M., Matsushita, M., Endo, Y., Fujita, T., Fontecilla-Camps, J. C., Arlaud, G. J., Thielens, N. M. and Gaboriaud, C.
Year: 2007
Title: Structural insights into the innate immune recognition specificities of L- and H-ficolins
Journal: Embo j
Volume: 26
Issue: 2
Pages: 623-33
Epub Date: 2007/01/12
Date: Jan 24
Short Title: Structural insights into the innate immune recognition specificities of L- and H-ficolins
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7601500
PMCID: PMC1783469
Accession Number: 17215869
Keywords: Acetylation
Acetyltransferases/metabolism
Amino Acid Sequence
Binding Sites
Galactose/metabolism
Glycoproteins/*chemistry/genetics/metabolism
Humans
Immunity, Innate
Lectins/*chemistry/genetics/metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Promoter Regions, Genetic
Protein Subunits/chemistry/metabolism
Sequence Homology, Amino Acid
Substrate Specificity
beta-Glucans/metabolism
Abstract: Innate immunity relies critically upon the ability of a few pattern recognition molecules to sense molecular markers on pathogens, but little is known about these interactions at the atomic level. Human L- and H-ficolins are soluble oligomeric defence proteins with lectin-like activity, assembled from collagen fibers prolonged by fibrinogen-like recognition domains. The X-ray structures of their trimeric recognition domains, alone and in complex with various ligands, have been solved to resolutions up to 1.95 and 1.7 A, respectively. Both domains have three-lobed structures with clefts separating the distal parts of the protomers. Ca(2+) ions are found at sites homologous to those described for tachylectin 5A (TL5A), an invertebrate lectin. Outer binding sites (S1) homologous to the GlcNAc-binding pocket of TL5A are present in the ficolins but show different structures and specificities. In L-ficolin, three additional binding sites (S2-S4) surround the cleft. Together, they define an unpredicted continuous recognition surface able to sense various acetylated and neutral carbohydrate markers in the context of extended polysaccharides such as 1,3-beta-D-glucan, as found on microbial or apoptotic surfaces.
Notes: 1460-2075
Garlatti, Virginie
Belloy, Nicolas
Martin, Lydie
Lacroix, Monique
Matsushita, Misao
Endo, Yuichi
Fujita, Teizo
Fontecilla-Camps, Juan Carlos
Arlaud, Gérard J
Thielens, Nicole M
Gaboriaud, Christine
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2007 Jan 24;26(2):623-33. doi: 10.1038/sj.emboj.7601500. Epub 2007 Jan 11.
Author Address: Laboratoire de Cristallographie et Cristallogénèse des Protéines, Grenoble, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 927
Author: Garman, S. C. and Garboczi, D. N.
Year: 2004
Title: The molecular defect leading to Fabry disease: structure of human alpha-galactosidase
Journal: J Mol Biol
Volume: 337
Issue: 2
Pages: 319-35
Epub Date: 2004/03/09
Date: Mar 19
Short Title: The molecular defect leading to Fabry disease: structure of human alpha-galactosidase
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2004.01.035
Accession Number: 15003450
Keywords: Carbohydrate Metabolism
Carbohydrates/chemistry
Catalytic Domain
Codon, Nonsense
Crystallography, X-Ray
Dimerization
Evolution, Molecular
*Fabry Disease/*enzymology/*genetics
Humans
Lysosomes/enzymology
Male
Models, Molecular
Mutation, Missense
Phylogeny
Protein Conformation
Static Electricity
Substrate Specificity
alpha-Galactosidase/chemistry/*genetics
Abstract: Fabry disease is an X-linked lysosomal storage disease afflicting 1 in 40,000 males with chronic pain, vascular degeneration, cardiac impairment, and other symptoms. Deficiency in the lysosomal enzyme alpha-galactosidase (alpha-GAL) causes an accumulation of its substrate, which ultimately leads to Fabry disease symptoms. Here, we present the structure of the human alpha-GAL glycoprotein determined by X-ray crystallography. The structure is a homodimer with each monomer containing a (beta/alpha)8 domain with the active site and an antiparallel beta domain. N-linked carbohydrate appears at six sites in the glycoprotein dimer, revealing the basis for lysosomal transport via the mannose-6-phosphate receptor. To understand how the enzyme cleaves galactose from glycoproteins and glycolipids, we also determined the structure of the complex of alpha-GAL with its catalytic product. The catalytic mechanism of the enzyme is revealed by the location of two aspartic acid residues (D170 and D231), which act as a nucleophile and an acid/base, respectively. As a point mutation in alpha-GAL can lead to Fabry disease, we have catalogued and plotted the locations of 245 missense and nonsense mutations in the three-dimensional structure. The structure of human alpha-GAL brings Fabry disease into the realm of molecular diseases, where insights into the structural basis of the disease phenotypes might help guide the clinical treatment of patients.
Notes: Garman, Scott C
Garboczi, David N
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
J Mol Biol. 2004 Mar 19;337(2):319-35. doi: 10.1016/j.jmb.2004.01.035.
Author Address: Structural Biology Section, Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Twinbrook II, 12441 Parklawn Drive, Rockville, MD 20852, USA. sgarman@niaid.nih.gov
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 696
Author: Garman, S. C., Kinet, J. P. and Jardetzky, T. S.
Year: 1998
Title: Crystal structure of the human high-affinity IgE receptor
Journal: Cell
Volume: 95
Issue: 7
Pages: 951-61
Epub Date: 1999/01/06
Date: Dec 23
Short Title: Crystal structure of the human high-affinity IgE receptor
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/s0092-8674(00)81719-5
Accession Number: 9875849
Keywords: Amino Acid Sequence
Animals
Binding Sites
Cell Line
Crystallization
Crystallography, X-Ray/methods
Glycosylation
Humans
Immunoglobulin E/metabolism
Models, Molecular
Molecular Sequence Data
Molecular Weight
Protein Conformation
Protein Structure, Tertiary
Receptors, IgE/*chemistry/genetics/isolation & purification/metabolism
Recombinant Proteins/chemistry
Sequence Alignment
Solubility
Tryptophan
Abstract: Allergic responses result from the activation of mast cells by the human high-affinity IgE receptor. IgE-mediated allergic reactions may develop to a variety of environmental compounds, but the initiation of a response requires the binding of IgE to its high-affinity receptor. We have solved the X-ray crystal structure of the antibody-binding domains of the human IgE receptor at 2.4 A resolution. The structure reveals a highly bent arrangement of immunoglobulin domains that form an extended convex surface of interaction with IgE. A prominent loop that confers specificity for IgE molecules extends from the receptor surface near an unusual arrangement of four exposed tryptophans. The crystal structure of the IgE receptor provides a foundation for the development of new therapeutic approaches to allergy treatment.
Notes: Garman, S C
Kinet, J P
Jardetzky, T S
AI-38972/AI/NIAID NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Cell. 1998 Dec 23;95(7):951-61. doi: 10.1016/s0092-8674(00)81719-5.
Author Address: Department of Biochemistry, Molecular Biology, and Cell Biology, Northwestern University, Evanston, Illinois 60208, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 811
Author: Garman, S. C., Sechi, S., Kinet, J. P. and Jardetzky, T. S.
Year: 2001
Title: The analysis of the human high affinity IgE receptor Fc epsilon Ri alpha from multiple crystal forms
Journal: J Mol Biol
Volume: 311
Issue: 5
Pages: 1049-62
Epub Date: 2001/09/05
Date: Aug 31
Short Title: The analysis of the human high affinity IgE receptor Fc epsilon Ri alpha from multiple crystal forms
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1006/jmbi.2001.4929
Accession Number: 11531339
Keywords: Binding Sites
Carbohydrate Metabolism
Crystallization
Crystallography, X-Ray
Databases as Topic
Drug Design
Glycoproteins/chemistry/metabolism
Humans
Immunoglobulin E/chemistry/metabolism
Immunoglobulin Heavy Chains/chemistry/metabolism
Ligands
Models, Molecular
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, IgE/*chemistry/*metabolism
Thermodynamics
Abstract: We have solved the structure of the human high affinity IgE receptor, Fc epsilon RI alpha, in six different crystal forms, showing the structure in 15 different chemical environments. This database of structures shows no change in the overall shape of the molecule, as the angle between domains 1 and 2 (D1 and D2) varies little across the ensemble. However, the receptor has local conformational variability in the C' strand of D2 and in the BC loop of D1. In every crystal form, a residue inserts between tryptophan residues 87 and 110, mimicking the position of a proline from the IgE ligand. The different crystal forms reveal a distribution of carbohydrates lining the front and back surfaces of the structure. An analysis of crystal contacts in the different forms indicates regions where the molecule interacts with other proteins, and reveals a potential new binding site distal to the IgE binding site. The results of this study point to new directions for the design of molecules to inhibit the interaction of Fc epsilon RI alpha with its natural ligand and thus to prevent a primary step in the allergic response.
Notes: Garman, S C
Sechi, S
Kinet, J P
Jardetzky, T S
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
J Mol Biol. 2001 Aug 31;311(5):1049-62. doi: 10.1006/jmbi.2001.4929.
Author Address: Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Twinbrook II, 12441 Parklawn Drive, Rockville, MD 20852, USA. garman@alpha.niaid.nih.gov
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 817
Author: Garner, B., Harvey, D. J., Royle, L., Frischmann, M., Nigon, F., Chapman, M. J. and Rudd, P. M.
Year: 2001
Title: Characterization of human apolipoprotein B100 oligosaccharides in LDL subfractions derived from normal and hyperlipidemic plasma: deficiency of alpha-N-acetylneuraminyllactosyl-ceramide in light and small dense LDL particles
Journal: Glycobiology
Volume: 11
Issue: 10
Pages: 791-802
Epub Date: 2001/10/06
Date: Oct
Short Title: Characterization of human apolipoprotein B100 oligosaccharides in LDL subfractions derived from normal and hyperlipidemic plasma: deficiency of alpha-N-acetylneuraminyllactosyl-ceramide in light and small dense LDL particles
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/11.10.791
Accession Number: 11588155
Keywords: Apolipoprotein B-100
Apolipoproteins B/*chemistry
Carbohydrate Sequence
Ceramides/*analysis
Humans
Hyperlipidemias/*blood
Lipoproteins, LDL/blood/*chemistry
Molecular Sequence Data
Oligosaccharides/*analysis
Polysaccharides/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: The carbohydrate composition of apolipoprotein (apo) B100, particularly its degree of sialylation, may contribute to the atherogenic properties of low-density lipoprotein (LDL). We analyzed LDL apoB100 glycans derived from normolipidemic, hypercholesterolemic, and hypertriglyceridemic diabetic subjects. Using exoglycosidase carbohydrate sequencing and matrix-assisted laser desorption/ionization mass spectrometry to analyze fluorescently labeled oligosaccharides, we report evidence for several carbohydrates not previously identified on apoB100, including truncated complex biantennary N-glycans and hybrid N-glycans. The distribution and diversity of the apoB100 glycans isolated from all individuals was highly conserved. The N-glycan composition of apoB100 derived from five LDL subpopulations (LDL1, d = 1.018-1.023; LDL2, d = 1.023-1.030; LDL3, d = 1.030-1.040; LDL4, d = 1.040-1.051; LDL5, d = 1.051-1.065 g/ml) did not vary in normolipidemic or hypercholesterolemic subjects. Furthermore, we found no evidence for "desialylated" apoB100 glycans in any of the samples analyzed. Analysis of the most abundant LDL ganglioside, alpha-N-acetylneuraminyllactosyl-ceramide, revealed a deficiency in small dense LDL and in the most buoyant subpopulation. These data provide a novel explanation for the apparent deficiency of sialic acid in small dense LDL and indicate that the global apoB100 N-glycan composition is invariable in the patient groups studied.
Notes: Garner, B
Harvey, D J
Royle, L
Frischmann, M
Nigon, F
Chapman, M J
Rudd, P M
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2001 Oct;11(10):791-802. doi: 10.1093/glycob/11.10.791.
Author Address: Oxford Glycobiology Institute, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 795
Author: Garner, B., Merry, A. H., Royle, L., Harvey, D. J., Rudd, P. M. and Thillet, J.
Year: 2001
Title: Structural elucidation of the N- and O-glycans of human apolipoprotein(a): role of o-glycans in conferring protease resistance
Journal: J Biol Chem
Volume: 276
Issue: 25
Pages: 22200-8
Epub Date: 2001/04/11
Date: Jun 22
Short Title: Structural elucidation of the N- and O-glycans of human apolipoprotein(a): role of o-glycans in conferring protease resistance
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M102150200
Accession Number: 11294842
Keywords: Apolipoproteins/chemistry/*metabolism
Apoprotein(a)
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Endopeptidases/*metabolism
Glycoside Hydrolases/metabolism
Glycosylation
Lipoprotein(a)/chemistry/*metabolism
Mass Spectrometry/methods
Molecular Sequence Data
Polysaccharides/chemistry/*metabolism
Protein Conformation
Abstract: Apolipoprotein(a) (apo(a)) is a multikringle domain glycoprotein that exists covalently linked to apolipoprotein B100 of low density lipoprotein, to form the lipoprotein(a) (Lp(a)) particle, or as proteolytic fragments. Elevated plasma concentrations of apo(a) and its fragments may promote atherosclerosis, but the underlying mechanisms are incompletely understood. The factors influencing apo(a) proteolysis are also uncertain. Here we have used exoglycosidase digestion and mass spectrometry to sequence the Asn (N)-linked and Ser/Thr (O)-linked oligosaccharides of human apo(a). We also assessed the potential role of apo(a) O-glycans in protecting thermolysin-sensitive regions of the polypeptide. Apo(a) contained two major N-glycans that accounted for 17% of the total oligosaccharide structures. The N-glycans were complex biantennary structures present in either a mono- or disialylated state. The O-glycans were mostly (80%) represented by the monosialylated core type 1 structure, NeuNAcalpha2-3Galbeta1-3GalNAc, with smaller amounts of disialylated and non-sialylated O-glycans also detected. Removal of apo(a) O-glycans by sialidase and O-glycosidase treatment dramatically increased the sensitivity of the polypeptide to thermolysin digestion. These studies provide the first direct sequencing data for apo(a) glycans and indicate a novel function for apo(a) O-glycans that is potentially related to the atherogenicity of Lp(a).
Notes: Garner, B
Merry, A H
Royle, L
Harvey, D J
Rudd, P M
Thillet, J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2001 Jun 22;276(25):22200-8. doi: 10.1074/jbc.M102150200. Epub 2001 Apr 6.
Author Address: Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, United Kingdom. brett@glycob.ox.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 852
Author: Garrett, T. P., McKern, N. M., Lou, M., Elleman, T. C., Adams, T. E., Lovrecz, G. O., Zhu, H. J., Walker, F., Frenkel, M. J., Hoyne, P. A., Jorissen, R. N., Nice, E. C., Burgess, A. W. and Ward, C. W.
Year: 2002
Title: Crystal structure of a truncated epidermal growth factor receptor extracellular domain bound to transforming growth factor alpha
Journal: Cell
Volume: 110
Issue: 6
Pages: 763-73
Epub Date: 2002/09/26
Date: Sep 20
Short Title: Crystal structure of a truncated epidermal growth factor receptor extracellular domain bound to transforming growth factor alpha
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/s0092-8674(02)00940-6
Accession Number: 12297049
Keywords: 3T3 Cells
Amino Acid Sequence
Amino Acid Substitution
Animals
Binding Sites
Cell Line
Conserved Sequence
Crystallization
Crystallography, X-Ray
Dimerization
Disulfides/chemistry
ErbB Receptors/*chemistry/*metabolism
Humans
Ligands
Mice
*Models, Molecular
Molecular Sequence Data
Molecular Structure
Mutation
Protein Structure, Secondary
Protein Structure, Tertiary
Sequence Alignment
Transforming Growth Factor alpha/*chemistry/genetics/*metabolism
Abstract: We report the crystal structure, at 2.5 A resolution, of a truncated human EGFR ectodomain bound to TGFalpha. TGFalpha interacts with both L1 and L2 domains of EGFR, making many main chain contacts with L1 and interacting with L2 via key conserved residues. The results indicate how EGFR family members can bind a family of highly variable ligands. In the 2:2 TGFalpha:sEGFR501 complex, each ligand interacts with only one receptor molecule. There are two types of dimers in the asymmetric unit: a head-to-head dimer involving contacts between the L1 and L2 domains and a back-to-back dimer dominated by interactions between the CR1 domains of each receptor. Based on sequence conservation, buried surface area, and mutagenesis experiments, the back-to-back dimer is favored to be biologically relevant.
Notes: Garrett, Thomas P J
McKern, Neil M
Lou, Meizhen
Elleman, Thomas C
Adams, Timothy E
Lovrecz, George O
Zhu, Hong-Jian
Walker, Francesca
Frenkel, Morry J
Hoyne, Peter A
Jorissen, Robert N
Nice, Edouard C
Burgess, Antony W
Ward, Colin W
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell. 2002 Sep 20;110(6):763-73. doi: 10.1016/s0092-8674(02)00940-6.
Author Address: Walter and Eliza Hall Institute of Medical Research, Post Office Royal Melbourne Hospital, Parkville, 3050, Victoria, Australia. tgarrett@wehi.edu.au
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 681
Author: Garrett, T. P., McKern, N. M., Lou, M., Frenkel, M. J., Bentley, J. D., Lovrecz, G. O., Elleman, T. C., Cosgrove, L. J. and Ward, C. W.
Year: 1998
Title: Crystal structure of the first three domains of the type-1 insulin-like growth factor receptor
Journal: Nature
Volume: 394
Issue: 6691
Pages: 395-9
Epub Date: 1998/08/05
Date: Jul 23
Short Title: Crystal structure of the first three domains of the type-1 insulin-like growth factor receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/28668
Accession Number: 9690478
Keywords: Alanine/metabolism
Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Cysteine/metabolism
Humans
Insulin/metabolism
Insulin-Like Growth Factor I/metabolism
Models, Molecular
Molecular Sequence Data
Peptide Fragments/chemistry
Protein Conformation
Receptor, IGF Type 1/*chemistry/metabolism
Abstract: The type-1 insulin-like growth-factor receptor (IGF-1R) and insulin receptor (IR) are closely related members of the tyrosine-kinase receptor superfamily. IR is essential for glucose homeostasis, whereas IGF-1R is involved in both normal growth and development and malignant transformation. Homologues of these receptors are found in animals as simple as cnidarians. The epidermal growth-factor receptor (EGFR) family is closely related to the IR family and has significant sequence identity to the extracellular portion we describe here. We now present the structure of the first three domains of IGF-IR (L1-Cys-rich-L2) determined to 2.6 A resolution. The L domains each consist of a single-stranded right-handed beta-helix. The Cys-rich region is composed of eight disulphide-bonded modules, seven of which form a rod-shaped domain with modules associated in an unusual manner. The three domains surround a central space of sufficient size to accommodate a ligand molecule. Although the fragment (residues 1-462) does not bind ligand, many of the determinants responsible for hormone binding and ligand specificity map to this central site. This structure therefore shows how the IR subfamily might interact with their ligands.
Notes: Garrett, T P
McKern, N M
Lou, M
Frenkel, M J
Bentley, J D
Lovrecz, G O
Elleman, T C
Cosgrove, L J
Ward, C W
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 1998 Jul 23;394(6691):395-9. doi: 10.1038/28668.
Author Address: Biomolecular Research Institute, Parkville, Victoria, Australia. tom.barrett@bioresi.com.au
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 851
Author: Garrigue-Antar, L., Hartigan, N. and Kadler, K. E.
Year: 2002
Title: Post-translational modification of bone morphogenetic protein-1 is required for secretion and stability of the protein
Journal: J Biol Chem
Volume: 277
Issue: 45
Pages: 43327-34
Epub Date: 2002/09/10
Date: Nov 8
Short Title: Post-translational modification of bone morphogenetic protein-1 is required for secretion and stability of the protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M207342200
Accession Number: 12218058
Keywords: Amino Acid Substitution
Base Sequence
Bone Morphogenetic Protein 1
Bone Morphogenetic Proteins/chemistry/*metabolism
Cell Line
DNA Primers
Enzyme Stability
Fibrosarcoma
Glycosylation
Humans
Kidney
Kinetics
Metalloendopeptidases/chemistry/*metabolism
Microscopy, Fluorescence
Mutagenesis, Site-Directed
Polymerase Chain Reaction
*Protein Processing, Post-Translational
Substrate Specificity
Transfection
Tumor Cells, Cultured
Abstract: Bone morphogenetic protein (BMP)-1 is a glycosylated metalloproteinase that is fundamental to the synthesis of a normal extracellular matrix because it cleaves type I procollagen, as well as other precursor proteins. Sequence analysis suggests that BMP-1 has six potential N-linked glycosylation sites (i.e. NXS/T) namely: Asn(91) (prodomain), Asn(142) (metalloproteinase domain), Asn(332) and Asn(363) (CUB1 domain), Asn(599) (CUB3 domain), and Asn(726) in the C-terminal-specific domain. In this study we showed that all these sites are N-glycosylated with complex-type oligosaccharides containing sialic acid, except Asn(726) presumably because proline occurs immediately C-terminal of threonine in the consensus sequence. Recombinant BMP-1 molecules lacking all glycosylation sites or the three CUB-specific sites were not secreted. BMP-1 lacking CUB glycosylation was translocated to the proteasome for degradation. BMP-1 molecules lacking individual glycosylation sites were efficiently secreted and exhibited full procollagen C-proteinase activity, but N332Q and N599Q exhibited a slower rate of cleavage. BMP-1 molecules lacking any one of the CUB-specific glycosylation sites were sensitive to thermal denaturation. The study showed that the glycosylation sites in the CUB domains of BMP-1 are important for secretion and stability of the molecule.
Notes: Garrigue-Antar, Laure
Hartigan, Nichola
Kadler, Karl E
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2002 Nov 8;277(45):43327-34. doi: 10.1074/jbc.M207342200. Epub 2002 Sep 5.
Author Address: Wellcome Trust Centre for Cell-Matrix Research, University of Manchester, School of Biological Sciences, Stopford Building 2.205, Oxford Road, Manchester M13 9PT, United Kingdom. laure.garrigue-antar@man.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1496
Author: Gawlowski, T., Suarez, J., Scott, B., Torres-Gonzalez, M., Wang, H., Schwappacher, R., Han, X., Yates, J. R., 3rd, Hoshijima, M. and Dillmann, W.
Year: 2012
Title: Modulation of dynamin-related protein 1 (DRP1) function by increased O-linked-β-N-acetylglucosamine modification (O-GlcNAc) in cardiac myocytes
Journal: J Biol Chem
Volume: 287
Issue: 35
Pages: 30024-34
Epub Date: 2012/06/30
Date: Aug 24
Short Title: Modulation of dynamin-related protein 1 (DRP1) function by increased O-linked-β-N-acetylglucosamine modification (O-GlcNAc) in cardiac myocytes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.390682
PMCID: PMC3436129
Accession Number: 22745122
Keywords: Acetylation
Acetylglucosamine/genetics/*metabolism
Acetylglucosaminidase/genetics/metabolism
Animals
Cytoplasm/genetics/metabolism/pathology
Diabetes Complications/genetics/*metabolism/pathology
Diabetes Mellitus, Experimental/genetics/*metabolism/pathology
Dynamins/genetics/*metabolism
Humans
Membrane Potential, Mitochondrial/genetics
Mice
Mitochondria, Heart/genetics/*metabolism/pathology
Mitochondrial Diseases/genetics/*metabolism/pathology
Muscle Proteins/genetics/*metabolism
Myocytes, Cardiac/*metabolism/pathology
Phosphorylation/genetics
Protein Transport/genetics
Abstract: O-linked-N-acetyl-glucosamine glycosylation (O-GlcNAcylation) of the serine and threonine residues of cellular proteins is a dynamic process and affects phosphorylation. Prolonged O-GlcNAcylation has been linked to diabetes-related complications, including mitochondrial dysfunction. Mitochondria are dynamically remodeling organelles, that constantly fuse (fusion) and divide (fission). An imbalance of this process affects mitochondrial function. In this study, we found that dynamin-related protein 1 (DRP1) is O-GlcNAcylated in cardiomyocytes at threonine 585 and 586. O-GlcNAcylation was significantly enhanced by the chemical inhibition of N-acetyl-glucosaminidase. Increased O-GlcNAcylation decreases the phosphorylation of DRP1 at serine 637, which is known to regulate DRP1 function. In fact, increased O-GlcNAcylation augments the level of the GTP-bound active form of DRP1 and induces translocation of DRP1 from the cytoplasm to mitochondria. Mitochondrial fragmentation and decreased mitochondrial membrane potential also accompany the increased O-GlcNAcylation. In conclusion, this report shows, for the first time, that O-GlcNAcylation modulates DRP1 functionality in cardiac muscle cells.
Notes: 1083-351x
Gawlowski, Thomas
Suarez, Jorge
Scott, Brian
Torres-Gonzalez, Moises
Wang, Hong
Schwappacher, Raphaela
Han, Xuemei
Yates, John R 3rd
Hoshijima, Masahiko
Dillmann, Wolfgang
P41 GM103533/GM/NIGMS NIH HHS/United States
P30 NS047101/NS/NINDS NIH HHS/United States
P60 MD000220/MD/NIMHD NIH HHS/United States
P41 RR011823/RR/NCRR NIH HHS/United States
P60 MD00220/MD/NIMHD NIH HHS/United States
HL066917/HL/NHLBI NIH HHS/United States
R01 HL066917/HL/NHLBI NIH HHS/United States
3R01HL066917-10S1/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Aug 24;287(35):30024-34. doi: 10.1074/jbc.M112.390682. Epub 2012 Jun 28.
Author Address: Department of Medicine, University of California, San Diego, La Jolla, CA 92039, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1934
Author: Gebai, A., Gorelik, A., Li, Z., Illes, K. and Nagar, B.
Year: 2018
Title: Structural basis for the activation of acid ceramidase
Journal: Nat Commun
Volume: 9
Issue: 1
Pages: 1621
Epub Date: 2018/04/26
Date: Apr 24
Short Title: Structural basis for the activation of acid ceramidase
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-018-03844-2
PMCID: PMC5915598
Accession Number: 29692406
Keywords: Acid Ceramidase/*chemistry/genetics/*metabolism
Binding Sites
Biocatalysis
Ceramides/chemistry/metabolism
Enzyme Activation
Farber Lipogranulomatosis/*enzymology/genetics
Humans
Hydrophobic and Hydrophilic Interactions
Muscular Atrophy, Spinal/*enzymology/genetics
Mutation
Protein Folding
Saposins/genetics/metabolism
Sphingolipids/chemistry/metabolism
Abstract: Acid ceramidase (aCDase, ASAH1) hydrolyzes lysosomal membrane ceramide into sphingosine, the backbone of all sphingolipids, to regulate many cellular processes. Abnormal function of aCDase leads to Farber disease, spinal muscular atrophy with progressive myoclonic epilepsy, and is associated with Alzheimer's, diabetes, and cancer. Here, we present crystal structures of mammalian aCDases in both proenzyme and autocleaved forms. In the proenzyme, the catalytic center is buried and protected from solvent. Autocleavage triggers a conformational change exposing a hydrophobic channel leading to the active site. Substrate modeling suggests distinct catalytic mechanisms for substrate hydrolysis versus autocleavage. A hydrophobic surface surrounding the substrate binding channel appears to be a site of membrane attachment where the enzyme accepts substrates facilitated by the accessory protein, saposin-D. Structural mapping of disease mutations reveals that most would destabilize the protein fold. These results will inform the rational design of aCDase inhibitors and recombinant aCDase for disease therapeutics.
Notes: 2041-1723
Gebai, Ahmad
Gorelik, Alexei
Orcid: 0000-0001-6236-4633
Li, Zixian
Illes, Katalin
Nagar, Bhushan
Orcid: 0000-0001-9869-2110
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2018 Apr 24;9(1):1621. doi: 10.1038/s41467-018-03844-2.
Author Address: Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, QC, H3G 0B1, Canada.
Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, QC, H3G 0B1, Canada. bhushan.nagar@mcgill.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 479
Author: Gejyo, F., Chang, J. L., Bürgi, W., Schmid, K., Offner, G. D., Troxler, R. F., Van Halbeek, H., Dorland, L., Gerwig, G. J. and Vliegenthart, J. F.
Year: 1983
Title: Characterization of the B-chain of human plasma alpha 2HS-glycoprotein. The complete amino acid sequence and primary structure of its heteroglycan
Journal: J Biol Chem
Volume: 258
Issue: 8
Pages: 4966-71
Epub Date: 1983/04/25
Date: Apr 25
Short Title: Characterization of the B-chain of human plasma alpha 2HS-glycoprotein. The complete amino acid sequence and primary structure of its heteroglycan
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6833285
Keywords: Amino Acid Sequence
Blood Proteins/*analysis
Carbohydrate Sequence
Circular Dichroism
Humans
Macromolecular Substances
Molecular Weight
alpha-2-HS-Glycoprotein
Abstract: alpha 2HS-Glycoprotein, a normal human plasma protein, was recently shown to consist of two polypeptide chains. In the present study, we have separated these two chains from one another and have elucidated the complete primary structure of the B-chain. Employing automated Edman degradation, the polypeptide moiety of this chain was shown to consist of 27 amino acid residues with an unequal distribution of the neutral and charged amino acid residues. The first 20 residues are uncharged, whereas the carboxyl-terminal heptapeptide contains all charged residues. Utilizing 500-MHz 1H-NMR spectroscopy, the carbohydrate unit proved to be a trisaccharide consisting of sialic acid, galactose, and N-acetylgalactosamine O-glycosidically linked to serine (residue 6). The structure of the B-chain was found to be as follows. (formula; see text) Thus, the molecular weight of the B-chain is 3386. Evaluation of the polypeptide chain by the procedure of Chou and Fasman (Chou, P.Y., and Fasman, G.D. (1979) Adv. Enzymol. 47, 45-148) predicts that the B-chain has two beta-turns. Thereby, the carbohydrate unit which is linked to the Ser residue located in the first beta-turn appears to be directed away from the protein. The second beta-turn probably includes the Cys residue which links the B- to the A-chain. In agreement with the CD analysis, the B-chain lacks beta-conformation but possesses a short alpha-helical region.
Notes: Gejyo, F
Chang, J L
Bürgi, W
Schmid, K
Offner, G D
Troxler, R F
Van Halbeek, H
Dorland, L
Gerwig, G J
Vliegenthart, J F
GM-10374/GM/NIGMS NIH HHS/United States
HL-13262/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1983 Apr 25;258(8):4966-71.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1471
Author: Gellynck, E., Andressen, K. W., Lintermans, B., Haegeman, G., Levy, F. O., Vanhoenacker, P. and Van Craenenbroeck, K.
Year: 2012
Title: Biochemical and pharmacological study of N-linked glycosylation of the human serotonin 5-HT₇a receptor
Journal: Febs j
Volume: 279
Issue: 11
Pages: 1994-2003
Epub Date: 2012/03/28
Date: Jun
Short Title: Biochemical and pharmacological study of N-linked glycosylation of the human serotonin 5-HT₇a receptor
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/j.1742-4658.2012.08581.x
Accession Number: 22448645
Keywords: Adenylyl Cyclases/metabolism
Asparagine/chemistry/metabolism
Glutamine/chemistry/metabolism
Glycosylation
HEK293 Cells
Humans
*Mutation
Plasmids
Protein Binding
Radioligand Assay
Receptors, Serotonin/chemistry/genetics/*metabolism
Serotonin/analogs & derivatives/metabolism
Transfection
Abstract: The 5-hydroxytryptamine (5-HT)(7(a)) receptor is a G-protein-coupled receptor critically involved in human psychiatric and neurological disorders. In the present study, we evaluate the presence and the functional role of N-glycosylation of the human 5-HT(7) receptor. Western blot analysis of HEK293T cells transiently expressing the 5-HT(7(a)) receptor in the presence of tunicamycin gave rise to a band shift, indicating the existence of an N-glycosylated form of the 5-HT(7(a)) receptor. To further investigate this, we mutated the two predicted N-glycosylation sites (N5Q and N66Q) and compared the molecular mass of the immunoreactive bands with those of the wild-type receptor, indicating that both asparagines were N-glycosylated. The mutant receptors had the same binding affinity for [(3) H]5-CT and the same potency and efficacy with regard to 5-HT-induced activation of adenylyl cyclase. However, there was a reduction in maximal ligand binding for the single and double mutants compared to the wild-type receptor. Next, membrane labelling and immunocytochemical studies demonstrated that the N-glycosylation mutants were expressed at the cell surface. We conclude that N-glycosylation is not important for cell surface expression of the 5-HT(7) receptor.
Notes: 1742-4658
Gellynck, Evelien
Andressen, Kjetil W
Lintermans, Béatrice
Haegeman, Guy
Levy, Finn O
Vanhoenacker, Peter
Van Craenenbroeck, Kathleen
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2012 Jun;279(11):1994-2003. doi: 10.1111/j.1742-4658.2012.08581.x. Epub 2012 Apr 25.
Author Address: Laboratory for Eukaryotic Gene Expression and Signal Transduction (LEGEST), Ghent University, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1466
Author: Geng, F., Zhu, W., Anderson, R. A., Leber, B. and Andrews, D. W.
Year: 2012
Title: Multiple post-translational modifications regulate E-cadherin transport during apoptosis
Journal: J Cell Sci
Volume: 125
Issue: Pt 11
Pages: 2615-25
Epub Date: 2012/03/01
Date: Jun 1
Short Title: Multiple post-translational modifications regulate E-cadherin transport during apoptosis
Alternate Journal: Journal of cell science
ISSN: 0021-9533 (Print)
0021-9533
DOI: 10.1242/jcs.096735
PMCID: PMC3706076
Accession Number: 22375065
Keywords: Acetylglucosamine/metabolism
Animals
Antigens, CD
*Apoptosis/drug effects
Cadherins/chemistry/*metabolism
Cell Membrane/metabolism
Dogs
Endoplasmic Reticulum/drug effects/metabolism
Glycosylation/drug effects
HEK293 Cells
Humans
MCF-7 Cells
Madin Darby Canine Kidney Cells
Models, Biological
Peptides/metabolism
Phosphotransferases (Alcohol Group Acceptor)/metabolism
*Protein Processing, Post-Translational/drug effects
Protein Transport/drug effects
Sequence Deletion/genetics
Stress, Physiological/drug effects
Thapsigargin/pharmacology
Transfection
Abstract: E-cadherin is synthesized as a precursor and then undergoes cleavage by proprotein convertases. This processing is essential for E-cadherin maturation and cell adhesion. Loss of cell adhesion causes detachment-induced apoptosis, which is called anoikis. Anoikis can be inhibited despite loss of cell-matrix interactions by preserving E-cadherin-mediated cell-cell adhesion. Conversely, acute loss of E-cadherin sensitizes cells to apoptosis by unknown post-translational mechanisms. After treatment of breast cancer cells with drugs, we found that two independent modifications of E-cadherin inhibit its cell surface transport. First, O-linked β-N-acetylglucosamine (O-GlcNAc) modification of the cytoplasmic domain retains E-cadherin in the endoplasmic reticulum. Second, incomplete processing by proprotein convertases arrests E-cadherin transport late in the secretory pathway. We demonstrated these E-cadherin modifications (detected by specific lectins and antibodies) do not affect binding to α-catenin, β-catenin or γ-catenin. However, binding of E-cadherin to Type I gamma phosphatidylinositol phosphate kinase (PIPKIγ), a protein required for recruitment of E-cadherin to adhesion sites, was blocked by O-GlcNAc glycosylation (O-GlcNAcylation). Consequently, E-cadherin trafficking to the plasma membrane was inhibited. However, deletion mutants that cannot be O-GlcNAcylated continued to bind PIPKIγ, trafficked to the cell surface and delayed apoptosis, confirming the biological significance of the modifications and PIPKIγ binding. Thus, O-GlyNAcylation of E-cadherin accelerates apoptosis. Furthermore, cell-stress-induced inactivation of proprotein convertases, inhibited E-cadherin maturation, further exacerbating apoptosis. The modifications of E-cadherin by O-GlcNAcylation and lack of pro-region processing represent novel mechanisms for rapid regulation of cell surface transport of E-cadherin in response to intoxication.
Notes: 1477-9137
Geng, Fei
Zhu, Weijia
Anderson, Richard A
Leber, Brian
Andrews, David W
R01 CA104708/CA/NCI NIH HHS/United States
R01 GM057549/GM/NIGMS NIH HHS/United States
FRN 10490/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
J Cell Sci. 2012 Jun 1;125(Pt 11):2615-25. doi: 10.1242/jcs.096735. Epub 2012 Feb 28.
Author Address: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1613
Author: Geng, Y., Bush, M., Mosyak, L., Wang, F. and Fan, Q. R.
Year: 2013
Title: Structural mechanism of ligand activation in human GABA(B) receptor
Journal: Nature
Volume: 504
Issue: 7479
Pages: 254-9
Epub Date: 2013/12/07
Date: Dec 12
Short Title: Structural mechanism of ligand activation in human GABA(B) receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature12725
PMCID: PMC3865065
NIHMSID: NIHMS528775
Accession Number: 24305054
Keywords: Apoproteins/chemistry/metabolism
Binding Sites
Crystallography, X-Ray
Disulfides/chemistry/metabolism
GABA-B Receptor Agonists/pharmacology
GABA-B Receptor Antagonists/pharmacology
Humans
Ligands
Models, Molecular
Protein Multimerization
Protein Structure, Tertiary
Protein Subunits/chemistry/metabolism
Receptors, GABA-B/*chemistry/*metabolism
Substrate Specificity
Abstract: Human GABA(B) (γ-aminobutyric acid class B) receptor is a G-protein-coupled receptor central to inhibitory neurotransmission in the brain. It functions as an obligatory heterodimer of the subunits GBR1 and GBR2. Here we present the crystal structures of a heterodimeric complex between the extracellular domains of GBR1 and GBR2 in the apo, agonist-bound and antagonist-bound forms. The apo and antagonist-bound structures represent the resting state of the receptor; the agonist-bound complex corresponds to the active state. Both subunits adopt an open conformation at rest, and only GBR1 closes on agonist-induced receptor activation. The agonists and antagonists are anchored in the interdomain crevice of GBR1 by an overlapping set of residues. An antagonist confines GBR1 to the open conformation of the inactive state, whereas an agonist induces its domain closure for activation. Our data reveal a unique activation mechanism for GABA(B) receptor that involves the formation of a novel heterodimer interface between subunits.
Notes: 1476-4687
Geng, Yong
Bush, Martin
Mosyak, Lidia
Wang, Feng
Fan, Qing R
R01 GM088454/GM/NIGMS NIH HHS/United States
R01GM088454/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2013 Dec 12;504(7479):254-9. doi: 10.1038/nature12725. Epub 2013 Dec 4.
Author Address: Department of Pharmacology, Columbia University, New York, New York 10032, USA.
1] Department of Pharmacology, Columbia University, New York, New York 10032, USA [2] Department of Pathology & Cell Biology, Columbia University, New York, New York 10032, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1797
Author: Geng, Y., Mosyak, L., Kurinov, I., Zuo, H., Sturchler, E., Cheng, T. C., Subramanyam, P., Brown, A. P., Brennan, S. C., Mun, H. C., Bush, M., Chen, Y., Nguyen, T. X., Cao, B., Chang, D. D., Quick, M., Conigrave, A. D., Colecraft, H. M., McDonald, P. and Fan, Q. R.
Year: 2016
Title: Structural mechanism of ligand activation in human calcium-sensing receptor
Journal: Elife
Volume: 5
Epub Date: 2016/07/20
Date: Jul 19
Short Title: Structural mechanism of ligand activation in human calcium-sensing receptor
Alternate Journal: eLife
ISSN: 2050-084x
DOI: 10.7554/eLife.13662
PMCID: PMC4977154
Accession Number: 27434672
Keywords: Binding Sites
Calcium/*metabolism
Crystallography, X-Ray
Humans
Models, Molecular
Phosphates/metabolism
Protein Binding
Protein Conformation
Protein Multimerization
Receptors, Calcium-Sensing/*agonists/*chemistry
Tryptophan/*chemistry/*metabolism
*amino acids
*biochemistry
*biophysics
*calcium-sensing receptor
*extracellular calcium homeostasis
*extracellular domain structure
*human
*principal agonist
*receptor activation mechanism
*structural biology
Abstract: Human calcium-sensing receptor (CaSR) is a G-protein-coupled receptor (GPCR) that maintains extracellular Ca(2+) homeostasis through the regulation of parathyroid hormone secretion. It functions as a disulfide-tethered homodimer composed of three main domains, the Venus Flytrap module, cysteine-rich domain, and seven-helix transmembrane region. Here, we present the crystal structures of the entire extracellular domain of CaSR in the resting and active conformations. We provide direct evidence that L-amino acids are agonists of the receptor. In the active structure, L-Trp occupies the orthosteric agonist-binding site at the interdomain cleft and is primarily responsible for inducing extracellular domain closure to initiate receptor activation. Our structures reveal multiple binding sites for Ca(2+) and PO4(3-) ions. Both ions are crucial for structural integrity of the receptor. While Ca(2+) ions stabilize the active state, PO4(3-) ions reinforce the inactive conformation. The activation mechanism of CaSR involves the formation of a novel dimer interface between subunits.
Notes: 2050-084x
Geng, Yong
Mosyak, Lidia
Kurinov, Igor
Zuo, Hao
Sturchler, Emmanuel
Cheng, Tat Cheung
Subramanyam, Prakash
Brown, Alice P
Brennan, Sarah C
Mun, Hee-Chang
Bush, Martin
Chen, Yan
Nguyen, Trang X
Cao, Baohua
Chang, Donald D
Quick, Matthias
Conigrave, Arthur D
Colecraft, Henry M
McDonald, Patricia
Fan, Qing R
Orcid: 0000-0002-9330-0963
P41 GM103403/GM/NIGMS NIH HHS/United States
R01 GM112973/GM/NIGMS NIH HHS/United States
S10 RR029205/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Elife. 2016 Jul 19;5:e13662. doi: 10.7554/eLife.13662.
Author Address: Department of Pharmacology, Columbia University, New York, United States.
Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China.
Department of Chemistry and Chemical Biology, Cornell University, Ithaca, United States.
Department of Molecular Therapeutics, The Scripps Translational Science Institute, Jupiter, United States.
Department of Physiology and Cellular Biophysics, Columbia University, New York, United States.
School of Life and Environmental Sciences, University of Sydney, New South Wales, Australia.
Department of Psychiatry, Columbia University, New York, United States.
Department of Pathology and Cell Biology, Columbia University, New York, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1487
Author: Geng, Y., Xiong, D., Mosyak, L., Malito, D. L., Kniazeff, J., Chen, Y., Burmakina, S., Quick, M., Bush, M., Javitch, J. A., Pin, J. P. and Fan, Q. R.
Year: 2012
Title: Structure and functional interaction of the extracellular domain of human GABA(B) receptor GBR2
Journal: Nat Neurosci
Volume: 15
Issue: 7
Pages: 970-8
Epub Date: 2012/06/05
Date: Jun 3
Short Title: Structure and functional interaction of the extracellular domain of human GABA(B) receptor GBR2
Alternate Journal: Nature neuroscience
ISSN: 1097-6256 (Print)
1097-6256
DOI: 10.1038/nn.3133
PMCID: PMC3374333
NIHMSID: NIHMS377158
Accession Number: 22660477
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Extracellular Fluid/*chemistry/drug effects/physiology
HEK293 Cells
Humans
Molecular Sequence Data
Protein Conformation
Protein Structure, Tertiary/genetics
Receptors, GABA-B/*chemistry/genetics/*metabolism
Structure-Activity Relationship
Abstract: Inhibitory neurotransmission is mediated primarily by GABA. The metabotropic GABA(B) receptor is a G protein-coupled receptor central to mammalian brain function. Malfunction of GABA(B) receptor has been implicated in several neurological disorders. GABA(B) receptor functions as a heterodimeric assembly of GBR1 and GBR2 subunits, where GBR1 is responsible for ligand-binding and GBR2 is responsible for G protein coupling. Here we demonstrate that the GBR2 ectodomain directly interacts with the GBR1 ectodomain to increase agonist affinity by selectively stabilizing the agonist-bound conformation of GBR1. We present the crystal structure of the GBR2 ectodomain, which reveals a polar heterodimeric interface. We also identify specific heterodimer contacts from both subunits, and GBR1 residues involved in ligand recognition. Lastly, our structural and functional data indicate that the GBR2 ectodomain adopts a constitutively open conformation, suggesting a structural asymmetry in the active state of GABA(B) receptor that is unique to the GABAergic system.
Notes: 1546-1726
Geng, Yong
Xiong, Dazhi
Mosyak, Lidia
Malito, David L
Kniazeff, Julie
Chen, Yan
Burmakina, Svetlana
Quick, Matthias
Bush, Martin
Javitch, Jonathan A
Pin, Jean-Philippe
Fan, Qing R
K05 DA022413/DA/NIDA NIH HHS/United States
R01 GM088454/GM/NIGMS NIH HHS/United States
R01GM088454/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Neurosci. 2012 Jun 3;15(7):970-8. doi: 10.1038/nn.3133.
Author Address: Department of Pharmacology, Columbia University, New York, New York, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 567
Author: Gerlitz, B., Hassell, T., Vlahos, C. J., Parkinson, J. F., Bang, N. U. and Grinnell, B. W.
Year: 1993
Title: Identification of the predominant glycosaminoglycan-attachment site in soluble recombinant human thrombomodulin: potential regulation of functionality by glycosyltransferase competition for serine474
Journal: Biochem J
Volume: 295 ( Pt 1)
Issue: Pt 1
Pages: 131-40
Epub Date: 1993/10/01
Date: Oct 1
Short Title: Identification of the predominant glycosaminoglycan-attachment site in soluble recombinant human thrombomodulin: potential regulation of functionality by glycosyltransferase competition for serine474
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2950131
PMCID: PMC1134829
Accession Number: 8216207
Keywords: Amino Acid Sequence
Animals
Base Sequence
Binding Sites
Cell Transformation, Viral
Cells, Cultured
Cricetinae
DNA Mutational Analysis
Glycosaminoglycans/*metabolism
Glycosyltransferases/*metabolism
Humans
Molecular Sequence Data
Mutagenesis, Site-Directed
N-Acetylgalactosaminyltransferases/metabolism
Pentosyltransferases/metabolism
Peptide Fragments/genetics
*Protein Processing, Post-Translational
Recombinant Proteins/metabolism
Sequence Alignment
Sequence Analysis
Sequence Homology, Amino Acid
Serine/*metabolism
Thrombomodulin/genetics/*metabolism
Abstract: Thrombomodulin (TM) is an endothelial cell thrombin receptor that converts thrombin from a procoagulant to an anticoagulant enzyme. It has previously been shown that TM is expressed in both a high-M(r) form containing chondroitin sulphate and a low-M(r) form lacking this modification. Site-directed mutagenesis of a soluble human TM derivative (TMD1) was employed to determine the attachment site(s) of this functionally important oligosaccharide on the core protein. Although there are four serine residues within the Ser/Thr-rich domain of TMD1 that might support glycosaminoglycan assembly, our analysis demonstrates that the primary site of attachment is at Ser474, and evidence is presented for low levels of attachment at Ser472. It was possible to improve the overall degree of attachment by mutating Ser472 to glutamic acid (so as to conform Ser474 to the xylosyltransferase acceptor consensus acidic-Gly-Ser-Gly-acidic); however, a significant proportion (approx. 35%) of the total TM still lacked a glycosaminoglycan moiety. Mutants that possess a substitution for Ser474 show an increased mobility of their low-M(r) form on SDS/PAGE compared with native TMD1. Isolation and sequencing of a C-terminal peptide demonstrated that this serine is modified in the low-M(r) form of native TMD1. An apparent 'acceptor consensus overlap' at Ser474 suggests that the mechanism behind the glycosaminoglycan split of TM may involve a competition for substrate between xylosyltransferase and N-acetylgalactosaminyltransferase.
Notes: 1470-8728
Gerlitz, B
Hassell, T
Vlahos, C J
Parkinson, J F
Bang, N U
Grinnell, B W
Comparative Study
Journal Article
Biochem J. 1993 Oct 1;295 ( Pt 1)(Pt 1):131-40. doi: 10.1042/bj2950131.
Author Address: Department of Cardiovascular Research, Lilly Research Laboratories, Indianapolis, IN 46285.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 911
Author: Gewinner, C., Hart, G., Zachara, N., Cole, R., Beisenherz-Huss, C. and Groner, B.
Year: 2004
Title: The coactivator of transcription CREB-binding protein interacts preferentially with the glycosylated form of Stat5
Journal: J Biol Chem
Volume: 279
Issue: 5
Pages: 3563-72
Epub Date: 2003/11/05
Date: Jan 30
Short Title: The coactivator of transcription CREB-binding protein interacts preferentially with the glycosylated form of Stat5
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M306449200
Accession Number: 14597631
Keywords: Acetylglucosamine/pharmacology
Active Transport, Cell Nucleus
Alanine/chemistry
Animals
CREB-Binding Protein
Cell Differentiation
Cell Division
Cell Line
Cell Nucleus/metabolism
Cytoplasm/metabolism
DNA-Binding Proteins/metabolism/*physiology
Dimerization
Epithelial Cells/cytology
Galactosyltransferases/metabolism
Glycosylation
Growth Substances/metabolism
Humans
Insecta
Interferons/metabolism
Lectins/metabolism
Ligands
Luciferases/metabolism
Mass Spectrometry
Microscopy, Fluorescence
*Milk Proteins
Mutation
Nuclear Proteins/chemistry/*metabolism
Phosphorylation
Prolactin/metabolism
Promoter Regions, Genetic
Protein Binding
Protein Structure, Tertiary
Recombinant Proteins/chemistry
STAT5 Transcription Factor
Serine/chemistry
Threonine/chemistry
Trans-Activators/chemistry/*metabolism/*physiology
Transcription, Genetic
Transfection
Tumor Suppressor Proteins
Tyrosine/chemistry
Abstract: The signal transducer and activator of transcription (Stat) gene family comprises seven members with similarities in their domain structure and a common mode of activation. Members of this gene family mediate interferon induction of gene transcription and the response to a large number of growth factors and hormones. Extracellular ligand binding to transmembrane receptors causes the intracellular activation of associated tyrosine kinases, phosphorylation of Stat molecules, dimerization, and translocation to the nucleus. Prolactin-induced phosphorylation of Stat5 is a key event in the development and differentiation of mammary epithelial cells. In addition to the crucial phosphorylation at tyrosine 694, we have identified an O-linked N-acetylglucosamine (O-GlcNAc) as another secondary modification essential for the transcriptional induction by Stat5. This modification was only found on nuclear Stat5 after cytokine activation. Similar observations were made with Stat1, Stat3, and Stat6. Glycosylation of Stat5, however, does not seem to be a prerequisite for nuclear translocation. Mass spectrometric analysis revealed a glycosylated peptide in the N-terminal region of Stat5. Replacement of threonine 92 by an alanine residue (Stat5a-T92A) strongly reduced the prolactin induction of Stat5a glycosylation and abolished transactivation of a target gene promoter. Only the glycosylated form of Stat5 was able to bind the coactivator of transcription CBP, an essential interaction for Stat5-mediated gene transcription.
Notes: Gewinner, Christina
Hart, Gerald
Zachara, Natasha
Cole, Robert
Beisenherz-Huss, Christian
Groner, Bernd
Journal Article
United States
J Biol Chem. 2004 Jan 30;279(5):3563-72. doi: 10.1074/jbc.M306449200. Epub 2003 Nov 3.
Author Address: Georg-Speyer-Haus, Institute for Biomedical Research, Paul-Ehrlich Strasse 42-44, D-60596 Frankfurt am Main, Germany. cgewinne@caregroup.harvard.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 725
Author: Geyer, H., Geyer, R., Odenthal-Schnittler, M. and Schnittler, H. J.
Year: 1999
Title: Characterization of human vascular endothelial cadherin glycans
Journal: Glycobiology
Volume: 9
Issue: 9
Pages: 915-25
Epub Date: 1999/08/26
Date: Sep
Short Title: Characterization of human vascular endothelial cadherin glycans
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/9.9.915
PMCID: PMC7108604
Accession Number: 10460833
Keywords: Actins/isolation & purification
Antigens, CD
Cadherins/*chemistry
Carbohydrate Sequence
Cells, Cultured
Endothelium, Vascular/*chemistry
Glycoproteins/*chemistry
Humans
Intercellular Junctions/chemistry
Molecular Sequence Data
N-Acetylneuraminic Acid/isolation & purification
Oligosaccharides/*chemistry
Umbilical Veins/cytology
Abstract: The glycosylation pattern of human vascular endothelial cadherin (VE-cadherin), purified from cultured human umbilical cord vein endothelial cells, was analyzed. VE-cadherin was metabolically radiolabeled with d-[6-(3)H]glucosamine, isolated by immunoprecipitation, purified by SDS-PAGE and in-gel digested with endoproteinase Asp N. Oligosaccharides were sequentially released from resulting glycopeptides and analyzed by chromatographic profiling. The results revealed that VE-cadherin carries predominantly sialylated diantennary and hybrid-type glycans in addition to some triantennary and high mannose-type species. Highly branched, tetraantennary oligosaccharides were found in trace amounts only. Immunohistochemical labeling of VE-cadherin and sialic acids displayed a codistribution along the intercellular junctions in endothelial cells of human umbilical arteries, veins, and cultured endothelial monolayers. Ca(2+)-depletion, performed on cultured endothelial cells, resulted in a reversible complete disappearance of VE-cadherin and of almost all sialic acid staining from the junctions. Sialidase treatment of whole cells caused a change of VE-cadherin immunofluorescence from a continuous and netlike superstructural organization to a scattered inconsistent one. Hence, cell surface sialic acids might play a role in VE-cadherin organization.
Notes: 1460-2423
Geyer, H
Geyer, R
Odenthal-Schnittler, M
Schnittler, H J
Journal Article
Research Support, Non-U.S. Gov't
Glycobiology. 1999 Sep;9(9):915-25. doi: 10.1093/glycob/9.9.915.
Author Address: Institute of Biochemistry, Justus-Liebig-Universität Giessen, Friedrichstrasse 24, D-35392 Giessen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1070
Author: Ghesquière, B., Van Damme, J., Martens, L., Vandekerckhove, J. and Gevaert, K.
Year: 2006
Title: Proteome-wide characterization of N-glycosylation events by diagonal chromatography
Journal: J Proteome Res
Volume: 5
Issue: 9
Pages: 2438-47
Epub Date: 2006/09/02
Date: Sep
Short Title: Proteome-wide characterization of N-glycosylation events by diagonal chromatography
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr060186m
Accession Number: 16944957
Keywords: Amino Acid Sequence
Animals
Binding Sites
Biomarkers/*analysis
Chromatography/*methods
Glycoproteins/*analysis/genetics
Glycosylation
Humans
Mice
Molecular Sequence Data
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Proteomics/*methods
Abstract: A procedure to map N-glycosylation sites is presented here. It can be applied to purified proteins as well as to highly complex mixtures. The method exploits deglycosylation by PNGase F in a diagonal, reverse-phase chromatographic setup. When applied to 10 microL of mouse serum, affinity-depleted for its three most abundant components, 117 known or predicted sites were mapped in addition to 10 novel sites. Several sites were detected on soluble membrane or receptor components. Our method furthermore senses the nature of glycan structures and can detect differential glycosylation on a given site. These properties--high sensitivity and dependence on glycan imprinting--can be exploited for glycan-biomarker analysis.
Notes: Ghesquière, Bart
Van Damme, Jozef
Martens, Lennart
Vandekerckhove, Joël
Gevaert, Kris
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2006 Sep;5(9):2438-47. doi: 10.1021/pr060186m.
Author Address: Department of Biochemistry and Medical Protein Research, Faculty of Medicine and Health Sciences, Ghent University, A. Baertsoenkaai 3, B9000 Ghent, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1587
Author: Ghosh, K., Mazumder Tagore, D., Anumula, R., Lakshmaiah, B., Kumar, P. P., Singaram, S., Matan, T., Kallipatti, S., Selvam, S., Krishnamurthy, P. and Ramarao, M.
Year: 2013
Title: Crystal structure of rat intestinal alkaline phosphatase--role of crown domain in mammalian alkaline phosphatases
Journal: J Struct Biol
Volume: 184
Issue: 2
Pages: 182-92
Epub Date: 2013/10/01
Date: Nov
Short Title: Crystal structure of rat intestinal alkaline phosphatase--role of crown domain in mammalian alkaline phosphatases
Alternate Journal: Journal of structural biology
ISSN: 1047-8477
DOI: 10.1016/j.jsb.2013.09.017
Accession Number: 24076154
Keywords: Alkaline Phosphatase/*chemistry/genetics
Amino Acid Substitution
Animals
Catalytic Domain
Coordination Complexes/chemistry
Crystallography, X-Ray
Enzyme Stability
Hydrogen-Ion Concentration
Isoenzymes/*chemistry/genetics
Kinetics
Macaca fascicularis
Magnesium/chemistry
Models, Molecular
Mutagenesis, Site-Directed
Protein Structure, Secondary
Rats
Sf9 Cells
Spodoptera
Zinc/chemistry
APs
Crown domain
EcAP
Escherichia coli alkaline phosphatase
Gcap
Glycosylation
Iap
Metal ion requirements
N-acetyl glucosamine
Nag
Plap
Rat intestinal alkaline phosphatase structure
T(m)
Tm
Tnap
alkaline phosphatases
cIAP
cynomolgus monkey intestinal alkaline phosphatase
germ cell alkaline phosphatase
intestinal alkaline phosphatase
melting temperature
p-nitrophenyl-phosphate
pNPP
placental alkaline phosphatase
rIAP
rIAP-Ec
rIAP-Ic
rat intestinal alkaline phosphatase
rat intestinal alkaline phosphatase expressed in E. coli
rat intestinal alkaline phosphatase expressed in Sf9 insect cell
tissue non-specific alkaline phosphatase
triple mutant
Abstract: Intestinal alkaline phosphatases (IAPs) are involved in the cleavage of phosphate prodrugs to liberate the drug for absorption in the intestine. To facilitate in vitro characterization of phosphate prodrugs, we have cloned, expressed, purified and characterized IAPs from rat and cynomolgus monkey (rIAP and cIAP respectively) which are important pre-clinical species for drug metabolism studies. The recombinant rat and monkey enzymes expressed in Sf9 insect cells (IAP-Ic) were found to be glycosylated and active. Expression of rat IAP in Escherichia coli (rIAP-Ec) led to ~200-fold loss of activity that was partially recovered by the addition of external Zn(2+) and Mg(2+) ions. Crystal structures of rIAP-Ec and rIAP-Ic were determined and they provide rationale for the discrepancy in enzyme activities. Rat IAP-Ic retains its activity in presence of both Zn(2+) and Mg(2+) whereas activity of most other alkaline phosphatases (APs) including the cIAP was strongly inhibited by excess Zn(2+). Based on our crystal structure, we hypothesized the residue Q317 in rIAP, present within 7 Å of the Mg(2+) at M3, to be important for this difference in activity. The Q317H rIAP and H317Q cIAP mutants showed reversal in effect of Zn(2+), corroborating the hypothesis. Further analysis of the two structures indicated a close linkage between glycosylation and crown domain stability. A triple mutant of rIAP, where all the three putative N-linked glycosylation sites were mutated showed thermal instability and reduced activity.
Notes: 1095-8657
Ghosh, Kaushik
Mazumder Tagore, Debarati
Anumula, Rushith
Lakshmaiah, Basanth
Kumar, P P B S
Singaram, Senthuran
Matan, Thangavelu
Kallipatti, Sanjith
Selvam, Sabariya
Krishnamurthy, Prasad
Ramarao, Manjunath
Journal Article
United States
J Struct Biol. 2013 Nov;184(2):182-92. doi: 10.1016/j.jsb.2013.09.017. Epub 2013 Sep 27.
Author Address: Applied Biotechnology, Biocon Bristol-Myers Squibb Research and Development Center, Syngene International Ltd., Biocon Park, Bommasandra IV Phase, Jigani Link Road, Bangalore 560099, India. Electronic address: Kaushik.Ghosh@syngeneintl.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 502
Author: Ghosh, P., Amaya, M., Mellins, E. and Wiley, D. C.
Year: 1995
Title: The structure of an intermediate in class II MHC maturation: CLIP bound to HLA-DR3
Journal: Nature
Volume: 378
Issue: 6556
Pages: 457-62
Epub Date: 1995/11/30
Date: Nov 30
Short Title: The structure of an intermediate in class II MHC maturation: CLIP bound to HLA-DR3
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/378457a0
Accession Number: 7477400
Keywords: Amino Acid Sequence
Antigen Presentation
Antigens, Differentiation, B-Lymphocyte/*chemistry/isolation &
purification/metabolism
Binding Sites
Cell Line
Crystallography, X-Ray
HLA-D Antigens/chemistry/metabolism
HLA-DR1 Antigen/chemistry/metabolism
HLA-DR3 Antigen/*chemistry/isolation & purification/metabolism
Half-Life
Hemagglutinin Glycoproteins, Influenza Virus
Hemagglutinins, Viral/chemistry/metabolism
Histocompatibility Antigens Class II/*chemistry/isolation & purification/metabolism
Humans
Hydrogen Bonding
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Conformation
Abstract: A complex between HLA-DR3 and a fragment of invariant chain called CLIP was isolated from a human cell line defective in antigen presentation and its X-ray crystal structure determined. Previous data indicate that this complex is an intermediate in class II histocompatibility maturation, occurring between invariant chain-DR3 and antigenic peptide-DR3 complexes. The structure shows that the CLIP fragment binds to DR3 in a way almost identical to that in which antigenic peptides bind class II histocompatibility glycoproteins. The structure is the substrate for the loading of antigenic peptides by an exchange process catalysed by DM.
Notes: Ghosh, P
Amaya, M
Mellins, E
Wiley, D C
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Nature. 1995 Nov 30;378(6556):457-62. doi: 10.1038/378457a0.
Author Address: Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts 02138, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 995
Author: Giabbai, B., Sidobre, S., Crispin, M. D., Sanchez-Ruìz, Y., Bachi, A., Kronenberg, M., Wilson, I. A. and Degano, M.
Year: 2005
Title: Crystal structure of mouse CD1d bound to the self ligand phosphatidylcholine: a molecular basis for NKT cell activation
Journal: J Immunol
Volume: 175
Issue: 2
Pages: 977-84
Epub Date: 2005/07/09
Date: Jul 15
Short Title: Crystal structure of mouse CD1d bound to the self ligand phosphatidylcholine: a molecular basis for NKT cell activation
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
DOI: 10.4049/jimmunol.175.2.977
Accession Number: 16002697
Keywords: Animals
Antigens, CD1/*chemistry/*immunology/metabolism
Antigens, CD1d
Cell Line
Complementarity Determining Regions/metabolism
Crystallography, X-Ray
Drosophila melanogaster
Killer Cells, Natural/*immunology/metabolism
Ligands
Lymphocyte Activation/*immunology
Mice
Models, Molecular
Phosphatidylcholines/*chemistry/*immunology/metabolism
Protein Binding/immunology
Protein Isoforms/chemistry/immunology/metabolism
Receptors, Antigen, T-Cell, alpha-beta/metabolism
Recombinant Proteins/chemistry/immunology/metabolism
Surface Properties
T-Lymphocyte Subsets/*immunology/metabolism
Abstract: NKT cells are immunoregulatory lymphocytes whose activation is triggered by the recognition of lipid Ags in the context of the CD1d molecules by the TCR. In this study we present the crystal structure to 2.8 A of mouse CD1d bound to phosphatidylcholine. The interactions between the ligand acyl chains and the CD1d molecule define the structural and chemical requirements for the binding of lipid Ags to CD1d. The orientation of the polar headgroup toward the C terminus of the alpha1 helix provides a rationale for the structural basis for the observed Valpha chain bias in invariant NKT cells. The contribution of the ligand to the protein surface suggests a likely mode of recognition of lipid Ags by the NKT cell TCR.
Notes: Giabbai, Barbara
Sidobre, Stèphane
Crispin, M D Max
Sanchez-Ruìz, Yovan
Bachi, Angela
Kronenberg, Mitchell
Wilson, Ian A
Degano, Massimo
AI 45053/AI/NIAID NIH HHS/United States
CA 58896/CA/NCI NIH HHS/United States
GM 62116/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Immunol. 2005 Jul 15;175(2):977-84. doi: 10.4049/jimmunol.175.2.977.
Author Address: Biocrystallography Unit and Mass Spectrometry Unit, DIBIT San Raffaele Scientific Institute, via Olgettina 58, 20132 Milan, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1236
Author: Gil, G. C., Velander, W. H. and Van Cott, K. E.
Year: 2009
Title: N-glycosylation microheterogeneity and site occupancy of an Asn-X-Cys sequon in plasma-derived and recombinant protein C
Journal: Proteomics
Volume: 9
Issue: 9
Pages: 2555-67
Epub Date: 2009/04/04
Date: May
Short Title: N-glycosylation microheterogeneity and site occupancy of an Asn-X-Cys sequon in plasma-derived and recombinant protein C
Alternate Journal: Proteomics
ISSN: 1615-9853 (Print)
1615-9853
DOI: 10.1002/pmic.200800775
PMCID: PMC2785108
NIHMSID: NIHMS146826 Progenetics LLC, a company that develops transgenic animal bioreactors for production of therapeutic proteins.
Accession Number: 19343721
Keywords: Animals
Animals, Genetically Modified
Asparagine/chemistry/metabolism
Bioreactors
Chromatography, High Pressure Liquid
Cysteine/chemistry/metabolism
Epithelium/chemistry
Glycopeptides/*chemistry/genetics/metabolism
Glycosylation
Humans
Mammary Glands, Animal/cytology
Mass Spectrometry
N-Acetylneuraminic Acid/chemistry
Plasma/chemistry
Protein C/*chemistry/genetics/metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Swine
Abstract: Human protein C (hPC) is glycosylated at three Asn-X-Ser/Thr and one atypical Asn-X-Cys sequons. We have characterized the micro- and macro-heterogeneity of plasma-derived hPC and compared the glycosylation features with recombinant protein C (tg-PC) produced in a transgenic pig bioreactor from two animals having approximately tenfold different expression levels. The N-glycans of hPC are complex di- and tri-sialylated structures, and we measured 78% site occupancy at Asn-329 (the Asn-X-Cys sequon). The N-glycans of tg-PC are complex sialylated structures, but less branched and partially sialylated. The porcine mammary epithelial cells glycosylate the Asn-X-Cys sequon with a similar efficiency as human hepatocytes even at these high expression levels, and site occupancy at this sequon was not affected by expression level. A distinct bias for particular structures was present at each of the four glycosylation sites for both hPC and tg-PC. Interestingly, glycans with GalNAc in the antennae were predominant at the Asn-329 site. The N-glycan structures found for tg-PC are very similar to those reported for a recombinant Factor IX produced in transgenic pig milk, and similar to the endogenous milk protein lactoferrin, which may indicate that N-glycan processing in the porcine mammary epithelial cells is more uniform than in other tissues.
Notes: 1615-9861
Gil, Geun-Cheol
Velander, William H
Van Cott, Kevin E
R01 HL078944/HL/NHLBI NIH HHS/United States
R01 HL078944-01/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proteomics. 2009 May;9(9):2555-67. doi: 10.1002/pmic.200800775.
Author Address: Department of Chemical and Biomolecular Engineering, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1685
Author: Glukhova, A., Hinkovska-Galcheva, V., Kelly, R., Abe, A., Shayman, J. A. and Tesmer, J. J.
Year: 2015
Title: Structure and function of lysosomal phospholipase A2 and lecithin:cholesterol acyltransferase
Journal: Nat Commun
Volume: 6
Pages: 6250
Epub Date: 2015/03/03
Date: Mar 2
Short Title: Structure and function of lysosomal phospholipase A2 and lecithin:cholesterol acyltransferase
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms7250
PMCID: PMC4397983
NIHMSID: NIHMS654078
Accession Number: 25727495
Keywords: Amino Acid Sequence
Cholesterol/metabolism
Chromatography, Gel
Crystallography, X-Ray
Humans
Lysosomes/*enzymology
*Models, Molecular
Molecular Sequence Data
Phosphatidylcholine-Sterol O-Acyltransferase/*chemistry/*metabolism
Phospholipases A2/*chemistry/*metabolism
Protein Conformation
Pulmonary Surfactants/metabolism
Sequence Alignment
Abstract: Lysosomal phospholipase A2 (LPLA2) and lecithin:cholesterol acyltransferase (LCAT) belong to a structurally uncharacterized family of key lipid-metabolizing enzymes responsible for lung surfactant catabolism and for reverse cholesterol transport, respectively. Whereas LPLA2 is predicted to underlie the development of drug-induced phospholipidosis, somatic mutations in LCAT cause fish eye disease and familial LCAT deficiency. Here we describe several high-resolution crystal structures of human LPLA2 and a low-resolution structure of LCAT that confirms its close structural relationship to LPLA2. Insertions in the α/β hydrolase core of LPLA2 form domains that are responsible for membrane interaction and binding the acyl chains and head groups of phospholipid substrates. The LCAT structure suggests the molecular basis underlying human disease for most of the known LCAT missense mutations, and paves the way for rational development of new therapeutics to treat LCAT deficiency, atherosclerosis and acute coronary syndrome.
Notes: 2041-1723
Glukhova, Alisa
Hinkovska-Galcheva, Vania
Kelly, Robert
Abe, Akira
Shayman, James A
Tesmer, John J G
R01 AR056991/AR/NIAMS NIH HHS/United States
HL086865/HL/NHLBI NIH HHS/United States
AR056991/AR/NIAMS NIH HHS/United States
I01 BX002021/BX/BLRD VA/United States
R01 HL086865/HL/NHLBI NIH HHS/United States
R01 HL122416/HL/NHLBI NIH HHS/United States
R01 HD076004/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Nat Commun. 2015 Mar 2;6:6250. doi: 10.1038/ncomms7250.
Author Address: 1] Life Sciences Institute and the Departments of Pharmacology and Biological Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA [2] Program in Chemical Biology, University of Michigan, Ann Arbor, Michigan 48109, USA.
Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan 48109, USA.
1] Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan 48109, USA [2] Department of Ophthalmology, School of Medicine, Sapporo Medical University, Sapporo, Hokkaido 060-8543, Japan.
Life Sciences Institute and the Departments of Pharmacology and Biological Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1060
Author: Goettsch, S., Badea, R. A., Mueller, J. W., Wotzlaw, C., Schoelermann, B., Schulz, L., Rabiller, M., Bayer, P. and Hartmann-Fatu, C.
Year: 2006
Title: Human TPST1 transmembrane domain triggers enzyme dimerisation and localisation to the Golgi compartment
Journal: J Mol Biol
Volume: 361
Issue: 3
Pages: 436-49
Epub Date: 2006/07/25
Date: Aug 18
Short Title: Human TPST1 transmembrane domain triggers enzyme dimerisation and localisation to the Golgi compartment
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2006.06.023
Accession Number: 16859706
Keywords: Amino Acid Sequence
Animals
Asparagine/genetics/metabolism
Autoantigens/metabolism
COS Cells
Chlorocebus aethiops
Dimerization
Fluorescence Resonance Energy Transfer
Glycosylation
Golgi Apparatus/*metabolism
Golgi Matrix Proteins
HeLa Cells
Humans
Intracellular Membranes/metabolism
Molecular Sequence Data
Mutation
Oligosaccharides/metabolism
Protein Binding
Protein Processing, Post-Translational
Protein Structure, Tertiary
Sulfotransferases/genetics/*physiology
Abstract: TPST1 is a human tyrosylprotein sulfotransferase that uses 3'phosphoadenosine-5'phosphosulfate (PAPS) to transfer the sulfate moiety to proteins predominantly designated for secretion. To achieve a general understanding of the cellular role of human tyrosine-directed sulfotransferases, we investigated targeting, structure and posttranslational modification of TPST1. Golgi localisation of the enzyme in COS-7 and HeLa cells was visualised by fluorescence imaging techniques. PNGase treatment and mutational studies determined that TPST1 bears N-linked glycosyl residues exclusively at position Asn60 and Asn262. By alanine mutation of these asparagine residues, we could determine that the N-linked oligosaccharides do not have an influence on Golgi retention of TPST1. In concert with N and C-terminal flanking residues, the transmembrane domain of TPST1 was determined to act in targeting and retention of the enzyme to the trans-Golgi compartment. This domain exhibits a pronounced secondary structure in a lipid environment. Further in vivo FRET studies using the transmembrane domain suggest that the human tyrosylprotein sulfotransferase may be functional as homodimer/oligomer in the trans-Golgi compartment.
Notes: Goettsch, Sandra
Badea, Rodica A
Mueller, Jonathan W
Wotzlaw, Christoph
Schoelermann, Beate
Schulz, Lars
Rabiller, Matthias
Bayer, Peter
Hartmann-Fatu, Cristina
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2006 Aug 18;361(3):436-49. doi: 10.1016/j.jmb.2006.06.023. Epub 2006 Jun 30.
Author Address: Department of Structural and Medicinal Biochemistry, University of Duisburg-Essen and Centre for Medicinal Biotechnology, Universitätsstr. 2-5, 45117 Essen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 738
Author: Goetz, D. H., Willie, S. T., Armen, R. S., Bratt, T., Borregaard, N. and Strong, R. K.
Year: 2000
Title: Ligand preference inferred from the structure of neutrophil gelatinase associated lipocalin
Journal: Biochemistry
Volume: 39
Issue: 8
Pages: 1935-41
Epub Date: 2000/02/24
Date: Feb 29
Short Title: Ligand preference inferred from the structure of neutrophil gelatinase associated lipocalin
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi992215v
Accession Number: 10684642
Keywords: *Acute-Phase Proteins
Binding Sites
Carrier Proteins/*chemistry/metabolism
Crystallography, X-Ray
Ligands
Lipocalin-2
Lipocalins
Models, Molecular
*Oncogene Proteins
Protein Binding
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Proto-Oncogene Proteins
Abstract: Neutrophil gelatinase associated lipocalin (NGAL), a constituent of neutrophil granules, is a member of the lipocalin family of binding proteins. NGAL can also be highly induced in epithelial cells in both inflammatory and neoplastic colorectal disease. NGAL is proposed to mediate inflammatory responses by sequestering neutrophil chemoattractants, particularly N-formylated tripeptides and possibly leukotriene B(4) and platelet activating factor. The crystal structures of NGAL display a typical lipocalin fold, albeit with an unusually large and atypically polar binding site, or calyx. The fold of NGAL is most similar to the epididymal retinoic acid-binding protein, another lipocalin, though the overall architecture of the calyces are very different. The crystal structures also reveal either sulfate ions or an adventitiously copurified fatty acid bound in the binding site. Neither ligand is displaced by added N-formylated tripeptides. The size, shape, and character of the NGAL calyx, as well as the low relative affinity for N-formylated tripeptides, suggest that neither the copurified fatty acid nor any of the proposed ligands are likely to be the preferred ligand of this protein. Comparisons between the crystal structures and the recently reported solution structure of NGAL reveal significant differences, in terms of both the details of the structure and the overall flexibility of the fold.
Notes: Goetz, D H
Willie, S T
Armen, R S
Bratt, T
Borregaard, N
Strong, R K
AI42200/AI/NIAID NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2000 Feb 29;39(8):1935-41. doi: 10.1021/bi992215v.
Author Address: Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 764
Author: Gohlke, M., Mach, U., Nuck, R., Zimmermann-Kordmann, M., Grunow, D., Fieger, C., Volz, B., Tauber, R., Petri, T., Debus, N. and Reutter, W.
Year: 2000
Title: Carbohydrate structures of soluble human L-selectin recombinantly expressed in baby-hamster kidney cells
Journal: Biotechnol Appl Biochem
Volume: 32
Issue: 1
Pages: 41-51
Epub Date: 2000/08/05
Date: Aug
Short Title: Carbohydrate structures of soluble human L-selectin recombinantly expressed in baby-hamster kidney cells
Alternate Journal: Biotechnology and applied biochemistry
ISSN: 0885-4513 (Print)
0885-4513
DOI: 10.1042/ba19990105
Accession Number: 10918037
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/*genetics
Cell Line
Cricetinae
Humans
Kidney/*cytology
L-Selectin/chemistry/*genetics/metabolism
Oligosaccharides/*analysis/chemistry
Polysaccharides/chemistry/isolation & purification
Recombinant Proteins/genetics/metabolism
Sequence Analysis
Solubility
Abstract: A soluble form of L-selectin was recombinantly produced, which might be an effective therapeutic agent in inflammatory disorders, acting as an inhibitor for leucocyte endothelium adhesion. In the present study the oligosaccharide structures of soluble human L-selectin, recombinantly expressed in baby-hamster kidney cells, were determined. The N-linked glycans were enzymically released and fluorescently labelled with 2-aminobenzamide. Sialylation of the N-glycans was analysed by anion-exchange chromatography followed by rechromatography of the resulting fractions on amino-phase HPLC after release of the sialic acid residues. Desialylated oligosaccharides were separated using two-dimensional HPLC and characterized by digestion with exoglycosidases and MS. More than 30 oligosaccharide structures representing at least 95% of the overall glycosylation of this protein were determined. The results revealed that recombinant soluble human L-selectin carries bi-, tri- and tetra-antennary sugar chains, which are fucosylated on the innermost residue of N-acetylglucosamine. The number of sialic acid residues linked to these glycans ranges from 0 (neutral glycans) to 4 (tetrasialylated oligosaccharides). The sialic acid is found exclusively in the alpha 2-3 linkage to galactose. In addition to the main glycans, different minor structures containing terminal N-acetylgalactosamine, or the H (O) blood-group determinant were also identified. O-Glycosylation of mucin-type sugar chains was not detected in recombinant soluble human L-selectin.
Notes: Gohlke, M
Mach, U
Nuck, R
Zimmermann-Kordmann, M
Grunow, D
Fieger, C
Volz, B
Tauber, R
Petri, T
Debus, N
Reutter, W
Journal Article
Research Support, Non-U.S. Gov't
United States
Biotechnol Appl Biochem. 2000 Aug;32(1):41-51. doi: 10.1042/ba19990105.
Author Address: Institut für Diagnostikforschung GmbH an der Freien Universität Berlin, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1132
Author: Golks, A., Tran, T. T., Goetschy, J. F. and Guerini, D.
Year: 2007
Title: Requirement for O-linked N-acetylglucosaminyltransferase in lymphocytes activation
Journal: Embo j
Volume: 26
Issue: 20
Pages: 4368-79
Epub Date: 2007/09/21
Date: Oct 17
Short Title: Requirement for O-linked N-acetylglucosaminyltransferase in lymphocytes activation
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7601845
PMCID: PMC2034663
Accession Number: 17882263
Keywords: Alternative Splicing
Antigens, CD/biosynthesis
Antigens, Differentiation, T-Lymphocyte/biosynthesis
B-Lymphocytes/metabolism
Glycosylation
Humans
Interleukin-2/metabolism
Lectins, C-Type
*Lymphocyte Activation
Lymphocytes/*metabolism
Models, Biological
N-Acetylglucosaminyltransferases/chemistry/*physiology
NF-kappa B/metabolism
NFATC Transcription Factors/metabolism
Protein Processing, Post-Translational
RNA, Small Interfering/metabolism
T-Lymphocytes/metabolism
Abstract: The dynamic modification of nuclear and cytoplasmic proteins with O-linked beta-N-acetylglucosamine (O-GlcNAc) by the O-linked N-acetylglucosaminyltransferase (OGT) is a regulatory post-translational modification that is responsive to various stimuli. Here, we demonstrate that OGT is a central factor for T- and B-lymphocytes activation. SiRNA-mediated knockdown of OGT in T cells leads to an impaired activation of the transcription factors NFAT and NFkappaB. This results in a reduction of IL-2 production consistent with prevention of T-cell activation. OGT is also required for the early activation of B cells mediated by stimulation of the B-cell receptor. Mechanistically, we demonstrate that NFkappaB as well as NFAT are glycosylated with O-GlcNAc after direct binding to OGT. Moreover, kinetic experiments show that O-GlcNAc modification prominently increased shortly after activation of lymphoid cells and it might be required for nuclear translocation of the transcription factors NFkappaB and NFAT.
Notes: 1460-2075
Golks, Alexander
Tran, Thi-Thanh Thao
Goetschy, Jean Francois
Guerini, Danilo
Journal Article
EMBO J. 2007 Oct 17;26(20):4368-79. doi: 10.1038/sj.emboj.7601845. Epub 2007 Sep 20.
Author Address: Autoimmunity and Transplantation, Novartis Pharma AG, Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1338
Author: Gomez, A., Nekvindova, J., Travica, S., Lee, M. Y., Johansson, I., Edler, D., Mkrtchian, S. and Ingelman-Sundberg, M.
Year: 2010
Title: Colorectal cancer-specific cytochrome P450 2W1: intracellular localization, glycosylation, and catalytic activity
Journal: Mol Pharmacol
Volume: 78
Issue: 6
Pages: 1004-11
Epub Date: 2010/09/02
Date: Dec
Short Title: Colorectal cancer-specific cytochrome P450 2W1: intracellular localization, glycosylation, and catalytic activity
Alternate Journal: Molecular pharmacology
ISSN: 0026-895x
DOI: 10.1124/mol.110.067652
Accession Number: 20805301
Keywords: Aflatoxin B1/metabolism/toxicity
Asparagine/metabolism
Biotransformation
Caco-2 Cells
Catalytic Domain/*physiology
Cell Line, Tumor
Colorectal Neoplasms/*enzymology/metabolism
Cytochrome P-450 Enzyme System/biosynthesis/genetics/*metabolism
Cytochrome P450 Family 2
Electron-Transferring Flavoproteins/metabolism
Glycosylation
HEK293 Cells
Hep G2 Cells
Humans
Indoles/metabolism
Intracellular Fluid/*enzymology/metabolism
Abstract: Cytochrome P450 2W1 (CYP2W1) is expressed at high levels in colorectal cancer cells. Moreover, we have shown previously that a higher tumor expression is associated with less survival. In this study, we characterize post-translational modification, inverted endoplasmic reticulum (ER) topology, and catalytic activity of CYP2W1. The analysis of colorectal normal and cancer tissues and CYP2W1 overexpressing human embryonic kidney (HEK) 293 cells showed that a fraction of CYP2W1 is modified by N-glycosylation. Bioinformatic analysis identified Asn177 as the only possible glycosylation site of CYP2W1, which was supported by the inability of an N177A mutant to be glycosylated in HEK 293 cells. Analysis of the membrane topology indicated that unlike other cytochromes P450, CYP2W1 in HEK 293-transfected cells and in nontransfected Caco2TC7 and HepG2 cells is oriented toward the lumen of the ER, a topology making CYP2W1 available to the ER glycosylation machinery. Immunofluorescence microscopy and cell surface biotinylation experiments revealed approximately 8% of the CYP2W1 on the cell surface. Despite the reverse orientation of CYP2W1 in the ER membrane, apparently making functional interactions with NADPH-cytochrome P450 reductase impossible, CYP2W1 in HEK 293 cells was active in the metabolism of indoline substrates and was able to activate aflatoxin B1 into cytotoxic products. The study identifies for the first time a cytochrome P450 enzyme with a luminal ER orientation and still retaining catalytic activity. Together, these results suggest the possibility of using CYP2W1 as a drug target in the treatment of colon cancer using antibodies and/or specific CYP2W1 activated prodrugs.
Notes: 1521-0111
Gomez, Alvin
Nekvindova, Jana
Travica, Sandra
Lee, Mi-Young
Johansson, Inger
Edler, David
Mkrtchian, Souren
Ingelman-Sundberg, Magnus
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Mol Pharmacol. 2010 Dec;78(6):1004-11. doi: 10.1124/mol.110.067652. Epub 2010 Aug 30.
Author Address: Section of Pharmacogenetics, Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 840
Author: Gong, Q., Anderson, C. L., January, C. T. and Zhou, Z.
Year: 2002
Title: Role of glycosylation in cell surface expression and stability of HERG potassium channels
Journal: Am J Physiol Heart Circ Physiol
Volume: 283
Issue: 1
Pages: H77-84
Epub Date: 2002/06/14
Date: Jul
Short Title: Role of glycosylation in cell surface expression and stability of HERG potassium channels
Alternate Journal: American journal of physiology. Heart and circulatory physiology
ISSN: 0363-6135 (Print)
0363-6135
DOI: 10.1152/ajpheart.00008.2002
Accession Number: 12063277
Keywords: Amino Acid Sequence
Binding Sites/genetics/physiology
Blotting, Western
*Cation Transport Proteins
Cell Line
Cell Membrane/*metabolism
*DNA-Binding Proteins
ERG1 Potassium Channel
Ether-A-Go-Go Potassium Channels
Glycosylation/drug effects
Green Fluorescent Proteins
Humans
Kidney/cytology/drug effects/metabolism
Luminescent Proteins/genetics
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Patch-Clamp Techniques
Potassium Channels/drug effects/genetics/*metabolism
*Potassium Channels, Voltage-Gated
Protein Transport/physiology
Recombinant Fusion Proteins/genetics/metabolism
*Trans-Activators
Transcriptional Regulator ERG
Transfection
Tunicamycin/pharmacology
Abstract: The human ether-à-go-go-related gene (HERG) encodes the pore-forming subunit of the rapidly activating delayed rectifier potassium channel in the heart. We previously showed that HERG channel protein is modified by N-linked glycosylation. HERG protein sequence contains two extracellular consensus sites for N-linked glycosylation (N598, N629). In this study, we used the approaches of site-directed mutagenesis and biochemical modification to inhibit N-linked glycosylation and studied the role of glycosylation in the cell surface expression and turnover of HERG channels. Our results show that N598 is the only site for N-linked glycosylation and that glycosylation is not required for the cell surface expression of functional HERG channels. In contrast, N629 is not used for glycosylation, but mutation of this site (N629Q) causes a protein trafficking defect, which results in its intracellular retention. Pulse-chase experiments show that the turnover rate of nonglycosylated HERG channel is faster than that of the glycosylated form, suggesting that N-linked glycosylation plays an important role in HERG channel stability.
Notes: Gong, Qiuming
Anderson, Corey L
January, Craig T
Zhou, Zhengfeng
R01 HL068854/HL/NHLBI NIH HHS/United States
R01 HL068854-01/HL/NHLBI NIH HHS/United States
HL-60723/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Am J Physiol Heart Circ Physiol. 2002 Jul;283(1):H77-84. doi: 10.1152/ajpheart.00008.2002.
Author Address: Division of Molecular Medicine, Department of Medicine, Oregon Health and Science University, Portland, Oregon 97201, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1784
Author: Gong, X., Qian, H., Zhou, X., Wu, J., Wan, T., Cao, P., Huang, W., Zhao, X., Wang, X., Wang, P., Shi, Y., Gao, G. F., Zhou, Q. and Yan, N.
Year: 2016
Title: Structural Insights into the Niemann-Pick C1 (NPC1)-Mediated Cholesterol Transfer and Ebola Infection
Journal: Cell
Volume: 165
Issue: 6
Pages: 1467-1478
Epub Date: 2016/05/31
Date: Jun 2
Short Title: Structural Insights into the Niemann-Pick C1 (NPC1)-Mediated Cholesterol Transfer and Ebola Infection
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2016.05.022
PMCID: PMC7111323
Accession Number: 27238017
Keywords: Amino Acid Sequence
Carrier Proteins/chemistry/*metabolism/ultrastructure
Cholesterol/*metabolism
Cryoelectron Microscopy
Ebolavirus/*metabolism
Glycoproteins/chemistry/metabolism
Hemorrhagic Fever, Ebola/*metabolism/virology
Humans
Intracellular Signaling Peptides and Proteins
Membrane Glycoproteins/chemistry/*metabolism/ultrastructure
Models, Molecular
Niemann-Pick Diseases/metabolism
Protein Conformation
Structure-Activity Relationship
Vesicular Transport Proteins
Viral Envelope Proteins/chemistry/*metabolism/ultrastructure
Abstract: Niemann-Pick disease type C (NPC) is associated with mutations in NPC1 and NPC2, whose gene products are key players in the endosomal/lysosomal egress of low-density lipoprotein-derived cholesterol. NPC1 is also the intracellular receptor for Ebola virus (EBOV). Here, we present a 4.4 Å structure of full-length human NPC1 and a low-resolution reconstruction of NPC1 in complex with the cleaved glycoprotein (GPcl) of EBOV, both determined by single-particle electron cryomicroscopy. NPC1 contains 13 transmembrane segments (TMs) and three distinct lumenal domains A (also designated NTD), C, and I. TMs 2-13 exhibit a typical resistance-nodulation-cell division fold, among which TMs 3-7 constitute the sterol-sensing domain conserved in several proteins involved in cholesterol metabolism and signaling. A trimeric EBOV-GPcl binds to one NPC1 monomer through the domain C. Our structural and biochemical characterizations provide an important framework for mechanistic understanding of NPC1-mediated intracellular cholesterol trafficking and Ebola virus infection.
Notes: 1097-4172
Gong, Xin
Qian, Hongwu
Zhou, Xinhui
Wu, Jianping
Wan, Tao
Cao, Pingping
Huang, Weiyun
Zhao, Xin
Wang, Xudong
Wang, Peiyi
Shi, Yi
Gao, George F
Zhou, Qiang
Yan, Nieng
HHMI/HHMI/United States
Journal Article
Research Support, Non-U.S. Gov't
Cell. 2016 Jun 2;165(6):1467-1478. doi: 10.1016/j.cell.2016.05.022. Epub 2016 May 26.
Author Address: State Key Laboratory of Membrane Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China; Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China; Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China.
CAS Key Laboratory of Pathogenic Microbiology and Immunology (CASPMI), Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
Faculty of Biological Sciences, University of Leeds, Leeds LS2 9JT, UK.
CAS Key Laboratory of Pathogenic Microbiology and Immunology (CASPMI), Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China; Savaid Medical School, University of Chinese Academy of Sciences, Beijing 100049, China.
State Key Laboratory of Membrane Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China; Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China; Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China. Electronic address: zhouqiang00@tsinghua.org.cn.
State Key Laboratory of Membrane Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China; Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China; Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China. Electronic address: nyan@tsinghua.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 841
Author: Gonzalez de Peredo, A., Klein, D., Macek, B., Hess, D., Peter-Katalinic, J. and Hofsteenge, J.
Year: 2002
Title: C-mannosylation and o-fucosylation of thrombospondin type 1 repeats
Journal: Mol Cell Proteomics
Volume: 1
Issue: 1
Pages: 11-8
Epub Date: 2002/07/04
Date: Jan
Short Title: C-mannosylation and o-fucosylation of thrombospondin type 1 repeats
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.m100011-mcp200
Accession Number: 12096136
Keywords: Amino Acid Motifs
Extracellular Matrix Proteins
Fucose/*metabolism
Glycopeptides/*chemistry
Glycosylation
*Growth Substances
Humans
Mannose/*metabolism
Mass Spectrometry/*methods
Neural Cell Adhesion Molecules/*metabolism
*Peptides
Protein Processing, Post-Translational
Recombinant Fusion Proteins/chemistry
Thrombospondin 1/chemistry/*metabolism
Abstract: The final chemical structure of a newly synthesized protein is often only attained after further covalent modification. Ideally, a comprehensive proteome analysis includes this aspect, a task that is complicated by our incomplete knowledge of the range of possible modifications and often by the lack of suitable analysis methods. Here we present two recently discovered, unusual forms of protein glycosylation, i.e. C-mannosylation and O-fucosylation. Their analysis by a combined mass spectrometric approach is illustrated with peptides from the thrombospondin type 1 repeats (TSRs) of the recombinant axonal guidance protein F-spondin. Nano-electrospray ionization tandem-mass spectrometry of isolated peptides showed that eight of ten Trp residues in the TSRs of F-spondin are C-mannosylated. O-Fucosylation sites were determined by a recently established nano-electrospray ionization quadrupole time-of-flight tandem-mass spectrometry approach. Four of five TSRs carry the disaccharide Hex-dHex-O-Ser/Thr in close proximity to the C-mannosylation sites. In analogy to thrombospondin-1, we assume this to be Glc-Fuc-O-Ser/Thr. Our current knowledge of these glycosylations will be discussed.
Notes: Gonzalez de Peredo, Anne
Klein, Dominique
Macek, Boris
Hess, Daniel
Peter-Katalinic, Jasna
Hofsteenge, Jan
Journal Article
United States
Mol Cell Proteomics. 2002 Jan;1(1):11-8. doi: 10.1074/mcp.m100011-mcp200.
Author Address: Friedrich Miescher Institute for Biomedical Research, Novartis Research Foundation, Maulbeerstrasse 66, CH-4058 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1779
Author: Goodman, K. M., Rubinstein, R., Thu, C. A., Bahna, F., Mannepalli, S., Ahlsén, G., Rittenhouse, C., Maniatis, T., Honig, B. and Shapiro, L.
Year: 2016
Title: Structural Basis of Diverse Homophilic Recognition by Clustered α- and β-Protocadherins
Journal: Neuron
Volume: 90
Issue: 4
Pages: 709-23
Epub Date: 2016/05/11
Date: May 18
Short Title: Structural Basis of Diverse Homophilic Recognition by Clustered α- and β-Protocadherins
Alternate Journal: Neuron
ISSN: 0896-6273 (Print)
0896-6273
DOI: 10.1016/j.neuron.2016.04.004
PMCID: PMC4873334
NIHMSID: NIHMS776597
Accession Number: 27161523
Keywords: Amino Acid Sequence/*physiology
Cadherins/*chemistry/*metabolism
Cells, Cultured
Humans
Nerve Net/metabolism
Neurons/*metabolism
Protein Binding/physiology
Protein Interaction Domains and Motifs
Protein Isoforms/metabolism
Abstract: Clustered protocadherin proteins (α-, β-, and γ-Pcdhs) provide a high level of cell-surface diversity to individual vertebrate neurons, engaging in highly specific homophilic interactions to mediate important roles in mammalian neural circuit development. How Pcdhs bind homophilically through their extracellular cadherin (EC) domains among dozens of highly similar isoforms has not been determined. Here, we report crystal structures for extracellular regions from four mouse Pcdh isoforms (α4, α7, β6, and β8), revealing a canonical head-to-tail interaction mode for homophilic trans dimers comprising primary intermolecular EC1:EC4 and EC2:EC3 interactions. A subset of trans interface residues exhibit isoform-specific conservation, suggesting roles in recognition specificity. Mutation of these residues, along with trans-interacting partner residues, altered the specificities of Pcdh interactions. Together, these data show how sequence variation among Pcdh isoforms encodes their diverse strict homophilic recognition specificities, which are required for their key roles in neural circuit assembly.
Notes: 1097-4199
Goodman, Kerry Marie
Rubinstein, Rotem
Thu, Chan Aye
Bahna, Fabiana
Mannepalli, Seetha
Ahlsén, Göran
Rittenhouse, Chelsea
Maniatis, Tom
Honig, Barry
Shapiro, Lawrence
P41 GM103403/GM/NIGMS NIH HHS/United States
R01 GM062270/GM/NIGMS NIH HHS/United States
R01 GM107571/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Neuron. 2016 May 18;90(4):709-23. doi: 10.1016/j.neuron.2016.04.004. Epub 2016 May 5.
Author Address: Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA; Department of Systems Biology, Columbia University, New York, NY 10032, USA.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA; Howard Hughes Medical Institute, Columbia University, New York, NY 10032, USA.
Department of Systems Biology, Columbia University, New York, NY 10032, USA; Howard Hughes Medical Institute, Columbia University, New York, NY 10032, USA.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA; Zuckerman Mind Brain and Behavior Institute, Columbia University, New York, NY 10032, USA.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA; Department of Systems Biology, Columbia University, New York, NY 10032, USA; Howard Hughes Medical Institute, Columbia University, New York, NY 10032, USA; Department of Medicine, Columbia University, New York, NY 10032, USA; Zuckerman Mind Brain and Behavior Institute, Columbia University, New York, NY 10032, USA. Electronic address: bh6@cumc.columbia.edu.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA; Department of Systems Biology, Columbia University, New York, NY 10032, USA; Zuckerman Mind Brain and Behavior Institute, Columbia University, New York, NY 10032, USA. Electronic address: shapiro@convex.hhmi.columbia.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 294
Author: Gooley, A. A., Classon, B. J., Marschalek, R. and Williams, K. L.
Year: 1991
Title: Glycosylation sites identified by detection of glycosylated amino acids released from Edman degradation: the identification of Xaa-Pro-Xaa-Xaa as a motif for Thr-O-glycosylation
Journal: Biochem Biophys Res Commun
Volume: 178
Issue: 3
Pages: 1194-201
Epub Date: 1991/08/15
Date: Aug 15
Short Title: Glycosylation sites identified by detection of glycosylated amino acids released from Edman degradation: the identification of Xaa-Pro-Xaa-Xaa as a motif for Thr-O-glycosylation
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(91)91019-9
Accession Number: 1908233
Keywords: Amino Acid Sequence
Animals
Cell Line
Glycopeptides/*isolation & purification
Glycosylation
Humans
Molecular Sequence Data
Peptide Fragments/isolation & purification
Proline
Proteins/*genetics
Rats
Serine
Threonine
Transfection
Abstract: Here we report the use of automated Edman degradation of covalently linked glycopeptides to identify positively the sites of O- and N-glycosylation. The O-glycosidic linkage of carbohydrate to the hydroxy amino acids Ser and Thr is a major form of post-translational modification. However, unlike Asn-linked glycosylation, which is identified by the consensus sequence Asn-Xaa-Thr/Ser, no simple motif conferring O-linkage to Thr and Ser has been described. After sequencing glycopeptides derived from two cell surface glycoproteins, a Thr-O-glycosylation motif of Xaa-Pro-Xaa-Xaa, where at least one Xaa = Thr(Sac), has been defined. This motif predicts the site(s) of Pro- associated Thr-O-glycosylation in O-glycosylated proteins, although it is clear that there are also other forms of Thr-O-glycosylation not associated with Pro.
Notes: Gooley, A A
Classon, B J
Marschalek, R
Williams, K L
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 1991 Aug 15;178(3):1194-201. doi: 10.1016/0006-291x(91)91019-9.
Author Address: School of Biological Sciences, Macquarie University, Sydney, N.S.W. Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 540
Author: Gooley, A. A., Pisano, A., Packer, N. H., Ball, M., Jones, A., Alewood, P. F., Redmond, J. W. and Williams, K. L.
Year: 1994
Title: Characterization of a single glycosylated asparagine site on a glycopeptide using solid-phase Edman degradation
Journal: Glycoconj J
Volume: 11
Issue: 3
Pages: 180-6
Epub Date: 1994/06/01
Date: Jun
Short Title: Characterization of a single glycosylated asparagine site on a glycopeptide using solid-phase Edman degradation
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/bf00731216
Accession Number: 7841792
Keywords: Amino Acid Sequence
Asparagine/chemistry/*metabolism
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Glycopeptides/chemistry/*metabolism
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/metabolism
Phenytoin/analogs & derivatives
Serum Albumin/metabolism
Abstract: The characterization of site-specific glycosylation is traditionally dependent on the availability of suitable proteolytic cleavage sites between each glycosylated residue, so that peptides containing individual glycosylation sites are recovered. In the case of heavily glycosylated domains such as the O-glycosylated mucins, which have no available protease sites, this approach is not possible. Here we introduce a new method to gain site-specific compositional data on the oligosaccharides attached to a single amino acid. Using a model glycopeptide from a mutant human albumin Casebrook, glycosylated PTH-Asn was recovered after sequential solid-phase Edman degradation, subjected to acid hydrolysis and the sugars were identified by high performance anion exchange chromatography with pulsed amperometric detection. The PTH-Asn(Sac) derivative was further characterized by ionspray mass spectrometry. Comparison between an endoproteinase Glu-C glycopeptide and a tryptic glycopeptide showed that the oligosaccharide attached to Asn494 was stable after at least 10 cycles of Edman degradation.
Notes: Gooley, A A
Pisano, A
Packer, N H
Ball, M
Jones, A
Alewood, P F
Redmond, J W
Williams, K L
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 1994 Jun;11(3):180-6. doi: 10.1007/BF00731216.
Author Address: Macquarie University Centre for Analytical Biotechnology (MUCAB), Sydney, NSW, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1972
Author: Gorelik, A., Gebai, A., Illes, K., Piomelli, D. and Nagar, B.
Year: 2018
Title: Molecular mechanism of activation of the immunoregulatory amidase NAAA
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 43
Pages: E10032-e10040
Epub Date: 2018/10/12
Date: Oct 23
Short Title: Molecular mechanism of activation of the immunoregulatory amidase NAAA
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1811759115
PMCID: PMC6205464
Accession Number: 30301806
Keywords: Amides
Amidohydrolases/*metabolism
Analgesics/pharmacology
Animals
Catalytic Domain/drug effects
Cell Line
Drug Discovery/methods
Enzyme Inhibitors/pharmacology
Ethanolamines/metabolism
Humans
Inflammation/metabolism
Ligands
Mice
Pain/drug therapy/metabolism
Palmitic Acids/metabolism
Rabbits
Sf9 Cells
Structure-Activity Relationship
*N-acylethanolamine acid amidase
*N-terminal nucleophile hydrolase
*endocannabinoid
*interfacial activation
*palmitoylethanolamide
Abstract: Palmitoylethanolamide is a bioactive lipid that strongly alleviates pain and inflammation in animal models and in humans. Its signaling activity is terminated through degradation by N-acylethanolamine acid amidase (NAAA), a cysteine hydrolase expressed at high levels in immune cells. Pharmacological inhibitors of NAAA activity exert profound analgesic and antiinflammatory effects in rodent models, pointing to this protein as a potential target for therapeutic drug discovery. To facilitate these efforts and to better understand the molecular mechanism of action of NAAA, we determined crystal structures of this enzyme in various activation states and in complex with several ligands, including both a covalent and a reversible inhibitor. Self-proteolysis exposes the otherwise buried active site of NAAA to allow catalysis. Formation of a stable substrate- or inhibitor-binding site appears to be conformationally coupled to the interaction of a pair of hydrophobic helices in the enzyme with lipid membranes, resulting in the creation of a linear hydrophobic cavity near the active site that accommodates the ligand's acyl chain.
Notes: 1091-6490
Gorelik, Alexei
Orcid: 0000-0001-6236-4633
Gebai, Ahmad
Illes, Katalin
Piomelli, Daniele
Nagar, Bhushan
Orcid: 0000-0001-9869-2110
MOP-133535/Canadian Institutes of Health Research/International
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2018 Oct 23;115(43):E10032-E10040. doi: 10.1073/pnas.1811759115. Epub 2018 Oct 9.
Author Address: Department of Biochemistry, McGill University, Montreal, H3G0B1, Canada.
Department of Anatomy and Neurobiology, University of California, Irvine, CA 92697.
Department of Biochemistry, University of California, Irvine, CA 92697.
Department of Pharmacology, University of California, Irvine, CA 92697.
Department of Biochemistry, McGill University, Montreal, H3G0B1, Canada; bhushan.nagar@mcgill.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1811
Author: Gorelik, A., Heinz, L. X., Illes, K., Superti-Furga, G. and Nagar, B.
Year: 2016
Title: Crystal Structure of the Acid Sphingomyelinase-like Phosphodiesterase SMPDL3B Provides Insights into Determinants of Substrate Specificity
Journal: J Biol Chem
Volume: 291
Issue: 46
Pages: 24054-24064
Epub Date: 2016/10/01
Date: Nov 11
Short Title: Crystal Structure of the Acid Sphingomyelinase-like Phosphodiesterase SMPDL3B Provides Insights into Determinants of Substrate Specificity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.755801
PMCID: PMC5104931
Accession Number: 27687724
Keywords: Adenosine Diphosphate/chemistry/genetics/metabolism
Adenosine Triphosphate/chemistry/genetics/metabolism
Animals
Choline/chemistry/genetics/metabolism
Crystallography, X-Ray
Cyclic Nucleotide Phosphodiesterases, Type 3/*chemistry/genetics/metabolism
Mice
Protein Domains
Protein Structure, Secondary
Sf9 Cells
Sphingomyelins/chemistry/genetics/metabolism
Spodoptera
Substrate Specificity
*Smpdl3b
*cell surface enzyme
*crystal structure
*lipid metabolism
*sphingomyelinase
*substrate specificity
Abstract: The enzyme acid sphingomyelinase-like phosphodiesterase 3B (SMPDL3B) was shown to act as a negative regulator of innate immune signaling, affecting cellular lipid composition and membrane fluidity. Furthermore, several reports identified this enzyme as an off target of the therapeutic antibody rituximab, with implications in kidney disorders. However, structural information for this protein is lacking. Here we present the high resolution crystal structure of murine SMPDL3B, which reveals a substrate binding site strikingly different from its paralogs. The active site is located in a narrow boot-shaped cavity. We identify a unique loop near the active site that appears to impose size constraints on incoming substrates. A structure in complex with phosphocholine indicates that the protein recognizes this head group via an aromatic box, a typical choline-binding motif. Although a potential substrate for SMPDL3B is sphingomyelin, we identify other possible substrates such as CDP-choline, ATP, and ADP. Functional experiments employing structure-guided mutagenesis in macrophages highlight amino acid residues potentially involved in recognition of endogenous substrates. Our study is an important step toward elucidating the specific function of this poorly characterized enzyme.
Notes: 1083-351x
Gorelik, Alexei
Heinz, Leonhard X
Illes, Katalin
Superti-Furga, Giulio
Nagar, Bhushan
P41 GM103485/GM/NIGMS NIH HHS/United States
Journal Article
J Biol Chem. 2016 Nov 11;291(46):24054-24064. doi: 10.1074/jbc.M116.755801. Epub 2016 Sep 28.
Author Address: From the Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Quebec H3G 0B1, Canada.
the CeMM Research Center for Molecular Medicine, Austrian Academy of Sciences, 1090 Vienna, Austria, and.
the Center for Physiology and Pharmacology, Medical University of Vienna, 1090 Vienna, Austria.
From the Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Quebec H3G 0B1, Canada, bhushan.nagar@mcgill.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1798
Author: Gorelik, A., Illes, K., Heinz, L. X., Superti-Furga, G. and Nagar, B.
Year: 2016
Title: Crystal structure of mammalian acid sphingomyelinase
Journal: Nat Commun
Volume: 7
Pages: 12196
Epub Date: 2016/07/21
Date: Jul 20
Short Title: Crystal structure of mammalian acid sphingomyelinase
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms12196
PMCID: PMC4961792
Accession Number: 27435900
Keywords: Animals
Catalytic Domain
Crystallography, X-Ray
Enzyme Assays
Enzyme Inhibitors/chemistry/metabolism
Humans
Hydrolysis
Mammals/*metabolism
Membrane Lipids/metabolism
Mice
Models, Molecular
Molecular Docking Simulation
Mutation/genetics
Niemann-Pick Diseases/genetics
Protein Structure, Secondary
Saposins/chemistry
Sphingomyelin Phosphodiesterase/*chemistry/genetics/isolation &
purification/metabolism
Static Electricity
Substrate Specificity
Abstract: Acid sphingomyelinase (ASMase, ASM, SMPD1) converts sphingomyelin into ceramide, modulating membrane properties and signal transduction. Inactivating mutations in ASMase cause Niemann-Pick disease, and its inhibition is also beneficial in models of depression and cancer. To gain a better understanding of this critical therapeutic target, we determined crystal structures of mammalian ASMase in various conformations. The catalytic domain adopts a calcineurin-like fold with two zinc ions and a hydrophobic track leading to the active site. Strikingly, the membrane interacting saposin domain assumes either a closed globular conformation independent from the catalytic domain, or an open conformation, which establishes an interface with the catalytic domain essential for activity. Structural mapping of Niemann-Pick mutations reveals that most of them likely destabilize the protein's fold. This study sheds light on the molecular mechanism of ASMase function, and provides a platform for the rational development of ASMase inhibitors and therapeutic use of recombinant ASMase.
Notes: 2041-1723
Gorelik, Alexei
Illes, Katalin
Heinz, Leonhard X
Superti-Furga, Giulio
Nagar, Bhushan
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2016 Jul 20;7:12196. doi: 10.1038/ncomms12196.
Author Address: Department of Biochemistry, McGill University, Montreal, Quebec, Canada H3G 0B1.
Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Quebec, Canada H3G 0B1.
CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria.
Center for Physiology and Pharmacology, Medical University of Vienna, 1090 Vienna, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1907
Author: Gorelik, A., Illes, K. and Nagar, B.
Year: 2018
Title: Crystal structure of the mammalian lipopolysaccharide detoxifier
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 5
Pages: E896-e905
Epub Date: 2018/01/19
Date: Jan 30
Short Title: Crystal structure of the mammalian lipopolysaccharide detoxifier
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1719834115
PMCID: PMC5798384
Accession Number: 29343645
Keywords: Animals
Calcium/chemistry
Carboxylic Ester Hydrolases/*chemistry
Catalytic Domain
Cell Membrane/metabolism
Crystallography, X-Ray
Endosomes/metabolism
Humans
Hydrophobic and Hydrophilic Interactions
Immune System
Lipopolysaccharides/*chemistry
Mice
Protein Binding
Protein Domains
Protein Structure, Secondary
Rabbits
Saposins/chemistry
Scattering, Radiation
Surface Properties
X-Rays
*acyloxyacyl hydrolase
*calcium-binding protein
*immune tolerance
*lipopolysaccharide
*saposin
Abstract: LPS is a potent bacterial endotoxin that triggers the innate immune system. Proper recognition of LPS by pattern-recognition receptors requires a full complement of typically six acyl chains in the lipid portion. Acyloxyacyl hydrolase (AOAH) is a host enzyme that removes secondary (acyloxyacyl-linked) fatty acids from LPS, rendering it immunologically inert. This activity is critical for recovery from immune tolerance that follows Gram-negative infection. To understand the molecular mechanism of AOAH function, we determined its crystal structure and its complex with LPS. The substrate's lipid moiety is accommodated in a large hydrophobic pocket formed by the saposin and catalytic domains with a secondary acyl chain inserted into a narrow lateral hydrophobic tunnel at the active site. The enzyme establishes dispensable contacts with the phosphate groups of LPS but does not interact with its oligosaccharide portion. Proteolytic processing allows movement of an amphipathic helix possibly involved in substrate access at membranes.
Notes: 1091-6490
Gorelik, Alexei
Illes, Katalin
Nagar, Bhushan
Orcid: 0000-0001-9869-2110
P41 GM103485/GM/NIGMS NIH HHS/United States
MOP-133535/CIHR/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2018 Jan 30;115(5):E896-E905. doi: 10.1073/pnas.1719834115. Epub 2018 Jan 17.
Author Address: Department of Biochemistry, McGill University, Montreal, H3G0B1, Canada.
Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, H3G0B1, Canada.
Department of Biochemistry, McGill University, Montreal, H3G0B1, Canada; bhushan.nagar@mcgill.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1751
Author: Gorelik, A., Illes, K., Superti-Furga, G. and Nagar, B.
Year: 2016
Title: Structural Basis for Nucleotide Hydrolysis by the Acid Sphingomyelinase-like Phosphodiesterase SMPDL3A
Journal: J Biol Chem
Volume: 291
Issue: 12
Pages: 6376-85
Epub Date: 2016/01/23
Date: Mar 18
Short Title: Structural Basis for Nucleotide Hydrolysis by the Acid Sphingomyelinase-like Phosphodiesterase SMPDL3A
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M115.711085
PMCID: PMC4813564
Accession Number: 26792860
Keywords: Adenosine Monophosphate/*chemistry
Adenosine Triphosphate/chemistry
Amino Acid Sequence
Animals
Biocatalysis
Catalytic Domain
Cell Line
Crystallography, X-Ray
Humans
Hydrolysis
Kinetics
Mice
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Secondary
Sphingomyelin Phosphodiesterase/*chemistry
Substrate Specificity
Zinc/chemistry
Smpdl3a
crystal structure
enzyme catalysis
nucleoside/nucleotide metabolism
sphingomyelinase
zinc
Abstract: Sphingomyelin phosphodiesterase, acid-like 3A (SMPDL3A) is a member of a small family of proteins founded by the well characterized lysosomal enzyme, acid sphingomyelinase (ASMase). ASMase converts sphingomyelin into the signaling lipid, ceramide. It was recently discovered that, in contrast to ASMase, SMPDL3A is inactive against sphingomyelin and, surprisingly, can instead hydrolyze nucleoside diphosphates and triphosphates, which may play a role in purinergic signaling. As none of the ASMase-like proteins has been structurally characterized to date, the molecular basis for their substrate preferences is unknown. Here we report crystal structures of murine SMPDL3A, which represent the first structures of an ASMase-like protein. The catalytic domain consists of a central mixed β-sandwich surrounded by α-helices. Additionally, SMPDL3A possesses a unique C-terminal domain formed from a cluster of four α-helices that appears to distinguish this protein family from other phosphoesterases. We show that SMDPL3A is a di-zinc-dependent enzyme with an active site configuration that suggests a mechanism of phosphodiester hydrolysis by a metal-activated water molecule and protonation of the leaving group by a histidine residue. Co-crystal structures of SMPDL3A with AMP and α,β-methylene ADP (AMPCP) reveal that the substrate binding site accommodates nucleotides by establishing interactions with their base, sugar, and phosphate moieties, with the latter the major contributor to binding affinity. Our study provides the structural basis for SMPDL3A substrate specificity and sheds new light on the function of ASMase-like proteins.
Notes: 1083-351x
Gorelik, Alexei
Illes, Katalin
Superti-Furga, Giulio
Nagar, Bhushan
P41 GM103485/GM/NIGMS NIH HHS/United States
GM-103485/GM/NIGMS NIH HHS/United States
MOP-133535/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2016 Mar 18;291(12):6376-85. doi: 10.1074/jbc.M115.711085. Epub 2016 Jan 20.
Author Address: From the Department of Biochemistry and Groupe de Recherche Axe sur la Structure des Proteines, Faculty of Medicine, McGill University, Montreal, Quebec H3G 0B1, Canada and.
CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, A-1090 Vienna, Austria.
From the Department of Biochemistry and Groupe de Recherche Axe sur la Structure des Proteines, Faculty of Medicine, McGill University, Montreal, Quebec H3G 0B1, Canada and bhushan.nagar@mcgill.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1838
Author: Gorelik, A., Liu, F., Illes, K. and Nagar, B.
Year: 2017
Title: Crystal structure of the human alkaline sphingomyelinase provides insights into substrate recognition
Journal: J Biol Chem
Volume: 292
Issue: 17
Pages: 7087-7094
Epub Date: 2017/03/16
Date: Apr 28
Short Title: Crystal structure of the human alkaline sphingomyelinase provides insights into substrate recognition
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.769273
PMCID: PMC5409475
Accession Number: 28292932
Keywords: Animals
Bile Acids and Salts/chemistry
Catalysis
Catalytic Domain
Cations
Cell Line
Crystallography, X-Ray
DNA Mutational Analysis
Detergents/chemistry
Humans
Hydrolysis
Hydrophobic and Hydrophilic Interactions
Insecta
Micelles
Phosphorylcholine/chemistry
Protein Binding
Protein Sorting Signals
Salts/chemistry
Sphingomyelin Phosphodiesterase/*chemistry
Substrate Specificity
Tyrosine/chemistry
*Enpp7
*Npp7
*alk-SMase
*bile acid
*choline
*micelle
*sphingolipid
*sphingomyelinase
this article
Abstract: Absorption of dietary sphingomyelin (SM) requires its initial degradation into ceramide, a process catalyzed by the intestinal enzyme alkaline sphingomyelinase (alk-SMase, NPP7, ENPP7). alk-SMase belongs to the nucleotide pyrophosphatase/phosphodiesterase (NPP) family, the members of which hydrolyze nucleoside phosphates, phospholipids, and other related molecules. NPP7 is the only paralog that can cleave SM, and its activity requires the presence of bile salts, a class of physiological anionic detergents. To elucidate the mechanism of substrate recognition, we determined the crystal structure of human alk-SMase in complex with phosphocholine, a reaction product. Although the overall fold and catalytic center are conserved relative to other NPPs, alk-SMase recognizes the choline moiety of its substrates via an NPP7-specific aromatic box composed of tyrosine residues. Mutational analysis and enzymatic activity assays identified features on the surface of the protein-a cationic patch and a unique hydrophobic loop-that are essential for accessing SM in bile salt micelles. These results shed new light on substrate specificity determinants within the NPP enzyme family.
Notes: 1083-351x
Gorelik, Alexei
Liu, Fangyu
Illes, Katalin
Nagar, Bhushan
MOP-133535/CIHR/Canada
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2017 Apr 28;292(17):7087-7094. doi: 10.1074/jbc.M116.769273. Epub 2017 Mar 14.
Author Address: From the Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Quebec H3G 0B1, Canada.
From the Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Quebec H3G 0B1, Canada bhushan.nagar@mcgill.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1936
Author: Gorelik, A., Randriamihaja, A., Illes, K. and Nagar, B.
Year: 2018
Title: Structural basis for nucleotide recognition by the ectoenzyme CD203c
Journal: Febs j
Volume: 285
Issue: 13
Pages: 2481-2494
Epub Date: 2018/05/03
Date: Jul
Short Title: Structural basis for nucleotide recognition by the ectoenzyme CD203c
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/febs.14489
Accession Number: 29717535
Keywords: Adenosine Triphosphate/chemistry/metabolism
Amino Acid Sequence
Animals
Crystallography, X-Ray
Humans
Models, Molecular
Nucleotides/*chemistry/genetics/metabolism
Phosphoric Diester Hydrolases/*chemistry/genetics/metabolism
Protein Binding
*Protein Domains
Protein Multimerization
Pyrophosphatases/*chemistry/genetics/metabolism
Sequence Homology, Amino Acid
Sf9 Cells
Spodoptera
Substrate Specificity
*CD203c
*Enpp3
*Npp3
*nucleotide
*phosphodiesterase
Abstract: The ecto-nucleotide pyrophosphatase/phosphodiesterase (NPP) enzyme family modulates purinergic signaling by degrading extracellular nucleotides. CD203c (NPP3, ENPP3) regulates the inflammatory response of basophils via ATP hydrolysis and is a marker for allergen sensitivity on the surface of these cells. Multiple other roles and substrates have also been proposed for this protein. In order to gain insight into its molecular functions, we determined the crystal structure of human NPP3 as well as its complex with an ATP analog. The enzyme exhibits little preference for nucleobase type, and forms specific contacts with the alpha and beta phosphate groups of its ligands. Dimerization of the protein does not affect its catalytic activity. These findings expand our understanding of substrate recognition within the NPP family. DATABASE: Structural data are available in the Protein Data Bank under the accession numbers 6C01 (human NPP3) and 6C02 (human NPP3 T205A N594S with AMPCPP).
Notes: 1742-4658
Gorelik, Alexei
Orcid: 0000-0001-6236-4633
Randriamihaja, Antsa
Illes, Katalin
Nagar, Bhushan
MOP‐133535/CIHR/Canada
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2018 Jul;285(13):2481-2494. doi: 10.1111/febs.14489. Epub 2018 May 9.
Author Address: Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Quebec, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 579
Author: Grabenhorst, E., Hofer, B., Nimtz, M., Jäger, V. and Conradt, H. S.
Year: 1993
Title: Biosynthesis and secretion of human interleukin 2 glycoprotein variants from baculovirus-infected Sf21 cells. Characterization of polypeptides and posttranslational modifications
Journal: Eur J Biochem
Volume: 215
Issue: 1
Pages: 189-97
Epub Date: 1993/07/01
Date: Jul 1
Short Title: Biosynthesis and secretion of human interleukin 2 glycoprotein variants from baculovirus-infected Sf21 cells. Characterization of polypeptides and posttranslational modifications
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb18022.x
Accession Number: 8344280
Keywords: Amino Acid Sequence
Animals
Baculoviridae/metabolism
Carbohydrate Sequence
Cell Line
Glycosylation
Humans
Interleukin-2/*biosynthesis/chemistry/metabolism
Molecular Sequence Data
Moths
Peptide Mapping
*Protein Processing, Post-Translational
Recombinant Proteins/*biosynthesis
Abstract: Human interleukin 2 (IL-2) and human IL-2 mutant proteins, with artificially introduced N-glycosylation or O-glycosylation sites, have been expressed in a lepidopteran cell line (Sf21, Spodoptera frugiperda) using recombinant baculovirus vectors. Only approximately 25% of the total recombinant IL-2 protein synthesized by Sf21 cells was secreted into the culture medium. Significant N-terminal truncations were detected in the secreted polypeptides (up to 85% of the molecules). Alanine and proline were absent in the major truncated forms; the first 3-5 amino acids were also absent in a small proportion of the purified proteins. The introduction of potential artificial O-glycosylation peptide sequences (..GGKAPTPPPK..), to the C-terminus or between positions 80 and 81 of the IL-2 polypeptide chain, resulted in the secretion of unglycosylated and O-glycosylated variant forms. Fast atom bombardment mass spectrometry, compositional analysis and methylation analysis, of the tryptic glycopeptide APTPPPK, revealed the presence of either GalNAc or the disaccharide Gal(beta 1-3)GalNAc as the only carbohydrate constituents attached exclusively to Thr in this peptide, in a specific ratio for each individual IL-2 mutant protein. The Gal(beta 1-3)GalNAc protein forms could be partially altered in vitro to mammalian-type glycoforms by porcine liver beta-galactoside alpha-2,3-sialyltransferase in the presence of CMP-N-acetylneuraminic acid. An IL-2 mutant form, with an 11-amino-acid peptide of human interferon-beta at position 4, which includes its only N-glycosylation site, had exclusively truncated proximally fucosylated oligomannosidic glycans; Man3GlcNAc[Fuc(alpha 1-6)]GlcNAc or Man2GlcNAc[Fuc(alpha 1-6)]GlcNAc structures, in a ratio of 3:1, were detected in the secreted proteins. No evidence was obtained for the presence of secreted proteins with complex oligosaccharide chains, irrespective of the cell-culture conditions used or the harvesting time, for infected cells with recombinant baculovirus constructs.
Notes: Grabenhorst, E
Hofer, B
Nimtz, M
Jäger, V
Conradt, H S
Journal Article
England
Eur J Biochem. 1993 Jul 1;215(1):189-97. doi: 10.1111/j.1432-1033.1993.tb18022.x.
Author Address: Department of Cell Biology and Genetics, Gesellschaft für Biotechnologische Forschung, Braunschweig, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 931
Author: Graf, G. A., Cohen, J. C. and Hobbs, H. H.
Year: 2004
Title: Missense mutations in ABCG5 and ABCG8 disrupt heterodimerization and trafficking
Journal: J Biol Chem
Volume: 279
Issue: 23
Pages: 24881-8
Epub Date: 2004/04/01
Date: Jun 4
Short Title: Missense mutations in ABCG5 and ABCG8 disrupt heterodimerization and trafficking
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M402634200
Accession Number: 15054092
Keywords: ATP Binding Cassette Transporter, Subfamily G, Member 5
ATP Binding Cassette Transporter, Subfamily G, Member 8
ATP-Binding Cassette Transporters/*genetics
Amino Acid Sequence
Animals
Asparagine/chemistry
Biological Transport
CHO Cells
Calnexin/chemistry
Cricetinae
Dimerization
Electrophoresis, Polyacrylamide Gel
Endoplasmic Reticulum/metabolism
Glycosylation
Golgi Apparatus/metabolism
Humans
Lectins/chemistry
Lipoproteins/*genetics
Mice
Models, Biological
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutation
*Mutation, Missense
Precipitin Tests
Protein Binding
Protein Folding
Protein Transport
Sequence Homology, Amino Acid
Transfection
Abstract: Mutations in ABCG5 (G5) or ABCG8 (G8) cause sitosterolemia, an autosomal recessive disease characterized by sterol accumulation and premature atherosclerosis. G5 and G8 are ATP-binding cassette (ABC) half-transporters that must heterodimerize to move to the apical surface of cells. We examined the role of N-linked glycans in the formation of the G5/G8 heterodimer to gain insight into the determinants of folding and trafficking of these proteins. Site-directed mutagenesis revealed that two asparagine residues (Asn(585) and Asn(592)) are glycosylated in G5 and that G8 has a single N-linked glycan attached to Asn(619). N-Linked glycosylation of G8 was required for efficient trafficking of the G5/G8 heterodimer, but mutations that abolished glycosylation of G5 did not prevent trafficking of the heterodimer. Both G5 and G8 are bound by the lectin chaperone, calnexin, suggesting that the calnexin cycle may facilitate folding of the G5/G8 heterodimer. To determine the effects of 13 disease-causing missense mutations in G5 and G8 on formation and trafficking of the G5/G8 heterodimer, mutant forms of the half-transporters were expressed in CHO-K1 cells. All 13 mutations reduced trafficking of the G5/G8 heterodimer from the endoplasmic reticulum to the Golgi complex, and most prevented the formation of stable heterodimers between G5 and G8. We conclude that the majority of the molecular defects in G5 and G8 that cause sitosterolemia impair transport of the sterol transporter to the cell surface.
Notes: Graf, Gregory A
Cohen, Jonathan C
Hobbs, Helen H
HL20948/HL/NHLBI NIH HHS/United States
HL72304/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2004 Jun 4;279(23):24881-8. doi: 10.1074/jbc.M402634200. Epub 2004 Mar 30.
Author Address: McDermott Center for Human Growth and Development and Department of Molecular Genetics, University of Texas Southwestern Medical Center at Dallas, 75390, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1389
Author: Graham, M. E., Thaysen-Andersen, M., Bache, N., Craft, G. E., Larsen, M. R., Packer, N. H. and Robinson, P. J.
Year: 2011
Title: A novel post-translational modification in nerve terminals: O-linked N-acetylglucosamine phosphorylation
Journal: J Proteome Res
Volume: 10
Issue: 6
Pages: 2725-33
Epub Date: 2011/04/20
Date: Jun 3
Short Title: A novel post-translational modification in nerve terminals: O-linked N-acetylglucosamine phosphorylation
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr1011153
Accession Number: 21500857
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Conserved Sequence
Molecular Sequence Data
Monomeric Clathrin Assembly Proteins/chemistry/metabolism
Peptide Fragments/chemistry
Phosphorylation
*Protein Processing, Post-Translational
Rats
Sugar Phosphates/*metabolism
Synapses/*metabolism
Tandem Mass Spectrometry
Abstract: Protein phosphorylation and glycosylation are the most common post-translational modifications observed in biology, frequently on the same protein. Assembly protein AP180 is a synapse-specific phosphoprotein and O-linked beta-N-acetylglucosamine (O-GlcNAc) modified glycoprotein. AP180 is involved in the assembly of clathrin coated vesicles in synaptic vesicle endocytosis. Unlike other types of O-glycosylation, O-GlcNAc is nucleocytoplasmic and reversible. It was thought to be a terminal modification, that is, the O-GlcNAc was not found to be additionally modified in any way. We now show that AP180 purified from rat brain contains a phosphorylated O-GlcNAc (O-GlcNAc-P) within a highly conserved sequence. O-GlcNAc or O-GlcNAc-P, but not phosphorylation alone, was found at Thr-310. Analysis of synthetic GlcNAc-6-P produced identical fragmentation products to GlcNAc-P from AP180. Direct O-linkage of GlcNAc-P to a Thr residue was confirmed by electron transfer dissociation MS. A second AP180 tryptic peptide was also glycosyl phosphorylated, but the site of modification was not assigned. Sequence similarities suggest there may be a common motif within AP180 involving glycosyl phosphorylation and dual flanking phosphorylation sites within 4 amino acid residues. This novel type of protein glycosyl phosphorylation adds a new signaling mechanism to the regulation of neurotransmission and more complexity to the study of O-GlcNAc modification.
Notes: 1535-3907
Graham, Mark E
Thaysen-Andersen, Morten
Bache, Nicolai
Craft, George E
Larsen, Martin R
Packer, Nicolle H
Robinson, Phillip J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2011 Jun 3;10(6):2725-33. doi: 10.1021/pr1011153. Epub 2011 May 2.
Author Address: Cell Signalling Unit, Children's Medical Research Institute, The University of Sydney, Westmead, Australia. mgraham@cmri.org.au
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1000
Author: Grayson, C. and Molday, R. S.
Year: 2005
Title: Dominant negative mechanism underlies autosomal dominant Stargardt-like macular dystrophy linked to mutations in ELOVL4
Journal: J Biol Chem
Volume: 280
Issue: 37
Pages: 32521-30
Epub Date: 2005/07/23
Date: Sep 16
Short Title: Dominant negative mechanism underlies autosomal dominant Stargardt-like macular dystrophy linked to mutations in ELOVL4
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M503411200
Accession Number: 16036915
Keywords: Amino Acid Motifs
Animals
Antibodies, Monoclonal/chemistry
COS Cells
Cell Line
Cell Nucleus/metabolism
Cells, Cultured
Centrifugation, Density Gradient
Chromatography, Gel
DNA, Complementary/metabolism
Electrophoresis, Polyacrylamide Gel
Endoplasmic Reticulum/metabolism
Eye Proteins/*genetics
*Genes, Dominant
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Immunoprecipitation
Macular Degeneration/*genetics/metabolism
Membrane Proteins/*genetics
Microscopy, Fluorescence
*Mutation
Oligosaccharides/chemistry
Protein Structure, Tertiary
Retina/metabolism
Retinal Diseases/*genetics/metabolism
Sucrose/pharmacology
Transfection
Vision, Ocular
Abstract: ELOVL4 (elongation of very long chain fatty acids 4) is a member of the ELO family of proteins involved in the biosynthesis of very long chain fatty acids. Protein truncation mutations in ELOVL4 have been identified in patients with autosomal dominant Stargardt-like macular degeneration. To determine whether a dominant negative mechanism is responsible for the autosomal dominant inheritance pattern of this disease, we studied the subcellular localization and interaction of wild type and mutant ELOVL4 in COS-7 and HEK 293T cultured cells by immunofluorescence and co-immunoprecipitation. Wild type ELOVL4 containing an endoplasmic reticulum retention sequence was localized to the endoplasmic reticulum as expected. In contrast, disease-associated C-terminal truncation ELOVL4 mutants accumulated as large inclusions exhibiting aggresome-like characteristics in a juxtanuclear position within COS-7 cells. When the wild type and mutant proteins were co-expressed incultured cells, wild type ELOVL4 co-purified with mutant ELOVL4 on an immunoaffinity column and co-localized with the mutant protein in aggresome-like inclusions adjacent to the nucleus. These results indicate that wild type and mutant ELOVL4 form a complex that exhibits an abnormal subcellular localization found for individually expressed mutant ELOVL4. From these studies, we conclude that disease-linked C-terminal truncation mutants of ELOVL4 exert a dominant negative effect on wild type ELOVL4, altering its subcellular localization. This dominant negative mechanism contributes to the autosomal dominant inheritance of Stargardt-like macular dystrophy.
Notes: Grayson, Celene
Molday, Robert S
EY 02422/EY/NEI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2005 Sep 16;280(37):32521-30. doi: 10.1074/jbc.M503411200. Epub 2005 Jul 21.
Author Address: Department of Biochemistry and Molecular Biology, Centre for Macular Research, University of British Columbia, Vancouver, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 885
Author: Gregory, L. A., Thielens, N. M., Arlaud, G. J., Fontecilla-Camps, J. C. and Gaboriaud, C.
Year: 2003
Title: X-ray structure of the Ca2+-binding interaction domain of C1s. Insights into the assembly of the C1 complex of complement
Journal: J Biol Chem
Volume: 278
Issue: 34
Pages: 32157-64
Epub Date: 2003/06/06
Date: Aug 22
Short Title: X-ray structure of the Ca2+-binding interaction domain of C1s. Insights into the assembly of the C1 complex of complement
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M305175200
Accession Number: 12788922
Keywords: Amino Acid Sequence
Animals
Base Sequence
Binding Sites
Calcium/*metabolism
Cell Line
Complement C1/chemistry/*metabolism
Crystallography, X-Ray
DNA Primers
Dimerization
Electrophoresis, Polyacrylamide Gel
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Conformation
Recombinant Proteins/chemistry/metabolism
Sequence Homology, Amino Acid
Spodoptera
Abstract: C1, the complex that triggers the classical pathway of complement, is assembled from two modular proteases C1r and C1s and a recognition protein C1q. The N-terminal CUB1-EGF segments of C1r and C1s are key elements of the C1 architecture, because they mediate both Ca2+-dependent C1r-C1s association and interaction with C1q. The crystal structure of the interaction domain of C1s has been solved and refined to 1.5 A resolution. The structure reveals a head-to-tail homodimer involving interactions between the CUB1 module of one monomer and the epidermal growth factor (EGF) module of its counterpart. A Ca2+ ion is bound to each EGF module and stabilizes both the intra- and inter-monomer interfaces. Unexpectedly, a second Ca2+ ion is bound to the distal end of each CUB1 module, through six ligands contributed by Glu45, Asp53, Asp98, and two water molecules. These acidic residues and Tyr17 are conserved in approximately two-thirds of the CUB repertoire and define a novel, Ca2+-binding CUB module subset. The C1s structure was used to build a model of the C1r-C1s CUB1-EGF heterodimer, which in C1 connects C1r to C1s and mediates interaction with C1q. A structural model of the C1q/C1r/C1s interface is proposed, where the rod-like collagen triple helix of C1q is accommodated into a groove along the transversal axis of the C1r-C1s heterodimer.
Notes: Gregory, Lynn A
Thielens, Nicole M
Arlaud, Gérard J
Fontecilla-Camps, Juan Carlos
Gaboriaud, Christine
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2003 Aug 22;278(34):32157-64. doi: 10.1074/jbc.M305175200. Epub 2003 Jun 4.
Author Address: Laboratoire de Cristallographie et Cristallogénèse des Protéines, Institut de Biologie Structurale Jean-Pierre Ebel, 41 rue Jules Horowitz, 38027 Grenoble Cedex 1, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1822
Author: Grieben, M., Pike, A. C., Shintre, C. A., Venturi, E., El-Ajouz, S., Tessitore, A., Shrestha, L., Mukhopadhyay, S., Mahajan, P., Chalk, R., Burgess-Brown, N. A., Sitsapesan, R., Huiskonen, J. T. and Carpenter, E. P.
Year: 2017
Title: Structure of the polycystic kidney disease TRP channel Polycystin-2 (PC2)
Journal: Nat Struct Mol Biol
Volume: 24
Issue: 2
Pages: 114-122
Epub Date: 2016/12/20
Date: Feb
Short Title: Structure of the polycystic kidney disease TRP channel Polycystin-2 (PC2)
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.3343
Accession Number: 27991905
Keywords: Animals
Cryoelectron Microscopy
Humans
Models, Molecular
Mutation, Missense
Polycystic Kidney, Autosomal Dominant/genetics
Protein Conformation, alpha-Helical
Protein Domains
Sf9 Cells
Spodoptera
TRPP Cation Channels/*chemistry/genetics
Abstract: Mutations in either polycystin-1 (PC1 or PKD1) or polycystin-2 (PC2, PKD2 or TRPP1) cause autosomal-dominant polycystic kidney disease (ADPKD) through unknown mechanisms. Here we present the structure of human PC2 in a closed conformation, solved by electron cryomicroscopy at 4.2-Å resolution. The structure reveals a novel polycystin-specific 'tetragonal opening for polycystins' (TOP) domain tightly bound to the top of a classic transient receptor potential (TRP) channel structure. The TOP domain is formed from two extensions to the voltage-sensor-like domain (VSLD); it covers the channel's endoplasmic reticulum lumen or extracellular surface and encloses an upper vestibule, above the pore filter, without blocking the ion-conduction pathway. The TOP-domain fold is conserved among the polycystins, including the homologous channel-like region of PC1, and is the site of a cluster of ADPKD-associated missense variants. Extensive contacts among the TOP-domain subunits, the pore and the VSLD provide ample scope for regulation through physical and chemical stimuli.
Notes: 1545-9985
Grieben, Mariana
Pike, Ashley C W
Orcid: 0000-0001-9661-2607
Shintre, Chitra A
Venturi, Elisa
El-Ajouz, Sam
Tessitore, Annamaria
Shrestha, Leela
Mukhopadhyay, Shubhashish
Mahajan, Pravin
Chalk, Rod
Burgess-Brown, Nicola A
Sitsapesan, Rebecca
Huiskonen, Juha T
Orcid: 0000-0002-0348-7323
Carpenter, Elisabeth P
Orcid: 0000-0001-9138-2937
RG/10/14/28576/British Heart Foundation/United Kingdom
092809/Z/10/Z/Wellcome Trust/United Kingdom
060208/Z/00/Z/Wellcome Trust/United Kingdom
093305/Z/10/Z/Wellcome Trust/United Kingdom
090532/Z/09/Z/Wellcome Trust/United Kingdom
P41 GM103311/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2017 Feb;24(2):114-122. doi: 10.1038/nsmb.3343. Epub 2016 Dec 19.
Author Address: Structural Genomics Consortium, University of Oxford, Oxford, UK.
Department of Pharmacology, University of Oxford, Oxford, UK.
Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1311
Author: Gruswitz, F., Chaudhary, S., Ho, J. D., Schlessinger, A., Pezeshki, B., Ho, C. M., Sali, A., Westhoff, C. M. and Stroud, R. M.
Year: 2010
Title: Function of human Rh based on structure of RhCG at 2.1 A
Journal: Proc Natl Acad Sci U S A
Volume: 107
Issue: 21
Pages: 9638-43
Epub Date: 2010/05/12
Date: May 25
Short Title: Function of human Rh based on structure of RhCG at 2.1 A
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1003587107
PMCID: PMC2906887
Accession Number: 20457942
Keywords: Ammonia/metabolism
Biological Transport
Cation Transport Proteins/*chemistry/*metabolism
Cell Line
Crystallography, X-Ray
Erythrocytes/chemistry/metabolism
Humans
Hydrogen-Ion Concentration
Membrane Glycoproteins/*chemistry/*metabolism
Models, Molecular
Protein Structure, Quaternary
Protein Structure, Tertiary
Rh-Hr Blood-Group System/*chemistry/*metabolism
Abstract: In humans, NH(3) transport across cell membranes is facilitated by the Rh (rhesus) family of proteins. Human Rh C glycoprotein (RhCG) forms a trimeric complex that plays an essential role in ammonia excretion and renal pH regulation. The X-ray crystallographic structure of human RhCG, determined at 2.1 A resolution, reveals the mechanism of ammonia transport. Each monomer contains 12 transmembrane helices, one more than in the bacterial homologs. Reconstituted into proteoliposomes, RhCG conducts NH(3) to raise internal pH. Models of the erythrocyte Rh complex based on our RhCG structure suggest that the erythrocytic Rh complex is composed of stochastically assembled heterotrimers of RhAG, RhD, and RhCE.
Notes: 1091-6490
Gruswitz, Franz
Chaudhary, Sarika
Ho, Joseph D
Schlessinger, Avner
Pezeshki, Bobak
Ho, Chi-Min
Sali, Andrej
Westhoff, Connie M
Stroud, Robert M
R37 GM024485/GM/NIGMS NIH HHS/United States
R01 GM024485/GM/NIGMS NIH HHS/United States
GM74929/GM/NIGMS NIH HHS/United States
GM73210/GM/NIGMS NIH HHS/United States
R01 GM24485/GM/NIGMS NIH HHS/United States
P50 GM073210/GM/NIGMS NIH HHS/United States
U54 GM074929/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Proc Natl Acad Sci U S A. 2010 May 25;107(21):9638-43. doi: 10.1073/pnas.1003587107. Epub 2010 May 10.
Author Address: Department of Biochemistry and Biophysics, S412C Genentech Hall, Center for the Structure of Membrane Proteins, and Membrane Protein Expression Center, University of California, San Francisco, CA 94158, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1906
Author: Gu, H., Song, M., Boonanantanasarn, K., Baek, K., Woo, K. M., Ryoo, H. M. and Baek, J. H.
Year: 2018
Title: Conditions Inducing Excessive O-GlcNAcylation Inhibit BMP2-Induced Osteogenic Differentiation of C2C12 Cells
Journal: Int J Mol Sci
Volume: 19
Issue: 1
Epub Date: 2018/01/10
Date: Jan 9
Short Title: Conditions Inducing Excessive O-GlcNAcylation Inhibit BMP2-Induced Osteogenic Differentiation of C2C12 Cells
Alternate Journal: International journal of molecular sciences
ISSN: 1422-0067
DOI: 10.3390/ijms19010202
PMCID: PMC5796151
Accession Number: 29315243
Keywords: Alkaline Phosphatase/genetics/metabolism
Animals
Bone Morphogenetic Protein 2/*pharmacology
Cell Differentiation/*drug effects
Cells, Cultured
Collagen Type I/genetics/metabolism
Core Binding Factor Alpha 1 Subunit/genetics/metabolism
Enzyme Inhibitors/pharmacology
Glucosamine/pharmacology
Glucose/pharmacology
Glycosylation/drug effects
Homeodomain Proteins/genetics/metabolism
Humans
Mice
Myoblasts/cytology/drug effects/metabolism
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/metabolism
Osteogenesis/*drug effects
Periodontal Ligament/cytology/drug effects/metabolism
Recombinant Proteins/pharmacology
Sp7 Transcription Factor/genetics/metabolism
Transcription, Genetic/drug effects
Transforming Growth Factor beta/*pharmacology
Runx2
excessive O-GlcNAcylation
hyperglycemia
osteogenic differentiation
Abstract: Hyperglycemic conditions in diabetic patients can affect various cellular functions, including the modulation of osteogenic differentiation. However, the molecular mechanisms by which hyperglycemia affects osteogenic differentiation are yet to be clarified. This study aimed to investigate whether the aberrant increase in protein O-linked-β-N-acetylglucosamine glycosylation (O-GlcNAcylation) contributes to the suppression of osteogenic differentiation due to hyperglycemia. To induce osteogenic differentiation, C2C12 cells were cultured in the presence of recombinant human bone morphogenetic protein 2 (BMP2). Excessive protein O-GlcNAcylation was induced by treating C2C12 cells with high glucose, glucosamine, or N-acetylglucosamine concentrations or by O-GlcNAc transferase (OGT) overexpression. The effect of O-GlcNAcylation on osteoblast differentiation was then confirmed by examining the expression levels of osteogenic marker gene mRNAs, activity of alkaline phosphatase, and transcriptional activity of Runx2, a critical transcription factor for osteoblast differentiation and bone formation. Cell treatment with high glucose, glucosamine or N-acetylglucosamine increased O-GlcNAcylation of Runx2 and the total levels of O-GlcNAcylated proteins, which led to a decrease in the transcriptional activity of Runx2, expression levels of osteogenic marker genes (Runx2, osterix, alkaline phosphatase, and type I collagen), and activity of alkaline phosphatase. These inhibitory effects were rescued by lowering protein O-GlcNAcylation levels by adding STO45849, an OGT inhibitor, or by overexpressing β-N-acetylglucosaminidase. Our findings suggest that excessive protein O-GlcNAcylation contributes to high glucose-suppressed osteogenic differentiation.
Notes: 1422-0067
Gu, Hanna
Song, Mina
Boonanantanasarn, Kanitsak
Baek, Kyunghwa
Woo, Kyung Mi
Orcid: 0000-0002-2782-1189
Ryoo, Hyun-Mo
Baek, Jeong-Hwa
Orcid: 0000-0002-2032-1683
Journal Article
Int J Mol Sci. 2018 Jan 9;19(1):202. doi: 10.3390/ijms19010202.
Author Address: Department of Molecular Genetics, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 08826, Korea. hnbn9@snu.ac.kr.
Department of Molecular Genetics, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 08826, Korea. 0508mina@gmail.com.
Department of Molecular Genetics, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 08826, Korea. kb57@buffalo.edu.
Department of Pharmacology, College of Dentistry and Research Institute of Oral Science, Gangneung-Wonju National University, Chuncheon 25457, Gangwon-do, Korea. daedanhae@gmail.com.
Department of Molecular Genetics, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 08826, Korea. kmwoo@snu.ac.kr.
Department of Molecular Genetics, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 08826, Korea. hmryoo@snu.ac.kr.
Department of Molecular Genetics, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 08826, Korea. baekjh@snu.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1212
Author: Guhaniyogi, J., Sohar, I., Das, K., Stock, A. M. and Lobel, P.
Year: 2009
Title: Crystal structure and autoactivation pathway of the precursor form of human tripeptidyl-peptidase 1, the enzyme deficient in late infantile ceroid lipofuscinosis
Journal: J Biol Chem
Volume: 284
Issue: 6
Pages: 3985-97
Epub Date: 2008/11/29
Date: Feb 6
Short Title: Crystal structure and autoactivation pathway of the precursor form of human tripeptidyl-peptidase 1, the enzyme deficient in late infantile ceroid lipofuscinosis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M806943200
PMCID: PMC2635056
Accession Number: 19038967
Keywords: Aminopeptidases
Binding Sites/genetics
Cell Line
Crystallography, X-Ray
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases
Endopeptidases/*chemistry/genetics/metabolism
Enzyme Activation/genetics
Enzyme Precursors/*chemistry/genetics/metabolism
Glycosylation
Humans
Models, Molecular
Mutation
Neuronal Ceroid-Lipofuscinoses/*enzymology/genetics
*Protein Folding
Protein Structure, Secondary/genetics
Protein Structure, Tertiary/genetics
Serine Proteases
Structural Homology, Protein
Structure-Activity Relationship
Abstract: Late infantile neuronal ceroid lipofuscinosis is a fatal childhood neurological disorder caused by a deficiency in the lysosomal protease tripeptidyl-peptidase 1 (TPP1). TPP1 represents the only known mammalian member of the S53 family of serine proteases, a group characterized by a subtilisin-like fold, a Ser-Glu-Asp catalytic triad, and an acidic pH optimum. TPP1 is synthesized as an inactive proenzyme (pro-TPP1) that is proteolytically processed into the active enzyme after exposure to low pH in vitro or targeting to the lysosome in vivo. In this study, we describe an endoglycosidase H-deglycosylated form of TPP1 containing four Asn-linked N-acetylglucosamines that is indistinguishable from fully glycosylated TPP1 in terms of autocatalytic processing of the proform and enzymatic properties of the mature protease. The crystal structure of deglycosylated pro-TPP1 was determined at 1.85 angstroms resolution. A large 151-residue C-shaped prodomain makes extensive contacts as it wraps around the surface of the catalytic domain with the two domains connected by a 24-residue flexible linker that passes through the substrate-binding groove. The proenzyme structure reveals suboptimal catalytic triad geometry with its propiece linker partially blocking the substrate-binding site, which together serve to prevent premature activation of the protease. Finally, we have identified numerous processing intermediates and propose a structural model that explains the pathway for TPP1 activation in vitro. These data provide new insights into TPP1 function and represent a valuable resource for constructing improved TPP1 variants for treatment of late infantile neuronal ceroid lipofuscinosis.
Notes: 1083-351x
Guhaniyogi, Jayita
Sohar, Istvan
Das, Kalyan
Stock, Ann M
Lobel, Peter
R01 NS037918/NS/NINDS NIH HHS/United States
S10 RR017992/RR/NCRR NIH HHS/United States
Howard Hughes Medical Institute/United States
NS037918/NS/NINDS NIH HHS/United States
S10 RR020932/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2009 Feb 6;284(6):3985-97. doi: 10.1074/jbc.M806943200. Epub 2008 Nov 26.
Author Address: Center for Advanced Biotechnology and Medicine, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, New Jersey, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 940
Author: Guinez, C., Lemoine, J., Michalski, J. C. and Lefebvre, T.
Year: 2004
Title: 70-kDa-heat shock protein presents an adjustable lectinic activity towards O-linked N-acetylglucosamine
Journal: Biochem Biophys Res Commun
Volume: 319
Issue: 1
Pages: 21-6
Epub Date: 2004/05/26
Date: Jun 18
Short Title: 70-kDa-heat shock protein presents an adjustable lectinic activity towards O-linked N-acetylglucosamine
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/j.bbrc.2004.04.144
Accession Number: 15158436
Keywords: Acetylglucosamine/*chemistry
Blotting, Western
Cell Line
Cell Line, Tumor
Electrophoresis, Polyacrylamide Gel
Glucose/chemistry
Glycosylation
HSP70 Heat-Shock Proteins/*metabolism
Humans
Lectins/*chemistry
Models, Chemical
Precipitin Tests
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Stress, Physiological
Abstract: Numerous works demonstrated that the dynamic O-GlcNAc glycosylation could protect against the proteasomal degradation by modifying the target proteins and the proteasome itself. Considering that Hsp70 is a crucial component in the quality control of protein conformation in the proteasomal pathway, we investigated the possibility that Hsp70 physically interacts with O-GlcNAc proteins through a lectinic activity. First, we demonstrate that in HepG2 cells, Hsp70 can specifically bind to O-GlcNAc residues but also is itself modified by O-GlcNAc. Second, when cells were deprived of glucose (nutrient stress), Hsp70 lectinic activity markedly increased whereas its glycosylation dramatically decreased. On the other hand, a 42 degrees C thermic stress did not affect any of these features. Lastly, the nature of O-GlcNAc modified proteins co-immunoprecipitating with Hsp70 was similar for cells submitted to the thermic and to nutrient stress. These results strongly suggest that O-GlcNAc influences protein stability through specific interaction with 70-kDa-heat shock protein members.
Notes: Guinez, Céline
Lemoine, Jérôme
Michalski, Jean-Claude
Lefebvre, Tony
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2004 Jun 18;319(1):21-6. doi: 10.1016/j.bbrc.2004.04.144.
Author Address: UMR 8576 du CNRS, IFR 118, Unité de Glycobiologie Structurale et Fonctionnelle, Bâtiment C9, Cité Scientifique, 59655 Villeneuve d'Ascq cedex, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1165
Author: Guinez, C., Mir, A. M., Dehennaut, V., Cacan, R., Harduin-Lepers, A., Michalski, J. C. and Lefebvre, T.
Year: 2008
Title: Protein ubiquitination is modulated by O-GlcNAc glycosylation
Journal: Faseb j
Volume: 22
Issue: 8
Pages: 2901-11
Epub Date: 2008/04/25
Date: Aug
Short Title: Protein ubiquitination is modulated by O-GlcNAc glycosylation
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638
DOI: 10.1096/fj.07-102509
Accession Number: 18434435
Keywords: Acetylglucosamine/*metabolism
Base Sequence
Cell Line
Cell Survival
*Glycosylation
HSP70 Heat-Shock Proteins/metabolism
HeLa Cells
Heat-Shock Response
Humans
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/metabolism
Proteasome Endopeptidase Complex/metabolism
Protein Processing, Post-Translational
Proteins/*chemistry/*metabolism
RNA Interference
RNA, Small Interfering/genetics
Ubiquitin-Activating Enzymes/metabolism
*Ubiquitination
Abstract: During the past two decades, O-GlcNAc modification of cytosolic and nuclear proteins has been intensively studied. Nevertheless, the function of this post-translational modification remains unclear. It has been recently speculated that O-GlcNAc could act as a protective signal against proteasomal degradation, both by modifying target substrates and/or by inhibiting the proteasome itself. In this work, we have investigated the putative relation between O-GlcNAc and the ubiquitin pathway. First, we showed that the level of both modifications increased rapidly after thermal stress but, unlike ubiquitinated proteins, O-GlcNAc-modified proteins failed to be stabilized by inhibiting proteasome function. Increasing O-GlcNAc levels, using glucosamine or PUGNAc, enhanced ubiquitination. Inversely, when O-GlcNAc levels were reduced, using forskolin or glucose deprivation, ubiquitination decreased. Targeted-RNA interference of O-GlcNAc transferase also reduced ubiquitination and moreover halved cell thermotolerance. Finally, we demonstrated that the ubiquitin-activating enzyme E1 was O-GlcNAc modified and that its glycosylation and its interaction with Hsp70 varied according to the conditions of cell culture. Altogether, these results show that O-GlcNAc and ubiquitin are not strictly antagonistic post-translational modifications, but rather that the former might regulate the latter, and also suggest that E1 could be one of the common links between the two pathways.
Notes: 1530-6860
Guinez, Céline
Mir, Anne-Marie
Dehennaut, Vanessa
Cacan, René
Harduin-Lepers, Anne
Michalski, Jean-Claude
Lefebvre, Tony
Journal Article
Research Support, Non-U.S. Gov't
United States
FASEB J. 2008 Aug;22(8):2901-11. doi: 10.1096/fj.07-102509. Epub 2008 Apr 23.
Author Address: UMR USTL/CNRS 8576, Laboratoire de Glycobiologie Structurale et Fonctionnelle, IFR 147, 59655 Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 699
Author: Guncar, G., Pungercic, G., Klemencic, I., Turk, V. and Turk, D.
Year: 1999
Title: Crystal structure of MHC class II-associated p41 Ii fragment bound to cathepsin L reveals the structural basis for differentiation between cathepsins L and S
Journal: Embo j
Volume: 18
Issue: 4
Pages: 793-803
Epub Date: 1999/02/18
Date: Feb 15
Short Title: Crystal structure of MHC class II-associated p41 Ii fragment bound to cathepsin L reveals the structural basis for differentiation between cathepsins L and S
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/18.4.793
PMCID: PMC1171172
Accession Number: 10022822
Keywords: Amino Acid Sequence
Antigens, Differentiation, B-Lymphocyte/*chemistry
Binding Sites
Cathepsin L
Cathepsins/*chemistry
Crystallography, X-Ray
Cysteine Endopeptidases
*Endopeptidases
Histocompatibility Antigens Class II/*chemistry
Humans
Kidney/enzymology
Molecular Sequence Data
Peptide Fragments/chemistry
Protease Inhibitors/chemistry
Protein Conformation
Protein Folding
Protein Structure, Secondary
Sequence Alignment
Substrate Specificity
Thyroglobulin/chemistry
Abstract: The lysosomal cysteine proteases cathepsins S and L play crucial roles in the degradation of the invariant chain during maturation of MHC class II molecules and antigen processing. The p41 form of the invariant chain includes a fragment which specifically inhibits cathepsin L but not S. The crystal structure of the p41 fragment, a homologue of the thyroglobulin type-1 domains, has been determined at 2.0 A resolution in complex with cathepsin L. The structure of the p41 fragment demonstrates a novel fold, consisting of two subdomains, each stabilized by disulfide bridges. The first subdomain is an alpha-helix-beta-strand arrangement, whereas the second subdomain has a predominantly beta-strand arrangement. The wedge shape and three-loop arrangement of the p41 fragment bound to the active site cleft of cathepsin L are reminiscent of the inhibitory edge of cystatins, thus demonstrating the first example of convergent evolution observed in cysteine protease inhibitors. However, the different fold of the p41 fragment results in additional contacts with the top of the R-domain of the enzymes, which defines the specificity-determining S2 and S1' substrate-binding sites. This enables inhibitors based on the thyroglobulin type-1 domain fold, in contrast to the rather non-selective cystatins, to exhibit specificity for their target enzymes.
Notes: 1460-2075
Guncar, G
Pungercic, G
Klemencic, I
Turk, V
Turk, D
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1999 Feb 15;18(4):793-803. doi: 10.1093/emboj/18.4.793.
Author Address: Department of Biochemistry and Molecular Biology, Jozcaronef Stefan Institute, Jamova 39, SLO-1000 Ljubljana, Slovenia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1258
Author: Gundry, R. L., Raginski, K., Tarasova, Y., Tchernyshyov, I., Bausch-Fluck, D., Elliott, S. T., Boheler, K. R., Van Eyk, J. E. and Wollscheid, B.
Year: 2009
Title: The mouse C2C12 myoblast cell surface N-linked glycoproteome: identification, glycosite occupancy, and membrane orientation
Journal: Mol Cell Proteomics
Volume: 8
Issue: 11
Pages: 2555-69
Epub Date: 2009/08/07
Date: Nov
Short Title: The mouse C2C12 myoblast cell surface N-linked glycoproteome: identification, glycosite occupancy, and membrane orientation
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M900195-MCP200
PMCID: PMC2773721
Accession Number: 19656770
Keywords: Animals
Aquaporin 1/chemistry
Binding Sites
Cell Differentiation
Cell Membrane/metabolism
Glycoproteins/chemistry
Glycosylation
Membrane Proteins/*metabolism
Mice
Models, Biological
Myoblasts/*metabolism
Proteome/*chemistry
Proteomics/*methods
Sarcoglycans/chemistry
Abstract: Endogenous regeneration and repair mechanisms are responsible for replacing dead and damaged cells to maintain or enhance tissue and organ function, and one of the best examples of endogenous repair mechanisms involves skeletal muscle. Although the molecular mechanisms that regulate the differentiation of satellite cells and myoblasts toward myofibers are not fully understood, cell surface proteins that sense and respond to their environment play an important role. The cell surface capturing technology was used here to uncover the cell surface N-linked glycoprotein subproteome of myoblasts and to identify potential markers of myoblast differentiation. 128 bona fide cell surface-exposed N-linked glycoproteins, including 117 transmembrane, four glycosylphosphatidylinositol-anchored, five extracellular matrix, and two membrane-associated proteins were identified from mouse C2C12 myoblasts. The data set revealed 36 cluster of differentiation-annotated proteins and confirmed the occupancy for 235 N-linked glycosylation sites. The identification of the N-glycosylation sites on the extracellular domain of the proteins allowed for the determination of the orientation of the identified proteins within the plasma membrane. One glycoprotein transmembrane orientation was found to be inconsistent with Swiss-Prot annotations, whereas ambiguous annotations for 14 other proteins were resolved. Several of the identified N-linked glycoproteins, including aquaporin-1 and beta-sarcoglycan, were found in validation experiments to change in overall abundance as the myoblasts differentiate toward myotubes. Therefore, the strategy and data presented shed new light on the complexity of the myoblast cell surface subproteome and reveal new targets for the clinically important characterization of cell intermediates during myoblast differentiation into myotubes.
Notes: 1535-9484
Gundry, Rebekah L
Raginski, Kimberly
Tarasova, Yelena
Tchernyshyov, Irina
Bausch-Fluck, Damaris
Elliott, Steven T
Boheler, Kenneth R
Van Eyk, Jennifer E
Wollscheid, Bernd
R01 HL085434-01A2/HL/NHLBI NIH HHS/United States
P50 HL084946/HL/NHLBI NIH HHS/United States
N01-HV-28180/HV/NHLBI NIH HHS/United States
P50-HL-084946-01/HL/NHLBI NIH HHS/United States
K99 HL094708/HL/NHLBI NIH HHS/United States
P50 HL084946-019003/HL/NHLBI NIH HHS/United States
P50 HL084946-039003/HL/NHLBI NIH HHS/United States
R01 HL085434/HL/NHLBI NIH HHS/United States
K99 HL094708-01/HL/NHLBI NIH HHS/United States
K99-L094708-01/PHS HHS/United States
K99 HL094708-02/HL/NHLBI NIH HHS/United States
N01 HV028180/HV/NHLBI NIH HHS/United States
N01HV28180/HL/NHLBI NIH HHS/United States
P50 HL084946-029003/HL/NHLBI NIH HHS/United States
R00 HL094708/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2009 Nov;8(11):2555-69. doi: 10.1074/mcp.M900195-MCP200. Epub 2009 Aug 4.
Author Address: Department of Medicine, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21224, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1672
Author: Guo, B., Liang, Q., Li, L., Hu, Z., Wu, F., Zhang, P., Ma, Y., Zhao, B., Kovács, A. L., Zhang, Z., Feng, D., Chen, S. and Zhang, H.
Year: 2014
Title: O-GlcNAc-modification of SNAP-29 regulates autophagosome maturation
Journal: Nat Cell Biol
Volume: 16
Issue: 12
Pages: 1215-26
Epub Date: 2014/11/25
Date: Dec
Short Title: O-GlcNAc-modification of SNAP-29 regulates autophagosome maturation
Alternate Journal: Nature cell biology
ISSN: 1465-7392
DOI: 10.1038/ncb3066
Accession Number: 25419848
Keywords: Acetylglucosamine/*metabolism
Animals
Autophagy/*physiology
Autophagy-Related Proteins
Caenorhabditis elegans/metabolism
Caenorhabditis elegans Proteins/genetics/metabolism
Endosomes/*physiology
Green Fluorescent Proteins/genetics
HeLa Cells
Humans
Lysosome-Associated Membrane Glycoproteins
Lysosomes/*physiology
*Membrane Fusion
Mutation
N-Acetylglucosaminyltransferases/genetics/*metabolism
Phagosomes/*physiology
Protein Binding
Proteins/genetics
Q-SNARE Proteins/genetics/metabolism
Qa-SNARE Proteins/genetics
Qb-SNARE Proteins/genetics/*metabolism
Qc-SNARE Proteins/genetics/*metabolism
R-SNARE Proteins/genetics
RNA Interference
RNA, Small Interfering
Signal Transduction/genetics
Vesicular Transport Proteins
Abstract: The mechanism by which nutrient status regulates the fusion of autophagosomes with endosomes/lysosomes is poorly understood. Here, we report that O-linked β-N-acetylglucosamine (O-GlcNAc) transferase (OGT) mediates O-GlcNAcylation of the SNARE protein SNAP-29 and regulates autophagy in a nutrient-dependent manner. In mammalian cells, OGT knockdown, or mutating the O-GlcNAc sites in SNAP-29, promotes the formation of a SNAP-29-containing SNARE complex, increases fusion between autophagosomes and endosomes/lysosomes, and promotes autophagic flux. In Caenorhabditis elegans, depletion of ogt-1 has a similar effect on autophagy; moreover, expression of an O-GlcNAc-defective SNAP-29 mutant facilitates autophagic degradation of protein aggregates. O-GlcNAcylated SNAP-29 levels are reduced during starvation in mammalian cells and in C. elegans. Our study reveals a mechanism by which O-GlcNAc-modification integrates nutrient status with autophagosome maturation.
Notes: 1476-4679
Guo, Bin
Liang, Qianqian
Li, Lin
Hu, Zhe
Wu, Fan
Zhang, Peipei
Ma, Yongfen
Zhao, Bin
Kovács, Attila L
Zhang, Zhiyuan
Feng, Du
Chen, She
Zhang, Hong
P40 OD010440/OD/NIH HHS/United States
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
England
Nat Cell Biol. 2014 Dec;16(12):1215-26. doi: 10.1038/ncb3066. Epub 2014 Nov 24.
Author Address: State Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
National Institute of Biological Sciences, Beijing 102206, China.
Institute of Neurology, Key Laboratory of Age-Associated Cardiac-Cerebral Vascular Disease of Guangdong Province, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, China.
Department of Anatomy, Cell and Developmental Biology, Eötvös Loránd University, Budapest 1117, Hungary.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1489
Author: Guo, K., Gan, L., Zhang, S., Cui, F. J., Cun, W., Li, Y., Kang, N. X., Gao, M. D. and Liu, K. Y.
Year: 2012
Title: Translocation of HSP27 into liver cancer cell nucleus may be associated with phosphorylation and O-GlcNAc glycosylation
Journal: Oncol Rep
Volume: 28
Issue: 2
Pages: 494-500
Epub Date: 2012/06/06
Date: Aug
Short Title: Translocation of HSP27 into liver cancer cell nucleus may be associated with phosphorylation and O-GlcNAc glycosylation
Alternate Journal: Oncology reports
ISSN: 1021-335x
DOI: 10.3892/or.2012.1844
Accession Number: 22664592
Keywords: Acetylglucosamine/*metabolism
Carcinoma, Hepatocellular/genetics/*metabolism
Cell Line, Tumor
Cell Nucleus/metabolism
Enzyme-Linked Immunosorbent Assay
Glycosylation
HSP27 Heat-Shock Proteins/genetics/*metabolism
Heat-Shock Proteins
Humans
Immunoblotting
Liver Neoplasms/genetics/*metabolism
Molecular Chaperones
Phosphorylation
Transfection
Abstract: It has been reported that the dynamic interplay between O-GlcNAcylation and O-phosphorylation is responsible for altering the activity or localization of heat-shock proteins. The aim of this study was to determine whether dynamic interplay between O-GlcNAcylation and O-phosphorylation of HSP27 in hepatocellular cancer (HCC) cells affect its entry into the nucleus. We demonstrate that the entry of HSP27 into the nucleus correlated with its phosphorylation through transfecting HCC cells with plasmids coding for wild-type HSP27 (HSP27-WT), its non-phosphorylatable (HSP27-3A) and pseudophosphorylated (HSP27-3D) mutants, however, not all of the endogenous or exogenous nuclear HSP27 was modified by phosphorylation. We observed that HSP27 was modified with O-GlcNAc glycosylation in HCC cells and report that at conserved Ser residues of HSP27, alternative phosphorylation and O-GlcNAc modification can be predicted by the YinOYang 1.2 method. Furthermore, after P79350 or combined SB203580 and PUGNAc treatment, increased nuclear import of HSP27-WT and HSP27-3D implied that the entry of HSP27 into the nucleus was not only correlated with phosphorylation, but also with O-GlcNAc glycosylation. Collectively, O-GlcNAcylation of HSP27 in HCC cells may be a novel regulatory mode of HSP27 function, particularly for its entry into the nucleus. Crosstalk or interplay between glycosylation and phosphorylation of HSP27 could regulate its subcellular localization and biological functions in liver cancer.
Notes: 1791-2431
Guo, Kun
Gan, Lin
Zhang, Shu
Cui, Feng Jie
Cun, Wang
Li, Yan
Kang, Nan Xiao
Gao, Mei Dong
Liu, Kun Yin
Journal Article
Research Support, Non-U.S. Gov't
Greece
Oncol Rep. 2012 Aug;28(2):494-500. doi: 10.3892/or.2012.1844. Epub 2012 Jun 1.
Author Address: Liver Cancer Institute, Zhongshan Hospital, and Research Center for Cancer, Institute of Biomedical Science, Fudan University, Shanghai 200032, PR China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1161
Author: Gurcel, C., Vercoutter-Edouart, A. S., Fonbonne, C., Mortuaire, M., Salvador, A., Michalski, J. C. and Lemoine, J.
Year: 2008
Title: Identification of new O-GlcNAc modified proteins using a click-chemistry-based tagging
Journal: Anal Bioanal Chem
Volume: 390
Issue: 8
Pages: 2089-97
Epub Date: 2008/03/29
Date: Apr
Short Title: Identification of new O-GlcNAc modified proteins using a click-chemistry-based tagging
Alternate Journal: Analytical and bioanalytical chemistry
ISSN: 1618-2642
DOI: 10.1007/s00216-008-1950-y
Accession Number: 18369606
Keywords: Acetylglucosamine/analogs & derivatives/*analysis/metabolism
Biotin/chemistry
Biotinylation/methods
Cell Line, Tumor
Female
Glycosylation
Humans
*Molecular Probe Techniques
Molecular Probes/*chemistry
Molecular Structure
Protein Processing, Post-Translational
Proteins/*analysis/chemistry/metabolism
Proteomics/*methods
Streptavidin/chemistry
Time Factors
Abstract: The O-linked beta-N-acetylglucosamine (O-GlcNAc) modification is an abundant post-translational modification in eukaryotic cells. This dynamic glycosylation plays a fundamental role in the activity of many nuclear and cytoplasmic proteins and is associated with pathologies like type II diabetes, Alzheimer's disease or some cancers. However the exact link between O-GlcNAc-modified proteins and their function in cells is largely undefined for most cases. Here we report a strategy based on the 1,3-dipolar cycloaddition, called click chemistry, between unnatural N-acetylglucosamine (GlcNAc) analogues (substituted with an azido or alkyne group) and the corresponding biotinylated probe to specifically detect, enrich and identify O-GlcNAc-modified proteins. This bio-orthogonal conjugation confirms that only azido analogue of GlcNAc is metabolized by the cell. Thanks to the biotin probe, affinity purification on streptavidin beads allowed us to identify 32 O-GlcNAc-azido-tagged proteins by LC-MS/MS analysis in an MCF-7 cellular model, 14 of which were previously unreported. This work illustrates the use of the click-chemistry-based strategy combined with a proteomic approach to get further insight into the pattern of O-GlcNAc-modified proteins and the biological significance of this post-translational modification. [figure: see text]
Notes: 1618-2650
Gurcel, Caroline
Vercoutter-Edouart, Anne-Sophie
Fonbonne, Catherine
Mortuaire, Marlène
Salvador, Arnaud
Michalski, Jean-Claude
Lemoine, Jérôme
Journal Article
Germany
Anal Bioanal Chem. 2008 Apr;390(8):2089-97. doi: 10.1007/s00216-008-1950-y. Epub 2008 Mar 28.
Author Address: UMR CNRS 5180, Sciences Analytiques, Université de Lyon, Lyon 1, 43 Bld du 11 novembre 1918, 69622, Villeurbanne cedex, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1990
Author: Gytz, H., Liang, J., Liang, Y., Gorelik, A., Illes, K. and Nagar, B.
Year: 2019
Title: The structure of mammalian β-mannosidase provides insight into β-mannosidosis and nystagmus
Journal: Febs j
Volume: 286
Issue: 7
Pages: 1319-1331
Epub Date: 2018/12/16
Date: Apr
Short Title: The structure of mammalian β-mannosidase provides insight into β-mannosidosis and nystagmus
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/febs.14731
Accession Number: 30552791
Keywords: Amino Acid Sequence
Animals
Catalytic Domain
Glycosylation
Humans
Mannose/*metabolism
Mice
*Mutation
Nystagmus, Pathologic/*enzymology/genetics/pathology
Phenotype
Protein Conformation
Sequence Homology
Substrate Specificity
beta-Mannosidase/*chemistry/genetics/metabolism
beta-Mannosidosis/*enzymology/genetics/pathology
*crystal structure
*enzyme stability
*lysosomal storage disease
*nystagmus
*β-mannosidase
Abstract: β-Mannosidase is a lysosomal enzyme from the glycosyl hydrolase family 2 that cleaves the single β(1-4)-linked mannose at the nonreducing end of N-glycosylated proteins, and plays an important role in the polysaccharide degradation pathway. Mutations in the MANBA gene, which encodes the β-mannosidase, can lead to the lysosomal storage disease β-mannosidosis, as well as nystagmus, an eye condition characterized by involuntary eye movements. Here, we present the first structures of a mammalian β-mannosidase in both the apo- and mannose-bound forms. The structure is similar to previously determined β-mannosidase structures with regard to domain organization and fold, however, there are important differences that underlie substrate specificity between species. Additionally, in contrast to most other ligand-bound β-mannosidases from bacterial and fungal sources where bound sugars were in a boat-like conformation, we find the mannose in the chair conformation. Evaluation of known disease mutations in the MANBA gene provides insight into their impact on disease phenotypes. Together, these results will be important for the design of therapeutics for treating diseases caused by β-mannosidase deficiency. DATABASE: Structural data are available in the Protein Data Bank under the accession numbers 6DDT and 6DDU.
Notes: 1742-4658
Gytz, Heidi
Liang, Jason
Liang, Yingke
Gorelik, Alexei
Orcid: 0000-0001-6236-4633
Illes, Katalin
Nagar, Bhushan
MOP-133535/CIHR/Canada
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2019 Apr;286(7):1319-1331. doi: 10.1111/febs.14731. Epub 2019 Jan 3.
Author Address: Department of Biochemistry, Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1739
Author: Ha, C. and Lim, K.
Year: 2015
Title: O-GlcNAc modification of Sp3 and Sp4 transcription factors negatively regulates their transcriptional activities
Journal: Biochem Biophys Res Commun
Volume: 467
Issue: 2
Pages: 341-7
Epub Date: 2015/10/04
Date: Nov 13
Short Title: O-GlcNAc modification of Sp3 and Sp4 transcription factors negatively regulates their transcriptional activities
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2015.09.155
Accession Number: 26431879
Keywords: Acetylglucosamine/*metabolism
Escherichia coli
Genes, Reporter
HEK293 Cells
HeLa Cells
Humans
Luciferases/genetics/metabolism
N-Acetylglucosaminyltransferases/genetics/metabolism
Plasmids/chemistry/metabolism
*Protein Processing, Post-Translational
Protein Structure, Tertiary
Recombinant Proteins/genetics/metabolism
Serine/metabolism
Signal Transduction
Sp2 Transcription Factor/genetics/metabolism
Sp3 Transcription Factor/genetics/*metabolism
Sp4 Transcription Factor/genetics/*metabolism
Threonine/metabolism
*Transcription, Genetic
Interaction
O-GlcNAc
Sp3
Sp4
Transcription
Abstract: The addition of O-linked N-acetylglucosamine (O-GlcNAc) on serine or threonine modifies a myriad of proteins and regulates their function, stability and localization. O-GlcNAc modification is common among chromosome-associated proteins, such as transcription factors, suggesting its extensive involvement in gene expression regulation. In this study, we demonstrate the O-GlcNAc status of the Sp family members of transcription factors and the functional impact on their transcriptional activities. We highlight the presence of O-GlcNAc residues in Sp3 and Sp4, but not Sp2, as demonstrated by their enrichment in GlcNAc positive protein fractions and by detection of O-GlcNAc residues on Sp3 and Sp4 co-expressed in Escherichia coli together with O-GlcNAc transferase (OGT) using an O-GlcNAc-specific antibody. Deletion mutants of Sp3 and Sp4 indicate that the majority of O-GlcNAc sites reside in their N-terminal transactivation domain. Overall, using reporter gene assays and co-immunoprecipitations, we demonstrate a functional inhibitory role of O-GlcNAc modifications in Sp3 and Sp4 transcription factors. Thereby, our study strengthens the current notion that O-GlcNAc modification is an important regulator of protein interactome.
Notes: 1090-2104
Ha, Changhoon
Lim, Kihong
Journal Article
United States
Biochem Biophys Res Commun. 2015 Nov 13;467(2):341-7. doi: 10.1016/j.bbrc.2015.09.155. Epub 2015 Oct 6.
Author Address: ASAN Institute for Life Science, ASAN Medical Center, Seoul, Republic of Korea.
Center for Vaccine Biology and Immunology, University of Rochester Medical Center, NY 14642, USA. Electronic address: Kihong_Lim@urmc.rochester.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1617
Author: Ha, J. R., Hao, L., Venkateswaran, G., Huang, Y. H., Garcia, E. and Persad, S.
Year: 2014
Title: β-catenin is O-GlcNAc glycosylated at Serine 23: implications for β-catenin's subcellular localization and transactivator function
Journal: Exp Cell Res
Volume: 321
Issue: 2
Pages: 153-66
Epub Date: 2013/12/18
Date: Feb 15
Short Title: β-catenin is O-GlcNAc glycosylated at Serine 23: implications for β-catenin's subcellular localization and transactivator function
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2013.11.021
Accession Number: 24342833
Keywords: Acetylglucosamine/*metabolism
Glycosylation
Humans
Phosphorylation
Protein Binding
*Protein Processing, Post-Translational
Protein Structure, Tertiary
Protein Transport
Serine/*metabolism
Tissue Distribution
Trans-Activators/chemistry/genetics/*metabolism
*Transcriptional Activation/physiology
Tumor Cells, Cultured
beta Catenin/chemistry/genetics/*metabolism
O-GlcNAcylation
Subcellular localization
Transcriptional function
β-Catenin
Abstract: BACKGROUND: We have previously reported that β-catenin is post-translationally modified with a single O-linked attachment of β-N-acetyl-glucosamine (O-GlcNAc). We showed that O-GlcNAc regulated β-catenin's subcellular localization and transcriptional activity. OBJECTIVE: The objectives of this investigation were to identify the putative O-GlcNAc sites of β-catenin and the relevance of identified sites in the regulation of β-catenin's localization and transcriptional activity. METHOD: Missense mutations were introduced to potential O-GlcNAc sites of pEGFP-C2-N-Terminal- or pEGFP-C2-Wild Type-β-catenin by site-directed mutagenesis. We determined the levels of O-GlcNAc-β-catenin, subcellular localization, interaction with binding partners and transcriptional activity of the various constructs. RESULTS: Serine 23 of β-catenin was determined as a site for O-GlcNAc modification which regulated its subcellular distribution, its interactions with cellular partners and consequently its transcriptional activity. SIGNIFICANCE: O-GlcNAcylation of Serine 23 is a novel regulatory modification for β-catenin's subcellular localization and transcriptional activity. This study is the first report to characterize site specific regulation of β-catenin by the O-GlcNAc modification.
Notes: 1090-2422
Ha, Jacqueline R
Hao, Li
Venkateswaran, Geetha
Huang, Yu Hao
Garcia, Elizabeth
Persad, Sujata
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
United States
Exp Cell Res. 2014 Feb 15;321(2):153-66. doi: 10.1016/j.yexcr.2013.11.021. Epub 2013 Dec 14.
Author Address: Department of Pediatrics, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada T6G 2E1.
Department of Pediatrics, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada T6G 2E1. Electronic address: sujata.persad@ualberta.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 943
Author: Hägglund, P., Bunkenborg, J., Elortza, F., Jensen, O. N. and Roepstorff, P.
Year: 2004
Title: A new strategy for identification of N-glycosylated proteins and unambiguous assignment of their glycosylation sites using HILIC enrichment and partial deglycosylation
Journal: J Proteome Res
Volume: 3
Issue: 3
Pages: 556-66
Epub Date: 2004/07/16
Date: May-Jun
Short Title: A new strategy for identification of N-glycosylated proteins and unambiguous assignment of their glycosylation sites using HILIC enrichment and partial deglycosylation
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr034112b
Accession Number: 15253437
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Asparagine/*metabolism
Chromatography, Liquid
Glycosylation
Humans
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase/*metabolism
Mass Spectrometry
Molecular Sequence Data
Plasma/*chemistry
Abstract: Characterization of glycoproteins using mass spectrometry ranges from determination of carbohydrate-protein linkages to the full characterization of all glycan structures attached to each glycosylation site. In a novel approach to identify N-glycosylation sites in complex biological samples, we performed an enrichment of glycosylated peptides through hydrophilic interaction liquid chromatography (HILIC) followed by partial deglycosylation using a combination of endo-beta-N-acetylglucosaminidases (EC 3.2.1.96). After hydrolysis with these enzymes, a single N-acetylglucosamine (GlcNAc) residue remains linked to the asparagine residue. The removal of the major part of the glycan simplifies the MS/MS fragment ion spectra of glycopeptides, while the remaining GlcNAc residue enables unambiguous assignment of the glycosylation site together with the amino acid sequence. We first tested our approach on a mixture of known glycoproteins, and subsequently the method was applied to samples of human plasma obtained by lectin chromatography followed by 1D gel-electrophoresis for determination of 62 glycosylation sites in 37 glycoproteins.
Notes: Hägglund, Per
Bunkenborg, Jakob
Elortza, Felix
Jensen, Ole Nørregaard
Roepstorff, Peter
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2004 May-Jun;3(3):556-66. doi: 10.1021/pr034112b.
Author Address: Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, DK-5230 Odense M, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1488
Author: Hahne, H., Gholami, A. M. and Kuster, B.
Year: 2012
Title: Discovery of O-GlcNAc-modified proteins in published large-scale proteome data
Journal: Mol Cell Proteomics
Volume: 11
Issue: 10
Pages: 843-50
Epub Date: 2012/06/05
Date: Oct
Short Title: Discovery of O-GlcNAc-modified proteins in published large-scale proteome data
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M112.019463
PMCID: PMC3494142
Accession Number: 22661428
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Cell Line
Cell Nucleus/metabolism
Cytoplasm/metabolism
Databases, Protein
Humans
Molecular Sequence Data
Peptides/*analysis
Phosphoproteins/*analysis
Phosphorylation
*Protein Processing, Post-Translational
Proteome/analysis/*metabolism
Serine/metabolism
*Software
Tandem Mass Spectrometry
Threonine/metabolism
Abstract: The attachment of N-acetylglucosamine to serine or threonine residues (O-GlcNAc) is a post-translational modification on nuclear and cytoplasmic proteins with emerging roles in numerous cellular processes, such as signal transduction, transcription, and translation. It is further presumed that O-GlcNAc can exhibit a site-specific, dynamic and possibly functional interplay with phosphorylation. O-GlcNAc proteins are commonly identified by tandem mass spectrometry following some form of biochemical enrichment. In the present study, we assessed if, and to which extent, O-GlcNAc-modified proteins can be discovered from existing large-scale proteome data sets. To this end, we conceived a straightforward O-GlcNAc identification strategy based on our recently developed Oscore software that automatically analyzes tandem mass spectra for the presence and intensity of O-GlcNAc diagnostic fragment ions. Using the Oscore, we discovered hundreds of O-GlcNAc peptides not initially identified in these studies, and most of which have not been described before. Merely re-searching this data extended the number of known O-GlcNAc proteins by almost 100 suggesting that this modification exists even more widely than previously anticipated and the modification is often sufficiently abundant to be detected without enrichment. However, a comparison of O-GlcNAc and phospho-identifications from the very same data indicates that the O-GlcNAc modification is considerably less abundant than phosphorylation. The discovery of numerous doubly modified peptides (i.e. peptides with one or multiple O-GlcNAc or phosphate moieties), suggests that O-GlcNAc and phosphorylation are not necessarily mutually exclusive, but can occur simultaneously at adjacent sites.
Notes: 1535-9484
Hahne, Hannes
Gholami, Amin Moghaddas
Kuster, Bernhard
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2012 Oct;11(10):843-50. doi: 10.1074/mcp.M112.019463. Epub 2012 Jun 1.
Author Address: Center of Life and Food Sciences Weihenstephan, Technische Universität München, Emil-Erlenmeyer-Forum 5, 85354 Freising, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1501
Author: Hahne, H. and Kuster, B.
Year: 2012
Title: Discovery of O-GlcNAc-6-phosphate modified proteins in large-scale phosphoproteomics data
Journal: Mol Cell Proteomics
Volume: 11
Issue: 10
Pages: 1063-9
Epub Date: 2012/07/25
Date: Oct
Short Title: Discovery of O-GlcNAc-6-phosphate modified proteins in large-scale phosphoproteomics data
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M112.019760
PMCID: PMC3494138
Accession Number: 22826440
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Brain/metabolism
Calcium-Calmodulin-Dependent Protein Kinase Type 4/genetics/metabolism
DNA-Binding Proteins/genetics/*metabolism
Embryonic Stem Cells/cytology/metabolism
Humans
Mice
Molecular Sequence Data
N-Acetylglucosaminyltransferases/metabolism
Nerve Tissue Proteins/genetics/*metabolism
Peptides/analysis
Phosphorylation
Pluripotent Stem Cells/cytology/metabolism
*Protein Processing, Post-Translational
Rats
*Software
Synaptic Transmission/genetics
Tandem Mass Spectrometry
Transcription Factors/genetics/*metabolism
Zinc Fingers/genetics
Abstract: Phosphorylated O-GlcNAc is a novel post-translational modification that has so far only been found on the neuronal protein AP180 from the rat (Graham et al., J. Proteome Res. 2011, 10, 2725-2733). Upon collision induced dissociation, the modification generates a highly mass deficient fragment ion (m/z 284.0530) that can be used as a reporter for the identification of phosphorylated O-GlcNAc. Using a publically available mouse brain phosphoproteome data set, we employed our recently developed Oscore software to re-evaluate high resolution/high accuracy tandem mass spectra and discovered the modification on 23 peptides corresponding to 11 mouse proteins. The systematic analysis of 220 candidate phosphoGlcNAc tandem mass spectra as well as a synthetic standard enabled the dissection of the major phosphoGlcNAc fragmentation pathways, suggesting that the modification is O-GlcNAc-6-phosphate. We find that the classical O-GlcNAc modification often exists on the same peptides indicating that O-GlcNAc-6-phosphate may biosynthetically arise in two steps involving the O-GlcNAc transferase and a currently unknown kinase. Many of the identified proteins are involved in synaptic transmission and for Ca(2+)/calmodulin kinase IV, the O-GlcNAc-6-phosphate modification was found in the vicinity of two autophosphorylation sites required for full activation of the kinase suggesting a potential regulatory role for O-GlcNAc-6-phosphate. By re-analyzing mass spectrometric data from human embryonic and induced pluripotent stem cells, our study also identified Zinc finger protein 462 (ZNF462) as the first human O-GlcNAc-6-phosphate modified protein. Collectively, the data suggests that O-GlcNAc-6-phosphate is a general post-translation modification of mammalian proteins with a variety of possible cellular functions.
Notes: 1535-9484
Hahne, Hannes
Kuster, Bernhard
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2012 Oct;11(10):1063-9. doi: 10.1074/mcp.M112.019760. Epub 2012 Jul 23.
Author Address: Center of Life and Food Sciences Weihenstephan, Technische Universität München, Emil-Erlenmeyer-Forum 5, 85354 Freising, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1538
Author: Hahne, H., Sobotzki, N., Nyberg, T., Helm, D., Borodkin, V. S., van Aalten, D. M., Agnew, B. and Kuster, B.
Year: 2013
Title: Proteome wide purification and identification of O-GlcNAc-modified proteins using click chemistry and mass spectrometry
Journal: J Proteome Res
Volume: 12
Issue: 2
Pages: 927-36
Epub Date: 2013/01/11
Date: Feb 1
Short Title: Proteome wide purification and identification of O-GlcNAc-modified proteins using click chemistry and mass spectrometry
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr300967y
PMCID: PMC4946622
NIHMSID: EMS53535
Accession Number: 23301498
Keywords: Acetylglucosamine/*metabolism
Alkynes/chemistry
Azides/chemistry
Chromatography, Liquid
Click Chemistry
Enzyme Inhibitors/pharmacology
HEK293 Cells
Humans
Immobilized Proteins/*isolation & purification/*metabolism
Peptide Fragments/*analysis
*Protein Processing, Post-Translational
Proteome/*isolation & purification/*metabolism
Tandem Mass Spectrometry
Trypsin/chemistry
beta-N-Acetylhexosaminidases/antagonists & inhibitors/metabolism
Abstract: The post-translational modification of proteins with N-acetylglucosamine (O-GlcNAc) is involved in the regulation of a wide variety of cellular processes and associated with a number of chronic diseases. Despite its emerging biological significance, the systematic identification of O-GlcNAc proteins is still challenging. In the present study, we demonstrate a significantly improved O-GlcNAc protein enrichment procedure, which exploits metabolic labeling of cells by azide-modified GlcNAc and copper-mediated Click chemistry for purification of modified proteins on an alkyne-resin. On-resin proteolysis using trypsin followed by LC-MS/MS afforded the identification of around 1500 O-GlcNAc proteins from a single cell line. Subsequent elution of covalently resin bound O-GlcNAc peptides using selective β-elimination enabled the identification of 185 O-GlcNAc modification sites on 80 proteins. To demonstrate the practical utility of the developed approach, we studied the global effects of the O-GlcNAcase inhibitor GlcNAcstatin G on the level of O-GlcNAc modification of cellular proteins. About 200 proteins including several key players involved in the hexosamine signaling pathway showed significantly increased O-GlcNAcylation levels in response to the drug, which further strengthens the link of O-GlcNAc protein modification to cellular nutrient sensing and response.
Notes: 1535-3907
Hahne, Hannes
Sobotzki, Nadine
Nyberg, Tamara
Helm, Dominic
Borodkin, Vladimir S
van Aalten, Daan M F
Agnew, Brian
Kuster, Bernhard
087590/Wellcome Trust/United Kingdom
G0900138/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
J Proteome Res. 2013 Feb 1;12(2):927-36. doi: 10.1021/pr300967y. Epub 2013 Jan 18.
Author Address: Chair for Proteomics and Bioanalytics, Center of Life and Food Sciences, Weihenstephan, Technische Universität München, Freising, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1408
Author: Halim, A., Brinkmalm, G., Rüetschi, U., Westman-Brinkmalm, A., Portelius, E., Zetterberg, H., Blennow, K., Larson, G. and Nilsson, J.
Year: 2011
Title: Site-specific characterization of threonine, serine, and tyrosine glycosylations of amyloid precursor protein/amyloid beta-peptides in human cerebrospinal fluid
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 29
Pages: 11848-53
Epub Date: 2011/06/30
Date: Jul 19
Short Title: Site-specific characterization of threonine, serine, and tyrosine glycosylations of amyloid precursor protein/amyloid beta-peptides in human cerebrospinal fluid
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1102664108
PMCID: PMC3141957
Accession Number: 21712440
Keywords: Alzheimer Disease/metabolism/*physiopathology
Amino Acid Sequence
Amyloid beta-Peptides/genetics/*metabolism
Amyloid beta-Protein Precursor/*metabolism
Chromatography, Liquid
Glycosylation
Humans
Molecular Sequence Data
Proteomics
Serine/*metabolism
Tandem Mass Spectrometry
Threonine/*metabolism
Tyrosine/*metabolism
Abstract: The proteolytic processing of human amyloid precursor protein (APP) into shorter aggregating amyloid β (Aβ)-peptides, e.g., Aβ1-42, is considered a critical step in the pathogenesis of Alzheimer's disease (AD). Although APP is a well-known membrane glycoprotein carrying both N- and O-glycans, nothing is known about the occurrence of released APP/Aβ glycopeptides in cerebrospinal fluid (CSF). We used the 6E10 antibody and immunopurified Aβ peptides and glycopeptides from CSF samples and then liquid chromatography-tandem mass spectrometry for structural analysis using collision-induced dissociation and electron capture dissociation. In addition to 33 unglycosylated APP/Aβ peptides, we identified 37 APP/Aβ glycopeptides with sialylated core 1 like O-glycans attached to Thr(-39, -21, -20, and -13), in a series of APP/AβX-15 glycopeptides, where X was -63, -57, -52, and -45, in relation to Asp1 of the Aβ sequence. Unexpectedly, we also identified a series of 27 glycopeptides, the Aβ1-X series, where X was 20 (DAEFRHDSGYEVHHQKLVFF), 19, 18, 17, 16, and 15, which were all uniquely glycosylated on Tyr10. The Tyr10 linked O-glycans were (Neu5Ac)(1-2)Hex(Neu5Ac)HexNAc-O- structures with the disialylated terminals occasionally O-acetylated or lactonized, indicating a terminal Neu5Acα2,8Neu5Ac linkage. We could not detect any glycosylation of the Aβ1-38/40/42 isoforms. We observed an increase of up to 2.5 times of Tyr10 glycosylated Aβ peptides in CSF in six AD patients compared to seven non-AD patients. APP/Aβ sialylated O-glycans, including that of a Tyr residue, the first in a mammalian protein, may modulate APP processing, inhibiting the amyloidogenic pathway associated with AD.
Notes: 1091-6490
Halim, Adnan
Brinkmalm, Gunnar
Rüetschi, Ulla
Westman-Brinkmalm, Ann
Portelius, Erik
Zetterberg, Henrik
Blennow, Kaj
Larson, Göran
Nilsson, Jonas
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Jul 19;108(29):11848-53. doi: 10.1073/pnas.1102664108. Epub 2011 Jun 28.
Author Address: Department of Clinical Chemistry and Transfusion Medicine, Institute of Biomedicine, University of Gothenburg, SE-413 45 Gothenburg, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1446
Author: Halim, A., Nilsson, J., Rüetschi, U., Hesse, C. and Larson, G.
Year: 2012
Title: Human urinary glycoproteomics; attachment site specific analysis of N- and O-linked glycosylations by CID and ECD
Journal: Mol Cell Proteomics
Volume: 11
Issue: 4
Pages: M111.013649
Epub Date: 2011/12/16
Date: Apr
Short Title: Human urinary glycoproteomics; attachment site specific analysis of N- and O-linked glycosylations by CID and ECD
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M111.013649
PMCID: PMC3322569
Accession Number: 22171320
Keywords: Chromatography, Liquid
Dialysis
Glycopeptides/chemistry/*urine
Glycoproteins/chemistry/*urine
Glycosylation
Humans
Male
N-Acetylneuraminic Acid/chemistry
Proteomics/*methods
Tandem Mass Spectrometry
Abstract: Urine is a complex mixture of proteins and waste products and a challenging biological fluid for biomarker discovery. Previous proteomic studies have identified more than 2800 urinary proteins but analyses aimed at unraveling glycan structures and glycosylation sites of urinary glycoproteins are lacking. Glycoproteomic characterization remains difficult because of the complexity of glycan structures found mainly on asparagine (N-linked) or serine/threonine (O-linked) residues. We have developed a glycoproteomic approach that combines efficient purification of urinary glycoproteins/glycopeptides with complementary MS-fragmentation techniques for glycopeptide analysis. Starting from clinical sample size, we eliminated interfering urinary compounds by dialysis and concentrated the purified urinary proteins by lyophilization. Sialylated urinary glycoproteins were conjugated to a solid support by hydrazide chemistry and trypsin digested. Desialylated glycopeptides, released through mild acid hydrolysis, were characterized by tandem MS experiments utilizing collision induced dissociation (CID) and electron capture dissociation fragmentation techniques. In CID-MS(2), Hex(5)HexNAc(4)-N-Asn and HexHexNAc-O-Ser/Thr were typically observed, in agreement with known N-linked biantennary complex-type and O-linked core 1-like structures, respectively. Additional glycoforms for specific N- and O-linked glycopeptides were also identified, e.g. tetra-antennary N-glycans and fucosylated core 2-like O-glycans. Subsequent CID-MS(3), of selected fragment-ions from the CID-MS(2) analysis, generated peptide specific b- and y-ions that were used for peptide identification. In total, 58 N- and 63 O-linked glycopeptides from 53 glycoproteins were characterized with respect to glycan- and peptide sequences. The combination of CID and electron capture dissociation techniques allowed for the exact identification of Ser/Thr attachment site(s) for 40 of 57 putative O-glycosylation sites. We defined 29 O-glycosylation sites which have, to our knowledge, not been previously reported. This is the first study of human urinary glycoproteins where "intact" glycopeptides were studied, i.e. the presence of glycans and their attachment sites were proven without doubt.
Notes: 1535-9484
Halim, Adnan
Nilsson, Jonas
Rüetschi, Ulla
Hesse, Camilla
Larson, Göran
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2012 Apr;11(4):M111.013649. doi: 10.1074/mcp.M111.013649. Epub 2011 Dec 14.
Author Address: Department of Clinical Chemistry and Transfusion Medicine, Institute of Biomedicine, Sahlgrenska Academy at the University of Gothenburg, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1531
Author: Halim, A., Rüetschi, U., Larson, G. and Nilsson, J.
Year: 2013
Title: LC-MS/MS characterization of O-glycosylation sites and glycan structures of human cerebrospinal fluid glycoproteins
Journal: J Proteome Res
Volume: 12
Issue: 2
Pages: 573-84
Epub Date: 2012/12/14
Date: Feb 1
Short Title: LC-MS/MS characterization of O-glycosylation sites and glycan structures of human cerebrospinal fluid glycoproteins
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr300963h
Accession Number: 23234360
Keywords: Amino Acid Sequence
Apolipoproteins E/chemistry
Cerebrospinal Fluid/*chemistry
Chromatography, Liquid
Glycomics
Glycoproteins/*chemistry
Glycosylation
Hemopexin/chemistry
Humans
Membrane Proteins/chemistry
Molecular Conformation
Molecular Sequence Data
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/chemistry
Polysaccharides/*chemistry
Receptor Protein-Tyrosine Kinases/chemistry
Receptors, G-Protein-Coupled
Serine/*chemistry
Spectrometry, Mass, Electrospray Ionization
Threonine/*chemistry
Trypsin/chemistry
Abstract: The GalNAc O-glycosylation on Ser/Thr residues of extracellular proteins has not been well characterized from a proteomics perspective. We previously reported a sialic acid capture-and-release protocol to enrich tryptic N- and O-glycopeptides from human cerebrospinal fluid glycoproteins using nano-LC-ESI-MS/MS with collision-induced dissociation (CID) for glycopeptide characterization. Here, we have introduced peptide N-glycosidase F (PNGase F) pretreatment of CSF samples to remove the N-glycans facilitating the selective characterization of O-glycopeptides and enabling the use of an automated CID-MS(2)/MS(3) search protocol for glycopeptide identification. We used electron-capture and -transfer dissociation (ECD/ETD) to pinpoint the glycosylation site(s) of the glycopeptides, identified as predominantly core-1-like HexHexNAc-O- structure attached to one to four Ser/Thr residues. We characterized 106 O-glycosylations and found Pro residues preferentially in the n - 1, n + 1, and/or n + 3 positions in relation to the Ser/Thr attachment site (n). The characterization of glycans and glycosylation sites in glycoproteins from human clinical samples provides a basis for future studies addressing the biological and diagnostic importance of specific protein glycosylations in relation to human disease.
Notes: 1535-3907
Halim, Adnan
Rüetschi, Ulla
Larson, Göran
Nilsson, Jonas
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2013 Feb 1;12(2):573-84. doi: 10.1021/pr300963h. Epub 2013 Jan 11.
Author Address: Department of Clinical Chemistry and Transfusion Medicine, Institute of Biomedicine, Sahlgrenska Academy at the University of Gothenburg, 413 45 Gothenburg, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1602
Author: Hall, M. K., Weidner, D. A., Bernetski, C. J. and Schwalbe, R. A.
Year: 2014
Title: N-Linked glycan site occupancy impacts the distribution of a potassium channel in the cell body and outgrowths of neuronal-derived cells
Journal: Biochim Biophys Acta
Volume: 1840
Issue: 1
Pages: 595-604
Epub Date: 2013/10/29
Date: Jan
Short Title: N-Linked glycan site occupancy impacts the distribution of a potassium channel in the cell body and outgrowths of neuronal-derived cells
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbagen.2013.10.025
Accession Number: 24161696
Keywords: Blotting, Western
Cell Adhesion
Cell Membrane/*metabolism
Cell Proliferation
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Neuroblastoma/metabolism/*pathology
Neurons/cytology/*metabolism
Polysaccharides/*metabolism
Shaw Potassium Channels/genetics/*metabolism
Subcellular Fractions
Tumor Cells, Cultured
Cb
Cdg
Dic
Excitable membrane
High density potassium channel cluster
Kv channel
Membrane glycoprotein
Membrane trafficking
N-glycosylation
Neu
Og
Potassium channel
Tirf
cell body
congenital disorder of glycosylation
differential interference contrast
neuraminidase
outgrowth
total internal reflection fluorescence
voltage-gating potassium channel
Abstract: BACKGROUND: Vacancy of occupied N-glycosylation sites of glycoproteins is quite disruptive to a multicellular organism, as underlined by congenital disorders of glycosylation. Since a neuronal component is typically associated with this disease, we evaluated the impact of N-glycosylation processing of a neuronal voltage gated potassium channel, Kv3.1b, expressed in a neuronal-derived cell line, B35 neuroblastoma cells. METHODS: Total internal reflection fluorescence and differential interference contrast microscopy measurements of live B35 cells expressing wild type and glycosylation mutant Kv3.1b proteins were used to evaluate the distribution of the various forms of the Kv3.1b protein in the cell body and outgrowths. Cell adhesion assays were also employed. RESULTS: Microscopy images revealed that occupancy of both N-glycosylation sites of Kv3.1b had relatively similar amounts of Kv3.1b in the outgrowth and cell body while vacancy of one or both sites led to increased accumulation of Kv3.1b in the cell body. Further both the fully glycosylated and partially glycosylated N229Q Kv3.1b proteins formed higher density particles in outgrowths compared to cell body. Cellular assays demonstrated that the distinct spatial arrangements altered cell adhesion properties. CONCLUSIONS: Our findings provide direct evidence that occupancy of the N-glycosylation sites of Kv3.1b contributes significantly to its lateral heterogeneity in membranes of neuronal-derived cells, and in turn alters cellular properties. GENERAL SIGNIFICANCE: Our study demonstrates that N-glycans of Kv3.1b contain information regarding the association, clustering, and distribution of Kv3.1b in the cell membrane, and furthermore that decreased occupancy caused by congenital disorders of glycosylation may alter the biological activity of Kv3.1b.
Notes: Hall, M K
Weidner, D A
Bernetski, C J
Schwalbe, R A
Journal Article
Netherlands
Biochim Biophys Acta. 2014 Jan;1840(1):595-604. doi: 10.1016/j.bbagen.2013.10.025. Epub 2013 Oct 23.
Author Address: Department of Biochemistry and Molecular Biology, Brody School of Medicine at East Carolina University, Greenville, NC 27834, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 660
Author: Haltiwanger, R. S., Grove, K. and Philipsberg, G. A.
Year: 1998
Title: Modulation of O-linked N-acetylglucosamine levels on nuclear and cytoplasmic proteins in vivo using the peptide O-GlcNAc-beta-N-acetylglucosaminidase inhibitor O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino-N-phenylcarbamate
Journal: J Biol Chem
Volume: 273
Issue: 6
Pages: 3611-7
Epub Date: 1998/03/07
Date: Feb 6
Short Title: Modulation of O-linked N-acetylglucosamine levels on nuclear and cytoplasmic proteins in vivo using the peptide O-GlcNAc-beta-N-acetylglucosaminidase inhibitor O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino-N-phenylcarbamate
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.6.3611
Accession Number: 9452489
Keywords: Acetylglucosamine/*analogs & derivatives/*metabolism/pharmacology
Acetylglucosaminidase/*antagonists & inhibitors
Cell Line
Cell Nucleus/*metabolism
Cytoplasm/*metabolism
Enzyme Inhibitors/*pharmacology
Glycosylation
HeLa Cells
Histone Acetyltransferases
Humans
Multienzyme Complexes
Oximes/*pharmacology
*Phenylcarbamates
beta-N-Acetylhexosaminidases
Abstract: O-Linked N-acetylglucosamine (O-GlcNAc) is a ubiquitous and abundant post-translational modification found on nuclear and cytoplasmic proteins and is thought to be a dynamically regulated modification much like phosphorylation. In this study we have demonstrated that O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino-N-phenylcarbama te (PUGNAc), a potent in vitro inhibitor of the enzyme responsible for the removal of O-GlcNAc from proteins (peptide O-GlcNAc-beta-N-acetylglucosaminidase), can be used to increase O-GlcNAc levels on nuclear and cytoplasmic proteins in vivo. Overall, PUGNAc caused approximately a 2-fold increase in O-GlcNAc levels in the human colon cancer cells, HT29, although the effects on individual proteins varied. The increase appeared to be the result of the direct inhibition of the peptide O-GlcNAc-beta-N-acetylglucosaminidase since neither the O-GlcNAc transferase nor UDP-GlcNAc levels were affected by the treatment. O-GlcNAc levels in other cell lines tested (NIH 3T3, CV-1, and HeLa) were also affected by PUGNAc, although the effects on HeLa cells were minimal. At the concentrations tested, PUGNAc was non-toxic and had no affect on the growth rate of any of the cell lines examined. Interestingly, we demonstrated that an increase in O-GlcNAc levels on the transcription factor Sp1 resulted in a reciprocal decrease in its level of phosphorylation, supporting the hypothesis that O-GlcNAc competes with phosphate on some proteins. These studies demonstrate that PUGNAc is an effective inhibitor of O-GlcNAc turnover within cells and can be used to selectively alter the extent of O-GlcNAc on cellular proteins.
Notes: Haltiwanger, R S
Grove, K
Philipsberg, G A
GM 48666/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1998 Feb 6;273(6):3611-7. doi: 10.1074/jbc.273.6.3611.
Author Address: Department of Biochemistry and Cell Biology, Institute for Cell and Developmental Biology, State University of New York, Stony Brook, New York 11794-5215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 639
Author: Haltiwanger, R. S. and Philipsberg, G. A.
Year: 1997
Title: Mitotic arrest with nocodazole induces selective changes in the level of O-linked N-acetylglucosamine and accumulation of incompletely processed N-glycans on proteins from HT29 cells
Journal: J Biol Chem
Volume: 272
Issue: 13
Pages: 8752-8
Epub Date: 1997/03/28
Date: Mar 28
Short Title: Mitotic arrest with nocodazole induces selective changes in the level of O-linked N-acetylglucosamine and accumulation of incompletely processed N-glycans on proteins from HT29 cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.13.8752
Accession Number: 9079710
Keywords: Acetylglucosamine/*metabolism
Amidohydrolases/metabolism
Antineoplastic Agents/*pharmacology
Electrophoresis, Polyacrylamide Gel
Glycosylation
HT29 Cells
Humans
Keratins/metabolism
Membrane Glycoproteins/metabolism
Mitosis/*drug effects
Neoplasm Proteins/*metabolism
Nocodazole/*pharmacology
Nuclear Pore Complex Proteins
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Polysaccharides/*metabolism
Sp1 Transcription Factor/metabolism
Abstract: O-Linked N-acetylglucosamine (O-GlcNAc) is a ubiquitous and abundant protein modification found on nuclear and cytoplasmic proteins. Several lines of evidence suggest that it is a highly dynamic modification and that the levels of this sugar on proteins may be regulated. Previous workers (Chou, C. F., and Omary, M. B. (1993) J. Biol. Chem. 268, 4465-4472) have shown that mitotic arrest with microtubule-destabilizing agents such as nocodazole causes an increase in the O-GlcNAc levels on keratins in the human colon cancer cell line HT29. We have sought to determine whether this increase in glycosylation is a general (i.e. occurring on many proteins) or a limited (i.e. occurring only on the keratins) process. A general increase would suggest that the microtubule-destabilizing agents were somehow affecting the enzymes responsible for addition and/or removal of O-GlcNAc. Our results suggest that the changes in O-GlcNAc induced by nocodazole are selective for the keratins. The levels of O-GlcNAc on other proteins, including the nuclear pore protein p62 and the transcription factor Sp1, are not significantly affected by this treatment. In agreement with these findings, nocodazole treatment caused no change in the activity of the enzymes responsible for addition or removal of O-GlcNAc as determined by direct in vitro assay. Interestingly, nocodazole treatment did cause a dramatic increase in modification of N-glycans with terminal GlcNAc residues on numerous proteins. Potential mechanisms for this and the change in glycosylation of the keratins are discussed.
Notes: Haltiwanger, R S
Philipsberg, G A
GM 48666/GM/NIGMS NIH HHS/United States
RR02427/RR/NCRR NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1997 Mar 28;272(13):8752-8. doi: 10.1074/jbc.272.13.8752.
Author Address: Department of Biochemistry and Cell Biology, Institute for Cell and Developmental Biology, State University of New York, Stony Brook, New York 11794-5215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 802
Author: Hamilton, S. R., Yao, S. Y., Ingram, J. C., Hadden, D. A., Ritzel, M. W., Gallagher, M. P., Henderson, P. J., Cass, C. E., Young, J. D. and Baldwin, S. A.
Year: 2001
Title: Subcellular distribution and membrane topology of the mammalian concentrative Na+-nucleoside cotransporter rCNT1
Journal: J Biol Chem
Volume: 276
Issue: 30
Pages: 27981-8
Epub Date: 2001/05/29
Date: Jul 27
Short Title: Subcellular distribution and membrane topology of the mammalian concentrative Na+-nucleoside cotransporter rCNT1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M100518200
Accession Number: 11375981
Keywords: Amino Acid Motifs
Animals
Asparagine/chemistry
Biological Transport
Blotting, Western
CHO Cells
Carrier Proteins/*chemistry/*physiology
Cell Membrane/metabolism
Cricetinae
DNA, Complementary/metabolism
Gene Deletion
Glycosylation
Humans
Immunohistochemistry
Intestinal Mucosa/metabolism
Kidney/metabolism
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase/pharmacology
*Membrane Transport Proteins
Mutagenesis, Site-Directed
Mutation
Oocytes/metabolism
Protein Structure, Secondary
Protein Structure, Tertiary
Rats
Recombinant Proteins/metabolism
Tissue Distribution
Transcription, Genetic
Transfection
Uridine/metabolism
Xenopus
Abstract: The rat transporter rCNT1 is the archetype of a family of concentrative nucleoside transporters (CNTs) found both in eukaryotes and in prokaryotes. In the present study we have used antibodies to investigate the subcellular distribution and membrane topology of this protein. rCNT1 was found to be expressed predominantly in the brush-border membranes of the polarized epithelial cells of rat jejunum and renal cortical tubules and in the bile canalicular membranes of liver parenchymal cells, consistent with roles in the absorption of dietary nucleosides, of nucleosides in the glomerular filtrate, or of nucleosides arising from the action of extracellular nucleotidases, respectively. The effect of endoglycosidase F treatment on wild-type and mutant rCNT1 expressed in Xenopus oocytes revealed that the recombinant transporter could be glycosylated at either or both of Asn605 and Asn643, indicating that its C terminus is extracellular. In contrast, potential N-glycosylation sites introduced near the N terminus, or between putative transmembrane (TM) helices 4 and 5, were not glycosylated. The deduced orientation of the N terminus in the cytoplasm was confirmed by immunocytochemistry on intact and saponin-permeabilized Chinese hamster ovary cells expressing recombinant rCNT1. These results, in conjunction with extensive analyses of CNT family protein sequences using predictive algorithms, lead us to propose a revised topological model, in which rCNT1 possesses 13 TM helices with the hydrophilic N-terminal and C-terminal domains on the cytoplasmic and extracellular sides of the membrane, respectively. Furthermore, we show that the first three TM helices, which are absent from prokaryote CNTs, are not essential for transporter function; truncated proteins lacking these helices, derived either from rCNT1 or from its human homolog hCNT1, were found to retain significant sodium-dependent uridine transport activity when expressed in oocytes.
Notes: Hamilton, S R
Yao, S Y
Ingram, J C
Hadden, D A
Ritzel, M W
Gallagher, M P
Henderson, P J
Cass, C E
Young, J D
Baldwin, S A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2001 Jul 27;276(30):27981-8. doi: 10.1074/jbc.M100518200. Epub 2001 May 25.
Author Address: School of Biochemistry and Molecular Biology, University of Leeds, Leeds LS2 9JT, the United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1450
Author: Hamming, O. J., Kang, L., Svensson, A., Karlsen, J. L., Rahbek-Nielsen, H., Paludan, S. R., Hjorth, S. A., Bondensgaard, K. and Hartmann, R.
Year: 2012
Title: Crystal structure of interleukin-21 receptor (IL-21R) bound to IL-21 reveals that sugar chain interacting with WSXWS motif is integral part of IL-21R
Journal: J Biol Chem
Volume: 287
Issue: 12
Pages: 9454-60
Epub Date: 2012/01/12
Date: Mar 16
Short Title: Crystal structure of interleukin-21 receptor (IL-21R) bound to IL-21 reveals that sugar chain interacting with WSXWS motif is integral part of IL-21R
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.311084
PMCID: PMC3308775
Accession Number: 22235133
Keywords: Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Glycosylation
Humans
Interleukins/*chemistry/genetics/*metabolism
Mannose/metabolism
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Secondary
Receptors, Interleukin-21/*chemistry/genetics/*metabolism
Abstract: IL-21 is a class I cytokine that exerts pleiotropic effects on both innate and adaptive immune responses. It signals through a heterodimeric receptor complex consisting of the IL-21 receptor (IL-21R) and the common γ-chain. A hallmark of the class I cytokine receptors is the class I cytokine receptor signature motif (WSXWS). The exact role of this motif has not been determined yet; however, it has been implicated in diverse functions, including ligand binding, receptor internalization, proper folding, and export, as well as signal transduction. Furthermore, the WXXW motif is known to be a consensus sequence for C-mannosylation. Here, we present the crystal structure of IL-21 bound to IL-21R and reveal that the WSXWS motif of IL-21R is C-mannosylated at the first tryptophan. We furthermore demonstrate that a sugar chain bridges the two fibronectin domains that constitute the extracellular domain of IL-21R and anchors at the WSXWS motif through an extensive hydrogen bonding network, including mannosylation. The glycan thus transforms the V-shaped receptor into an A-frame. This finding offers a novel structural explanation of the role of the class I cytokine signature motif.
Notes: 1083-351x
Hamming, Ole J
Kang, Lishan
Svensson, Anders
Karlsen, Jesper L
Rahbek-Nielsen, Henrik
Paludan, Søren R
Hjorth, Siv A
Bondensgaard, Kent
Hartmann, Rune
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Mar 16;287(12):9454-60. doi: 10.1074/jbc.M111.311084. Epub 2012 Jan 10.
Author Address: Department of Molecular Biology and Genetics, Aarhus University, Aarhus C, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1892
Author: Han, C., Gu, Y., Shan, H., Mi, W., Sun, J., Shi, M., Zhang, X., Lu, X., Han, F., Gong, Q. and Yu, W.
Year: 2017
Title: O-GlcNAcylation of SIRT1 enhances its deacetylase activity and promotes cytoprotection under stress
Journal: Nat Commun
Volume: 8
Issue: 1
Pages: 1491
Epub Date: 2017/11/15
Date: Nov 14
Short Title: O-GlcNAcylation of SIRT1 enhances its deacetylase activity and promotes cytoprotection under stress
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-017-01654-6
PMCID: PMC5684413
Accession Number: 29133780
Keywords: Acetylation
Acetylglucosamine/*metabolism
Animals
Cell Line
Cell Survival
*Cytoprotection
Enzyme Activation
Female
Humans
Life Expectancy
Mice, Inbred BALB C
*Oxidative Stress
Protein Binding
Serine/metabolism
Sirtuin 1/chemistry/*metabolism
Abstract: SIRT1 is the most evolutionarily conserved mammalian sirtuin, and it plays a vital role in the regulation of metabolism, stress responses, genome stability, and ageing. As a stress sensor, SIRT1 deacetylase activity is significantly increased during stresses, but the molecular mechanisms are not yet fully clear. Here, we show that SIRT1 is dynamically modified with O-GlcNAc at Ser 549 in its carboxy-terminal region, which directly increases its deacetylase activity both in vitro and in vivo. The O-GlcNAcylation of SIRT1 is elevated during genotoxic, oxidative, and metabolic stress stimuli in cellular and mouse models, thereby increasing SIRT1 deacetylase activity and protecting cells from stress-induced apoptosis. Our findings demonstrate a new mechanism for the activation of SIRT1 under stress conditions and suggest a novel potential therapeutic target for preventing age-related diseases and extending healthspan.
Notes: 2041-1723
Han, Cuifang
Gu, Yuchao
Orcid: 0000-0002-8723-3899
Shan, Hui
Mi, Wenyi
Sun, Jiahui
Shi, Minghui
Zhang, Xinling
Lu, Xinzhi
Han, Feng
Orcid: 0000-0001-5706-7028
Gong, Qianhong
Yu, Wengong
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2017 Nov 14;8(1):1491. doi: 10.1038/s41467-017-01654-6.
Author Address: School of Medicine and Pharmacy, Ocean University of China, 5 Yushan Road, Qingdao, 266003, China.
Key Laboratory of Marine Drugs, Ministry of Education, Qingdao, 266003, China.
Key Laboratory of Glycoscience & Glycotechnology of Shandong Province, Qingdao, 266003, China.
School of Medicine and Pharmacy, Ocean University of China, 5 Yushan Road, Qingdao, 266003, China. guych@ouc.edu.cn.
Laboratory for Marine Drugs and Bioproducts of Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266200, China. guych@ouc.edu.cn.
Key Laboratory of Marine Drugs, Ministry of Education, Qingdao, 266003, China. guych@ouc.edu.cn.
Key Laboratory of Glycoscience & Glycotechnology of Shandong Province, Qingdao, 266003, China. guych@ouc.edu.cn.
Laboratory for Marine Drugs and Bioproducts of Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266200, China.
School of Medicine and Pharmacy, Ocean University of China, 5 Yushan Road, Qingdao, 266003, China. yuwg66@ouc.edu.cn.
Laboratory for Marine Drugs and Bioproducts of Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266200, China. yuwg66@ouc.edu.cn.
Key Laboratory of Marine Drugs, Ministry of Education, Qingdao, 266003, China. yuwg66@ouc.edu.cn.
Key Laboratory of Glycoscience & Glycotechnology of Shandong Province, Qingdao, 266003, China. yuwg66@ouc.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 640
Author: Han, I. and Kudlow, J. E.
Year: 1997
Title: Reduced O glycosylation of Sp1 is associated with increased proteasome susceptibility
Journal: Mol Cell Biol
Volume: 17
Issue: 5
Pages: 2550-8
Epub Date: 1997/05/01
Date: May
Short Title: Reduced O glycosylation of Sp1 is associated with increased proteasome susceptibility
Alternate Journal: Molecular and cellular biology
ISSN: 0270-7306 (Print)
0270-7306
DOI: 10.1128/mcb.17.5.2550
PMCID: PMC232104
Accession Number: 9111324
Keywords: Animals
Cell Line
Colforsin/pharmacology
Cysteine Endopeptidases/*metabolism
DNA/metabolism
Glucosamine/biosynthesis
Glucose/metabolism
Glycosylation
Multienzyme Complexes/*metabolism
Protease Inhibitors/pharmacology
Proteasome Endopeptidase Complex
Rats
Sp1 Transcription Factor/*metabolism
Abstract: Sp1 is a ubiquitously expressed transcription factor that is particularly important for the regulation of TATA-less genes that encode housekeeping proteins. Most growth factors and receptors are also encoded by such genes. Sp1 is multiply O glycosylated by covalent linkage of the monosaccharide N-acetylglucosamine (O-GlcNAc) to serine and threonine residues. Based on an earlier observation that growth factor gene transcription can be regulated by glucose and glucosamine in vascular smooth muscle cells, we determined whether Sp1 glycosylation could be regulated and if this modification altered Sp1 function. We found that Sp1 becomes hyperglycosylated when cells are exposed to 5 mM glucosamine, whereas under glucose starvation, stimulation with cyclic AMP (cAMP) results in nearly complete deglycosylation of this protein. Correlating with this hypoglycosylated state, Sp1 is rapidly proteolytically degraded by an enzyme(s) that can be inhibited by specific proteasome inhibitors, lactacystin and LLnL. Treatment of cells with glucose or glucosamine protects Sp1 from cAMP-mediated degradation, whereas blockade of glucosamine synthesis abrogates glucose but not glucosamine protection. This effect on Sp1 is specific, in that the Stat-3 and E2F transcription factors did not undergo degradation under these conditions. The O-GlcNAc modification of Sp1 may play a role as a nutritional checkpoint. In the absence of adequate nutrition, Sp1 becomes hypoglycosylated and thereby subject to proteasome degradation. This process could potentially result in reduced general transcription, thereby conserving nutrients.
Notes: 1098-5549
Han, I
Kudlow, J E
DK43652/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Mol Cell Biol. 1997 May;17(5):2550-8. doi: 10.1128/MCB.17.5.2550.
Author Address: Department of Medicine, University of Alabama at Birmingham, 35294, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 767
Author: Han, I., Oh, E. S. and Kudlow, J. E.
Year: 2000
Title: Responsiveness of the state of O-linked N-acetylglucosamine modification of nuclear pore protein p62 to the extracellular glucose concentration
Journal: Biochem J
Volume: 350 Pt 1
Issue: Pt 1
Pages: 109-14
Epub Date: 2000/08/06
Date: Aug 15
Short Title: Responsiveness of the state of O-linked N-acetylglucosamine modification of nuclear pore protein p62 to the extracellular glucose concentration
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
PMCID: PMC1221231
Accession Number: 10926833
Keywords: Acetylglucosamine/*metabolism
Animals
Cell Line
Extracellular Space/metabolism
Glucosamine/metabolism
Glucose/*metabolism
Glycosylation
Membrane Glycoproteins/*metabolism
Nuclear Pore Complex Proteins
Rats
Abstract: O-linked N-acetylglucosamine (O-GlcNAc) modification has been described in many proteins, including nuclear pore glycoproteins. In the present study we investigated the effect of extracellular glucose on the level of modification of nuclear pore protein p62 by O-GlcNAc. We found that exposure of cells to a high concentration of glucose caused an increased level of modification of p62 with O-GlcNAc, whereas the modification of other proteins did not change detectably. The increased O-GlcNAc modification of p62 in response to glucose required the metabolism of glucose to glucosamine. The exposure of the cells to glucosamine itself also caused increased O-GlcNAc modification, whereas mannosamine and galactosamine did not. Furthermore, changes in glucose concentrations within the physiological range induced the O-GlcNAc modification in p62 in rat aortic smooth-muscle cells, indicating that this modification of p62 might occur in an intact animal. These results imply that the ambient glucose concentration has an impact on the level of O-GlcNAc modification of proteins such as p62, and that functional changes in some of these proteins might ensue.
Notes: 1470-8728
Han, I
Oh, E S
Kudlow, J E
DK55262/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Biochem J. 2000 Aug 15;350 Pt 1(Pt 1):109-14.
Author Address: Departments of Medicine and Cell Biology, Division of Endocrinology and Metabolism, University of Alabama at Birmingham, Birmingham AL 35294, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1461
Author: Han, J., Kim, H. J., Lee, S. C., Hong, S., Park, K., Jeon, Y. H., Kim, D., Cheong, H. K. and Kim, H. S.
Year: 2012
Title: Structure-based rational design of a Toll-like receptor 4 (TLR4) decoy receptor with high binding affinity for a target protein
Journal: PLoS One
Volume: 7
Issue: 2
Pages: e30929
Epub Date: 2012/03/01
Short Title: Structure-based rational design of a Toll-like receptor 4 (TLR4) decoy receptor with high binding affinity for a target protein
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0030929
PMCID: PMC3281905
Accession Number: 22363519
Keywords: Crystallography, X-Ray
*Drug Design
Humans
Hydrogen Bonding
Hydrophobic and Hydrophilic Interactions
Lymphocyte Antigen 96/chemistry/*metabolism
Molecular Dynamics Simulation
Mutant Proteins/metabolism
Mutation/genetics
Protein Binding
Recombinant Fusion Proteins/*chemistry/*metabolism
Thermodynamics
Abstract: Repeat proteins are increasingly attracting much attention as alternative scaffolds to immunoglobulin antibodies due to their unique structural features. Nonetheless, engineering interaction interface and understanding molecular basis for affinity maturation of repeat proteins still remain a challenge. Here, we present a structure-based rational design of a repeat protein with high binding affinity for a target protein. As a model repeat protein, a Toll-like receptor4 (TLR4) decoy receptor composed of leucine-rich repeat (LRR) modules was used, and its interaction interface was rationally engineered to increase the binding affinity for myeloid differentiation protein 2 (MD2). Based on the complex crystal structure of the decoy receptor with MD2, we first designed single amino acid substitutions in the decoy receptor, and obtained three variants showing a binding affinity (K(D)) one-order of magnitude higher than the wild-type decoy receptor. The interacting modes and contributions of individual residues were elucidated by analyzing the crystal structures of the single variants. To further increase the binding affinity, single positive mutations were combined, and two double mutants were shown to have about 3000- and 565-fold higher binding affinities than the wild-type decoy receptor. Molecular dynamics simulations and energetic analysis indicate that an additive effect by two mutations occurring at nearby modules was the major contributor to the remarkable increase in the binding affinities.
Notes: 1932-6203
Han, Jieun
Kim, Hyun Jung
Lee, Sang-Chul
Hong, Seungpyo
Park, Keunwan
Jeon, Young Ho
Kim, Dongsup
Cheong, Hae-Kap
Kim, Hak-Sung
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2012;7(2):e30929. doi: 10.1371/journal.pone.0030929. Epub 2012 Feb 17.
Author Address: Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Yuseong-gu, Daejeon, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1863
Author: Han, J. W., Valdez, J. L., Ho, D. V., Lee, C. S., Kim, H. M., Wang, X., Huang, L. and Chan, J. Y.
Year: 2017
Title: Nuclear factor-erythroid-2 related transcription factor-1 (Nrf1) is regulated by O-GlcNAc transferase
Journal: Free Radic Biol Med
Volume: 110
Pages: 196-205
Epub Date: 2017/06/20
Date: Sep
Short Title: Nuclear factor-erythroid-2 related transcription factor-1 (Nrf1) is regulated by O-GlcNAc transferase
Alternate Journal: Free radical biology & medicine
ISSN: 0891-5849
DOI: 10.1016/j.freeradbiomed.2017.06.008
Accession Number: 28625484
Keywords: Acetylglucosamine/analogs & derivatives/pharmacology
Amino Acid Sequence
Binding Sites
Cloning, Molecular
Gene Expression
Glycosylation
HEK293 Cells
Host Cell Factor C1/chemistry/genetics/*metabolism
Humans
N-Acetylglucosaminyltransferases/chemistry/genetics/*metabolism
Nuclear Respiratory Factor 1/chemistry/genetics/*metabolism
Oximes/pharmacology
Phenylcarbamates/pharmacology
Plasmids/chemistry/metabolism
Protein Binding
Protein Interaction Domains and Motifs
*Protein Processing, Post-Translational
Protein Stability
Proteolysis
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Transcriptional Activation/drug effects
Transfection
Ubiquitination
*O-GlcNAcylation
*Ogt
*Oxidative stress
*Protein stability
*Transcription factor
Abstract: The Nrf1 (Nuclear factor E2-related factor 1) transcription factor performs a critical role in regulating cellular homeostasis. Using a proteomic approach, we identified Host Cell Factor-1 (HCF1), a co-regulator of transcription, and O-GlcNAc transferase (OGT), the enzyme that mediates protein O-GlcNAcylation, as cellular partners of Nrf1a, an isoform of Nrf1. Nrf1a directly interacts with HCF1 through the HCF1 binding motif (HBM), while interaction with OGT is mediated through HCF1. Overexpression of HCF1 and OGT leads to increased Nrf1a protein stability. Addition of O-GlcNAc decreases ubiquitination and degradation of Nrf1a. Transcriptional activation by Nrf1a is increased by OGT overexpression and treatment with PUGNAc. Together, these data suggest that OGT can act as a regulator of Nrf1a.
Notes: 1873-4596
Han, Jeong Woo
Valdez, Joshua L
Ho, Daniel V
Lee, Candy S
Kim, Hyun Min
Wang, Xiaorong
Huang, Lan
Chan, Jefferson Y
Journal Article
Research Support, N.I.H., Extramural
United States
Free Radic Biol Med. 2017 Sep;110:196-205. doi: 10.1016/j.freeradbiomed.2017.06.008. Epub 2017 Jun 15.
Author Address: Department of Laboratory Medicine and Pathology, University of California, Irvine, D440 Medical Sciences, Irvine, CA 92697, USA.
Departments of Physiology and Biophysics, University of California, Irvine, D440 Medical Sciences, Irvine, CA 92697, USA.
Department of Laboratory Medicine and Pathology, University of California, Irvine, D440 Medical Sciences, Irvine, CA 92697, USA. Electronic address: jchan@uci.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1760
Author: Han, L., Nishimura, K., Sadat Al Hosseini, H., Bianchi, E., Wright, G. J. and Jovine, L.
Year: 2016
Title: Divergent evolution of vitamin B9 binding underlies Juno-mediated adhesion of mammalian gametes
Journal: Curr Biol
Volume: 26
Issue: 3
Pages: R100-1
Epub Date: 2016/02/10
Date: Feb 8
Short Title: Divergent evolution of vitamin B9 binding underlies Juno-mediated adhesion of mammalian gametes
Alternate Journal: Current biology : CB
ISSN: 0960-9822 (Print)
0960-9822
DOI: 10.1016/j.cub.2015.12.034
PMCID: PMC4751342
Accession Number: 26859261
Keywords: Animals
*Evolution, Molecular
Folic Acid/*metabolism
Male
Mice/*genetics/metabolism
Oocytes/metabolism
Receptors, Cell Surface/*genetics/metabolism
*Sperm-Ovum Interactions
Spermatozoa/metabolism
Vitamin B Complex/*metabolism
Abstract: The interaction between egg and sperm is the first necessary step of fertilization in all sexually reproducing organisms. A decade-long search for a protein pair mediating this event in mammals culminated in the identification of the glycosylphosphatidylinositol (GPI)-anchored glycoprotein Juno as the egg plasma membrane receptor of sperm Izumo1 [1,2]. The Juno-Izumo1 interaction was shown to be essential for fertilization since mice lacking either gene exhibit sex-specific sterility, making these proteins promising non-hormonal contraceptive targets [1,3]. No structural information is available on how gamete membranes interact at fertilization, and it is unclear how Juno - which was previously named folate receptor (FR) 4, based on sequence similarity considerations - triggers membrane adhesion by binding Izumo1. Here, we report the crystal structure of Juno and find that the overall fold is similar to that of FRα and FRβ but with significant flexibility within the area that corresponds to the rigid ligand-binding site of these bona fide folate receptors. This explains both the inability of Juno to bind vitamin B9/folic acid [1], and why mutations within the flexible region can either abolish or change the species specificity of this interaction. Furthermore, structural similarity between Juno and the cholesterol-binding Niemann-Pick disease type C1 protein (NPC1) suggests how the modified binding surface of Juno may recognize the helical structure of the amino-terminal domain of Izumo1. As Juno appears to be a mammalian innovation, our study indicates that a key evolutionary event in mammalian reproduction originated from the neofunctionalization of the vitamin B9-binding pocket of an ancestral folate receptor molecule.
Notes: 1879-0445
Han, Ling
Nishimura, Kaoru
Sadat Al Hosseini, Hamed
Bianchi, Enrica
Wright, Gavin J
Jovine, Luca
260759/European Research Council/International
098051/Wellcome Trust/United Kingdom
MR/M012468/1/Medical Research Council/United Kingdom
Letter
Research Support, Non-U.S. Gov't
Curr Biol. 2016 Feb 8;26(3):R100-1. doi: 10.1016/j.cub.2015.12.034.
Author Address: Department of Biosciences and Nutrition & Center for Innovative Medicine, Karolinska Institutet, Huddinge, SE-141 83, Sweden.
Cell Surface Signalling Laboratory, Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, UK.
Department of Biosciences and Nutrition & Center for Innovative Medicine, Karolinska Institutet, Huddinge, SE-141 83, Sweden. Electronic address: luca.jovine@ki.se.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 674
Author: Hanisch, F. G., Green, B. N., Bateman, R. and Peter-Katalinic, J.
Year: 1998
Title: Localization of O-glycosylation sites of MUC1 tandem repeats by QTOF ESI mass spectrometry
Journal: J Mass Spectrom
Volume: 33
Issue: 4
Pages: 358-62
Epub Date: 1998/05/23
Date: Apr
Short Title: Localization of O-glycosylation sites of MUC1 tandem repeats by QTOF ESI mass spectrometry
Alternate Journal: Journal of mass spectrometry : JMS
ISSN: 1076-5174 (Print)
1076-5174
DOI: 10.1002/(sici)1096-9888(199804)33:4<358::aid-jms642>3.0.co;2-3
Accession Number: 9597769
Keywords: Amino Acid Sequence
Glycosylation
Molecular Sequence Data
Mucin-1/*analysis/*chemistry/metabolism
Protein Processing, Post-Translational
*Spectrometry, Mass, Secondary Ion
Abstract: The potential of electrospray mass spectrometry (ESMS) for the sequencing of glycopeptides was evaluated using quadrupole time-of-flight (QTOF) technology in the MS/MS mode. The location of O-glycosylation sites was possible in the positive ion (+) mode by detection of prominent y- and b-fragment ions from the underivatized TAP25-2 [T1APPAHGVT9S10APDT14RPAPGS20T21APPA], an overlapping sequence of MUC1 tandem repeats which had been glycosylated in vitro by two GalNAc residues in the positions T9 and T21. The high mass resolution and accuracy of QTOF-(+)ESMS allowed reliable structural assignments. The reduced complexity of the fragment spectra and the higher signal-to-noise ratio render QTOF-(+)ESMS an alternative mass spectrometric approach to the identification of O-glycosylation sites when compared with sequencing by post-source decay matrix-assisted laser desorption/ionization MS. Diagnostic ions from the N-terminus in the b-series offered direct evidence, which was supported by indirect evidence from the C-terminus ions of the y-series. The higher glycosylated GalNAc2-substituted fragments were mainly observed as multiply ionized species.
Notes: Hanisch, F G
Green, B N
Bateman, R
Peter-Katalinic, J
Journal Article
Research Support, Non-U.S. Gov't
England
J Mass Spectrom. 1998 Apr;33(4):358-62. doi: 10.1002/(SICI)1096-9888(199804)33:4<358::AID-JMS642>3.0.CO;2-3.
Author Address: Institut für Biochemie, Universität Köln, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 692
Author: Haniu, M., Arakawa, T., Bures, E. J., Young, Y., Hui, J. O., Rohde, M. F., Welcher, A. A. and Horan, T.
Year: 1998
Title: Human leptin receptor. Determination of disulfide structure and N-glycosylation sites of the extracellular domain
Journal: J Biol Chem
Volume: 273
Issue: 44
Pages: 28691-9
Epub Date: 1998/10/24
Date: Oct 30
Short Title: Human leptin receptor. Determination of disulfide structure and N-glycosylation sites of the extracellular domain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.44.28691
Accession Number: 9786864
Keywords: Amino Acid Sequence
Animals
CHO Cells
Carrier Proteins/chemistry/isolation & purification/*metabolism
Cricetinae
Cross-Linking Reagents
Disulfides/*chemistry
Glycosylation
Humans
Ligands
Molecular Sequence Data
Pepsin A/metabolism
Peptide Mapping
*Receptors, Cell Surface
Receptors, Leptin
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: The leptin receptor (OB-R) is a member of the class I cytokine receptor family and mediates the weight regulatory effects of its ligand through interaction with cytoplasmic kinases. The extracellular domain of this receptor is comprised of two immunoglobulin-like and cytokine-receptor homology domains each and type III fibronectin domains. The extracellular domain of human leptin receptor was expressed in and purified from Chinese hamster ovary cells and was found to contain extensive N-glycosylation (approximately 36% of the total protein). The purified protein had a molecular weight of approximately 145,000 and exhibited ligand binding ability as evidenced by formation of ligand-receptor complex, followed by chemical cross-linking. The determined disulfide motif of the soluble leptin receptor contained several distinct cystine knots as well as 10 free cysteines. The N-glycosylation analysis revealed that Asn624 of the WSXWS motif (residues 622-626) within the C-terminal cytokine receptor homology domain was glycosylated, indicating that this region is solvent-exposed. On the other hand, the N-terminal WSXWS motif was not glycosylated.
Notes: Haniu, M
Arakawa, T
Bures, E J
Young, Y
Hui, J O
Rohde, M F
Welcher, A A
Horan, T
Journal Article
United States
J Biol Chem. 1998 Oct 30;273(44):28691-9. doi: 10.1074/jbc.273.44.28691.
Author Address: Departments of Protein Structure, Protein Chemistry, and Immunology, Amgen Inc., Thousand Oaks, California 91320-1789, USA. mhaniu@amgen.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1418
Author: Haniu, M., Horan, T., Spahr, C., Hui, J., Fan, W., Chen, C., Richards, W. G. and Lu, H. S.
Year: 2011
Title: Human Dickkopf-1 (huDKK1) protein: characterization of glycosylation and determination of disulfide linkages in the two cysteine-rich domains
Journal: Protein Sci
Volume: 20
Issue: 11
Pages: 1802-13
Epub Date: 2011/08/02
Date: Nov
Short Title: Human Dickkopf-1 (huDKK1) protein: characterization of glycosylation and determination of disulfide linkages in the two cysteine-rich domains
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1002/pro.705
PMCID: PMC3267945
Accession Number: 21805521
Keywords: Amino Acid Sequence
Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Cysteine/chemistry/metabolism
Disulfides
Electrophoresis, Polyacrylamide Gel
Glycosylation
Intercellular Signaling Peptides and Proteins/chemistry/*metabolism
Molecular Sequence Data
*Peptide Mapping
Polysaccharides/chemistry/metabolism
Protein Structure, Secondary
Protein Structure, Tertiary
Sequence Analysis, Protein
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Wnt Signaling Pathway
Abstract: Human Dickkopf-1 (huDKK1), an inhibitor of the canonical Wnt-signaling pathway that has been implicated in bone metabolism and other diseases, was expressed in engineered Chinese hamster ovary cells and purified. HuDKK1 is biologically active in a TCF/lef-luciferase reporter gene assay and is able to bind LRP6 coreceptor. In SDS-PAGE, huDKK1 exhibits molecular weights of 27-28 K and 30 K at ∼ 1:9 ratio. By MALDI-MS analysis, the observed molecular weights of 27.4K and 29.5K indicate that the low molecular weight form may contain O-linked glycans while the high molecular weight form contains both N- and O-linked glycans. LC-MS/MS peptide mapping indicates that ∼ 92% of huDKK1 is glycosylated at Asn²²⁵ with three N-linked glycans composed of two biantennary forms with 1 and 2 sialic acid (23% and 60%, respectively), and one triantennary structure with 2 sialic acids (9%). HuDKK1 contains two O-linked glycans, GalNAc (sialic acid)-Gal-sialic acid (65%) and GalNAc-Gal[sialic acid] (30%), attached at Ser³⁰ as confirmed by β-elimination and targeted LC-MS/MS. The 10 intramolecular disulfide bonds at the N- and C-terminal cysteine-rich domains were elucidated by analyses including multiple proteolytic digestions, isolation and characterization of disulfide-containing peptides, and secondary digestion and characterization of selected disulfide-containing peptides. The five disulfide bonds within the huDKK1 N-terminal domain are unique to the DKK family proteins; there are no exact matches in disulfide positioning when compared to other known disulfide clusters. The five disulfide bonds assigned in the C-terminal domain show the expected homology with those found in colipase and other reported disulfide clusters.
Notes: 1469-896x
Haniu, Mitsuru
Horan, Tom
Spahr, Chris
Hui, John
Fan, Wei
Chen, Ching
Richards, William G
Lu, Hsieng S
Journal Article
Protein Sci. 2011 Nov;20(11):1802-13. doi: 10.1002/pro.705. Epub 2011 Sep 27.
Author Address: Department of Protein Sciences, Amgen, Inc., One Amgen Center Dr., Thousand Oaks, California 91320, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 515
Author: Haniu, M., Talvenheimo, J., Le, J., Katta, V., Welcher, A. and Rohde, M. F.
Year: 1995
Title: Extracellular domain of neurotrophin receptor trkB: disulfide structure, N-glycosylation sites, and ligand binding
Journal: Arch Biochem Biophys
Volume: 322
Issue: 1
Pages: 256-64
Epub Date: 1995/09/10
Date: Sep 10
Short Title: Extracellular domain of neurotrophin receptor trkB: disulfide structure, N-glycosylation sites, and ligand binding
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1995.1460
Accession Number: 7574684
Keywords: Amino Acid Sequence
Animals
Binding Sites
CHO Cells
Cricetinae
Cross-Linking Reagents
Disulfides/chemistry
Glycosylation
Humans
Ligands
Molecular Sequence Data
Molecular Structure
Molecular Weight
Peptide Fragments/chemistry/genetics
Peptide Mapping
Receptor, Ciliary Neurotrophic Factor
Receptors, Nerve Growth Factor/*chemistry/genetics/metabolism
Solubility
Abstract: An extracellular domain of a human neurotrophin receptor trkB was expressed in Chinese hamster ovary cells and isolated as a glycoprotein possessing binding activity for brain-derived neurotrophic factor. The extracellular domain contains 398 amino acids and has a molecular weight of 60.6 kDa according to laser desorption mass spectrometry, indicating that the extracellular domain of trkB contains 33.3% carbohydrate moieties. Six disulfide linkages were determined to be Cys1-Cys7, Cys5-Cys14, Cys121-Cys145, Cys123-Cys163, Cys187-Cys235, and Cys271-Cys314, respectively. Cys300 was detected as a free sulfhydryl residue. Cysteine clusters 1 and 2 located in the N-terminal domain possess a similar type of disulfide structure and two other disulfide bonds in the C-terminal region are homologous to that of the Ig-like C2 domain. Among 12 potential N-linked glycosylation sites proposed in the soluble domain of trkB, 10 sites are actually glycosylated.
Notes: Haniu, M
Talvenheimo, J
Le, J
Katta, V
Welcher, A
Rohde, M F
Journal Article
United States
Arch Biochem Biophys. 1995 Sep 10;322(1):256-64. doi: 10.1006/abbi.1995.1460.
Author Address: Amgen, Inc., Thousand Oaks, California 91320, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 318
Author: Hannum, C. H., Wilcox, C. J., Arend, W. P., Joslin, F. G., Dripps, D. J., Heimdal, P. L., Armes, L. G., Sommer, A., Eisenberg, S. P. and Thompson, R. C.
Year: 1990
Title: Interleukin-1 receptor antagonist activity of a human interleukin-1 inhibitor
Journal: Nature
Volume: 343
Issue: 6256
Pages: 336-40
Epub Date: 1990/01/25
Date: Jan 25
Short Title: Interleukin-1 receptor antagonist activity of a human interleukin-1 inhibitor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/343336a0
Accession Number: 2137200
Keywords: Amidohydrolases
Amino Acid Sequence
Binding, Competitive
Cells, Cultured
Chromatography
Chromatography, High Pressure Liquid
Dinoprostone/biosynthesis
Electrophoresis, Polyacrylamide Gel
Fibroblasts/metabolism
Glycosylation
Humans
Interleukin 1 Receptor Antagonist Protein
Interleukin-1/*antagonists & inhibitors
Molecular Sequence Data
Monocytes/*metabolism
Peptide Fragments
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Proteins/*isolation & purification/metabolism/pharmacology
Receptors, Immunologic/*antagonists & inhibitors/metabolism
Receptors, Interleukin-1
Recombinant Proteins/metabolism/pharmacology
*Sialoglycoproteins
Abstract: Three interleukin-1 inhibitors have been purified to homogeneity from medium conditioned by human monocytes. Partial sequence analysis and digestion with N-glycanase indicate that these are glycosylation forms of a single protein. The protein binds to the interleukin-1 receptor but has no interleukin-1-like activity, even at very high concentrations, and is therefore a pure receptor antagonist.
Notes: Hannum, C H
Wilcox, C J
Arend, W P
Joslin, F G
Dripps, D J
Heimdal, P L
Armes, L G
Sommer, A
Eisenberg, S P
Thompson, R C
Comparative Study
Journal Article
England
Nature. 1990 Jan 25;343(6256):336-40. doi: 10.1038/343336a0.
Author Address: Synergen Inc., Boulder, Colorado 80301.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1184
Author: Hansen, G., Hercus, T. R., McClure, B. J., Stomski, F. C., Dottore, M., Powell, J., Ramshaw, H., Woodcock, J. M., Xu, Y., Guthridge, M., McKinstry, W. J., Lopez, A. F. and Parker, M. W.
Year: 2008
Title: The structure of the GM-CSF receptor complex reveals a distinct mode of cytokine receptor activation
Journal: Cell
Volume: 134
Issue: 3
Pages: 496-507
Epub Date: 2008/08/12
Date: Aug 8
Short Title: The structure of the GM-CSF receptor complex reveals a distinct mode of cytokine receptor activation
Alternate Journal: Cell
ISSN: 0092-8674
DOI: 10.1016/j.cell.2008.05.053
Accession Number: 18692472
Keywords: Amino Acid Sequence
Crystallography
Granulocyte-Macrophage Colony-Stimulating Factor/*chemistry/*metabolism
Humans
Models, Molecular
Molecular Sequence Data
Receptors, Granulocyte-Macrophage Colony-Stimulating Factor/*chemistry/*metabolism
Abstract: Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic cytokine that controls the production and function of blood cells, is deregulated in clinical conditions such as rheumatoid arthritis and leukemia, yet offers therapeutic value for other diseases. Its receptors are heterodimers consisting of a ligand-specific alpha subunit and a betac subunit that is shared with the interleukin (IL)-3 and IL-5 receptors. How signaling is initiated remains an enigma. We report here the crystal structure of the human GM-CSF/GM-CSF receptor ternary complex and its assembly into an unexpected dodecamer or higher-order complex. Importantly, mutagenesis of the GM-CSF receptor at the dodecamer interface and functional studies reveal that dodecamer formation is required for receptor activation and signaling. This unusual form of receptor assembly likely applies also to IL-3 and IL-5 receptors, providing a structural basis for understanding their mechanism of activation and for the development of therapeutics.
Notes: 1097-4172
Hansen, Guido
Hercus, Timothy R
McClure, Barbara J
Stomski, Frank C
Dottore, Mara
Powell, Jason
Ramshaw, Hayley
Woodcock, Joanna M
Xu, Yibin
Guthridge, Mark
McKinstry, William J
Lopez, Angel F
Parker, Michael W
R01-AI50744-02/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
Cell. 2008 Aug 8;134(3):496-507. doi: 10.1016/j.cell.2008.05.053.
Author Address: Biota Structural Biology Laboratory, St. Vincent's Institute of Medical Research, 41 Victoria Parade, Fitzroy, Victoria 3065, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1459
Author: Hanson, M. A., Roth, C. B., Jo, E., Griffith, M. T., Scott, F. L., Reinhart, G., Desale, H., Clemons, B., Cahalan, S. M., Schuerer, S. C., Sanna, M. G., Han, G. W., Kuhn, P., Rosen, H. and Stevens, R. C.
Year: 2012
Title: Crystal structure of a lipid G protein-coupled receptor
Journal: Science
Volume: 335
Issue: 6070
Pages: 851-5
Epub Date: 2012/02/22
Date: Feb 17
Short Title: Crystal structure of a lipid G protein-coupled receptor
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1215904
PMCID: PMC3338336
NIHMSID: NIHMS368267
Accession Number: 22344443
Keywords: Anilides/chemistry
Binding Sites
Crystallography, X-Ray
Models, Molecular
Muramidase/chemistry
Mutagenesis
Organophosphonates/chemistry
Protein Conformation
Receptors, Lysosphingolipid/agonists/antagonists & inhibitors/*chemistry/genetics
Recombinant Fusion Proteins/chemistry/genetics
Abstract: The lyso-phospholipid sphingosine 1-phosphate modulates lymphocyte trafficking, endothelial development and integrity, heart rate, and vascular tone and maturation by activating G protein-coupled sphingosine 1-phosphate receptors. Here, we present the crystal structure of the sphingosine 1-phosphate receptor 1 fused to T4-lysozyme (S1P(1)-T4L) in complex with an antagonist sphingolipid mimic. Extracellular access to the binding pocket is occluded by the amino terminus and extracellular loops of the receptor. Access is gained by ligands entering laterally between helices I and VII within the transmembrane region of the receptor. This structure, along with mutagenesis, agonist structure-activity relationship data, and modeling, provides a detailed view of the molecular recognition and requirement for hydrophobic volume that activates S1P(1), resulting in the modulation of immune and stromal cell responses.
Notes: 1095-9203
Hanson, Michael A
Roth, Christopher B
Jo, Euijung
Griffith, Mark T
Scott, Fiona L
Reinhart, Greg
Desale, Hans
Clemons, Bryan
Cahalan, Stuart M
Schuerer, Stephan C
Sanna, M Germana
Han, Gye Won
Kuhn, Peter
Rosen, Hugh
Stevens, Raymond C
Y1-GM-1104/GM/NIGMS NIH HHS/United States
U54 GM094618-02/GM/NIGMS NIH HHS/United States
U54 MH084512/MH/NIMH NIH HHS/United States
R01 AI055509-04/AI/NIAID NIH HHS/United States
P50 GM073197/GM/NIGMS NIH HHS/United States
U54 GM094618/GM/NIGMS NIH HHS/United States
U01 AI074564-04/AI/NIAID NIH HHS/United States
R01 AI055509/AI/NIAID NIH HHS/United States
U54 MH084512-04/MH/NIMH NIH HHS/United States
Y1-CO-1020/CO/NCI NIH HHS/United States
AI074564/AI/NIAID NIH HHS/United States
AI055509/AI/NIAID NIH HHS/United States
P50 GM073197-08/GM/NIGMS NIH HHS/United States
U01 AI074564/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Science. 2012 Feb 17;335(6070):851-5. doi: 10.1126/science.1215904.
Author Address: Receptos, 10835 Road to the Cure, San Diego, CA 92121, USA. mhanson@receptos.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 589
Author: Hara, H., Nakae, Y., Sogabe, T., Ihara, I., Ueno, S., Sakai, H., Inoue, H., Shimizu, S., Nakamura, T. and Shimizu, N.
Year: 1993
Title: Structural study of the N-linked oligosaccharides of hepatocyte growth factor by two-dimensional sugar mapping
Journal: J Biochem
Volume: 114
Issue: 1
Pages: 76-82
Epub Date: 1993/07/01
Date: Jul
Short Title: Structural study of the N-linked oligosaccharides of hepatocyte growth factor by two-dimensional sugar mapping
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a124143
Accession Number: 8407880
Keywords: Aminopyridines/chemistry
Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Cricetinae
Cricetulus
Hepatocyte Growth Factor/*chemistry
Liver/*chemistry
Molecular Sequence Data
Oligosaccharides/*chemistry
Rats
Recombinant Proteins/chemistry
Spectrometry, Fluorescence
Abstract: The structures of the N-linked oligosaccharides on recombinant human hepatocyte growth factor (rh-HGF) expressed by Chinese hamster ovary (CHO) cells were studied by two-dimensional sugar mapping. The oligosaccharides released from the glycopeptides by peptide: N-glycosidase F (PNGase F) treatment were tagged with 2-aminopyridine at the reducing ends. The alpha-chain was linked by biantennary, triantennary, and tetraantennary oligosaccharides, but the dominant oligosaccharides linking the beta-chain were biantennary (> 85%). There was no significant difference in oligosaccharide structures between the two glycosylation sites on each chain, that is, Asn263 and Asn371 on the alpha-chain, and Asn535 and Asn622 on the beta-chain. The linkage of sialic acid to the non-reducing terminal galactose was identified as NeuAc alpha(2-3) by 1H-NMR spectrometry. The structures of the N-linked oligosaccharides from rat HGF were also studied. Triantennary oligosaccharides were obtained from the alpha-chain and a biantennary oligosaccharide was obtained from the beta-chain. This result indicates that the alpha-chain is also linked by higher branched oligosaccharides than the beta-chain in rat HGF.
Notes: Hara, H
Nakae, Y
Sogabe, T
Ihara, I
Ueno, S
Sakai, H
Inoue, H
Shimizu, S
Nakamura, T
Shimizu, N
Journal Article
England
J Biochem. 1993 Jul;114(1):76-82. doi: 10.1093/oxfordjournals.jbchem.a124143.
Author Address: Pharmaceuticals Research Center of Toyobo Co., Ltd., Otsu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1434
Author: Hara, Y., Kanagawa, M., Kunz, S., Yoshida-Moriguchi, T., Satz, J. S., Kobayashi, Y. M., Zhu, Z., Burden, S. J., Oldstone, M. B. and Campbell, K. P.
Year: 2011
Title: Like-acetylglucosaminyltransferase (LARGE)-dependent modification of dystroglycan at Thr-317/319 is required for laminin binding and arenavirus infection
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 42
Pages: 17426-31
Epub Date: 2011/10/12
Date: Oct 18
Short Title: Like-acetylglucosaminyltransferase (LARGE)-dependent modification of dystroglycan at Thr-317/319 is required for laminin binding and arenavirus infection
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1114836108
PMCID: PMC3198351
Accession Number: 21987822
Keywords: Amino Acid Sequence
Animals
Arenaviridae Infections/*etiology/*metabolism
Binding Sites
Cell Line
Dystroglycans/chemistry/genetics/*metabolism
Glycosylation
HEK293 Cells
Humans
Laminin/*metabolism
*Lymphocytic choriomeningitis virus/pathogenicity
Mice
Models, Biological
Molecular Sequence Data
Mutagenesis
Myoblasts/metabolism
N-Acetylglucosaminyltransferases/*metabolism
Phosphorylation
Protein Binding
Protein Processing, Post-Translational
Recombinant Proteins/chemistry/genetics/metabolism
Threonine/chemistry
Virus Internalization
Abstract: α-dystroglycan is a highly O-glycosylated extracellular matrix receptor that is required for anchoring of the basement membrane to the cell surface and for the entry of Old World arenaviruses into cells. Like-acetylglucosaminyltransferase (LARGE) is a key molecule that binds to the N-terminal domain of α-dystroglycan and attaches ligand-binding moieties to phosphorylated O-mannose on α-dystroglycan. Here we show that the LARGE modification required for laminin- and virus-binding occurs on specific Thr residues located at the extreme N terminus of the mucin-like domain of α-dystroglycan. Deletion and mutation analyses demonstrate that the ligand-binding activity of α-dystroglycan is conferred primarily by LARGE modification at Thr-317 and -319, within the highly conserved first 18 amino acids of the mucin-like domain. The importance of these paired residues in laminin-binding and clustering activity on myoblasts and in arenavirus cell entry is confirmed by mutational analysis with full-length dystroglycan. We further demonstrate that a sequence of five amino acids, Thr(317)ProThr(319)ProVal, contains phosphorylated O-glycosylation and, when modified by LARGE is sufficient for laminin-binding. Because the N-terminal region adjacent to the paired Thr residues is removed during posttranslational maturation of dystroglycan, our results demonstrate that the ligand-binding activity resides at the extreme N terminus of mature α-dystroglycan and is crucial for α-dystroglycan to coordinate the assembly of extracellular matrix proteins and to bind arenaviruses on the cell surface.
Notes: 1091-6490
Hara, Yuji
Kanagawa, Motoi
Kunz, Stefan
Yoshida-Moriguchi, Takako
Satz, Jakob S
Kobayashi, Yvonne M
Zhu, Zihan
Burden, Steven J
Oldstone, Michael B A
Campbell, Kevin P
U54 NS053672/NS/NINDS NIH HHS/United States
3R01AR051199-05S1/AR/NIAMS NIH HHS/United States
AI09484/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 AI009484/AI/NIAID NIH HHS/United States
P30 DK054759/DK/NIDDK NIH HHS/United States
1U54NS053672/NS/NINDS NIH HHS/United States
P30 DK54759/DK/NIDDK NIH HHS/United States
R01 AR051199/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, American Recovery and Reinvestment Act
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Oct 18;108(42):17426-31. doi: 10.1073/pnas.1114836108. Epub 2011 Oct 10.
Author Address: The Howard Hughes Medical Institute, The Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, IA 52242, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1323
Author: Harada, H., Ohto, U. and Satow, Y.
Year: 2010
Title: Crystal structure of mouse MD-1 with endogenous phospholipid bound in its cavity
Journal: J Mol Biol
Volume: 400
Issue: 4
Pages: 838-46
Epub Date: 2010/07/03
Date: Jul 23
Short Title: Crystal structure of mouse MD-1 with endogenous phospholipid bound in its cavity
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2010.05.063
Accession Number: 20595044
Keywords: Animals
Antigens, Surface/*chemistry/*metabolism
Binding Sites
Crystallography, X-Ray
Dimerization
Membrane Glycoproteins/*chemistry/*metabolism
Mice
Phospholipids/*chemistry/*metabolism
Protein Binding
Protein Structure, Quaternary
Protein Structure, Secondary
Protein Structure, Tertiary
Abstract: MD-1 is a glycoprotein that associates with a B-cell-specific RP105 protein and has a low sequence identity of 16% to MD-2 that associates with Toll-like receptor 4 and recognizes endotoxic lipopolysaccharide. MD-1 and RP105 are supposed to mediate lipopolysaccharide recognition; however, little is known about their structures and functions. Here, the crystal structure of mouse MD-1 is determined at 1.65 A resolution. MD-1 has a hydrophobic cavity sandwiched by two beta-sheets as is MD-2. The cavity is 25 A long, 5 A wide, and 10 A deep: longer, narrower, and shallower than that of MD-2. No charged residues are located on the cavity entrance. MD-1 is primarily monomeric in solution but shows a dimeric assembly in the crystal lattices, with their cavity entrances facing each other. In the cavity, electron densities attributable to phosphatidylcholine are located. Together with the binding assay with tetra-acylated lipid IVa, MD-1 is shown to be a lipid-binding coreceptor.
Notes: 1089-8638
Harada, Hitomi
Ohto, Umeharu
Satow, Yoshinori
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2010 Jul 23;400(4):838-46. doi: 10.1016/j.jmb.2010.05.063. Epub 2010 Jun 1.
Author Address: Graduate School of Pharmaceutical Sciences, University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1017
Author: Harazono, A., Kawasaki, N., Itoh, S., Hashii, N., Ishii-Watabe, A., Kawanishi, T. and Hayakawa, T.
Year: 2006
Title: Site-specific N-glycosylation analysis of human plasma ceruloplasmin using liquid chromatography with electrospray ionization tandem mass spectrometry
Journal: Anal Biochem
Volume: 348
Issue: 2
Pages: 259-68
Epub Date: 2005/12/03
Date: Jan 15
Short Title: Site-specific N-glycosylation analysis of human plasma ceruloplasmin using liquid chromatography with electrospray ionization tandem mass spectrometry
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/j.ab.2005.10.036
Accession Number: 16321355
Keywords: Amino Acid Sequence
Asparagine/*analysis/metabolism
Ceruloplasmin/*analysis/chemistry/metabolism
Chromatography, High Pressure Liquid/*methods
Glycopeptides/*analysis
Glycosylation
Humans
Molecular Sequence Data
Oligosaccharides/analysis
Peptide Mapping
Spectrometry, Mass, Electrospray Ionization/*methods
Trypsin/chemistry
Abstract: Ceruloplasmin has ferroxidase activity and plays an essential role in iron metabolism. In this study, a site-specific glycosylation analysis of human ceruloplasmin (CP) was carried out using reversed-phase high-performance liquid chromatography with electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). A tryptic digest of carboxymethylated CP was subjected to LC-ESI-MS/MS. Product ion spectra acquired data-dependently were used for both distinction of the glycopeptides from the peptides using the carbohydrate B-ions, such as m/z 204 (HexNAc) and m/z 366 (HexHexNAc), and identification of the peptide moiety of the glycopeptide based on the presence of the b- and y-series ions derived from the peptide. Oligosaccharide composition was deduced from the molecular weight calculated from the observed mass of the glycopeptide and theoretical mass of the peptide. Of the seven potential N-glycosylation sites, four (Asn119, Asn339, Asn378, and Asn743) were occupied by a sialylated biantennary or triantennary oligosaccharide with fucose residues (0, 1, or 2). A small amount of sialylated tetraantennary oligosaccharide was detected. Exoglycosidase digestion suggested that fucose residues were linked to reducing end GlcNAc in biantennary oligosaccharides and to reducing end and/or alpha1-3 to outer arms GlcNAc in triantennary oligosaccharides and that roughly one of the antennas in triantennary oligosaccharides was alpha2-3 sialylated and occasionally alpha1-3 fucosylated at GlcNAc.
Notes: Harazono, Akira
Kawasaki, Nana
Itoh, Satsuki
Hashii, Noritaka
Ishii-Watabe, Akiko
Kawanishi, Toru
Hayakawa, Takao
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Biochem. 2006 Jan 15;348(2):259-68. doi: 10.1016/j.ab.2005.10.036. Epub 2005 Nov 10.
Author Address: National Institute of Health Sciences, Division of Biological Chemistry and Biologicals, 1-18-1 Kami-yoga, Tokyo 158-8501, Japan. harazono@nihs.go.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1171
Author: Harazono, A., Kawasaki, N., Itoh, S., Hashii, N., Matsuishi-Nakajima, Y., Kawanishi, T. and Yamaguchi, T.
Year: 2008
Title: Simultaneous glycosylation analysis of human serum glycoproteins by high-performance liquid chromatography/tandem mass spectrometry
Journal: J Chromatogr B Analyt Technol Biomed Life Sci
Volume: 869
Issue: 1-2
Pages: 20-30
Epub Date: 2008/06/03
Date: Jun 15
Short Title: Simultaneous glycosylation analysis of human serum glycoproteins by high-performance liquid chromatography/tandem mass spectrometry
Alternate Journal: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences
ISSN: 1570-0232 (Print)
1570-0232
DOI: 10.1016/j.jchromb.2008.05.006
Accession Number: 18514042
Keywords: Chromatography, High Pressure Liquid/*methods
Databases, Protein
Glycopeptides/analysis/isolation & purification
Glycoproteins/*blood/*chemistry/metabolism
Glycosylation
Humans
Protein Processing, Post-Translational
Tandem Mass Spectrometry/*methods
Abstract: Changes in the glycosylation of some serum proteins are associated with certain diseases. In this study, we performed simultaneous site-specific glycosylation analysis of abundant serum glycoproteins by LC/Qq-TOF MS of human serum tryptic digest, the albumin of which was depleted. The glycopeptide peaks on the chromatogram were basically assigned by database searching with modified peak-list text files of MS/MS spectra and then based on mass differences of glycan units from characterized glycopeptides. Glycopeptide of IgG, haptoglobin and ceruloplasmin were confirmed by means of a comparison of their retention times and m/z values with those obtained by LC/MS of commercially available glycoproteins. Mass spectrometric carbohydrate heterogeneity in the assigned glycopeptides was analyzed by an additional LC/MS. We successfully demonstrated site-specific glycosylation of 23 sites in abundant serum glycoproteins.
Notes: Harazono, Akira
Kawasaki, Nana
Itoh, Satsuki
Hashii, Noritaka
Matsuishi-Nakajima, Yukari
Kawanishi, Toru
Yamaguchi, Teruhide
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Chromatogr B Analyt Technol Biomed Life Sci. 2008 Jun 15;869(1-2):20-30. doi: 10.1016/j.jchromb.2008.05.006. Epub 2008 May 10.
Author Address: Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, Tokyo, Japan. harazono@nihs.go.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 232
Author: Hård, K., Damm, J. B., Spruijt, M. P., Bergwerff, A. A., Kamerling, J. P., Van Dedem, G. W. and Vliegenthart, J. F.
Year: 1992
Title: The carbohydrate chains of the beta subunit of human chorionic gonadotropin produced by the choriocarcinoma cell line BeWo. Novel O-linked and novel bisecting-GlcNAc-containing N-linked carbohydrates
Journal: Eur J Biochem
Volume: 205
Issue: 2
Pages: 785-98
Epub Date: 1992/04/15
Date: Apr 15
Short Title: The carbohydrate chains of the beta subunit of human chorionic gonadotropin produced by the choriocarcinoma cell line BeWo. Novel O-linked and novel bisecting-GlcNAc-containing N-linked carbohydrates
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1992.tb16843.x
Accession Number: 1374031
Keywords: Acetylglucosamine/*analysis
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Choriocarcinoma
Chorionic Gonadotropin/biosynthesis/*chemistry/urine
Chorionic Gonadotropin, beta Subunit, Human
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Electrophoresis, Polyacrylamide Gel
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/biosynthesis/*chemistry/urine
Abstract: The N-linked carbohydrate chains of the beta subunit of human chorionic gonadotropin (hCG-beta) isolated from the culture fluid of the choriocarcinoma cell line BeWo were released enzymatically by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. Subsequently, the O-linked oligosaccharides were split off from the N-deglycosylated protein by mild alkaline borohydride treatment. The carbohydrate chains were purified in their intact sialylated forms by FPLC anion-exchange chromatography on Mono Q, HPLC on Lichrosorb-NH2, and high-pH anion-exchange chromatography on CarboPac PA1. 1H-NMR spectroscopic analysis of the major fractions demonstrates the occurrence of the following sialylated diantennary and triantennary N-linked oligosaccharides. Residues not written in bold letters are variably present. [formula: see text] The incidence of triantennary carbohydrate chains is much higher than in normal urinary hCG-beta (26% vs 2%). The same holds for the alpha 1-6-fucosylation of the asparagine-bound GlcNAc (95% vs 42%). The presence of a bisecting GlcNAc and the occurrence of alpha 2-6-linked Neu5Ac in the most abundant N-glycans, are new features for hCG-beta. The major O-linked carbohydrate chains identified are the tetrasaccharide Neu5Ac alpha 2-3Gal beta 1-3(Neu5Ac alpha 2-6)GalNAc-ol and the hexasaccharide Neu5Ac alpha 2-3Gal beta 1-4GlcNAc beta 1-6(Neu5Ac alpha 2-3Gal beta 1-3)GalNAc-ol, both also found in normal urinary hCG. In addition, two novel O-glycans were characterized: [formula: see text]
Notes: Hård, K
Damm, J B
Spruijt, M P
Bergwerff, A A
Kamerling, J P
Van Dedem, G W
Vliegenthart, J F
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1992 Apr 15;205(2):785-98. doi: 10.1111/j.1432-1033.1992.tb16843.x.
Author Address: Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 314
Author: Hård, K., Mekking, A., Damm, J. B., Kamerling, J. P., de Boer, W., Wijnands, R. A. and Vliegenthart, J. F.
Year: 1990
Title: Isolation and structure determination of the intact sialylated N-linked carbohydrate chains of recombinant human follitropin expressed in Chinese hamster ovary cells
Journal: Eur J Biochem
Volume: 193
Issue: 1
Pages: 263-71
Epub Date: 1990/10/05
Date: Oct 5
Short Title: Isolation and structure determination of the intact sialylated N-linked carbohydrate chains of recombinant human follitropin expressed in Chinese hamster ovary cells
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1990.tb19332.x
Accession Number: 2121479
Keywords: Animals
Carbohydrate Sequence
Cell Line
Cricetinae
Cricetulus
Follicle Stimulating Hormone/*chemistry/genetics/metabolism
Glycosylation
Molecular Sequence Data
Monosaccharides/analysis
Protein Processing, Post-Translational
Recombinant Proteins/chemistry/metabolism
Sialic Acids/analysis
Abstract: Biologically active recombinant human follitropin has been expressed in Chinese hamster ovary cells. The carbohydrate chains of the recombinant glycoprotein hormone were enzymatically released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. The oligosaccharides were separated from the N-deglycosylated protein by gel-permeation chromatography on Bio-Gel P-100, and fractionated by a combination of FPLC on Mono Q and HPLC on Lichrosorb-NH2. The structures of the carbohydrate chains were determined by 500- or 600-MHz 1H-NMR spectroscopy. The following types of carbohydrates occur: monosialylated diantennary (10%), disialylated diantennary (43%), disialylated tri-antennary (5%), trisialylated tri-antennary (13%), trisialylated tri'-antennary (8%), and tetrasialylated tetraantennary (12%) N-acetyllactosamine type of carbohydrate chains, all bearing exclusively alpha 2-3-linked N-acetylneuraminic acid (Neu5Ac). Previously, for pituitary follitropin mono-, di-, tri-, tri'-, and tetra-antennary oligosaccharides containing alpha 2-3- as well as alpha 2-6-linked Neu5Ac residues were reported. The bisecting GlcNAc residues present in native follitropin were not detected in the recombinant glycoprotein. Of the oligosaccharides 29% have an alpha 1-6-linked Fuc residue at the asparagine-bound GlcNAc, whereas this amount is about 50% in pituitary follitropin. In some of the tri-, tri'- and tetra-antennary oligosaccharide fractions small amounts (less than 5%) of compounds were detected having one or more additional N-acetyllactosamine units.
Notes: Hård, K
Mekking, A
Damm, J B
Kamerling, J P
de Boer, W
Wijnands, R A
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1990 Oct 5;193(1):263-71. doi: 10.1111/j.1432-1033.1990.tb19332.x.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 240
Author: Hård, K., Van Zadelhoff, G., Moonen, P., Kamerling, J. P. and Vliegenthart, F. G.
Year: 1992
Title: The Asn-linked carbohydrate chains of human Tamm-Horsfall glycoprotein of one male. Novel sulfated and novel N-acetylgalactosamine-containing N-linked carbohydrate chains
Journal: Eur J Biochem
Volume: 209
Issue: 3
Pages: 895-915
Epub Date: 1992/11/01
Date: Nov 1
Short Title: The Asn-linked carbohydrate chains of human Tamm-Horsfall glycoprotein of one male. Novel sulfated and novel N-acetylgalactosamine-containing N-linked carbohydrate chains
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1992.tb17362.x
Accession Number: 1425697
Keywords: Acetylgalactosamine/*chemistry
Asparagine/*chemistry
Carbohydrate Sequence
Carbohydrates/*chemistry
Chromatography, Ion Exchange
Chromatography, Liquid
Humans
Magnetic Resonance Spectroscopy
Male
Molecular Sequence Data
Mucoproteins/*chemistry
Sulfates/*chemistry
Uromodulin
Abstract: Human Tamm-Horsfall glycoprotein has been purified from the urine of one male. The Asn-linked carbohydrate chains were enzymically released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F, and separated from the remaining protein by gel-permeation chromatography on Bio-Gel P-100. Fractionation of the intact (sulfated) sialylated carbohydrate chains was achieved by a combination of three liquid-chromatographic techniques, namely, anion-exchange FPLC on Q-Sepharose, amine-adsorption HPLC on Lichrospher-NH2, and high-pH anion-exchange chromatography on CarboPac PA1. In total, more than 150 carbohydrate-containing fractions were obtained, some of which still contained mixtures of oligosaccharides. The primary structure of 30 N-glycans, including 10 novel oligosaccharides, were determined by one- and two-dimensional 1H-NMR spectroscopy at 500 MHz or 600 MHz. The types of compounds identified range from non-fucosylated, monosialylated, diantennary to fucosylated, tetrasialylated, tetraantennary carbohydrate chains, possessing the following terminal structural elements: [formula: see text]
Notes: Hård, K
Van Zadelhoff, G
Moonen, P
Kamerling, J P
Vliegenthart, F G
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1992 Nov 1;209(3):895-915. doi: 10.1111/j.1432-1033.1992.tb17362.x.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2050
Author: Hardivillé, S., Banerjee, P. S., Selen Alpergin, E. S., Smith, D. M., Han, G., Ma, J., Talbot, C. C., Jr., Hu, P., Wolfgang, M. J. and Hart, G. W.
Year: 2020
Title: TATA-Box Binding Protein O-GlcNAcylation at T114 Regulates Formation of the B-TFIID Complex and Is Critical for Metabolic Gene Regulation
Journal: Mol Cell
Volume: 77
Issue: 5
Pages: 1143-1152.e7
Epub Date: 2019/12/24
Date: Mar 5
Short Title: TATA-Box Binding Protein O-GlcNAcylation at T114 Regulates Formation of the B-TFIID Complex and Is Critical for Metabolic Gene Regulation
Alternate Journal: Molecular cell
ISSN: 1097-2765 (Print)
1097-2765
DOI: 10.1016/j.molcel.2019.11.022
PMCID: PMC7061307
NIHMSID: NIHMS1545887
Accession Number: 31866147
Keywords: Animals
Chromatin/genetics/metabolism
Diabetes Mellitus, Experimental/genetics/metabolism
Gene Expression Regulation
Glucose/*metabolism
Glycosylation
HEK293 Cells
HeLa Cells
Humans
Lipid Droplets/*metabolism
*Lipid Metabolism/genetics
Male
Multiprotein Complexes
Rats, Sprague-Dawley
Signal Transduction
TATA-Binding Protein Associated Factors/genetics/*metabolism
TATA-Box Binding Protein/genetics/*metabolism
Time Factors
Transcription Factor TFIID/genetics/*metabolism
Transcription, Genetic
Transcriptome
*B-tfiid
*Btaf1
*O-GlcNAc
*O-GlcNAcylation
*TATA-box binding protein
*Tbp
*lipid droplets
*metabolism
*nutrient sensing
*transcription
Hopkins University (JHU) on sales of the CTD110.6 antibody, which is managed by JHU.
Abstract: In eukaryotes, gene expression is performed by three RNA polymerases that are targeted to promoters by molecular complexes. A unique common factor, the TATA-box binding protein (TBP), is thought to serve as a platform to assemble pre-initiation complexes competent for transcription. Here, we describe a novel molecular mechanism of nutrient regulation of gene transcription by dynamic O-GlcNAcylation of TBP. We show that O-GlcNAcylation at T114 of TBP blocks its interaction with BTAF1, hence the formation of the B-TFIID complex, and its dynamic cycling on and off of DNA. Transcriptomic and metabolomic analyses of TBP(T114A) CRISPR/Cas9-edited cells showed that loss of O-GlcNAcylation at T114 increases TBP binding to BTAF1 and directly impacts expression of 408 genes. Lack of O-GlcNAcylation at T114 is associated with a striking reprogramming of cellular metabolism induced by a profound modification of the transcriptome, leading to gross alterations in lipid storage.
Notes: 1097-4164
Hardivillé, Stéphan
Banerjee, Partha S
Selen Alpergin, Ebru S
Smith, Danielle M
Han, Guanghui
Ma, Junfeng
Talbot, C Conover Jr
Hu, Ping
Wolfgang, Michael J
Hart, Gerald W
R01 DK116746/DK/NIDDK NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
R01 GM116891/GM/NIGMS NIH HHS/United States
R01 NS072241/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cell. 2020 Mar 5;77(5):1143-1152.e7. doi: 10.1016/j.molcel.2019.11.022. Epub 2019 Dec 19.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. Electronic address: stephan.hardiville@univ-lille.fr.
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Institute for Basic Biomedical Sciences, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. Electronic address: gerald.hart@uga.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1304
Author: Hardivillé, S., Hoedt, E., Mariller, C., Benaïssa, M. and Pierce, A.
Year: 2010
Title: O-GlcNAcylation/phosphorylation cycling at Ser10 controls both transcriptional activity and stability of delta-lactoferrin
Journal: J Biol Chem
Volume: 285
Issue: 25
Pages: 19205-18
Epub Date: 2010/04/21
Date: Jun 18
Short Title: O-GlcNAcylation/phosphorylation cycling at Ser10 controls both transcriptional activity and stability of delta-lactoferrin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.080572
PMCID: PMC2885199
Accession Number: 20404350
Keywords: Acetylglucosamine/*chemistry
Aspartic Acid/chemistry
Carbohydrates/chemistry
Cell Line
DNA/chemistry
Genes, Reporter
Humans
Lactoferrin/chemistry/*physiology
Microscopy, Fluorescence/methods
Mutagenesis, Site-Directed
Phosphorylation
Promoter Regions, Genetic
Serine/*chemistry
*Transcription, Genetic
Transcriptional Activation
Abstract: Delta-lactoferrin (DeltaLf) is a transcription factor that up-regulates DcpS, Skp1, and Bax genes, provoking cell cycle arrest and apoptosis. It is post-translationally modified either by O-GlcNAc or phosphate, but the effects of the O-GlcNAc/phosphorylation interplay on DeltaLf function are not yet understood. Here, using a series of glycosylation mutants, we showed that Ser(10) is O-GlcNAcylated and that this modification is associated with increased DeltaLf stability, achieved by blocking ubiquitin-dependent proteolysis, demonstrating that O-GlcNAcylation protects against polyubiquitination. We highlighted the (391)KSQQSSDPDPNCVD(404) sequence as a functional PEST motif responsible for DeltaLf degradation and defined Lys(379) as the main polyubiquitin acceptor site. We next investigated the control of DeltaLf transcriptional activity by the O-GlcNAc/phosphorylation interplay. Reporter gene analyses using the Skp1 promoter fragment containing a DeltaLf response element showed that O-GlcNAcylation at Ser(10) negatively regulates DeltaLf transcriptional activity, whereas phosphorylation activates it. Using a chromatin immunoprecipitation assay, we showed that O-GlcNAcylation inhibits DNA binding. Deglycosylation leads to DNA binding and transactivation of the Skp1 promoter at a basal level. Basal transactivation was markedly enhanced by 2-3-fold when phosphorylation was mimicked at Ser(10) by aspartate. Moreover, using double chromatin immunoprecipitation assays, we showed that the DeltaLf transcriptional complex binds to the DeltaLf response element and is phosphorylated and/or ubiquitinated, suggesting that DeltaLf transcriptional activity and degradation are concomitant events. Collectively, our results indicate that reciprocal occupancy of Ser(10) by either O-phosphate or O-GlcNAc coordinately regulates DeltaLf stability and transcriptional activity.
Notes: 1083-351x
Hardivillé, Stéphan
Hoedt, Esthelle
Mariller, Christophe
Benaïssa, Monique
Pierce, Annick
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Jun 18;285(25):19205-18. doi: 10.1074/jbc.M109.080572. Epub 2010 Apr 19.
Author Address: Unité de Glycobiologie Structurale et Fonctionnelle, Unité Mixte de Recherche 8576 CNRS, Université des Sciences et Technologies de Lille, IFR 147, 59655 Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 946
Author: Haridas, M., Anderson, B. F. and Baker, E. N.
Year: 1995
Title: Structure of human diferric lactoferrin refined at 2.2 A resolution
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 51
Issue: Pt 5
Pages: 629-46
Epub Date: 1995/09/01
Date: Sep 1
Short Title: Structure of human diferric lactoferrin refined at 2.2 A resolution
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s0907444994013521
Accession Number: 15299793
Abstract: The three-dimensional structure of the diferric form of human lactoferrin has been refined at 2.2 A resolution, using synchrotron data combined with a lower resolution (3.2 A) diffractometer data set. Following restrained least-squares refinement and model rebuilding the final model comprises 5330 protein atoms (691 residues), 2Fe(3+) and 2CO(3)(2-) ions, 469 solvent molecules and 98 carbohydrate atoms (eight sugar residues). Root-mean-square deviations from standard geometry are 0.015 A for bond lengths and 0.038 A for angle (1-3) distances, and the final crystallographic R-factor is 0.179 for all 39 113 reflections in the resolution range 8.0-2.2 A. A close structural similarity is seen between the two lobes of the molecule, with differences mainly in loops and turns. The two binding sites are extremely similar, the only apparent differences being a slightly more asymmetric bidentate binding of the carbonate ion to the metal, and a slightly longer Fe-O bond to one of the Tyr ligands, in the N-lobe site relative to the C-lobe site. Distinct differences are seen in the interactions made by two cationic groups, Arg210 and Lys546, behind the iron site, and these may influence the stability of the two metal sites. Analysis of interdomain and interlobe interactions shows that these are few in number which is consistent with the known flexibility of the molecule with respect to domain and lobe movements. Internal water molecules are found in discrete sites and in two large clusters (in the two interdomain clefts) and one tightly bound water molecule is present 3.8 A from the Fe atom in each lobe. The carbohydrate is weakly defined and has been modelled to a limited extent; two sugar residues of the N-lobe glycan and six of the C-lobe glycan. Only one direct protein-carbohydrate contact can be found.
Notes: Haridas, M
Anderson, B F
Baker, E N
Journal Article
United States
Acta Crystallogr D Biol Crystallogr. 1995 Sep 1;51(Pt 5):629-46. doi: 10.1107/S0907444994013521.
Author Address: Department of Chemistry and Biochemistry, Massey University, Palmerston North, New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1234
Author: Harkiolaki, M., Holmes, S. L., Svendsen, P., Gregersen, J. W., Jensen, L. T., McMahon, R., Friese, M. A., van Boxel, G., Etzensperger, R., Tzartos, J. S., Kranc, K., Sainsbury, S., Harlos, K., Mellins, E. D., Palace, J., Esiri, M. M., van der Merwe, P. A., Jones, E. Y. and Fugger, L.
Year: 2009
Title: T cell-mediated autoimmune disease due to low-affinity crossreactivity to common microbial peptides
Journal: Immunity
Volume: 30
Issue: 3
Pages: 348-57
Epub Date: 2009/03/24
Date: Mar 20
Short Title: T cell-mediated autoimmune disease due to low-affinity crossreactivity to common microbial peptides
Alternate Journal: Immunity
ISSN: 1074-7613
DOI: 10.1016/j.immuni.2009.01.009
Accession Number: 19303388
Keywords: Animals
Autoimmune Diseases/*immunology
Bacterial Proteins/*immunology
Cells, Cultured
Cerebellum/pathology
Cross Reactions/immunology
Drosophila
Escherichia coli/immunology
HLA-D Antigens/metabolism
HLA-DR2 Antigen/metabolism
Humans
Immunohistochemistry
Mice
Mice, Transgenic
Models, Molecular
Molecular Mimicry/*immunology
Multiple Sclerosis/immunology
Peptides/*immunology/metabolism
Receptors, Antigen, T-Cell/chemistry/metabolism
Spinal Cord/pathology
T-Lymphocytes/*immunology/physiology
Abstract: Environmental factors account for 75% of the risk of developing multiple sclerosis (MS). Numerous infections have been suspected as environmental disease triggers, but none of them has consistently been incriminated, and it is unclear how so many different infections may play a role. We show that a microbial peptide, common to several major classes of bacteria, can induce MS-like disease in humanized mice by crossreacting with a T cell receptor (TCR) that also recognizes a peptide from myelin basic protein, a candidate MS autoantigen. Structural analysis demonstrates this crossreactivity is due to structural mimicry of a binding hotspot shared by self and microbial antigens, rather than to degenerate TCR recognition. Biophysical studies reveal that the autoreactive TCR binding affinity is markedly lower for the microbial (mimicry) peptide than for the autoantigenic peptide. Thus, these data suggest a possible explanation for the difficulty in incriminating individual infections in the development of MS.
Notes: 1097-4180
Harkiolaki, Maria
Holmes, Samantha L
Svendsen, Pia
Gregersen, Jon W
Jensen, Lise T
McMahon, Roisin
Friese, Manuel A
van Boxel, Gijs
Etzensperger, Ruth
Tzartos, John S
Kranc, Kamil
Sainsbury, Sarah
Harlos, Karl
Mellins, Elizabeth D
Palace, Jackie
Esiri, Margaret M
van der Merwe, P Anton
Jones, E Yvonne
Fugger, Lars
G0500365/Medical Research Council/United Kingdom
G9722488/Medical Research Council/United Kingdom
G9900061/Medical Research Council/United Kingdom
MC_U137881016/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Immunity. 2009 Mar 20;30(3):348-57. doi: 10.1016/j.immuni.2009.01.009.
Author Address: Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Roosevelt Drive, Oxford OX37BN, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 291
Author: Harris, R. J., Leonard, C. K., Guzzetta, A. W. and Spellman, M. W.
Year: 1991
Title: Tissue plasminogen activator has an O-linked fucose attached to threonine-61 in the epidermal growth factor domain
Journal: Biochemistry
Volume: 30
Issue: 9
Pages: 2311-4
Epub Date: 1991/03/05
Date: Mar 5
Short Title: Tissue plasminogen activator has an O-linked fucose attached to threonine-61 in the epidermal growth factor domain
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00223a004
Accession Number: 1900431
Keywords: Amino Acid Sequence
Animals
Cell Line
Chromatography, High Pressure Liquid
Chymotrypsin
Epidermal Growth Factor/genetics
Fucose/*analysis
Glycosylation
Humans
Melanoma/enzymology
Molecular Sequence Data
Peptide Fragments/isolation & purification
Peptide Mapping
Recombinant Proteins/chemistry/isolation & purification
*Threonine
Tissue Plasminogen Activator/*chemistry/genetics/isolation & purification
Trypsin
Abstract: An unusual type of glycosylation has been observed for tissue plasminogen activator (t-PA). The monosaccharide fucose is glycosidically linked to threonine-61 in the epidermal growth factor region of t-PA. The presence of O-linked fucose was demonstrated by carbohydrate analysis and mass spectrometry of tryptic and chymotryptic peptides that contain this site. The susceptibility of the fucose residue to alpha-fucosidase indicated that it was in the alpha-anomeric configuration. Fucosylation of threonine-61 was observed in t-PA isolated from the Bowes melanoma cell line and from recombinant expression systems using Chinese hamster ovary or human embryonic kidney cells. Fucosylation of the homologous residue in prourokinase has also been reported recently. Our results indicate that this novel type of glycosylation may be common to the epidermal growth factor domains found in coagulation and fibrinolytic proteins and, therefore, suggest that the modification may have functional significance.
Notes: Harris, R J
Leonard, C K
Guzzetta, A W
Spellman, M W
Journal Article
United States
Biochemistry. 1991 Mar 5;30(9):2311-4. doi: 10.1021/bi00223a004.
Author Address: Department of Medicinal and Analytical Chemistry, Genentech, Inc., South San Francisco, California 94080.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 256
Author: Harris, R. J., Ling, V. T. and Spellman, M. W.
Year: 1992
Title: O-linked fucose is present in the first epidermal growth factor domain of factor XII but not protein C
Journal: J Biol Chem
Volume: 267
Issue: 8
Pages: 5102-7
Epub Date: 1992/03/15
Date: Mar 15
Short Title: O-linked fucose is present in the first epidermal growth factor domain of factor XII but not protein C
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1544894
Keywords: Amino Acid Sequence
Chromatography, High Pressure Liquid
Cyanogen Bromide
Epidermal Growth Factor/*chemistry
Factor XII/*chemistry
Fucose/*analysis
Humans
Mass Spectrometry
Molecular Sequence Data
Peptide Fragments/isolation & purification
Peptide Mapping
Protein C/*chemistry
Sequence Homology, Nucleic Acid
Abstract: Epidermal growth factor (EGF) domains are found in many proteins, particularly those of the coagulation/fibrinolytic system. We and others have demonstrated that tissue plasminogen activator (t-PA) and prourokinase are modified by the attachment of fucose to equivalent threonine residues within their EGF domains. Factor XII and protein C each contain two EGF domains; in both proteins, the EGF domain nearest the N terminus has a threonine residue in a position homologous to that which is fucosylated in t-PA. In protein C, this site is 3 residues from the position of another post-translational modification, beta-hydroxylation of Asp-71. We isolated peptides containing these sites to determine, primarily by mass spectrometric analysis, the presence of O-linked fucose and/or beta-hydroxyaspartate. We found that factor XII is fully fucosylated at Thr-90. Protein C is unmodified at the equivalent site (Thr-68) and is completely beta-hydroxylated at Asp-71. It has been recently reported that the first EGF domain of human factor VII has O-linked fucose at the equivalent position (Ser-60) (Bjoern, S., Foster, D. C., Thim, L., Wiberg, F. C., Christensen, M., Komiyama, Y., Pedersen, A. H., and Kisiel, W. (1991) J. Biol. Chem. 266, 11051-11057), while it is unmodified at Asp-63 despite having the consensus sequence for beta-hydroxylation at the latter site. These observations raise the possibility that O-linked fucosylation and beta-hydroxylation of EGF domains are mutually exclusive post-translational modifications.
Notes: Harris, R J
Ling, V T
Spellman, M W
Comparative Study
Journal Article
United States
J Biol Chem. 1992 Mar 15;267(8):5102-7.
Author Address: Department of Medicinal and Analytical Chemistry, Genentech Inc., South San Francisco, California 94080.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 575
Author: Harris, R. J., van Halbeek, H., Glushka, J., Basa, L. J., Ling, V. T., Smith, K. J. and Spellman, M. W.
Year: 1993
Title: Identification and structural analysis of the tetrasaccharide NeuAc alpha(2-->6)Gal beta(1-->4)GlcNAc beta(1-->3)Fuc alpha 1-->O-linked to serine 61 of human factor IX
Journal: Biochemistry
Volume: 32
Issue: 26
Pages: 6539-47
Epub Date: 1993/07/06
Date: Jul 6
Short Title: Identification and structural analysis of the tetrasaccharide NeuAc alpha(2-->6)Gal beta(1-->4)GlcNAc beta(1-->3)Fuc alpha 1-->O-linked to serine 61 of human factor IX
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00077a007
Accession Number: 8329384
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Factor IX/*chemistry
Fucose/analysis
Glycopeptides/isolation & purification
Glycoside Hydrolases
Humans
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/isolation & purification
*Serine
Threonine
Abstract: O-Linked fucose has been found attached to Thr/Ser residues within the sequence Cys-X-X-Gly-Gly-Thr/Ser-Cys in the N-terminal EGF domains of several coagulation/fibrinolytic proteins. Carbohydrate composition and mass spectrometric analyses of tryptic and thermolytic peptides containing the corresponding site (Ser-61) in the first EGF domain of human factor IX indicated the presence of a tetrasaccharide containing one residue each of sialic acid, galactose, N-acetylglucosamine, and fucose. The Ser-61 tetrasaccharide was not susceptible to alpha-fucosidase digestion. Fragments generated during mass spectrometric analysis indicated that fucose was the attachment sugar residue. The involvement of fucose in the carbohydrate-peptide linkage was confirmed by two-dimensional 1H NMR spectroscopic analysis of the glycopeptide containing factor IX residues 57-65. The complete structure of the tetrasaccharide was obtained by methylation analysis and two-dimensional 1H TOCSY and ROESY experiments as NeuAc alpha(2-->6)Gal beta(1-->4)GlcNAc beta(1-->3)Fuc alpha 1-->O-Ser61.
Notes: Harris, R J
van Halbeek, H
Glushka, J
Basa, L J
Ling, V T
Smith, K J
Spellman, M W
P41-RR-05351/RR/NCRR NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1993 Jul 6;32(26):6539-47. doi: 10.1021/bi00077a007.
Author Address: Department of Medicinal and Analytical Chemistry, Genentech, Inc., South San Francisco, California 94080.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1396
Author: Harrison, K. D., Park, E. J., Gao, N., Kuo, A., Rush, J. S., Waechter, C. J., Lehrman, M. A. and Sessa, W. C.
Year: 2011
Title: Nogo-B receptor is necessary for cellular dolichol biosynthesis and protein N-glycosylation
Journal: Embo j
Volume: 30
Issue: 12
Pages: 2490-500
Epub Date: 2011/05/17
Date: May 13
Short Title: Nogo-B receptor is necessary for cellular dolichol biosynthesis and protein N-glycosylation
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2011.147
PMCID: PMC3116281
Accession Number: 21572394
Keywords: Alkyl and Aryl Transferases/antagonists & inhibitors/deficiency/metabolism
Animals
COS Cells
Carrier Proteins/metabolism
Chlorocebus aethiops
Dolichol Phosphates/biosynthesis/deficiency
Dolichols/*biosynthesis/deficiency
Enzyme Activation/genetics
Glycoproteins/metabolism
Humans
Protein Conformation
Receptors, Cell Surface/chemistry/deficiency/*physiology
Vesicular Transport Proteins
Abstract: Dolichol monophosphate (Dol-P) functions as an obligate glycosyl carrier lipid in protein glycosylation reactions. Dol-P is synthesized by the successive condensation of isopentenyl diphosphate (IPP), with farnesyl diphosphate catalysed by a cis-isoprenyltransferase (cis-IPTase) activity. Despite the recognition of cis-IPTase activity 40 years ago and the molecular cloning of the human cDNA encoding the mammalian enzyme, the molecular machinery responsible for regulating this activity remains incompletely understood. Here, we identify Nogo-B receptor (NgBR) as an essential component of the Dol-P biosynthetic machinery. Loss of NgBR results in a robust deficit in cis-IPTase activity and Dol-P production, leading to diminished levels of dolichol-linked oligosaccharides and a broad reduction in protein N-glycosylation. NgBR interacts with the previously identified cis-IPTase hCIT, enhances hCIT protein stability, and promotes Dol-P production. Identification of NgBR as a component of the cis-IPTase machinery yields insights into the regulation of dolichol biosynthesis.
Notes: 1460-2075
Harrison, Kenneth D
Park, Eon Joo
Gao, Ningguo
Kuo, Andrew
Rush, Jeffrey S
Waechter, Charles J
Lehrman, Mark A
Sessa, William C
P01 HL070295/HL/NHLBI NIH HHS/United States
R01 HL61371/HL/NHLBI NIH HHS/United States
R01 HL64793/HL/NHLBI NIH HHS/United States
R01 HL064793/HL/NHLBI NIH HHS/United States
R01 GM36065/GM/NIGMS NIH HHS/United States
R01 GM38545/GM/NIGMS NIH HHS/United States
R01 GM038545/GM/NIGMS NIH HHS/United States
R01 HL096670/HL/NHLBI NIH HHS/United States
R01 GM036065/GM/NIGMS NIH HHS/United States
R01 GM038545-23/GM/NIGMS NIH HHS/United States
P01 HL70295/HL/NHLBI NIH HHS/United States
R01 HL081190/HL/NHLBI NIH HHS/United States
R01 HL061371/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
EMBO J. 2011 May 13;30(12):2490-500. doi: 10.1038/emboj.2011.147.
Author Address: Department of Pharmacology and Vascular Biology and Therapeutics Program, Yale University School of Medicine, New Haven, CT, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1376
Author: Harrison, O. J., Jin, X., Hong, S., Bahna, F., Ahlsen, G., Brasch, J., Wu, Y., Vendome, J., Felsovalyi, K., Hampton, C. M., Troyanovsky, R. B., Ben-Shaul, A., Frank, J., Troyanovsky, S. M., Shapiro, L. and Honig, B.
Year: 2011
Title: The extracellular architecture of adherens junctions revealed by crystal structures of type I cadherins
Journal: Structure
Volume: 19
Issue: 2
Pages: 244-56
Epub Date: 2011/02/09
Date: Feb 9
Short Title: The extracellular architecture of adherens junctions revealed by crystal structures of type I cadherins
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2010.11.016
PMCID: PMC3070544
NIHMSID: NIHMS274056
Accession Number: 21300292
Keywords: Adherens Junctions/*metabolism/*ultrastructure
Animals
Binding Sites
Cadherins/chemistry/genetics/*metabolism
Cell Adhesion
Cells, Cultured
Crystallography, X-Ray
Dimerization
Escherichia coli
Gene Expression
Humans
Liposomes/*metabolism
Mice
Models, Molecular
Mutagenesis, Site-Directed
Protein Binding
Protein Structure, Tertiary
Recombinant Proteins/genetics/*metabolism
Stereoisomerism
Abstract: Adherens junctions, which play a central role in intercellular adhesion, comprise clusters of type I classical cadherins that bind via extracellular domains extended from opposing cell surfaces. We show that a molecular layer seen in crystal structures of E- and N-cadherin ectodomains reported here and in a previous C-cadherin structure corresponds to the extracellular architecture of adherens junctions. In all three ectodomain crystals, cadherins dimerize through a trans adhesive interface and are connected by a second, cis, interface. Assemblies formed by E-cadherin ectodomains coated on liposomes also appear to adopt this structure. Fluorescent imaging of junctions formed from wild-type and mutant E-cadherins in cultured cells confirm conclusions derived from structural evidence. Mutations that interfere with the trans interface ablate adhesion, whereas cis interface mutations disrupt stable junction formation. Our observations are consistent with a model for junction assembly involving strong trans and weak cis interactions localized in the ectodomain.
Notes: 1878-4186
Harrison, Oliver J
Jin, Xiangshu
Hong, Soonjin
Bahna, Fabiana
Ahlsen, Goran
Brasch, Julia
Wu, Yinghao
Vendome, Jeremie
Felsovalyi, Klara
Hampton, Cheri M
Troyanovsky, Regina B
Ben-Shaul, Avinoam
Frank, Joachim
Troyanovsky, Sergey M
Shapiro, Lawrence
Honig, Barry
R01 GM062270/GM/NIGMS NIH HHS/United States
T32 GM082797/GM/NIGMS NIH HHS/United States
R56 AR044016/AR/NIAMS NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 GM062270-09/GM/NIGMS NIH HHS/United States
R01 AR044016/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Structure. 2011 Feb 9;19(2):244-56. doi: 10.1016/j.str.2010.11.016.
Author Address: Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1507
Author: Harrison, O. J., Vendome, J., Brasch, J., Jin, X., Hong, S., Katsamba, P. S., Ahlsen, G., Troyanovsky, R. B., Troyanovsky, S. M., Honig, B. and Shapiro, L.
Year: 2012
Title: Nectin ectodomain structures reveal a canonical adhesive interface
Journal: Nat Struct Mol Biol
Volume: 19
Issue: 9
Pages: 906-15
Epub Date: 2012/08/21
Date: Sep
Short Title: Nectin ectodomain structures reveal a canonical adhesive interface
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb.2366
PMCID: PMC3443293
NIHMSID: NIHMS395153
Accession Number: 22902367
Keywords: Animals
Antigens, Neoplasm/*metabolism
Cell Adhesion
Cell Adhesion Molecules/*chemistry/*metabolism
Cell Line
Crystallography, X-Ray
Humans
Mice
Models, Molecular
Nectins
Neoplasm Proteins/*metabolism
Protein Binding
Protein Multimerization
Protein Structure, Tertiary
Receptors, Virus/*metabolism
Abstract: Nectins are immunoglobulin superfamily glycoproteins that mediate intercellular adhesion in many vertebrate tissues. Homophilic and heterophilic interactions between nectin family members help mediate tissue patterning. We determined the homophilic binding affinities and heterophilic specificities of all four nectins and the related protein nectin-like 5 (Necl-5) from human and mouse, revealing a range of homophilic interaction strengths and a defined heterophilic specificity pattern. To understand the molecular basis of their adhesion and specificity, we determined the crystal structures of natively glycosylated full ectodomains or adhesive fragments of all four nectins and Necl-5. All of the crystal structures revealed dimeric nectins bound through a stereotyped interface that was previously proposed to represent a cis dimer. However, conservation of this interface and the results of targeted cross-linking experiments showed that this dimer probably represents the adhesive trans interaction. The structure of the dimer provides a simple molecular explanation for the adhesive binding specificity of nectins.
Notes: 1545-9985
Harrison, Oliver J
Vendome, Jeremie
Brasch, Julia
Jin, Xiangshu
Hong, Soonjin
Katsamba, Phinikoula S
Ahlsen, Goran
Troyanovsky, Regina B
Troyanovsky, Sergey M
Honig, Barry
Shapiro, Lawrence
R01 GM062270/GM/NIGMS NIH HHS/United States
AR44016/AR/NIAMS NIH HHS/United States
R01 AR057992/AR/NIAMS NIH HHS/United States
AR057992/AR/NIAMS NIH HHS/United States
R56 AR044016/AR/NIAMS NIH HHS/United States
R01 AR044016/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Nat Struct Mol Biol. 2012 Sep;19(9):906-15. doi: 10.1038/nsmb.2366. Epub 2012 Aug 19.
Author Address: Department of Biochemistry and Molecular Biophysics, Columbia University, New York, New York, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 758
Author: Hartmann, S. and Hofsteenge, J.
Year: 2000
Title: Properdin, the positive regulator of complement, is highly C-mannosylated
Journal: J Biol Chem
Volume: 275
Issue: 37
Pages: 28569-74
Epub Date: 2000/07/06
Date: Sep 15
Short Title: Properdin, the positive regulator of complement, is highly C-mannosylated
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M001732200
Accession Number: 10878002
Keywords: Amino Acid Sequence
Humans
Mannose/metabolism
Molecular Sequence Data
Properdin/*chemistry
Abstract: Properdin is the positive regulator of the alternative pathway of complement activation. The 53-kDa protein is essentially composed of six thrombospondin type 1 repeats, all of which contain the WXXW motif, the recognition sequence for C-mannosylation. C-Mannosylation is a post-translational modification of tryptophan residues in which, in contrast to the well known N- and O-glycosylation, the carbohydrate is attached via a C-C bond to C-2 of the indole moiety of tryptophan. C-Mannosylation was first found in human RNase 2 and interleukin-12. The terminal complement proteins C6-C9 also carry this modification as part of their thrombospondin type 1 repeats. We studied the C-mannosylation pattern of human properdin by mass spectrometry and Edman degradation. Properdin contains 20 tryptophans of which 17 are part of a WXXW motif. Fourteen tryptophans were found to be modified 100%. This is the first example of a protein in which the majority of tryptophan residues occurs in the C-mannosylated form. These results show that C-mannosylated proteins occur at several steps along the complement activation cascade. Therefore, this system would be ideal to investigate the function of C-mannosylation.
Notes: Hartmann, S
Hofsteenge, J
Journal Article
United States
J Biol Chem. 2000 Sep 15;275(37):28569-74. doi: 10.1074/jbc.M001732200.
Author Address: Friedrich-Miescher Institut, CH-4058 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 436
Author: Hase, S., Kikuchi, N., Ikenaka, T. and Inoue, K.
Year: 1985
Title: Structures of sugar chains of the third component of human complement
Journal: J Biochem
Volume: 98
Issue: 4
Pages: 863-74
Epub Date: 1985/10/01
Date: Oct
Short Title: Structures of sugar chains of the third component of human complement
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a135366
Accession Number: 4077844
Keywords: Acetates
Acetic Acid
Amino Acids/analysis
Carbohydrate Sequence
Carbohydrates/analysis
Chemical Phenomena
Chemistry
Chromatography, High Pressure Liquid
*Complement C3
Humans
Abstract: Human C3, the third component of human complement, contained mannose and N-acetylglucosamine as sugar components. The sugar chains were liberated from the polypeptide chains by hydrazinolysis, and the free amino groups were N-acetylated. The reducing end residues of the sugar chains thus obtained were tagged with 2-aminopyridine, and the pyridylamino (PA-) derivatives of sugar chains were separated by high-performance liquid chromatography. The structures of purified PA-sugar chains were analyzed by a combination of stepwise exoglycosidase digestions, size determination by paper electrophoresis, methylation analysis, Smith degradation, and partial acetolysis. These results showed that C3 contained two high-mannose type sugar chains ranging from Man5GlcNAc2 to Man9GlcNAc2. Analyses of the sugar chains of alpha- and beta-chains of C3 indicated that the alpha-chain contained mainly Man8GlcNAc2 and Man9GlcNAc2, while the beta-chain contained mainly Man5GlcNAc2 and Man6GlcNAc2.
Notes: Hase, S
Kikuchi, N
Ikenaka, T
Inoue, K
Journal Article
England
J Biochem. 1985 Oct;98(4):863-74. doi: 10.1093/oxfordjournals.jbchem.a135366.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 398
Author: Hase, S., Natsuka, S., Oku, H. and Ikenaka, T.
Year: 1987
Title: Identification method for twelve oligomannose-type sugar chains thought to be processing intermediates of glycoproteins
Journal: Anal Biochem
Volume: 167
Issue: 2
Pages: 321-6
Epub Date: 1987/12/01
Date: Dec
Short Title: Identification method for twelve oligomannose-type sugar chains thought to be processing intermediates of glycoproteins
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/0003-2697(87)90171-0
Accession Number: 3442327
Keywords: Carbohydrate Sequence
Chromatography, High Pressure Liquid
Complement C3/isolation & purification
Glycoproteins/*isolation & purification
Glycosides/*isolation & purification
Humans
Mannosides/*isolation & purification
Molecular Sequence Data
Oligosaccharides/*isolation & purification
Abstract: A mixture of the pyridylamino (PA) derivatives of 12 oligomannose-type sugar chains was fractionated into five fractions (mannose5N-acetylglucosamine2-PA approximately mannose9N-acetylglucosamine2-PA) by size-fractionation HPLC with a MicroPak AX-5 column. Each fraction thus obtained was then analyzed by reversed-phase HPLC with a Cosmosil 5C18-P column. In this way, the 12 PA-oligomannose-type sugar chains were completely separated from each other. The method was used to identify the structures of oligomannose-type sugar chains of human C3, the third component of human complement.
Notes: Hase, S
Natsuka, S
Oku, H
Ikenaka, T
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Biochem. 1987 Dec;167(2):321-6. doi: 10.1016/0003-2697(87)90171-0.
Author Address: Department of Chemistry, Osaka University College of Science, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1204
Author: Hassan, M. I., Bilgrami, S., Kumar, V., Singh, N., Yadav, S., Kaur, P. and Singh, T. P.
Year: 2008
Title: Crystal structure of the novel complex formed between zinc alpha2-glycoprotein (ZAG) and prolactin-inducible protein (PIP) from human seminal plasma
Journal: J Mol Biol
Volume: 384
Issue: 3
Pages: 663-72
Epub Date: 2008/10/22
Date: Dec 19
Short Title: Crystal structure of the novel complex formed between zinc alpha2-glycoprotein (ZAG) and prolactin-inducible protein (PIP) from human seminal plasma
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2008.09.072
Accession Number: 18930737
Keywords: Adipokines
Binding Sites
Carrier Proteins/*chemistry
Crystallography, X-Ray
Glycoproteins/*chemistry
Glycosylation
Humans
Hydrogen Bonding
Ions
Membrane Transport Proteins
Models, Molecular
Molecular Conformation
Protein Binding
Protein Interaction Mapping
Protein Structure, Tertiary
Semen/*metabolism
Abstract: This is the first report on the formation of a complex between zinc alpha2-glycoprotein (ZAG) and prolactin-inducible protein (PIP). The complex was purified from human seminal plasma and crystallized using 20% polyethylene glycol 9000 and 5% hexaethylene glycol. The structure of the complex has been determined using X-ray crystallographic method and refined to an R(cryst) of 0.199 (R(free)=0.239). The structure of ZAG is broadly similar to the structure of serum ZAG. The scaffolding of PIP consists of seven beta-strands that are organized in the form of two antiparallel beta-pleated sheets, resulting in the formation of a sandwiched beta-sheet. The amino acid sequence of PIP contains one potential N-glycosylation site at Asn77, and the same is found glycosylated with four sugar residues. The structure of the complex shows that the beta-structure of PIP is ideally aligned with the beta-structure of domain alpha3 of ZAG to form a long interface between two proteins. The proximal beta-strands at the long interface are arranged in an antiparallel manner. There are 12 hydrogen bonds and three salt bridges between ZAG and PIP. At the two ends of vertical interface, two salt bridges are formed between pairs of Lys41-Asp233 and Lys68-Glu229. On the perpendicular interface involving alpha1-alpha2 domains of ZAG and a loop of PIP, another salt bridge is formed. The internal space at the corner of the L-shaped structure is filled with solvent molecules including a carbonate ion. The overall buried area in the complex is approximately 914 A(2), which is considerably higher than the 660 A(2) reported for the class I major histocompatibility complex structures.
Notes: 1089-8638
Hassan, Md Imtaiyaz
Bilgrami, Sameeta
Kumar, Vijay
Singh, Nagendra
Yadav, Savita
Kaur, Punit
Singh, T P
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2008 Dec 19;384(3):663-72. doi: 10.1016/j.jmb.2008.09.072. Epub 2008 Oct 9.
Author Address: Department of Biophysics, All India Institute of Medical Sciences, New Delhi 110029, India.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1181
Author: Hatherley, D., Graham, S. C., Turner, J., Harlos, K., Stuart, D. I. and Barclay, A. N.
Year: 2008
Title: Paired receptor specificity explained by structures of signal regulatory proteins alone and complexed with CD47
Journal: Mol Cell
Volume: 31
Issue: 2
Pages: 266-77
Epub Date: 2008/07/29
Date: Jul 25
Short Title: Paired receptor specificity explained by structures of signal regulatory proteins alone and complexed with CD47
Alternate Journal: Molecular cell
ISSN: 1097-2765
DOI: 10.1016/j.molcel.2008.05.026
Accession Number: 18657508
Keywords: Amino Acid Sequence
Animals
CD47 Antigen/*chemistry
CHO Cells
Cell Communication
Cricetinae
Cricetulus
Crystallography, X-Ray
Macrophages/cytology
Models, Molecular
Molecular Sequence Data
Polymorphism, Genetic
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Immunologic/*chemistry/*metabolism
Abstract: CD47 is a widely distributed cell-surface protein that acts a marker of self through interactions of myeloid and neural cells. We describe the high-resolution X-ray crystallographic structures of the immunoglobulin superfamily domain of CD47 alone and in complex with the N-terminal ligand-binding domain of signal regulatory protein alpha (SIRPalpha). The unusual and convoluted interacting face of CD47, comprising the N terminus and loops at the end of the domain, intercalates with the corresponding regions in SIRPalpha. We have also determined structures of the N-terminal domains of SIRPbeta, SIRPbeta(2), and SIRPgamma; proteins that are closely related to SIRPalpha but bind CD47 with negligible or reduced affinity. These results explain the specificity of CD47 for the SIRP family of paired receptors in atomic detail. Analysis of SIRPalpha polymorphisms suggests that these, as well as the activating SIRPs, may have evolved to counteract pathogen binding to the inhibitory SIRPalpha receptor.
Notes: 1097-4164
Hatherley, Deborah
Graham, Stephen C
Turner, Jessie
Harlos, Karl
Stuart, David I
Barclay, A Neil
G0500365/Medical Research Council/United Kingdom
G0500367/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Mol Cell. 2008 Jul 25;31(2):266-77. doi: 10.1016/j.molcel.2008.05.026.
Author Address: Sir William Dunn School of Pathology, University of Oxford, Oxford, OX1 3RE, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1554
Author: Hatherley, D., Lea, S. M., Johnson, S. and Barclay, A. N.
Year: 2013
Title: Structures of CD200/CD200 receptor family and implications for topology, regulation, and evolution
Journal: Structure
Volume: 21
Issue: 5
Pages: 820-32
Epub Date: 2013/04/23
Date: May 7
Short Title: Structures of CD200/CD200 receptor family and implications for topology, regulation, and evolution
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2013.03.008
PMCID: PMC3664923
NIHMSID: EMS53259
Accession Number: 23602662
Keywords: Amino Acid Sequence
Antigens, CD/*chemistry/metabolism
Antigens, Surface/*chemistry/metabolism
Biological Evolution
Crystallography, X-Ray
Membrane Glycoproteins/chemistry/metabolism
Myeloid Cells/metabolism
Surface Plasmon Resonance
Abstract: CD200 is a widely distributed membrane glycoprotein that regulates myeloid cell activity through its interaction with an inhibitory receptor (CD200R). The interaction is of interest as a target for treating excessive inflammation and for treating leukemia. There are closely related proteins to CD200R that give activating signals making this a "paired receptor." We report X-ray crystallography structures for the inhibitory CD200R, the activating receptor CD200RLa, and a complex between CD200R and CD200. Both CD200 and CD200R contain two Ig-like domains and interact through their NH₂ terminal domains compatible with immunological synapse-like interactions occurring between myeloid cells and other CD200-expressing cells. The failure of the activating receptor to bind CD200 resides in subtle changes around the interface. CD200 has been acquired by herpes viruses to mimic the host interaction. CD200R has evolved rapidly presumably driven by pathogen pressure but it may also be important in homeostasis through interactions with commensal bacteria.
Notes: 1878-4186
Hatherley, Deborah
Lea, Susan M
Johnson, Steven
Barclay, A Neil
G0900888/Medical Research Council/United Kingdom
G9826026/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Structure. 2013 May 7;21(5):820-32. doi: 10.1016/j.str.2013.03.008. Epub 2013 Apr 18.
Author Address: Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 892
Author: Hatsell, S., Medina, L., Merola, J., Haltiwanger, R. and Cowin, P.
Year: 2003
Title: Plakoglobin is O-glycosylated close to the N-terminal destruction box
Journal: J Biol Chem
Volume: 278
Issue: 39
Pages: 37745-52
Epub Date: 2003/07/09
Date: Sep 26
Short Title: Plakoglobin is O-glycosylated close to the N-terminal destruction box
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M301346200
Accession Number: 12847106
Keywords: Amino Acid Sequence
Base Sequence
Cytoskeletal Proteins/*chemistry/physiology
Desmoplakins
Glycogen Synthase Kinase 3/chemistry
Glycogen Synthase Kinase 3 beta
Glycosylation
Humans
Molecular Sequence Data
Trans-Activators/chemistry
beta Catenin
gamma Catenin
Abstract: Plakoglobin provides a key linkage in protein chains that connect desmosomal and classical cadherins to the cytoskeleton. It is also present in a significant cytosolic pool that has the capacity to impact on canonical Wnt signaling by competing for interaction with partner proteins of beta-catenin. The closely related protein, beta-catenin, is rapidly targeted for proteasomal degradation by phosphorylation of a "destruction box" within the N-terminal domain. Inhibition of this process forms the basis of Wnt signaling. This destruction box is also found in the N-terminal domain of plakoglobin. We report that plakoglobin is modified by the addition of O-GlcNAc at a single site in close proximity to the destruction box. O-GlcNAc modification has been proposed to counteract phosphorylation, provide protection from proteasomal degradation, mediate signal transduction, silence transcription, and regulate multimolecular protein assembly. This finding has potential implications for understanding the roles of plakoglobin.
Notes: Hatsell, Sarah
Medina, Lillian
Merola, Joe
Haltiwanger, Robert
Cowin, Pamela
GM47429/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2003 Sep 26;278(39):37745-52. doi: 10.1074/jbc.M301346200. Epub 2003 Jul 7.
Author Address: Department of Cell Biology, New York University Medical School, New York, New York 10016, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1370
Author: Hausmann, J., Kamtekar, S., Christodoulou, E., Day, J. E., Wu, T., Fulkerson, Z., Albers, H. M., van Meeteren, L. A., Houben, A. J., van Zeijl, L., Jansen, S., Andries, M., Hall, T., Pegg, L. E., Benson, T. E., Kasiem, M., Harlos, K., Kooi, C. W., Smyth, S. S., Ovaa, H., Bollen, M., Morris, A. J., Moolenaar, W. H. and Perrakis, A.
Year: 2011
Title: Structural basis of substrate discrimination and integrin binding by autotaxin
Journal: Nat Struct Mol Biol
Volume: 18
Issue: 2
Pages: 198-204
Epub Date: 2011/01/18
Date: Feb
Short Title: Structural basis of substrate discrimination and integrin binding by autotaxin
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb.1980
PMCID: PMC3064516
NIHMSID: NIHMS251225
Accession Number: 21240271
Keywords: Amino Acid Sequence
Animals
Binding Sites
Catalytic Domain
Cell Line
Crystallography, X-Ray
Humans
Integrins/*metabolism
Lysophospholipids/metabolism
Molecular Sequence Data
Mutation
Phosphoric Diester Hydrolases/*chemistry/genetics/*metabolism
Protein Binding
Protein Structure, Tertiary
Pyrophosphatases/*chemistry/genetics/*metabolism
Rats
Substrate Specificity
Abstract: Autotaxin (ATX, also known as ectonucleotide pyrophosphatase/phosphodiesterase-2, ENPP2) is a secreted lysophospholipase D that generates the lipid mediator lysophosphatidic acid (LPA), a mitogen and chemoattractant for many cell types. ATX-LPA signaling is involved in various pathologies including tumor progression and inflammation. However, the molecular basis of substrate recognition and catalysis by ATX and the mechanism by which it interacts with target cells are unclear. Here, we present the crystal structure of ATX, alone and in complex with a small-molecule inhibitor. We have identified a hydrophobic lipid-binding pocket and mapped key residues for catalysis and selection between nucleotide and phospholipid substrates. We have shown that ATX interacts with cell-surface integrins through its N-terminal somatomedin B-like domains, using an atypical mechanism. Our results define determinants of substrate discrimination by the ENPP family, suggest how ATX promotes localized LPA signaling and suggest new approaches for targeting ATX with small-molecule therapeutic agents.
Notes: 1545-9985
Hausmann, Jens
Kamtekar, Satwik
Christodoulou, Evangelos
Day, Jacqueline E
Wu, Tao
Fulkerson, Zachary
Albers, Harald M H G
van Meeteren, Laurens A
Houben, Anna J S
van Zeijl, Leonie
Jansen, Silvia
Andries, Maria
Hall, Troii
Pegg, Lyle E
Benson, Timothy E
Kasiem, Mobien
Harlos, Karl
Kooi, Craig W Vander
Smyth, Susan S
Ovaa, Huib
Bollen, Mathieu
Morris, Andrew J
Moolenaar, Wouter H
Perrakis, Anastassis
5T32HL0727-43-06/HL/NHLBI NIH HHS/United States
GM50388/GM/NIGMS NIH HHS/United States
F30 HL099272/HL/NHLBI NIH HHS/United States
P20 RR021954-03/RR/NCRR NIH HHS/United States
R01 HL078663/HL/NHLBI NIH HHS/United States
HL078663/HL/NHLBI NIH HHS/United States
R01 GM094155/GM/NIGMS NIH HHS/United States
R01 GM050388-18/GM/NIGMS NIH HHS/United States
GM094155/GM/NIGMS NIH HHS/United States
P20RR021954/RR/NCRR NIH HHS/United States
S10 RR024598/RR/NCRR NIH HHS/United States
R01 GM050388/GM/NIGMS NIH HHS/United States
R01 HL078663-06/HL/NHLBI NIH HHS/United States
P20 RR021954/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Struct Mol Biol. 2011 Feb;18(2):198-204. doi: 10.1038/nsmb.1980. Epub 2011 Jan 16.
Author Address: Division of Biochemistry, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1888
Author: Hayakawa, K., Hirosawa, M., Tani, R., Yoneda, C., Tanaka, S. and Shiota, K.
Year: 2017
Title: H2A O-GlcNAcylation at serine 40 functions genomic protection in association with acetylated H2AZ or γH2AX
Journal: Epigenetics Chromatin
Volume: 10
Issue: 1
Pages: 51
Epub Date: 2017/11/01
Date: Oct 30
Short Title: H2A O-GlcNAcylation at serine 40 functions genomic protection in association with acetylated H2AZ or γH2AX
Alternate Journal: Epigenetics & chromatin
ISSN: 1756-8935
DOI: 10.1186/s13072-017-0157-x
PMCID: PMC5663087
Accession Number: 29084613
Keywords: Acetylation
Animals
Cell Line
DNA Repair
Embryonic Stem Cells/metabolism
*Genomic Instability
Histones/genetics/*metabolism
Mice
Mutation
Phosphorylation
Protein Isoforms/genetics/metabolism
*Protein Processing, Post-Translational
*Canonical histone
*DNA damage
*DNA repair
*Histone modification
*Histone variant
*O-GlcNAcylation
Abstract: BACKGROUND: We have previously reported a novel O-GlcNAc modification at serine 40 (S40) of H2A (H2AS40Gc). S40-type H2A isoforms susceptible to O-GlcNAcylation are evolutionarily new and restricted to the viviparous animals; however, the biological function of H2AS40Gc is largely unknown. H2A isoforms are consisted of S40 and alanine 40 (A40) type and this residue on H2A is located in the L1 of the globular domain, which is also known as a variable portion that distinguishes between the canonical and non-canonical H2A variants. In this study, by considering the similarity between the S40-type H2A and histone H2A variants, we explored the function of H2AS40Gc in mouse embryonic stem cells (mESCs). RESULTS: We found several similarities between the S40-type H2A isoforms and histone H2A variants such H2AZ and H2AX. mRNA of S40-type H2A isoforms (H2A1 N and H2A3) had a poly(A) tail and was produced throughout the cell cycle in contrast to that of A40-type. Importantly, H2AS40Gc level increased owing to chemical-induced DNA damage, similar to phosphorylated H2AX (γH2AX) and acetylated H2AZ (AcH2AZ). H2AS40Gc was accumulated at the restricted area (± 1.5 kb) of DNA damage sites induced by CRISPR/CAS9 system in contrast to accumulation of γH2AX, which was widely scattered. Overexpression of the wild-type (WT) H2A3, but not the S40 to A40 mutation (S40A-mutant), protected the mESC genome against chemical-induced DNA damage. Furthermore, 3 h after the DNA damage treatment, the genome was almost recovered in WT mESCs, whereas the damage advanced further in the S40A-mutant mESCs, suggesting functions of H2AS40Gc in the DNA repair mechanism. Furthermore, the S40A mutant prevented the accumulation of the DNA repair apparatus such as DNA-PKcs and Rad51 at the damage site. Co-immunoprecipitation experiment in WT and S40A-mutant mESCs revealed that H2AS40Gc physiologically bound to AcH2AZ at the initial phase upon DNA damage, followed by binding with γH2AX during the DNA damage repair process. CONCLUSIONS: These data suggest that H2AS40Gc functions to maintain genome integrity through the DNA repair mechanism in association with AcH2AZ and γH2AX.
Notes: 1756-8935
Hayakawa, Koji
Orcid: 0000-0003-0453-1303
Hirosawa, Mitsuko
Tani, Ruiko
Yoneda, Chikako
Tanaka, Satoshi
Shiota, Kunio
Journal Article
Research Support, Non-U.S. Gov't
Epigenetics Chromatin. 2017 Oct 30;10(1):51. doi: 10.1186/s13072-017-0157-x.
Author Address: Laboratory of Cellular Biochemistry, Department of Animal Resource Sciences/Veterinary Medical Sciences, The University of Tokyo, Tokyo, 113-8657, Japan.
Laboratory of Cellular Biochemistry, Department of Animal Resource Sciences/Veterinary Medical Sciences, The University of Tokyo, Tokyo, 113-8657, Japan. ashiota@mail.ecc.u-tokyo.ac.jp.
Waseda Research Institute for Science and Engineering, Waseda University, Tokyo, 169-8555, Japan. ashiota@mail.ecc.u-tokyo.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1444
Author: Hayashi, H. and Yamashita, Y.
Year: 2012
Title: Role of N-glycosylation in cell surface expression and protection against proteolysis of the intestinal anion exchanger SLC26A3
Journal: Am J Physiol Cell Physiol
Volume: 302
Issue: 5
Pages: C781-95
Epub Date: 2011/12/14
Date: Mar 1
Short Title: Role of N-glycosylation in cell surface expression and protection against proteolysis of the intestinal anion exchanger SLC26A3
Alternate Journal: American journal of physiology. Cell physiology
ISSN: 0363-6143
DOI: 10.1152/ajpcell.00165.2011
Accession Number: 22159084
Keywords: Amino Acid Substitution
Animals
Asparagine/genetics/metabolism
CHO Cells
Cell Membrane/genetics/*metabolism
Chloride-Bicarbonate Antiporters/genetics/*metabolism
Congenital Disorders of Glycosylation/*metabolism
Cricetinae
Cricetulus
Glutamine/genetics/metabolism
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Hydrogen-Ion Concentration
Intestinal Absorption/physiology
Intestinal Mucosa/metabolism
Mutagenesis, Site-Directed
Oligosaccharides/*metabolism
*Proteolysis
Sulfate Transporters
Abstract: SLC26A3 is a Cl(-)/HCO(3)(-) exchanger that plays a major role in Cl(-) absorption from the intestine. Its mutation causes congenital chloride-losing diarrhea. It has been shown that SLC26A3 are glycosylated, with the attached carbohydrate being extracellular and perhaps modulating function. However, the role of glycosylation has yet to be clearly determined. We used the approaches of biochemical modification and site-directed mutagenesis to prevent glycosylation. Deglycosylation experiments with glycosidases indicated that the mature glycosylated form of SLC26A3 exists at the plasma membrane, and a putative large second extracellular loop contains all of the N-linked carbohydrates. Deglycosylation of SLC26A3 causes depression of transport activity compared with wild-type, although robust intracellular pH changes were still observed, suggesting that N-glycosylation is not absolutely necessary for transport activity. To localize glycosylation sites, we mutated the five consensus sites by replacing asparagine (N) with glutamine. Immnoblotting suggests that SLC26A3 is glycosylated at N153, N161, and N165. Deglycosylation of SLC26A3 causes a defect in cell surface processing with decreased cell surface expression. We also assessed whether SLC26A3 is protected from tryptic digestion. While the mature glycosylated SLC26A3 showed little breakdown after treatment with trypsin, deglycosylated SLC26A3 exhibited increased susceptibility to trypsin, suggesting that the oligosaccharides protect SLC26A3 from tryptic digestion. In conclusion, our data indicate that N-glycosylation of SLC26A3 is important for cell surface expression and for protection from proteolytic degradation that may contribute to the understanding of pathogenesis of congenital disorders of glycosylation.
Notes: 1522-1563
Hayashi, Hisayoshi
Yamashita, Yukari
Journal Article
Research Support, Non-U.S. Gov't
United States
Am J Physiol Cell Physiol. 2012 Mar 1;302(5):C781-95. doi: 10.1152/ajpcell.00165.2011. Epub 2011 Dec 7.
Author Address: Laboratory of Physiology, School of Food and Nutritional Sciences, Univ. of Shizuoka, Suruga-ku, Shizuoka, Japan. hayashih@smail.u-shizuoka-ken.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 239
Author: Hayes, G. R., Enns, C. A. and Lucas, J. J.
Year: 1992
Title: Identification of the O-linked glycosylation site of the human transferrin receptor
Journal: Glycobiology
Volume: 2
Issue: 4
Pages: 355-9
Epub Date: 1992/08/01
Date: Aug
Short Title: Identification of the O-linked glycosylation site of the human transferrin receptor
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/2.4.355
Accession Number: 1421757
Keywords: Amino Acid Sequence
Binding Sites
Carbohydrate Conformation
Female
Glycoproteins/*chemistry
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Peptide Fragments/chemistry
Placenta/chemistry
Receptors, Transferrin/*chemistry
Threonine/metabolism
Trypsin
Abstract: The human transferrin receptor is a glycoprotein containing three N-linked and one O-linked glycosylation sites. Tryptic digestion of the receptor, followed by chromatography on BioGel P-2 and reverse-phase HPLC, yields a glycopeptide (amino acids 101-120) containing the O-linked site. Amino acid sequence analysis reveals that the site of O-glycosylation is Thr-104. Mass spectral analysis is consistent with the presence of a Gal-GalNAc core with predominantly two sialic acid residues.
Notes: Hayes, G R
Enns, C A
Lucas, J J
GM 43111/GM/NIGMS NIH HHS/United States
RR0317/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1992 Aug;2(4):355-9. doi: 10.1093/glycob/2.4.355.
Author Address: Department of Biochemistry and Molecular Biology, SUNY Health Science Center, Syracuse 13210.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 537
Author: Hayes, G. R., Williams, A., Costello, C. E., Enns, C. A. and Lucas, J. J.
Year: 1995
Title: The critical glycosylation site of human transferrin receptor contains a high-mannose oligosaccharide
Journal: Glycobiology
Volume: 5
Issue: 2
Pages: 227-32
Epub Date: 1995/03/01
Date: Mar
Short Title: The critical glycosylation site of human transferrin receptor contains a high-mannose oligosaccharide
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/5.2.227
Accession Number: 7780197
Keywords: 3T3 Cells
Amino Acid Sequence
Animals
Asparagine/analysis
Binding Sites
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Glycosylation
Hexosaminidases
Humans
Mannose/*analysis
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
Mass Spectrometry
Mice
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Mapping
Receptors, Transferrin/biosynthesis/*chemistry/*metabolism
Recombinant Proteins/biosynthesis/chemistry/metabolism
Transfection
Trypsin
Abstract: The human transferrin receptor (TfR) contains three N-linked oligosaccharides and glycosylation is required for the proper folding and function of the molecule. Earlier studies demonstrated that the oligosaccharide at Asn-727 is vital for the production of fully active TfR. The oligosaccharide(s) present at this site have been analysed using a combination of site-directed mutagenesis and chemical analysis. Wild-type TfR and mutants containing only the Asn-727 site or missing all three sites were transfected into mouse 3T3 cells and receptors were analysed by endo-N-acetylglucosaminidase H (Endo-H) digestion, SDS-PAGE and immunoblotting. These studies suggested that the Asn-727 site contains high-mannose or Endo-H-sensitive hybrid oligosaccharides. Glycosylation of Asn-727 found in the TfR purified from human placentae was analysed by high-pH anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) and mass spectrometry following tryptic digestion, peptide purification via reverse-phase high-performance liquid chromatography (RP-HPLC) and peptide sequencing. HPAE-PAD showed the presence of a series of high-mannose oligosaccharides. Mass spectrometry confirmed these observations, but also showed the presence of an 80 Da anionic moiety on a fraction of the oligosaccharides.
Notes: Hayes, G R
Williams, A
Costello, C E
Enns, C A
Lucas, J J
DK 40608/DK/NIDDK NIH HHS/United States
GM 43111/GM/NIGMS NIH HHS/United States
RR00317/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1995 Mar;5(2):227-32. doi: 10.1093/glycob/5.2.227.
Author Address: Department of Biochemistry and Molecular Biology, SUNY Health Science Center at Syracuse 13210, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 642
Author: Hayes, G. R., Williams, A. M., Lucas, J. J. and Enns, C. A.
Year: 1997
Title: Structure of human transferrin receptor oligosaccharides: conservation of site-specific processing
Journal: Biochemistry
Volume: 36
Issue: 17
Pages: 5276-84
Epub Date: 1997/04/29
Date: Apr 29
Short Title: Structure of human transferrin receptor oligosaccharides: conservation of site-specific processing
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi962884y
Accession Number: 9136890
Keywords: 3T3 Cells
Animals
Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/chemistry
Cells, Cultured
Hexosaminidases/metabolism
Humans
Mice
Models, Molecular
Molecular Sequence Data
Oligosaccharides/*chemistry
Receptors, Transferrin/*chemistry
Structure-Activity Relationship
Abstract: The human transferrin receptor (TfR) has three N-linked oligosaccharides. A combination of site-directed mutagenesis and carbohydrate and protein chemistry was used to characterize the structures of the N-linked oligosaccharides and to map their locations. We find that the type of oligosaccharide at each position was unique for that particular site. Human TfR isolated from placentae was used to characterize the structure of the oligosaccharides found in the native TfR. Following digestion of purified TfR with trypsin, individual peptides were obtained via RP-HPLC and were assayed for monosaccharides by strong acid hydrolysis and HPAE-PAD. Peptides containing carbohydrate were subjected to amino acid sequencing to identify the specific Asn residue. The oligosaccharides at Asn 251 are of the complex type. HPAE-PAD and FACE analysis suggests that they are triantennary and trisialylated with core fucosylation. The glycopeptide containing the site at Asn 317 was obtained after limited tryptic digestion and RP-HPLC. FACE analysis reveals predominantly a family of sialylated hybrid oligosaccharides. The consensus sequences for each N-linked site were mutated in various combinations and the resultant TfRs expressed in mouse 3T3 cells. Endoglycosidase H digestion of the mutated TfRs indicates that the pattern of oligosaccharides is consistent with the type of oligosaccharides found at each position in human tissue and the glycosylation of one site does not directly affect the glycosylation of other sites. Previous studies indicated that the oliosaccharide at Asn 727 was high-mannose type [Hayes, G. R., et al. (1995) Glycobiology 5, 227-232]. These results indicate that the type of oligosaccharide found at each site is most dependent on the environment surrounding it.
Notes: Hayes, G R
Williams, A M
Lucas, J J
Enns, C A
DK40608/DK/NIDDK NIH HHS/United States
GM43111/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1997 Apr 29;36(17):5276-84. doi: 10.1021/bi962884y.
Author Address: Department of Biochemistry and Molecular Biology, State University of New York Health Sciences Center Syracuse, New York 13201, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 222
Author: Hayes, M. L. and Castellino, F. J.
Year: 1979
Title: Carbohydrate of the human plasminogen variants. III. Structure of the O-glycosidically linked oligosaccharide unit
Journal: J Biol Chem
Volume: 254
Issue: 18
Pages: 8777-80
Epub Date: 1979/09/25
Date: Sep 25
Short Title: Carbohydrate of the human plasminogen variants. III. Structure of the O-glycosidically linked oligosaccharide unit
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 479158
Keywords: Amino Acid Sequence
Amino Acids/analysis
Carbohydrate Sequence
Carbohydrates/analysis
*Genetic Variation
Glycopeptides/analysis
Glycosides/*analysis
Mass Spectrometry
Oligosaccharides/analysis
*Plasminogen
Abstract: The preceding two manuscripts in this issue (Hayes, M. L., and Castellino, F. J. (1979) J. Biol. Chem. 254, 8768-8771, 8772-8776) describe the isolation and characterization of glycopeptides from human plasminogen affinity chromatography variants 1 and 2. Plasminogen variant 1 contains an asparagine288-based branched carbohydrate structure, which has been established in the immediately preceding manuscript. This structure is absent in variant 2. Plasminogen variants 1 and 2 contain a threonine-based glycoconjugate. This latter structure has been established by combination of methylation data, glycosidase digestions, periodate oxidations, and Smith degradations of the beta-eliminated reduced oligosaccharides. One glycopeptide unit, isolated from both plasminogen variants 1 (1D) and 2 (2D) possessed the following structure: Sia alpha 2 yields 3Gal beta 1 yields 3GalNAc-Thr. The threonine was found to be residue 345 in the Glu-plasminogen sequence. Another glycopeptide unit was also found to be present, in lower yields in both variants 1 (1E) and 2 (2E). The structure of this unit was: Sia alpha 2 yields 3Gal beta1 yields 3GalNAc-Thr. : formula: (see text), alpha 2,6 Sia. Again Thr 345 was the glycosylated amino acid. The amino acid sequence around the glycosylated threonine was found to be NH2-Ala.Pro.Thr(CHO).Ala.Pro.Pro.Glu.
Notes: Hayes, M L
Castellino, F J
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1979 Sep 25;254(18):8777-80.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 812
Author: He, Xl, Chow, Dc, Martick, M. M. and Garcia, K. C.
Year: 2001
Title: Allosteric activation of a spring-loaded natriuretic peptide receptor dimer by hormone
Journal: Science
Volume: 293
Issue: 5535
Pages: 1657-62
Epub Date: 2001/09/05
Date: Aug 31
Short Title: Allosteric activation of a spring-loaded natriuretic peptide receptor dimer by hormone
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1062246
Accession Number: 11533490
Keywords: Allosteric Regulation
Amino Acid Sequence
Animals
Atrial Natriuretic Factor/metabolism
Binding Sites
Calorimetry
Cell Line
Chlorides/metabolism
Crystallization
Crystallography, X-Ray
Dimerization
Drosophila
Glycosylation
Guanylate Cyclase/*chemistry/*metabolism
Humans
Hydrogen Bonding
Ligands
Models, Molecular
Molecular Sequence Data
Natriuretic Peptide, Brain/metabolism
Natriuretic Peptide, C-Type/chemistry/*metabolism
Protein Conformation
Protein Folding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Atrial Natriuretic Factor/*chemistry/*metabolism
Thermodynamics
Abstract: Natriuretic peptides (NPs) are vasoactive cyclic-peptide hormones important in blood pressure regulation through interaction with natriuretic cell-surface receptors. We report the hormone-binding thermodynamics and crystal structures at 2.9 and 2.0 angstroms, respectively, of the extracellular domain of the unliganded human NP receptor (NPR-C) and its complex with CNP, a 22-amino acid NP. A single CNP molecule is bound in the interface of an NPR-C dimer, resulting in asymmetric interactions between the hormone and the symmetrically related receptors. Hormone binding induces a 20 angstrom closure between the membrane-proximal domains of the dimer. In each monomer, the opening of an interdomain cleft, which is tethered together by a linker peptide acting as a molecular spring, is likely a conserved allosteric trigger for intracellular signaling by the natriuretic receptor family.
Notes: He Xl
Chow Dc
Martick, M M
Garcia, K C
Journal Article
United States
Science. 2001 Aug 31;293(5535):1657-62. doi: 10.1126/science.1062246.
Author Address: Departments of Microbiology and Immunology and Structural Biology, Stanford University School of Medicine, Fairchild D319, 299 Campus Drive, Stanford, CA 93405-5124, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 878
Author: He, X. L., Bazan, J. F., McDermott, G., Park, J. B., Wang, K., Tessier-Lavigne, M., He, Z. and Garcia, K. C.
Year: 2003
Title: Structure of the Nogo receptor ectodomain: a recognition module implicated in myelin inhibition
Journal: Neuron
Volume: 38
Issue: 2
Pages: 177-85
Epub Date: 2003/04/30
Date: Apr 24
Short Title: Structure of the Nogo receptor ectodomain: a recognition module implicated in myelin inhibition
Alternate Journal: Neuron
ISSN: 0896-6273 (Print)
0896-6273
DOI: 10.1016/s0896-6273(03)00232-0
Accession Number: 12718853
Keywords: Binding Sites
Conserved Sequence
Crystallography, X-Ray
GPI-Linked Proteins
Humans
Ligands
Models, Molecular
Molecular Sequence Data
Myelin Proteins/*chemistry/metabolism
Myelin Sheath/*physiology
Neurites/physiology
Nogo Receptor 1
Protein Structure, Tertiary/physiology
Receptor, Nerve Growth Factor
Receptors, Cell Surface/*chemistry/metabolism
Receptors, Nerve Growth Factor/metabolism
Repetitive Sequences, Amino Acid/physiology
Sequence Homology, Amino Acid
Abstract: Failure of axon regeneration in the adult mammalian central nervous system (CNS) is at least partly due to inhibitory molecules associated with myelin. Recent studies suggest that an axon surface protein, the Nogo receptor (NgR), may play a role in this process through an unprecedented degree of crossreactivity with myelin-associated inhibitory ligands. Here, we report the 1.5 A crystal structure and functional characterization of a soluble extracellular domain of the human Nogo receptor. Nogo receptor adopts a leucine-rich repeat (LRR) module whose concave exterior surface contains a broad region of evolutionarily conserved patches of aromatic residues, possibly suggestive of degenerate ligand binding sites. A deep cleft at the C-terminal base of the LRR may play a role in NgR association with the p75 coreceptor. These results now provide a detailed framework for focused structure-function studies aimed at assessing the physiological relevance of NgR-mediated protein-protein interactions to axon regeneration inhibition.
Notes: He, Xiaolin L
Bazan, J Fernando
McDermott, Gerry
Park, Jong Bae
Wang, Kevin
Tessier-Lavigne, Marc
He, Zhigang
Garcia, K Christopher
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Neuron. 2003 Apr 24;38(2):177-85. doi: 10.1016/s0896-6273(03)00232-0.
Author Address: Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1061
Author: He, X. L., Dukkipati, A. and Garcia, K. C.
Year: 2006
Title: Structural determinants of natriuretic peptide receptor specificity and degeneracy
Journal: J Mol Biol
Volume: 361
Issue: 4
Pages: 698-714
Epub Date: 2006/07/28
Date: Aug 25
Short Title: Structural determinants of natriuretic peptide receptor specificity and degeneracy
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2006.06.060
Accession Number: 16870210
Keywords: Amino Acid Sequence
Animals
Atrial Natriuretic Factor/chemistry/*metabolism
Binding Sites
Crystallography, X-Ray
Dimerization
Drosophila melanogaster
Humans
Hydrophobic and Hydrophilic Interactions
Ligands
Models, Molecular
Molecular Sequence Data
Natriuretic Peptide, Brain/chemistry/*metabolism
Natriuretic Peptide, C-Type/chemistry/*metabolism
Phenylalanine/chemistry
Protein Conformation
Receptors, Atrial Natriuretic Factor/*chemistry/genetics/*metabolism
Sequence Alignment
Abstract: Cardiovascular homeostasis and blood pressure regulation are reliant, in part, on interactions between natriuretic peptide (NP) hormones and natriuretic peptide receptors (NPR). The C-type NPR (NPR-C) is responsible for clearance of NP hormones from the circulation, and displays a cross-reactivity for all NP hormones (ANP, BNP, and CNP), in contrast to other NPRs, which are more restricted in their specificity. In order to elucidate the structural determinants for the binding specificity and cross-reactivity of NPR-C with NP hormones, we have determined the crystal structures of the complexes of NPR-C with atrial natriuretic peptide (ANP), and with brain natriuretic peptide (BNP). A structural comparison of these complexes, with the previous structure of the NPR-C/CNP complex, reveals that NPR-C uses a conformationally inflexible surface to bind three different, highly flexible, NP ligands. The complex structures support a mechanism of rigid promiscuity rather than conformational plasticity by the receptor. While ANP and BNP appear to adopt similar receptor-bound conformations, the CNP structure diverges, yet shares sets of common receptor contacts with the other ligands. The degenerate versus selective hormone recognition properties of different NPRs appears to derive largely from two cavities on the receptor surfaces, pocket I and pocket II, that serve as anchoring sites for hormone side-chains and modulate receptor selectivity.
Notes: He, Xiao-lin
Dukkipati, Abhiram
Garcia, K Christopher
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2006 Aug 25;361(4):698-714. doi: 10.1016/j.jmb.2006.06.060. Epub 2006 Jul 10.
Author Address: Department of Microbiology & Immunology, Howard Hughes Medical Institute, Stanford University School of Medicine, Fairchild D319, 299 Campus Drive, Stanford, CA 94305-5124, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1776
Author: Heard, M. E., Besio, R., Weis, M., Rai, J., Hudson, D. M., Dimori, M., Zimmerman, S. M., Kamykowski, J. A., Hogue, W. R., Swain, F. L., Burdine, M. S., Mackintosh, S. G., Tackett, A. J., Suva, L. J., Eyre, D. R. and Morello, R.
Year: 2016
Title: Sc65-Null Mice Provide Evidence for a Novel Endoplasmic Reticulum Complex Regulating Collagen Lysyl Hydroxylation
Journal: PLoS Genet
Volume: 12
Issue: 4
Pages: e1006002
Epub Date: 2016/04/28
Date: Apr
Short Title: Sc65-Null Mice Provide Evidence for a Novel Endoplasmic Reticulum Complex Regulating Collagen Lysyl Hydroxylation
Alternate Journal: PLoS genetics
ISSN: 1553-7390 (Print)
1553-7390
DOI: 10.1371/journal.pgen.1006002
PMCID: PMC4847768
Accession Number: 27119146
Keywords: Animals
Autoantigens/genetics/*metabolism
Bone and Bones/physiology
Cell Line
Collagen/*biosynthesis/metabolism
Cyclophilins/metabolism
Endoplasmic Reticulum/*metabolism
Extracellular Matrix/metabolism
Hydroxylation/genetics
Mice
Mice, Inbred C57BL
Mice, Knockout
Osteogenesis Imperfecta/genetics/pathology
Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase/genetics/*metabolism
Procollagen-Proline Dioxygenase/*metabolism
Protein Processing, Post-Translational/*physiology
Abstract: Collagen is a major component of the extracellular matrix and its integrity is essential for connective tissue and organ function. The importance of proteins involved in intracellular collagen post-translational modification, folding and transport was recently highlighted from studies on recessive forms of osteogenesis imperfecta (OI). Here we describe the critical role of SC65 (Synaptonemal Complex 65, P3H4), a leprecan-family member, as part of an endoplasmic reticulum (ER) complex with prolyl 3-hydroxylase 3. This complex affects the activity of lysyl-hydroxylase 1 potentially through interactions with the enzyme and/or cyclophilin B. Loss of Sc65 in the mouse results in instability of this complex, altered collagen lysine hydroxylation and cross-linking leading to connective tissue defects that include low bone mass and skin fragility. This is the first indication of a prolyl-hydroxylase complex in the ER controlling lysyl-hydroxylase activity during collagen synthesis.
Notes: 1553-7404
Heard, Melissa E
Besio, Roberta
Orcid: 0000-0003-1430-2934
Weis, MaryAnn
Rai, Jyoti
Hudson, David M
Dimori, Milena
Zimmerman, Sarah M
Kamykowski, Jeffrey A
Orcid: 0000-0003-4738-3814
Hogue, William R
Swain, Frances L
Burdine, Marie S
Mackintosh, Samuel G
Tackett, Alan J
Suva, Larry J
Eyre, David R
Morello, Roy
Orcid: 0000-0003-1604-4879
AR060823/AR/NIAMS NIH HHS/United States
P30GM103450/GM/NIGMS NIH HHS/United States
HD070394/HD/NICHD NIH HHS/United States
UL1TR000039/TR/NCATS NIH HHS/United States
R01 AR060823/AR/NIAMS NIH HHS/United States
R01 AR036794/AR/NIAMS NIH HHS/United States
R33CA173264/CA/NCI NIH HHS/United States
AR036794/AR/NIAMS NIH HHS/United States
P20GM103429/GM/NIGMS NIH HHS/United States
P30 GM103450/GM/NIGMS NIH HHS/United States
R33 CA173264/CA/NCI NIH HHS/United States
P01 HD070394/HD/NICHD NIH HHS/United States
UL1 TR000039/TR/NCATS NIH HHS/United States
AR037318/AR/NIAMS NIH HHS/United States
R01GM106024/GM/NIGMS NIH HHS/United States
R01 GM106024/GM/NIGMS NIH HHS/United States
R01 AR037318/AR/NIAMS NIH HHS/United States
R37 AR036794/AR/NIAMS NIH HHS/United States
R37 AR037318/AR/NIAMS NIH HHS/United States
P20 GM103429/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
PLoS Genet. 2016 Apr 27;12(4):e1006002. doi: 10.1371/journal.pgen.1006002. eCollection 2016 Apr.
Author Address: Department of Physiology & Biophysics, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.
Department of Orthopaedics and Sports Medicine, University of Washington, Seattle, Washington, United States of America.
Department of Orthopaedic Surgery, Center for Orthopaedic Research, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.
Department of Biochemistry & Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.
Division of Genetics, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 762
Author: Heise, C. T., Nicholls, J. R., Leamy, C. E. and Wallis, R.
Year: 2000
Title: Impaired secretion of rat mannose-binding protein resulting from mutations in the collagen-like domain
Journal: J Immunol
Volume: 165
Issue: 3
Pages: 1403-9
Epub Date: 2000/07/21
Date: Aug 1
Short Title: Impaired secretion of rat mannose-binding protein resulting from mutations in the collagen-like domain
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
DOI: 10.4049/jimmunol.165.3.1403
Accession Number: 10903744
Keywords: Amino Acid Substitution/genetics
Animals
Arginine/genetics
Aspartic Acid/genetics
CHO Cells
Carrier Proteins/*genetics/*metabolism
Collagen/genetics/metabolism
Collectins
Complement Fixation Tests
Cricetinae
Cysteine/genetics
Endoplasmic Reticulum/metabolism
Glutamic Acid/genetics
Glycine/genetics
Humans
Lectins/genetics/metabolism
Lysine/metabolism
Mannans/metabolism
Mutagenesis, Site-Directed
Protein Processing, Post-Translational
Protein Structure, Tertiary/genetics
Proteins/*genetics/metabolism
Rats
Time Factors
Transfection
Abstract: Serum mannose-binding protein (MBP) or mannose-binding lectin initiates the lectin branch of the innate immune response by binding to the surface of potentially pathogenic microorganisms and initiating complement fixation through an N-terminal collagen-like domain. Mutations in this region of human MBP are associated with immunodeficiency resulting from a reduction in the ability of the mutant MBPs to fix complement as well as from reduced serum concentrations. Inefficient secretion of the mutant proteins, which is one possible cause of the reduced serum levels, has been investigated using a mammalian expression system in which each of the naturally occurring human mutations has been recreated in rat serum MBP. The mutations Gly25-->Asp and Gly28-->Glu disrupt the disulfide-bonding arrangement of the protein and cause at least a 5-fold increase in the half-time of secretion of MBP compared with wild-type rat serum MBP. A similar phenotype, including a 3-fold increase in the half-time of secretion, disruption of the disulfide bonding arrangement, and inefficient complement fixation, is observed when nearby glucosylgalactosyl hydroxylysine residues at positions 27 and 30 are replaced with arginine residues. The results suggest that defective secretion resulting from structural changes in the collagen-like domain is likely to be a contributory factor for MBP immunodeficiency.
Notes: Heise, C T
Nicholls, J R
Leamy, C E
Wallis, R
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 2000 Aug 1;165(3):1403-9. doi: 10.4049/jimmunol.165.3.1403.
Author Address: Glycobiology Institute, Department of Biochemistry, University of Oxford, Oxford, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 888
Author: Henrich, S., Cameron, A., Bourenkov, G. P., Kiefersauer, R., Huber, R., Lindberg, I., Bode, W. and Than, M. E.
Year: 2003
Title: The crystal structure of the proprotein processing proteinase furin explains its stringent specificity
Journal: Nat Struct Biol
Volume: 10
Issue: 7
Pages: 520-6
Epub Date: 2003/06/10
Date: Jul
Short Title: The crystal structure of the proprotein processing proteinase furin explains its stringent specificity
Alternate Journal: Nature structural biology
ISSN: 1072-8368 (Print)
1072-8368
DOI: 10.1038/nsb941
Accession Number: 12794637
Keywords: Amino Acid Sequence
Animals
Catalytic Domain
Crystallography, X-Ray
Enzyme Inhibitors/metabolism
Furin
Mice
Models, Molecular
Molecular Sequence Data
Protein Conformation
Sequence Homology, Amino Acid
Substrate Specificity
Subtilisins/antagonists & inhibitors/chemistry/*metabolism
Abstract: In eukaryotes, many essential secreted proteins and peptide hormones are excised from larger precursors by members of a class of calcium-dependent endoproteinases, the prohormone-proprotein convertases (PCs). Furin, the best-characterized member of the mammalian PC family, has essential functions in embryogenesis and homeostasis but is also implicated in various pathologies such as tumor metastasis, neurodegeneration and various bacterial and viral diseases caused by such pathogens as anthrax and pathogenic Ebola virus strains. Furin cleaves protein precursors with narrow specificity following basic Arg-Xaa-Lys/Arg-Arg-like motifs. The 2.6 A crystal structure of the decanoyl-Arg-Val-Lys-Arg-chloromethylketone (dec-RVKR-cmk)-inhibited mouse furin ectodomain, the first PC structure, reveals an eight-stranded jelly-roll P domain associated with the catalytic domain. Contoured surface loops shape the active site by cleft, thus explaining furin's stringent requirement for arginine at P1 and P4, and lysine at P2 sites by highly charge-complementary pockets. The structure also explains furin's preference for basic residues at P3, P5 and P6 sites. This structure will aid in the rational design of antiviral and antibacterial drugs.
Notes: Henrich, Stefan
Cameron, Angus
Bourenkov, Gleb P
Kiefersauer, Reiner
Huber, Robert
Lindberg, Iris
Bode, Wolfram
Than, Manuel E
R01 DA005084/DA/NIDA NIH HHS/United States
R56 DA005084/DA/NIDA NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Nat Struct Biol. 2003 Jul;10(7):520-6. doi: 10.1038/nsb941.
Author Address: Max-Planck-Institut für Biochemie, Abt. Strukturforschung, Am Klopferspitz 18A, 82152 Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 825
Author: Hepbildikler, S. T., Sandhoff, R., Kolzer, M., Proia, R. L. and Sandhoff, K.
Year: 2002
Title: Physiological substrates for human lysosomal beta -hexosaminidase S
Journal: J Biol Chem
Volume: 277
Issue: 4
Pages: 2562-72
Epub Date: 2001/11/15
Date: Jan 25
Short Title: Physiological substrates for human lysosomal beta -hexosaminidase S
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M105457200
Accession Number: 11707436
Keywords: Animals
Carbohydrate Sequence
Cell Line
Chondroitin Sulfates/chemistry
Chromatography, Thin Layer
Dermatan Sulfate/metabolism
G(M2) Activator Protein
Glycolipids
Hexosaminidase A
Hexosaminidase B
Humans
Hydrogen-Ion Concentration
Hydrolysis
Insecta
Kidney/metabolism
Kinetics
Lysophospholipids/chemistry
Lysosomes/*enzymology
Mice
Mice, Knockout
Models, Chemical
Molecular Sequence Data
Monoglycerides
Polysaccharides/metabolism
Protein Binding
Protein Processing, Post-Translational
Proteins/metabolism
Rats
Recombinant Proteins/chemistry/metabolism
Spectrometry, Mass, Electrospray Ionization
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Substrate Specificity
Time Factors
Water/metabolism
beta-N-Acetylhexosaminidases/*chemistry
Abstract: Human lysosomal beta-hexosaminidases remove terminal beta-glycosidically bound N-acetylhexosamine residues from a number of glycoconjugates. Three different isozymes composed of two noncovalently linked subunits alpha and beta exist: Hex A (alphabeta), Hex B (betabeta), and Hex S (alphaalpha). While the role of Hex A and B for the degradation of several anionic and neutral glycoconjugates has been well established, the physiological significance of labile Hex S has remained unclear. However, the striking accumulation of anionic oligosaccharides in double knockout mice totally deficient in hexosaminidase activity but not in mice expressing Hex S (Sango, K., McDonald, M. P., Crawley, J. N., Mack, M. L., Tifft, C.J., Skop, E., Starr, C. M., Hoffmann, A., Sandhoff, K., Suzuki, K., and Proia, R. L., (1996) Nat. Genet. 14, 348-352) prompted us to reinvestigate the substrate specificity of Hex S. To identify physiological substrates of Hex S, anionic and neutral oligosaccharides excreted in the urine of the double knockout mice were isolated and analyzed. Using ESI-MS/MS and glycosidase digestion the anionic glycans were identified as products of incomplete dermatan sulfate degradation whereas the neutral storage oligosaccharides were found to be fragments of N-glycan degradation. In vitro, recombinant Hex S was highly active on water-soluble and amphiphilic glycoconjugates including artificial substrates, sulfated GAG fragments, and the sulfated glycosphingolipid SM2. Hydrolysis of membrane-bound SM2 by the recombinant Hex S was synergistically stimulated by the GM2 activator protein and the lysosomal anionic phospholipid bis(monoacylglycero)phosphate.
Notes: Hepbildikler, Stefan T
Sandhoff, Roger
Kolzer, Melanie
Proia, Richard L
Sandhoff, Konrad
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2002 Jan 25;277(4):2562-72. doi: 10.1074/jbc.M105457200. Epub 2001 Nov 13.
Author Address: Kekulé-Institut für Organische Chemie und Biochemie, Universität Bonn, Gerhard-Domagk-Str. 1, 53121 Bonn, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 429
Author: Herkt, F., Parente, J. P., Leroy, Y., Fournet, B., Blanchard, D., Cartron, J. P., van Halbeek, H. and Vliegenthart, J. F.
Year: 1985
Title: Structure determination of oligosaccharides isolated from Cad erythrocyte membranes by permethylation analysis and 500-MHz 1H-NMR spectroscopy
Journal: Eur J Biochem
Volume: 146
Issue: 1
Pages: 125-9
Epub Date: 1985/01/02
Date: Jan 2
Short Title: Structure determination of oligosaccharides isolated from Cad erythrocyte membranes by permethylation analysis and 500-MHz 1H-NMR spectroscopy
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1985.tb08628.x
Accession Number: 3967649
Keywords: *Blood Group Antigens
Chemical Phenomena
Chemistry
Chromatography, High Pressure Liquid
Erythrocyte Membrane/*analysis
Glycophorins/isolation & purification
Humans
Magnetic Resonance Spectroscopy
Methylation
Oligosaccharides/blood/*isolation & purification
Abstract: Alkaline borohydride reductive cleavage (beta-elimination) of glycophorin A isolated from one individual of the rare blood group Cad, resulted in the release of six acidic oligosaccharide-alditols which were separated by high-performance liquid chromatography (HPLC) on an alkyl amine silicagel column. The structure of four of them has been determined by the application of methanolysis, methylation analysis and 1H-NMR spectroscopy at 500 MHz. The structures and relative amounts were as follows: oligosaccharide 1: NeuAc(alpha 2-3)Gal(beta 1-3)GalNAc-ol (3.5%); oligosaccharide 3: GalNAc(beta 1-4)[NeuAc(alpha 2-3)]Gal(beta 1-3)GalNAc-ol (10.5%); oligosaccharide 5: NeuAc(alpha 2-3)Gal(beta 1-3)[NeuAc(alpha 2-6)]GalNAc-ol (10.4%); oligosaccharide 6: GalNAc(beta 1-4)[NeuAc(alpha 2-3)]Gal(beta 1-3)[NeuAc(alpha 2-6)]GalNAc-ol (71.2%). The two other oligosaccharides (2 and 4) were obtained in very low amount. The major pentasaccharide (oligosaccharide 6) carries the blood group Cad determinant and is a potent inhibitor of human anti-Sda antibody.
Notes: Herkt, F
Parente, J P
Leroy, Y
Fournet, B
Blanchard, D
Cartron, J P
van Halbeek, H
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1985 Jan 2;146(1):125-9. doi: 10.1111/j.1432-1033.1985.tb08628.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 592
Author: Hermans, M. M., Wisselaar, H. A., Kroos, M. A., Oostra, B. A. and Reuser, A. J.
Year: 1993
Title: Human lysosomal alpha-glucosidase: functional characterization of the glycosylation sites
Journal: Biochem J
Volume: 289 ( Pt 3)
Issue: Pt 3
Pages: 681-6
Epub Date: 1993/02/01
Date: Feb 1
Short Title: Human lysosomal alpha-glucosidase: functional characterization of the glycosylation sites
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2890681
PMCID: PMC1132229
Accession Number: 8435067
Keywords: Animals
Base Sequence
Biological Transport
Cell Compartmentation
Cells, Cultured
DNA Mutational Analysis
Glycosylation
Humans
Immunohistochemistry
Lysosomes/*enzymology
Molecular Sequence Data
Mutagenesis, Site-Directed
Phosphorylation
*Protein Processing, Post-Translational
Recombinant Proteins/metabolism
Transcription, Genetic
alpha-Glucosidases/genetics/isolation & purification/*metabolism
Abstract: N-linked glycosylation is one of the important events in the post-translational modification of human lysosomal alpha-glucosidase. Phosphorylation of mannose residues ensures efficient transport of the enzyme to the lysosomes via the mannose 6-phosphate receptor. The primary structure of lysosomal alpha-glucosidase, as deduced from the cDNA sequence, indicates that there are seven potential glycosylation sites. We have eliminated these sites individually by site-directed mutagenesis and thereby demonstrated that all seven sites are glycosylated. The sites at Asn-882 and Asn-925 were found to be located in a C-terminal propeptide which is cleaved off during maturation. Evidence is presented that at least two of the oligosaccharide side chains of human lysosomal alpha-glucosidase are phosphorylated. Elimination of six of the seven sites does not disturb enzyme synthesis or function. However, removal of the second glycosylation site at Asn-233 interferes dramatically with the formation of mature enzyme. The mutant precursor is synthesized normally and assembles in the endoplasmic reticulum, but immunoelectron microscopy reveals a deficiency of alpha-glucosidase in the Golgi complex and in the more distal compartments of the lysosomal transport pathway.
Notes: 1470-8728
Hermans, M M
Wisselaar, H A
Kroos, M A
Oostra, B A
Reuser, A J
Journal Article
Biochem J. 1993 Feb 1;289 ( Pt 3)(Pt 3):681-6. doi: 10.1042/bj2890681.
Author Address: Department of Cell Biology and Genetics, Erasmus University, Rotterdam, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 510
Author: Hermentin, P., Doenges, R., Witzel, R., Hokke, C. H., Vliegenthart, J. F., Kamerling, J. P., Conradt, H. S., Nimtz, M. and Brazel, D.
Year: 1994
Title: A strategy for the mapping of N-glycans by high-performance capillary electrophoresis
Journal: Anal Biochem
Volume: 221
Issue: 1
Pages: 29-41
Epub Date: 1994/08/15
Date: Aug 15
Short Title: A strategy for the mapping of N-glycans by high-performance capillary electrophoresis
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1006/abio.1994.1374
Accession Number: 7527189
Keywords: Electrophoresis/*methods
Erythropoietin/chemistry
Information Systems
Orosomucoid/chemistry
Polysaccharides/*chemistry
alpha-Fetoproteins/chemistry
Abstract: We have evaluated high-performance capillary electrophoresis (HPCE) with respect to its suitability for use in establishing a carbohydrate-mapping database that would enable a carbohydrate structural analysis by mere comparison of migration times. The suitability of HPCE for carbohydrate structural assignments was ascertained by validation experiments. The migration times of distinct N-glycans, prepared and measured on different days, were shown to be highly reproducible, with a coefficient of variation of usually less than 0.20%, requiring only femtomoles of N-glycan per injection for reliable measurements. By including mesityl oxide and sialic acid as internal standards and a triple-correction method, HPCE fulfills the analytical requirements with respect to accuracy, precision, reproducibility, and sensitivity. The N-glycan-mapping database was established using a newly developed and optimized buffer system containing 1,5-diaminopentane as an organic modifier. Approximately 80 different sialylated N-glycans of known structure, which have thus far been measured and characterized, have been entered into our Lotus 1-2-3 mapping database. The database for structural determinations was tested using the N-linked carbohydrates released from recombinant human urinary erythropoietin (baby hamster kidney) by PNGase F treatment and from bovine serum fetuin and alpha 1-acid glycoprotein by automated and manual (large-scale) hydrazinolysis, respectively. The efficiency of the database and of the triple-correction method was further confirmed by HPCE measurements performed in a different laboratory and by a different analyst who used the HPCE system of a different manufacturer.
Notes: Hermentin, P
Doenges, R
Witzel, R
Hokke, C H
Vliegenthart, J F
Kamerling, J P
Conradt, H S
Nimtz, M
Brazel, D
Journal Article
United States
Anal Biochem. 1994 Aug 15;221(1):29-41. doi: 10.1006/abio.1994.1374.
Author Address: Research Laboratories of Behringwerke AG, Marburg, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 876
Author: Hernandez-Guzman, F. G., Higashiyama, T., Pangborn, W., Osawa, Y. and Ghosh, D.
Year: 2003
Title: Structure of human estrone sulfatase suggests functional roles of membrane association
Journal: J Biol Chem
Volume: 278
Issue: 25
Pages: 22989-97
Epub Date: 2003/03/27
Date: Jun 20
Short Title: Structure of human estrone sulfatase suggests functional roles of membrane association
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M211497200
Accession Number: 12657638
Keywords: Amino Acid Sequence
Binding Sites
Catalytic Domain
Crystallography, X-Ray
Endoplasmic Reticulum/enzymology
Female
Humans
Intracellular Membranes/*enzymology
Microscopy, Electron
Microsomes/enzymology
Models, Molecular
Molecular Sequence Data
Placenta/*enzymology
Pregnancy
Protein Conformation
Protein Structure, Secondary
Sequence Alignment
Sequence Homology, Amino Acid
Sulfatases/*chemistry/isolation & purification/*metabolism/ultrastructure
Abstract: Estrone sulfatase (ES; 562 amino acids), one of the key enzymes responsible for maintaining high levels of estrogens in breast tumor cells, is associated with the membrane of the endoplasmic reticulum (ER). The structure of ES, purified from the microsomal fraction of human placentas, has been determined at 2.60-A resolution by x-ray crystallography. This structure shows a domain consisting of two antiparallel alpha-helices that protrude from the roughly spherical molecule, thereby giving the molecule a "mushroom-like" shape. These highly hydrophobic helices, each about 40 A long, are capable of traversing the membrane, thus presumably anchoring the functional domain on the membrane surface facing the ER lumen. The location of the transmembrane domain is such that the opening to the active site, buried deep in a cavity of the "gill" of the "mushroom," rests near the membrane surface, thereby suggesting a role of the lipid bilayer in catalysis. This simple architecture could be a prototype utilized by the ER membrane in dictating the form and the function of ER-resident enzymes.
Notes: Hernandez-Guzman, Francisco G
Higashiyama, Tadayoshi
Pangborn, Walter
Osawa, Yoshio
Ghosh, Debashis
GM62794/GM/NIGMS NIH HHS/United States
RR-01646/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2003 Jun 20;278(25):22989-97. doi: 10.1074/jbc.M211497200. Epub 2003 Mar 25.
Author Address: Department of Structural Biology, Hauptman-Woodward Medical Research Institute, Buffalo, New York 14203, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 883
Author: Herr, A. B., Ballister, E. R. and Bjorkman, P. J.
Year: 2003
Title: Insights into IgA-mediated immune responses from the crystal structures of human FcalphaRI and its complex with IgA1-Fc
Journal: Nature
Volume: 423
Issue: 6940
Pages: 614-20
Epub Date: 2003/05/28
Date: Jun 5
Short Title: Insights into IgA-mediated immune responses from the crystal structures of human FcalphaRI and its complex with IgA1-Fc
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature01685
Accession Number: 12768205
Keywords: Amino Acid Sequence
Antigens, CD/*chemistry/*immunology
Crystallography, X-Ray
Humans
Immunoglobulin A/*chemistry/*immunology
Immunoglobulin E/chemistry
Immunoglobulin Fragments/chemistry/immunology
Immunoglobulin G/chemistry
Molecular Sequence Data
Protein Structure, Quaternary
Receptors, Fc/*chemistry/*immunology
Signal Transduction
Abstract: Immunoglobulin-alpha (IgA)-bound antigens induce immune effector responses by activating the IgA-specific receptor FcalphaRI (CD89) on immune cells. Here we present crystal structures of human FcalphaRI alone and in a complex with the Fc region of IgA1 (Fcalpha). FcalphaRI has two immunoglobulin-like domains that are oriented at approximately right angles to each other. Fcalpha resembles the Fcs of immunoglobulins IgG and IgE, but has differently located interchain disulphide bonds and external rather than interdomain N-linked carbohydrates. Unlike 1:1 FcgammaRIII:IgG and Fc epsilon RI:IgE complexes, two FcalphaRI molecules bind each Fcalpha dimer, one at each Calpha2-Calpha3 junction. The FcalphaRI-binding site on IgA1 overlaps the reported polymeric immunoglobulin receptor (pIgR)-binding site, which might explain why secretory IgA cannot initiate phagocytosis or bind to FcalphaRI-expressing cells in the absence of an integrin co-receptor.
Notes: Herr, Andrew B
Ballister, Edward R
Bjorkman, Pamela J
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2003 Jun 5;423(6940):614-20. doi: 10.1038/nature01685. Epub 2003 May 21.
Author Address: Division of Biology 114-96, California Institute of Technology, Pasadena, California 91125, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1855
Author: Hieronimus, B., Pfohl, J., Busch, C. and Graeve, L.
Year: 2017
Title: Expression and Characterization of Membrane-Type 4 Matrix Metalloproteinase (MT4-MMP) and its Different Forms in Melanoma
Journal: Cell Physiol Biochem
Volume: 42
Issue: 1
Pages: 198-210
Epub Date: 2017/05/23
Short Title: Expression and Characterization of Membrane-Type 4 Matrix Metalloproteinase (MT4-MMP) and its Different Forms in Melanoma
Alternate Journal: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology
ISSN: 1015-8987
DOI: 10.1159/000477311
Accession Number: 28531887
Keywords: Cell Line, Tumor
Cell Membrane/metabolism
Endoplasmic Reticulum Stress
Glycosylation
Golgi Apparatus/metabolism
Humans
Matrix Metalloproteinases, Membrane-Associated/genetics/*metabolism
Melanoma/metabolism/*pathology
Mutagenesis, Site-Directed
Protein Isoforms/genetics/metabolism
RNA, Messenger/metabolism
Skin Neoplasms/metabolism/*pathology
Glycosylphosphatidylinositol (GPI) - linked Proteins
N-glycosylation
Plasma membrane
Protein processing
RNA-protein relationship
Subcellular fractionation
Abstract: BACKGROUND/AIMS: Membrane-type matrix metalloproteinases (MT-MMPs) are expressed on the cell surface and hydrolyze extracellular matrix components and signaling molecules by which they influence cancer cell migration and metastasis. Two of the six known MT-MMPs are anchored to the plasma membrane via a GPI anchor, one of which is MT4-MMP. Only little is known about MT4-MMP expression, synthesis, regulation and degradation. METHODS: We analyzed several human cancer cell lines as well as tissue homogenates using Western blotting and quantitative PCR for the expression of MT4-MMP. Organelles of SK-Mel-28 cells were separated using continuous Iodixanol gradients. Glycosylation of the SK-Mel-28 protein was studied via glucosidases and site directed mutagenesis of the MT4-MMP cDNA prior to transfection. RESULTS: We found the MT4-MMP highly expressed in human melanoma cell lines as well as skin and melanoma tissue samples. Three forms of MT4-MMP with molecular masses of 45 kDa, 58 kDa and 69 kDa were detected. Further, we demonstrate that the 58 kDa form is the mature protein in the cell membrane, while the 69 kDa form is its precursor found in intracellular compartments. The 69 kDa forms are processed by furin cleavage in the Golgi apparatus. Moreover, we identified Asn318 as the single N-glycosylation site of MT4-MMP. CONCLUSION: We demonstrate the novel expression of MT4-MMP in melanocytic tissues and propose a precursor/product-relationship of the different forms of MT4-MMP in melanoma cells.
Notes: 1421-9778
Hieronimus, Bettina
Pfohl, Julian
Busch, Christian
Graeve, Lutz
Journal Article
Germany
Cell Physiol Biochem. 2017;42(1):198-210. doi: 10.1159/000477311. Epub 2017 May 25.
Author Address: Institute of Biological Chemistry and Nutrition, University of Hohenheim, Stuttgart, Germany.
Section of Dermatolo-Oncology, Department of Dermatology and Allergology, University of Tuebingen, Tuebingen, Germany.
Dermateam, Winterthur, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 259
Author: Higashiyama, S., Lau, K., Besner, G. E., Abraham, J. A. and Klagsbrun, M.
Year: 1992
Title: Structure of heparin-binding EGF-like growth factor. Multiple forms, primary structure, and glycosylation of the mature protein
Journal: J Biol Chem
Volume: 267
Issue: 9
Pages: 6205-12
Epub Date: 1992/03/25
Date: Mar 25
Short Title: Structure of heparin-binding EGF-like growth factor. Multiple forms, primary structure, and glycosylation of the mature protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1556128
Keywords: 3T3 Cells
Amino Acid Sequence
Animals
Antibodies
Biological Assay
Cell Line
Chromatography, Affinity
Chromatography, High Pressure Liquid
DNA Replication/drug effects
Electrophoresis, Polyacrylamide Gel
Epidermal Growth Factor/drug effects/*isolation &
purification/*metabolism/pharmacology
Glycosylation
Growth Substances/*isolation & purification/pharmacology
Heparin/*isolation & purification/pharmacology
Heparin-binding EGF-like Growth Factor
Humans
Intercellular Signaling Peptides and Proteins
Isoelectric Focusing
Mice
Molecular Sequence Data
Molecular Weight
Peptide Fragments/isolation & purification
Peptide Mapping
Platelet-Derived Growth Factor/analysis/pharmacology
Sequence Homology, Nucleic Acid
Abstract: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a newly described member of the epidermal growth factor (EGF) family that is mitogenic for BALB/c 3T3 cells, inhibits the binding of 125I-EGF to its receptor, and triggers autophosphorylation of the EGF receptor. HB-EGF was purified from the conditioned medium of U-937 cells using cation exchange, copper affinity, heparin affinity, and two rounds of C4 reversed phase liquid chromatography. The elution profile of the first round of C4 column chromatography contained four growth factor activity peaks with similar specific biological activities. N-terminal and tryptic fragment microsequencing demonstrated that these peaks contained different structural forms of the HB-EGF protein. Some of the differences in the various forms of HB-EGF were found to be due to N-terminal heterogeneity. Microsequencing of tryptic fragments indicated that the mature HB-EGF polypeptide can contain at least 86 of the 208 amino acids predicted by nucleotide sequence to be the HB-EGF precursor molecule. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis indicated that the various forms of HB-EGF have apparent molecular masses of 19-23 kDa. Further analysis of the most predominant form of HB-EGF found in U-937 cell conditioned medium indicated that it has a pI of 7.2-7.8 and is O-glycosylated.
Notes: Higashiyama, S
Lau, K
Besner, G E
Abraham, J A
Klagsbrun, M
CA37392/CA/NCI NIH HHS/United States
CA45548/CA/NCI NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Mar 25;267(9):6205-12.
Author Address: Department of Surgical Research, Children's Hospital, Boston, Massachusetts.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1188
Author: Hilvo, M., Baranauskiene, L., Salzano, A. M., Scaloni, A., Matulis, D., Innocenti, A., Scozzafava, A., Monti, S. M., Di Fiore, A., De Simone, G., Lindfors, M., Jänis, J., Valjakka, J., Pastoreková, S., Pastorek, J., Kulomaa, M. S., Nordlund, H. R., Supuran, C. T. and Parkkila, S.
Year: 2008
Title: Biochemical characterization of CA IX, one of the most active carbonic anhydrase isozymes
Journal: J Biol Chem
Volume: 283
Issue: 41
Pages: 27799-27809
Epub Date: 2008/08/16
Date: Oct 10
Short Title: Biochemical characterization of CA IX, one of the most active carbonic anhydrase isozymes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M800938200
Accession Number: 18703501
Keywords: Animals
Antigens, Neoplasm/*chemistry/genetics/metabolism
Baculoviridae
Carbonic Anhydrase IX
Carbonic Anhydrases/*chemistry/genetics/metabolism
Catalysis
Cell Adhesion/genetics
Cell Line
Cell Proliferation
Dimerization
Disulfides/chemistry/metabolism
Glycosylation
Humans
Hydrogen-Ion Concentration
Isoenzymes/chemistry/genetics/metabolism
Neoplasms/enzymology/genetics
Polysaccharides/chemistry/genetics/metabolism
Protein Structure, Tertiary/genetics
Recombinant Proteins/chemistry/genetics/metabolism
Spodoptera
Abstract: Carbonic anhydrase IX (CA IX) is an exceptional member of the CA protein family; in addition to its classical role in pH regulation, it has also been proposed to participate in cell proliferation, cell adhesion, and tumorigenic processes. To characterize the biochemical properties of this membrane protein, two soluble recombinant forms were produced using the baculovirus-insect cell expression system. The recombinant proteins consisted of either the CA IX catalytic domain only (CA form) or the extracellular domain, which included both the proteoglycan and catalytic domains (PG + CA form). The produced proteins lacked the small transmembrane and intracytoplasmic regions of CA IX. Stopped-flow spectrophotometry experiments on both proteins demonstrated that in the excess of certain metal ions the PG + CA form exhibited the highest catalytic activity ever measured for any CA isozyme. Investigations on the oligomerization and stability of the enzymes revealed that both recombinant proteins form dimers that are stabilized by intermolecular disulfide bond(s). Mass spectrometry experiments showed that CA IX contains an intramolecular disulfide bridge (Cys(119)-Cys(299)) and a unique N-linked glycosylation site (Asn(309)) that bears high mannose-type glycan structures. Parallel experiments on a recombinant protein obtained by a mammalian cell expression system demonstrated the occurrence of an additional O-linked glycosylation site (Thr(78)) and characterized the nature of the oligosaccharide structures. This study provides novel information on the biochemical properties of CA IX and may help characterize the various cellular and pathophysiological processes in which this unique enzyme is involved.
Notes: Hilvo, Mika
Baranauskiene, Lina
Salzano, Anna Maria
Scaloni, Andrea
Matulis, Daumantas
Innocenti, Alessio
Scozzafava, Andrea
Monti, Simona Maria
Di Fiore, Anna
De Simone, Giuseppina
Lindfors, Mikaela
Jänis, Janne
Valjakka, Jarkko
Pastoreková, Silvia
Pastorek, Jaromir
Kulomaa, Markku S
Nordlund, Henri R
Supuran, Claudiu T
Parkkila, Seppo
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2008 Oct 10;283(41):27799-27809. doi: 10.1074/jbc.M800938200. Epub 2008 Aug 13.
Author Address: Institute of Medical Technology, FI-33014 Tampere, Finland. Electronic address: Mika.Hilvo@uta.fi.
Laboratory of Biothermodynamics and Drug Design, Institute of Biotechnology, LT-02241 Vilnius, Lithuania.
Proteomics and Mass Spectrometry Laboratory, ISPAAM, National Research Council, 80147 Naples, Italy.
Bioinorganic Chemistry Laboratory, University of Florence, 50019 Sesto Fiorentino (Florence), Italy.
Institute of Biostructures and Bioimages, National Research Council, 80134 Naples, Italy.
Institute of Medical Technology, FI-33014 Tampere, Finland.
Department of Chemistry, University of Joensuu, FI-80101 Joensuu, Finland.
Centre of Molecular Medicine, Institute of Virology, Slovak Academy of Sciences, 84505 Bratislava, Slovak Republic.
Institute of Medical Technology, FI-33014 Tampere, Finland; School of Medicine, University of Tampere and Tampere University Hospital, FI-33014 Tampere, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 937
Author: Himanen, J. P., Chumley, M. J., Lackmann, M., Li, C., Barton, W. A., Jeffrey, P. D., Vearing, C., Geleick, D., Feldheim, D. A., Boyd, A. W., Henkemeyer, M. and Nikolov, D. B.
Year: 2004
Title: Repelling class discrimination: ephrin-A5 binds to and activates EphB2 receptor signaling
Journal: Nat Neurosci
Volume: 7
Issue: 5
Pages: 501-9
Epub Date: 2004/04/27
Date: May
Short Title: Repelling class discrimination: ephrin-A5 binds to and activates EphB2 receptor signaling
Alternate Journal: Nature neuroscience
ISSN: 1097-6256 (Print)
1097-6256
DOI: 10.1038/nn1237
Accession Number: 15107857
Keywords: Alkaline Phosphatase/metabolism
Animals
Animals, Newborn
Cell Line
Chromatography, Gel/methods
Chromatography, Ion Exchange/methods
Cricetinae
Cricetulus
Crystallography/methods
Electrophoresis/methods
Ephrin-A5/chemistry/*metabolism
Ephrin-B2/*metabolism
Fluorescent Antibody Technique/methods
Green Fluorescent Proteins
Humans
Infections
Luminescent Proteins/metabolism
Mice
Neurites/physiology
Neuroblastoma
Phosphorylation
Protein Binding/physiology
Receptor, EphA3/metabolism
Receptor, EphB2/chemistry/*metabolism
Signal Transduction/*physiology
Sindbis Virus
Spectrometry, Fluorescence/methods
Surface Plasmon Resonance/methods
Time Factors
Transfection/methods
Video Recording
Abstract: The interactions between Eph receptor tyrosine kinases and their ephrin ligands regulate cell migration and axon pathfinding. The EphA receptors are generally thought to become activated by ephrin-A ligands, whereas the EphB receptors interact with ephrin-B ligands. Here we show that two of the most widely studied of these molecules, EphB2 and ephrin-A5, which have never been described to interact with each other, do in fact bind one another with high affinity. Exposure of EphB2-expressing cells to ephrin-A5 leads to receptor clustering, autophosphorylation and initiation of downstream signaling. Ephrin-A5 induces EphB2-mediated growth cone collapse and neurite retraction in a model system. We further show, using X-ray crystallography, that the ephrin-A5-EphB2 complex is a heterodimer and is architecturally distinct from the tetrameric EphB2-ephrin-B2 structure. The structural data reveal the molecular basis for EphB2-ephrin-A5 signaling and provide a framework for understanding the complexities of functional interactions and crosstalk between A- and B-subclass Eph receptors and ephrins.
Notes: Himanen, Juha-Pekka
Chumley, Michael J
Lackmann, Martin
Li, Chen
Barton, William A
Jeffrey, Phillip D
Vearing, Christopher
Geleick, Detlef
Feldheim, David A
Boyd, Andrew W
Henkemeyer, Mark
Nikolov, Dimitar B
R01 MH066332/MH/NIMH NIH HHS/United States
R01-MH66332/MH/NIMH NIH HHS/United States
R01-NS38486/NS/NINDS NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Nat Neurosci. 2004 May;7(5):501-9. doi: 10.1038/nn1237. Epub 2004 Apr 25.
Author Address: Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1315
Author: Himanen, J. P., Yermekbayeva, L., Janes, P. W., Walker, J. R., Xu, K., Atapattu, L., Rajashankar, K. R., Mensinga, A., Lackmann, M., Nikolov, D. B. and Dhe-Paganon, S.
Year: 2010
Title: Architecture of Eph receptor clusters
Journal: Proc Natl Acad Sci U S A
Volume: 107
Issue: 24
Pages: 10860-5
Epub Date: 2010/05/28
Date: Jun 15
Short Title: Architecture of Eph receptor clusters
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1004148107
PMCID: PMC2890748
Accession Number: 20505120
Keywords: Amino Acid Sequence
Binding Sites
Cell Line
Crystallography, X-Ray
Ephrin-A1/chemistry/genetics/metabolism
Ephrin-A5/chemistry/genetics/metabolism
Humans
Models, Molecular
Molecular Sequence Data
Multiprotein Complexes
Protein Structure, Secondary
Protein Structure, Tertiary
Receptor, EphA1/*chemistry/genetics/metabolism
Receptor, EphA2/chemistry/genetics/metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Signal Transduction
Abstract: Eph receptor tyrosine kinases and their ephrin ligands regulate cell navigation during normal and oncogenic development. Signaling of Ephs is initiated in a multistep process leading to the assembly of higher-order signaling clusters that set off bidirectional signaling in interacting cells. However, the structural and mechanistic details of this assembly remained undefined. Here we present high-resolution structures of the complete EphA2 ectodomain and complexes with ephrin-A1 and A5 as the base unit of an Eph cluster. The structures reveal an elongated architecture with novel Eph/Eph interactions, both within and outside of the Eph ligand-binding domain, that suggest the molecular mechanism underlying Eph/ephrin clustering. Structure-function analysis, by using site-directed mutagenesis and cell-based signaling assays, confirms the importance of the identified oligomerization interfaces for Eph clustering.
Notes: 1091-6490
Himanen, Juha P
Yermekbayeva, Laila
Janes, Peter W
Walker, John R
Xu, Kai
Atapattu, Lakmali
Rajashankar, Kanagalaghatta R
Mensinga, Anneloes
Lackmann, Martin
Nikolov, Dimitar B
Dhe-Paganon, Sirano
P41 RR015301/RR/NCRR NIH HHS/United States
RR-15301/RR/NCRR NIH HHS/United States
R01 NS038486/NS/NINDS NIH HHS/United States
GM75886/GM/NIGMS NIH HHS/United States
R21 GM075886/GM/NIGMS NIH HHS/United States
NS38486/NS/NINDS NIH HHS/United States
Wellcome Trust/United Kingdom
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2010 Jun 15;107(24):10860-5. doi: 10.1073/pnas.1004148107. Epub 2010 May 26.
Author Address: Structural Biology Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 648
Author: Hipfner, D. R., Almquist, K. C., Leslie, E. M., Gerlach, J. H., Grant, C. E., Deeley, R. G. and Cole, S. P.
Year: 1997
Title: Membrane topology of the multidrug resistance protein (MRP). A study of glycosylation-site mutants reveals an extracytosolic NH2 terminus
Journal: J Biol Chem
Volume: 272
Issue: 38
Pages: 23623-30
Epub Date: 1997/09/20
Date: Sep 19
Short Title: Membrane topology of the multidrug resistance protein (MRP). A study of glycosylation-site mutants reveals an extracytosolic NH2 terminus
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.38.23623
Accession Number: 9295302
Keywords: ATP-Binding Cassette Transporters/chemistry/*metabolism
Amines/chemistry
Animals
COS Cells
Cell Membrane/metabolism
Cytosol/metabolism
Drug Resistance, Multiple/genetics
Glycosylation
HeLa Cells
Humans
Multidrug Resistance-Associated Proteins
Mutagenesis, Site-Directed
Trypsin/chemistry
Abstract: Multidrug resistance protein, MRP, is a 190-kDa integral membrane phosphoglycoprotein that belongs to the ATP-binding cassette superfamily of transport proteins and is capable of conferring resistance to multiple chemotherapeutic agents. Previous studies have indicated that MRP consists of two membrane spanning domains (MSD) each followed by a nucleotide binding domain, plus an additional extremely hydrophobic NH2-terminal MSD. Computer-assisted hydropathy analyses and multiple sequence alignments suggest several topological models for MRP. To aid in determining the topology most likely to be correct, we have identified which of the 14 N-glycosylation sequons in this protein are utilized. Limited proteolysis of MRP-enriched membranes and deglycosylation of intact MRP and its tryptic fragments with PNGase F was carried out followed by immunoblotting with antibodies known to react with specific regions of MRP. The results obtained indicated that the sequon at Asn354 in the middle MSD is not utilized and suggested approximate sites of N-glycosylation. Subsequent site-directed mutagenesis studies established that Asn19 and Asn23 in the NH2-terminal MSD and Asn1006 in the COOH-terminal MSD are the only sites in MRP that are modified with N-linked oligosaccharides. N-Glycosylation of Asn19 and Asn23 provides the first direct experimental evidence that MRP has an extracytosolic NH2 terminus. This finding, together with those of previous studies, strongly suggests that the NH2-terminal MSD of MRP contains an odd number of transmembrane helices. These results may have important implications for the further understanding of the interaction of drugs with MRP.
Notes: Hipfner, D R
Almquist, K C
Leslie, E M
Gerlach, J H
Grant, C E
Deeley, R G
Cole, S P
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1997 Sep 19;272(38):23623-30. doi: 10.1074/jbc.272.38.23623.
Author Address: Department of Pathology, Queen's University, Kingston, Ontario, Canada K7L 3N6.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1014
Author: Hirai, H., Pang, Z., Bao, D., Miyazaki, T., Li, L., Miura, E., Parris, J., Rong, Y., Watanabe, M., Yuzaki, M. and Morgan, J. I.
Year: 2005
Title: Cbln1 is essential for synaptic integrity and plasticity in the cerebellum
Journal: Nat Neurosci
Volume: 8
Issue: 11
Pages: 1534-41
Epub Date: 2005/10/20
Date: Nov
Short Title: Cbln1 is essential for synaptic integrity and plasticity in the cerebellum
Alternate Journal: Nature neuroscience
ISSN: 1097-6256 (Print)
1097-6256
DOI: 10.1038/nn1576
Accession Number: 16234806
Keywords: Age Factors
Analysis of Variance
Animals
Animals, Newborn
Ataxia/genetics
Behavior, Animal
Blotting, Northern/methods
Blotting, Western/methods
Cells, Cultured
Cerebellum/*cytology
Cloning, Molecular/methods
Dendritic Spines
Dose-Response Relationship, Radiation
Electric Stimulation/methods
Excitatory Postsynaptic Potentials/physiology/radiation effects
Gene Expression Regulation
Humans
In Situ Hybridization/methods
In Vitro Techniques
Membrane Potentials/physiology/radiation effects
Mice
Mice, Inbred ICR
Mice, Transgenic
Microscopy, Electron, Transmission/methods
Motor Activity/genetics
Mutagenesis/physiology
Nerve Tissue Proteins/genetics/*physiology
Neuronal Plasticity/*physiology
Neurons/*physiology/ultrastructure
Patch-Clamp Techniques/methods
Protein Precursors/genetics/*physiology
RNA, Messenger/metabolism
Radioimmunoassay/methods
Reverse Transcriptase Polymerase Chain Reaction/methods
Synapses/*metabolism/ultrastructure
Transfection/methods
Vesicular Glutamate Transport Protein 2/metabolism
Abstract: Cbln1 is a cerebellum-specific protein of previously unknown function that is structurally related to the C1q and tumor necrosis factor families of proteins. We show that Cbln1 is a glycoprotein secreted from cerebellar granule cells that is essential for three processes in cerebellar Purkinje cells: the matching and maintenance of pre- and postsynaptic elements at parallel fiber-Purkinje cell synapses, the establishment of the proper pattern of climbing fiber-Purkinje cell innervation, and induction of long-term depression at parallel fiber-Purkinje cell synapses. Notably, the phenotype of cbln1-null mice mimics loss-of-function mutations in the orphan glutamate receptor, GluR delta2, a gene selectively expressed in Purkinje neurons. Therefore, Cbln1 secreted from presynaptic granule cells may be a component of a transneuronal signaling pathway that controls synaptic structure and plasticity.
Notes: Hirai, Hirokazu
Pang, Zhen
Bao, Dashi
Miyazaki, Taisuke
Li, Leyi
Miura, Eriko
Parris, Jennifer
Rong, Yongqi
Watanabe, Masahiko
Yuzaki, Michisuke
Morgan, James I
CA 21765/CA/NCI NIH HHS/United States
ES10772/ES/NIEHS NIH HHS/United States
NS040361/NS/NINDS NIH HHS/United States
NS040749/NS/NINDS NIH HHS/United States
NS042828/NS/NINDS NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Nat Neurosci. 2005 Nov;8(11):1534-41. doi: 10.1038/nn1576. Epub 2005 Oct 23.
Author Address: Department of Developmental Neurobiology, St. Jude Children's Research Hospital, 332 North Lauderdale Street, MS 323, Memphis, Tennessee 38105-2794, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2035
Author: Hiraizumi, M., Yamashita, K., Nishizawa, T. and Nureki, O.
Year: 2019
Title: Cryo-EM structures capture the transport cycle of the P4-ATPase flippase
Journal: Science
Volume: 365
Issue: 6458
Pages: 1149-1155
Epub Date: 2019/08/17
Date: Sep 13
Short Title: Cryo-EM structures capture the transport cycle of the P4-ATPase flippase
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.aay3353
Accession Number: 31416931
Keywords: Adenosine Triphosphatases/*chemistry
Adenosine Triphosphate/chemistry
Amino Acid Sequence
Binding Sites
Cryoelectron Microscopy
HEK293 Cells
Humans
Membrane Proteins/*chemistry
Models, Molecular
Phosphatidylserines
Phospholipid Transfer Proteins/*chemistry
Phospholipids/*metabolism
Phosphorylation
Protein Binding
Protein Interaction Domains and Motifs
Protein Structure, Quaternary
Sequence Alignment
Abstract: In eukaryotic membranes, type IV P-type adenosine triphosphatases (P4-ATPases) mediate the translocation of phospholipids from the outer to the inner leaflet and maintain lipid asymmetry, which is critical for membrane trafficking and signaling pathways. Here, we report the cryo-electron microscopy structures of six distinct intermediates of the human ATP8A1-CDC50a heterocomplex at resolutions of 2.6 to 3.3 angstroms, elucidating the lipid translocation cycle of this P4-ATPase. ATP-dependent phosphorylation induces a large rotational movement of the actuator domain around the phosphorylation site in the phosphorylation domain, accompanied by lateral shifts of the first and second transmembrane helices, thereby allowing phosphatidylserine binding. The phospholipid head group passes through the hydrophilic cleft, while the acyl chain is exposed toward the lipid environment. These findings advance our understanding of the flippase mechanism and the disease-associated mutants of P4-ATPases.
Notes: 1095-9203
Hiraizumi, Masahiro
Orcid: 0000-0002-4340-2937
Yamashita, Keitaro
Orcid: 0000-0002-5442-7582
Nishizawa, Tomohiro
Orcid: 0000-0001-7463-8398
Nureki, Osamu
Orcid: 0000-0003-1813-7008
Journal Article
Research Support, Non-U.S. Gov't
United States
Science. 2019 Sep 13;365(6458):1149-1155. doi: 10.1126/science.aay3353. Epub 2019 Aug 15.
Author Address: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
Discovery Technology Laboratories, Innovative Research Division, Mitsubishi Tanabe Pharma Corporation, 1000 Kamoshida, Aoba-ku, Yokohama, 227-0033, Japan.
RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan.
Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan. t-2438@bs.s.u-tokyo.ac.jp nureki@bs.s.u-tokyo.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 873
Author: Hiramatsu, H., Kyono, K., Higashiyama, Y., Fukushima, C., Shima, H., Sugiyama, S., Inaka, K., Yamamoto, A. and Shimizu, R.
Year: 2003
Title: The structure and function of human dipeptidyl peptidase IV, possessing a unique eight-bladed beta-propeller fold
Journal: Biochem Biophys Res Commun
Volume: 302
Issue: 4
Pages: 849-54
Epub Date: 2003/03/21
Date: Mar 21
Short Title: The structure and function of human dipeptidyl peptidase IV, possessing a unique eight-bladed beta-propeller fold
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/s0006-291x(03)00258-4
Accession Number: 12646248
Keywords: Amino Acid Sequence
Catalytic Domain
Dipeptidyl Peptidase 4/*chemistry/genetics
Humans
Models, Molecular
Molecular Sequence Data
Prolyl Oligopeptidases
*Protein Structure, Secondary
Protein Structure, Tertiary
Sequence Alignment
Serine Endopeptidases/chemistry/genetics
Abstract: Dipeptidyl peptidase IV (DPPIV) is a serine protease, a member of the prolyl oligopeptidase (POP) family, and has been implicated in several diseases. Therefore, the development of DPPIV selective inhibitors, which are able to control the biological function of DPPIV, is important. We determined the crystal structure of human DPPIV at 2.6A resolution. The molecule consists of a unique eight-bladed beta-propeller domain in the N-terminal region and a serine protease domain in the C-terminal region. Also, the large "cave" structure, which is thought to control the access of the substrate, is found on the side of the beta-propeller fold. Comparison of the overall amino acid sequence between human DPPIV and POP shows low homology (12.9%). In this paper, we report the structure of human DPPIV, especially focusing on a unique eight-bladed beta-propeller domain. We also discuss the way for the access of the substrate to this domain.
Notes: Hiramatsu, Hajime
Kyono, Kiyoshi
Higashiyama, Yutaka
Fukushima, Chiaki
Shima, Hideaki
Sugiyama, Shigeru
Inaka, Koji
Yamamoto, Atsushi
Shimizu, Ryo
Journal Article
United States
Biochem Biophys Res Commun. 2003 Mar 21;302(4):849-54. doi: 10.1016/s0006-291x(03)00258-4.
Author Address: Discovery Research Laboratory, Tanabe Seiyaku Co. Ltd., 3-16-86 Kashima, Yodogawa-ku, Osaka 532-8505, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1910
Author: Hirata, T., Mishra, S. K., Nakamura, S., Saito, K., Motooka, D., Takada, Y., Kanzawa, N., Murakami, Y., Maeda, Y., Fujita, M., Yamaguchi, Y. and Kinoshita, T.
Year: 2018
Title: Identification of a Golgi GPI-N-acetylgalactosamine transferase with tandem transmembrane regions in the catalytic domain
Journal: Nat Commun
Volume: 9
Issue: 1
Pages: 405
Epub Date: 2018/01/28
Date: Jan 26
Short Title: Identification of a Golgi GPI-N-acetylgalactosamine transferase with tandem transmembrane regions in the catalytic domain
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-017-02799-0
PMCID: PMC5785973
Accession Number: 29374258
Keywords: Acetylgalactosamine/biosynthesis/*chemistry
Amino Acid Motifs
Animals
CHO Cells
Catalytic Domain
Cricetulus
Crystallography, X-Ray
Gene Expression
Genetic Vectors/chemistry/metabolism
Glycosylphosphatidylinositols/*chemistry/metabolism
Golgi Apparatus/*metabolism/ultrastructure
Humans
Mice
Mice, Knockout
Models, Molecular
Mutation
N-Acetylgalactosaminyltransferases/*chemistry/genetics/metabolism
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Structural Homology, Protein
Substrate Specificity
Abstract: Many eukaryotic proteins are anchored to the cell surface via the glycolipid glycosylphosphatidylinositol (GPI). Mammalian GPIs have a conserved core but exhibit diverse N-acetylgalactosamine (GalNAc) modifications, which are added via a yet unresolved process. Here we identify the Golgi-resident GPI-GalNAc transferase PGAP4 and show by mass spectrometry that PGAP4 knockout cells lose GPI-GalNAc structures. Furthermore, we demonstrate that PGAP4, in contrast to known Golgi glycosyltransferases, is not a single-pass membrane protein but contains three transmembrane domains, including a tandem transmembrane domain insertion into its glycosyltransferase-A fold as indicated by comparative modeling. Mutational analysis reveals a catalytic site, a DXD-like motif for UDP-GalNAc donor binding, and several residues potentially involved in acceptor binding. We suggest that a juxtamembrane region of PGAP4 accommodates various GPI-anchored proteins, presenting their acceptor residue toward the catalytic center. In summary, we present insights into the structure of PGAP4 and elucidate the initial step of GPI-GalNAc biosynthesis.
Notes: 2041-1723
Hirata, Tetsuya
Mishra, Sushil K
Orcid: 0000-0002-3080-9754
Nakamura, Shota
Saito, Kazunobu
Motooka, Daisuke
Takada, Yoko
Kanzawa, Noriyuki
Murakami, Yoshiko
Maeda, Yusuke
Fujita, Morihisa
Yamaguchi, Yoshiki
Kinoshita, Taroh
Orcid: 0000-0001-7166-7257
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2018 Jan 26;9(1):405. doi: 10.1038/s41467-017-02799-0.
Author Address: Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, 565-0871, Japan.
WPI Immunology Frontier Research Center, Osaka University, Suita, Osaka, 565-0871, Japan.
National Institute for Physiological Sciences, National Institutes of Natural Sciences, Okazaki, Aichi, 444-8787, Japan.
Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN Global Research Cluster, Wako, Saitama, 351-0198, Japan.
Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, Jiangsu, 214122, China.
Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, 565-0871, Japan. tkinoshi@biken.osaka-u.ac.jp.
WPI Immunology Frontier Research Center, Osaka University, Suita, Osaka, 565-0871, Japan. tkinoshi@biken.osaka-u.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 865
Author: Hiromura, M., Choi, C. H., Sabourin, N. A., Jones, H., Bachvarov, D. and Usheva, A.
Year: 2003
Title: YY1 is regulated by O-linked N-acetylglucosaminylation (O-glcNAcylation)
Journal: J Biol Chem
Volume: 278
Issue: 16
Pages: 14046-52
Epub Date: 2003/02/18
Date: Apr 18
Short Title: YY1 is regulated by O-linked N-acetylglucosaminylation (O-glcNAcylation)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M300789200
Accession Number: 12588874
Keywords: Acetylglucosamine/*metabolism
Amidohydrolases/pharmacology
Animals
Arteries/metabolism
Blotting, Western
Carbohydrates/chemistry
Cell Nucleus/metabolism
Cells, Cultured
DNA-Binding Proteins/*metabolism
Erythroid-Specific DNA-Binding Factors
Galactosyltransferases/metabolism
*Gene Expression Regulation
Glucose/pharmacology
Glycosylation
Golgi Apparatus/metabolism
HeLa Cells
Humans
Insecta
Muscle, Smooth/cytology
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Promoter Regions, Genetic
Protein Binding
Retinoblastoma Protein/metabolism
Time Factors
Transcription Factors/*metabolism
Transcription, Genetic
YY1 Transcription Factor
Abstract: YY1 is a zinc finger DNA-binding transcription factor that influences expression of a wide variety of cellular and viral genes. YY1 is essential for the development of mammalian embryos. It regulates the expression of genes with important functions in DNA replication, protein synthesis, and cellular response to external stimuli during cell growth and differentiation. How YY1 accomplishes such a variety of functions is unknown. Here, we show that a subset of the nuclear YY1 appears to be O-GlcNAcylated regardless of the differentiation status of the cells. We found that glucose strongly stimulates O-linked N-acetylglucosaminylation (O-GlcNAcylation) on YY1. Glycosylated YY1 no longer binds the retinoblastoma protein (Rb). Upon dissociation from Rb, the glycosylated YY1 is free to bind DNA. The ability of the O-glycosylation on YY1 to disrupt the complex with Rb leads us to propose that O-glycosylation might have a profound effect on cell cycle transitions that regulate the YY1-Rb heterodimerization and promote the activity of YY1. Our observations provide strong evidence that YY1-regulated transcription is very likely connected to the pathway of glucose metabolism that culminates in the O-GlcNAcylation on YY1, changing its function in transcription.
Notes: Hiromura, Makoto
Choi, Chu H
Sabourin, Nicaulas A
Jones, Heath
Bachvarov, Dimcho
Usheva, Anny
HL62458/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2003 Apr 18;278(16):14046-52. doi: 10.1074/jbc.M300789200. Epub 2003 Feb 13.
Author Address: Beth Israel Deaconess Medical Center, Department of Medicine, Endocrinology, Harvard Medical School, Boston, Massachussetts 02215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 253
Author: Hitsumoto, Y., Nakano, A., Ohnishi, H., Hamada, F., Saheki, S. and Takeuchi, N.
Year: 1992
Title: Purification of the murine heat-stable antigen from erythrocytes
Journal: Biochem Biophys Res Commun
Volume: 187
Issue: 2
Pages: 773-7
Epub Date: 1992/09/16
Date: Sep 16
Short Title: Purification of the murine heat-stable antigen from erythrocytes
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(92)91262-o
Accession Number: 1530634
Keywords: Amino Acid Sequence
Animals
Antibodies, Monoclonal
*Antigens, CD
Antigens, Differentiation/*blood/chemistry/immunology
CD24 Antigen
Cholic Acids
Electrophoresis, Polyacrylamide Gel
Erythrocyte Membrane/*immunology
Glycosylation
*Membrane Glycoproteins
Mice
Mice, Inbred C57BL
Molecular Sequence Data
Rats
Rats, Inbred Strains
Abstract: The rat anti-mouse erythrocyte (MRBC) monoclonal antibody (mAb), R13, has been developed. The MRBC membrane protein recognized by R13 (R13-Ag) can be purified by loading the butanol-extracted MRBC membrane solution on a R13-conjugated Cellulofine column in the presence of 0.1% CHAPS followed by elution with 1% CHAPS. The amino acid sequence of the affinity-purified R13-Ag corresponded to that predicted from the cDNA for the murine heat-stable antigen. It was revealed that the actual heat-stable antigen was composed of 27 amino acids.
Notes: Hitsumoto, Y
Nakano, A
Ohnishi, H
Hamada, F
Saheki, S
Takeuchi, N
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 1992 Sep 16;187(2):773-7. doi: 10.1016/0006-291x(92)91262-o.
Author Address: Department of Clinical Laboratory Medicine, Ehime University School of Medicine, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 246
Author: Hiyama, J., Weisshaar, G. and Renwick, A. G.
Year: 1992
Title: The asparagine-linked oligosaccharides at individual glycosylation sites in human thyrotrophin
Journal: Glycobiology
Volume: 2
Issue: 5
Pages: 401-9
Epub Date: 1992/10/01
Date: Oct
Short Title: The asparagine-linked oligosaccharides at individual glycosylation sites in human thyrotrophin
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/2.5.401
Accession Number: 1457969
Keywords: Asparagine/chemistry
Carbohydrate Sequence
Glycosylation
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Molecular Structure
Oligosaccharides/chemistry
Thyrotropin/*chemistry
Abstract: The asparagine-linked carbohydrate structures at each of the three glycosylation sites of human thyrotrophin were investigated by 400 MHz 1H-NMR spectroscopy. Highly purified, biologically active human thyrotrophin (hTSH) was dissociated into its subunits hTSH alpha (glycosylated at Asn 52 and Asn 78) and hTSH beta (glycosylated at Asn 23). The alpha-subunit was further treated with trypsin which gave two glycopeptides that were subsequently separated by reverse-phase HPLC and identified by amino acid sequence analysis. The oligosaccharides were liberated from hTSH alpha glycopeptides and from intact hTSH beta by hydrazinolysis, and were fractionated as alditols by anion-exchange and ion-suppression amine-adsorption HPLC preparatory to structural analysis. The N-glycans present on hTSH were mainly diantennary complex-type structures with a common Man alpha 1-3 branch that terminated with 4-O-sulphated GalNAc. The Man alpha 1-6 branch displayed structural heterogeneity in the terminal sequence, with chiefly alpha 2-3-sialylated Gal and/or 4-O-sulphated GalNAc. The relative amounts of the two major complete diantennary oligosaccharides and their core fucosylation differed according to glycosylation site; the sulphated/sialylated diantennary oligosaccharide was most abundant at the two sites on the alpha-subunit, whereas the disulphated, core-fucosylated oligosaccharide was more plentiful on the beta-subunit. Some interesting structural features, not previously reported for the N-glycans of hTSH, included 3-O-sulphated galactose (SO4-3Gal) and peripheral fucose (Fuc alpha 1-3GlcNAc) in the Man alpha 1-6 branch of some diantennary structures; the former suggests the presence of a hitherto uncharacterized galactose-3-O-sulphotransferase in thyrotroph cells of the human anterior pituitary gland.
Notes: Hiyama, J
Weisshaar, G
Renwick, A G
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1992 Oct;2(5):401-9. doi: 10.1093/glycob/2.5.401.
Author Address: Department of Biochemistry, University of Auckland, New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1262
Author: Hlouchova, K., Barinka, C., Konvalinka, J. and Lubkowski, J.
Year: 2009
Title: Structural insight into the evolutionary and pharmacologic homology of glutamate carboxypeptidases II and III
Journal: Febs j
Volume: 276
Issue: 16
Pages: 4448-62
Epub Date: 2009/08/15
Date: Aug
Short Title: Structural insight into the evolutionary and pharmacologic homology of glutamate carboxypeptidases II and III
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/j.1742-4658.2009.07152.x
Accession Number: 19678840
Keywords: Antigens, Surface/*chemistry
Binding Sites
Carboxypeptidases/*chemistry
Catalytic Domain
Crystallography, X-Ray
Glutamate Carboxypeptidase II/*chemistry
Glutamic Acid/chemistry
Humans
Ligands
Molecular Mimicry
Phosphopeptides/chemistry
Protein Conformation
Protein Structure, Quaternary
Quisqualic Acid/chemistry
Abstract: Glutamate carboxypeptidase III (GCPIII) is a metalloenzyme that belongs to the transferrin receptor/glutamate carboxypeptidase II (GCPII; EC 3.4.17.21) superfamily. GCPIII has been studied mainly because of its evolutionary relationship to GCPII, an enzyme involved in a variety of neuropathologies and malignancies, such as glutamatergic neurotoxicity and prostate cancer. Given the potential functional and pharmacological overlap between GCPIII and GCPII, studies addressing the structural and physiological properties of GCPIII are crucial for obtaining a deeper understanding of the GCPII/GCPIII system. In the present study, we report high-resolution crystal structures of the human GCPIII ectodomain in a 'pseudo-unliganded' state and in a complex with: (a) L-glutamate (a product of hydrolysis); (b) a phosphapeptide transition state mimetic, namely (2S,3'S)-{[(3'-amino-3'-carboxy-propyl)-hydroxyphosphinoyl]methyl}-pentanedioic acid; and (c) quisqualic acid, a glutamate biostere. Our data reveal the overall fold and quaternary arrangement of the GCPIII molecule, define the architecture of the GCPIII substrate-binding cavity, and offer an experimental evidence for the presence of Zn(2+) ions in the bimetallic active site. Furthermore, the structures allow us to detail interactions between the enzyme and its ligands and to characterize the functional flexibility of GCPIII, which is essential for substrate recognition. A comparison of these GCPIII structures with the equivalent GCPII complexes reveals differences in the organization of specificity pockets, in surface charge distribution, and in the occupancy of the co-catalytic zinc sites. The data presented here provide information that should prove to be essential for the structurally-aided design of GCPIII-specific inhibitors and might comprise guidelines for future comparative GCPII/GCPIII studies.
Notes: 1742-4658
Hlouchova, Klara
Barinka, Cyril
Konvalinka, Jan
Lubkowski, Jacek
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
England
FEBS J. 2009 Aug;276(16):4448-62. doi: 10.1111/j.1742-4658.2009.07152.x.
Author Address: Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1279
Author: Ho, S. R., Wang, K., Whisenhunt, T. R., Huang, P., Zhu, X., Kudlow, J. E. and Paterson, A. J.
Year: 2010
Title: O-GlcNAcylation enhances FOXO4 transcriptional regulation in response to stress
Journal: FEBS Lett
Volume: 584
Issue: 1
Pages: 49-54
Epub Date: 2009/11/26
Date: Jan 4
Short Title: O-GlcNAcylation enhances FOXO4 transcriptional regulation in response to stress
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2009.11.059
PMCID: PMC2802345
NIHMSID: NIHMS162690
Accession Number: 19932102
Keywords: Acetylglucosamine/*metabolism
Acylation
Cell Cycle Proteins
Cell Line
Forkhead Transcription Factors
*Gene Expression Regulation
Humans
N-Acetylglucosaminyltransferases/metabolism
Oxidative Stress/*genetics
Transcription Factors/*metabolism
*Transcription, Genetic
beta-N-Acetylhexosaminidases/metabolism
Abstract: The FOXO4 transcription factor plays an important role in cell survival in response to oxidative stress. The regulation of FOXO4 is orchestrated by post-translational modifications including phosphorylation, acetylation, and ubiquitination. Here, we demonstrate that O-GlcNAcylation also contributes to the FOXO4-dependent oxidative stress response. We show that hydrogen peroxide treatment of HEK293 cells increases FOXO4 association with OGT, the enzyme that adds O-GlcNAc to proteins, causing FOXO4 O-GlcNAcylation and enhanced transcriptional activity under acute oxidative stress. O-GlcNAcylation is known to be protective for cells under stress conditions, including oxidative stress. Our data provide a mechanism of FOXO4 anti-oxidative protection through O-GlcNAcylation.
Notes: 1873-3468
Ho, Shiuh-Rong
Wang, Kai
Whisenhunt, Thomas R
Huang, Ping
Zhu, Xiaolin
Kudlow, Jeffrey E
Paterson, Andrew J
R01 DK043652/DK/NIDDK NIH HHS/United States
R01 DK043652-17/DK/NIDDK NIH HHS/United States
DK043652/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
FEBS Lett. 2010 Jan 4;584(1):49-54. doi: 10.1016/j.febslet.2009.11.059.
Author Address: Department of Cell Biology, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 450
Author: Hochstrasser, K., Schönberger, O. L., Rossmanith, I. and Wachter, E.
Year: 1981
Title: Kunitz-type proteinase inhibitors derived by limited proteolysis of the inter-alpha-trypsin inhibitor, V. Attachments of carbohydrates in the human urinary trypsin inhibitor isolated by affinity chromatography
Journal: Hoppe Seylers Z Physiol Chem
Volume: 362
Issue: 10
Pages: 1357-62
Epub Date: 1981/10/01
Date: Oct
Short Title: Kunitz-type proteinase inhibitors derived by limited proteolysis of the inter-alpha-trypsin inhibitor, V. Attachments of carbohydrates in the human urinary trypsin inhibitor isolated by affinity chromatography
Alternate Journal: Hoppe-Seyler's Zeitschrift fur physiologische Chemie
ISSN: 0018-4888 (Print)
0018-4888
DOI: 10.1515/bchm2.1981.362.2.1357
Accession Number: 6171497
Keywords: Alpha-Globulins/isolation & purification/*urine
Amino Acid Sequence
Carbohydrates/analysis
Chromatography, Affinity
Glycopeptides/analysis
Glycoproteins/isolation & purification/*urine
Humans
Protein Conformation
Trypsin Inhibitors/isolation & purification/*urine
Abstract: The inhibitory active part of the inter-alpha-trypsin inhibitor with a known amino acid sequence is present as an acid-resistant inhibitor in human serum, in urine, in bronchial and in nasal mucus. The inhibitor molecule has a 50% carbohydrate content. Carbohydrate side chains are attached in two positions. One chain is linked to the polypeptide O-glycosidically via the serine residue in position 10 in the N-terminal extension peptide. The second side chain is attached N-glycosidically via the asparagine residue in position 24, located in the inactive "inhibitory" Kunitz-type domain of the inhibitor. The composition of the carbohydrate side chains were determined.
Notes: Hochstrasser, K
Schönberger, O L
Rossmanith, I
Wachter, E
Journal Article
Research Support, Non-U.S. Gov't
Germany
Hoppe Seylers Z Physiol Chem. 1981 Oct;362(10):1357-62. doi: 10.1515/bchm2.1981.362.2.1357.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 626
Author: Hoffman, R. C., Andersen, H., Walker, K., Krakover, J. D., Patel, S., Stamm, M. R. and Osborn, S. G.
Year: 1996
Title: Peptide, disulfide, and glycosylation mapping of recombinant human thrombopoietin from ser1 to Arg246
Journal: Biochemistry
Volume: 35
Issue: 47
Pages: 14849-61
Epub Date: 1996/11/26
Date: Nov 26
Short Title: Peptide, disulfide, and glycosylation mapping of recombinant human thrombopoietin from ser1 to Arg246
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi961075b
Accession Number: 8942648
Keywords: Amino Acid Sequence
Arginine/*metabolism
Carbohydrate Sequence
Chromatography/methods
Disulfides/metabolism
Glycosylation
Humans
Molecular Sequence Data
Peptide Mapping
Polysaccharides/metabolism
Protein Processing, Post-Translational
Recombinant Proteins/genetics/metabolism
Serine/*metabolism
Spectrophotometry/methods
Thrombopoietin/genetics/*metabolism
Abstract: Thrombopoietin (TPO) is a hematopoietic factor involved in the regulation of megakaryocytopoiesis. Full length recombinant human TPO (332 residues) has been expressed in BHK cells and purified to homogeneity using conventional means. Peptide, disulfide, and glycosylation mapping of human TPO from residues 1 to 246 has been carried out using liquid chromatography-electrospray mass spectrometry (LC-ESMS). A modification of the ramped orifice method of Carr and co-workers [Carr et al. (1993) Protein Sci. 2, 183-196] is employed, providing additional information for assignment of the LC-ESMS chromatograms. With the modification, b- and y-series peptide ions are produced via front-end CID which confirms the mass-based assignments. The results of our analysis of TPO indicate that the amino acid sequence of TPO 1-246 is as expected from the transfected cDNA with complete cleavage of the signal peptide. Two unique disulfides are formed between the four cysteines in the cytokine domain of TPO: Cys7-Cys151 and Cys29-Cys85. The glycosylation map indicates the position, occupancy, and structures of the N- and O-glycans in TPO 1-246. In addition, site specific structural characterization of the PNGase F-liberated N-glycans has been performed following purification by high-pH anionic exchange chromatography with pulsed amperometric detection (HPAEC-PAD); the results corroborate the LC-ESMS data. The N-glycans are of the complex type with the core-fucosylated disialylated biantennary and trisialylated triantennary structures predominating. The O-glycans are of the mucin type with the monosialylated and disialylated GalGalNAc-S/T structures predominating. Furthermore, we propose that the C-terminal domain of TPO be further divided into two domains on the basis of sequence homology among the cloned sequences and glycosylation/structural features: an N-glycan domain (154-246) and an O-glycan domain (247-332).
Notes: Hoffman, R C
Andersen, H
Walker, K
Krakover, J D
Patel, S
Stamm, M R
Osborn, S G
Journal Article
United States
Biochemistry. 1996 Nov 26;35(47):14849-61. doi: 10.1021/bi961075b.
Author Address: Department of Biological Structure, ZymoGenetics, Inc., Seattle, Washington 98102, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 924
Author: Hoffman, R. C., Jennings, L. L., Tsigelny, I., Comoletti, D., Flynn, R. E., Sudhof, T. C. and Taylor, P.
Year: 2004
Title: Structural characterization of recombinant soluble rat neuroligin 1: mapping of secondary structure and glycosylation by mass spectrometry
Journal: Biochemistry
Volume: 43
Issue: 6
Pages: 1496-506
Epub Date: 2004/02/11
Date: Feb 17
Short Title: Structural characterization of recombinant soluble rat neuroligin 1: mapping of secondary structure and glycosylation by mass spectrometry
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi035278t
Accession Number: 14769026
Keywords: Amino Acid Sequence
Animals
Asparagine/chemistry
Cell Adhesion Molecules, Neuronal/chemistry
Chromatography, High Pressure Liquid
Disulfides/chemistry
Extracellular Space/chemistry
Glycosylation
Mass Spectrometry
Membrane Proteins/*chemistry/metabolism
Molecular Sequence Data
Mucins/chemistry
Nerve Tissue Proteins/*chemistry/metabolism
Oligosaccharides/chemistry
Peptide Fragments/chemistry
*Peptide Mapping/methods
Protein Structure, Secondary
Rats
Recombinant Proteins/*chemistry
Solubility
Trypsin/chemistry
Abstract: Neuroligins (NLs) are a family of transmembrane proteins that function in synapse formation and/or remodeling by interacting with beta-neurexins (beta-NXs) to form heterophilic cell adhesions. The large N-terminal extracellular domain of NLs, required for beta-NX interactions, has sequence homology to the alpha/beta hydrolase fold superfamily of proteins. By peptide mapping and mass spectrometric analysis of a soluble recombinant form of NL1, several structural features of the extracellular domain have been established. Of the nine cysteine residues in NL1, eight are shown to form intramolecular disulfide bonds. Disulfide pairings of Cys 117 to Cys 153 and Cys 342 to Cys 353 are consistent with disulfide linkages that are conserved among the family of alpha/beta hydrolase proteins. The disulfide bond between Cys 172 and Cys 181 occurs within a region of the protein encoded by an alternatively spliced exon. The disulfide pairing of Cys 512 and Cys 546 in NL1 yields a structural motif unique to the NLs, since these residues are highly conserved. The potential N-glycosylation sequons in NL1 at Asn 109, Asn 303, Asn 343, and Asn 547 are shown occupied by carbohydrate. An additional consensus sequence for N-glycosylation at Asn 662 is likely occupied. Analysis of N-linked oligosaccharide content by mass matching paradigms reveals significant microheterogeneous populations of complex glycosyl moieties. In addition, O-linked glycosylation is observed in the predicted stalk region of NL1, prior to the transmembrane spanning domain. From predictions based on sequence homology of NL1 to acetylcholinesterase and the molecular features of NL1 established from mass spectrometric analysis, a novel topology model for NL three-dimensional structure has been constructed.
Notes: Hoffman, Ross C
Jennings, Lori L
Tsigelny, Igor
Comoletti, Davide
Flynn, Robyn E
Sudhof, Thomas C
Taylor, Palmer
P42-ES10337/ES/NIEHS NIH HHS/United States
R37 GM-18360/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2004 Feb 17;43(6):1496-506. doi: 10.1021/bi035278t.
Author Address: Department of Pharmacology and Howard Hughes Medical Institute Mass Spectrometry Facility, University of California, San Diego, La Jolla 92093, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 577
Author: Hoffmann, A., Conradt, H. S., Gross, G., Nimtz, M., Lottspeich, F. and Wurster, U.
Year: 1993
Title: Purification and chemical characterization of beta-trace protein from human cerebrospinal fluid: its identification as prostaglandin D synthase
Journal: J Neurochem
Volume: 61
Issue: 2
Pages: 451-6
Epub Date: 1993/08/01
Date: Aug
Short Title: Purification and chemical characterization of beta-trace protein from human cerebrospinal fluid: its identification as prostaglandin D synthase
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042 (Print)
0022-3042
DOI: 10.1111/j.1471-4159.1993.tb02145.x
Accession Number: 8336134
Keywords: Amino Acid Sequence
Binding Sites
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
*Intramolecular Oxidoreductases
Isomerases/*cerebrospinal fluid/chemistry
Lipocalins
Methylation
Molecular Sequence Data
Molecular Weight
Peptide Fragments/chemistry/metabolism
Peptide Mapping
Protein Processing, Post-Translational
Trypsin/metabolism
Abstract: beta-Trace protein from pooled human CSF was purified to homogeneity. An apparent molecular mass of 23-29 kDa was determined for the polypeptide on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Amino-terminal sequencing of the polypeptide yielded the unique amino acid sequence APEAQVSVQPNFQQDKFLGRWFSA. Alignment of amino acid sequences obtained from tryptic peptides with the sequence previously deduced from a cDNA clone isolated by other investigators allowed the identification of beta-trace protein as prostaglandin D synthase [prostaglandin-H2 D-isomerase; (5Z, 13E)-(15S)-9 alpha, 11 alpha-epidioxy-15-hydroxyprosta-5,13-dienoate D-isomerase; EC 5.3.99.2]. A conservative amino acid exchange (Thr instead of Ser) was detected at amino acid position 154 of the beta-trace polypeptide chain in the corresponding tryptic peptide. The two N-glycosylation sites of the polypeptide were shown to be almost quantitatively occupied by carbohydrate. Carbohydrate compositional as well as methylation analysis indicated that Asn29 and Asn56 bear exclusively complex-type oligosaccharide structures (partially sialylated with alpha 2-3- and/or alpha 2-6-linked N-acetylneuraminic acid) that are almost quantitatively alpha 1-6 fucosylated at the proximal N-acetylglucosamine; approximately 70% of these molecules contain a bisecting N-acetylglucosamine. Agalacto structures as well as those with a peripheral fucose are also present.
Notes: Hoffmann, A
Conradt, H S
Gross, G
Nimtz, M
Lottspeich, F
Wurster, U
Journal Article
England
J Neurochem. 1993 Aug;61(2):451-6. doi: 10.1111/j.1471-4159.1993.tb02145.x.
Author Address: CSF-Laboratory, Department of Neurology, Medizinische Hochschule Hannover, Hannover, F.R.G.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 541
Author: Hoffmann, A., Nimtz, M., Getzlaff, R. and Conradt, H. S.
Year: 1995
Title: 'Brain-type' N-glycosylation of asialo-transferrin from human cerebrospinal fluid
Journal: FEBS Lett
Volume: 359
Issue: 2-3
Pages: 164-8
Epub Date: 1995/02/13
Date: Feb 13
Short Title: 'Brain-type' N-glycosylation of asialo-transferrin from human cerebrospinal fluid
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(95)00034-7
Accession Number: 7867791
Keywords: Amino Acid Sequence
Asialoglycoproteins/*cerebrospinal fluid/chemistry
Carbohydrate Sequence
Glycosylation
Humans
Molecular Sequence Data
Oligosaccharides/analysis
Polysaccharides/analysis
Transferrin/*analogs & derivatives/cerebrospinal fluid/chemistry
Abstract: Asialo-transferrin from human cerebrospinal fluid was purified to homogeneity. Investigation of the structural characteristics of its oligosaccharides support our hypothesis of 'brain-type' glycosylation of intrathecally synthesized cerebrospinal fluid proteins. For carbohydrate structural analysis, high-pH anion-exchange chromatography, methylation analysis, liquid secondary ion- and matrix-assisted laser desorption/ ionization mass spectrometry of the permethylated derivatives were used. The major structure turned out to be a complex-type agalactodiantennary oligosaccharide with bisecting N-acetylglucosamine and proximal fucose. Analysis of a second transferrin preparation containing both asialo- and sialo-transferrin revealed another major glycan species derived from the sialylated transferrin variant which is galactosylated and lacks bisecting N-acetylglucosamine and fucose.
Notes: Hoffmann, A
Nimtz, M
Getzlaff, R
Conradt, H S
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1995 Feb 13;359(2-3):164-8. doi: 10.1016/0014-5793(95)00034-7.
Author Address: Department of Molecular Biology, GBF - Gesellschaft für Biotechnologische Forschung, Braunschweig, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 509
Author: Hoffmann, A., Nimtz, M., Wurster, U. and Conradt, H. S.
Year: 1994
Title: Carbohydrate structures of beta-trace protein from human cerebrospinal fluid: evidence for "brain-type" N-glycosylation
Journal: J Neurochem
Volume: 63
Issue: 6
Pages: 2185-96
Epub Date: 1994/12/01
Date: Dec
Short Title: Carbohydrate structures of beta-trace protein from human cerebrospinal fluid: evidence for "brain-type" N-glycosylation
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042 (Print)
0022-3042
DOI: 10.1046/j.1471-4159.1994.63062185.x
Accession Number: 7525874
Keywords: Acetylglucosamine/analysis
Amidohydrolases/metabolism
Beta-Globulins/*cerebrospinal fluid/*chemistry
Binding Sites
Carbohydrate Conformation
Carbohydrate Metabolism
Carbohydrate Sequence
Carbohydrates/*chemistry
Chromatography, High Pressure Liquid
Fucose/metabolism
Galactose/analysis
Glycosylation
Humans
*Intramolecular Oxidoreductases
Lipocalins
Molecular Sequence Data
N-Acetylneuraminic Acid
Neuraminidase/metabolism
Oligosaccharides/chemistry
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Repetitive Sequences, Nucleic Acid
Sialic Acids/analysis/chemistry
Trypsin/metabolism
Abstract: The carbohydrate structures of beta-trace protein from human cerebrospinal fluid have been elucidated. This protein carries exclusively N-linked oligosaccharides at two sites (Asn29 and Asn56). Enzymatically released N-glycans were studied by compositional and methylation analyses, high-pH anion-exchange chromatography, and liquid secondary ion mass spectrometry. All glycans were found to be of the complex type, and most (90%) of them were biantennary with no (40%), one (40%), or two (20%) N-acetylneuraminic acid residues. The rest were triantennary chains or biantennary chains with intact or truncated lactosamine repeats. The innermost N-acetylglucosamine residues of nearly all structures were found to be alpha 1,6-fucosylated. Peripheral fucose (about 20% alpha 1,3-linked to N-acetylglucosamine) was also detected. Seventy percent of the oligosaccharides contained a bisecting N-acetylglucosamine. Especially in the neutral, but also in the monosialylated oligosaccharide fractions, many incomplete antennae consisting of N-acetylglucosamine only were present. At least 20 different N-glycans were identified. Analysis of the site-specific glycosylation patterns at Asn29 and Asn56 revealed only minor differences. According to the structural features (a high degree of fucosylation, high amounts of bisecting N-acetylglucosamine, as well as terminal N-acetylglucosamine and galactose residues, and significant amounts of N-acetylneuraminic acid in alpha 2,3 linkage), this protein can be classified as "brain-type" glycosylated.
Notes: Hoffmann, A
Nimtz, M
Wurster, U
Conradt, H S
Journal Article
Research Support, Non-U.S. Gov't
England
J Neurochem. 1994 Dec;63(6):2185-96. doi: 10.1046/j.1471-4159.1994.63062185.x.
Author Address: Department of Neurology, Medizinische Hochschule Hannover, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1469
Author: Hoffmann, B. R., Liu, Y. and Mosher, D. F.
Year: 2012
Title: Modification of EGF-like module 1 of thrombospondin-1, an animal extracellular protein, by O-linked N-acetylglucosamine
Journal: PLoS One
Volume: 7
Issue: 3
Pages: e32762
Epub Date: 2012/03/10
Short Title: Modification of EGF-like module 1 of thrombospondin-1, an animal extracellular protein, by O-linked N-acetylglucosamine
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0032762
PMCID: PMC3293841
Accession Number: 22403705
Keywords: Acetylglucosamine/*chemistry/*metabolism
Amino Acid Sequence
Animals
Consensus Sequence
Drosophila melanogaster
Epidermal Growth Factor/*chemistry
Extracellular Space/*metabolism
Humans
Molecular Sequence Data
*Oxygen
Sequence Analysis, DNA
Thrombospondin 1/*chemistry/*metabolism
Abstract: Thrombospondin-1 (TSP-1) is known to be subject to three unusual carbohydrate modifications: C-mannosylation, O-fucosylation, and O-glucosylation. We now describe a fourth: O-β-N-acetylglucosaminylation. Previously, O-β-N-acetylglucosamine (O-β-GlcNAc) was found on a threonine in the loop between the fifth and sixth cysteines of the 20(th) epidermal growth factor (EGF)-like module of Drosophila Notch. A BLAST search based on the Drosophila Notch loop sequence identified a number of human EGF-like modules that contain a similar sequence, including EGF-like module 1 of TSP-1 and its homolog, TSP-2. TSP-1, which has a potentially modifiable serine in the loop, reacted in immuno-blots with the CTD110.6 anti-O-GlcNAc antibody. Antibody reactivity was diminished by treatment of TSP-1 with β-N-acetylhexosaminidase. TSP-2, which lacks a potentially modifiable serine/threonine in the loop, did not react with CTD110.6. Analysis of tandem modules of TSP-1 localized reactivity of CTD110.6 to EGF-like module 1. Top-down mass spectrometric analysis of EGF-like module 1 demonstrated the expected modifications with glucose (+162 Da) and xylose (+132 Da) separately from modification with N-acetyl hexosamine (+203 Da). Mass spectrometric sequence analysis localized the +203-Da modification to Ser580 in the sequence (575)CPPGYSGNGIQC(586). These results demonstrate that O-β-N-acetylglucosaminylation can occur on secreted extracellular matrix proteins as well as on cell surface proteins.
Notes: 1932-6203
Hoffmann, Brian R
Liu, Yuanyuan
Mosher, Deane F
R01 HL054462/HL/NHLBI NIH HHS/United States
T32 HL007899/HL/NHLBI NIH HHS/United States
R01HL054462/HL/NHLBI NIH HHS/United States
T32-HL007899/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
PLoS One. 2012;7(3):e32762. doi: 10.1371/journal.pone.0032762. Epub 2012 Mar 5.
Author Address: Departments of Medicine and Biomolecular Chemistry, University of Wisconsin, Madison, Wisconsin, United States of America.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1746
Author: Hoffmann, M., Marx, K., Reichl, U., Wuhrer, M. and Rapp, E.
Year: 2016
Title: Site-specific O-Glycosylation Analysis of Human Blood Plasma Proteins
Journal: Mol Cell Proteomics
Volume: 15
Issue: 2
Pages: 624-41
Epub Date: 2015/11/26
Date: Feb
Short Title: Site-specific O-Glycosylation Analysis of Human Blood Plasma Proteins
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M115.053546
PMCID: PMC4739677
Accession Number: 26598643
Keywords: Acute-Phase Proteins/biosynthesis/*genetics
Antigens, Viral, Tumor/blood/*genetics
Chromatography, Reverse-Phase
Glycopeptides/blood/*genetics
Glycosylation
Healthy Volunteers
Humans
Hydrophobic and Hydrophilic Interactions
Mass Spectrometry
Mucin-1/blood/genetics
Polysaccharides/blood/*genetics
Abstract: Site-specific glycosylation analysis is key to investigate structure-function relationships of glycoproteins, e.g. in the context of antigenicity and disease progression. The analysis, though, is quite challenging and time consuming, in particular for O-glycosylated proteins. In consequence, despite their clinical and biopharmaceutical importance, many human blood plasma glycoproteins have not been characterized comprehensively with respect to their O-glycosylation. Here, we report on the site-specific O-glycosylation analysis of human blood plasma glycoproteins. To this end pooled human blood plasma of healthy donors was proteolytically digested using a broad-specific enzyme (Proteinase K), followed by a precipitation step, as well as a glycopeptide enrichment and fractionation step via hydrophilic interaction liquid chromatography, the latter being optimized for intact O-glycopeptides carrying short mucin-type core-1 and -2 O-glycans, which represent the vast majority of O-glycans on human blood plasma proteins. Enriched O-glycopeptide fractions were subjected to mass spectrometric analysis using reversed-phase liquid chromatography coupled online to an ion trap mass spectrometer operated in positive-ion mode. Peptide identity and glycan composition were derived from low-energy collision-induced dissociation fragment spectra acquired in multistage mode. To pinpoint the O-glycosylation sites glycopeptides were fragmented using electron transfer dissociation. Spectra were annotated by database searches as well as manually. Overall, 31 O-glycosylation sites and regions belonging to 22 proteins were identified, the majority being acute-phase proteins. Strikingly, also 11 novel O-glycosylation sites and regions were identified. In total 23 O-glycosylation sites could be pinpointed. Interestingly, the use of Proteinase K proved to be particularly beneficial in this context. The identified O-glycan compositions most probably correspond to mono- and disialylated core-1 mucin-type O-glycans (T-antigen). The developed workflow allows the identification and characterization of the major population of the human blood plasma O-glycoproteome and our results provide new insights, which can help to unravel structure-function relationships. The data were deposited to ProteomeXchange PXD003270.
Notes: 1535-9484
Hoffmann, Marcus
Marx, Kristina
Reichl, Udo
Wuhrer, Manfred
Rapp, Erdmann
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2016 Feb;15(2):624-41. doi: 10.1074/mcp.M115.053546. Epub 2015 Nov 23.
Author Address: From the ‡Max Planck Institute for Dynamics of Complex Technical Systems, Bioprocess Engineering, 39106 Magdeburg, Germany;
§Bruker Daltonik GmbH, 28359 Bremen, Germany;
From the ‡Max Planck Institute for Dynamics of Complex Technical Systems, Bioprocess Engineering, 39106 Magdeburg, Germany; ¶Otto von Guericke University Magdeburg, Chair of Bioprocess Engineering, 39106 Magdeburg, Germany;
‖Center for Proteomics and Metabolomics, Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands;
From the ‡Max Planck Institute for Dynamics of Complex Technical Systems, Bioprocess Engineering, 39106 Magdeburg, Germany; **glyXera GmbH, Leipziger Strasse 44 (Zenit), 39120 Magdeburg, Germany rapp@mpi-magdeburg.mpg.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 816
Author: Hofmann, B. A., Sydow, S., Jahn, O., van Werven, L., Liepold, T., Eckart, K. and Spiess, J.
Year: 2001
Title: Functional and protein chemical characterization of the N-terminal domain of the rat corticotropin-releasing factor receptor 1
Journal: Protein Sci
Volume: 10
Issue: 10
Pages: 2050-62
Epub Date: 2001/09/22
Date: Oct
Short Title: Functional and protein chemical characterization of the N-terminal domain of the rat corticotropin-releasing factor receptor 1
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.12101
PMCID: PMC2374224
Accession Number: 11567096
Keywords: Amino Acid Sequence
Animals
Cells, Cultured
Disulfides/*chemistry
Gene Deletion
Glycosylation
Humans
Ligands
Mass Spectrometry
Molecular Sequence Data
Protein Sorting Signals/physiology
Protein Structure, Secondary
Protein Structure, Tertiary/physiology
Rats
Receptors, Corticotropin-Releasing Hormone/chemistry/genetics/metabolism/*physiology
Abstract: Rat corticotropin-releasing factor receptor 1 (rCRFR1) was produced either in transfected HEK 293 cells as a complex glycosylated protein or in the presence of the mannosidase I inhibitor kifunensine as a high mannose glycosylated protein. The altered glycosylation did not influence the biological function of rCRFR1 as demonstrated by competitive binding of rat urocortin (rUcn) or human/rat corticotropin-releasing factor (h/rCRF) and agonist-induced cAMP accumulation. The low production rate of the N-terminal domain of rCRFR1 (rCRFR1-NT) by transfected HEK 293 cells, was increased by a factor of 100 in the presence of kifunensine. The product, rCRFR1-NT-Kif, bound rUcn specifically (K(D) = 27 nM) and astressin (K(I) = 60 nM). This affinity was 10-fold lower than the affinity of full length rCRFR1. However, it was sufficiently high for rCRFR1-NT-Kif to serve as a model for the N-terminal domain of rCRFR1. With protein fragmentation, Edman degradation, and mass spectrometric analysis, evidence was found for the signal peptide cleavage site C-terminally to Thr(23) and three disulfide bridges between precursor residues 30 and 54, 44 and 87, and 68 and 102. Of all putative N-glycosylation sites in positions 32, 38, 45, 78, 90, and 98, all Asn residues except for Asn(32) were glycosylated to a significant extent. No O-glycosylation was observed.
Notes: 1469-896x
Hofmann, B A
Sydow, S
Jahn, O
van Werven, L
Liepold, T
Eckart, K
Spiess, J
Journal Article
Protein Sci. 2001 Oct;10(10):2050-62. doi: 10.1110/ps.12101.
Author Address: Department of Molecular Neuroendocrinology, Max Planck Institute for Experimental Medicine, D-37073 Göttingen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1845
Author: Hofmann, L., Alexander, N. S., Sun, W., Zhang, J., Orban, T. and Palczewski, K.
Year: 2017
Title: Hydrogen/Deuterium Exchange Mass Spectrometry of Human Green Opsin Reveals a Conserved Pro-Pro Motif in Extracellular Loop 2 of Monostable Visual G Protein-Coupled Receptors
Journal: Biochemistry
Volume: 56
Issue: 17
Pages: 2338-2348
Epub Date: 2017/04/13
Date: May 2
Short Title: Hydrogen/Deuterium Exchange Mass Spectrometry of Human Green Opsin Reveals a Conserved Pro-Pro Motif in Extracellular Loop 2 of Monostable Visual G Protein-Coupled Receptors
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/acs.biochem.7b00165
PMCID: PMC5501310
NIHMSID: NIHMS873298
Accession Number: 28402104
Keywords: Amino Acid Motifs
Amino Acid Substitution
Asparagine/metabolism
Binding Sites
Computational Biology
Cone Opsins/*chemistry/genetics/metabolism/radiation effects
Conserved Sequence
Deuterium Exchange Measurement
Glycosylation
Humans
Ligands
Light
Point Mutation
Proline/chemistry
Protein Conformation
Protein Refolding/radiation effects
Protein Structure, Tertiary
Receptors, G-Protein-Coupled/*chemistry/genetics/metabolism/radiation effects
Recombinant Proteins
Rod Opsins/*chemistry/genetics/metabolism/radiation effects
Spectrometry, Mass, Electrospray Ionization
Tandem Mass Spectrometry
Abstract: Opsins comprise the protein component of light sensitive G protein-coupled receptors (GPCRs) in the retina of the eye that are responsible for the transduction of light into a biochemical signal. Here, we used hydrogen/deuterium (H/D) exchange coupled with mass spectrometry to map conformational changes in green cone opsin upon light activation. We then compared these findings with those reported for rhodopsin. The extent of H/D exchange in green cone opsin was greater than in rhodopsin in the dark and bleached states, suggesting a higher structural heterogeneity for green cone opsin. Further analysis revealed that green cone opsin exists as a dimer in both dark (inactive) and bleached (active) states, and that the predicted glycosylation sites at N(32) and N(34) are indeed glycosylated. Comparison of deuterium uptake between inactive and active states of green cone opsin also disclosed a reduced solvent accessibility of the extracellular N-terminal region and an increased accessibility of the chromophore binding site. Increased H/D exchange at the extracellular side of transmembrane helix four (TM4) combined with an analysis of sequence alignments revealed a conserved Pro-Pro motif in extracellular loop 2 (EL2) of monostable visual GPCRs. These data present new insights into the locus of chromophore release at the extracellular side of TM4 and TM5 and provide a foundation for future functional evaluation.
Notes: 1520-4995
Hofmann, Lukas
Alexander, Nathan S
Sun, Wenyu
Zhang, Jianye
Orban, Tivadar
Palczewski, Krzysztof
Orcid: 0000-0002-0788-545x
R01 EY009339/EY/NEI NIH HHS/United States
R24 EY027283/EY/NEI NIH HHS/United States
U01 EY025451/EY/NEI NIH HHS/United States
K99 EY025007/EY/NEI NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochemistry. 2017 May 2;56(17):2338-2348. doi: 10.1021/acs.biochem.7b00165. Epub 2017 Apr 21.
Author Address: Department of Pharmacology and Cleveland Center for Membrane and Structural Biology, School of Medicine, Case Western Reserve University , 10900 Euclid Avenue, Cleveland, Ohio 44106, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 730
Author: Hofsteenge, J., Blommers, M., Hess, D., Furmanek, A. and Miroshnichenko, O.
Year: 1999
Title: The four terminal components of the complement system are C-mannosylated on multiple tryptophan residues
Journal: J Biol Chem
Volume: 274
Issue: 46
Pages: 32786-94
Epub Date: 1999/11/07
Date: Nov 12
Short Title: The four terminal components of the complement system are C-mannosylated on multiple tryptophan residues
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.274.46.32786
Accession Number: 10551839
Keywords: Complement Membrane Attack Complex/*chemistry
Glycopeptides/chemistry
Glycoproteins/chemistry
Glycosylation
Humans
Macromolecular Substances
Magnetic Resonance Spectroscopy
Mannose/*chemistry
Mass Spectrometry
Peptide Fragments/chemistry
Thrombospondins/chemistry
Tryptophan/*analogs & derivatives/chemistry
Abstract: C-Mannosylation is a unique form of protein glycosylation, involving the C-glycosidic attachment of a mannosyl residue to the indole moiety of Trp. In the two examples found so far, human RNase 2 and interleukin-12, only the first Trp in the recognition motif WXXW is specifically C-mannosylated. To establish the generality of protein C-mannosylation, and to learn more about its mechanism, the terminal components of the human complement system (C6, C7, C8,and C9), which contain multiple and complex recognition motifs, were examined. Together with C5b they form the cytolytic agent, the membrane attack complex. These are the first proteins that are C-mannosylated on more than one Trp residue as follows: six in C6, four in C7, C8alpha, and C8beta, and two in C9. Thus, from the 113 Trp residues in the complete membrane attack complex, 50 were found to undergo C-mannosylation. The other important finding is that in C6, C7, C8, and C9 Trp residues without a second Trp (or another aromatic residue) at the +3 position can be C-mannosylated. This shows that they must contain an additional C-mannosylation signal. Whether this is encoded in the primary or tertiary structure is presently unknown. Finally, all modified Trp residues are part of the highly conserved core of the thrombospondin type 1 repeats present in these proteins. Since this module has been found in a large number of other proteins, the results suggest further candidates for C-mannosylation.
Notes: Hofsteenge, J
Blommers, M
Hess, D
Furmanek, A
Miroshnichenko, O
Journal Article
United States
J Biol Chem. 1999 Nov 12;274(46):32786-94. doi: 10.1074/jbc.274.46.32786.
Author Address: Friedrich Miescher-Institut, Switzerland. hofsteen@fmi.ch
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 775
Author: Hofsteenge, J., Huwiler, K. G., Macek, B., Hess, D., Lawler, J., Mosher, D. F. and Peter-Katalinic, J.
Year: 2001
Title: C-mannosylation and O-fucosylation of the thrombospondin type 1 module
Journal: J Biol Chem
Volume: 276
Issue: 9
Pages: 6485-98
Epub Date: 2000/11/09
Date: Mar 2
Short Title: C-mannosylation and O-fucosylation of the thrombospondin type 1 module
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M008073200
Accession Number: 11067851
Keywords: Amino Acid Motifs
Amino Acid Sequence
Fucose/*metabolism
Glycosylation
Humans
Mannose/*metabolism
Molecular Sequence Data
Thrombospondins/chemistry/*metabolism
Abstract: Thrombospondin-1 (TSP-1) is a multidomain protein that has been implicated in cell adhesion, motility, and growth. Some of these functions have been localized to the three thrombospondin type 1 repeats (TSRs), modules of approximately 60 amino acids in length with conserved Cys and Trp residues. The Trp residues occur in WXXW patterns, which are the recognition motifs for protein C-mannosylation. This modification involves the attachment of an alpha-mannosyl residue to the C-2 atom of the first tryptophan. Analysis of human platelet TSP-1 revealed that Trp-368, -420, -423, and -480 are C-mannosylated. Mannosylation also occurred in recombinant, baculovirally expressed TSR modules from Sf9 and "High Five" cells, contradictory to earlier reports that such cells do not carry out this reaction. In the course of these studies it was appreciated that the TSRs in TSP-1 undergo a second form of unusual glycosylation. By using a novel mass spectrometric approach, it was found that Ser-377, Thr-432, and Thr-489 in the motif CSX(S/T)CG carry the O-linked disaccharide Glc-Fuc-O-Ser/Thr. This is the first protein in which such a disaccharide has been identified, although protein O-fucosylation is well described in epidermal growth factor-like modules. Both C- and O-glycosylations take place on residues that have been implicated in the interaction of TSP-1 with glycosaminoglycans or other cellular receptors.
Notes: Hofsteenge, J
Huwiler, K G
Macek, B
Hess, D
Lawler, J
Mosher, D F
Peter-Katalinic, J
HL28749/HL/NHLBI NIH HHS/United States
HL54462/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2001 Mar 2;276(9):6485-98. doi: 10.1074/jbc.M008073200. Epub 2000 Nov 6.
Author Address: Friedrich Miescher-Institut, P. O. Box 2543, CH-4002 Basel, Switzerland. hofsteen@fmi.ch
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 547
Author: Hofsteenge, J., Müller, D. R., de Beer, T., Löffler, A., Richter, W. J. and Vliegenthart, J. F.
Year: 1994
Title: New type of linkage between a carbohydrate and a protein: C-glycosylation of a specific tryptophan residue in human RNase Us
Journal: Biochemistry
Volume: 33
Issue: 46
Pages: 13524-30
Epub Date: 1994/11/22
Date: Nov 22
Short Title: New type of linkage between a carbohydrate and a protein: C-glycosylation of a specific tryptophan residue in human RNase Us
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00250a003
Accession Number: 7947762
Keywords: Amino Acid Sequence
Endoribonucleases/*chemistry
Glycosylation
Hexoses/*chemistry
Humans
Indoles/chemistry
Magnetic Resonance Spectroscopy
Mass Spectrometry
Molecular Sequence Data
Tryptophan/*chemistry
Abstract: We report a new type of linkage between a carbohydrate and a protein, involving the rarely modified side chain of a tryptophan residue. An aldohexopyranosyl residue was found to be linked via a C-C bond to the indole ring of the tryptophan residue at position 7 of human RNase Us. Mass spectrometric analysis of peptides containing this residue showed a molecular mass 162 Da higher than that expected for tryptophan. The fragmentation pattern of the modified amino acid side chain was reminiscent of that of aromatic C-glycosides, suggesting a direct attachment of a hexose residue to a C-position of the tryptophan indole moiety. 1H and 13C NMR spectroscopic data confirmed this inference and unequivocally demonstrated the substituent to be an aldohexopyranosyl residue, C-glycosidically linked to the C2 atom of the indole. This mode of attachment differs from the ones known so far, in which carbohydrates are linked to an amino acid side chain by N- or O-glycosidic bonds.
Notes: Hofsteenge, J
Müller, D R
de Beer, T
Löffler, A
Richter, W J
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1994 Nov 22;33(46):13524-30. doi: 10.1021/bi00250a003.
Author Address: Friedrich Miescher-Institut, Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 645
Author: Hohenester, E., Maurer, P. and Timpl, R.
Year: 1997
Title: Crystal structure of a pair of follistatin-like and EF-hand calcium-binding domains in BM-40
Journal: Embo j
Volume: 16
Issue: 13
Pages: 3778-86
Epub Date: 1997/07/01
Date: Jul 1
Short Title: Crystal structure of a pair of follistatin-like and EF-hand calcium-binding domains in BM-40
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/16.13.3778
PMCID: PMC1170001
Accession Number: 9233787
Keywords: Agrin/chemistry
Amino Acid Sequence
Animals
Binding Sites
Calcium/*metabolism
Crystallography, X-Ray
Epidermal Growth Factor/chemistry
Factor IX/chemistry
Follistatin
Glycoproteins/*chemistry/metabolism
Humans
Models, Molecular
Molecular Sequence Data
Osteonectin/*chemistry/metabolism
Ovomucin/chemistry
*Protein Structure, Secondary
Rats
Sequence Homology, Amino Acid
Structure-Activity Relationship
Trypsin Inhibitor, Kazal Pancreatic/chemistry
Abstract: BM-40 (also known as SPARC or osteonectin) is an anti-adhesive secreted glycoprotein involved in tissue remodelling. Apart from an acidic N-terminal segment, BM-40 consists of a follistatin-like (FS) domain and an EF-hand calcium-binding (EC) domain. Here we report the crystal structure at 3.1 A resolution of the FS-EC domain pair of human BM-40. The two distinct domains interact through a small interface that involves the EF-hand pair of the EC domain. Residues implicated in cell binding, inhibition of cell spreading and disassembly of focal adhesions cluster on one face of BM-40, opposite the binding epitope for collagens and the N-linked carbohydrate. The elongated FS domain is structurally related to serine protease inhibitors of the Kazal family. Notable differences are an insertion into the inhibitory loop in BM-40 and a protruding N-terminal beta-hairpin with striking similarities to epidermal growth factor. This hairpin is likely to act as a rigid spacer in proteins containing tandemly repeated FS domains, such as follistatin and agrin, and forms the heparin-binding site in follistatin.
Notes: 1460-2075
Hohenester, E
Maurer, P
Timpl, R
Journal Article
EMBO J. 1997 Jul 1;16(13):3778-86. doi: 10.1093/emboj/16.13.3778.
Author Address: Department of Crystallography, Birkbeck College, London, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 749
Author: Hoja-Lukowicz, D., Ciołczyk, D., Bergquist, J., Lityńska, A. and Laidler, P.
Year: 2000
Title: High-mannose-type oligosaccharides from human placental arylsulfatase A are core fucosylated as confirmed by MALDI MS
Journal: Glycobiology
Volume: 10
Issue: 6
Pages: 551-7
Epub Date: 2000/05/18
Date: Jun
Short Title: High-mannose-type oligosaccharides from human placental arylsulfatase A are core fucosylated as confirmed by MALDI MS
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/10.6.551
Accession Number: 10814696
Keywords: Amidohydrolases/metabolism
Blotting, Western
Cerebroside-Sulfatase/*chemistry
Electrophoresis, Polyacrylamide Gel
Female
Fucose/*analysis/metabolism
Glycosylation
Humans
Mannose/*analysis
Oligosaccharides/*analysis/metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Placenta/*enzymology
Polysaccharides/analysis/metabolism
Pregnancy
*Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Despite numerous studies on arylsulfatase A, the structure of its glycans is not well understood. It has been shown that the concentration of arylsulfatase A increases in the body fluids of patients with some forms of cancer, and the carbohydrate component of arylsulfatase A synthesized in tumor tissues and transformed cells undergoes increased sialylation, phosphorylation and sulfation. To understand the significance of any changes in the glycosylation of arylsulfatase A in cancer, it is important to know the structure of its carbohydrate component in normal tissue. In the present study we have analyzed carbohydrate moieties of human placental arylsylfatase A using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by Western blotting on Immobilon P and on-blot deglycosylation using PNGase F for glycan release. Profiles of N-glycans were obtained by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Oligosaccharides were sequenced using specific exoglycosidases, and digestion products were analyzed by MALDI MS and the computer matching of the resulting masses with those derived from a sequence database. Fifty picomoles (6 microg) of arylsulfatase A applied to the gel were sufficient to characterize its oligosaccharide content. The results indicated that human placental arylsulfatase A possesses only high-mannose-type oligosaccharides, of which almost half are core fucosylated. In addition, there was a minor species of high-mannose-type glycan bearing six mannose residues with a core fucose. This structure was not expected since high-mannose-type oligosaccharides basically have not been recognized as a substrate for the alpha1,6-fucosyltransferase.
Notes: Hoja-Lukowicz, D
Ciołczyk, D
Bergquist, J
Lityńska, A
Laidler, P
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2000 Jun;10(6):551-7. doi: 10.1093/glycob/10.6.551.
Author Address: Department of Animal Physiology, Institute of Zoology, Jagiellonian University, Kraków, Poland
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 531
Author: Hokke, C. H., Bergwerff, A. A., Van Dedem, G. W., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1995
Title: Structural analysis of the sialylated N- and O-linked carbohydrate chains of recombinant human erythropoietin expressed in Chinese hamster ovary cells. Sialylation patterns and branch location of dimeric N-acetyllactosamine units
Journal: Eur J Biochem
Volume: 228
Issue: 3
Pages: 981-1008
Epub Date: 1995/03/15
Date: Mar 15
Short Title: Structural analysis of the sialylated N- and O-linked carbohydrate chains of recombinant human erythropoietin expressed in Chinese hamster ovary cells. Sialylation patterns and branch location of dimeric N-acetyllactosamine units
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1995.tb20350.x
Accession Number: 7737204
Keywords: Amino Sugars/*chemistry
Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Cricetinae
Cricetulus
Erythropoietin/*chemistry
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
N-Acetylneuraminic Acid
Recombinant Proteins/chemistry
Sialic Acids/*chemistry
Abstract: The N-linked carbohydrate chains of recombinant human erythropoietin expressed in CHO cells were quantitatively released with peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F, separated from the remaining O-glycoprotein by gel-permeation chromatography, and subsequently fractionated via FPLC on Mono Q, HPLC on Lichrosorb-NH2 and high-pH anion-exchange chromatography on CarboPac PA1. The purified sialylated oligosaccharides were analyzed by one-dimensional and two-dimensional 500-MHz 1H-NMR spectroscopy. When necessary, oligosaccharides were treated with endo-beta-galactosidase (and N-acetyl-beta-glucosaminidase) followed by 1H-NMR analysis of the incubation products, to obtain additional structural information. Di-, tri-, tri'- and tetraantennary N-acetyllactosamine-type oligosaccharides occur which can be completely (major) or partially (minor) sialylated. Three different types of alpha 2-3-linked sialic acids are present, namely, N-acetylneuraminic acid (95%), N-glycolylneuraminic acid (2%) and N-acetyl-9-O-acetylneuraminic acid (3%). In the case of partial sialylation, a non-random distribution of the sialic acids over the branches is observed. One or two extra N-acetyllactosamine units, being exclusively located in the branches attached to the alpha 1-6-linked Man residue, can be present in completely or partially sialylated di-, tri'-, and tetraantennary oligosaccharides. Tetraantennary oligosaccharides with N-acetyllactosamine repeats could be digested quantitatively with endo-beta-galactosidase from Bacteroides fragilis, whereas under the same conditions tri' antennary oligosaccharides hardly reacted (< 15%). Using endo-beta-galactosidase from Escherichia freundii, these tri'antennary oligosaccharides could be digested more extensively (> 75%). The O-linked carbohydrate chains were released from the O-glycoprotein by alkaline borohydride treatment, and purified via FPLC on Mono Q and HPLC on Lichrosorb-NH2. Two O-glycans were found, namely, Neu5Ac alpha 2-3Gal beta 1-3GalNAc-ol and Neu5Ac alpha 2-3Gal beta 1-3(Neu5Ac alpha 2-6)GalNAc-ol.
Notes: Hokke, C H
Bergwerff, A A
Van Dedem, G W
Kamerling, J P
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1995 Mar 15;228(3):981-1008. doi: 10.1111/j.1432-1033.1995.tb20350.x.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 315
Author: Hokke, C. H., Bergwerff, A. A., van Dedem, G. W., van Oostrum, J., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1990
Title: Sialylated carbohydrate chains of recombinant human glycoproteins expressed in Chinese hamster ovary cells contain traces of N-glycolylneuraminic acid
Journal: FEBS Lett
Volume: 275
Issue: 1-2
Pages: 9-14
Epub Date: 1990/11/26
Date: Nov 26
Short Title: Sialylated carbohydrate chains of recombinant human glycoproteins expressed in Chinese hamster ovary cells contain traces of N-glycolylneuraminic acid
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(90)81427-p
Accession Number: 2124546
Keywords: Animals
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Cricetinae
Cricetulus
Erythropoietin/chemistry
Female
Glycoproteins/*chemistry
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Neuraminic Acids/*analysis
Ovary
Recombinant Proteins/*chemistry
Tissue Plasminogen Activator/chemistry
Abstract: HPLC analysis of sialic acids released from recombinant variants of human tissue plasminogen activator, human chimeric plasminogen activator, human erythropoietin, and human follitropin, expressed in Chinese hamster ovary cells, demonstrates for each glycoprotein the presence of N-acetylneuraminic acid and N-glycolylneuraminic acid in a ratio of 97:3. Structural analysis by 500 MHz1H-NMR spectroscopy, of the enzymatically released N-linked carbohydrate chains of chimeric plasminogen activator and of erythropoietin, showed that alpha 2-3 linked N-glycolylneuraminic acid can occur in different N-acetyllactosamine type antennary structures.
Notes: Hokke, C H
Bergwerff, A A
van Dedem, G W
van Oostrum, J
Kamerling, J P
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1990 Nov 26;275(1-2):9-14. doi: 10.1016/0014-5793(90)81427-p.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 293
Author: Hokke, C. H., Kamerling, J. P., van Dedem, G. W. and Vliegenthart, J. F.
Year: 1991
Title: Determination of the branch location of extra N-acetyllactosamine units in sialo N-linked tetraantennary oligosaccharides
Journal: FEBS Lett
Volume: 286
Issue: 1-2
Pages: 18-24
Epub Date: 1991/07/29
Date: Jul 29
Short Title: Determination of the branch location of extra N-acetyllactosamine units in sialo N-linked tetraantennary oligosaccharides
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(91)80931-r
Accession Number: 1907570
Keywords: Acetylglucosaminidase/metabolism
Amino Sugars/*chemistry/metabolism
Animals
Carbohydrate Sequence
Cell Line
Cricetinae
Erythropoietin/chemistry
*Glycoside Hydrolases
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/*chemistry/metabolism
beta-Galactosidase/metabolism
Abstract: An approach is presented for the determination of the branch location of 1 or 2 extra N-acetyllactosamine units in sialo N-linked carbohydrate chains from glycoproteins. Tetraantennary oligosaccharides containing extra N-acetyllactosamine units were digested with endo-beta-galactosidase, followed by treatment with N-acetyl-beta-glucosaminidase, yielding products which could be analysed by 1H-NMR spectroscopy, thereby giving conclusive data about the location of the extra units in the intact structures.
Notes: Hokke, C H
Kamerling, J P
van Dedem, G W
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1991 Jul 29;286(1-2):18-24. doi: 10.1016/0014-5793(91)80931-r.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1709
Author: Holemans, T., Sørensen, D. M., van Veen, S., Martin, S., Hermans, D., Kemmer, G. C., Van den Haute, C., Baekelandt, V., Günther Pomorski, T., Agostinis, P., Wuytack, F., Palmgren, M., Eggermont, J. and Vangheluwe, P.
Year: 2015
Title: A lipid switch unlocks Parkinson's disease-associated ATP13A2
Journal: Proc Natl Acad Sci U S A
Volume: 112
Issue: 29
Pages: 9040-5
Epub Date: 2015/07/03
Date: Jul 21
Short Title: A lipid switch unlocks Parkinson's disease-associated ATP13A2
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1508220112
PMCID: PMC4517210
Accession Number: 26134396
Keywords: Amino Acid Sequence
Cell Line
Cell Membrane/drug effects/metabolism
Cytosol/metabolism
Endosomes/drug effects/metabolism
Green Fluorescent Proteins/metabolism
HeLa Cells
Humans
Lipids/*chemistry
Lysosomes/drug effects/metabolism
Manganese/pharmacology
Mitochondria/drug effects/metabolism
Models, Biological
Models, Molecular
Molecular Sequence Data
Mutation/genetics
Parkinson Disease/*metabolism
Phosphatidic Acids/metabolism
Phosphatidylinositol Phosphates/metabolism
Phosphorylation/drug effects
Protein Binding/drug effects
Proton-Translocating ATPases/chemistry/genetics/*metabolism
Structural Homology, Protein
Zinc/pharmacology
P5-type ATPase
flippase
lysosome
mitochondria
α-synuclein
Abstract: ATP13A2 is a lysosomal P-type transport ATPase that has been implicated in Kufor-Rakeb syndrome and Parkinson's disease (PD), providing protection against α-synuclein, Mn(2+), and Zn(2+) toxicity in various model systems. So far, the molecular function and regulation of ATP13A2 remains undetermined. Here, we demonstrate that ATP13A2 contains a unique N-terminal hydrophobic extension that lies on the cytosolic membrane surface of the lysosome, where it interacts with the lysosomal signaling lipids phosphatidic acid (PA) and phosphatidylinositol(3,5)bisphosphate [PI(3,5)P2]. We further demonstrate that ATP13A2 accumulates in an inactive autophosphorylated state and that PA and PI(3,5)P2 stimulate the autophosphorylation of ATP13A2. In a cellular model of PD, only catalytically active ATP13A2 offers cellular protection against rotenone-induced mitochondrial stress, which relies on the availability of PA and PI(3,5)P2. Thus, the N-terminal binding of PA and PI(3,5)P2 emerges as a key to unlock the activity of ATP13A2, which may offer a therapeutic strategy to activate ATP13A2 and thereby reduce α-synuclein toxicity or mitochondrial stress in PD or related disorders.
Notes: 1091-6490
Holemans, Tine
Sørensen, Danny Mollerup
van Veen, Sarah
Martin, Shaun
Hermans, Diane
Kemmer, Gerdi Christine
Orcid: 0000-0002-4686-5531
Van den Haute, Chris
Baekelandt, Veerle
Günther Pomorski, Thomas
Agostinis, Patrizia
Wuytack, Frank
Palmgren, Michael
Eggermont, Jan
Vangheluwe, Peter
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2015 Jul 21;112(29):9040-5. doi: 10.1073/pnas.1508220112. Epub 2015 Jul 1.
Author Address: Laboratory of Cellular Transport Systems, Department of Cellular and Molecular Medicine, ON1 Campus Gasthuisberg, KU Leuven, B3000 Leuven, Belgium;
Laboratory of Cellular Transport Systems, Department of Cellular and Molecular Medicine, ON1 Campus Gasthuisberg, KU Leuven, B3000 Leuven, Belgium; Laboratory of Cell Death Research and Therapy, Department of Cellular and Molecular Medicine, ON1 Campus Gasthuisberg, KU Leuven, B3000 Leuven, Belgium;
Centre for Membrane Pumps in Cells and Disease-PUMPkin, Department of Plant and Environmental Sciences, University of Copenhagen, DK-1871 Frederiksberg C, Denmark;
Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, KU Leuven, B3000 Leuven, Belgium; Leuven Viral Vector Core, KU Leuven, B3000 Leuven, Belgium.
Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, KU Leuven, B3000 Leuven, Belgium;
Laboratory of Cell Death Research and Therapy, Department of Cellular and Molecular Medicine, ON1 Campus Gasthuisberg, KU Leuven, B3000 Leuven, Belgium;
Laboratory of Cellular Transport Systems, Department of Cellular and Molecular Medicine, ON1 Campus Gasthuisberg, KU Leuven, B3000 Leuven, Belgium; peter.vangheluwe@med.kuleuven.be.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 373
Author: Holt, G. D., Haltiwanger, R. S., Torres, C. R. and Hart, G. W.
Year: 1987
Title: Erythrocytes contain cytoplasmic glycoproteins. O-linked GlcNAc on Band 4.1
Journal: J Biol Chem
Volume: 262
Issue: 31
Pages: 14847-50
Epub Date: 1987/11/05
Date: Nov 5
Short Title: Erythrocytes contain cytoplasmic glycoproteins. O-linked GlcNAc on Band 4.1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3117790
Keywords: Acetylglucosamine/*analysis
Blood Proteins/isolation & purification/*metabolism
Cytoplasm/analysis
*Cytoskeletal Proteins
Erythrocytes/*analysis
Galactosyltransferases
Glucosamine/*analogs & derivatives
Glycoproteins/*blood/isolation & purification
Humans
*Membrane Proteins
Molecular Weight
*Neuropeptides
Abstract: Previously we reported that the novel protein-saccharide linkage, O-linked N-acetylglucosamine (GlcNAc), is found in abundance on proteins associated with the cytoplasmic and nucleoplasmic faces of the nuclear pore complex. Here we demonstrate that O-GlcNAc moieties are also added to human erythrocyte cytoplasmic proteins. Intact or permeabilized erythrocytes, as well as subcellular fractions, were labeled with bovine milk galactosyltransferase and UDP-[3H] galactose. The proportion of the incorporated label found on O-GlcNAc was determined by a variety of chemical and enzymatic techniques. The bulk of the O-GlcNAc residues are found in the cytoplasm of erythrocytes, the majority of which are on an as yet unidentified 65-kDa protein. In addition, we have determined that Band 4.1, a protein which serves as a bridge joining the cytoskeleton to the inner surface of the plasma membrane in erythrocytes, also contains O-GlcNAc moieties. One of the sites of O-GlcNAc addition has been localized to the last 117 amino acids of the carboxy terminus of Band 4.1.
Notes: Holt, G D
Haltiwanger, R S
Torres, C R
Hart, G W
CA-42486/CA/NCI NIH HHS/United States
GM-07445/GM/NIGMS NIH HHS/United States
HD-13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1987 Nov 5;262(31):14847-50.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 917
Author: Hong, M., Luo, S., Baumeister, P., Huang, J. M., Gogia, R. K., Li, M. and Lee, A. S.
Year: 2004
Title: Underglycosylation of ATF6 as a novel sensing mechanism for activation of the unfolded protein response
Journal: J Biol Chem
Volume: 279
Issue: 12
Pages: 11354-63
Epub Date: 2003/12/31
Date: Mar 19
Short Title: Underglycosylation of ATF6 as a novel sensing mechanism for activation of the unfolded protein response
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M309804200
Accession Number: 14699159
Keywords: Activating Transcription Factor 6
Animals
Base Sequence
Calcium/metabolism
Cell Line
DNA Primers
DNA-Binding Proteins/genetics/*metabolism
Endoplasmic Reticulum/metabolism
Fluorescent Antibody Technique
Glycosylation
Homeostasis
Humans
Mutation
Protein Denaturation
Reverse Transcriptase Polymerase Chain Reaction
Transcription Factors/genetics/*metabolism
Abstract: ATF6 is a key transcriptional activator of the unfolded protein response (UPR), which allows mammalian cells to maintain cellular homeostasis when they are subjected to a variety of environmental and physiological stresses that target the endoplasmic reticulum (ER). ATF6, a 90-kDa ER transmembrane protein, contains three evolutionarily conserved N-linked glycosylation sites within its carboxyl luminal domain. Although it is well established that p90ATF6 activation requires transit from the ER to the Golgi, where it is cleaved by the S1P/S2P protease system to generate a nuclear form p60ATF6 that acts as a transcriptional activator, the functional significance of p90ATF6 N-linked glycosylation is unknown. Here we show that ER Ca(2+) depletion stress, a triggering mechanism for the UPR, induces the formation of ATF6(f), which represents de novo partial glycosylation of newly synthesized p90ATF6. By mutating a single amino acid within the N-linked glycosylation site closest to the carboxyl terminus of p90ATF6, we recreated ATF6(f). This mutation sharply reduces p90ATF6 association with calreticulin, a major Ca(2+)-binding chaperone for N-glycoprotein. We further determined that ATF6(f) exhibits a faster rate of constitutive transport to the Golgi, resulting in a higher level of p60ATF6 in the nucleus and stronger transactivating activity in the absence of ER stress. Additional analysis of p90ATF6 mutants targeting single or multiple N-glycosylation sites also showed higher constitutive transactivating activity than wild type ATF6. Because accumulation of underglycosylated proteins in the ER is a potent inducer for the UPR, these studies uncover a novel mechanism whereby the glycosylation status of p90ATF6 can serve as a sensor for ER homeostasis, resulting in ATF6 activation to trigger the UPR.
Notes: Hong, Min
Luo, Shengzhan
Baumeister, Peter
Huang, Jen-Ming
Gogia, Raveen K
Li, Mingqing
Lee, Amy S
CA27607/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2004 Mar 19;279(12):11354-63. doi: 10.1074/jbc.M309804200. Epub 2003 Dec 29.
Author Address: Department of Biochemistry and Molecular Biology and the University of Southern California/Norris Comprehensive Cancer Center, Keck School of Medicine of the University of Southern California, Los Angeles, California 90089-9176, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1744
Author: Hong, Q., Ruhaak, L. R., Stroble, C., Parker, E., Huang, J., Maverakis, E. and Lebrilla, C. B.
Year: 2015
Title: A Method for Comprehensive Glycosite-Mapping and Direct Quantitation of Serum Glycoproteins
Journal: J Proteome Res
Volume: 14
Issue: 12
Pages: 5179-92
Epub Date: 2015/10/30
Date: Dec 4
Short Title: A Method for Comprehensive Glycosite-Mapping and Direct Quantitation of Serum Glycoproteins
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/acs.jproteome.5b00756
PMCID: PMC4670571
NIHMSID: NIHMS734402
Accession Number: 26510530
Keywords: Amino Acid Sequence
Chromatography, Liquid/methods
Glycopeptides/chemistry
Glycoproteins/*blood/chemistry
Glycosylation
Humans
Immunoglobulins/blood/chemistry
Mass Spectrometry/methods
Molecular Sequence Data
Peptide Mapping/*methods
Reproducibility of Results
alpha-Macroglobulins/chemistry
absolute quantitation
glycopeptide
immunoglobulin
site specific glycan analysis
Abstract: A comprehensive glycan map was constructed for the top eight abundant glycoproteins in plasma using both specific and nonspecific enzyme digestions followed by nano liquid chromatography (LC)-chip/quadrupole time-of-flight mass spectrometry (MS) analysis. Glycopeptides were identified using an in-house software tool, GPFinder. A sensitive and reproducible multiple reaction monitoring (MRM) technique on a triple quadrupole MS was developed and applied to quantify immunoglobulins G, A, M, and their site-specific glycans simultaneously and directly from human serum/plasma without protein enrichments. A total of 64 glycopeptides and 15 peptides were monitored for IgG, IgA, and IgM in a 20 min ultra high performance (UP)LC gradient. The absolute protein contents were quantified using peptide calibration curves. The glycopeptide ion abundances were normalized to the respective protein abundances to separate protein glycosylation from protein expression. This technique yields higher method reproducibility and less sample loss when compared with the quantitation method that involves protein enrichments. The absolute protein quantitation has a wide linear range (3-4 orders of magnitude) and low limit of quantitation (femtomole level). This rapid and robust quantitation technique, which provides quantitative information for both proteins and glycosylation, will further facilitate disease biomarker discoveries.
Notes: 1535-3907
Hong, Qiuting
Ruhaak, L Renee
Stroble, Carol
Parker, Evan
Huang, Jincui
Maverakis, Emanual
Lebrilla, Carlito B
R01 CA136647/CA/NCI NIH HHS/United States
R01AT008759/AT/NCCIH NIH HHS/United States
R01HD061923/HD/NICHD NIH HHS/United States
R01 HD061923/HD/NICHD NIH HHS/United States
R01 GM049077/GM/NIGMS NIH HHS/United States
1DP2OD008752/OD/NIH HHS/United States
R01GM049077/GM/NIGMS NIH HHS/United States
DP2 OD008752/OD/NIH HHS/United States
R01 AT008759/AT/NCCIH NIH HHS/United States
R01 AT007079/AT/NCCIH NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Proteome Res. 2015 Dec 4;14(12):5179-92. doi: 10.1021/acs.jproteome.5b00756. Epub 2015 Nov 9.
Author Address: Department of Chemistry and ‡Department of Dermatology, School of Medicine, University of California , Davis, California 95616, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1727
Author: Hong, S. Y. and Hagen, T.
Year: 2015
Title: 2-Deoxyglucose induces the expression of thioredoxin interacting protein (TXNIP) by increasing O-GlcNAcylation - Implications for targeting the Warburg effect in cancer cells
Journal: Biochem Biophys Res Commun
Volume: 465
Issue: 4
Pages: 838-44
Epub Date: 2015/09/01
Date: Oct 2
Short Title: 2-Deoxyglucose induces the expression of thioredoxin interacting protein (TXNIP) by increasing O-GlcNAcylation - Implications for targeting the Warburg effect in cancer cells
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2015.08.097
Accession Number: 26315267
Keywords: Adenosine Triphosphate/metabolism
Animals
Carrier Proteins/chemistry/genetics/*metabolism
Cell Line, Tumor
Deoxyglucose/*pharmacology
Dogs
Endoplasmic Reticulum Stress
Glycolysis/*drug effects
Glycosylation/drug effects
HEK293 Cells
Hep G2 Cells
Humans
Madin Darby Canine Kidney Cells
Mice
Neoplasm Proteins/chemistry/genetics/*metabolism
Neoplasms/drug therapy/genetics/metabolism
Thioredoxins/chemistry/genetics/*metabolism
Tumor Microenvironment/drug effects/genetics/physiology
2-Deoxyglucose
ChoRE
MondoA
O-GlcNAcylation
Txnip
Abstract: The high proliferation rate of cancer cells and the microenvironment in the tumor tissue require the reprogramming of tumor cell metabolism. The major mechanism of metabolic reprogramming in cancer cells is the Warburg effect, defined as the preferential utilization of glucose via glycolysis even in the presence of oxygen. Targeting the Warburg effect is considered as a promising therapeutic strategy in cancer therapy. In this regard, the glycolytic inhibitor 2-deoxyglucose (2DG) has been evaluated clinically. 2DG exerts its effect by directly inhibiting glycolysis at the level of hexokinase and phosphoglucoisomerase. In addition, 2DG is also known to induce the expression of thioredoxin interacting protein (TXNIP), a tumor suppressor protein and an important negative regulator of cellular glucose uptake. Hence, characterization of the mechanism through which 2DG regulates TXNIP expression may reveal novel approaches to target the Warburg effect in cancer cells. Therefore, in this study we sought to test various hypotheses for the mechanistic basis of the 2DG dependent TXNIP regulation. We have shown that 2DG induced TXNIP expression is independent of carbohydrate response element mediated transcription. Furthermore, the induction of TXNIP is neither dependent on the ability of 2DG to deplete cellular ATP nor to cause endoplasmic reticulum stress. We found that the 2DG induced TXNIP expression is at least in part dependent on the inhibition of the O-GlcNAcase enzyme and the accumulation of O-GlcNAc modified proteins. These results have implications for the identification of therapeutic targets to increase TXNIP expression in cancer.
Notes: 1090-2104
Hong, Shin Yee
Hagen, Thilo
Journal Article
United States
Biochem Biophys Res Commun. 2015 Oct 2;465(4):838-44. doi: 10.1016/j.bbrc.2015.08.097. Epub 2015 Aug 24.
Author Address: Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, 117597 Singapore, Singapore.
Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, 117597 Singapore, Singapore. Electronic address: bchth@nus.edu.sg.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1939
Author: Horowitz, B., Javitt, G., Ilani, T., Gat, Y., Morgenstern, D., Bard, F. A. and Fass, D.
Year: 2018
Title: Quiescin sulfhydryl oxidase 1 (QSOX1) glycosite mutation perturbs secretion but not Golgi localization
Journal: Glycobiology
Volume: 28
Issue: 8
Pages: 580-591
Epub Date: 2018/05/15
Date: Aug 1
Short Title: Quiescin sulfhydryl oxidase 1 (QSOX1) glycosite mutation perturbs secretion but not Golgi localization
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwy044
Accession Number: 29757379
Keywords: Cell Line
Fibroblasts/cytology/*metabolism
Glycosylation
Golgi Apparatus/genetics/*metabolism
Humans
Oxidoreductases Acting on Sulfur Group Donors/genetics/*metabolism
Protein Transport/physiology
Abstract: Quiescin sulfhydryl oxidase 1 (QSOX1) catalyzes the formation of disulfide bonds in protein substrates. Unlike other enzymes with related activities, which are commonly found in the endoplasmic reticulum, QSOX1 is localized to the Golgi apparatus or secreted. QSOX1 is upregulated in quiescent fibroblast cells and secreted into the extracellular environment, where it contributes to extracellular matrix assembly. QSOX1 is also upregulated in adenocarcinomas, though the extent to which it is secreted in this context is currently unknown. To achieve a better understanding of factors that dictate QSOX1 localization and function, we aimed to determine how post-translational modifications affect QSOX1 trafficking and activity. We found a highly conserved N-linked glycosylation site to be required for QSOX1 secretion from fibroblasts and other cell types. Notably, QSOX1 lacking a glycan at this site arrives at the Golgi, suggesting that it passes endoplasmic reticulum quality control but is not further transported to the cell surface for secretion. The QSOX1 transmembrane segment is dispensable for Golgi localization and secretion, as fully luminal and transmembrane variants displayed the same trafficking behavior. This study provides a key example of the effect of glycosylation on Golgi exit and contributes to an understanding of late secretory sorting and quality control.
Notes: 1460-2423
Horowitz, Ben
Javitt, Gabriel
Ilani, Tal
Gat, Yair
Morgenstern, David
Bard, Frederic A
Fass, Deborah
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2018 Aug 1;28(8):580-591. doi: 10.1093/glycob/cwy044.
Author Address: Department of Structural Biology, Weizmann Institute of Science, Rehovot, Israel.
The Nancy and Stephen Grand Israel National Center for Personalized Medicine, Weizmann Institute of Science, Rehovot, Israel.
Institute of Molecular and Cell Biology, Agency for Science, Technology and Research, 61 Biopolis Drive, Proteos, Singapore.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 380
Author: Hostikka, S. L. and Tryggvason, K.
Year: 1988
Title: The complete primary structure of the alpha 2 chain of human type IV collagen and comparison with the alpha 1(IV) chain
Journal: J Biol Chem
Volume: 263
Issue: 36
Pages: 19488-93
Epub Date: 1988/12/25
Date: Dec 25
Short Title: The complete primary structure of the alpha 2 chain of human type IV collagen and comparison with the alpha 1(IV) chain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3198637
Keywords: Amino Acid Sequence
Base Sequence
Cloning, Molecular
Collagen/*genetics
DNA/genetics
Female
Humans
Macromolecular Substances
Molecular Sequence Data
Placenta/metabolism
Pregnancy
Restriction Mapping
Sequence Homology, Nucleic Acid
Abstract: The complete primary structure of the human type IV collagen alpha 2(IV) chain has been determined by nucleotide sequencing of cDNA clones. The overlapping cDNA clones cover 6,257 base pairs with a 5'-untranslated region of 283 base pairs, the 5,136-base pair open reading frame, and the 3'-untranslated region of 838 base pairs. The predicted amino acid sequence demonstrates that the complete translation product consists of 1,712 residues corresponding in molecular weight to 167,560. The translated polypeptide has a signal peptide of 36 amino acids, an amino-terminal noncollagenous part of 21 residues, a 1,428-residue collagenous domain with 23 interruptions, and a carboxyl-terminal noncollagenous (NC) domain of 227 residues. The calculated molecular mass of the mature human alpha 2(IV) chain is 163,774 Da.
Notes: Hostikka, S L
Tryggvason, K
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1988 Dec 25;263(36):19488-93.
Author Address: Department of Biochemistry, University of Oulu, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1732
Author: Hou, C. W., Mohanan, V., Zachara, N. E. and Grimes, C. L.
Year: 2016
Title: Identification and biological consequences of the O-GlcNAc modification of the human innate immune receptor, Nod2
Journal: Glycobiology
Volume: 26
Issue: 1
Pages: 13-8
Epub Date: 2015/09/16
Date: Jan
Short Title: Identification and biological consequences of the O-GlcNAc modification of the human innate immune receptor, Nod2
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwv076
PMCID: PMC4672147
Accession Number: 26369908
Keywords: Acetylglucosamine/*metabolism
Glycosylation
HCT116 Cells
HEK293 Cells
Humans
Mutation
NF-kappa B/metabolism
Nod2 Signaling Adaptor Protein/genetics/*metabolism
*Protein Processing, Post-Translational
Crohn's disease
Nf-κb
Nod2
O-GlcNAcylation
Ogt
Abstract: Nucleotide-binding oligomerization domain 2 (Nod2) is an intracellular receptor that can sense the bacterial peptidoglycan component, muramyl dipeptide. Upon activation, Nod2 induces the production of various inflammatory molecules such as cytokines and chemokines. Genetic linkage analysis identified and revealed three major mutations in Nod2 that are associated with the development of Crohn's disease. The objective of this study is to further characterize this protein by determining whether Nod2 is posttranslationally modified by O-N-acetylglucosamine (O-GlcNAc). O-GlcNAcylation is one type of posttranslational modification in which the O-GlcNAc transferase transfers GlcNAc from UDP-GlcNAc to selected serine and threonine residues of intracellular proteins. We found that wild-type Nod2 and a Nod2 Crohn's-associated variant are O-GlcNAcylated and this modification affects Nod2's ability to signal via the nuclear factor kappa B pathway.
Notes: 1460-2423
Hou, Ching-Wen
Mohanan, Vishnu
Zachara, Natasha E
Grimes, Catherine Leimkuhler
P01 HL107153/HL/NHLBI NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
P01HL107153/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Glycobiology. 2016 Jan;26(1):13-8. doi: 10.1093/glycob/cwv076. Epub 2015 Sep 14.
Author Address: Department of Chemistry and Biochemistry.
Department of Biological Sciences, University of Delaware, Newark, DE 19716, USA.
Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Department of Chemistry and Biochemistry Department of Biological Sciences, University of Delaware, Newark, DE 19716, USA cgrimes@udel.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1163
Author: Housley, M. P., Rodgers, J. T., Udeshi, N. D., Kelly, T. J., Shabanowitz, J., Hunt, D. F., Puigserver, P. and Hart, G. W.
Year: 2008
Title: O-GlcNAc regulates FoxO activation in response to glucose
Journal: J Biol Chem
Volume: 283
Issue: 24
Pages: 16283-92
Epub Date: 2008/04/19
Date: Jun 13
Short Title: O-GlcNAc regulates FoxO activation in response to glucose
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M802240200
PMCID: PMC2423255
Accession Number: 18420577
Keywords: Acetylglucosamine/*pharmacology
Animals
Catalase/metabolism
Diabetes Mellitus, Experimental/metabolism
Forkhead Box Protein O1
Forkhead Transcription Factors/*metabolism
*Gene Expression Regulation
Glucose/*metabolism
Glucose-6-Phosphatase/metabolism
Humans
Models, Biological
Nerve Tissue Proteins/*metabolism
Phosphoenolpyruvate Carboxykinase (ATP)/metabolism
Rats
Reactive Oxygen Species
Superoxide Dismutase/metabolism
Abstract: FoxO proteins are key transcriptional regulators of nutrient homeostasis and stress response. The transcription factor FoxO1 activates expression of gluconeogenic, including phosphoenolpyruvate carboxykinase and glucose-6-phosphatase, and also activates the expression of the oxidative stress response enzymes catalase and manganese superoxide dismutase. Hormonal and stress-dependent regulation of FoxO1 via acetylation, ubiquitination, and phosphorylation, are well established, but FoxOs have not been studied in the context of the glucose-derived O-linked beta-N-acetylglucosamine (O-GlcNAc) modification. Here we show that O-GlcNAc on hepatic FoxO1 is increased in diabetes. Furthermore, O-GlcNAc regulates FoxO1 activation in response to glucose, resulting in the paradoxically increased expression of gluconeogenic genes while concomitantly inducing expression of genes encoding enzymes that detoxify reactive oxygen species. GlcNAcylation of FoxO provides a new mechanism for direct nutrient control of transcription to regulate metabolism and stress response through control of FoxO1 activity.
Notes: 1083-351x
Housley, Michael P
Rodgers, Joseph T
Udeshi, Namrata D
Kelly, Timothy J
Shabanowitz, Jeffrey
Hunt, Donald F
Puigserver, Pere
Hart, Gerald W
DK61671/DK/NIDDK NIH HHS/United States
GM37537/GM/NIGMS NIH HHS/United States
R37 HD013563/HD/NICHD NIH HHS/United States
HD13563/HD/NICHD NIH HHS/United States
R01 GM037537/GM/NIGMS NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2008 Jun 13;283(24):16283-92. doi: 10.1074/jbc.M802240200. Epub 2008 Apr 17.
Author Address: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 884
Author: Houston, D. R., Recklies, A. D., Krupa, J. C. and van Aalten, D. M.
Year: 2003
Title: Structure and ligand-induced conformational change of the 39-kDa glycoprotein from human articular chondrocytes
Journal: J Biol Chem
Volume: 278
Issue: 32
Pages: 30206-12
Epub Date: 2003/05/31
Date: Aug 8
Short Title: Structure and ligand-induced conformational change of the 39-kDa glycoprotein from human articular chondrocytes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M303371200
Accession Number: 12775711
Keywords: Adipokines
Chitinase-3-Like Protein 1
Chondrocytes/*metabolism
Crystallography, X-Ray
Electrons
Glycoproteins
Humans
Lectins/*chemistry/*metabolism
*Ligands
Models, Molecular
Oligosaccharides/chemistry
Protein Binding
Protein Conformation
Signal Transduction
Abstract: The 39-kDa human cartilage glycoprotein (HCGP39), a member of a novel family of chitinase-like lectins (Chilectins), is overexpressed in articular chondrocytes and certain cancers. Proposed functions of this protein include a role in connective tissue remodeling and defense against pathogens. Similar to other Chi-lectins, HCGP39 promotes the growth of connective tissue cells. The ability of HCGP39 to activate cytoplasmic signaling pathways suggests the presence of a ligand for this protein at the cell surface. There is currently no information regarding the identity of any physiological or pathological ligands of the Chi-lectins or the nature of the protein-ligand interaction. Here, we show that HCGP39 is able to bind chitooligosaccharides with micromolar affinity. Crystal structures of the native protein and a complex with GlcNAc8 show that the ligand is bound in identical fashion to family 18 chitinases. However, unlike the chitinases, binding of the oligosaccharide ligand to HCGP39 induces a large conformational change. Thus, HCGP39 could be a lectin that binds chitin-like oligosaccharide ligands and possibly plays a role in innate responses to chitinous pathogens, such as fungi and nematodes.
Notes: Houston, Douglas R
Recklies, Anneliese D
Krupa, Joanne C
van Aalten, Daan M F
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2003 Aug 8;278(32):30206-12. doi: 10.1074/jbc.M303371200. Epub 2003 May 29.
Author Address: Division of Biological Chemistry & Molecular Microbiology, School of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2046
Author: Hsieh, Y. L., Su, F. Y., Tsai, L. K., Huang, C. C., Ko, Y. L., Su, L. W., Chen, K. Y., Shih, H. M., Hu, C. M. and Lee, W. H.
Year: 2019
Title: NPGPx-Mediated Adaptation to Oxidative Stress Protects Motor Neurons from Degeneration in Aging by Directly Modulating O-GlcNAcase
Journal: Cell Rep
Volume: 29
Issue: 8
Pages: 2134-2143.e7
Epub Date: 2019/11/21
Date: Nov 19
Short Title: NPGPx-Mediated Adaptation to Oxidative Stress Protects Motor Neurons from Degeneration in Aging by Directly Modulating O-GlcNAcase
Alternate Journal: Cell reports
DOI: 10.1016/j.celrep.2019.10.053
Accession Number: 31747588
Keywords: Aging/physiology
Amyotrophic Lateral Sclerosis/metabolism
Animals
Female
Humans
Mice
Mice, Mutant Strains
Motor Neurons/*metabolism
Muscle Denervation
Oxidative Stress/genetics/*physiology
Paralysis/metabolism
beta-N-Acetylhexosaminidases/*metabolism
Als
NPGPx
O-GlcNAcylation
Oga
aging
motor neuron
oxidative stress
Abstract: Amyotrophic lateral sclerosis (ALS), the most common motor neuron disease, usually occurs in middle-aged people. However, the molecular basis of age-related cumulative stress in ALS pathogenesis remains elusive. Here, we found that mice deficient in NPGPx (GPx7), an oxidative stress sensor, develop ALS-like phenotypes, including paralysis, muscle denervation, and motor neurons loss. Unlike normal spinal motor neurons that exhibit elevated O-GlcNAcylation against age-dependent oxidative stress, NPGPx-deficient spinal motor neurons fail to boost O-GlcNAcylation and exacerbate ROS accumulation, leading to cell death. Mechanistically, stress-activated NPGPx inhibits O-GlcNAcase (OGA) through disulfide bonding to fine-tune global O-GlcNAcylation. Pharmacological inhibition of OGA rescues spinal motor neuron loss in aged NPGPx-deficient mice. Furthermore, expression of NPGPx in ALS patients is significantly lower than in unaffected adults. These results suggest that NPGPx modulates O-GlcNAcylation by inhibiting OGA to cope with age-dependent oxidative stress and protect motor neurons from degeneration, providing a potential therapeutic axis for ALS.
Notes: 2211-1247
Hsieh, Yung-Lin
Su, Fang-Yi
Tsai, Li-Kai
Huang, Chien-Chang
Ko, Yi-Ling
Su, Li-Wen
Chen, Kai-Yun
Shih, Hsiu-Ming
Hu, Chun-Mei
Lee, Wen-Hwa
Journal Article
United States
Cell Rep. 2019 Nov 19;29(8):2134-2143.e7. doi: 10.1016/j.celrep.2019.10.053.
Author Address: Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan; Institute of Biochemistry and Molecular Biology, National Yang-Ming University, Taipei 112, Taiwan.
Department of Neurology, National Taiwan University Hospital, Taipei 100, Taiwan.
Graduate Institute of Neural Regenerative Medicine, College of Medical Science and Technology, Taipei Medical University, Taipei 110, Taiwan.
Institute of Biomedical Sciences, Academia Sinica, Taipei 115, Taiwan; Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli 350, Taiwan.
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan; Department of Biological Chemistry, University of California, Irvine, Irvine, CA 92697, USA; Drug Development Center, China Medical University, Taichung 404, Taiwan. Electronic address: whlee@uci.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2012
Author: Hu, C. M., Tien, S. C., Hsieh, P. K., Jeng, Y. M., Chang, M. C., Chang, Y. T., Chen, Y. J., Chen, Y. J., Lee, E. Y. P. and Lee, W. H.
Year: 2019
Title: High Glucose Triggers Nucleotide Imbalance through O-GlcNAcylation of Key Enzymes and Induces KRAS Mutation in Pancreatic Cells
Journal: Cell Metab
Volume: 29
Issue: 6
Pages: 1334-1349.e10
Epub Date: 2019/03/12
Date: Jun 4
Short Title: High Glucose Triggers Nucleotide Imbalance through O-GlcNAcylation of Key Enzymes and Induces KRAS Mutation in Pancreatic Cells
Alternate Journal: Cell metabolism
ISSN: 1550-4131
DOI: 10.1016/j.cmet.2019.02.005
Accession Number: 30853214
Keywords: Acetylation/drug effects
Acetylglucosamine/*metabolism
Acetyltransferases/metabolism
Adult
Aged
Animals
Carcinoma, Pancreatic Ductal/chemically induced/genetics/metabolism/pathology
Cell Transformation, Neoplastic/*chemically induced/genetics/metabolism
Cells, Cultured
DNA Damage/genetics
Dose-Response Relationship, Drug
Enzymes/genetics/*metabolism
Female
Glucose/adverse effects/*pharmacology
HEK293 Cells
Humans
Infant, Newborn
Male
Metabolic Networks and Pathways/drug effects/genetics
Mice
Mice, Inbred C57BL
Middle Aged
Mutagenesis/drug effects
Mutation/drug effects
Nucleotides/*metabolism
Pancreas/*drug effects/metabolism
Pancreatic Neoplasms/chemically induced/genetics/metabolism/pathology
Protein Processing, Post-Translational/drug effects
Proto-Oncogene Proteins p21(ras)/*genetics/metabolism
Young Adult
*KRAS mutation
*O-GlcNAcylation
*Pfk
*Rnr
*Rrm1
*high glucose
*nucleotide imbalance
Abstract: KRAS mutations are the earliest events found in approximately 90% of pancreatic ductal adenocarcinomas (PDACs). However, little is known as to why KRAS mutations preferentially occur in PDACs and what processes/factors generate these mutations. While abnormal carbohydrate metabolism is associated with a high risk of pancreatic cancer, it remains elusive whether a direct relationship between KRAS mutations and sugar metabolism exists. Here, we show that under high-glucose conditions, cellular O-GlcNAcylation is significantly elevated in pancreatic cells that exhibit lower phosphofructokinase (PFK) activity than other cell types. This post-translational modification specifically compromises the ribonucleotide reductase (RNR) activity, leading to deficiency in dNTP pools, genomic DNA alterations with KRAS mutations, and cellular transformation. These results establish a mechanistic link between a perturbed sugar metabolism and genomic instability that induces de novo oncogenic KRAS mutations preferentially in pancreatic cells.
Notes: 1932-7420
Hu, Chun-Mei
Tien, Sui-Chih
Hsieh, Ping-Kun
Jeng, Yung-Ming
Chang, Ming-Chu
Chang, Yu-Ting
Chen, Yi-Ju
Chen, Yu-Ju
Lee, Eva Y-H P
Lee, Wen-Hwa
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell Metab. 2019 Jun 4;29(6):1334-1349.e10. doi: 10.1016/j.cmet.2019.02.005. Epub 2019 Mar 7.
Author Address: Genomics Research Center, Academia Sinica, Taipei 11529, Taiwan. Electronic address: cmhu1220@gate.sinica.edu.tw.
Genomics Research Center, Academia Sinica, Taipei 11529, Taiwan.
Department of Pathology, National Taiwan University Hospital, Taipei 10041, Taiwan.
Department of Internal Medicine, National Taiwan University Hospital, Taipei 10041, Taiwan.
Institute of Chemistry, Academia Sinica, Taipei 11529, Taiwan.
Department of Biological Chemistry, University of California, Irvine, Irvine, CA 92697, USA.
Genomics Research Center, Academia Sinica, Taipei 11529, Taiwan; Drug Development Center, China Medical University, Taichung 40402, Taiwan. Electronic address: whlee@uci.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1937
Author: Hu, Y., Shah, P., Clark, D. J., Ao, M. and Zhang, H.
Year: 2018
Title: Reanalysis of Global Proteomic and Phosphoproteomic Data Identified a Large Number of Glycopeptides
Journal: Anal Chem
Volume: 90
Issue: 13
Pages: 8065-8071
Epub Date: 2018/05/10
Date: Jul 3
Short Title: Reanalysis of Global Proteomic and Phosphoproteomic Data Identified a Large Number of Glycopeptides
Alternate Journal: Analytical chemistry
ISSN: 0003-2700 (Print)
0003-2700
DOI: 10.1021/acs.analchem.8b01137
PMCID: PMC6440470
NIHMSID: NIHMS1014734
Accession Number: 29741879
Keywords: Animals
Cell Line, Tumor
Cell Transformation, Neoplastic
Chromatography, Affinity
Female
Glycopeptides/*metabolism
Humans
Mammary Neoplasms, Experimental/metabolism
Mice
Phosphopeptides/*metabolism
Proteomics/*methods
Abstract: Protein glycosylation plays fundamental roles in many cellular processes, and previous reports have shown dysregulation to be associated with several human diseases, including diabetes, cancer, and neurodegenerative disorders. Despite the vital role of glycosylation for proper protein function, the analysis of glycoproteins has been lagged behind to other protein modifications. In this study, we describe the reanalysis of global proteomic data from breast cancer xenograft tissues using recently developed software package GPQuest 2.0, revealing a large number of previously unidentified N-linked glycopeptides. More importantly, we found that using immobilized metal affinity chromatography (IMAC) technology for the enrichment of phosphopeptides had coenriched a substantial number of sialoglycopeptides, allowing for a large-scale analysis of sialoglycopeptides in conjunction with the analysis of phosphopeptides. Collectively, combined tandem mass spectrometry (MS/MS) analyses of global proteomic and phosphoproteomic data sets resulted in the identification of 6 724 N-linked glycopeptides from 617 glycoproteins derived from two breast cancer xenograft tissues. Next, we utilized GPQuest 2.0 for the reanalysis of global and phosphoproteomic data generated from 108 human breast cancer tissues that were previously analyzed by Clinical Proteomic Analysis Consortium (CPTAC). Reanalysis of the CPTAC data set resulted in the identification of 2 683 glycopeptides from the global proteomic data set and 4 554 glycopeptides from phosphoproteomic data set, respectively. Together, 11 292 N-linked glycopeptides corresponding to 1 731 N-linked glycosites from 883 human glycoproteins were identified from the two data sets. This analysis revealed an extensive number of glycopeptides hidden in the global and enriched in IMAC-based phosphopeptide-enriched proteomic data, information which would have remained unknown from the original study otherwise. The reanalysis described herein can be readily applied to identify glycopeptides from already existing data sets, providing insight into many important facets of protein glycosylation in different biological, physiological, and pathological processes.
Notes: 1520-6882
Hu, Yingwei
Orcid: 0000-0002-4629-0985
Shah, Punit
Clark, David J
Orcid: 0000-0003-0527-8469
Ao, Minghui
Zhang, Hui
Orcid: 0000-0001-8726-7098
U01 CA152813/CA/NCI NIH HHS/United States
U24 CA210985/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Anal Chem. 2018 Jul 3;90(13):8065-8071. doi: 10.1021/acs.analchem.8b01137. Epub 2018 Jun 11.
Author Address: Department of Pathology , Johns Hopkins University , Baltimore , Maryland 21287 , United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1030
Author: Huai, Q., Mazar, A. P., Kuo, A., Parry, G. C., Shaw, D. E., Callahan, J., Li, Y., Yuan, C., Bian, C., Chen, L., Furie, B., Furie, B. C., Cines, D. B. and Huang, M.
Year: 2006
Title: Structure of human urokinase plasminogen activator in complex with its receptor
Journal: Science
Volume: 311
Issue: 5761
Pages: 656-9
Epub Date: 2006/02/04
Date: Feb 3
Short Title: Structure of human urokinase plasminogen activator in complex with its receptor
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1121143
Accession Number: 16456079
Keywords: Antibodies/chemistry/metabolism
Crystallography, X-Ray
Humans
Hydrogen Bonding
Hydrophobic and Hydrophilic Interactions
Ligands
Models, Molecular
Peptide Fragments/chemistry/metabolism
Protein Binding
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Cell Surface/*chemistry/immunology/metabolism
Receptors, Urokinase Plasminogen Activator
Urokinase-Type Plasminogen Activator/*chemistry/metabolism
Abstract: The urokinase plasminogen activator binds to its cellular receptor with high affinity and initiates signaling cascades that are implicated in pathological processes including tumor growth, metastasis, and inflammation. We report the crystal structure at 1.9 angstroms of the urokinase receptor complexed with the urokinase amino-terminal fragment and an antibody against the receptor. The three domains of urokinase receptor form a concave shape with a central cone-shaped cavity where the urokinase fragment inserts. The structure provides insight into the flexibility of the urokinase receptor that enables its interaction with a wide variety of ligands and a basis for the design of urokinase-urokinase receptor antagonists.
Notes: 1095-9203
Huai, Qing
Mazar, Andrew P
Kuo, Alice
Parry, Graham C
Shaw, David E
Callahan, Jennifer
Li, Yongdong
Yuan, Cai
Bian, Chuanbing
Chen, Liqing
Furie, Bruce
Furie, Barbara C
Cines, Douglas B
Huang, Mingdong
R01 HL086584/HL/NHLBI NIH HHS/United States
R01 HL086584-01/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Science. 2006 Feb 3;311(5761):656-9. doi: 10.1126/science.1121143.
Author Address: Division of Hemostasis and Thrombosis, Center for Vascular Biology Research, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA 02215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1622
Author: Huang, J., Lee, H., Zivkovic, A. M., Smilowitz, J. T., Rivera, N., German, J. B. and Lebrilla, C. B.
Year: 2014
Title: Glycomic analysis of high density lipoprotein shows a highly sialylated particle
Journal: J Proteome Res
Volume: 13
Issue: 2
Pages: 681-91
Epub Date: 2014/01/15
Date: Feb 7
Short Title: Glycomic analysis of high density lipoprotein shows a highly sialylated particle
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr4012393
PMCID: PMC3975653
NIHMSID: NIHMS557294
Accession Number: 24417605
Keywords: Amino Acid Sequence
Carbohydrates/*chemistry
Chromatography, High Pressure Liquid
Gangliosides/chemistry
Lipoproteins, HDL/*chemistry
Molecular Sequence Data
N-Acetylneuraminic Acid/*chemistry
Tandem Mass Spectrometry
Abstract: Many of the functional proteins and lipids in high density lipoprotein (HDL) particles are potentially glycosylated, yet very little is known about the glycoconjugates of HDL. In this study, HDL was isolated from plasma by sequential micro-ultracentrifugation, followed by glycoprotein and glycolipid analysis. N-Glycans, glycopeptides, and gangliosides were extracted and purified followed by analysis with nano-HPLC Chip quadrupole time of flight mass spectrometry and MS/MS. HDL particles were found to be highly sialylated. Most of the N-glycans (∼90%) from HDL glycoproteins were sialylated with one or two neuraminic acids (Neu5Ac). The most abundant N-glycan was a biantennary complex type glycan with two sialic acids (Hexose5HexNAc4Neu5Ac2) and was found in multiple glycoproteins using site-specific glycosylation analysis. The observed O-glycans were all sialylated, and most contained a core 1 structure with two Neu5Acs, including those that were associated with apolipoprotein CIII (ApoC-III) and fetuin A. GM3 (monosialoganglioside, NeuAc2-3Gal1-4Glc-Cer) and GD3 (disialoganglioside, NeuAc2-8NeuAc2-3Gal1-4Glc-Cer) were the major gangliosides in HDL. A 60% GM3 and 40% GD3 distribution was observed. Both GM3 and GD3 were composed of heterogeneous ceramide lipid tails, including d18:1/16:0 and d18:1/23:0. This report describes for the first time a glycomic approach for analyzing HDL, highlighting that HDL are highly sialylated particles.
Notes: 1535-3907
Huang, Jincui
Lee, Hyeyoung
Zivkovic, Angela M
Smilowitz, Jennifer T
Rivera, Nancy
German, J Bruce
Lebrilla, Carlito B
R01 HD061923/HD/NICHD NIH HHS/United States
R01 GM049077/GM/NIGMS NIH HHS/United States
S10 RR027639/RR/NCRR NIH HHS/United States
R01GM049077/GM/NIGMS NIH HHS/United States
R01 AT007079/AT/NCCIH NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Proteome Res. 2014 Feb 7;13(2):681-91. doi: 10.1021/pr4012393. Epub 2014 Jan 21.
Author Address: Department of Chemistry, ‡Department of Food Science and Technology, and §Foods for Health Institute, University of California , Davis, California 95616, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1673
Author: Huang, L. J., Lin, J. H., Tsai, J. H., Chu, Y. Y., Chen, Y. W., Chen, S. L. and Chen, S. H.
Year: 2014
Title: Identification of protein O-glycosylation site and corresponding glycans using liquid chromatography-tandem mass spectrometry via mapping accurate mass and retention time shift
Journal: J Chromatogr A
Volume: 1371
Pages: 136-45
Epub Date: 2014/12/03
Date: Dec 5
Short Title: Identification of protein O-glycosylation site and corresponding glycans using liquid chromatography-tandem mass spectrometry via mapping accurate mass and retention time shift
Alternate Journal: Journal of chromatography. A
ISSN: 0021-9673
DOI: 10.1016/j.chroma.2014.10.046
Accession Number: 25456591
Keywords: Amino Acid Sequence
Animals
Caseins/*chemistry
Cattle
Chromatography, High Pressure Liquid/*methods
Glycopeptides/*chemistry
Glycosylation
Molecular Sequence Data
Molecular Weight
Phosphorylation
Polysaccharides/*chemistry
Tandem Mass Spectrometry/*methods
Enbrel(®)
Lc–ms
O-glycosylation
O-glycosylation site
β-Elimination
Abstract: We reported an improved combinatorial approach for identifying site-specific O-glycosylation using both glycan cleaved and non-cleaved methods. In this approach, a non-reducing β-elimination kit coupled with non-specific enzymes performed efficient digestion, O-glycan cleavage, and partial dephosphorylation without significant side reactions, thus enabling an automatic database search for the cleaved O-glycosylation or serine/threonine (S/T) phosphorylation sites. From the same sample concurrently prepared without β-elimination, the corresponding intact O-glycopeptides were mapped by accurate precursor ion mass using an in-house glycan database majorly composed of GalNAc (mucin-type) core and the retention-time shift (ΔRt). Each glycopeptide assignment was verified by the detection of glycan-specific fragments using collision-induced dissociation (CID) to estimate False Discovery Rate (FDR). Using fetuin as a model, all identified S/T elimination sites were matched to multiple intact glycopeptides with a 31% FDR. This considerably reduced to 0% FDR by ΔRt filtering. This approach was then applied to a protein mixture composed of therapeutic Factor IX and Enbrel(®) mixed with fetuin and kappa-casein. A total of 26 glycosylation sites each of which corresponds to 1-4 glycans were positively mapped and confirmed. The FDR decreased from 33% to 3.3% by ΔRt filtering and exclusion of repeated peptide tags that covered the same glycosylation sites. Moreover, the phosphorylation and O-glycosylation on the same site such as T159 of Factor IX and T170 of kappa-casein were able to be unambiguously differentiated. Thus, our approach is useful for in-depth characterization of site-specific O-glycosylation of a simple mixture such as protein-based therapeutics.
Notes: 1873-3778
Huang, Li-Juan
Lin, Jen-Hui
Tsai, Jung-Heng
Chu, Yen-Yin
Chen, Yen-Wen
Chen, Shun-Li
Chen, Shu-Hui
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Chromatogr A. 2014 Dec 5;1371:136-45. doi: 10.1016/j.chroma.2014.10.046. Epub 2014 Nov 1.
Author Address: Department of Chemistry, National Cheng Kung University, Tainan, Taiwan.
Department of Communication Engineering, National Central University, Jhongli, Taiwan.
Department of Chemistry, National Cheng Kung University, Tainan, Taiwan. Electronic address: shchen@mail.ncku.edu.tw.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1738
Author: Huang, X., Chen, H., Michelsen, K., Schneider, S. and Shaffer, P. L.
Year: 2015
Title: Crystal structure of human glycine receptor-α3 bound to antagonist strychnine
Journal: Nature
Volume: 526
Issue: 7572
Pages: 277-80
Epub Date: 2015/09/30
Date: Oct 8
Short Title: Crystal structure of human glycine receptor-α3 bound to antagonist strychnine
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature14972
Accession Number: 26416729
Keywords: Binding Sites
Crystallography, X-Ray
Humans
Ion Channel Gating/drug effects
Models, Molecular
Protein Multimerization/drug effects
Protein Structure, Tertiary/drug effects
Receptors, Glycine/antagonists & inhibitors/*chemistry/*metabolism
Strychnine/chemistry/*metabolism/pharmacology
Substrate Specificity
Abstract: Neurotransmitter-gated ion channels of the Cys-loop receptor family are essential mediators of fast neurotransmission throughout the nervous system and are implicated in many neurological disorders. Available X-ray structures of prokaryotic and eukaryotic Cys-loop receptors provide tremendous insights into the binding of agonists, the subsequent opening of the ion channel, and the mechanism of channel activation. Yet the mechanism of inactivation by antagonists remains unknown. Here we present a 3.0 Å X-ray structure of the human glycine receptor-α3 homopentamer in complex with a high affinity, high-specificity antagonist, strychnine. Our structure allows us to explore in detail the molecular recognition of antagonists. Comparisons with previous structures reveal a mechanism for antagonist-induced inactivation of Cys-loop receptors, involving an expansion of the orthosteric binding site in the extracellular domain that is coupled to closure of the ion pore in the transmembrane domain.
Notes: 1476-4687
Huang, Xin
Chen, Hao
Michelsen, Klaus
Schneider, Stephen
Shaffer, Paul L
Journal Article
England
Nature. 2015 Oct 8;526(7572):277-80. doi: 10.1038/nature14972. Epub 2015 Sep 28.
Author Address: Department of Molecular Structure and Characterization, Amgen Inc., 360 Binney Street, Cambridge, Massachusetts 02142, USA.
Department of Protein Technologies, Amgen Inc., 360 Binney Street, Cambridge, Massachusetts 02142, USA.
Department of Neuroscience, Amgen Inc., 360 Binney Street, Cambridge, Massachusetts 02142, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1309
Author: Huang, X., Dementiev, A., Olson, S. T. and Gettins, P. G.
Year: 2010
Title: Basis for the specificity and activation of the serpin protein Z-dependent proteinase inhibitor (ZPI) as an inhibitor of membrane-associated factor Xa
Journal: J Biol Chem
Volume: 285
Issue: 26
Pages: 20399-409
Epub Date: 2010/04/30
Date: Jun 25
Short Title: Basis for the specificity and activation of the serpin protein Z-dependent proteinase inhibitor (ZPI) as an inhibitor of membrane-associated factor Xa
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.112748
PMCID: PMC2888451
Accession Number: 20427285
Keywords: Binding Sites/genetics
Blood Proteins/*chemistry/genetics/metabolism
Calcium/chemistry/metabolism
Cell Membrane/metabolism
Crystallography, X-Ray
Cysteine/chemistry/genetics/metabolism
Factor Xa/*chemistry/metabolism
Glutamic Acid/chemistry/genetics/metabolism
Humans
Kinetics
Light
Models, Molecular
Mutation
Phospholipids/chemistry/metabolism
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Scattering, Radiation
Serpins/*chemistry/genetics/metabolism
Tyrosine/chemistry/genetics/metabolism
Abstract: The serpin ZPI is a protein Z (PZ)-dependent specific inhibitor of membrane-associated factor Xa (fXa) despite having an unfavorable P1 Tyr. PZ accelerates the inhibition reaction approximately 2000-fold in the presence of phospholipid and Ca(2+). To elucidate the role of PZ, we determined the x-ray structure of Gla-domainless PZ (PZ(DeltaGD)) complexed with protein Z-dependent proteinase inhibitor (ZPI). The PZ pseudocatalytic domain bound ZPI at a novel site through ionic and polar interactions. Mutation of four ZPI contact residues eliminated PZ binding and membrane-dependent PZ acceleration of fXa inhibition. Modeling of the ternary Michaelis complex implicated ZPI residues Glu-313 and Glu-383 in fXa binding. Mutagenesis established that only Glu-313 is important, contributing approximately 5-10-fold to rate acceleration of fXa and fXIa inhibition. Limited conformational change in ZPI resulted from PZ binding, which contributed only approximately 2-fold to rate enhancement. Instead, template bridging from membrane association, together with previously demonstrated interaction of the fXa and ZPI Gla domains, resulted in an additional approximately 1000-fold rate enhancement. To understand why ZPI has P1 tyrosine, we examined a P1 Arg variant. This reacted at a diffusion-limited rate with fXa, even without PZ, and predominantly as substrate, reflecting both rapid acylation and deacylation. P1 tyrosine thus ensures that reaction with fXa or most other arginine-specific proteinases is insignificant unless PZ binds and localizes ZPI and fXa on the membrane, where the combined effects of Gla-Gla interaction, template bridging, and interaction of fXa with Glu-313 overcome the unfavorability of P1 Tyr and ensure a high rate of reaction as an inhibitor.
Notes: 1083-351x
Huang, Xin
Dementiev, Alexey
Olson, Steven T
Gettins, Peter G W
R37 HL039888/HL/NHLBI NIH HHS/United States
HL78827/HL/NHLBI NIH HHS/United States
R01 HL078827/HL/NHLBI NIH HHS/United States
HL79430/HL/NHLBI NIH HHS/United States
R37 HL049234/HL/NHLBI NIH HHS/United States
R01 HL079430/HL/NHLBI NIH HHS/United States
HL49234/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Jun 25;285(26):20399-409. doi: 10.1074/jbc.M110.112748. Epub 2010 Apr 28.
Author Address: Center for Molecular Biology of Oral Diseases, University of Illinois at Chicago, Chicago, Illinois 60607, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 261
Author: Hudgins, W. R., Hampton, B., Burgess, W. H. and Perdue, J. F.
Year: 1992
Title: The identification of O-glycosylated precursors of insulin-like growth factor II
Journal: J Biol Chem
Volume: 267
Issue: 12
Pages: 8153-60
Epub Date: 1992/04/25
Date: Apr 25
Short Title: The identification of O-glycosylated precursors of insulin-like growth factor II
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1569071
Keywords: Amino Acid Sequence
Chromatography, Affinity
Chromatography, Liquid
Electrophoresis, Polyacrylamide Gel
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Insulin-Like Growth Factor II/*metabolism
Isoelectric Point
Lectins/metabolism
Molecular Sequence Data
Protein Precursors/*metabolism
Protein Processing, Post-Translational
Abstract: A procedure that combined ion exchange, gel permeation, and insulin-like growth factor-binding protein 3 (IGF-BP-3) affinity chromatography with chromatofocusing and reversed-phase high pressure liquid chromatography was used to isolate high molecular weight precursors of human insulin-like growth factor II (IGF-II) from acetic acid extracts of Cohn fraction IV1. Two precursors had isoelectric points (pI) of 5.1 and 5.4 and apparent Mr values of 15,000 and 11,500, respectively. An apparent Mr = 16,000 RLPG/Ser29 variant of IGF-II was also identified in the acetic acid extracts. Amino-terminal amino acid sequencing of the major E domain-containing peptide that had been isolated from apparent Mr = 15,000 IGF-II (pI 5.1), following its digestion with the endoprotease Lys-C, indicated the carboxyl terminus of this precursor was near or at Lys88. During the sequencing of this peptide, a sharply reduced yield of derivatized amino acid occurred at cycle 10, indicating that Thr75 had been posttranslationally modified, possibly by O-glycosylation. To evaluate this possibility, the 125I-labeled high molecular weight IGF-IIs and their endoprotease-generated peptides were treated with glycosidases, and their effects were determined from the change in relative mobilities of the polypeptide and peptides during sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Neuraminidase treatment of apparent Mr = 15,000 and 11,500 IGF-II reduced their Mr values to a common value of 10,500. When the desialylated precursors of IGF-II were treated with O-glycosidase, but not when treated with N-glycosidase, the Mr values were reduced further to about 10,000. This was the Mr value that would be predicted for an unglycosylated form of precursor IGF-II that had a carboxyl-terminal end at or near Lys88. When the Ser66-Lys88 endoprotease-generated E domain peptides from pI5.1 and 5.4 high Mr IGF-II were treated with the glycosidases, they had relative mobility changes during sodium dodecyl sulfate-polyacrylamide gel electrophoresis that were similar to those of the intact precursors. Finally, the association of O-linked oligosaccharide with the E domain peptide of IGF-II was confirmed by demonstrating the specificity of binding of the Ser66-Lys88 asialoglycopeptide to jackfruit lectin.
Notes: Hudgins, W R
Hampton, B
Burgess, W H
Perdue, J F
CA-47150/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Apr 25;267(12):8153-60.
Author Address: Jerome H. Holland Laboratory for the Biomedical Sciences, American Red Cross, Rockville, Maryland 20855.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1952
Author: Hulse, R. E., Li, Z., Huang, R. K., Zhang, J. and Clapham, D. E.
Year: 2018
Title: Cryo-EM structure of the polycystin 2-l1 ion channel
Journal: Elife
Volume: 7
Epub Date: 2018/07/14
Date: Jul 13
Short Title: Cryo-EM structure of the polycystin 2-l1 ion channel
Alternate Journal: eLife
ISSN: 2050-084x
DOI: 10.7554/eLife.36931
PMCID: PMC6056229
Accession Number: 30004384
Keywords: Calcium
Calcium Channels/chemistry/metabolism/*ultrastructure
Cations/metabolism
*Cryoelectron Microscopy
Humans
Models, Molecular
Protein Conformation
Receptors, Cell Surface/chemistry/metabolism/*ultrastructure
*Pc-2l1
*TRP channel
*calcium
*cryo-EM
*human
*molecular biophysics
*primary cilia
*structural biology
Abstract: We report the near atomic resolution (3.3 Å) of the human polycystic kidney disease 2-like 1 (polycystin 2-l1) ion channel. Encoded by PKD2L1, polycystin 2-l1 is a calcium and monovalent cation-permeant ion channel in primary cilia and plasma membranes. The related primary cilium-specific polycystin-2 protein, encoded by PKD2, shares a high degree of sequence similarity, yet has distinct permeability characteristics. Here we show that these differences are reflected in the architecture of polycystin 2-l1.
Notes: 2050-084x
Hulse, Raymond E
Orcid: 0000-0002-0110-3752
Li, Zongli
Huang, Rick K
Zhang, Jin
Clapham, David E
Orcid: 0000-0002-4459-9428
T32 HL007572/HL/NHLBI NIH HHS/United States
HHMI/Howard Hughes Medical Institute/United States
Journal Article
Research Support, Non-U.S. Gov't
Elife. 2018 Jul 13;7:e36931. doi: 10.7554/eLife.36931.
Author Address: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States.
Department of Cardiology, Howard Hughes Medical Institute, Boston Children's Hospital, Boston, United States.
Department of Biological Chemistry and Molecular Pharmacology, Howard Hughes Medical Institute, Harvard Medical School, Boston, United States.
School of Basic Medical Sciences, Nanchang University, Nanchang, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 864
Author: Huntington, J. A., Kjellberg, M. and Stenflo, J.
Year: 2003
Title: Crystal structure of protein C inhibitor provides insights into hormone binding and heparin activation
Journal: Structure
Volume: 11
Issue: 2
Pages: 205-15
Epub Date: 2003/02/11
Date: Feb
Short Title: Crystal structure of protein C inhibitor provides insights into hormone binding and heparin activation
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/s0969-2126(02)00944-9
Accession Number: 12575940
Keywords: Binding Sites
Crystallography, X-Ray
Heparin/*metabolism
Protein Binding
Protein C Inhibitor/*chemistry/metabolism
Protein Structure, Secondary
Protein Structure, Tertiary
Retinoids/*metabolism
Abstract: Protein C inhibitor (PCI) is a member of the serpin family that has many biological functions. In blood it acts as a procoagulant, and, in the seminal vesicles, it is required for spermatogenesis. The activity of PCI is affected by heparin binding in a manner unique among the heparin binding serpins, and, in addition, PCI binds hydrophobic hormones with apparent specificity for retinoids. Here we present the 2.4 A crystallographic structure of reactive center loop (RCL) cleaved PCI. A striking feature of the structure is a two-turn N-terminal shortening of helix A, which creates a large hydrophobic pocket that docking studies indicate to be the retinoid binding site. On the basis of surface electrostatic properties, a novel mechanism for heparin activation is proposed.
Notes: Huntington, James A
Kjellberg, Margareta
Stenflo, Johan
G117/444/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Structure. 2003 Feb;11(2):205-15. doi: 10.1016/s0969-2126(02)00944-9.
Author Address: Department of Haematology, Division of Structural Medicine, University of Cambridge, Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, CB2 2XY, Cambridge, United Kingdom. jah52@cam.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1122
Author: Huyton, T., Zhang, J. G., Luo, C. S., Lou, M. Z., Hilton, D. J., Nicola, N. A. and Garrett, T. P.
Year: 2007
Title: An unusual cytokine:Ig-domain interaction revealed in the crystal structure of leukemia inhibitory factor (LIF) in complex with the LIF receptor
Journal: Proc Natl Acad Sci U S A
Volume: 104
Issue: 31
Pages: 12737-42
Epub Date: 2007/07/27
Date: Jul 31
Short Title: An unusual cytokine:Ig-domain interaction revealed in the crystal structure of leukemia inhibitory factor (LIF) in complex with the LIF receptor
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0705577104
PMCID: PMC1937536
Accession Number: 17652170
Keywords: Animals
Crystallography, X-Ray
Cytokine Receptor gp130/chemistry/metabolism
Humans
Immunoglobulins/*chemistry/genetics/immunology/*metabolism
Interleukin-6/chemistry/metabolism
Leukemia Inhibitory Factor/*chemistry/genetics/immunology/*metabolism
Ligands
Mice
Models, Molecular
Protein Binding
Protein Structure, Quaternary
Protein Structure, Tertiary
Receptors, OSM-LIF/*chemistry/genetics/immunology/*metabolism
Signal Transduction
Abstract: Leukemia inhibitory factor (LIF) receptor is a cell surface receptor that mediates the actions of LIF and other IL-6 type cytokines through the formation of high-affinity signaling complexes with gp130. Here we present the crystal structure of a complex of mouse LIF receptor with human LIF at 4.0 A resolution. The structure is, to date, the largest cytokine receptor fragment determined by x-ray crystallography. The binding of LIF to its receptor via the central Ig-like domain is unlike other cytokine receptor complexes that bind ligand predominantly through their cytokine-binding modules. This structure, in combination with previous crystallographic studies, also provides a structural template to understand the formation and orientation of the high-affinity signaling complex between LIF, LIF receptor, and gp130.
Notes: 1091-6490
Huyton, Trevor
Zhang, Jian-Guo
Luo, Cindy S
Lou, Mei-Zhen
Hilton, Douglas J
Nicola, Nicos A
Garrett, Thomas P J
R01 CA022556/CA/NCI NIH HHS/United States
R01 CA 22556/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2007 Jul 31;104(31):12737-42. doi: 10.1073/pnas.0705577104. Epub 2007 Jul 24.
Author Address: The Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Parkville, Victoria 3050, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1669
Author: Hwang, H., Lee, J. Y., Lee, H. K., Park, G. W., Jeong, H. K., Moon, M. H., Kim, J. Y. and Yoo, J. S.
Year: 2014
Title: In-depth analysis of site-specific N-glycosylation in vitronectin from human plasma by tandem mass spectrometry with immunoprecipitation
Journal: Anal Bioanal Chem
Volume: 406
Issue: 30
Pages: 7999-8011
Epub Date: 2014/11/07
Date: Dec
Short Title: In-depth analysis of site-specific N-glycosylation in vitronectin from human plasma by tandem mass spectrometry with immunoprecipitation
Alternate Journal: Analytical and bioanalytical chemistry
ISSN: 1618-2642
DOI: 10.1007/s00216-014-8226-5
Accession Number: 25374123
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Carcinoma, Hepatocellular/blood/chemistry
Glycopeptides/*analysis
Glycosylation
Humans
Immunoprecipitation/*methods
Liver Neoplasms/blood/chemistry
Molecular Sequence Data
Tandem Mass Spectrometry/*methods
Vitronectin/*blood/*chemistry
Abstract: The characterization of site-specific microheterogeneity in glycoprotein is very important for understanding cell biology and disease processes. Vitronectin is well known to be a multifunctional glycoprotein in the blood and the extracellular matrix, which is related to hepatocellular carcinoma (HCC). Here, we systematically analyzed the site-specific N-glycopeptides of vitronectin in human plasma by tandem mass spectrometry combined with immunoprecipitation and hydrophilic interaction liquid chromatography (HILIC) enrichment. Vitronectin was purified with immunoprecipitation by monoclonal antibody from plasma and digested to tryptic N-glycopeptides.Then, enrichment with HILIC materials was used and followed by analysis with nano-LC/MS/MS. The sequences of N-glycopeptides were identified from the mass spectra by high-energy C-trap dissociation (HCD) and collision-induced dissociation (CID). In HCD mode, oxonium ions were used for recognizing glycopeptides and y ions for sequencing the peptide backbone. In CID mode, Y ions were used for characterizing their glycoforms. As a result, a total of 17 site-specific N-glycopeptides were completely identified in all of the three N-glycosylation sites of vitronectin in human plasma, including 12 N-glycopeptides first reported. Finally, we specifically found that three hybrid and four complex glycopeptides of triantennary forms with outer fucosylation increased in HCC human plasma.
Notes: 1618-2650
Hwang, Heeyoun
Lee, Ju Yeon
Lee, Hyun Kyoung
Park, Gun Wook
Jeong, Hoi Keun
Moon, Myeong Hee
Kim, Jin Young
Yoo, Jong Shin
Journal Article
Research Support, Non-U.S. Gov't
Germany
Anal Bioanal Chem. 2014 Dec;406(30):7999-8011. doi: 10.1007/s00216-014-8226-5. Epub 2014 Nov 6.
Author Address: Mass Spectrometry Research Center, Korea Basic Science Institute, Chungbuk, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 956
Author: Hyuga, M., Itoh, S., Kawasaki, N., Ohta, M., Ishii, A., Hyuga, S. and Hayakawa, T.
Year: 2004
Title: Analysis of site-specific glycosylation in recombinant human follistatin expressed in Chinese hamster ovary cells
Journal: Biologicals
Volume: 32
Issue: 2
Pages: 70-7
Epub Date: 2004/09/30
Date: Jun
Short Title: Analysis of site-specific glycosylation in recombinant human follistatin expressed in Chinese hamster ovary cells
Alternate Journal: Biologicals : journal of the International Association of Biological Standardization
ISSN: 1045-1056 (Print)
1045-1056
DOI: 10.1016/j.biologicals.2004.04.001
Accession Number: 15454184
Keywords: Amino Acid Sequence
Animals
Asparagine/chemistry
Binding Sites
CHO Cells
Carbohydrates
Cell Line
Chromatography, Liquid
Cricetinae
Electrophoresis, Polyacrylamide Gel
Follistatin/*chemistry
Glycopeptides/chemistry
Glycosylation
Humans
Isoelectric Focusing
Mass Spectrometry
Molecular Sequence Data
Monosaccharides/chemistry
Oligosaccharides/chemistry
Peptides/chemistry
Recombinant Proteins/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Time Factors
Abstract: Follistatin (FS), a glycoprotein, plays an important role in cell growth and differentiation through the neutralization of the biological activities of activins. In this study, we analyzed the glycosylation of recombinant human FS (rhFS) produced in Chinese hamster ovary cells. The results of SDS-PAGE and MALDI-TOF MS revealed the presence of both non-glycosylated and glycosylated forms. FS contains two potential N-glycosylation sites, Asn95 and Asn259. Using mass spectrometric peptide/glycopeptide mapping and precursor-ion scanning, we found that both N-glycosylation sites were partially glycosylated. Monosaccharide composition analyses suggested the linkages of fucosylated bi- and triantennary complex-type oligosaccharides on rhFS. This finding was supported by mass spectrometric oligosaccharide profiling, in which the m/z values and elution times of some of the oligosaccharides from rhFS were in good agreement with those of standard oligosaccharides. Site-specific glycosylation was deduced on the basis of the mass spectra of the glycopeptides. It was suggested that biantennary oligosaccharides are major oligosaccharides located at both Asn95 and Asn259, whereas the triantennary structures are present mainly at Asn95.
Notes: Hyuga, Masashi
Itoh, Satsuki
Kawasaki, Nana
Ohta, Miyako
Ishii, Akiko
Hyuga, Sumiko
Hayakawa, Takao
Journal Article
Research Support, Non-U.S. Gov't
England
Biologicals. 2004 Jun;32(2):70-7. doi: 10.1016/j.biologicals.2004.04.001.
Author Address: Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, 1-18-1, Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan. mhyuga@nihs.go.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1152
Author: Iacob, R. E., Perdivara, I., Przybylski, M. and Tomer, K. B.
Year: 2008
Title: Mass spectrometric characterization of glycosylation of hepatitis C virus E2 envelope glycoprotein reveals extended microheterogeneity of N-glycans
Journal: J Am Soc Mass Spectrom
Volume: 19
Issue: 3
Pages: 428-44
Epub Date: 2008/01/12
Date: Mar
Short Title: Mass spectrometric characterization of glycosylation of hepatitis C virus E2 envelope glycoprotein reveals extended microheterogeneity of N-glycans
Alternate Journal: Journal of the American Society for Mass Spectrometry
ISSN: 1044-0305 (Print)
1044-0305
DOI: 10.1016/j.jasms.2007.11.022
PMCID: PMC2287207
NIHMSID: NIHMS43788
Accession Number: 18187336
Keywords: Acetylglucosamine/analysis
Amino Acid Sequence
Chymotrypsin/chemistry
Disaccharides/chemistry
Fucose/analysis
Glycopeptides/analysis/chemistry
Glycosylation
Mannose/analysis
Mass Spectrometry/*methods
Models, Molecular
Oligosaccharides, Branched-Chain/chemistry
Polysaccharides/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
Tandem Mass Spectrometry/methods
Viral Envelope Proteins/*chemistry
Abstract: Hepatitis C virus (HCV) causes acute and chronic liver disease in humans, including chronic hepatitis, cirrhosis, and hepatocellular carcinoma. The polyprotein encoded in the HCV genome is co- and post-translationally processed by host and viral peptidases, generating the structural proteins Core, E1, E2, and p7, and five nonstructural proteins. The two envelope proteins E1 and E2 are heavily glycosylated. Studying the glycan moieties attached to the envelope E2 glycoprotein is important because the N-linked glycans on E2 envelope protein are involved in the interaction with some human neutralizing antibodies, and may also have a direct or indirect effect on protein folding. In the present study, we report the mass spectrometric characterization of the glycan moieties attached to the E2 glycoprotein. The mass spectrometric analysis clearly identified the nature, composition, and microheterogeneity of the sugars attached to the E2 glycopeptides. All 11 sites of glycosylation on E2 protein were characterized, and the majority of these sites proved to be occupied by high mannose glycans. However, complex type oligosaccharides, which have not been previously identified, were exclusively observed at two N-linked sites, and their identity and heterogeneity were determined.
Notes: Iacob, Roxana E
Perdivara, Irina
Przybylski, Michael
Tomer, Kenneth B
Z01 ES050171-08/Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
J Am Soc Mass Spectrom. 2008 Mar;19(3):428-44. doi: 10.1016/j.jasms.2007.11.022. Epub 2007 Dec 8.
Author Address: Laboratory of Structural Biology, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, North Carolina 27709, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1255
Author: Igarashi, M., Osuga, J., Isshiki, M., Sekiya, M., Okazaki, H., Takase, S., Takanashi, M., Ohta, K., Kumagai, M., Nishi, M., Fujita, T., Nagai, R., Kadowaki, T. and Ishibashi, S.
Year: 2010
Title: Targeting of neutral cholesterol ester hydrolase to the endoplasmic reticulum via its N-terminal sequence
Journal: J Lipid Res
Volume: 51
Issue: 2
Pages: 274-85
Epub Date: 2009/07/14
Date: Feb
Short Title: Targeting of neutral cholesterol ester hydrolase to the endoplasmic reticulum via its N-terminal sequence
Alternate Journal: Journal of lipid research
ISSN: 0022-2275 (Print)
0022-2275
DOI: 10.1194/jlr.M900201-JLR200
PMCID: PMC2803229
Accession Number: 19592704
Keywords: Animals
Biocatalysis
Catalytic Domain
Cattle
Cell Line
Endoplasmic Reticulum/*metabolism
Glucose/metabolism
Glycosylation
Humans
Hydrophobic and Hydrophilic Interactions
Intracellular Space/metabolism
Lipid Metabolism
Mice
Mice, Inbred C57BL
Protein Transport
Sterol Esterase/*chemistry/*metabolism
Abstract: Neutral cholesterol ester hydrolase (NCEH) accounts for a large part of the nCEH activity in macrophage foam cells, a hallmark of atherosclerosis, but its subcellular localization and structure-function relationship are unknown. Here, we determined subcellular localization, glycosylation, and nCEH activity of a series of NCEH mutants expressed in macrophages. NCEH is a single-membrane-spanning type II membrane protein comprising three domains: N-terminal, catalytic, and lipid-binding domains. The N-terminal domain serves as a type II signal anchor sequence to recruit NCEH to the endoplasmic reticulum (ER) with its catalytic domain within the lumen. All of the putative N-linked glycosylation sites (Asn(270), Asn(367), and Asn(389)) of NCEH are glycosylated. Glycosylation at Asn(270), which is located closest to the catalytic serine motif, is important for the enzymatic activity. Cholesterol loading by incubation with acetyl-LDL does not change the ER localization of NCEH. In conclusion, NCEH is targeted to the ER of macrophages, where it hydrolyzes CE to deliver cholesterol for efflux out of the cells.
Notes: 1539-7262
Igarashi, Masaki
Osuga, Jun-Ichi
Isshiki, Masashi
Sekiya, Motohiro
Okazaki, Hiroaki
Takase, Satoru
Takanashi, Mikio
Ohta, Keisuke
Kumagai, Masayoshi
Nishi, Makiko
Fujita, Toshiro
Nagai, Ryozo
Kadowaki, Takashi
Ishibashi, Shun
Journal Article
Research Support, Non-U.S. Gov't
J Lipid Res. 2010 Feb;51(2):274-85. doi: 10.1194/jlr.M900201-JLR200. Epub 2009 Jul 10.
Author Address: Department of Metabolic Diseases, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1108
Author: Ignoul, S., Simaels, J., Hermans, D., Annaert, W. and Eggermont, J.
Year: 2007
Title: Human ClC-6 is a late endosomal glycoprotein that associates with detergent-resistant lipid domains
Journal: PLoS One
Volume: 2
Issue: 5
Pages: e474
Epub Date: 2007/05/31
Date: May 30
Short Title: Human ClC-6 is a late endosomal glycoprotein that associates with detergent-resistant lipid domains
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0000474
PMCID: PMC1868598
Accession Number: 17534424
Keywords: Amino Acid Sequence
Animals
Antibodies/chemistry/immunology
Blotting, Western
COS Cells
Cell Line, Tumor
Chloride Channels/immunology/*metabolism
Chlorocebus aethiops
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Immunohistochemistry
*Lipid Metabolism
Molecular Sequence Data
Sequence Homology, Amino Acid
Abstract: BACKGROUND: The mammalian CLC protein family comprises nine members (ClC-1 to -7 and ClC-Ka, -Kb) that function either as plasma membrane chloride channels or as intracellular chloride/proton antiporters, and that sustain a broad spectrum of cellular processes, such as membrane excitability, transepithelial transport, endocytosis and lysosomal degradation. In this study we focus on human ClC-6, which is structurally most related to the late endosomal/lysomal ClC-7. PRINCIPAL FINDINGS: Using a polyclonal affinity-purified antibody directed against a unique epitope in the ClC-6 COOH-terminal tail, we show that human ClC-6, when transfected in COS-1 cells, is N-glycosylated in a region that is evolutionary poorly conserved between mammalian CLC proteins and that is located between the predicted helices K and M. Three asparagine residues (N410, N422 and N432) have been defined by mutagenesis as acceptor sites for N-glycosylation, but only two of the three sites seem to be simultaneously N-glycosylated. In a differentiated human neuroblastoma cell line (SH-SY5Y), endogenous ClC-6 colocalizes with LAMP-1, a late endosomal/lysosomal marker, but not with early/recycling endosomal markers such as EEA-1 and transferrin receptor. In contrast, when transiently expressed in COS-1 or HeLa cells, human ClC-6 mainly overlaps with markers for early/recycling endosomes (transferrin receptor, EEA-1, Rab5, Rab4) and not with late endosomal/lysosomal markers (LAMP-1, Rab7). Analogously, overexpression of human ClC-6 in SH-SY5Y cells also leads to an early/recycling endosomal localization of the exogenously expressed ClC-6 protein. Finally, in transiently transfected COS-1 cells, ClC-6 copurifies with detergent-resistant membrane fractions, suggesting its partitioning in lipid rafts. Mutating a juxtamembrane string of basic amino acids (amino acids 71-75: KKGRR) disturbs the association with detergent-resistant membrane fractions and also affects the segregation of ClC-6 and ClC-7 when cotransfected in COS-1 cells. CONCLUSIONS: We conclude that human ClC-6 is an endosomal glycoprotein that partitions in detergent resistant lipid domains. The differential sorting of endogenous (late endosomal) versus overexpressed (early and recycling endosomal) ClC-6 is reminiscent of that of other late endosomal/lysosomal membrane proteins (e.g. LIMP II), and is consistent with a rate-limiting sorting step for ClC-6 between early endosomes and its final destination in late endosomes.
Notes: 1932-6203
Ignoul, Sofie
Simaels, Jeannine
Hermans, Diane
Annaert, Wim
Eggermont, Jan
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2007 May 30;2(5):e474. doi: 10.1371/journal.pone.0000474.
Author Address: Laboratory of Membrane Transport, Department of Molecular Cell Biology, University of Leuven, Leuven, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1119
Author: Iida, R., Yasuda, T. and Kishi, K.
Year: 2007
Title: Identification of novel fibronectin fragments detected specifically in juvenile urine
Journal: Febs j
Volume: 274
Issue: 15
Pages: 3939-47
Epub Date: 2007/07/07
Date: Aug
Short Title: Identification of novel fibronectin fragments detected specifically in juvenile urine
Alternate Journal: The FEBS journal
ISSN: 1742-464X (Print)
1742-464x
DOI: 10.1111/j.1742-4658.2007.05926.x
Accession Number: 17614963
Keywords: Adolescent
Adult
Amino Acid Sequence
Animals
Cell Extracts
Cell Line
Child
Child, Preschool
Chromatography, High Pressure Liquid
Fibronectins/chemistry/*classification/genetics/*urine
Gene Expression Regulation
Humans
Isoelectric Focusing
Male
Molecular Sequence Data
Osteoclasts/metabolism
Peptide Fragments/chemistry/*classification/genetics/*urine
Sensitivity and Specificity
Sequence Alignment
Abstract: Ugl-Y (young age-related urinary glycoprotein) is an age-specific protein that we have previously identified in urine from healthy subjects under 18 years of age. Isoelectric focusing analysis of Ugl-Y gives a set of three bands, Y1, Y2 and Y3, in the pH region around 3. To determine the complete structure of Ugl-Y, purified Y1 and Y2 from pediatric urine were enzymatically cleaved, and the resulting peptides were analyzed by protein sequencing and/or MALDI-TOF MS. As a result, it was demonstrated that Y1 consists of 189 amino acid residues, and is identical to the region from A723 to R911 of fibronectin, whereas Y2 consists of 181 amino acid residues, and is identical to the region from A723 to R903. Electrophoretic analysis of the lysate prepared from COS-7 cells transfected with Y1- or Y2-expressing vectors gave specific bands corresponding to Y1 or Y2, respectively, showing the validity of the sequences determined. Partial purification of pediatric serum followed by western blotting revealed that Ugl-Y is derived from plasma. Furthermore, Ugl-Y was generated by in vitro digestion of fibronectin by acid protease in extracts of osteoclast cells. These findings suggest that Ugl-Y is probably produced by degradation of fibronectin comprising bone matrix during the process of vigorous bone resorption in children and adolescents. This is the first report on the identification and characterization of juvenile-specific fibronectin fragments excreted into urine.
Notes: Iida, Reiko
Yasuda, Toshihiro
Kishi, Koichiro
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2007 Aug;274(15):3939-47. doi: 10.1111/j.1742-4658.2007.05926.x. Epub 2007 Jul 5.
Author Address: Division of Forensic Medicine, Faculty of Medical Sciences, University of Fukui, Fukui, Japan. riida@u-fukui.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1096
Author: Iijima, T., Miura, E., Matsuda, K., Kamekawa, Y., Watanabe, M. and Yuzaki, M.
Year: 2007
Title: Characterization of a transneuronal cytokine family Cbln--regulation of secretion by heteromeric assembly
Journal: Eur J Neurosci
Volume: 25
Issue: 4
Pages: 1049-57
Epub Date: 2007/03/03
Date: Feb
Short Title: Characterization of a transneuronal cytokine family Cbln--regulation of secretion by heteromeric assembly
Alternate Journal: The European journal of neuroscience
ISSN: 0953-816X (Print)
0953-816x
DOI: 10.1111/j.1460-9568.2007.05361.x
Accession Number: 17331201
Keywords: Amino Acid Sequence
Animals
Animals, Newborn
Biological Transport
Cells, Cultured
Cerebellum/cytology
Culture Media/chemistry
Endoplasmic Reticulum/metabolism/ultrastructure
Gene Expression Regulation/physiology
Glycoproteins/*metabolism
Humans
Immunoprecipitation/methods
Mice
Mice, Knockout
Models, Biological
Nerve Tissue Proteins/deficiency/*physiology
Neurons/cytology/metabolism/ultrastructure
*Protein Precursors/deficiency
Protein Structure, Tertiary/physiology
Sequence Alignment
Transfection/methods
Abstract: Cbln1, a member of the C1q and tumor necrosis factor superfamily, plays crucial roles as a cerebellar granule cell-derived transneuronal regulator of synapse integrity and plasticity in Purkinje cells. Although other Cbln family members, Cbln2-Cbln4, have distinct spatial and temporal patterns of expression throughout the CNS, their biochemical and biological properties have remained largely uncharacterized. Here, we demonstrated that in mammalian heterologous cells, Cbln2 and Cbln4 were secreted as N-linked glycoproteins, like Cbln1. In contrast, despite the presence of a functional signal sequence, Cbln3 was not secreted when expressed alone but was retained in the endoplasmic reticulum (ER) or cis-Golgi because of its N-terminal domain. All members of the Cbln family formed not only homomeric but also heteromeric complexes with each other in vitro. Accordingly, when Cbln1 and Cbln3 were co-expressed in heterologous cells, a proportion of the Cbln1 proteins was retained in the ER or cis-Golgi; conversely, some Cbln3 proteins were secreted together with Cbln1. Similarly, in wild-type granule cells expressing Cbln1 and Cbln3, Cbln3 proteins were partially secreted and reached postsynaptic sites on Purkinje cell dendrites, while Cbln3 was almost completely degraded in cbln1-null granule cells. These results indicate that like Cbln1, Cbln2 and Cbln4 may also serve as transneuronal regulators of synaptic functions in various brain regions. Furthermore, heteromer formation between Cbln1 and Cbln3 in cerebellar granule cells may modulate each other's trafficking and signaling pathways; similarly, heteromerization of other Cbln family proteins may also have biological significance in other neurons.
Notes: Iijima, Takatoshi
Miura, Eriko
Matsuda, Keiko
Kamekawa, Yuichi
Watanabe, Masahiko
Yuzaki, Michisuke
Journal Article
Research Support, Non-U.S. Gov't
France
Eur J Neurosci. 2007 Feb;25(4):1049-57. doi: 10.1111/j.1460-9568.2007.05361.x.
Author Address: Department of Physiology, School of Medicine, Keio University, Tokyo 160-8582, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 363
Author: Inaba, M. and Maede, Y.
Year: 1989
Title: O-N-acetyl-D-glucosamine moiety on discrete peptide of multiple protein 4.1 isoforms regulated by alternative pathways
Journal: J Biol Chem
Volume: 264
Issue: 30
Pages: 18149-55
Epub Date: 1989/10/25
Date: Oct 25
Short Title: O-N-acetyl-D-glucosamine moiety on discrete peptide of multiple protein 4.1 isoforms regulated by alternative pathways
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2808371
Keywords: Acetylglucosamine/*analysis
Amino Acid Sequence
Animals
Chromatography, Ion Exchange
Chymotrypsin
*Cytoskeletal Proteins
Electrophoresis, Polyacrylamide Gel
Erythrocyte Membrane/analysis/metabolism
Galactosemias/blood
Glucosamine/*analogs & derivatives
Glycopeptides/isolation & purification
Glycosylation
Humans
*Membrane Proteins/isolation & purification
Molecular Sequence Data
Molecular Weight
*Neuropeptides
Peptide Fragments/isolation & purification
Peptide Mapping
Abstract: Erythrocyte protein 4.1 is a cytoplasmic protein that possesses a protein-saccharide modification structure, an O-N-acetyl-D-glucosamine (GlcNAc) moiety. We determined the amino acid sequence of the proteolytic fragment containing the O-GlcNAc moiety after labeling the saccharide with [3H]galactose in the presence of bovine milk galactosyltransferase. Glycosylation appears to occur on one or more serine or threonine residues in the following sequence: Thr-Ala-Gln-Thr-Ile-Thr-Ser-Glu-Thr-Pro-Ser-Ser-Thr-Thr-Thr-Thr-Gln-Ile-Thr-Lys . This sequence corresponds to the carboxyl-terminal half of the 34-amino acid peptide in the 22/24-kDa carboxyl-terminal domain of protein 4.1, which is one of the discrete peptides regulated by alternative RNA splicing. Multiple protein 4.1 isoforms in erythroid and nonerythroid cells including major components of erythrocyte membrane proteins, 4.1a and 4.1b, appear to contain this sequence since most immunochemically reactive proteins were labeled with [3H]galactose, with the exception of several variant polypeptides. These results appear to suggest the functional or biological significance of the O-GlcNAc linkage in protein 4.1.
Notes: Inaba, M
Maede, Y
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1989 Oct 25;264(30):18149-55.
Author Address: Department of Veterinary Internal Medicine, Faculty of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1923
Author: Inoue, D., Fujino, T., Sheridan, P., Zhang, Y. Z., Nagase, R., Horikawa, S., Li, Z., Matsui, H., Kanai, A., Saika, M., Yamaguchi, R., Kozuka-Hata, H., Kawabata, K. C., Yokoyama, A., Goyama, S., Inaba, T., Imoto, S., Miyano, S., Xu, M., Yang, F. C., Oyama, M. and Kitamura, T.
Year: 2018
Title: A novel ASXL1-OGT axis plays roles in H3K4 methylation and tumor suppression in myeloid malignancies
Journal: Leukemia
Volume: 32
Issue: 6
Pages: 1327-1337
Epub Date: 2018/03/21
Date: Jun
Short Title: A novel ASXL1-OGT axis plays roles in H3K4 methylation and tumor suppression in myeloid malignancies
Alternate Journal: Leukemia
ISSN: 0887-6924
DOI: 10.1038/s41375-018-0083-3
Accession Number: 29556021
Keywords: Animals
Cell Differentiation
Female
HEK293 Cells
HL-60 Cells
Histones/*metabolism
Humans
Leukemia, Myeloid, Acute/*etiology/prevention & control
Methylation
Mice
Mice, Inbred C57BL
Myelodysplastic Syndromes/*etiology/prevention & control
N-Acetylglucosaminyltransferases/*physiology
Protein Stability
Repressor Proteins/chemistry/*physiology
Tumor Suppressor Proteins/physiology
Abstract: ASXL1 plays key roles in epigenetic regulation of gene expression through methylation of histone H3K27, and disruption of ASXL1 drives myeloid malignancies, at least in part, via derepression of posterior HOXA loci. However, little is known about the identity of proteins that interact with ASXL1 and about the functions of ASXL1 in modulation of the active histone mark, such as H3K4 methylation. In this study, we demonstrate that ASXL1 is a part of a protein complex containing HCFC1 and OGT; OGT directly stabilizes ASXL1 by O-GlcNAcylation. Disruption of this novel axis inhibited myeloid differentiation and H3K4 methylation as well as H2B glycosylation and impaired transcription of genes involved in myeloid differentiation, splicing, and ribosomal functions; this has implications for myelodysplastic syndrome (MDS) pathogenesis, as each of these processes are perturbed in the disease. This axis is responsible for tumor suppression in the myeloid compartment, as reactivation of OGT induced myeloid differentiation and reduced leukemogenecity both in vivo and in vitro. Our data also suggest that MLL5, a known HCFC1/OGT-interacting protein, is responsible for gene activation by the ASXL1-OGT axis. These data shed light on the novel roles of the ASXL1-OGT axis in H3K4 methylation and activation of transcription.
Notes: 1476-5551
Inoue, Daichi
Fujino, Takeshi
Sheridan, Paul
Zhang, Yao-Zhong
Nagase, Reina
Horikawa, Sayuri
Li, Zaomin
Matsui, Hirotaka
Kanai, Akinori
Saika, Makoto
Yamaguchi, Rui
Kozuka-Hata, Hiroko
Kawabata, Kimihito Cojin
Yokoyama, Akihiko
Goyama, Susumu
Inaba, Toshiya
Imoto, Seiya
Orcid: 0000-0002-2989-308x
Miyano, Satoru
Xu, Mingjiang
Yang, Feng-Chun
Oyama, Masaaki
Kitamura, Toshio
Journal Article
Research Support, Non-U.S. Gov't
England
Leukemia. 2018 Jun;32(6):1327-1337. doi: 10.1038/s41375-018-0083-3. Epub 2018 Mar 3.
Author Address: Division of Cellular Therapy, The Institute of Medical Science, The University of Tokyo, Tokyo, 1088639, Japan. inoued0612@gmail.com.
Division of Cellular Therapy, The Institute of Medical Science, The University of Tokyo, Tokyo, 1088639, Japan.
Laboratory of Genome Data Base, The Institute of Medical Science, The University of Tokyo, Tokyo, 1088639, Japan.
Department of Biochemistry and Molecular Biology, Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, Miami, FL, 33136, USA.
Department of Molecular Laboratory Medicine, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, 8608556, Japan.
Department of Molecular Oncology and Leukemia Program Project, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, 7348553, Japan.
Medical Proteomics Laboratory, The Institute of Medical Science, The University of Tokyo, Tokyo, 1088639, Japan.
Tsuruoka Metabolomics Laboratory, National Cancer Center, Tsuruoka, Japan.
Division of Cellular Therapy, The Institute of Medical Science, The University of Tokyo, Tokyo, 1088639, Japan. kitamura@ims.u-tokyo.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 569
Author: Inoue, K. and Morita, T.
Year: 1993
Title: Identification of O-linked oligosaccharide chains in the activation peptides of blood coagulation factor X. The role of the carbohydrate moieties in the activation of factor X
Journal: Eur J Biochem
Volume: 218
Issue: 1
Pages: 153-63
Epub Date: 1993/11/15
Date: Nov 15
Short Title: Identification of O-linked oligosaccharide chains in the activation peptides of blood coagulation factor X. The role of the carbohydrate moieties in the activation of factor X
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb18361.x
Accession Number: 8243461
Keywords: Amino Acid Sequence
Animals
Cattle
Factor X/*chemistry/metabolism
Glycosylation
Humans
Molecular Sequence Data
Oligosaccharides/*analysis
Peptide Fragments/*chemistry
Sequence Homology, Amino Acid
Abstract: Conversion of factor X to factor Xa results in release of a heavily glycosylated activation peptide. Analysis of protease-digested glycopeptides derived from the activation peptides of bovine and human blood coagulation factor X allowed the identification of sites of the O-linked oligosaccharide chains in these peptides. Glycopeptides were prepared from the activation peptides by digestion with chymotrypsin or Staphylococcus aureus V8 protease. By combined analysis of amino acid sequence and sialic acid content, we found that bovine factor X had an O-linked oligosaccharide chain linked to Thr26, and human factor X had four carbohydrate-attachment sites, namely, O-glycosidic linkages to Thr17 and Thr29, respectively, and N-glycosidic linkages to Asn39 and Asn49, respectively, in their activation peptides. The O-linked carbohydrate-attachment sites were identified since the yields of phenylthiohydantoin derivatives of amino acids that corresponded to their residues were increased during amino acid sequencing after deglycosylation of the glycopeptides with sialidase and O-glycanase. The effect of deglycosylation of bovine factor X1 was investigated with factor-X-activating enzyme from Russell's viper venom or extrinsic Xase (factor VIIa/tissue factor/phospholipid) by examining the activation rates of derivatives of factor X prepared using O-glycanase, sialidase, and/or N-glycanase. The removal of O-linked carbohydrate resulted in a decrease in the rate of activation. It appears that carbohydrate residues in factor X play an important role in the activation of the zymogen.
Notes: Inoue, K
Morita, T
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1993 Nov 15;218(1):153-63. doi: 10.1111/j.1432-1033.1993.tb18361.x.
Author Address: Department of Biochemistry, Meiji College of Pharmacy, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1205
Author: Inoue, N., Ikawa, M. and Okabe, M.
Year: 2008
Title: Putative sperm fusion protein IZUMO and the role of N-glycosylation
Journal: Biochem Biophys Res Commun
Volume: 377
Issue: 3
Pages: 910-4
Epub Date: 2008/10/28
Date: Dec 19
Short Title: Putative sperm fusion protein IZUMO and the role of N-glycosylation
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2008.10.073
Accession Number: 18952059
Keywords: Amino Acid Sequence
Animals
Cell Fusion
Dogs
Epididymis/cytology/growth & development/metabolism
Female
*Fertility/genetics
Glycosylation
Humans
Immunoglobulins/genetics/*metabolism
Male
Membrane Proteins/genetics/*metabolism
Mice
Mice, Transgenic
Molecular Sequence Data
Ovum/metabolism/physiology
Rats
Spermatozoa/metabolism/*physiology
Abstract: IZUMO is the mouse sperm protein proven to be essential for fusion with eggs. It contains one immunoglobulin-like domain with a conserved glycosylation site within. In the present paper, we produced transgenic mouse lines expressing unglycosylated IZUMO (N204Q-IZUMO) in Izumo1 -/- background. The expression of N204Q-IZUMO rescued the infertile phenotype of IZUMO disrupted mice, indicating glycosylation is not essential for fusion-facilitating activity of IZUMO. The N204Q-IZUMO was produced in testis in comparable amounts to wild-type IZUMO, but the amount of N204Q-IZUMO on sperm was significantly decreased by the time sperm reached the cauda epididymis. These data suggest that glycosylation is not essential for the function of IZUMO, but has a role in protecting it from fragmentation in cauda epididymis.
Notes: 1090-2104
Inoue, Naokazu
Ikawa, Masahito
Okabe, Masaru
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2008 Dec 19;377(3):910-4. doi: 10.1016/j.bbrc.2008.10.073. Epub 2008 Oct 24.
Author Address: Research Institute for Microbial Diseases, Osaka University, Yamadaoka 3-1, Suita, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 594
Author: Inoue, N., Takeuchi, M., Asano, K., Shimizu, R., Takasaki, S. and Kobata, A.
Year: 1993
Title: Structures of mucin-type sugar chains on human erythropoietins purified from urine and the culture medium of recombinant Chinese hamster ovary cells
Journal: Arch Biochem Biophys
Volume: 301
Issue: 2
Pages: 375-8
Epub Date: 1993/03/01
Date: Mar
Short Title: Structures of mucin-type sugar chains on human erythropoietins purified from urine and the culture medium of recombinant Chinese hamster ovary cells
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1993.1158
Accession Number: 8460945
Keywords: Anemia, Aplastic
Animals
CHO Cells
Carbohydrate Sequence
Cricetinae
Erythropoietin/*chemistry/genetics/urine
Humans
Molecular Sequence Data
Mucins/*chemistry/genetics/urine
Oligosaccharides/*chemistry/isolation & purification
Recombinant Proteins/chemistry
Urine/chemistry
Abstract: Less is known about the mucin-type sugar chains attached to human erythropoietin as compared with N-linked sugar chains which structures and function have been well studied. In this study, we purified urinary human erythropoietin from three independent groups of aplastic anemia patients, and analyzed the structures of mucin-type sugar chains as well as that obtained from recombinant human erythropoietin produced by Chinese hamster ovary cells. Unlike the N-linked sugar chains, the mucin-type sugar chains are totally different between the urinary and the recombinant erythropoietins. All of the three independent samples of urinary erythropoietin contained oligosaccharides with the structures of +/- Neu5Ac alpha 2-->6GalNAc, while recombinant human erythropoietin contained those with the structures of Neu5Ac alpha 2-->3Gal beta 1-->3(Neu5Ac alpha 2-->6)GalNAc, Neu5Ac alpha 2-->3Gal beta 1-->3GalNAc, and Gal beta 1-->3(Neu5Ac alpha 2-->6)GalNAc.
Notes: Inoue, N
Takeuchi, M
Asano, K
Shimizu, R
Takasaki, S
Kobata, A
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Arch Biochem Biophys. 1993 Mar;301(2):375-8. doi: 10.1006/abbi.1993.1158.
Author Address: Pharmaceutical Laboratory of Kirin Brewery Co. Ltd., Gumma, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1895
Author: Inoue, Y., Moriwaki, K., Ueda, Y., Takeuchi, T., Higuchi, K. and Asahi, M.
Year: 2018
Title: Elevated O-GlcNAcylation stabilizes FOXM1 by its reduced degradation through GSK-3β inactivation in a human gastric carcinoma cell line, MKN45 cells
Journal: Biochem Biophys Res Commun
Volume: 495
Issue: 2
Pages: 1681-1687
Epub Date: 2017/12/03
Date: Jan 8
Short Title: Elevated O-GlcNAcylation stabilizes FOXM1 by its reduced degradation through GSK-3β inactivation in a human gastric carcinoma cell line, MKN45 cells
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2017.11.179
Accession Number: 29196265
Keywords: Acetylglucosamine/metabolism
Acylation
Cell Line, Tumor
Cell Proliferation
Diabetes Complications/etiology/metabolism/pathology
Enzyme Inhibitors/pharmacology
Forkhead Box Protein M1/chemistry/*metabolism
Glucose/metabolism
Glycogen Synthase Kinase 3 beta/antagonists & inhibitors/chemistry/*metabolism
Humans
Protein Processing, Post-Translational
Protein Stability/drug effects
Proteolysis/drug effects
Pyrans/pharmacology
Stomach Neoplasms/etiology/*metabolism/pathology
Thiazoles/pharmacology
Ubiquitination/drug effects
Up-Regulation/drug effects
beta-N-Acetylhexosaminidases/antagonists & inhibitors/metabolism
*Foxm1
*Gsk-3β
*O-GlcNAcylation
*Ubiquitination
Abstract: O-GlcNAcylation is a dynamic post-translational modification of cytonuclear proteins for intracellular signaling. Elevated O-GlcNAcylation is a general feature of cancer and contributes to cancer progression, and recent studies indicate the contribution to increasing incidence of various types of cancer in diabetic patients. However, the role of O-GlcNAcylation in tumor progression is not fully elucidated. Forkhead box M1 (FOXM1), a master mitotic transcription factor, has been implicated in all major hallmarks of cancer, and is wildly expressed in solid tumors. Given that FOXM1 expression was reported to be elevated in gastric cancer, we examined the effect of high glucose or an inhibitor of O-GlcNAc hydrolase, Thiamet G (TMG), on FOXM1 protein expression in a human gastric cancer cell line, MKN45 cells, and confirmed that FOXM1 protein level and the cell proliferation were upregulated. To investigate the molecular mechanisms by which FOXM1 protein expression is regulated by O-GlcNAcylation, the effect of high glucose and TMG on FOXM1 ubiquitination was examined in MKN45 cells. As a result, the ubiquitination and degradation of FOXM1 protein were both suppressed by high glucose and TMG treatment. However, the O-GlcNAcylation was not detected on FOXM1 but not on GSK-3β. High glucose and TMG treatment increased phospho-serine 9 GSK-3β, an inactive form, and the degradation of FOXM1 protein was suppressed by treatment of GSK-3β inhibitors in MKN45 cells. Taken together, we suggest that high glucose and elevated O-GlcNAcylation stabilize FOXM1 protein by its reduced degradation via GSK-3β inactivation in MKN45 cells, suggesting that the higher risk of gastric cancer in diabetic patients could be partially due to O-GlcNAcylation-mediated FOXM1 stabilization.
Notes: 1090-2104
Inoue, Yosuke
Moriwaki, Kazumasa
Ueda, Yasuhiro
Takeuchi, Toshihisa
Higuchi, Kazuhide
Asahi, Michio
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2018 Jan 8;495(2):1681-1687. doi: 10.1016/j.bbrc.2017.11.179. Epub 2017 Dec 2.
Author Address: Department of Internal Medicine II, Faculty of Medicine, Osaka Medical College, Osaka 569-8686, Japan.
Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka 569-8686, Japan.
Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka 569-8686, Japan. Electronic address: masahi@osaka-med.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1884
Author: Ishimura, E., Nakagawa, T., Moriwaki, K., Hirano, S., Matsumori, Y. and Asahi, M.
Year: 2017
Title: Augmented O-GlcNAcylation of AMP-activated kinase promotes the proliferation of LoVo cells, a colon cancer cell line
Journal: Cancer Sci
Volume: 108
Issue: 12
Pages: 2373-2382
Epub Date: 2017/10/04
Date: Dec
Short Title: Augmented O-GlcNAcylation of AMP-activated kinase promotes the proliferation of LoVo cells, a colon cancer cell line
Alternate Journal: Cancer science
ISSN: 1347-9032 (Print)
1347-9032
DOI: 10.1111/cas.13412
PMCID: PMC5715261
Accession Number: 28973823
Keywords: AMP-Activated Protein Kinases/*metabolism
Acylation
Animals
Cell Line, Tumor
Cell Proliferation/*physiology
Colonic Neoplasms/*metabolism/*pathology
Heterografts
Humans
Mice
Mice, Inbred BALB C
Mice, Nude
N-Acetylglucosaminyltransferases/metabolism
Protein Processing, Post-Translational
mTOR
AMP-activated kinase
O-GlcNAcylation
colon cancer
tumor growth
Abstract: Increasing incidence of various cancers has been reported in diabetic patients. O-linked N-acetylglucosamine (O-GlcNAc) modification of proteins at serine/threonine residues (O-GlcNAcylation) is an essential post-translational modification that is upregulated in diabetic patients and has been implicated in tumor growth. However, the mechanisms by which O-GlcNAcylation promotes tumor growth remain unclear. Given that AMP-activated kinase (AMPK) has been thought to play important roles in suppressing tumor growth, we evaluated the involvement of AMPK O-GlcNAcylation on the growth of LoVo cells, a human colon cancer cell line. Results revealed that treatment with Thiamet G (TMG), an inhibitor of O-GlcNAc hydrolase, increased both anchorage-dependent and -independent growth of the cells. O-GlcNAc transferase overexpression also increased the growth. These treatments increased AMPK O-GlcNAcylation in a dose-dependent manner, which led to reduced AMPK phosphorylation and mTOR activation. Chemical inhibition or activation of AMPK led to increased or decreased growth, respectively, which was consistent with the data with genetic inhibition of AMPK. In addition, TMG-mediated acceleration of tumor growth was abolished by both chemical and genetic inhibition of AMPK. To examine the effects of AMPK O-GlcNAcylation in vivo, the LoVo cells were s.c. transplanted onto the backs of BALB/c-nu/nu mice. Injection of TMG promoted the growth and enhanced O-GlcNAcylation of the tumors of the mice. Consistent with in vitro data, AMPK O-GlcNAcylation was increased, which reduced AMPK phosphorylation and resulted in activation of mTOR. Collectively, the higher colon cancer risk of diabetic patients could be due to O-GlcNAcylation-mediated AMPK inactivation and subsequent activation of mTOR.
Notes: 1349-7006
Ishimura, Emi
Nakagawa, Takatoshi
Moriwaki, Kazumasa
Hirano, Seiichi
Matsumori, Yoshinobu
Asahi, Michio
Orcid: 0000-0002-3139-6373
Journal Article
Cancer Sci. 2017 Dec;108(12):2373-2382. doi: 10.1111/cas.13412. Epub 2017 Oct 21.
Author Address: Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Takatsuki, Japan.
Department of Gastroenterology, Amagasaki Daimotsu Hospital, Amagasahi, Hyogo, 660-0828, Japan.
Department of Gastroenterology, Takatsuki General Hospital, Takatsuki, Japan.
Amagasaki Daimotsu Hospital, Amagasahi, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 399
Author: Ishioka, N., Takahashi, N. and Putnam, F. W.
Year: 1986
Title: Amino acid sequence of human plasma alpha 1B-glycoprotein: homology to the immunoglobulin supergene family
Journal: Proc Natl Acad Sci U S A
Volume: 83
Issue: 8
Pages: 2363-7
Epub Date: 1986/04/01
Date: Apr
Short Title: Amino acid sequence of human plasma alpha 1B-glycoprotein: homology to the immunoglobulin supergene family
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.83.8.2363
PMCID: PMC323297
Accession Number: 3458201
Keywords: Amino Acid Sequence
Blood Proteins/*analysis
Carbohydrate Sequence
Disulfides
Genes
*Glycoproteins/genetics
Humans
Immunoglobulins/genetics
Molecular Weight
Receptors, Immunologic/genetics
Secretory Component/genetics
Abstract: The complete amino acid sequence has been determined for alpha 1B-glycoprotein (alpha 1B), a protein of unknown function present in human plasma. This protein (Mr approximately equal to 63,000) consists of a single polypeptide chain N-linked to four glucosamine oligosaccharides. The polypeptide has five intrachain disulfide bonds and contains 474 amino acid residues. Analysis of the amino acid sequence by several computer programs shows that alpha 1B exhibits internal duplication and consists of five repeating structural domains, each containing about 95 amino acids and one disulfide bond. alpha 1B has a unique amino acid sequence. However, several domains of alpha 1B, especially the third, show statistically significant homology to variable regions of certain immunoglobulin light and heavy chains. alpha 1B also exhibits sequence similarity to other members of the immunoglobulin supergene family such as the receptor for transepithelial transport of IgA and IgM and the secretory component of human IgA. Because of its internal duplication and its sequence homology to immunoglobulin-like proteins, alpha 1B appears to have evolved from an ancestral gene similar to that of the immunoglobulin supergene family.
Notes: 1091-6490
Ishioka, N
Takahashi, N
Putnam, F W
AM19221/AM/NIADDK NIH HHS/United States
CA08497/CA/NCI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1986 Apr;83(8):2363-7. doi: 10.1073/pnas.83.8.2363.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1077
Author: Ishmael, S. S., Ishmael, F. T., Jones, A. D. and Bond, J. S.
Year: 2006
Title: Protease domain glycans affect oligomerization, disulfide bond formation, and stability of the meprin A metalloprotease homo-oligomer
Journal: J Biol Chem
Volume: 281
Issue: 49
Pages: 37404-15
Epub Date: 2006/10/17
Date: Dec 8
Short Title: Protease domain glycans affect oligomerization, disulfide bond formation, and stability of the meprin A metalloprotease homo-oligomer
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M602769200
Accession Number: 17040911
Keywords: Animals
Base Sequence
Binding Sites/genetics
Cell Line
DNA, Complementary/genetics
Disulfides/chemistry
Enzyme Stability
Glycosylation
Humans
In Vitro Techniques
Metalloendopeptidases/*chemistry/genetics/metabolism
Mice
Mutagenesis, Site-Directed
Peptide Mapping
Polysaccharides/chemistry
Protein Structure, Quaternary
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Abstract: The meprin A homo-oligomer is a highly glycosylated, secreted zinc metalloprotease of the astacin family and metzincin superfamily. This isoform of meprin is composed of disulfide-bonded dimers of alpha subunits that further associate to form large, secreted megadalton complexes of 10 or more subunits. The aim of this study was to determine the sites of glycan attachment and to assess their ability to affect the formation and stability of the homo-oligomer. Nine of the ten potential N-linked glycosylation sites (Asn-41, Asn-152, Asn-234, Asn-270, Asn-330, Asn-426, Asn-452, Asn-546, and Asn-553) were found to be glycosylated in recombinant mouse meprin A using chemical and enzymatic deglycosylation methods and electrospray ionization mass spectrometry. Chemical cross-linking demonstrated that carbohydrates are at or near the noncovalent subunit interface. The removal of two glycans in the protease domain at Asn-234 and Asn-270, as well as one in the tumor necrosis factor receptor-associated factor domain at Asn-452, by a deglycosidase under nondenaturing conditions decreased the chemical and thermal stability of the homo-oligomer without affecting quaternary structure. Site-directed mutagenesis demonstrated that no single glycan was essential for oligomer formation; however, the combined absence of the glycans at Asn-152 and Asn-270 in the protease domain hindered intersubunit disulfide bond formation, prevented noncovalent associations, and abolished enzymatic activity. These studies provide insights into the role of glycans in the biosynthesis, activity, and stability of this extracellular protease.
Notes: Ishmael, Susan S
Ishmael, Faoud T
Jones, A Daniel
Bond, Judith S
DK19691/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
J Biol Chem. 2006 Dec 8;281(49):37404-15. doi: 10.1074/jbc.M602769200. Epub 2006 Oct 13.
Author Address: Department of Biochemistry and Molecular Biology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1392
Author: Isono, T.
Year: 2011
Title: O-GlcNAc-specific antibody CTD110.6 cross-reacts with N-GlcNAc2-modified proteins induced under glucose deprivation
Journal: PLoS One
Volume: 6
Issue: 4
Pages: e18959
Epub Date: 2011/04/29
Date: Apr 19
Short Title: O-GlcNAc-specific antibody CTD110.6 cross-reacts with N-GlcNAc2-modified proteins induced under glucose deprivation
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0018959
PMCID: PMC3079744
Accession Number: 21526146
Keywords: Acetylglucosamine/*metabolism
Antibodies, Monoclonal/*immunology
Antibody Specificity/drug effects/*immunology
Cell Line, Tumor
Cross Reactions/*immunology
Glucose/*deficiency/pharmacology
Glycosylation/drug effects
Humans
Mannosyltransferases/metabolism
N-Acetylglucosaminyltransferases/metabolism
Proteins/*metabolism
Reproducibility of Results
Abstract: Modification of serine and threonine residues in proteins by O-linked β-N-acetylglucosamine (O-GlcNAc) glycosylation is a feature of many cellular responses to the nutritional state and to stress. O-GlcNAc modification is reversibly regulated by O-linked β-N-acetylglucosamine transferase (OGT) and β-D-N-acetylglucosaminase (O-GlcNAcase). O-GlcNAc modification of proteins is dependent on the concentration of uridine 5'-diphospho-N-acetylglucosamine (UDP-GlcNAc), which is a substrate of OGT and is synthesized via the hexosamine biosynthetic pathway. Immunoblot analysis using the O-GlcNAc-specific antibody CTD110.6 has indicated that glucose deprivation increases protein O-GlcNAcylation in some cancer cells. The mechanism of this paradoxical phenomenon has remained unclear. Here we show that the increased glycosylation induced by glucose deprivation and detected by CTD110.6 antibodies is actually modification by N-GlcNAc(2), rather than by O-GlcNAc. We found that this induced glycosylation was not regulated by OGT and O-GlcNAcase, unlike typical O-GlcNAcylation, and it was inhibited by treatment with tunicamycin, an N-glycosylation inhibitor. Proteomics analysis showed that proteins modified by this induced glycosylation were N-GlcNAc(2)-modified glycoproteins. Furthermore, CTD110.6 antibodies reacted with N-GlcNAc(2)-modified glycoproteins produced by a yeast strain with a ts-mutant of ALG1 that could not add a mannose residue to dolichol-PP-GlcNAc(2). Our results demonstrated that N-GlcNAc(2)-modified glycoproteins were induced under glucose deprivation and that they cross-reacted with the O-GlcNAc-specific antibody CTD110.6. We therefore propose that the glycosylation status of proteins previously classified as O-GlcNAc-modified proteins according to their reactivity with CTD110.6 antibodies must be re-examined. We also suggest that the repression of mature N-linked glycoproteins due to increased levels of N-GlcNAc(2)-modified proteins is a newly recognized pathway for effective use of sugar under stress and deprivation conditions. Further research is needed to clarify the physiological and pathological roles of N-GlcNAc(2)-modified proteins.
Notes: 1932-6203
Isono, Takahiro
Journal Article
PLoS One. 2011 Apr 19;6(4):e18959. doi: 10.1371/journal.pone.0018959.
Author Address: Central Research Laboratory, Shiga University of Medical Science, Otsu, Shiga, Japan. isono@belle.shiga-med.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 578
Author: Ito, K., Takahashi, N., Takahashi, A., Shimada, I., Arata, Y., O'Brien, J. S. and Kishimoto, Y.
Year: 1993
Title: Structural study of the oligosaccharide moieties of sphingolipid activator proteins, saposins A, C and D obtained from the spleen of a Gaucher patient
Journal: Eur J Biochem
Volume: 215
Issue: 1
Pages: 171-9
Epub Date: 1993/07/01
Date: Jul 1
Short Title: Structural study of the oligosaccharide moieties of sphingolipid activator proteins, saposins A, C and D obtained from the spleen of a Gaucher patient
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb18020.x
Accession Number: 8344278
Keywords: Carbohydrate Sequence
Gaucher Disease/*metabolism
Glycoproteins/*chemistry
Molecular Sequence Data
Oligosaccharides/*chemistry
Saposins
Spleen/*chemistry
Abstract: We have determined and compared the structures of the oligosaccharide moieties of saposin A, C and D purified from the spleen of a patient with Gaucher disease. These saposins, together with saposin B, are small glycoproteins, derived from separate domains of a single precursor, prosaposin, and are required for the lysosomal hydrolysis of various sphingolipids. The characteristic features of the oligosaccharide moieties of saposin A are (a) the predominance of a fucosylated trimannosyl core structure and (b) the occurrence of several different oligomannose-type and N-acetyllactosamine-type oligosaccharides. Saposin C contains (a) a predominance of oligomannose-type oligosaccharides and monoantennary oligosaccharides and (b) the presence of four different oligosaccharides having bisecting N-acetylglucosamine residues (found only in this saposin). Saposin D is distinguished by the occurrence of oligomannose-type oligosaccharides, which comprise nearly 90% of its total oligosaccharides. The possible reasons for the unique glycosylation of each saposin is discussed.
Notes: Ito, K
Takahashi, N
Takahashi, A
Shimada, I
Arata, Y
O'Brien, J S
Kishimoto, Y
NS-08682/NS/NINDS NIH HHS/United States
NS-13559/NS/NINDS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Eur J Biochem. 1993 Jul 1;215(1):171-9. doi: 10.1111/j.1432-1033.1993.tb18020.x.
Author Address: Department of Neurology, Nagoya University School of Medicine, Aichi, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1612
Author: Ito, R., Katsura, S., Shimada, H., Tsuchiya, H., Hada, M., Okumura, T., Sugawara, A. and Yokoyama, A.
Year: 2014
Title: TET3-OGT interaction increases the stability and the presence of OGT in chromatin
Journal: Genes Cells
Volume: 19
Issue: 1
Pages: 52-65
Epub Date: 2013/12/07
Date: Jan
Short Title: TET3-OGT interaction increases the stability and the presence of OGT in chromatin
Alternate Journal: Genes to cells : devoted to molecular & cellular mechanisms
ISSN: 1356-9597
DOI: 10.1111/gtc.12107
Accession Number: 24304661
Keywords: Animals
Cell Culture Techniques
Chromatin/*chemistry/metabolism
DNA-Binding Proteins/*chemistry/metabolism
Dioxygenases/*chemistry/metabolism
Humans
Mice
N-Acetylglucosaminyltransferases/*chemistry/metabolism
Protein Stability
Proto-Oncogene Proteins/*chemistry/metabolism
Abstract: Gene expression is controlled by alterations in the epigenome, including DNA methylation and histone modification. Recently, it was reported that 5-methylcytosine (5mC) is converted to 5-hydroxymethylcytosine (5hmC) by proteins in the ten-eleven translocation (TET) family. This conversion is believed to be part of the mechanism by which methylated DNA is demethylated. Moreover, histones undergo modifications such as phosphorylation and acetylation. In addition, modification with O-linked-N-acetylglucosamine (O-GlcNAc) by O-GlcNAc transferase (OGT) was recently identified as a novel histone modification. Herein, we focused on TET3, the regulation of which is still unclear. We attempted to elucidate the mechanism of its regulation by biochemical approaches. First, we conducted mass spectrometric analysis in combination with affinity purification of FLAG-TET3, which identified OGT as an important partner of TET3. Co-immunoprecipitation assays using a series of deletion mutants showed that the C-terminal H domain of TET3 was required for its interaction with OGT. Furthermore, we showed that TET3 is GlcNAcylated by OGT, although the GlcNAcylation did not affect the global hydroxylation of methylcytosine by TET3. Moreover, we showed that TET3 enhanced its localization to chromatin through the stabilization of OGT protein. Taken together, we showed a novel function of TET3 that likely supports the function of OGT.
Notes: 1365-2443
Ito, Ryo
Katsura, Shogo
Shimada, Hiroki
Tsuchiya, Hikaru
Hada, Masashi
Okumura, Tomoko
Sugawara, Akira
Yokoyama, Atsushi
Journal Article
Research Support, Non-U.S. Gov't
England
Genes Cells. 2014 Jan;19(1):52-65. doi: 10.1111/gtc.12107. Epub 2013 Dec 4.
Author Address: Department of Molecular Endocrinology, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, 980-8575, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1191
Author: Itoh, S., Hachisuka, A., Kawasaki, N., Hashii, N., Teshima, R., Hayakawa, T., Kawanishi, T. and Yamaguchi, T.
Year: 2008
Title: Glycosylation analysis of IgLON family proteins in rat brain by liquid chromatography and multiple-stage mass spectrometry
Journal: Biochemistry
Volume: 47
Issue: 38
Pages: 10132-54
Epub Date: 2008/08/30
Date: Sep 23
Short Title: Glycosylation analysis of IgLON family proteins in rat brain by liquid chromatography and multiple-stage mass spectrometry
Alternate Journal: Biochemistry
ISSN: 0006-2960
DOI: 10.1021/bi8009778
Accession Number: 18729387
Keywords: Amino Acid Sequence
Animals
Brain/*metabolism
Carrier Proteins/analysis/metabolism
Cell Adhesion Molecules/analysis/metabolism
Cell Adhesion Molecules, Neuronal/metabolism
Chromatography, Liquid/methods
GPI-Linked Proteins
Glycosylation
Immunoglobulin Subunits/analysis/metabolism
Lysosome-Associated Membrane Glycoproteins/analysis/metabolism
Male
Membrane Glycoproteins/*analysis/*metabolism
Molecular Sequence Data
*Multigene Family
Neural Cell Adhesion Molecules/metabolism
Rats
Rats, Wistar
Tandem Mass Spectrometry/methods
Abstract: IgLON family proteins, including limbic-associated membrane protein (LAMP), opioid-binding cell adhesion molecule (OBCAM), neurotrimin, and Kilon, are immunoglobulin (Ig) superfamily cell adhesion molecules. These molecules are composed of three Ig domains and a glycosylphosphatidylinositol (GPI) anchor and contain six or seven potential N-glycosylation sites. Although their glycosylations are supposed to be associated with the development of the central nervous system like other Ig superfamily proteins, they are still unknown because of difficulty in isolating individual proteins with a high degree of homology in performing carbohydrate analysis. In this study, we conducted simultaneous site-specific glycosylation analysis of rat brain IgLON proteins by liquid chromatography and multiple-stage mass spectrometry (LC-MS ( n )). The rat brain GPI-linked proteins were enriched and separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The four proteins were extracted from the gel, and subjected to LC-MS ( n ) after proteinase digestions. A set of glycopeptide MS data, including the mass spectrum, the mass spectrum in the selected ion monitoring mode, and the product ion spectra, was selected from all data based on carbohydrate-related ions in the MS/MS spectrum. The peptide portion and the carbohydrate structure were identified on the basis of peptide-related ion and carbohydrate-related ions, and the accurate mass. The site-specific glycosylations of four proteins were elucidated as follows. N-Glycans near the N-terminal were disialic acid-conjugated complex- and hybrid-type oligosaccharides. The first Ig domains were occupied by Man-5-9. Diverse oligosaccharides, including Lewis a/x-modified glycans, a brain-specific glycan known as BA-2, and Man-5, were found to be attached to the third Ig domain. Three common structures of glycans were found in the GPI moiety of LAMP, OBCAM, and neurotrimin.
Notes: 1520-4995
Itoh, Satsuki
Hachisuka, Akiko
Kawasaki, Nana
Hashii, Noritaka
Teshima, Reiko
Hayakawa, Takao
Kawanishi, Toru
Yamaguchi, Teruhide
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2008 Sep 23;47(38):10132-54. doi: 10.1021/bi8009778. Epub 2008 Aug 26.
Author Address: Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, 1-18-1, Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 909
Author: Ivanenkov, V. V., Murphy-Piedmonte, D. M. and Kirley, T. L.
Year: 2003
Title: Bacterial expression, characterization, and disulfide bond determination of soluble human NTPDase6 (CD39L2) nucleotidase: implications for structure and function
Journal: Biochemistry
Volume: 42
Issue: 40
Pages: 11726-35
Epub Date: 2003/10/08
Date: Oct 14
Short Title: Bacterial expression, characterization, and disulfide bond determination of soluble human NTPDase6 (CD39L2) nucleotidase: implications for structure and function
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi035137r
Accession Number: 14529283
Keywords: Amino Acid Sequence/genetics
Circular Dichroism
Cloning, Molecular
Cysteine/chemistry/genetics
Disulfides/*chemistry
Escherichia coli/*enzymology/*genetics
*Gene Expression Regulation, Bacterial
Genetic Vectors
Humans
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Folding
Protein Structure, Secondary
Pyrophosphatases/biosynthesis/*chemistry/*genetics/isolation & purification
Solubility
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Structure-Activity Relationship
Substrate Specificity/genetics
Sulfhydryl Compounds/chemistry
Abstract: The ectonucleoside triphosphate diphosphohydrolases (NTPDases) control extracellular nucleotide concentrations, thereby modulating many important biological responses, including blood clotting and pain perception. NTPDases1-4 are oligomeric integral membrane proteins, whereas NTPDase5 (CD39L4) and NTPDase6 (CD39L2) are soluble monomeric enzymes, making them more amenable to thorough structural and functional analyses than the membrane-bound forms. Therefore, we report here the bacterial expression, refolding, purification, and biochemical characterization of the soluble portion of human NTPDase6. Consistent with the enzyme expressed in mammalian cells, this recombinant NTPDase6 efficiently hydrolyzes GDP, IDP, and UDP (specific activity of approximately 50000 micromol mg(-1) h(-1)), with slower hydrolysis of CDP, ITP, GTP, CTP, ADP, and UTP and virtually no hydrolysis of ATP. The K(m) for GDP (130 +/- 30 microM) is similar to that determined for the soluble rat NTPDase6 expressed in mammalian cells. The secondary structure of the refolded enzyme was determined by circular dichroism to be 33% alpha-helix, 18% beta-sheet, and 49% random coil, consistent with the secondary structure predicted from the amino acid sequence of soluble NTPDase6. Four of the five cysteine residues in the soluble NTPDase6 are highly conserved among all the NTPDases, while the fifth residue is not. Mutation of this nonconserved cysteine resulted in an enzyme very similar to wild type in its enzymology and secondary structure, indicating that this cysteine exists as a free sulfhydryl and is not essential for structure or function. The disulfide pairing of the other four cysteine residues was determined as Cys(249)-Cys(280) and Cys(340)-Cys(354) by HPLC and mass spectral analysis of tryptic peptides. Due to conservation of these cysteine residues, these two disulfide bonds are likely to exist in all NTPDases. A structural model for NTPDase6, incorporating these and other findings obtained with other NTPDases, is proposed.
Notes: Ivanenkov, Vasily V
Murphy-Piedmonte, Deirdre M
Kirley, Terence L
HL59915/HL/NHLBI NIH HHS/United States
HL72382/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2003 Oct 14;42(40):11726-35. doi: 10.1021/bi035137r.
Author Address: Department of Pharmacology and Cell Biophysics, College of Medicine, University of Cincinnati, P.O. Box 670575, Cincinnati, Ohio 45267-0575, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 316
Author: Iwanaga, S., Nishimura, H., Kawabata, S., Kisiel, W., Hase, S. and Ikenaka, T.
Year: 1990
Title: A new trisaccharide sugar chain linked to a serine residue in the first EGF-like domain of clotting factors VII and IX and protein Z
Journal: Adv Exp Med Biol
Volume: 281
Pages: 121-31
Epub Date: 1990/01/01
Short Title: A new trisaccharide sugar chain linked to a serine residue in the first EGF-like domain of clotting factors VII and IX and protein Z
Alternate Journal: Advances in experimental medicine and biology
ISSN: 0065-2598 (Print)
0065-2598
DOI: 10.1007/978-1-4615-3806-6_12
Accession Number: 2129367
Keywords: Amino Acid Sequence
Animals
Blood Proteins/*chemistry
Carbohydrate Sequence
Cattle
Chromatography, High Pressure Liquid
Epidermal Growth Factor/*chemistry
Factor IX/*chemistry
Factor VII/*chemistry
Humans
Models, Structural
Molecular Sequence Data
Oligosaccharides/chemistry/*isolation & purification
Peptide Fragments/isolation & purification
Protein Conformation
Sequence Homology, Nucleic Acid
*Serine
Spectrometry, Mass, Fast Atom Bombardment
Abstract: Recently, we determined the complete amino acid sequence of bovine factor VII (Takeya, H. et al. (1988) J. Biol. Chem. 263, 14868-14877). In the course of the studies, we found an unknown serine derivative at position 52 in the first epidermal growth factor-like domain of factor VII. A pentapeptide isolated from the S-aminoethylated factor VII contained Ser-52, which could not be identified with a gas-phase sequencer. The same results were also obtained for a pentapeptide containing Ser-53 of factor IX and protein Z. Component sugar analysis revealed that the peptide contained 1 mol of glucose and 2 mol of xylose. This sugar component was also confirmed by high-resolution fast atom bombardment mass spectrometric analysis of the pentapeptide. The trisaccharide was released from the peptides by means of beta-elimination reaction and its reducing end was identified as pyridylamino-glucose. These results indicate the existence of a (Xyl2)Glc-Ser structure in factors VII, IX and protein Z. Similar results were obtained for human factors VII, IX and protein Z. This is the first report of a (Xyl2)-Glc-Ser structure in glycoproteins to our knowledge. The presence of the unique trisaccharide structure in factors VII, IX and protein Z leads us to anticipate its biological role in the tissue factor pathway.
Notes: Iwanaga, S
Nishimura, H
Kawabata, S
Kisiel, W
Hase, S
Ikenaka, T
HL35246/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Adv Exp Med Biol. 1990;281:121-31. doi: 10.1007/978-1-4615-3806-6_12.
Author Address: Department of Biology, Faculty of Science, Kyushu University, Fukuoka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 228
Author: Iwase, H., Ishii-Karakasa, I., Fujii, E., Hotta, K., Hiki, Y. and Kobayashi, Y.
Year: 1992
Title: Analysis of glycoform of O-glycan from human myeloma immunoglobulin A1 by gas-phase hydrazinolysis following pyridylamination of oligosaccharides
Journal: Anal Biochem
Volume: 206
Issue: 1
Pages: 202-5
Epub Date: 1992/10/01
Date: Oct
Short Title: Analysis of glycoform of O-glycan from human myeloma immunoglobulin A1 by gas-phase hydrazinolysis following pyridylamination of oligosaccharides
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/s0003-2697(05)80034-x
Accession Number: 1280920
Keywords: Amination
Aminopyridines/metabolism
Animals
Carbohydrate Sequence
Cattle
Chromatography, Gas/methods
Chromatography, High Pressure Liquid/methods
Humans
*Hydrazines
Immunoglobulins/*analysis
Molecular Sequence Data
Multiple Myeloma/immunology
Myeloma Proteins/*analysis
Oligosaccharides/*analysis
Polysaccharides/*analysis
alpha-Fetoproteins/analysis
Abstract: A comparative study was made on the glycoform of O-glycan from human myeloma immunoglobulin A1. By gas-phase hydrazinolysis, O-glycan was released from its hinge portion. The released oligosaccharide was pyridylaminated and separated by a two-dimensional analytical method of gel filtration and reverse-phase HPLC. Four major pyridylamino derivatives (P1-P4) were obtained. The neutral component (P4) among them was identified as Gal beta 1,3GalNAc-PA by cochromatography with an authentic standard pyridylamino sugar. The desialylation of the other components indicated the largest P1 and middle size P2 components possibly corresponded to a disialylated structure, NeuAc alpha 2,3Gal beta 1,3(NeuAc alpha 2,6)GalNAc-PA, and a monosialylated component, NeuAc alpha 2,3Gal beta 1,3GalNAc-PA, respectively. The structural assignment of P3 is still incomplete. Four similar components were also detected in bovine fetuin whose relative content (P1:P2: P3:P:4) was 16:43:19:22. The relative content (%) of P1-P4 (glycoform) in IgA1 from the healthy control was 10.1 +/- 3.3, 48.2 +/- 4.6, 7.0 +/- 2.6, and 34.7 +/- 4.5. The glycoform of O-glycan on IgA1 thus appears the same for any individual. Analysis of IgA1 myeloma protein indicated glycoforms distinct from those of the healthy controls. The relative content of these component could be classed as 2:8:0:90 (Type I, only one case designated as Kita), 5:24:3:68 (Type II, seven cases), and 9:41:5:45 (Type III, four cases). Thus, the results for IgA1 myeloma protein indicate that at least three glycoforms of O-glycan are possible for the IgA1 hinge structure. However, only one glycoform was found in the healthy controls.
Notes: Iwase, H
Ishii-Karakasa, I
Fujii, E
Hotta, K
Hiki, Y
Kobayashi, Y
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Biochem. 1992 Oct;206(1):202-5. doi: 10.1016/s0003-2697(05)80034-x.
Author Address: Department of Biochemistry, School of Medicine and Nursing, Kitasato University, Kanagawa, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 376
Author: Jackson, S. P. and Tjian, R.
Year: 1988
Title: O-glycosylation of eukaryotic transcription factors: implications for mechanisms of transcriptional regulation
Journal: Cell
Volume: 55
Issue: 1
Pages: 125-33
Epub Date: 1988/10/07
Date: Oct 7
Short Title: O-glycosylation of eukaryotic transcription factors: implications for mechanisms of transcriptional regulation
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/0092-8674(88)90015-3
Accession Number: 3139301
Keywords: Animals
Cattle
DNA/metabolism
DNA-Binding Proteins/metabolism
Galactosyltransferases/metabolism
Glycosylation
RNA Polymerase II/metabolism
Sp1 Transcription Factor
Transcription Factors/*metabolism
*Transcription, Genetic
Wheat Germ Agglutinins/metabolism
Abstract: Glycosylation is often regarded as being restricted to proteins confined to the cell surface or within the lumen of intracellular organelles. Here we show that the human RNA polymerase II transcription factor Sp1 bears multiple O-linked N-acetylglucosamine (GlcNAc) monosaccharide residues. The lectin wheat germ agglutinin specifically inhibits the transcriptional activation but not the DNA binding function of Sp1. Furthermore, many other RNA polymerase II transcription factors also bear terminal GlcNAc residues, whereas most nuclear proteins, including RNA polymerase I and III transcription factors tested, do not. In some cases, only a subset of the polypeptide species within a particular family of closely related RNA polymerase II factors appears to be glycosylated. Our findings raise the possibility that O-linked GlcNAc residues play a role in the mechanism or regulation of transcriptional activation of RNA polymerase II.
Notes: Jackson, S P
Tjian, R
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Cell. 1988 Oct 7;55(1):125-33. doi: 10.1016/0092-8674(88)90015-3.
Author Address: Howard Hughes Medical Institute, Department of Biochemistry, University of California, Berkeley 94720.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1686
Author: Jackson, V. A., del Toro, D., Carrasquero, M., Roversi, P., Harlos, K., Klein, R. and Seiradake, E.
Year: 2015
Title: Structural basis of latrophilin-FLRT interaction
Journal: Structure
Volume: 23
Issue: 4
Pages: 774-81
Epub Date: 2015/03/03
Date: Apr 7
Short Title: Structural basis of latrophilin-FLRT interaction
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2015.01.013
PMCID: PMC4396693
Accession Number: 25728924
Keywords: Amino Acid Sequence
Animals
Binding Sites
HEK293 Cells
HeLa Cells
Humans
Membrane Glycoproteins/chemistry/*metabolism
Mice
Molecular Sequence Data
Protein Binding
Receptors, G-Protein-Coupled/*chemistry/metabolism
Receptors, Peptide/*chemistry/metabolism
Abstract: Latrophilins, receptors for spider venom α-latrotoxin, are adhesion type G-protein-coupled receptors with emerging functions in synapse development. The N-terminal region binds the endogenous cell adhesion molecule FLRT, a major regulator of cortical and synapse development. We present crystallographic data for the mouse Latrophilin3 lectin and olfactomedin-like (Olf) domains, thereby revealing the Olf β-propeller fold and conserved calcium-binding site. We locate the FLRT-Latrophilin binding surfaces by a combination of sequence conservation analysis, point mutagenesis, and surface plasmon resonance experiments. In stripe assays, we show that wild-type Latrophilin3 and its high-affinity interactor FLRT2, but not the binding-impaired mutants we generated, promote HeLa cell adhesion. In contrast, cortical neurons expressing endogenous FLRTs are repelled by wild-type Latrophilin3 and not by the binding-impaired mutant. Taken together, we present molecular level insights into Latrophilin structure, its FLRT-binding mechanism, and a role for Latrophilin and FLRT that goes beyond a simply adhesive interaction.
Notes: 1878-4186
Jackson, Verity A
del Toro, Daniel
Carrasquero, Maria
Roversi, Pietro
Harlos, Karl
Klein, Rüdiger
Seiradake, Elena
G1000099/Medical Research Council/United Kingdom
G0900084/Medical Research Council/United Kingdom
MR/M000141/1/Medical Research Council/United Kingdom
MR/N00065X/1/Medical Research Council/United Kingdom
MR/L018039/1/Medical Research Council/United Kingdom
G1100525/Medical Research Council/United Kingdom
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Structure. 2015 Apr 7;23(4):774-81. doi: 10.1016/j.str.2015.01.013. Epub 2015 Feb 26.
Author Address: Department of Biochemistry, Oxford University, South Parks Road, Oxford OX1 3QU, UK.
Max-Planck Institute of Neurobiology, Am Klopferspitz 18, 82152 Munich-Martinsried, Germany.
Division of Structural Biology, Oxford University, Roosevelt Drive, Oxford OX3 7BN, UK.
Max-Planck Institute of Neurobiology, Am Klopferspitz 18, 82152 Munich-Martinsried, Germany; Munich Cluster for Systems Neurology (SyNergy), Feodor-Lynen-Straße 17, 81377 Munich, Germany.
Department of Biochemistry, Oxford University, South Parks Road, Oxford OX1 3QU, UK. Electronic address: elena.seiradake@bioch.ox.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1365
Author: Jacobsen, K. T. and Iverfeldt, K.
Year: 2011
Title: O-GlcNAcylation increases non-amyloidogenic processing of the amyloid-β precursor protein (APP)
Journal: Biochem Biophys Res Commun
Volume: 404
Issue: 3
Pages: 882-6
Epub Date: 2010/12/25
Date: Jan 21
Short Title: O-GlcNAcylation increases non-amyloidogenic processing of the amyloid-β precursor protein (APP)
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2010.12.080
Accession Number: 21182826
Keywords: Acetylglucosamine/analogs & derivatives/genetics/*metabolism/pharmacology
Acylation/drug effects/genetics
Alzheimer Disease/*enzymology/therapy
Amyloid beta-Protein Precursor/*metabolism
Cell Line, Tumor
Gene Knockdown Techniques
Humans
N-Acetylglucosaminyltransferases/genetics/*metabolism
Oximes/pharmacology
Phenylcarbamates/pharmacology
RNA, Small Interfering/genetics
beta-N-Acetylhexosaminidases/antagonists & inhibitors/genetics/*metabolism
Abstract: The amyloid-β precursor protein (APP) was shown to be O-GlcNAcylated 15 years ago, but the effect of this modification on APP processing and formation of the Alzheimer's disease associated amyloid-β (Aβ) peptide has so far not been investigated. Here, we demonstrate with pharmacological tools or siRNA that O-GlcNAcase and O-GlcNAc transferase regulate the level of O-GlcNAcylated APP. We also show that O-GlcNAcylation increases non-amyloidogenic α-secretase processing, resulting in increased levels of the neuroprotective sAPPα fragment and decreased Aβ secretion. Our results implicate O-GlcNAcylation as a potential therapeutic target for Alzheimer's disease.
Notes: 1090-2104
Jacobsen, Kristin T
Iverfeldt, Kerstin
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2011 Jan 21;404(3):882-6. doi: 10.1016/j.bbrc.2010.12.080. Epub 2010 Dec 21.
Author Address: Department of Neurochemistry, Stockholm University, SE-10691 Stockholm, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 546
Author: Jaeken, J., Schachter, H., Carchon, H., De Cock, P., Coddeville, B. and Spik, G.
Year: 1994
Title: Carbohydrate deficient glycoprotein syndrome type II: a deficiency in Golgi localised N-acetyl-glucosaminyltransferase II
Journal: Arch Dis Child
Volume: 71
Issue: 2
Pages: 123-7
Epub Date: 1994/08/01
Date: Aug
Short Title: Carbohydrate deficient glycoprotein syndrome type II: a deficiency in Golgi localised N-acetyl-glucosaminyltransferase II
Alternate Journal: Archives of disease in childhood
ISSN: 0003-9888 (Print)
0003-9888
DOI: 10.1136/adc.71.2.123
PMCID: PMC1029941
Accession Number: 7944531
Keywords: Carbohydrate Metabolism, Inborn Errors/blood/*classification/enzymology
Carbohydrate Sequence
Child
Fibroblasts/enzymology
Glycoproteins/*metabolism
Glycosylation
Golgi Apparatus/*enzymology
Humans
Isoelectric Focusing
Male
Molecular Sequence Data
N-Acetylglucosaminyltransferases/*deficiency
Transferrin/analysis
Abstract: The carbohydrate deficient glycoprotein (CDG) syndromes are a family of genetic multisystemic disorders with severe nervous system involvement. This report is on a child with a CDG syndrome that differs from the classical picture but is very similar to a patient reported in 1991. Both these patients are therefore designated CDG syndrome type II. Compared with type I patients they have a more severe psychomotor retardation but no peripheral neuropathy nor cerebellar hypoplasia. The serum transferrin isoform pattern obtained by isoelectric focusing showed disialotransferrin as the major fraction. The serum disialotransferrin, studied in the present patient, contained two moles of truncated monoantennary Sialyl-Gal-GlcNAc-Man(alpha 1-->3)[Man(alpha 1-->6)]Man(beta 1-->4)GlcNAc (beta 1-->4)GlcNAc-Asn per mole of transferrin. A profoundly deficient activity of the Golgi enzyme N-acetylglucosaminyltransferase II (EC 2.4.1.143) was demonstrated in fibroblasts.
Notes: 1468-2044
Jaeken, J
Schachter, H
Carchon, H
De Cock, P
Coddeville, B
Spik, G
Case Reports
Journal Article
Research Support, Non-U.S. Gov't
Arch Dis Child. 1994 Aug;71(2):123-7. doi: 10.1136/adc.71.2.123.
Author Address: Department of Paediatrics, University Hospital Gasthuisberg, Leuven, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 822
Author: Jahnel, R., Dreger, M., Gillen, C., Bender, O., Kurreck, J. and Hucho, F.
Year: 2001
Title: Biochemical characterization of the vanilloid receptor 1 expressed in a dorsal root ganglia derived cell line
Journal: Eur J Biochem
Volume: 268
Issue: 21
Pages: 5489-96
Epub Date: 2001/10/31
Date: Nov
Short Title: Biochemical characterization of the vanilloid receptor 1 expressed in a dorsal root ganglia derived cell line
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1033.2001.02500.x
Accession Number: 11683872
Keywords: Animals
Asparagine/genetics
Cell Line
Cell Membrane/metabolism
Cloning, Molecular
Ganglia, Spinal/cytology/*physiology
Glycoproteins/chemistry/genetics/metabolism
Glycosylation
Mice
Mutagenesis, Site-Directed
Protein Denaturation
Protein Folding
Rats
Receptors, Drug/chemistry/*genetics/*metabolism
Transfection
Abstract: The vanilloid receptor VR1 is an ion channel predominantly expressed by primary sensory neurons involved in nociception. Here we describe its biochemical properties and assess the subcellular localization, the glycosylation state and the quaternary structure of VR1 expressed in HEK293 cells and in the DRG-derived cell line F-11 (N18TG2 mouse neuroblastoma x rat dorsal root ganglia, hybridoma). VR1 was found to be glycosylated in both cell types. Of the five potential N-glycosylation sites, the predicted transient receptor potential channel-like transmembrane folding proposes N604 is localized extracellularly. We used site-directed mutagenesis to mutate the Asn at position 604 to Thr. This mutated VR1 was not glycosylated, confirming the extracellular location of N604 and its role as the exclusive site of glycosylation of the VR1 protein. VR1 occured in high molecular mass complexes as assessed by blue native PAGE. In the presence of limited amounts of SDS dimers, trimers and tetramers of VR1 were observed, consistent with the predicted tetrameric quaternary structure of the receptor. Cross-linking with dimethyladipimidate yielded almost exclusively dimers. Whereas VR1 localized both to the plasma membrane and to intracellular membranes in HEK293 cells, it localized predominantly to the plasma membrane in F-11 cells. Using confocal laserscanning microscopy, we observed an enrichment of anti-VR1 immunoreactivity in neurite-like structures of F-11 cells. In the light of conflicting literature data on biochemical characteristics of VR1, our data suggest that dorsal root ganglion-derived F-11 cells provide a powerful experimental system for the study of VR1 biochemistry.
Notes: Jahnel, R
Dreger, M
Gillen, C
Bender, O
Kurreck, J
Hucho, F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 2001 Nov;268(21):5489-96. doi: 10.1046/j.1432-1033.2001.02500.x.
Author Address: Arbeitsgruppe Neurochemie and Arbeitsgruppe Molekulare Medizin, Institut für Chemie-Biochemie, Freie Universität Berlin, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 733
Author: Jakob, C. G., Lewinski, K., Kuciel, R., Ostrowski, W. and Lebioda, L.
Year: 2000
Title: Crystal structure of human prostatic acid phosphatase
Journal: Prostate
Volume: 42
Issue: 3
Pages: 211-8
Epub Date: 2000/01/19
Date: Feb 15
Short Title: Crystal structure of human prostatic acid phosphatase
Alternate Journal: The Prostate
ISSN: 0270-4137 (Print)
0270-4137
DOI: 10.1002/(sici)1097-0045(20000215)42:3<211::aid-pros7>3.0.co;2-u
Accession Number: 10639192
Keywords: Acid Phosphatase/*chemistry/metabolism
Binding Sites
Carbohydrates/chemistry
Crystallography, X-Ray
Glycosylation
Humans
Male
Models, Molecular
Prostate/*enzymology
Protein Conformation
Protein Processing, Post-Translational
Semen/enzymology
Abstract: BACKGROUND: Prostatic acid phosphatase (hPAP) is a major product of the human prostate gland, yet its physiological substrate remains unknown. METHODS: Human PAP, purified from semen, was crystallized using polyethylene glycol as the precipitant and its crystal structure was determined using X-ray diffraction. The structure was refined at 3.1 A resolution to R = 16% and R(free) = 27%. RESULTS: The structure of hPAP is similar to that of other known histidine phosphatases, and the positions of its catalytic residues are conserved. N-linked carbohydrates are present at each of the possible glycosylation sites. It appears that high-mannose chains are attached to Asn 62 and Asp 301, while complex chains are at Asn 188. CONCLUSIONS: The similarity of the three-dimensional structures of rat PAP and human PAP indicates that the mechanistic analyses of the catalytic mechanism proposed for the rat enzyme should be extended to the human enzyme without reservations. The crystallographic data allowed the correlation of attachment sites of N-linked carbohydrate chains with a given carbohydrate type. The carbohydrates of the protein produced in the prostate cells and in the baculovirus expression system appear to differ at the site of complex carbohydrates attachment.
Notes: Jakob, C G
Lewinski, K
Kuciel, R
Ostrowski, W
Lebioda, L
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
Prostate. 2000 Feb 15;42(3):211-8. doi: 10.1002/(sici)1097-0045(20000215)42:3<211::aid-pros7>3.0.co;2-u.
Author Address: Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1700
Author: Jang, I., Kim, H. B., Seo, H., Kim, J. Y., Choi, H., Yoo, J. S., Kim, J. W. and Cho, J. W.
Year: 2015
Title: O-GlcNAcylation of eIF2α regulates the phospho-eIF2α-mediated ER stress response
Journal: Biochim Biophys Acta
Volume: 1853
Issue: 8
Pages: 1860-9
Epub Date: 2015/05/06
Date: Aug
Short Title: O-GlcNAcylation of eIF2α regulates the phospho-eIF2α-mediated ER stress response
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbamcr.2015.04.017
Accession Number: 25937070
Keywords: Acetylglucosamine/*metabolism
Animals
*Endoplasmic Reticulum Stress/genetics
Eukaryotic Initiation Factor-2/genetics/*metabolism
HEK293 Cells
Hep G2 Cells
Humans
Mice
Mice, Inbred C57BL
N-Acetylglucosaminyltransferases/*metabolism
Phosphorylation
Protein Processing, Post-Translational/genetics
Unfolded Protein Response
eIF-2 Kinase/*metabolism
Apoptosis
Chop
ER stress
O-GlcNAc
eIF2α
p-eIF2α
Abstract: O-GlcNAcylation is highly involved in cellular stress responses including the endoplasmic reticulum (ER) stress response. For example, glucosamine-induced flux through the hexosamine biosynthetic pathway can promote ER stress and ER stress inducers can change the total cellular level of O-GlcNAcylation. However, it is largely unknown which component(s) of the unfolded protein response (UPR) is directly regulated by O-GlcNAcylation. In this study, eukaryotic translation initiation factor 2α (eIF2α), a major branch of the UPR, was O-GlcNAcylated at Ser 219, Thr 239, and Thr 241. Upon ER stress, eIF2α is phosphorylated at Ser 51 by phosphorylated PKR-like ER kinase and this inhibits global translation initiation, except for that of specific mRNAs, including activating transcription factor 4, that induce stress-responsive genes such as C/EBP homologous protein (CHOP). Hyper-O-GlcNAcylation induced by O-GlcNAcase inhibitor (thiamet-G) treatment or O-GlcNAc transferase (OGT) overexpression hindered phosphorylation of eIF2α at Ser 51. The level of O-GlcNAcylation of eIF2α was changed by dithiothreitol treatment dependent on its phosphorylation at Ser 51. Point mutation of the O-GlcNAcylation sites of eIF2α increased its phosphorylation at Ser 51 and CHOP expression and resulted in increased apoptosis upon ER stress. These results suggest that O-GlcNAcylation of eIF2α affects its phosphorylation at Ser 51 and influences CHOP-mediated cell death. This O-GlcNAcylation of eIF2α was reproduced in thiamet-G-injected mouse liver. In conclusion, proper regulation of O-GlcNAcylation and phosphorylation of eIF2α is important to maintain cellular homeostasis upon ER stress.
Notes: Jang, Insook
Kim, Han Byeol
Seo, Hojoong
Kim, Jin Young
Choi, Hyeonjin
Yoo, Jong Shin
Kim, Jae-woo
Cho, Jin Won
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2015 Aug;1853(8):1860-9. doi: 10.1016/j.bbamcr.2015.04.017. Epub 2015 Apr 30.
Author Address: Department of Integrated OMICS for Biomedical Science, Yonsei University, Seoul 120-749, Republic of Korea.
Division of Mass Spectrometry Research, Korea Basic Science Institute, 162 Yeongudanjiro, Ochang, Cheongju 363-883, Republic of Korea.
Department of Biochemistry and Molecular Biology, Integrated Genomic Research Center for Metabolic Regulation, Institute of Genetic Science, Yonsei University College of Medicine, Seoul 120-749, Republic of Korea.
Brain Korea 21 PLUS Project for Medical Science, Yonsei University, Seoul 120-749, Republic of Korea.
Department of Integrated OMICS for Biomedical Science, Yonsei University, Seoul 120-749, Republic of Korea. Electronic address: chojw311@yonsei.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1599
Author: Jank, T., Bogdanović, X., Wirth, C., Haaf, E., Spoerner, M., Böhmer, K. E., Steinemann, M., Orth, J. H., Kalbitzer, H. R., Warscheid, B., Hunte, C. and Aktories, K.
Year: 2013
Title: A bacterial toxin catalyzing tyrosine glycosylation of Rho and deamidation of Gq and Gi proteins
Journal: Nat Struct Mol Biol
Volume: 20
Issue: 11
Pages: 1273-80
Epub Date: 2013/10/22
Date: Nov
Short Title: A bacterial toxin catalyzing tyrosine glycosylation of Rho and deamidation of Gq and Gi proteins
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.2688
Accession Number: 24141704
Keywords: Amino Acid Sequence
Animals
Bacterial Toxins/*chemistry/*metabolism
Crystallography, X-Ray
Glycosylation
Humans
Magnetic Resonance Spectroscopy
Models, Molecular
Molecular Sequence Data
Photorhabdus/*enzymology
Protein Conformation
Tyrosine/*metabolism
Uridine Diphosphate N-Acetylglucosamine/*chemistry/*metabolism
rho GTP-Binding Proteins/*metabolism
Abstract: Entomopathogenic Photorhabdus asymbiotica is an emerging pathogen in humans. Here, we identified a P. asymbiotica protein toxin (PaTox), which contains a glycosyltransferase and a deamidase domain. PaTox mono-O-glycosylates Y32 (or Y34) of eukaryotic Rho GTPases by using UDP-N-acetylglucosamine (UDP-GlcNAc). Tyrosine glycosylation inhibits Rho activation and prevents interaction with downstream effectors, resulting in actin disassembly, inhibition of phagocytosis and toxicity toward insects and mammalian cells. The crystal structure of the PaTox glycosyltransferase domain in complex with UDP-GlcNAc determined at 1.8-Å resolution represents a canonical GT-A fold and is the smallest glycosyltransferase toxin known. (1)H-NMR analysis identifies PaTox as a retaining glycosyltransferase. The glutamine-deamidase domain of PaTox blocks GTP hydrolysis of heterotrimeric Gαq/11 and Gαi proteins, thereby activating RhoA. Thus, PaTox hijacks host GTPase signaling in a bidirectional manner by deamidation-induced activation and glycosylation-induced inactivation of GTPases.
Notes: 1545-9985
Jank, Thomas
Bogdanović, Xenia
Wirth, Christophe
Haaf, Erik
Spoerner, Michael
Böhmer, Kira E
Steinemann, Marcus
Orth, Joachim H C
Kalbitzer, Hans Robert
Orcid: 0000000265142355
Warscheid, Bettina
Hunte, Carola
Aktories, Klaus
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2013 Nov;20(11):1273-80. doi: 10.1038/nsmb.2688. Epub 2013 Oct 20.
Author Address: Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, Albert-Ludwigs-Universität Freiburg, Freiburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1523
Author: Jansen, S., Perrakis, A., Ulens, C., Winkler, C., Andries, M., Joosten, R. P., Van Acker, M., Luyten, F. P., Moolenaar, W. H. and Bollen, M.
Year: 2012
Title: Structure of NPP1, an ectonucleotide pyrophosphatase/phosphodiesterase involved in tissue calcification
Journal: Structure
Volume: 20
Issue: 11
Pages: 1948-59
Epub Date: 2012/10/09
Date: Nov 7
Short Title: Structure of NPP1, an ectonucleotide pyrophosphatase/phosphodiesterase involved in tissue calcification
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126
DOI: 10.1016/j.str.2012.09.001
Accession Number: 23041369
Keywords: Amino Acid Sequence
*Calcinosis
Models, Molecular
Molecular Sequence Data
Phosphoric Diester Hydrolases/*chemistry/metabolism
Protein Conformation
Pyrophosphatases/*chemistry/metabolism
Substrate Specificity
Abstract: Ectonucleotide pyrophosphatase/phosphodiesterase-1 (NPP1) converts extracellular nucleotides into inorganic pyrophosphate, whereas its close relative NPP2/autotaxin hydrolyzes lysophospholipids. NPP1 regulates calcification in mineralization-competent tissues, and a lack of NPP1 function underlies calcification disorders. Here, we show that NPP1 forms homodimers via intramembrane disulfide bonding, but is also processed intracellularly to a secreted monomer. The structure of secreted NPP1 reveals a characteristic bimetallic active site and a nucleotide-binding groove, but it lacks the lipid-binding pocket and open tunnel present in NPP2. A loop adjacent to the nucleotide-binding site, which is disordered in NPP2, is well ordered in NPP1 and might promote nucleotide binding. Remarkably, the N-terminal somatomedin B-like domains of NPP1, unlike those in NPP2, are flexible and do not contact the catalytic domain. Our results provide a structural basis for the nucleotide pyrophosphatase activity of NPP1 and help to understand how disease-causing mutations may affect NPP1 structure and function.
Notes: 1878-4186
Jansen, Silvia
Perrakis, Anastassis
Ulens, Chris
Winkler, Claudia
Andries, Maria
Joosten, Robbie P
Van Acker, Maarten
Luyten, Frank P
Moolenaar, Wouter H
Bollen, Mathieu
Journal Article
Research Support, Non-U.S. Gov't
United States
Structure. 2012 Nov 7;20(11):1948-59. doi: 10.1016/j.str.2012.09.001. Epub 2012 Oct 4.
Author Address: Laboratory of Biosignaling and Therapeutics, Department of Cellular and Molecular Medicine, University of Leuven, 3000 Leuven, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1254
Author: Janssen, B. J., Gomes, L., Koning, R. I., Svergun, D. I., Koster, A. J., Fritzinger, D. C., Vogel, C. W. and Gros, P.
Year: 2009
Title: Insights into complement convertase formation based on the structure of the factor B-cobra venom factor complex
Journal: Embo j
Volume: 28
Issue: 16
Pages: 2469-78
Epub Date: 2009/07/04
Date: Aug 19
Short Title: Insights into complement convertase formation based on the structure of the factor B-cobra venom factor complex
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2009.184
PMCID: PMC2735180
Accession Number: 19574954
Keywords: Animals
Cell Line
Complement C3-C5 Convertases/chemistry/*metabolism
Complement Factor B/*chemistry/genetics/*metabolism
Crystallography, X-Ray
Elapid Venoms/*chemistry/genetics/isolation & purification/*metabolism
Elapidae/*metabolism
Enzyme Activation
Gene Expression
Humans
Models, Molecular
Mutation
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Abstract: Immune protection by the complement system critically depends on assembly of C3 convertases on the surface of pathogens and altered host cells. These short-lived protease complexes are formed through pro-convertases, which for the alternative pathway consist of the complement component C3b and the pro-enzyme factor B (FB). Here, we present the crystal structure at 2.2-A resolution, small-angle X-ray scattering and electron microscopy (EM) data of the pro-convertase formed by human FB and cobra venom factor (CVF), a potent homologue of C3b that generates more stable convertases. FB is loaded onto CVF through its pro-peptide Ba segment by specific contacts, which explain the specificity for the homologous C3b over the native C3 and inactive products iC3b and C3c. The protease segment Bb binds the carboxy terminus of CVF through the metal-ion dependent adhesion site of the Von Willebrand factor A-type domain. A possible dynamic equilibrium between a 'loading' and 'activation' state of the pro-convertase may explain the observed difference between the crystal structure of CVFB and the EM structure of C3bB. These insights into formation of convertases provide a basis for further development of complement therapeutics.
Notes: 1460-2075
Janssen, Bert J C
Gomes, Lucio
Koning, Roman I
Svergun, Dmitri I
Koster, Abraham J
Fritzinger, David C
Vogel, Carl-Wilhelm
Gros, Piet
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
EMBO J. 2009 Aug 19;28(16):2469-78. doi: 10.1038/emboj.2009.184. Epub 2009 Jul 2.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1124
Author: Janssen, B. J., Halff, E. F., Lambris, J. D. and Gros, P.
Year: 2007
Title: Structure of compstatin in complex with complement component C3c reveals a new mechanism of complement inhibition
Journal: J Biol Chem
Volume: 282
Issue: 40
Pages: 29241-7
Epub Date: 2007/08/09
Date: Oct 5
Short Title: Structure of compstatin in complex with complement component C3c reveals a new mechanism of complement inhibition
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M704587200
Accession Number: 17684013
Keywords: Binding Sites
Complement C3c/*chemistry
Complement Inactivator Proteins/*pharmacology
Humans
Macroglobulins/chemistry
Models, Molecular
Molecular Conformation
Peptides/chemistry
Peptides, Cyclic/*chemistry/metabolism
Plasma/metabolism
Protein Binding
Protein Structure, Tertiary
Substrate Specificity
Abstract: Undesired complement activation is a major cause of tissue injury in various pathological conditions and contributes to several immune complex diseases. Compstatin, a 13-residue peptide, is an effective inhibitor of the activation of complement component C3 and thus blocks a central and crucial step in the complement cascade. The precise binding site on C3, the structure in the bound form, and the exact mode of action of compstatin are unknown. Here we present the crystal structure of compstatin in complex with C3c, a major proteolytic fragment of C3. The structure reveals that the compstatin-binding site is formed by the macroglobulin (MG) domains 4 and 5. This binding site is part of the structurally stable MG-ring formed by domains MG 1-6 and is far away from any other known binding site on C3. Compstatin does not alter the conformation of C3c, whereas compstatin itself undergoes a large conformational change upon binding. We propose a model in which compstatin sterically hinders the access of the substrate C3 to the convertase complexes, thus blocking complement activation and amplification. These insights are instrumental for further development of compstatin as a potential therapeutic.
Notes: Janssen, Bert J C
Halff, Els F
Lambris, John D
Gros, Piet
AI30040/AI/NIAID NIH HHS/United States
GM069736/GM/NIGMS NIH HHS/United States
GM62134/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2007 Oct 5;282(40):29241-7. doi: 10.1074/jbc.M704587200. Epub 2007 Aug 6.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Sciences, Utrecht University, 3584 CH Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1346
Author: Janssen, B. J., Robinson, R. A., Pérez-Brangulí, F., Bell, C. H., Mitchell, K. J., Siebold, C. and Jones, E. Y.
Year: 2010
Title: Structural basis of semaphorin-plexin signalling
Journal: Nature
Volume: 467
Issue: 7319
Pages: 1118-22
Epub Date: 2010/09/30
Date: Oct 28
Short Title: Structural basis of semaphorin-plexin signalling
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature09468
PMCID: PMC3587840
NIHMSID: EMS51603
Accession Number: 20877282
Keywords: Animals
Antigens, CD/chemistry/genetics/metabolism
Binding Sites
Cell Adhesion Molecules/*chemistry/genetics/*metabolism
Cell Communication
Crystallography, X-Ray
Humans
Ligands
Mice
Mice, Inbred C57BL
Models, Molecular
NIH 3T3 Cells
Nerve Tissue Proteins/*chemistry/genetics/*metabolism
Protein Binding
Protein Structure, Tertiary
Receptors, Cell Surface/chemistry/genetics/metabolism
Semaphorins/*chemistry/genetics/*metabolism
*Signal Transduction
Structure-Activity Relationship
Abstract: Cell-cell signalling of semaphorin ligands through interaction with plexin receptors is important for the homeostasis and morphogenesis of many tissues and is widely studied for its role in neural connectivity, cancer, cell migration and immune responses. SEMA4D and Sema6A exemplify two diverse vertebrate, membrane-spanning semaphorin classes (4 and 6) that are capable of direct signalling through members of the two largest plexin classes, B and A, respectively. In the absence of any structural information on the plexin ectodomain or its interaction with semaphorins the extracellular specificity and mechanism controlling plexin signalling has remained unresolved. Here we present crystal structures of cognate complexes of the semaphorin-binding regions of plexins B1 and A2 with semaphorin ectodomains (human PLXNB1(1-2)-SEMA4D(ecto) and murine PlxnA2(1-4)-Sema6A(ecto)), plus unliganded structures of PlxnA2(1-4) and Sema6A(ecto). These structures, together with biophysical and cellular assays of wild-type and mutant proteins, reveal that semaphorin dimers independently bind two plexin molecules and that signalling is critically dependent on the avidity of the resulting bivalent 2:2 complex (monomeric semaphorin binds plexin but fails to trigger signalling). In combination, our data favour a cell-cell signalling mechanism involving semaphorin-stabilized plexin dimerization, possibly followed by clustering, which is consistent with previous functional data. Furthermore, the shared generic architecture of the complexes, formed through conserved contacts of the amino-terminal seven-bladed β-propeller (sema) domains of both semaphorin and plexin, suggests that a common mode of interaction triggers all semaphorin-plexin based signalling, while distinct insertions within or between blades of the sema domains determine binding specificity.
Notes: 1476-4687
Janssen, Bert J C
Robinson, Ross A
Pérez-Brangulí, Francesc
Bell, Christian H
Mitchell, Kevin J
Siebold, Christian
Jones, E Yvonne
A3964/Cancer Research UK/United Kingdom
G0700232(82098)/Medical Research Council/United Kingdom
A10976/Cancer Research UK/United Kingdom
G0900084/Medical Research Council/United Kingdom
082301/Wellcome Trust/United Kingdom
G0700232/Medical Research Council/United Kingdom
Wellcome Trust/United Kingdom
10976/Cancer Research UK/United Kingdom
A5261/Cancer Research UK/United Kingdom
083111/Wellcome Trust/United Kingdom
G9900061/Medical Research Council/United Kingdom
G9900061(69203)/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Nature. 2010 Oct 28;467(7319):1118-22. doi: 10.1038/nature09468. Epub 2010 Sep 26.
Author Address: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford OX3 7BN, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 613
Author: Jaques, A. J., Opdenakker, G., Rademacher, T. W., Dwek, R. A. and Zamze, S. E.
Year: 1996
Title: The glycosylation of Bowes melanoma tissue plasminogen activator: lectin mapping, reaction with anti-L2/HNK-1 antibodies and the presence of sulphated/glucuronic acid containing glycans
Journal: Biochem J
Volume: 316 ( Pt 2)
Issue: Pt 2
Pages: 427-37
Epub Date: 1996/06/01
Date: Jun 1
Short Title: The glycosylation of Bowes melanoma tissue plasminogen activator: lectin mapping, reaction with anti-L2/HNK-1 antibodies and the presence of sulphated/glucuronic acid containing glycans
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj3160427
PMCID: PMC1217368
Accession Number: 8687384
Keywords: Antibodies, Monoclonal/immunology
Blotting, Western
CD57 Antigens/immunology
Carbohydrate Sequence
Chromatography, Affinity
Chromatography, Gel
Chromatography, Ion Exchange
Electrophoresis, Paper
Epitopes/chemistry
Gas Chromatography-Mass Spectrometry
Glucuronates/analysis
Glycosylation
Lectins/metabolism
Melanoma, Experimental/*chemistry
Molecular Sequence Data
N-Acetylneuraminic Acid
Oligosaccharides/analysis/chemistry
Polysaccharides/*analysis
Sialic Acids/metabolism
Sulfates/analysis
Tissue Plasminogen Activator/*chemistry/metabolism
Tumor Cells, Cultured
Abstract: The glycosylation of tissue plasminogen activator (t-PA) obtained from the Bowes melanoma cell line was re-examined using methods of serial lectin affinity chromatography coupled with Bio-Gel P-4 gel filtration chromatography and exoglycosidase sequencing. This study clarified an earlier discrepancy in the literature and confirmed that the major complex N-linked glycans on Bowes t-PA that carry sialic acid as their sole charged group are bi-antennary, core fucosylated, with terminal N-acetylgalactosamine residues. We also report the characterization of a series of related and previously unidentified sialylated glycans. Further we show that Bowes t-PA expresses glucuronic acid/sulphate containing N-linked glycans and is recognized by anti-carbohydrate L2/HNK-1 monoclonal antibodies. The presence on Bowes t-PA of glycans associated primarily with the nervous system is consistent with its expression in a cell line of neuroectodermal origin.
Notes: 1470-8728
Jaques, A J
Opdenakker, G
Rademacher, T W
Dwek, R A
Zamze, S E
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1996 Jun 1;316 ( Pt 2)(Pt 2):427-37. doi: 10.1042/bj3160427.
Author Address: Biochemistry Department, Oxford University, U.K.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 809
Author: Jassal, R., Jenkins, N., Charlwood, J., Camilleri, P., Jefferis, R. and Lund, J.
Year: 2001
Title: Sialylation of human IgG-Fc carbohydrate by transfected rat alpha2,6-sialyltransferase
Journal: Biochem Biophys Res Commun
Volume: 286
Issue: 2
Pages: 243-9
Epub Date: 2001/08/14
Date: Aug 17
Short Title: Sialylation of human IgG-Fc carbohydrate by transfected rat alpha2,6-sialyltransferase
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1006/bbrc.2001.5382
Accession Number: 11500028
Keywords: Animals
CHO Cells
Carbohydrates/analysis
Chromatography, High Pressure Liquid
Complement Activation
Cricetinae
Glycosylation
Humans
Immunoglobulin Fc Fragments/*chemistry
Immunoglobulin G/*chemistry/genetics/immunology
K562 Cells
Mutation
N-Acetylneuraminic Acid/analysis
Nitrohydroxyiodophenylacetate/immunology
Oligosaccharides/analysis
Rats
Sialyltransferases/*genetics
Superoxides/metabolism
Transfection
U937 Cells
Abstract: A recombinant IgG3 antibody with Phe-243 replaced by Ala (FA243) was expressed in a CHO-K1 parental cell line. The resulting IgG-Fc-linked carbohydrate was significantly alpha2,3-sialylated (53% of glycans), as indicated by normal- and reverse-phase HPLC analyses. Following transfection of a rat alpha2,6-sialyltransferase gene into this parental cell line, IgG-Fc-linked glycans were sialylated (60% of glycans) such that the ratio of alpha2,6- to alpha2,3-linked sialic acid was 0.9:1.0. By comparison, the wild-type IgG3 (F243) is minimally sialylated (2-3% alpha2,3-linked), thus suggesting that sialylation is controlled primarily by the protein structure local to the carbohydrate and that the two sialyltransferases compete to sialylate the nascent oligosaccharide. The additional alpha2,6-sialylation affected the function of the recombinant antibody. FA243 IgG3 having both alpha2,6 and alpha2,3-sialylation restored recognition to wild-type IgG3 levels for human FcgammaRI, FcgammaRII, and target cell lysis by complement. We discuss how sialylation linkage could modulate IgG function.
Notes: Jassal, R
Jenkins, N
Charlwood, J
Camilleri, P
Jefferis, R
Lund, J
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2001 Aug 17;286(2):243-9. doi: 10.1006/bbrc.2001.5382.
Author Address: Department of Biological Sciences, De Montfort University, Leicester, LE1 9BH, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 333
Author: Jefferis, R., Lund, J., Mizutani, H., Nakagawa, H., Kawazoe, Y., Arata, Y. and Takahashi, N.
Year: 1990
Title: A comparative study of the N-linked oligosaccharide structures of human IgG subclass proteins
Journal: Biochem J
Volume: 268
Issue: 3
Pages: 529-37
Epub Date: 1990/06/15
Date: Jun 15
Short Title: A comparative study of the N-linked oligosaccharide structures of human IgG subclass proteins
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2680529
PMCID: PMC1131471
Accession Number: 2363690
Keywords: Carbohydrate Sequence
Chromatography, High Pressure Liquid
Humans
Immunoglobulin G/*analysis/classification
Molecular Sequence Data
Multiple Myeloma/metabolism
Oligosaccharides/*analysis
Paraproteins/*analysis/classification
Abstract: Quantitative oligosaccharide profiles were determined for each of 18 human IgG paraproteins representing the four subclasses. Each paraprotein exhibits a unique profile that may be substantially different from that observed for polyclonal IgG. The IgG2 and some IgG3 proteins analysed exhibit a predominance of oligosaccharide moieties having galactose on the Man(alpha 1----3) arm rather than the Man(alpha 1----6) arm; it was previously held that galactosylation of the Man(alpha 1----6) arm is preferred, as observed for IgG1, IgG4 and polyclonal IgG. An IgG4 protein is reported that has galactosylated Man(alpha 1----3) and Man(alpha 1----6) arms on both Fc-localized carbohydrate moieties; previous findings suggested that such fully glycosylated structures could not be accommodated within the internal space of the C gamma 2 domains. Unusual monoantennary oligosaccharides present in IgG2 and IgG3 proteins were isolated and their structures determined.
Notes: 1470-8728
Jefferis, R
Lund, J
Mizutani, H
Nakagawa, H
Kawazoe, Y
Arata, Y
Takahashi, N
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1990 Jun 15;268(3):529-37. doi: 10.1042/bj2680529.
Author Address: Department of Immunology, University of Birmingham Medical School, Edgbaston, U.K.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1082
Author: Jensen, A. G., Chemali, M., Chapel, A., Kieffer-Jaquinod, S., Jadot, M., Garin, J. and Journet, A.
Year: 2007
Title: Biochemical characterization and lysosomal localization of the mannose-6-phosphate protein p76 (hypothetical protein LOC196463)
Journal: Biochem J
Volume: 402
Issue: 3
Pages: 449-58
Epub Date: 2006/11/16
Date: Mar 15
Short Title: Biochemical characterization and lysosomal localization of the mannose-6-phosphate protein p76 (hypothetical protein LOC196463)
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20061205
PMCID: PMC1863569
Accession Number: 17105447
Keywords: Animals
Biomarkers
Cell Line
Glycoproteins/*metabolism
Glycosylation
Humans
Liver/metabolism
Lysosomes/*metabolism
Mice
Protein Processing, Post-Translational
Rats
Abstract: Most soluble lysosomal proteins carry Man6P (mannose 6-phosphate), a specific carbohydrate marker that enables their binding to cellular MPRs (Man6P receptors) and their subsequent targeting towards the lysosome. This characteristic was exploited to identify novel soluble lysosomal proteins by proteomic analysis of Man6P proteins purified from a human cell line. Among the proteins identified during the course of the latter study [Journet, Chapel, Kieffer, Roux and Garin (2002) Proteomics, 2, 1026-1040], some had not been previously described as lysosomal proteins. We focused on a protein detected at 76 kDa by SDS/PAGE. We named this protein 'p76' and it appeared later in the NCBI protein database as the 'hypothetical protein LOC196463'. In the present paper, we describe the identification of p76 by MS and we analyse several of its biochemical characteristics. The presence of Man6P sugars was confirmed by an MPR overlay experiment, which showed the direct and Man6P-dependent interaction between p76 and the MPR. The presence of six N-glycosylation sites was validated by progressive peptide-N-glycosidase F deglycosylation. Experiments using N- and C-termini directed anti-p76 antibodies provided insights into p76 maturation. Most importantly, we were able to demonstrate the lysosomal localization of this protein, which was initially suggested by its Man6P tags, by both immunofluorescence and sub-cellular fractionation of mouse liver homogenates.
Notes: 1470-8728
Jensen, Anaïs G
Chemali, Magali
Chapel, Agnès
Kieffer-Jaquinod, Sylvie
Jadot, Michel
Garin, Jérôme
Journet, Agnès
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2007 Mar 15;402(3):449-58. doi: 10.1042/BJ20061205.
Author Address: Commissariat à l'Energie Atomique, Direction des Sciences du Vivant, Département Dynamique et Réponse Cellulaire, Laboratoire de Chimie des Protéines, Grenoble F-38054, France; INSERM, ERM 0201, Grenoble F-38054, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 544
Author: Jensen, C. H., Krogh, T. N., Højrup, P., Clausen, P. P., Skjødt, K., Larsson, L. I., Enghild, J. J. and Teisner, B.
Year: 1994
Title: Protein structure of fetal antigen 1 (FA1). A novel circulating human epidermal-growth-factor-like protein expressed in neuroendocrine tumors and its relation to the gene products of dlk and pG2
Journal: Eur J Biochem
Volume: 225
Issue: 1
Pages: 83-92
Epub Date: 1994/10/01
Date: Oct 1
Short Title: Protein structure of fetal antigen 1 (FA1). A novel circulating human epidermal-growth-factor-like protein expressed in neuroendocrine tumors and its relation to the gene products of dlk and pG2
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1994.00083.x
Accession Number: 7925474
Keywords: Amino Acid Sequence
Amniotic Fluid/*chemistry
Animals
Carcinoid Tumor/pathology
Epidermal Growth Factor/*chemistry
Female
Glycoproteins/biosynthesis/*chemistry/isolation & purification
Glycosylation
Humans
Immunohistochemistry
Intracellular Signaling Peptides and Proteins
*Membrane Proteins
Mice
Molecular Sequence Data
Neoplasm Proteins/biosynthesis/*chemistry
Neuroendocrine Tumors/*metabolism
Pancreas/cytology
Placenta/cytology
Pregnancy
Pregnancy Trimester, Second
RNA, Messenger/biosynthesis/*chemistry/metabolism
Sequence Homology, Amino Acid
Abstract: The present paper describes the primary structure, glycosylation and tissue localization of fetal antigen 1 (FA1) isolated from second-trimester human amniotic fluid. FA1 is a single-chained, heterogeneous glycoprotein of 225-262 amino acid residues. FA1 has six well conserved epidermal-growth-factor motifs and contains up to ten O-glycosylation and N-glycosylation sites, six of which are differentially glycosylated. Alignment to the translated sequences of Mus. musculus dlk and human dlk revealed 86% and 99% identity, respectively, to a 259-amino-acid residue overlap, and this high similarity extends with minor corrections to the human adrenal-specific mRNA, pG2 as well. Immunohistochemical analysis demonstrated the presence of FA1 in 10 out of 14 lung tumors containing neuroendocrine elements, and in the placental villi where FA1 was exclusively seen in stromal cells in close contact to the vascular structure. In the pancreas, FA1 co-localized with insulin in the insulin secretory granules of the beta cells within the islets of Langerhans. Our findings suggest that FA1 is synthesized as a membrane anchored protein and released into the circulation after enzymic cleavage, and that circulating FA1 represents the post-translationally modified gene product of human dlk which, in turn, is identical to human adrenal-specific mRNA pG2.
Notes: Jensen, C H
Krogh, T N
Højrup, P
Clausen, P P
Skjødt, K
Larsson, L I
Enghild, J J
Teisner, B
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1994 Oct 1;225(1):83-92. doi: 10.1111/j.1432-1033.1994.00083.x.
Author Address: Division of Immunology, Statens Seruminstitut, Copenhagen S, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1449
Author: Ji, S., Park, S. Y., Roth, J., Kim, H. S. and Cho, J. W.
Year: 2012
Title: O-GlcNAc modification of PPARγ reduces its transcriptional activity
Journal: Biochem Biophys Res Commun
Volume: 417
Issue: 4
Pages: 1158-63
Epub Date: 2012/01/10
Date: Jan 27
Short Title: O-GlcNAc modification of PPARγ reduces its transcriptional activity
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2011.12.086
Accession Number: 22226965
Keywords: Acetylglucosamine/antagonists & inhibitors/*metabolism
Adipocytes/*cytology/metabolism
Adipogenesis/drug effects/*genetics
Animals
Bridged Bicyclo Compounds, Heterocyclic/pharmacology
HeLa Cells
Humans
Mice
PPAR gamma/genetics/*metabolism
*Protein Processing, Post-Translational
*Transcription, Genetic
Abstract: The peroxisome proliferator-activated receptor γ (PPARγ), a member of the nuclear receptor superfamily, is a key regulator of adipogenesis and is important for the homeostasis of the adipose tissue. The β-O-linked N-acetylglucosamine (O-GlcNAc) modification, a posttranslational modification on various nuclear and cytoplasmic proteins, is involved in the regulation of protein function. Here, we report that PPARγ is modified by O-GlcNAc in 3T3-L1 adipocytes. Mass spectrometric analysis and mutant studies revealed that the threonine 54 of the N-terminal AF-1 domain of PPARγ is the major O-GlcNAc site. Transcriptional activity of wild type PPARγ was decreased 30% by treatment with the specific O-GlcNAcase (OGA) inhibitor, but the T54A mutant of PPARγ did not respond to inhibitor treatment. In 3T3-L1 cells, an increase in O-GlcNAc modification by OGA inhibitor reduced PPARγ transcriptional activity and terminal adipocyte differentiation. Our results suggest that the O-GlcNAc state of PPARγ influences its transcriptional activity and is involved in adipocyte differentiation.
Notes: 1090-2104
Ji, Suena
Park, Sang Yoon
Roth, Jürgen
Kim, Hoe Suk
Cho, Jin Won
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2012 Jan 27;417(4):1158-63. doi: 10.1016/j.bbrc.2011.12.086. Epub 2011 Dec 27.
Author Address: Department of Biology, Yonsei University, Seoul 120-749, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2065
Author: Jia, C., Li, H., Fu, D. and Lan, Y.
Year: 2020
Title: GFAT1/HBP/O-GlcNAcylation Axis Regulates β-Catenin Activity to Promote Pancreatic Cancer Aggressiveness
Journal: Biomed Res Int
Volume: 2020
Pages: 1921609
Epub Date: 2020/03/10
Short Title: GFAT1/HBP/O-GlcNAcylation Axis Regulates β-Catenin Activity to Promote Pancreatic Cancer Aggressiveness
Alternate Journal: BioMed research international
ISSN: 2314-6133 (Print)
DOI: 10.1155/2020/1921609
PMCID: PMC7048922
Accession Number: 32149084
Keywords: Cell Line, Tumor
Cell Proliferation/drug effects
Databases, Genetic
*Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing)/genetics/metabolism
Glycosylation
Hexosamines/*metabolism
Humans
*Pancreatic Neoplasms/genetics/metabolism/mortality/pathology
*beta Catenin/genetics/metabolism
Abstract: Reprogrammed glucose and glutamine metabolism are essential for tumor initiation and development. As a branch of glucose and metabolism, the hexosamine biosynthesis pathway (HBP) generates uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) and contributes to the O-GlcNAcylation process. However, the spectrum of HBP-dependent tumors and the mechanisms by which the HBP promotes tumor aggressiveness remain areas of active investigation. In this study, we analyzed the activity of the HBP and its prognostic value across 33 types of human cancers. Increased HBP activity was observed in pancreatic ductal adenocarcinoma (PDAC), and higher HBP activity predicted a poor prognosis in PDAC patients. Genetic silencing or pharmacological inhibition of the first and rate-limiting enzyme of the HBP, glutamine:fructose-6-phosphate amidotransferase 1 (GFAT1), inhibited PDAC cell proliferation, invasive capacity, and triggered cell apoptosis. Notably, these effects can be restored by addition of UDP-GlcNAc. Moreover, similar antitumor effects were noticed by pharmacological inhibition of GFAT1 with 6-diazo-5-oxo-l-norleucine (DON) or Azaserine. PDAC is maintained by oncogenic Wnt/β-catenin transcriptional activity. Our data showed that GFAT1 can regulate β-catenin expression via modulation of the O-GlcNAcylation process. TOP/FOP-Flash and real-time qPCR analysis showed that GFAT1 knockdown inhibited β-catenin activity and the transcription of its downstream target genes CCND1 and MYC. Ectopic expression of a stabilized form of β-catenin restored the suppressive roles of GFAT1 knockdown on PDAC cell proliferation and invasion. Collectively, our findings indicate that higher GFAT1/HBP/O-GlcNAcylation exhibits tumor-promoting roles by maintaining β-catenin activity in PDAC.
Notes: 2314-6141
Jia, Chunzeng
Orcid: 0000-0002-6741-1360
Li, Hengchao
Fu, Deliang
Orcid: 0000-0003-1832-2000
Lan, Yu
Orcid: 0000-0002-8396-1899
Journal Article
Biomed Res Int. 2020 Feb 15;2020:1921609. doi: 10.1155/2020/1921609. eCollection 2020.
Author Address: Department of Gastroenterology, Beijing Jishuitan Hospital, Beijing 100035, China.
Department of Pancreatic Surgery, Huashan Hospital, Fudan University, Shanghai 200040, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1218
Author: Jia, W., Lu, Z., Fu, Y., Wang, H. P., Wang, L. H., Chi, H., Yuan, Z. F., Zheng, Z. B., Song, L. N., Han, H. H., Liang, Y. M., Wang, J. L., Cai, Y., Zhang, Y. K., Deng, Y. L., Ying, W. T., He, S. M. and Qian, X. H.
Year: 2009
Title: A strategy for precise and large scale identification of core fucosylated glycoproteins
Journal: Mol Cell Proteomics
Volume: 8
Issue: 5
Pages: 913-23
Epub Date: 2009/01/14
Date: May
Short Title: A strategy for precise and large scale identification of core fucosylated glycoproteins
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M800504-MCP200
PMCID: PMC2689764
Accession Number: 19139490
Keywords: Acetylglucosamine/metabolism
Amino Acid Sequence
Biomedical Research
Fucose/*metabolism
Glycopeptides/blood/chemistry
Glycoproteins/*analysis
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Proteomics/*methods
Ultrafiltration
Abstract: Core fucosylation (CF) patterns of some glycoproteins are more sensitive and specific than evaluation of their total respective protein levels for diagnosis of many diseases, such as cancers. Global profiling and quantitative characterization of CF glycoproteins may reveal potent biomarkers for clinical applications. However, current techniques are unable to reveal CF glycoproteins precisely on a large scale. Here we developed a robust strategy that integrates molecular weight cutoff, neutral loss-dependent MS(3), database-independent candidate spectrum filtering, and optimization to effectively identify CF glycoproteins. The rationale for spectrum treatment was innovatively based on computation of the mass distribution in spectra of CF glycopeptides. The efficacy of this strategy was demonstrated by implementation for plasma from healthy subjects and subjects with hepatocellular carcinoma. Over 100 CF glycoproteins and CF sites were identified, and over 10,000 mass spectra of CF glycopeptide were found. The scale of identification results indicates great progress for finding biomarkers with a particular and attractive prospect, and the candidate spectra will be a useful resource for the improvement of database searching methods for glycopeptides.
Notes: 1535-9484
Jia, Wei
Lu, Zhuang
Fu, Yan
Wang, Hai-Peng
Wang, Le-Heng
Chi, Hao
Yuan, Zuo-Fei
Zheng, Zhao-Bin
Song, Li-Na
Han, Huan-Huan
Liang, Yi-Min
Wang, Jing-Lan
Cai, Yun
Zhang, Yu-Kui
Deng, Yu-Lin
Ying, Wan-Tao
He, Si-Min
Qian, Xiao-Hong
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2009 May;8(5):913-23. doi: 10.1074/mcp.M800504-MCP200. Epub 2009 Jan 12.
Author Address: State Key Laboratory of Proteomics-Beijing Proteome Research Center-Beijing Institute of Radiation Medicine, No. 33 Life Science Park Road, Changping District, Beijing 102206, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1734
Author: Jiang, K., Gao, Y., Hou, W., Tian, F., Ying, W., Li, L., Bai, B., Hou, G., Wang, P. G. and Zhang, L.
Year: 2016
Title: Proteomic analysis of O-GlcNAcylated proteins in invasive ductal breast carcinomas with and without lymph node metastasis
Journal: Amino Acids
Volume: 48
Issue: 2
Pages: 365-74
Epub Date: 2015/09/17
Date: Feb
Short Title: Proteomic analysis of O-GlcNAcylated proteins in invasive ductal breast carcinomas with and without lymph node metastasis
Alternate Journal: Amino acids
ISSN: 0939-4451
DOI: 10.1007/s00726-015-2089-8
Accession Number: 26374642
Keywords: Acetylglucosamine/*metabolism
Carcinoma, Ductal, Breast/*pathology
Chromatography, Liquid
Female
Glycosylation
Humans
Lymph Nodes/pathology
Lymphatic Metastasis/*pathology
N-Acetylglucosaminyltransferases/*metabolism
Protein Processing, Post-Translational
Proteomics
Tandem Mass Spectrometry
Breast cancer
Glycolysis
Invasive ductal carcinoma
Lymph node
Metastasis
O-GlcNAc
Abstract: The potential role of protein O-GlcNAcylation in cancer has been studied extensively, and the spread of cancer cells to regional lymph nodes is the first step in the dissemination of breast cancer. However, the correlation between O-GlcNAcylation and lymphatic metastasis in breast cancer remains elusive. In this study, we demonstrated that the overall O-GlcNAcylation as well as O-GlcNAc transferase (OGT) tends to decrease in response to the augmentation of lymph node metastasis (LNM) in invasive ductal breast carcinomas (IDCs). Although accumulating evidence indicates that individual O-GlcNAcylation may be important in the pathogenesis of breast cancer, O-GlcNAcylated proteins in IDCs are still largely unexplored. Herein, O-GlcNAcylated proteins of IDCs were chemo-enzymatically enriched and identified via liquid chromatography combined with tandem mass spectrometry. In total, 155 O-GlcNAcylated proteins were determined, of which 41 were only observed in LNM tissues, while 40 were unique in non-LNM samples. Gene ontology analysis showed that O-GlcNAc is primarily a nucleocytoplasmic post-translational modification, and most enriched functional terms were related to cancer development in both metastatic and non-metastatic IDCs. Moreover, several O-GlcNAcylated proteins involved in glycolysis and its accessory pathway were identified from LNM and non-LNM groups, respectively. These results indicate that the O-GlcNAcylation statuses of individual proteins were independent of the overall O-GlcNAcylation levels of metastatic and non-metastatic IDCs. Aberrant O-GlcNAc modification of these proteins might be associated with LNM progression.
Notes: 1438-2199
Jiang, Kuan
Gao, Yang
Hou, Weiwei
Tian, Fang
Ying, Wantao
Li, Ling
Bai, Bingyang
Hou, Gang
Wang, Peng George
Zhang, Lianwen
Journal Article
Research Support, Non-U.S. Gov't
Austria
Amino Acids. 2016 Feb;48(2):365-74. doi: 10.1007/s00726-015-2089-8. Epub 2015 Sep 15.
Author Address: State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy, Collaborative Innovation Center for Biotherapy, and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin, 300071, China.
School of Medicine, Nankai University, Tianjin, 300071, China.
Department of Pathology, The Central Hospital of Taian, 29 Longtan Road, Taian, 271000, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing, 102206, China.
Department of Pathology, The Central Hospital of Taian, 29 Longtan Road, Taian, 271000, China. hougang5586@126.com.
State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy, Collaborative Innovation Center for Biotherapy, and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin, 300071, China. pwang@nankai.edu.cn.
State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy, Collaborative Innovation Center for Biotherapy, and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin, 300071, China. lianwen@nankai.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1768
Author: Jiang, K., Li, W., Zhang, Q., Yan, G., Guo, K., Zhang, S. and Liu, Y.
Year: 2016
Title: GP73 N-glycosylation at Asn144 reduces hepatocellular carcinoma cell motility and invasiveness
Journal: Oncotarget
Volume: 7
Issue: 17
Pages: 23530-41
Epub Date: 2016/03/20
Date: Apr 26
Short Title: GP73 N-glycosylation at Asn144 reduces hepatocellular carcinoma cell motility and invasiveness
Alternate Journal: Oncotarget
ISSN: 1949-2553
DOI: 10.18632/oncotarget.8120
PMCID: PMC5029645
Accession Number: 26993603
Keywords: Apoptosis
Carcinoma, Hepatocellular/genetics/metabolism/*pathology
Cell Adhesion
*Cell Movement
Cell Proliferation
Glycosylation
Golgi Apparatus/metabolism
Humans
Lectins/metabolism
Liver Neoplasms/genetics/metabolism/*pathology
Membrane Proteins/genetics/*metabolism
Neoplasm Invasiveness
Polysaccharides/metabolism
Tumor Cells, Cultured
Golgi protein 73
N-glycan
hepatocellular carcinoma
Abstract: Golgi Protein 73 (GP73) is a potential liver disease glycobiomarker warranting comprehensive analyses of its glycan structure and glycosylation function. In this study, we used mass spectrometry to identify glycosylation sites and the glycan structure, high-throughput lectin microarray to provide rapid and sensitive profiling of glycoconjugates, and site-directed mutagenesis to clarify the impact of glycans on target glycoproteins in vivo. We identified three GP73 N-glycosylation sites: Asn109, Asn144 and Asn398. We found five glycoforms on Asn144, including biantennary, triantennary and fucosylated glycans. Removal of N-glycans at Asn144 enhanced the motility and invasiveness of hepatocellular carcinoma cells, possibly due to inhibition of cell adhesion related to the changes of cell membrane glycosylation. This study increases our understanding of the functional relevance of GP73 glycosylation and suggests that Asn144-deleted GP73 can influence the progression and metastasis of hepatocellular carcinoma.
Notes: 1949-2553
Jiang, Kai
Li, Wei
Zhang, Qinle
Yan, Guoquan
Guo, Kun
Zhang, Shu
Liu, Yinkun
Journal Article
Oncotarget. 2016 Apr 26;7(17):23530-41. doi: 10.18632/oncotarget.8120.
Author Address: Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Shanghai, China.
Interdisciplinary Research Center on Biology and Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai, China.
Cancer Research Center, Institutes of Biomedical Sciences, Fudan University, Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2034
Author: Jiang, M., Wu, N., Xu, B., Chu, Y., Li, X., Su, S., Chen, D., Li, W., Shi, Y., Gao, X., Zhang, H., Zhang, Z., Du, W., Nie, Y., Liang, J. and Fan, D.
Year: 2019
Title: Fatty acid-induced CD36 expression via O-GlcNAcylation drives gastric cancer metastasis
Journal: Theranostics
Volume: 9
Issue: 18
Pages: 5359-5373
Epub Date: 2019/08/15
Short Title: Fatty acid-induced CD36 expression via O-GlcNAcylation drives gastric cancer metastasis
Alternate Journal: Theranostics
ISSN: 1838-7640
DOI: 10.7150/thno.34024
PMCID: PMC6691574
Accession Number: 31410220
Keywords: Animals
Biosynthetic Pathways
CD36 Antigens/genetics/*metabolism
Cell Line, Tumor
Diet, High-Fat
Fatty Acids/*pharmacology
Female
Glycosylation
Hexosamines/metabolism
Humans
Male
Mice, Inbred BALB C
Mice, Nude
Middle Aged
N-Acetylglucosaminyltransferases/metabolism
NF-kappa B/metabolism
Neoplasm Metastasis
Signal Transduction/drug effects
Stomach Neoplasms/*metabolism/*pathology
Survival Analysis
Transcription, Genetic/drug effects
Up-Regulation/drug effects
*Cd36
*O-GlcNAcylation
*fatty acid
*gastric cancer
*metastasis
Abstract: Metastasis is the primary cause of death in patients with advanced cancer. Recently, a high-fat diet was shown to specifically promote the metastatic potential of specific cancer cells in a CD36-dependent manner. However, the molecular basis of the fatty acid (FA)-induced upregulation of CD36 has remained unclear. Methods: RT-qPCR, FACS analysis, immunoblotting and immunohistochemistry, as well as retrieving TCGA database, were carried out to quantitate CD36 expression in gastric cancer (GC) tissues and cell lines. Transwell assay and xenografts were used to assess cell metastasis abilities in vitro and in vivo after indicated treatment. Luciferase reporter assay was carried out to evaluate the changes in signaling pathways when O-GlcNAcylation level was increased in GC cells and in vitro O-GlcNAcylation assay was utilized for wild and mutant types of CD36 protein to explore the potential O-GlcNAcylation sites. Results: High CD36 expression is a predictor of poor survival and promotes metastasis of GC cells and the use of neutralizing antibodies to block CD36 inhibits GC metastasis in mice. FA or a HFD promotes the metastatic potential of GC cells by upregulating CD36 via increasing the O-GlcNAcylation level. Increased O-GlcNAcylation levels promote the transcription of CD36 by activating the NF-κB pathway and also increase its FA uptake activity by directly modifying CD36 at S468 and T470. Conclusion: FA-induced hyper-O-GlcNAcylation promotes the transcription and function of CD36 by activating the NF-κB pathway and directly modifying CD36 at S468 and T470, which drives GC metastasis.
Notes: 1838-7640
Jiang, Mingzuo
Wu, Nan
Xu, Bing
Chu, Yi
Li, Xiaowei
Su, Song
Chen, Di
Li, Wenjiao
Shi, Yanting
Gao, Xiaoliang
Zhang, Haohao
Zhang, Zhao
Du, Wei
Nie, Yongzhan
Liang, Jie
Fan, Daiming
Journal Article
Research Support, Non-U.S. Gov't
Theranostics. 2019 Jul 9;9(18):5359-5373. doi: 10.7150/thno.34024. eCollection 2019.
Author Address: State key Laboratory of Cancer Biology, National Clinical Research Center for Digestive Diseases and Xijing Hospital of Digestive Diseases, Air Force Military Medical University, 710032, Xi'an, China.
Lab of Tissue Engineering, Faculty of Life Science, Northwest University,710032, Xi'an, China.
Department of Gastroenterology, Second Affiliated Hospital of Xi'an Jiaotong University, 710032, Xi'an, China.
Department of Gastroenterology, Navy General Hospital, 100048, Beijing, China.
The School of Basic Medicine, Air Force Military Medical University, 710032, Xi'an, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1959
Author: Jiang, M., Xu, B., Li, X., Shang, Y., Chu, Y., Wang, W., Chen, D., Wu, N., Hu, S., Zhang, S., Li, M., Wu, K., Yang, X., Liang, J., Nie, Y. and Fan, D.
Year: 2019
Title: O-GlcNAcylation promotes colorectal cancer metastasis via the miR-101-O-GlcNAc/EZH2 regulatory feedback circuit
Journal: Oncogene
Volume: 38
Issue: 3
Pages: 301-316
Epub Date: 2018/08/11
Date: Jan
Short Title: O-GlcNAcylation promotes colorectal cancer metastasis via the miR-101-O-GlcNAc/EZH2 regulatory feedback circuit
Alternate Journal: Oncogene
ISSN: 0950-9232 (Print)
0950-9232
DOI: 10.1038/s41388-018-0435-5
PMCID: PMC6336687
Accession Number: 30093632
Keywords: Acetylglucosamine/*metabolism
Adenocarcinoma/metabolism/mortality/*secondary
Cell Line, Tumor
Colorectal Neoplasms/metabolism/mortality/*pathology
Enhancer of Zeste Homolog 2 Protein/*metabolism
Epithelial-Mesenchymal Transition/*physiology
Feedback, Physiological
Feeding Behavior
Gene Expression Regulation, Neoplastic
Humans
Kaplan-Meier Estimate
Lymphatic Metastasis/*physiopathology
MicroRNAs/*physiology
N-Acetylglucosaminyltransferases/*metabolism
Neoplasm Proteins/*metabolism
Prognosis
Promoter Regions, Genetic
Proportional Hazards Models
*Protein Processing, Post-Translational
RNA Interference
RNA, Small Interfering/genetics
Transcription, Genetic
Abstract: Advanced colorectal cancer (CRC) is one of the deadliest cancers, and the 5-year survival rate of patients with metastasis is extremely low. The epithelial-mesenchymal transition (EMT) is considered essential for metastatic CRC, but the fundamental molecular basis underlying this effect remains unknown. Here, we identified that O-GlcNAcylation, a unique posttranslational modification (PTM) involved in cancer metabolic reprogramming, increased the metastatic capability of CRC. The levels of O-GlcNAcylation were increased in the metastatic CRC tissues and cell lines, which likely promoted the EMT by enhancing EZH2 protein stability and function. The CRC patients with higher levels of O-GlcNAcylation exhibited greater lymph node metastasis potential and lower overall survival. Bioinformatic analysis and luciferase reporter assays revealed that both O-GlcNAcylation transferase (OGT) and EZH2 are posttranscriptionally inhibited by microRNA-101. In addition, O-GlcNAcylation and H3K27me3 modification in the miR-101 promoter region further inhibited the transcription of miR-101, resulting in the upregulation of OGT and EZH2 in metastatic CRC, thus forming a vicious cycle. In this study, we demonstrated that O-GlcNAcylation, which is negatively regulated by microRNA-101, likely promotes CRC metastasis by enhancing EZH2 protein stability and function. Reducing O-GlcNAcylation may be a potential therapeutic strategy for metastatic CRC.
Notes: 1476-5594
Jiang, Mingzuo
Xu, Bing
Li, Xiaowei
Shang, Yulong
Chu, Yi
Wang, Weijie
Chen, Di
Wu, Nan
Hu, Sijun
Zhang, Song
Li, Mengbin
Wu, Kaichun
Yang, Xiaoyong
Orcid: 0000-0002-5315-7285
Liang, Jie
Nie, Yongzhan
Fan, Daiming
81730016/National Natural Science Foundation of China (National Science Foundation of China)/International
81421003/National Natural Science Foundation of China (National Science Foundation of China)/International
81772650/National Natural Science Foundation of China (National Science Foundation of China)/International
81430072/National Natural Science Foundation of China (National Science Foundation of China)/International
Journal Article
Research Support, Non-U.S. Gov't
Oncogene. 2019 Jan;38(3):301-316. doi: 10.1038/s41388-018-0435-5. Epub 2018 Aug 9.
Author Address: State key Laboratory of Cancer Biology, National Clinical Research Center for Digestive Diseases and Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, China.
Department of Gastroenterology, Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, 710004, Shaanxi Province, China.
Lab of Tissue Engineering, Faculty of Life Science, Northwest University, Xi'an, China.
Department of molecular cellular and developmental biology, Yale University, New Haven, USA.
State key Laboratory of Cancer Biology, National Clinical Research Center for Digestive Diseases and Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, China. yongznie@fmmu.edu.cn.
State key Laboratory of Cancer Biology, National Clinical Research Center for Digestive Diseases and Xijing Hospital of Digestive Diseases, Fourth Military Medical University, Xi'an, China. daimingfan@fmmu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 634
Author: Jiang, M. S. and Hart, G. W.
Year: 1997
Title: A subpopulation of estrogen receptors are modified by O-linked N-acetylglucosamine
Journal: J Biol Chem
Volume: 272
Issue: 4
Pages: 2421-8
Epub Date: 1997/01/24
Date: Jan 24
Short Title: A subpopulation of estrogen receptors are modified by O-linked N-acetylglucosamine
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.4.2421
Accession Number: 8999954
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Cattle
Chromatography, Affinity
Chromatography, Gel
Chromatography, High Pressure Liquid
Female
Galactose/metabolism
Humans
Mice
Molecular Sequence Data
Receptors, Estrogen/*metabolism
Structure-Activity Relationship
Tumor Cells, Cultured
Abstract: Estrogen receptors (ER) are ligand-inducible transcription factors regulated by Ser(Thr)-O-phosphorylation. Many transcription factors and eukaryotic RNA polymerase II itself are also dynamically modified by Ser(Thr)-O-linked N-acetylglucosamine moieties (O-GlcNAc). Here we report that subpopulations of murine, bovine, and human estrogen receptors are modified by O-GlcNAc. O-GlcNAc moieties were detected on insect cell-expressed, mouse ER (mER) by probing with bovine milk galactosyltransferase, followed by structural analysis. Wheat germ agglutinin-Sepharose affinity chromatography also readily detected terminal GlcNAc residues on subpopulations of ER purified from calf uterus, from human breast cancer cells (MCF-7), or from mER produced by in vitro translation. These data suggest that greater than 10% of these populations of estrogen receptors bear O-GlcNAc. Site mapping of insect cell expressed mER localized one major site of O-GlcNAc addition to Thr-575, within a PEST region of the carboxyl-terminal F domain. Based upon their relative resistance to both hexosaminidase and to in vitro galactosylation, O-GlcNAc moieties appear to be largely buried on native mER. This dynamic saccharide modification, like phosphorylation, may play a role in modulating the dimerization, stability, or transactivation functions of estrogen receptors.
Notes: Jiang, M S
Hart, G W
5T3 GM07445/GM/NIGMS NIH HHS/United States
CA42486/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1997 Jan 24;272(4):2421-8. doi: 10.1074/jbc.272.4.2421.
Author Address: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1633
Author: Jiang, X., Fischer, D., Chen, X., McKenna, S. D., Liu, H., Sriraman, V., Yu, H. N., Goutopoulos, A., Arkinstall, S. and He, X.
Year: 2014
Title: Evidence for Follicle-stimulating Hormone Receptor as a Functional Trimer
Journal: J Biol Chem
Volume: 289
Issue: 20
Pages: 14273-82
Epub Date: 2014/04/03
Date: May 16
Short Title: Evidence for Follicle-stimulating Hormone Receptor as a Functional Trimer
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M114.549592
PMCID: PMC4022893
Accession Number: 24692546
Keywords: Allosteric Regulation
Animals
CHO Cells
Cricetinae
Cricetulus
Follicle Stimulating Hormone/metabolism
Humans
Intracellular Space/metabolism
Models, Molecular
Mutagenesis
Mutation
*Protein Multimerization
Protein Structure, Quaternary
Receptors, FSH/agonists/antagonists & inhibitors/*chemistry/*metabolism
Signal Transduction
Arrestin
Cysteine-knot Growth Factor
G Protein-coupled Receptors (GPCR)
Glycoprotein Hormones
Receptor Structure-function
Reproduction
Abstract: Follicle-stimulating hormone receptor (FSHR), a G-protein coupled receptor, is an important drug target in the development of novel therapeutics for reproductive indications. The FSHR extracellular domains were observed in the crystal structure as a trimer, which enabled us to propose a novel model for the receptor activation mechanism. The model predicts that FSHR binds Asnα(52)-deglycosylated FSH at a 3-fold higher capacity than fully glycosylated FSH. It also predicts that, upon dissociation of the FSHR trimer into monomers, the binding of glycosylated FSH, but not deglycosylated FSH, would increase 3-fold, and that the dissociated monomers would in turn enhance FSHR binding and signaling activities by 3-fold. This study presents evidence confirming these predictions and provides crystallographic and mutagenesis data supporting the proposed model. The model also provides a mechanistic explanation to the agonist and antagonist activities of thyroid-stimulating hormone receptor autoantibodies. We conclude that FSHR exists as a functional trimer.
Notes: 1083-351x
Jiang, Xuliang
Fischer, David
Chen, Xiaoyan
McKenna, Sean D
Liu, Heli
Sriraman, Venkataraman
Yu, Henry N
Goutopoulos, Andreas
Arkinstall, Steve
He, Xiaolin
P30 CA060553/CA/NCI NIH HHS/United States
Journal Article
J Biol Chem. 2014 May 16;289(20):14273-82. doi: 10.1074/jbc.M114.549592. Epub 2014 Apr 1.
Author Address: From the EMD Serono Research and Development Institute, Billerica, Massachusetts 01821 and xuliang.jiang@emdserono.com.
From the EMD Serono Research and Development Institute, Billerica, Massachusetts 01821 and.
the Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1548
Author: Jin, F. Z., Yu, C., Zhao, D. Z., Wu, M. J. and Yang, Z.
Year: 2013
Title: A correlation between altered O-GlcNAcylation, migration and with changes in E-cadherin levels in ovarian cancer cells
Journal: Exp Cell Res
Volume: 319
Issue: 10
Pages: 1482-90
Epub Date: 2013/03/26
Date: Jun 10
Short Title: A correlation between altered O-GlcNAcylation, migration and with changes in E-cadherin levels in ovarian cancer cells
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2013.03.013
Accession Number: 23524144
Keywords: Acetylglucosamine/analogs & derivatives/*metabolism/pharmacology
Antigens, CD
Blotting, Western
Cadherins/genetics/*metabolism
Catenins/genetics/metabolism
Cell Adhesion
Cell Line, Tumor
Cell Movement/*drug effects
Female
Gene Silencing
Glycosylation
Humans
Multiprotein Complexes/genetics/metabolism
N-Acetylglucosaminyltransferases/*metabolism
Ovarian Neoplasms/metabolism/*pathology
Oximes/pharmacology
Phenylcarbamates/pharmacology
Pyrans/pharmacology
RNA, Messenger/genetics/metabolism
RNA, Small Interfering/genetics/metabolism
Thiazoles/pharmacology
Transfection
beta Catenin/genetics/metabolism
Abstract: O-GlcNAcylation is a dynamic and reversible posttranslational modification of nuclear and cytoplasmic proteins. In recent years, the roles of O-GlcNAcylation in several human malignant tumors have been investigated, and O-GlcNAcylation was found to be linked to cellular features relevant to metastasis. In this study, we modeled four diverse ovarian cancer cells and investigated the effects of O-GlcNAcylation on ovarian cancer cell migration. We found that total O-GlcNAcylation level was elevated in HO-8910PM cells compared to OVCAR3 cells. Additionally, through altering the total O-GlcNAcylation level by OGT silencing or OGA inhibition, we found that the migration of OVCAR3 cells was dramatically enhanced by PUGNAc and Thiamet G treatment, and the migration ability of HO-8910PM cells was significantly inhibited by OGT silencing. Furthermore, we also found that the expression of E-cadherin, an O-GlcNAcylated protein in ovarian cancer cells, was reduced by OGA inhibition in OVCAR3 cells and elevated by OGT silencing in HO-8910PM cells. These results indicate that O-GlcNAcylation could enhance ovarian cancer cell migration and decrease the expression of E-cadherin. Our studies also suggest that O-GlcNAcylation might become another potential target for the therapy of ovarian cancer.
Notes: 1090-2422
Jin, Feng-zhen
Yu, Chao
Zhao, De-zhang
Wu, Ming-jun
Yang, Zhu
Journal Article
Research Support, Non-U.S. Gov't
United States
Exp Cell Res. 2013 Jun 10;319(10):1482-90. doi: 10.1016/j.yexcr.2013.03.013. Epub 2013 Mar 22.
Author Address: Department of Obstetrics and Gynecology, The Second Affiliated Hospital of Chongqing Medical University, 76 Lin Jiang Road, Chongqing 400010, PR China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2068
Author: Jin, L., Yuan, F., Dai, G., Yao, Q., Xiang, H., Wang, L., Xue, B., Shan, Y. and Liu, X.
Year: 2020
Title: Blockage of O-linked GlcNAcylation induces AMPK-dependent autophagy in bladder cancer cells
Journal: Cell Mol Biol Lett
Volume: 25
Pages: 17
Epub Date: 2020/03/17
Short Title: Blockage of O-linked GlcNAcylation induces AMPK-dependent autophagy in bladder cancer cells
Alternate Journal: Cellular & molecular biology letters
ISSN: 1425-8153 (Print)
1425-8153
DOI: 10.1186/s11658-020-00208-x
PMCID: PMC7063793
Accession Number: 32174982
Keywords: AMP-Activated Protein Kinases/genetics/*metabolism
Acylation/drug effects/genetics
Autophagy/drug effects/*genetics
Autophagy-Related Protein-1 Homolog/genetics/metabolism
Azo Compounds/pharmacology
Cell Line, Tumor
Humans
Intracellular Signaling Peptides and Proteins/genetics/metabolism
N-Acetylglucosaminyltransferases/genetics/*metabolism
Norleucine/analogs & derivatives/pharmacology
Phosphorylation
Protein Processing, Post-Translational/*drug effects/genetics
Pyrans/pharmacology
RNA, Small Interfering
Thiazoles/pharmacology
Urinary Bladder Neoplasms/enzymology/genetics/*metabolism
beta-N-Acetylhexosaminidases/genetics/*metabolism
Ampk
Autophagy
O-GlcNAcylation
Ulk1
Abstract: BACKGROUND: High levels of the post-translational modification O-GlcNAcylation (O-GlcNAc) are found in multiple cancers, including bladder cancer. Autophagy, which can be induced by stress from post-translational modifications, plays a critical role in maintaining cellular homeostasis and regulating tumorigenesis. The impact of O-GlcNAcylation on autophagy in bladder cancer remains unclear. Here, we evaluate the change in autophagic activity in response to O-GlcNAcylation and explore the potential mechanisms. METHODS: O-GlcNAcylation levels in bladder cancer cells were altered through pharmacological or genetic manipulations: treating with 6-diazo-5-oxo-norleucine (DON) or thiamet-G (TG) or up- and downregulation of O-GlcNAc transferase (OGT) or O-GlcNAcase (OGA). Autophagy was determined using fluorescence microscopy and western blotting. Co-immunoprecipitation (Co-IP) assays were performed to evaluate whether the autophagy regulator AMP-activated protein kinase (AMPK) was O-GlcNAc modified. RESULTS: Cellular autophagic flux was strikingly enhanced as a result of O-GlcNAcylation suppression, whereas it decreased at high O-GlcNAcylation levels. Phosphorylation of AMPK increased after the suppression of O-GlcNAcylation. We found that O-GlcNAcylation of AMPK suppressed the activity of this regulator, thereby inhibiting ULK1 activity and autophagy. CONCLUSION: We characterized a new function of O-GlcNAcylation in the suppression of autophagy via regulation of AMPK. GRAPHICAL ABSTRACT: Blockage of O-linked GlcNAcylation induces AMPK dependent autophagy in bladder cancer cells.
Notes: 1689-1392
Jin, Lu
Yuan, Feng
Dai, Guangcheng
Yao, Qiu
Xiang, Han
Wang, Lixia
Xue, Boxin
Shan, Yuxi
Liu, Xiaolong
Journal Article
Cell Mol Biol Lett. 2020 Mar 10;25:17. doi: 10.1186/s11658-020-00208-x. eCollection 2020.
Author Address: Department of Urology, The Second Affiliated Hospital of Soochow University, 1055 Sanxiang Road, Suzhou, 215004, R.P China. GRID: grid.452666.5. ISNI: 0000 0004 1762 8363
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1133
Author: Jin, M. S., Kim, S. E., Heo, J. Y., Lee, M. E., Kim, H. M., Paik, S. G., Lee, H. and Lee, J. O.
Year: 2007
Title: Crystal structure of the TLR1-TLR2 heterodimer induced by binding of a tri-acylated lipopeptide
Journal: Cell
Volume: 130
Issue: 6
Pages: 1071-82
Epub Date: 2007/09/25
Date: Sep 21
Short Title: Crystal structure of the TLR1-TLR2 heterodimer induced by binding of a tri-acylated lipopeptide
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2007.09.008
Accession Number: 17889651
Keywords: Acylation
Amino Acid Sequence
Animals
Binding Sites
Cell Line
Cloning, Molecular
Crystallography
Dimerization
Humans
Hydrogen Bonding
Hydrophobic and Hydrophilic Interactions
Immunity, Innate
Lipopeptides
Mice
Models, Molecular
Molecular Sequence Data
Peptides/*chemistry/metabolism
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Sequence Alignment
Signal Transduction
Toll-Like Receptor 1/*chemistry/genetics/metabolism
Toll-Like Receptor 2/*chemistry/genetics/metabolism
Toll-Like Receptor 6/chemistry
Abstract: TLR2 in association with TLR1 or TLR6 plays an important role in the innate immune response by recognizing microbial lipoproteins and lipopeptides. Here we present the crystal structures of the human TLR1-TLR2-lipopeptide complex and of the mouse TLR2-lipopeptide complex. Binding of the tri-acylated lipopeptide, Pam(3)CSK(4), induced the formation of an "m" shaped heterodimer of the TLR1 and TLR2 ectodomains whereas binding of the di-acylated lipopeptide, Pam(2)CSK(4), did not. The three lipid chains of Pam(3)CSK(4) mediate the heterodimerization of the receptor; the two ester-bound lipid chains are inserted into a pocket in TLR2, while the amide-bound lipid chain is inserted into a hydrophobic channel in TLR1. An extensive hydrogen-bonding network, as well as hydrophobic interactions, between TLR1 and TLR2 further stabilize the heterodimer. We propose that formation of the TLR1-TLR2 heterodimer brings the intracellular TIR domains close to each other to promote dimerization and initiate signaling.
Notes: Jin, Mi Sun
Kim, Sung Eun
Heo, Jin Young
Lee, Mi Eun
Kim, Ho Min
Paik, Sang-Gi
Lee, Hayyoung
Lee, Jie-Oh
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell. 2007 Sep 21;130(6):1071-82. doi: 10.1016/j.cell.2007.09.008.
Author Address: Department of Chemistry, Korea Advanced Institute of Science and Technology, Daejon, Korea 305-701.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1115
Author: Jin, Z., Tietjen, I., Bu, L., Liu-Yesucevitz, L., Gaur, S. K., Walsh, C. A. and Piao, X.
Year: 2007
Title: Disease-associated mutations affect GPR56 protein trafficking and cell surface expression
Journal: Hum Mol Genet
Volume: 16
Issue: 16
Pages: 1972-85
Epub Date: 2007/06/20
Date: Aug 15
Short Title: Disease-associated mutations affect GPR56 protein trafficking and cell surface expression
Alternate Journal: Human molecular genetics
ISSN: 0964-6906 (Print)
0964-6906
DOI: 10.1093/hmg/ddm144
Accession Number: 17576745
Keywords: Animals
Cell Line
Cell Membrane/*metabolism
Endoplasmic Reticulum/metabolism
Fluorescent Antibody Technique
Glycosylation
Golgi Apparatus/metabolism
Humans
Malformations of Cortical Development/*genetics
Mice
*Mutation, Missense
Protein Transport
Receptors, G-Protein-Coupled/*genetics/*metabolism
Transfection
Abstract: Bilateral frontoparietal polymicrogyria (BFPP) is a congenital brain malformation resulting in irregularities on the surface of the cortex, where normally convoluted gyri are replaced by numerous (poly) and noticeably smaller (micro) gyri. Individuals with BFPP suffer from epilepsy, mental retardation, language impairment and motor developmental delay. Mutations in the gene-encoding G protein-coupled receptor 56 (GPR56) cause BFPP; however, it remains unclear how these mutations affect GPR56 function. Here, we examine the biochemical properties and protein trafficking of wild-type and mutant GPR56. We demonstrate that GPR56 protein undergoes two major modifications, GPS domain-mediated protein cleavage and N-glycosylation, and that the N-terminal fragment can be released from the cell surface. In contrast to the wild-type protein, disease-associated GPR56 missense mutations in the tip of the N-terminal domain (R38Q, R38W, Y88C and C91S) produce proteins with reduced intracellular trafficking and poor cell surface expression, whereas the two mutations in the GPS domain (C346S and W349S) produce proteins with dramatically impaired cleavage that fail to traffic beyond the endoplasmic reticulum. Cell-trafficking impairments are abrogated in part by pharmacological chaperones that can partially rescue mutant GPR56 cell surface expression. These data demonstrate that some BFPP-associated mutations in GPR56 impair trafficking of the mutant protein to the plasma membrane, thus providing insights into how BFPP-associated mutations affect GPR56 function.
Notes: Jin, Zhaohui
Tietjen, Ian
Bu, Lihong
Liu-Yesucevitz, Liqun
Gaur, Shantanu K
Walsh, Christopher A
Piao, Xianhua
F32 HD048035/HD/NICHD NIH HHS/United States
K08 NS045762/NS/NINDS NIH HHS/United States
R37 NS35129/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
England
Hum Mol Genet. 2007 Aug 15;16(16):1972-85. doi: 10.1093/hmg/ddm144. Epub 2007 Jun 18.
Author Address: Division of Newborn Medicine, Department of Medicine, Children's Hospital and Harvard Medical School, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2000
Author: Jitschin, R., Böttcher, M., Saul, D., Lukassen, S., Bruns, H., Loschinski, R., Ekici, A. B., Reis, A., Mackensen, A. and Mougiakakos, D.
Year: 2019
Title: Inflammation-induced glycolytic switch controls suppressivity of mesenchymal stem cells via STAT1 glycosylation
Journal: Leukemia
Volume: 33
Issue: 7
Pages: 1783-1796
Epub Date: 2019/01/27
Date: Jul
Short Title: Inflammation-induced glycolytic switch controls suppressivity of mesenchymal stem cells via STAT1 glycosylation
Alternate Journal: Leukemia
ISSN: 0887-6924
DOI: 10.1038/s41375-018-0376-6
Accession Number: 30679801
Keywords: CD8-Positive T-Lymphocytes/*immunology/metabolism/pathology
Cell Proliferation
*Glycolysis
Glycosylation
HeLa Cells
Humans
Indoleamine-Pyrrole 2,3,-Dioxygenase/genetics/metabolism
Inflammation/*immunology/metabolism/pathology
Janus Kinase 1/genetics/metabolism
Mesenchymal Stem Cells/*immunology/metabolism/pathology
STAT1 Transcription Factor/genetics/*metabolism
Signal Transduction
Up-Regulation
Abstract: Mesenchymal stem cells (MSCs) represent key contributors to tissue homeostasis and promising therapeutics for hyperinflammatory conditions including graft-versus-host disease. Their immunomodulatory effects are controlled by microenvironmental signals. The MSCs' functional response towards inflammatory cues is known as MSC-"licensing" and includes indoleamine 2,3-dioxygenase (IDO) upregulation. MSCs use tryptophan-depleting IDO to suppress T-cells. Increasing evidence suggests that several functions are (co-)determined by the cells' metabolic commitment. MSCs are capable of both, high levels of glycolysis and of oxidative phosphorylation. Although several studies have addressed alterations of the immune regulatory phenotype elicited by inflammatory priming metabolic mechanisms controlling this process remain unknown. We demonstrate that inflammatory MSC-licensing causes metabolic shifts including enhanced glycolysis and increased fatty acid oxidation. Yet, only interfering with glycolysis impacts IDO upregulation and impedes T-cell-suppressivity. We identified the Janus kinase (JAK)/signal transducer and activator of transcription (STAT)1 pathway as a regulator of both glycolysis and IDO, and show that enhanced glucose turnover is linked to abundant STAT1 glycosylation. Inhibiting the responsible O-acetylglucosamine (O-GlcNAc) transferase abolishes STAT1 activity together with IDO upregulation. Our data suggest that STAT1-O-GlcNAcylation increases its stability towards degradation thus sustaining downstream effects. This pathway could represent a target for interventions aiming to enhance the MSCs' immunoregulatory potency.
Notes: 1476-5551
Jitschin, R
Böttcher, M
Orcid: 0000-0003-2911-8830
Saul, D
Lukassen, S
Orcid: 0000-0001-7045-6327
Bruns, H
Loschinski, R
Ekici, A B
Reis, A
Orcid: 0000-0002-6301-6363
Mackensen, A
Orcid: 0000-0002-0685-4483
Mougiakakos, D
Journal Article
Research Support, Non-U.S. Gov't
England
Leukemia. 2019 Jul;33(7):1783-1796. doi: 10.1038/s41375-018-0376-6. Epub 2019 Jan 24.
Author Address: Department of Medicine 5 for Hematology and Oncology, Friedrich-Alexander-Universität Erlangen-Nürnberg, 91054, Erlangen, Germany.
Institute of Human Genetics, Friedrich-Alexander-Universität Erlangen-Nürnberg, 91054, Erlangen, Germany.
Department of Medicine 5 for Hematology and Oncology, Friedrich-Alexander-Universität Erlangen-Nürnberg, 91054, Erlangen, Germany. dimitrios.mougiakakos@uk-erlangen.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1804
Author: Jo, Y. K., Park, N. Y., Park, S. J., Kim, B. G., Shin, J. H., Jo, D. S., Bae, D. J., Suh, Y. A., Chang, J. H., Lee, E. K., Kim, S. Y., Kim, J. C. and Cho, D. H.
Year: 2016
Title: O-GlcNAcylation of ATG4B positively regulates autophagy by increasing its hydroxylase activity
Journal: Oncotarget
Volume: 7
Issue: 35
Pages: 57186-57196
Epub Date: 2016/08/17
Date: Aug 30
Short Title: O-GlcNAcylation of ATG4B positively regulates autophagy by increasing its hydroxylase activity
Alternate Journal: Oncotarget
ISSN: 1949-2553
DOI: 10.18632/oncotarget.11083
PMCID: PMC5302982
Accession Number: 27527864
Keywords: Acetylglucosamine/analogs & derivatives/metabolism
Animals
*Autophagy
Autophagy-Related Proteins/*chemistry
Cell Line, Tumor
Cysteine Endopeptidases/*chemistry
Down-Regulation
Fibroblasts/metabolism
Fluorescent Dyes/chemistry
*Gene Expression Regulation, Enzymologic
Glucose/chemistry
Humans
Immunoprecipitation
Luciferases/metabolism
Mass Spectrometry
Mice
Mixed Function Oxygenases/*chemistry
N-Acetylglucosaminyltransferases/*metabolism
Oximes/metabolism
Phenylcarbamates/metabolism
Signal Transduction
beta-N-Acetylhexosaminidases/metabolism
Atg4b
O-GlcNAcylation
Ogt
SH-SY5Y cells
autophagy
Abstract: Autophagy is a catabolic degradation process and maintains cellular homeostasis. And autophagy is activated in response to various stress conditions. Although O-GlcNAcylation functions a sensor for nutrient and stress, the relationship between O-GlcNAcylation and autophagy is largely unknown. Here, we identified that ATG4B is novel target for O-GlcNAcylation under metabolic stress condition. Treatment with PugNAc, an O-GlcNAcase inhibitor increased activation of autophagy in SH-SY5Y cells. Both bimolecular fluorescence complementation and immunoprecipitation assay indicated that OGT directly interacts with ATG4B in SH-SY5Y cells. We also found that the O-GlcNAcylated ATG4B was increased in autophagy activation conditions, and down-regulation of OGT reduces O-GlcNAcylation of ATG4B under low glucose condition. Furthermore, the proteolytic activity of ATG4B for LC3 cleavage was enhanced in PugNAc-treated cells. Taken together, these results imply that O-GlcNAcylation of ATG4B regulates autophagy activation by increasing its proteolytic activity under metabolic stress condition.
Notes: 1949-2553
Jo, Yoon Kyung
Park, Na Yeon
Park, So Jung
Kim, Byung-Gyu
Shin, Ji Hyun
Jo, Doo Sin
Bae, Dong-Jun
Suh, Young-Ah
Chang, Jeong Ho
Lee, Eun Kyung
Kim, Sang-Yeob
Kim, Jin Cheon
Cho, Dong-Hyung
Journal Article
Oncotarget. 2016 Aug 30;7(35):57186-57196. doi: 10.18632/oncotarget.11083.
Author Address: Department of Gerontology, Graduate School of East-West Medical Science, Kyung Hee University, Yongin, South Korea.
Leading-edge Research Center for Drug Discovery and Development for Diabetes and Metabolic Disease, School of Medicine, Kyungpook National University Hospital, Daegu, South Korea.
Asan Institute for Life Sciences, Asan Medical Center, Seoul, South Korea.
Department of Biology Education, Kyungpook National University, Daegu, South Korea.
Department of Biochemistry, College of Medicine, Catholic University of Korea, Seoul, South Korea.
Department of Surgery, University of Ulsan College of Medicine, Asan Medical Center, Seoul, South Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 724
Author: John, H., Preissner, K. T., Forssmann, W. G. and Ständker, L.
Year: 1999
Title: Novel glycosylated forms of human plasma endostatin and circulating endostatin-related fragments of collagen XV
Journal: Biochemistry
Volume: 38
Issue: 32
Pages: 10217-24
Epub Date: 1999/08/11
Date: Aug 10
Short Title: Novel glycosylated forms of human plasma endostatin and circulating endostatin-related fragments of collagen XV
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi990787+
Accession Number: 10441114
Keywords: Amino Acid Sequence
Animals
Blotting, Western
Collagen/*blood/*chemistry/isolation & purification
Collagen Type XVIII
Endostatins
Female
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/analysis
Peptide Fragments/*blood/*chemistry/isolation & purification
Rabbits
Abstract: Circulating elongated forms of the angiogenesis inhibitor and potential anti-cancer drug endostatin were isolated from human blood filtrate. Immunoreactive endostatin was identified by a polyclonal rabbit antiserum raised against an N-terminal epitope of the polypeptide and purified by consecutive chromatographic steps and immunoblotting. N- and C-terminal sequence analyses of the isolated molecules revealed different forms of endostatin starting with V(117)HLRPAR. lacking the last and final three residues of the noncollagenous domain 1 (NC-1) of collagen XVIII, respectively. These polypetides are found to be O-glycosylated at T(125) (residue 9) with a glycan structure of the mucin type consisting of galactose N-acetylgalactosamine and N-acetylneuraminic acid residues. Carbohydrate analyses were performed via the semiquantitative HPLC-electrospray ionization mass spectrometry (ESMS) technique after exoglycosidase hydrolysis. Circulating endostatins are present as sialoglycoprotein (22 000 and 21 841 Da +/- 0.02%) and asialoglycoprotein structures (21 710 and 21 549 Da +/- 0.02%), while the two completely deglycosylated forms are obtained only after enzymatic incubation. The described glycosylated endostatins may represent intermediates in the proteolytic pathway of the NC-1 domain of collagen XVIII resulting in bioactive endostatins. Furthermore, immunoreactive endostatin-related C-terminal fragments of human collagen XV are found in the hemofiltrate. These polypeptides exhibit the N-terminal sequences P(66)HLLPPP. and Y(81)EKPALH. of the collagen XV NC-1 domain. ESMS and immunoblotting analyses reveal three glycosylated polypeptides with a molecular mass ranging from 16 to 21 kDa. Due to the high degree of homology between collagen XV and collagen XVIII as well as their analoqous proteolytic processing, functional similarities of collagen XVIII- and XV-related fragments should be revealed in future experiments.
Notes: John, H
Preissner, K T
Forssmann, W G
Ständker, L
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1999 Aug 10;38(32):10217-24. doi: 10.1021/bi990787+.
Author Address: Lower Saxony Institute for Peptide Research (IPF), Hannover, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1582
Author: Johnson, B., Opimba, M. and Bernier, J.
Year: 2014
Title: Implications of the O-GlcNAc modification in the regulation of nuclear apoptosis in T cells
Journal: Biochim Biophys Acta
Volume: 1840
Issue: 1
Pages: 191-8
Epub Date: 2013/09/17
Date: Jan
Short Title: Implications of the O-GlcNAc modification in the regulation of nuclear apoptosis in T cells
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbagen.2013.09.011
Accession Number: 24035784
Keywords: Acetylglucosamine/analogs & derivatives/pharmacology
*Apoptosis
Blotting, Western
Cell Cycle
Cell Nucleus/metabolism/*pathology
Electrophoresis, Gel, Two-Dimensional
Glycosylation/drug effects
Humans
Immunoprecipitation
Leukemia-Lymphoma, Adult T-Cell/drug therapy/metabolism/*pathology
Myocytes, Cardiac
N-Acetylglucosaminyltransferases/antagonists & inhibitors/*metabolism
Oximes/pharmacology
Phenylcarbamates/pharmacology
*Protein Processing, Post-Translational
Proteins/*metabolism
Signal Transduction/drug effects
Trialkyltin Compounds/pharmacology
Apoptosis
Dff
Dff45
O-(2-acetamidO-2-deoxy-D-glucopyranosylidene) amino-N-phenylcarbamate
O-linked β-N-acetylglucosamine
Tbt
Abstract: BACKGROUND: O-linked β-N-acetylglucosamine (O-GlcNAc) is a nutrient-/stress-sensitive post-translational modification that affects nucleocytoplasmic proteins. The enzyme O-N-acetylglucosamine transferase (OGT) catalyzes the addition of O-GlcNAc, whereas O-N-acetylglucosaminidase (OGA) removes it. O-GlcNAcylation plays a role in fundamental regulatory mechanisms through the modification of proteins involved in cell division, metabolism, transcription, cell signaling and apoptosis. The effects of O-GlcNAcylation on apoptosis appear to be cell-dependent, as elevated levels played a protective role in primary neonatal rat ventricular myocytes but had a cytotoxic effect in rat pancreatic β-cells. The aim of the current study was to determine the implications of the O-GlcNAc modification on T cell apoptosis. METHODS: Human T lymphoblastic HPB-ALL cells were treated with the OGA inhibitor O-(2-acetamido-2-deoxy-d-glucopyranosylidene) amino-N-phenylcarbamate (PUGNAc), or with glucosamine (GlcN), to increase O-GlcNAcylation. Apoptosis was induced in the presence of tributyltin (TBT). DNA fragmentation was observed by cell cycle analysis and corresponded to the sub G0/G1 population. O-GlcNAcylated proteins were detected by immunoblot using a specific antibody (ctd110.6) and were precipitated using succinylated wheat germ agglutinin (sWGA). RESULTS: HPB-ALL cells treated with PUGNAc displayed a significant reduction in DNA fragmentation after TBT-induced apoptosis. DFF45, the protein that inhibits the endonuclease DFF40, was identified to be O-GlcNAc modified. O-GlcNAcylated DFF45 appeared to be more resistant to caspase cleavage during apoptosis. Our results suggest that a decrease in the O-GlcNAc modification on DFF45 occurs before its cleavage by caspase. GENERAL SIGNIFICANCE: Our results indicate that the O-GlcNAcylation of DFF45 may represent a mechanism to control the accidental activation of DFF.
Notes: Johnson, Bruno
Opimba, Marlyse
Bernier, Jacques
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2014 Jan;1840(1):191-8. doi: 10.1016/j.bbagen.2013.09.011. Epub 2013 Sep 13.
Author Address: INRS-Institut Armand-Frappier, 531 Boul. des Prairies, Laval, QC H7V1B7, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1417
Author: Jokela, T. A., Makkonen, K. M., Oikari, S., Kärnä, R., Koli, E., Hart, G. W., Tammi, R. H., Carlberg, C. and Tammi, M. I.
Year: 2011
Title: Cellular content of UDP-N-acetylhexosamines controls hyaluronan synthase 2 expression and correlates with O-linked N-acetylglucosamine modification of transcription factors YY1 and SP1
Journal: J Biol Chem
Volume: 286
Issue: 38
Pages: 33632-40
Epub Date: 2011/07/29
Date: Sep 23
Short Title: Cellular content of UDP-N-acetylhexosamines controls hyaluronan synthase 2 expression and correlates with O-linked N-acetylglucosamine modification of transcription factors YY1 and SP1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.265637
PMCID: PMC3190925
Accession Number: 21795679
Keywords: Acetylglucosamine/*metabolism/pharmacology
Computational Biology
Gene Expression Regulation, Enzymologic/drug effects
Gene Silencing/drug effects
Glucuronosyltransferase/genetics/*metabolism
Humans
Hyaluronan Synthases
Hyaluronic Acid/biosynthesis
Keratinocytes/drug effects/metabolism
Mannose/metabolism/pharmacology
Nitrogenous Group Transferases/metabolism
Protein Binding/drug effects
RNA, Messenger/genetics/metabolism
RNA, Small Interfering/metabolism
Response Elements/genetics
Sp1 Transcription Factor/*metabolism
Time Factors
Uridine Diphosphate/*metabolism
YY1 Transcription Factor/*metabolism
Abstract: Hyaluronan, a high molecular mass polysaccharide on the vertebrate cell surface and extracellular matrix, is produced at the plasma membrane by hyaluronan synthases using UDP-GlcNAc and UDP-GlcUA as substrates. The availability of these UDP-sugar substrates can limit the synthesis rate of hyaluronan. In this study, we show that the cellular level of UDP-HexNAc also controls hyaluronan synthesis by modulating the expression of HAS2 (hyaluronan synthase 2). Increasing UDP-HexNAc in HaCaT keratinocytes by adding glucosamine down-regulated HAS2 gene expression, whereas a decrease in UDP-HexNAc, realized by mannose treatment or siRNA for GFAT1 (glutamine:fructose-6-phosphate amidotransferase 1), enhanced expression of the gene. Tracing the UDP-HexNAc-initiated signal to the HAS2 promoter revealed no change in the binding of STAT3, NF-κB, and cAMP response element-binding protein, shown previously to mediate growth factor and cytokine signals on HAS2 expression. Instead, altered binding of SP1 and YY1 to the promoter correlated with cellular UDP-HexNAc content and inhibition of HAS2 expression. siRNA silencing of YY1 and SP1 confirmed their inhibitory effects on HAS2 expression. Reduced and increased levels of O-GlcNAc-modified SP1 and YY1 proteins were associated with stimulation or inhibition of HAS2 expression, respectively. Our data are consistent with the hypothesis that, by regulating the level of protein O-GlcNAc modifications, cellular UDP-HexNAc content controls HAS2 transcription and decreases the effects on hyaluronan synthesis that would result from cellular fluctuations of this substrate.
Notes: 1083-351x
Jokela, Tiina A
Makkonen, Katri M
Oikari, Sanna
Kärnä, Riikka
Koli, Elina
Hart, Gerald W
Tammi, Raija H
Carlberg, Carsten
Tammi, Markku I
P01 HL107153/HL/NHLBI NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 Sep 23;286(38):33632-40. doi: 10.1074/jbc.M111.265637. Epub 2011 Jul 27.
Author Address: Institute of Biomedicine, University of Eastern Finland, FIN-70211 Kuopio, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1093
Author: Jongen, S. P., Gerwig, G. J., Leeflang, B. R., Koles, K., Mannesse, M. L., van Berkel, P. H., Pieper, F. R., Kroos, M. A., Reuser, A. J., Zhou, Q., Jin, X., Zhang, K., Edmunds, T. and Kamerling, J. P.
Year: 2007
Title: N-glycans of recombinant human acid alpha-glucosidase expressed in the milk of transgenic rabbits
Journal: Glycobiology
Volume: 17
Issue: 6
Pages: 600-19
Epub Date: 2007/02/13
Date: Jun
Short Title: N-glycans of recombinant human acid alpha-glucosidase expressed in the milk of transgenic rabbits
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwm015
Accession Number: 17293352
Keywords: Animals
Animals, Genetically Modified
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Female
Glycosylation
Humans
Mammary Glands, Animal/metabolism
Mass Spectrometry
Milk/*chemistry
Nuclear Magnetic Resonance, Biomolecular
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/pharmacology
Polysaccharides/*chemistry/isolation & purification/*metabolism
Rabbits
Recombinant Proteins/chemistry/isolation & purification/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
alpha-Glucosidases/*chemistry/genetics/*metabolism
Abstract: Pompe disease is a lysosomal glycogen storage disorder characterized by acid alpha-glucosidase (GAA) deficiency. More than 110 different pathogenic mutations in the gene encoding GAA have been observed. Patients with this disease are being treated by intravenous injection of recombinant forms of the enzyme. Focusing on recombinant approaches to produce the enzyme means that specific attention has to be paid to the generated glycosylation patterns. Here, human GAA was expressed in the mammary gland of transgenic rabbits. The N-linked glycans of recombinant human GAA (rhAGLU), isolated from the rabbit milk, were released by peptide-N(4)-(N-acetyl-beta-glucosaminyl)asparagine amidase F. The N-glycan pool was fractionated and purified into individual components by a combination of anion-exchange, normal-phase, and Sambucus nigra agglutinin-affinity chromatography. The structures of the components were analyzed by 500 MHz one-dimensional and 600 MHz cryo two-dimensional (total correlation spectroscopy [TOCSY] nuclear Overhauser enhancement spectroscopy) (1)H nuclear magnetic resonance spectroscopy, combined with two-dimensional (31)P-filtered (1)H-(1)H TOCSY spectroscopy, matrix-assisted laser desorption ionization time-of-flight mass spectrometry, and high-performance liquid chromatography (HPLC)-profiling of 2-aminobenzamide-labeled glycans combined with exoglycosidase digestions. The recombinant rabbit glycoprotein contained a broad array of different N-glycans, comprising oligomannose-, hybrid-, and complex-type structures. Part of the oligomannose-type glycans showed the presence of phospho-diester-bridged N-acetylglucosamine. For the complex-type glycans (partially) (alpha2-6)-sialylated (nearly only N-acetylneuraminic acid) diantennary structures were found; part of the structures were (alpha1-6)-core-fucosylated or (alpha1-3)-fucosylated in the upper antenna (Lewis x). Using HPLC-mass spectrometry of glycopeptides, information was generated with respect to the site-specific location of the various glycans.
Notes: Jongen, Susanne P
Gerwig, Gerrit J
Leeflang, Bas R
Koles, Kate
Mannesse, Maurice L M
van Berkel, Patrick H C
Pieper, Frank R
Kroos, Marian A
Reuser, Arnold J J
Zhou, Qun
Jin, Xiaoying
Zhang, Kate
Edmunds, Tim
Kamerling, Johannis P
Journal Article
England
Glycobiology. 2007 Jun;17(6):600-19. doi: 10.1093/glycob/cwm015. Epub 2007 Feb 9.
Author Address: Bijvoet Center for Biomolecular Research, Department of Bio-Organic Chemistry, Utrecht University, Padualaan 8, NL-3584 CH Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 753
Author: Jonsson, A. P., Griffiths, W. J., Bratt, P., Johansson, I., Strömberg, N., Jörnvall, H. and Bergman, T.
Year: 2000
Title: A novel Ser O-glucuronidation in acidic proline-rich proteins identified by tandem mass spectrometry
Journal: FEBS Lett
Volume: 475
Issue: 2
Pages: 131-4
Epub Date: 2000/06/20
Date: Jun 16
Short Title: A novel Ser O-glucuronidation in acidic proline-rich proteins identified by tandem mass spectrometry
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/s0014-5793(00)01645-8
Accession Number: 10858503
Keywords: Amino Acid Sequence
Chromatography, High Pressure Liquid
Glucuronic Acid/*metabolism
Humans
Mass Spectrometry/*methods
Molecular Sequence Data
Peptides/*chemistry/isolation & purification/metabolism
Proline/chemistry
Proline-Rich Protein Domains
Protein Processing, Post-Translational
Serine/*metabolism
Time Factors
Trypsin/metabolism
Abstract: Human acidic proline-rich salivary protein PRP-1 and its C-terminally truncated form PRP-3 were analyzed by electrospray tandem mass spectrometry. Post-translational modifications were detected and characterized. A pyroglutamic acid residue was demonstrated at the N-terminus, Ser-8 and Ser-22 were shown to be phosphorylated and an O-linked glucuronic acid conjugation was identified. The latter modification was located to Ser-17 and found to be present in approximately 40% of the polypeptides.
Notes: Jonsson, A P
Griffiths, W J
Bratt, P
Johansson, I
Strömberg, N
Jörnvall, H
Bergman, T
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2000 Jun 16;475(2):131-4. doi: 10.1016/s0014-5793(00)01645-8.
Author Address: Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1544
Author: Kaasik, K., Kivimäe, S., Allen, J. J., Chalkley, R. J., Huang, Y., Baer, K., Kissel, H., Burlingame, A. L., Shokat, K. M., Ptáček, L. J. and Fu, Y. H.
Year: 2013
Title: Glucose sensor O-GlcNAcylation coordinates with phosphorylation to regulate circadian clock
Journal: Cell Metab
Volume: 17
Issue: 2
Pages: 291-302
Epub Date: 2013/02/12
Date: Feb 5
Short Title: Glucose sensor O-GlcNAcylation coordinates with phosphorylation to regulate circadian clock
Alternate Journal: Cell metabolism
ISSN: 1550-4131 (Print)
1550-4131
DOI: 10.1016/j.cmet.2012.12.017
PMCID: PMC3597447
NIHMSID: NIHMS435920
Accession Number: 23395175
Keywords: Acetylglucosamine/*metabolism
Adenosine Triphosphate/analogs & derivatives
Amino Acid Sequence
Animals
CLOCK Proteins/chemistry/genetics/metabolism
*Circadian Clocks
Drosophila Proteins/chemistry/genetics/metabolism
Drosophila melanogaster/metabolism
Glucose/*metabolism
Glycogen Synthase Kinase 3/metabolism
Glycogen Synthase Kinase 3 beta
Glycosylation
Humans
Mice
Molecular Sequence Data
N-Acetylglucosaminyltransferases/chemistry/metabolism
Phosphorylation
Substrate Specificity
Transcription, Genetic
Transfection
Abstract: Posttranslational modifications play central roles in myriad biological pathways including circadian regulation. We employed a circadian proteomic approach to demonstrate that circadian timing of phosphorylation is a critical factor in regulating complex GSK3β-dependent pathways and identified O-GlcNAc transferase (OGT) as a substrate of GSK3β. Interestingly, OGT activity is regulated by GSK3β; hence, OGT and GSK3β exhibit reciprocal regulation. Modulating O-GlcNAcylation levels alter circadian period length in both mice and Drosophila; conversely, protein O-GlcNAcylation is circadianly regulated. Central clock proteins, Clock and Period, are reversibly modified by O-GlcNAcylation to regulate their transcriptional activities. In addition, O-GlcNAcylation of a region in PER2 known to regulate human sleep phase (S662-S674) competes with phosphorylation of this region, and this interplay is at least partly mediated by glucose levels. Together, these results indicate that O-GlcNAcylation serves as a metabolic sensor for clock regulation and works coordinately with phosphorylation to fine-tune circadian clock.
Notes: 1932-7420
Kaasik, Krista
Kivimäe, Saul
Allen, Jasmina J
Chalkley, Robert J
Huang, Yong
Baer, Kristin
Kissel, Holger
Burlingame, Alma L
Shokat, Kevan M
Ptáček, Louis J
Fu, Ying-Hui
MH074924/MH/NIMH NIH HHS/United States
S10 RR015804/RR/NCRR NIH HHS/United States
P41 GM103481/GM/NIGMS NIH HHS/United States
EB001987/EB/NIBIB NIH HHS/United States
RR015804/RR/NCRR NIH HHS/United States
R01 HL059596/HL/NHLBI NIH HHS/United States
R01 GM079180/GM/NIGMS NIH HHS/United States
GM103481/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
GM079180/GM/NIGMS NIH HHS/United States
HL059596/HL/NHLBI NIH HHS/United States
R01 NS072360/NS/NINDS NIH HHS/United States
R01 EB001987/EB/NIBIB NIH HHS/United States
P50 MH074924/MH/NIMH NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell Metab. 2013 Feb 5;17(2):291-302. doi: 10.1016/j.cmet.2012.12.017.
Author Address: Department of Neurology, University of California San Francisco, San Francisco, CA 94158, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1928
Author: Kadirvelraj, R., Yang, J. Y., Sanders, J. H., Liu, L., Ramiah, A., Prabhakar, P. K., Boons, G. J., Wood, Z. A. and Moremen, K. W.
Year: 2018
Title: Human N-acetylglucosaminyltransferase II substrate recognition uses a modular architecture that includes a convergent exosite
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 18
Pages: 4637-4642
Epub Date: 2018/04/19
Date: May 1
Short Title: Human N-acetylglucosaminyltransferase II substrate recognition uses a modular architecture that includes a convergent exosite
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1716988115
PMCID: PMC5939069
Accession Number: 29666272
Keywords: Humans
N-Acetylglucosaminyltransferases/*chemistry/metabolism
Protein Domains
*Protein Folding
Uridine Diphosphate N-Acetylglucosamine/*chemistry/metabolism
*N-glycan processing
*convergent evolution
*exosite
*glycosyltranferase
*substrate recognition
Abstract: Asn-linked oligosaccharides are extensively modified during transit through the secretory pathway, first by trimming of the nascent glycan chains and subsequently by initiating and extending multiple oligosaccharide branches from the trimannosyl glycan core. Trimming and branching pathway steps are highly ordered and hierarchal based on the precise substrate specificities of the individual biosynthetic enzymes. A key committed step in the synthesis of complex-type glycans is catalyzed by N-acetylglucosaminyltransferase II (MGAT2), an enzyme that generates the second GlcNAcβ1,2- branch from the trimannosyl glycan core using UDP-GlcNAc as the sugar donor. We determined the structure of human MGAT2 as a Mn(2+)-UDP donor analog complex and as a GlcNAcMan(3)GlcNAc(2)-Asn acceptor complex to reveal the structural basis for substrate recognition and catalysis. The enzyme exhibits a GT-A Rossmann-like fold that employs conserved divalent cation-dependent substrate interactions with the UDP-GlcNAc donor. MGAT2 interactions with the extended glycan acceptor are distinct from other related glycosyltransferases. These interactions are composed of a catalytic subsite that binds the Man-α1,6- monosaccharide acceptor and a distal exosite pocket that binds the GlcNAc-β1,2Man-α1,3Manβ- substrate "recognition arm." Recognition arm interactions are similar to the enzyme-substrate interactions for Golgi α-mannosidase II, a glycoside hydrolase that acts just before MGAT2 in the Asn-linked glycan biosynthetic pathway. These data suggest that substrate binding by MGAT2 employs both conserved and convergent catalytic subsite modules to provide substrate selectivity and catalysis. More broadly, the MGAT2 active-site architecture demonstrates how glycosyltransferases create complementary modular templates for regiospecific extension of glycan structures in mammalian cells.
Notes: 1091-6490
Kadirvelraj, Renuka
Yang, Jeong-Yeh
Sanders, Justin H
Liu, Lin
Ramiah, Annapoorani
Prabhakar, Pradeep Kumar
Boons, Geert-Jan
Wood, Zachary A
Moremen, Kelley W
Orcid: 0000-0003-1768-582x
P01 GM107012/GM/NIGMS NIH HHS/United States
P41 GM103390/GM/NIGMS NIH HHS/United States
R01 GM114298/GM/NIGMS NIH HHS/United States
R01 GM130915/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Proc Natl Acad Sci U S A. 2018 May 1;115(18):4637-4642. doi: 10.1073/pnas.1716988115. Epub 2018 Apr 16.
Author Address: Department of Biochemistry and Molecular Biology, University of Georgia, Athens, GA 30602.
Complex Carbohydrate Research Center, University of Georgia, Athens, GA 30602-4712.
Department of Biochemistry and Molecular Biology, University of Georgia, Athens, GA 30602; zaw@uga.edu moremen@uga.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1008
Author: Kadoya, T., Khurana, A., Tcherpakov, M., Bromberg, K. D., Didier, C., Broday, L., Asahara, T., Bhoumik, A. and Ronai, Z.
Year: 2005
Title: JAMP, a Jun N-terminal kinase 1 (JNK1)-associated membrane protein, regulates duration of JNK activity
Journal: Mol Cell Biol
Volume: 25
Issue: 19
Pages: 8619-30
Epub Date: 2005/09/17
Date: Oct
Short Title: JAMP, a Jun N-terminal kinase 1 (JNK1)-associated membrane protein, regulates duration of JNK activity
Alternate Journal: Molecular and cellular biology
ISSN: 0270-7306 (Print)
0270-7306
DOI: 10.1128/mcb.25.19.8619-8630.2005
PMCID: PMC1265750
Accession Number: 16166642
Keywords: 3T3 Cells
Amino Acid Sequence
Animals
Apoptosis
Carrier Proteins/biosynthesis/*physiology
Cell Line
Cell Line, Tumor
Cell Membrane/*metabolism
Cell Movement
DNA/metabolism
DNA, Complementary/metabolism
Dual-Specificity Phosphatases
*Gene Expression Regulation, Enzymologic
Glycosylation
Green Fluorescent Proteins/metabolism
HeLa Cells
Humans
Immunoprecipitation
MAP Kinase Signaling System
Membrane Glycoproteins/biosynthesis/*physiology
Mice
Microscopy, Confocal
Molecular Sequence Data
NIH 3T3 Cells
Phosphoprotein Phosphatases/metabolism
Protein Binding
Protein Structure, Tertiary
RNA Interference
Reverse Transcriptase Polymerase Chain Reaction
Subcellular Fractions/metabolism
Time Factors
Tissue Distribution
Transfection
Tunicamycin/pharmacology
Ultraviolet Rays
Abstract: We report the identification and characterization of JAMP (JNK1 [Jun N-terminal kinase 1]-associated membrane protein), a predicted seven-transmembrane protein that is localized primarily within the plasma membrane and associates with JNK1 through its C-terminal domain. JAMP association with JNK1 outcompetes JNK1 association with mitogen-activated protein kinase phosphatase 5, resulting in increased and prolonged JNK1 activity following stress. Elevated expression of JAMP following UV or tunicamycin treatment results in sustained JNK activity and a higher level of JNK-dependent apoptosis. Inhibition of JAMP expression by RNA interference reduces the degree and duration of JNK activation and concomitantly the level of stress-induced apoptosis. Through its regulation of JNK1 activity, JAMP emerges as a membrane-anchored regulator of the duration of JNK1 activity in response to diverse stress stimuli.
Notes: 1098-5549
Kadoya, Takayuki
Khurana, Ashwani
Tcherpakov, Marianna
Bromberg, Kenneth D
Didier, Christine
Broday, Limor
Asahara, Toshimasa
Bhoumik, Anindita
Ronai, Ze'ev
R01 CA097105/CA/NCI NIH HHS/United States
CA97105/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, P.H.S.
Mol Cell Biol. 2005 Oct;25(19):8619-30. doi: 10.1128/MCB.25.19.8619-8630.2005.
Author Address: Signal Transduction Program, The Burnham Institute, 10901 N. Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1433
Author: Kajander, T., Kuja-Panula, J., Rauvala, H. and Goldman, A.
Year: 2011
Title: Crystal structure and role of glycans and dimerization in folding of neuronal leucine-rich repeat protein AMIGO-1
Journal: J Mol Biol
Volume: 413
Issue: 5
Pages: 1001-15
Epub Date: 2011/10/11
Date: Nov 11
Short Title: Crystal structure and role of glycans and dimerization in folding of neuronal leucine-rich repeat protein AMIGO-1
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2011.09.032
Accession Number: 21983541
Keywords: Amino Acid Sequence
Animals
Blotting, Western
Crystallography, X-Ray
Mice
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutation/genetics
Neurons/cytology/*metabolism
Polysaccharides/chemistry/*metabolism
Protein Conformation
*Protein Folding
Protein Multimerization
Proteins/*chemistry/genetics/*metabolism
Sequence Homology, Amino Acid
Abstract: AMIGO-1 is the parent member of a novel family of three cell surface leucine-rich repeat (LRR) proteins. Its expression is induced by the binding of HMGB1 (high-mobility group box 1 protein) to RAGE (receptor for advanced glycation end products) on neurons. Binding of HMGB1 to RAGE is known to have a direct effect on cellular growth regulation and mobility, and AMIGO-1 directly supports growth of neuronal processes and fasciculation of neurites. In addition, the second member of the AMIGO-family, AMIGO-2, has been implicated in adhesion of tumor cells in adenocarcinoma and survival of neurons. We have determined the crystal structure of AMIGO-1 at 2.0 Å resolution, which reveals a typical cell surface LRR domain arrangement with N- and C-terminal capping domains with disulfide bridges, followed by a C2-type Ig domain. AMIGO-1 is a dimer, with the LRR regions forming the dimer interface, and sequence conservation analysis and static light-scattering measurements suggest that all three AMIGO family proteins form similar dimers. Based on the AMIGO-1 structure, we have also modeled AMIGO-2 and present small-angle X-ray scattering data on AMIGO-2 and AMIGO-3. Our mutagenesis studies show that AMIGO-1 dimerization is necessary for proper cell surface expression and thus probably for proper or stable folding in the endoplastic reticulum and for the function of the protein. Based on the data presented earlier, we also suggest that dimerization through the LRR-LRR interface is likely to be involved in cell-cell adhesion by AMIGO-1, while extensive glycosylation may have a role.
Notes: 1089-8638
Kajander, Tommi
Kuja-Panula, Juha
Rauvala, Heikki
Goldman, Adrian
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2011 Nov 11;413(5):1001-15. doi: 10.1016/j.jmb.2011.09.032. Epub 2011 Sep 29.
Author Address: Institute of Biotechnology, Structural Biology and Biophysics, University of Helsinki, 00014 Helsinki, Finland. tommi.kajander@helsinki.fi
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1772
Author: Kakade, P. S., Budnar, S., Kalraiya, R. D. and Vaidya, M. M.
Year: 2016
Title: Functional Implications of O-GlcNAcylation-dependent Phosphorylation at a Proximal Site on Keratin 18
Journal: J Biol Chem
Volume: 291
Issue: 23
Pages: 12003-13
Epub Date: 2016/04/10
Date: Jun 3
Short Title: Functional Implications of O-GlcNAcylation-dependent Phosphorylation at a Proximal Site on Keratin 18
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.728717
PMCID: PMC4933253
Accession Number: 27059955
Keywords: Acetylglucosamine/*metabolism
Acylation
Binding Sites/genetics
Cell Line
Cell Movement/genetics
Fibronectins/metabolism
Hepatocytes/cytology/metabolism
Humans
Immunoblotting
Keratin-18/genetics/*metabolism
Microscopy, Confocal
Mutation, Missense
Phosphorylation
Serine/genetics/*metabolism
O-GlcNAcylation
cytoskeleton
intermediate filament
keratin
migration
protein degradation
Abstract: Keratins 8/18 (K8/18) are phosphoglycoproteins and form the major intermediate filament network of simple epithelia. The three O-GlcNAcylation (Ser(29), Ser(30), and Ser(48)) and two phosphorylation (Ser(33) and Ser(52)) serine sites on K18 are well characterized. Both of these modifications have been reported to increase K18 solubility and regulate its filament organization. In this report, we investigated the site-specific interplay between these two modifications in regulating the functional properties of K18, like solubility, stability, and filament organization. An immortalized hepatocyte cell line (HHL-17) stably expressing site-specific single, double, and triple O-GlcNAc and phosphomutants of K18 were used to identify the site(s) critical for regulating these functions. Keratin 18 mutants where O-GlcNAcylation at Ser(30) was abolished (K18-S30A) exhibited reduced phosphorylation induced solubility, increased stability, defective filament architecture, and slower migration. Interestingly, K18-S30A mutants also showed loss of phosphorylation at Ser(33), a modification known to regulate the solubility of K18. Further to this, the K18 phosphomutant (K18-S33A) mimicked K18-S30A in its stability, filament organization, and cell migration. These results indicate that O-GlcNAcylation at Ser(30) promotes phosphorylation at Ser(33) to regulate the functional properties of K18 and also impact cellular processes like migration. O-GlcNAcylation and phosphorylation on the same or adjacent sites on most proteins antagonize each other in regulating protein functions. Here we report a novel, positive interplay between O-GlcNAcylation and phosphorylation at adjacent sites on K18 to regulate its fundamental properties.
Notes: 1083-351x
Kakade, Poonam S
Budnar, Srikanth
Kalraiya, Rajiv D
Vaidya, Milind M
Address
Research Support, Non-U.S. Gov't
J Biol Chem. 2016 Jun 3;291(23):12003-13. doi: 10.1074/jbc.M116.728717. Epub 2016 Apr 8.
Author Address: From the Advanced Centre for Treatment, Research, and Education in Cancer (ACTREC), Tata Memorial Centre, Sector 22, Kharghar, Navi Mumbai 410210, India.
From the Advanced Centre for Treatment, Research, and Education in Cancer (ACTREC), Tata Memorial Centre, Sector 22, Kharghar, Navi Mumbai 410210, India s.budnar@imb.uq.edu.au srikanth.budnar@gmail.com.
From the Advanced Centre for Treatment, Research, and Education in Cancer (ACTREC), Tata Memorial Centre, Sector 22, Kharghar, Navi Mumbai 410210, India mvaidya@actrec.gov.in.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1830
Author: Kakuda, S. and Haltiwanger, R. S.
Year: 2017
Title: Deciphering the Fringe-Mediated Notch Code: Identification of Activating and Inhibiting Sites Allowing Discrimination between Ligands
Journal: Dev Cell
Volume: 40
Issue: 2
Pages: 193-201
Epub Date: 2017/01/17
Date: Jan 23
Short Title: Deciphering the Fringe-Mediated Notch Code: Identification of Activating and Inhibiting Sites Allowing Discrimination between Ligands
Alternate Journal: Developmental cell
ISSN: 1534-5807 (Print)
1534-5807
DOI: 10.1016/j.devcel.2016.12.013
PMCID: PMC5263050
NIHMSID: NIHMS836856
Accession Number: 28089369
Keywords: Amino Acid Sequence
Animals
Calcium-Binding Proteins
Epidermal Growth Factor/metabolism
Fucose/metabolism
Glucosyltransferases
Glycosyltransferases/*metabolism
HEK293 Cells
Humans
Intercellular Signaling Peptides and Proteins/*metabolism
Jagged-1 Protein/*metabolism
Ligands
Mice
Models, Biological
NIH 3T3 Cells
Protein Binding
Proteins/*metabolism
Receptor, Notch1/chemistry/*metabolism
Repetitive Sequences, Amino Acid
*EGF repeats
*Fringe
*Notch
*O-fucose
*development
*glycosylation
*signal transduction
Abstract: Fringe proteins are β3-N-acetylglucosaminyltransferases that modulate Notch activity by modifying O-fucose residues on epidermal growth factor-like (EGF) repeats of Notch. Mammals have three Fringes: Lunatic, Manic, and Radical. While Lunatic and Manic Fringe inhibit Notch1 activation from Jagged1 and enhance activation from Delta-like 1, Radical Fringe enhances signaling from both. We used a mass spectrometry approach to determine whether the variable effects of Fringes on Notch1 result from generation of unique glycosylation patterns on Notch1. We found that Lunatic and Manic Fringe modified similar sites on Notch1, while Radical Fringe modified a subset. Fringe modifications at EGF8 and EGF12 enhanced Notch1 binding to and activation from Delta-like 1, while modifications at EGF6 and EGF36 (added by Manic and Lunatic but not Radical) inhibited Notch1 activation from Jagged1. Combined, these results suggest that Fringe modifications "mark" different regions in the Notch1 extracellular domain for activation or inhibition.
Notes: 1878-1551
Kakuda, Shinako
Haltiwanger, Robert S
R01 GM061126/GM/NIGMS NIH HHS/United States
Journal Article
Dev Cell. 2017 Jan 23;40(2):193-201. doi: 10.1016/j.devcel.2016.12.013. Epub 2017 Jan 12.
Author Address: Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY 11794, USA.
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY 11794, USA. Electronic address: rhalti@uga.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1817
Author: Kamemura, K.
Year: 2017
Title: O-GlcNAc glycosylation stoichiometry of the FET protein family: only EWS is glycosylated with a high stoichiometry
Journal: Biosci Biotechnol Biochem
Volume: 81
Issue: 3
Pages: 541-546
Epub Date: 2016/12/03
Date: Mar
Short Title: O-GlcNAc glycosylation stoichiometry of the FET protein family: only EWS is glycosylated with a high stoichiometry
Alternate Journal: Bioscience, biotechnology, and biochemistry
ISSN: 0916-8451
DOI: 10.1080/09168451.2016.1263148
Accession Number: 27903134
Keywords: Acetylglucosamine/metabolism
Animals
Cell Differentiation
Glycosylation
Humans
Mice
Mice, Inbred Strains
Neurons/metabolism/pathology
RNA-Binding Protein EWS/chemistry/*metabolism
RNA-Binding Protein FUS/chemistry/*metabolism
TATA-Binding Protein Associated Factors/chemistry/*metabolism
*Ewing sarcoma protein (EWS)
*FET protein family
*O-GlcNAc
*chemoenzymatic labeling
*glycosylation stoichiometry
Abstract: Of the FET (fused in sarcoma [FUS]/Ewing sarcoma protein [EWS]/TATA binding protein-associated factor 15 [TAF15]) family of heterogeneous nuclear ribonucleoprotein particle proteins, FUS and TAF15 are consistently and EWS variably found in inclusion bodies in neurodegenerative diseases such as frontotemporal lobar degeneration associated with FUS. It is speculated that dysregulation of FET proteins at the post-translational level is involved in their cytoplasmic deposition. Here, the O-linked β-N-acetylglucosamine (O-GlcNAc) glycosylation stoichiometry of the FET proteins was chemoenzymatically analyzed, and it was found that only EWS is dynamically glycosylated with a high stoichiometry in the neural cell lines tested and in mouse brain. It was also confirmed that EWS, but not FUS and TAF15, is glycosylated with a high stoichiometry not only in the neural cells but also in the non-neural cell lines tested. These results indicate that O-GlcNAc glycosylation imparts a physicochemical property on EWS that is distinct from that of the other FET proteins in most of cell lineages or tissues.
Notes: 1347-6947
Kamemura, Kazuo
Comparative Study
Journal Article
England
Biosci Biotechnol Biochem. 2017 Mar;81(3):541-546. doi: 10.1080/09168451.2016.1263148. Epub 2016 Dec 1.
Author Address: a Department of Bioscience, Nagahama Institute of Bio-Science and Technology , Nagahama , Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 832
Author: Kamemura, K., Hayes, B. K., Comer, F. I. and Hart, G. W.
Year: 2002
Title: Dynamic interplay between O-glycosylation and O-phosphorylation of nucleocytoplasmic proteins: alternative glycosylation/phosphorylation of THR-58, a known mutational hot spot of c-Myc in lymphomas, is regulated by mitogens
Journal: J Biol Chem
Volume: 277
Issue: 21
Pages: 19229-35
Epub Date: 2002/03/21
Date: May 24
Short Title: Dynamic interplay between O-glycosylation and O-phosphorylation of nucleocytoplasmic proteins: alternative glycosylation/phosphorylation of THR-58, a known mutational hot spot of c-Myc in lymphomas, is regulated by mitogens
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M201729200
Accession Number: 11904304
Keywords: Amino Acid Sequence
Antibodies/immunology
Base Sequence
Cell Line
Cytoplasm/*metabolism
DNA Primers
Glycosylation
Humans
Lymphoma/*genetics
Mitogens/*pharmacology
Molecular Sequence Data
Nuclear Proteins/chemistry/*metabolism
Phosphorylation
*Protein Processing, Post-Translational
Proto-Oncogene Proteins c-myc/genetics/immunology/*metabolism
Threonine/*metabolism
Abstract: Previously, we reported that c-Myc is glycosylated by O-linked N-acetylglucosamine at Thr-58, a known phosphorylation site and a mutational hot spot in lymphomas. In this paper, we describe the production and characterization of two Thr-58 site-specific antibodies and use them to examine the modification of Thr-58 in living cells. One antibody specifically reacts with the Thr-58-glycosylated form of c-Myc, and the other reacts only with unmodified Thr-58 in c-Myc. Using these antibodies together with a commercial anti-Thr-58-phosphorylated c-Myc antibody, we simultaneously detected three forms of c-Myc (Thr-58-unmodified, -phosphorylated, and -glycosylated). It has been reported that Thr-58 phosphorylation is dependent on a prior phosphorylation of Ser-62. Mutagenesis of Ser-62 to Ala showed a marked decrease of Thr-58 phosphorylation and a marked increase of Thr-58 glycosylation. Growth inhibition of HL60 cells by serum starvation increases Thr-58 glycosylation and correspondingly decreases its phosphorylation. Serum stimulation has the opposite effect upon the modification status of Thr-58. A candidate kinase responsible for Thr-58 phosphorylation is the glycogen synthase kinase 3 (GSK3). Lithium, a competitive inhibitor of GSK3, decreased Thr-58 phosphorylation and increased its glycosylation. Finally, we show that the Thr-58-phosphorylated form of c-Myc predominantly accumulates in the cytoplasm rather than the nucleus upon inhibition of proteasome activity. These data suggest that hierarchical phosphorylation of Ser-62 and Thr-58 and alternative glycosylation/phosphorylation of Thr-58 together regulate the myriad functions of c-Myc in cells.
Notes: Kamemura, Kazuo
Hayes, Bradley K
Comer, Frank I
Hart, Gerald W
CA42486/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2002 May 24;277(21):19229-35. doi: 10.1074/jbc.M201729200. Epub 2002 Mar 19.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205-2185, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 233
Author: Kameyoshi, Y., Dörschner, A., Mallet, A. I., Christophers, E. and Schröder, J. M.
Year: 1992
Title: Cytokine RANTES released by thrombin-stimulated platelets is a potent attractant for human eosinophils
Journal: J Exp Med
Volume: 176
Issue: 2
Pages: 587-92
Epub Date: 1992/08/01
Date: Aug 1
Short Title: Cytokine RANTES released by thrombin-stimulated platelets is a potent attractant for human eosinophils
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.176.2.587
PMCID: PMC2119329
Accession Number: 1380064
Keywords: Amino Acid Sequence
Blood Platelets/metabolism/*physiology
Chemokine CCL5
Chemotactic Factors, Eosinophil/metabolism/*physiology
Chemotaxis, Leukocyte/physiology
Cytokines/metabolism/*physiology
Eosinophils/*physiology
Humans
Lymphokines/metabolism/*physiology
Molecular Sequence Data
Platelet Activation/physiology
Thrombin/*physiology
Abstract: Thrombin stimulation of human platelets results in the release of a preformed proteinaceous human eosinophil (Eo)-chemotactic activity. By the use of different high-performance liquid chromatography techniques, two Eo-chemotactic polypeptides (EoCPs), tentatively termed EoCP-1 and EoCP-2, were purified to homogeneity. Upon SDS-PAGE analysis, these chemotaxins showed molecular masses near 8 kD. NH2-terminal amino acid sequence analysis revealed identical sequences for both EoCP-1 and EoCP-2, which are also identical to that of RANTES, a cytokine that structurally belongs to the interleukin 8 superfamily of leukocyte selective attractants, and that is known to be a "memory-type" T lymphocyte-selective attractant. In the major Eo chemotaxin, EoCP-1, the residues 4 and 5, which in EoCP-2 were found to be serine residues, could not be identified. Electrospray mass spectrometry (ESP-MS) of EoCPs revealed for EoCP-2 a molecular mass of 7,862.8 +/- 1.1 daltons, which is 15.8 mass units higher than the calculated value of RANTES, indicating that EoCP-2 is identical to the full-length cytokine, and oxygenation, probably at methionine residue number 64, has taken place. Upon ESP-MS, EoCP-1 showed an average molecular mass of 8,355 +/- 10 daltons, suggesting O-glycosylation at these serine residues. Both natural forms of RANTES showed strong Eo-chemotactic activity (ED50 = 2 nM) with optimal chemotactic migration at concentrations near 10 nM, however, there were no significant migratory responses with human neutrophils. Chemotactic activity of RANTES for human Eos could be confirmed using recombinant material, which has been found to be as active as the natural forms. Since RANTES gene expression has been detected in activated T lymphocytes, and recombinant RANTES was shown to be a "memory" T lymphocyte-selective attractant, it is now tempting to speculate about an important role of RANTES in clinical situations such as allergene-induced late-phase skin reactions in atopic subjects or asthma, where in affected tissues both memory T cells and Eos are characteristic.
Notes: 1540-9538
Kameyoshi, Y
Dörschner, A
Mallet, A I
Christophers, E
Schröder, J M
Journal Article
Research Support, Non-U.S. Gov't
J Exp Med. 1992 Aug 1;176(2):587-92. doi: 10.1084/jem.176.2.587.
Author Address: Department of Dermatology, University of Kiel, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1089
Author: Kaneko, M. K., Kato, Y., Kameyama, A., Ito, H., Kuno, A., Hirabayashi, J., Kubota, T., Amano, K., Chiba, Y., Hasegawa, Y., Sasagawa, I., Mishima, K. and Narimatsu, H.
Year: 2007
Title: Functional glycosylation of human podoplanin: glycan structure of platelet aggregation-inducing factor
Journal: FEBS Lett
Volume: 581
Issue: 2
Pages: 331-6
Epub Date: 2007/01/16
Date: Jan 23
Short Title: Functional glycosylation of human podoplanin: glycan structure of platelet aggregation-inducing factor
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2006.12.044
Accession Number: 17222411
Keywords: Amino Acid Sequence
Animals
Blood Platelets/drug effects
CHO Cells
Carbohydrate Sequence
Cricetinae
Cricetulus
Glycosylation
Humans
Lectins/chemistry
Mass Spectrometry
Membrane Glycoproteins/*chemistry/isolation & purification/pharmacology
Molecular Sequence Data
N-Acetylneuraminic Acid/analysis
Organophosphorus Compounds/chemistry
Peptide Fragments/chemistry
Platelet Aggregation
Polysaccharides/analysis/*chemistry
Protein Array Analysis
Recombinant Proteins/chemistry/isolation & purification/pharmacology
Threonine/chemistry
Abstract: Podoplanin (Aggrus) is a mucin-type sialoglycoprotein that plays a key role in tumor cell-induced platelet aggregation. Podoplanin possesses a platelet aggregation-stimulating (PLAG) domain, and Thr52 in the PLAG domain of human podoplanin is important for its activity. Endogenous or recombinant human podoplanin were purified, and total glycosylation profiles were surveyed by lectin microarray. Analyses of glycopeptides produced by Edman degradation and mass spectrometry revealed that the disialyl-corel (NeuAc alpha2-3Gal beta l-3(NeuAc alpha2-6)GalNAc alpha l-O-Thr) structure was primarily attached to a glycosylation site at residue Thr52. Sialic acid-deficient podoplanin recovered its activity after additional sialylation. These results indicated that the sialylated Corel at Thr52 is critical for podoplanin-induced platelet aggregation.
Notes: Kaneko, Mika Kato
Kato, Yukinari
Kameyama, Akihiko
Ito, Hiromi
Kuno, Atsushi
Hirabayashi, Jun
Kubota, Tomomi
Amano, Koh
Chiba, Yasunori
Hasegawa, Yasushi
Sasagawa, Isoji
Mishima, Kazuhiko
Narimatsu, Hisashi
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2007 Jan 23;581(2):331-6. doi: 10.1016/j.febslet.2006.12.044. Epub 2007 Jan 11.
Author Address: Research Center for Medical Glycoscience, National Institute of Advanced Industrial Science and Technology, C-2, 1-1-1 Umezono, Tsukuba, Ibaraki 305 8568, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1269
Author: Kang, J. G., Park, S. Y., Ji, S., Jang, I., Park, S., Kim, H. S., Kim, S. M., Yook, J. I., Park, Y. I., Roth, J. and Cho, J. W.
Year: 2009
Title: O-GlcNAc protein modification in cancer cells increases in response to glucose deprivation through glycogen degradation
Journal: J Biol Chem
Volume: 284
Issue: 50
Pages: 34777-84
Epub Date: 2009/10/17
Date: Dec 11
Short Title: O-GlcNAc protein modification in cancer cells increases in response to glucose deprivation through glycogen degradation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.026351
PMCID: PMC2787340
Accession Number: 19833729
Keywords: AMP-Activated Protein Kinases/metabolism
Acetylglucosamine/*metabolism
Animals
Cell Line
Glucose/*metabolism
Glycogen/*metabolism
Humans
N-Acetylglucosaminyltransferases/metabolism
Neoplasms/*metabolism
*Protein Processing, Post-Translational
Abstract: When cellular glucose concentrations fall below normal levels, in general the extent of protein O-GlcNAc modification (O-GlcNAcylation) decreases. However, recent reports demonstrated increased O-GlcNAcylation by glucose deprivation in HepG2 and Neuro-2a cells. Here, we report increased O-GlcNAcylation in non-small cell lung carcinoma A549 cells and various other cells in response to glucose deprivation. Although the level of O-GlcNAc transferase was unchanged, the enzyme contained less O-GlcNAc, and its activity was increased. Moreover, O-GlcNAcase activity was reduced. The studied cells contain glycogen, and we show that its degradation in response to glucose deprivation provides a source for UDP-GlcNAc required for increased O-GlcNAcylation under this condition. This required active glycogen phosphorylase and resulted in increased glutamine:fructose-6-phosphate amidotransferase, the first and rate-limiting enzyme in the hexosamine biosynthetic pathway. Interestingly, glucose deprivation reduced the amount of phosphofructokinase 1, a regulatory glycolytic enzyme, and blocked ATP synthesis. These findings suggest that glycogen is the source for increased O-GlcNAcylation but not for generating ATP in response to glucose deprivation and that this may be useful for cancer cells to survive.
Notes: 1083-351x
Kang, Jeong Gu
Park, Sang Yoon
Ji, Suena
Jang, Insook
Park, Sujin
Kim, Hyun Sil
Kim, Sung-Min
Yook, Jong In
Park, Yong-Il
Roth, Jürgen
Cho, Jin Won
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2009 Dec 11;284(50):34777-84. doi: 10.1074/jbc.M109.026351. Epub 2009 Oct 15.
Author Address: Department of Biology, Oral Cancer Research Institute, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1278
Author: Kang, J. Y., Nan, X., Jin, M. S., Youn, S. J., Ryu, Y. H., Mah, S., Han, S. H., Lee, H., Paik, S. G. and Lee, J. O.
Year: 2009
Title: Recognition of lipopeptide patterns by Toll-like receptor 2-Toll-like receptor 6 heterodimer
Journal: Immunity
Volume: 31
Issue: 6
Pages: 873-84
Epub Date: 2009/11/26
Date: Dec 18
Short Title: Recognition of lipopeptide patterns by Toll-like receptor 2-Toll-like receptor 6 heterodimer
Alternate Journal: Immunity
ISSN: 1074-7613
DOI: 10.1016/j.immuni.2009.09.018
Accession Number: 19931471
Keywords: Acylation
Animals
Binding Sites
Crystallography, X-Ray
Hagfishes
Humans
Ligands
Lipopeptides/chemistry/*immunology
Lipopolysaccharides/chemistry/immunology/metabolism
Mice
Phosphatidylethanolamines/chemistry/immunology
Protein Multimerization
Recombinant Fusion Proteins/chemistry/immunology
Teichoic Acids/chemistry/immunology/metabolism
Toll-Like Receptor 1/chemistry/immunology
Toll-Like Receptor 2/chemistry/*immunology
Toll-Like Receptor 6/chemistry/*immunology
Abstract: Toll-like receptor 2 (TLR2) initiates potent immune responses by recognizing diacylated and triacylated lipopeptides. Its ligand specificity is controlled by whether it heterodimerizes with TLR1 or TLR6. We have determined the crystal structures of TLR2-TLR6-diacylated lipopeptide, TLR2-lipoteichoic acid, and TLR2-PE-DTPA complexes. PE-DTPA, 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine-N-diethylenetriaminepentaacetic acid, is a synthetic phospholipid derivative. Two major factors contribute to the ligand specificity of TLR2-TLR1 or TLR2-TLR6 heterodimers. First, the lipid channel of TLR6 is blocked by two phenylalanines. Simultaneous mutation of these phenylalanines made TLR2-TLR6 fully responsive not only to diacylated but also to triacylated lipopeptides. Second, the hydrophobic dimerization interface of TLR2-TLR6 is increased by 80%, which compensates for the lack of amide lipid interaction between the lipopeptide and TLR2-TLR6. The structures of the TLR2-lipoteichoic acid and the TLR2-PE-DTPA complexes demonstrate that a precise interaction pattern of the head group is essential for a robust immune response by TLR2 heterodimers.
Notes: 1097-4180
Kang, Jin Young
Nan, Xuehua
Jin, Mi Sun
Youn, Suk-Jun
Ryu, Young Hee
Mah, Shinjee
Han, Seung Hyun
Lee, Hayyoung
Paik, Sang-Gi
Lee, Jie-Oh
Journal Article
Research Support, Non-U.S. Gov't
United States
Immunity. 2009 Dec 18;31(6):873-84. doi: 10.1016/j.immuni.2009.09.018.
Author Address: Department of Chemistry, KAIST, Daejon, 305-701, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1785
Author: Kang, K. A., Piao, M. J., Ryu, Y. S., Kang, H. K., Chang, W. Y., Keum, Y. S. and Hyun, J. W.
Year: 2016
Title: Interaction of DNA demethylase and histone methyltransferase upregulates Nrf2 in 5-fluorouracil-resistant colon cancer cells
Journal: Oncotarget
Volume: 7
Issue: 26
Pages: 40594-40620
Epub Date: 2016/10/27
Date: Jun 28
Short Title: Interaction of DNA demethylase and histone methyltransferase upregulates Nrf2 in 5-fluorouracil-resistant colon cancer cells
Alternate Journal: Oncotarget
ISSN: 1949-2553
DOI: 10.18632/oncotarget.9745
PMCID: PMC5130030
Accession Number: 27259240
Keywords: Cell Line, Tumor
Colonic Neoplasms/*drug therapy/*metabolism
*Drug Resistance, Neoplasm
Fluorouracil/*pharmacology
Gene Expression Regulation, Neoplastic
Heme Oxygenase-1/metabolism
Histone Methyltransferases
Histone-Lysine N-Methyltransferase/*metabolism
Histones/metabolism
Humans
Hydrogen Peroxide/chemistry
Lysine/chemistry
Mixed Function Oxygenases/*metabolism
Myeloid-Lymphoid Leukemia Protein/metabolism
N-Acetylglucosaminyltransferases/metabolism
NF-E2-Related Factor 2/*metabolism
Promoter Regions, Genetic
Proto-Oncogene Proteins/*metabolism
RNA, Small Interfering/metabolism
Reactive Oxygen Species/metabolism
Up-Regulation
5-fluorouracil-resistance
DNA demethylase
Nrf2 transcription factor
histone methyltransferase
oxidative stress
Abstract: We recently reported that DNA demethylase ten-eleven translocation 1 (TET1) upregulates nuclear factor erythroid 2-related factor 2 (Nrf2) in 5-fluorouracil-resistant colon cancer cells (SNUC5/5-FUR). In the present study, we examined the effect of histone modifications on Nrf2 transcriptional activation. Histone deacetylase (HDAC) and histone acetyltransferase (HAT) were respectively decreased and increased in SNUC5/5-FUR cells as compared to non-resistant parent cells. Mixed-lineage leukemia (MLL), a histone methyltransferase, was upregulated, leading to increased trimethylation of histone H3 lysine 4, while G9a was downregulated, leading to decreased dimethylation of histone H3 lysine 9. siRNA-mediated MLL knockdown decreased levels of Nrf2 and HO-1 to a greater extent than did silencing HAT1. Host cell factor 1 (HCF1) was upregulated in SNUC5/5-FUR cells, and we observed interaction between HCF1 and MLL. Upregulation of O-GlcNAc transferase (OGT), an activator of HCF1, was also associated with HCF1-MLL interaction. In SNUC5/5-FUR cells, a larger fraction of OGT was bound to TET1, which recruits OGT to the Nrf2 promoter region, than in SNUC5 cells. These findings indicate that SNUC5/5-FUR cells are under oxidative stress, which induces expression of histone methylation-related proteins as well as DNA demethylase, leading to upregulation of Nrf2 and 5-FU resistance.
Notes: 1949-2553
Kang, Kyoung Ah
Piao, Mei Jing
Ryu, Yea Seong
Kang, Hee Kyoung
Chang, Weon Young
Keum, Young Sam
Hyun, Jin Won
Journal Article
Oncotarget. 2016 Jun 28;7(26):40594-40620. doi: 10.18632/oncotarget.9745.
Author Address: School of Medicine, Jeju National University, Jeju 63243, Republic of Korea.
Department of Biochemistry, College of Pharmacy, Dongguk University, Goyang 10326, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 650
Author: Kapron, J. T., Hilliard, G. M., Lakins, J. N., Tenniswood, M. P., West, K. A., Carr, S. A. and Crabb, J. W.
Year: 1997
Title: Identification and characterization of glycosylation sites in human serum clusterin
Journal: Protein Sci
Volume: 6
Issue: 10
Pages: 2120-33
Epub Date: 1997/10/23
Date: Oct
Short Title: Identification and characterization of glycosylation sites in human serum clusterin
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1002/pro.5560061007
PMCID: PMC2143570
Accession Number: 9336835
Keywords: Amino Acid Sequence
Binding Sites
Chromatography, High Pressure Liquid
Clusterin
Glycoproteins/*blood/*chemistry
Glycosylation
Humans
Mass Spectrometry
*Molecular Chaperones
Molecular Sequence Data
Molecular Weight
Oligosaccharides/chemistry/*metabolism
Peptide Fragments/chemistry
Abstract: Clusterin is a ubiquitous, heterodimeric glycoprotein with multiple possible functions that are likely influenced by glycosylation. Identification of oligosaccharide attachment sites and structural characterization of oligosaccharides in human serum clusterin has been performed by mass spectrometry and Edman degradation. Matrix-assisted laser desorption ionization mass spectrometry revealed two molecular weight species of holoclusterin (58,505 +/- 250 and 63,507 +/- 200). Mass spectrometry also revealed molecular heterogeneity associated with both the alpha and beta subunits of clusterin, consistent with the presence of multiple glycoforms. The data indicate that clusterin contains 17-27% carbohydrate by weight, the alpha subunit contains 0-30% carbohydrate and the beta subunit contains 27-30% carbohydrate. Liquid chromatography electrospray mass spectrometry with stepped collision energy scanning was used to selectively identify and preparatively fractionate tryptic glycopeptides. Edman sequence analysis was then used to confirm the identities of the glycopeptides and to define the attachment sites within each peptide. A total of six N-linked glycosylation sites were identified, three in the alpha subunit (alpha 64N, alpha 81N, alpha 123N) and three in the beta subunit (beta 64N, beta 127N, and beta 147N). Seven different possible types of oligosaccharide structures were identified by mass including: a monosialobiantennary structure, bisialobiantennary structures without or with one fucose, trisialotriantennary structures without or with one fucose, and possibly a trisialotriantennary structure with two fucose and/or a tetrasialotriantennary structure. Site beta 64N exhibited the least glycosylation diversity, with two detected types of oligosaccharides, and site beta 147N exhibited the greatest diversity, with five or six detected types of oligosaccharides. Overall, the most abundant glycoforms detected were bisialobiantennary without fucose and the least abundant were monosialobiantennary, trisialotriantennary with two fucose and/or tetrasialotriantennary. Clusterin peptides accounting for 99% of the primary structure were identified from analysis of the isolated alpha and beta subunits, including all Ser- and Thr-containing peptides. No evidence was found for the presence of O-linked or sulfated oligosaccharides. The results provide a molecular basis for developing a better understanding of clusterin structure-function relationships and the role clusterin glycosylation plays in physiological function.
Notes: 1469-896x
Kapron, J T
Hilliard, G M
Lakins, J N
Tenniswood, M P
West, K A
Carr, S A
Crabb, J W
DK38639/DK/NIDDK NIH HHS/United States
EY06603/EY/NEI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Protein Sci. 1997 Oct;6(10):2120-33. doi: 10.1002/pro.5560061007.
Author Address: W. Alton Jones Cell Science Center, Lake Placid, New York 12946, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1771
Author: Kapuria, V., Röhrig, U. F., Bhuiyan, T., Borodkin, V. S., van Aalten, D. M., Zoete, V. and Herr, W.
Year: 2016
Title: Proteolysis of HCF-1 by Ser/Thr glycosylation-incompetent O-GlcNAc transferase:UDP-GlcNAc complexes
Journal: Genes Dev
Volume: 30
Issue: 8
Pages: 960-72
Epub Date: 2016/04/09
Date: Apr 15
Short Title: Proteolysis of HCF-1 by Ser/Thr glycosylation-incompetent O-GlcNAc transferase:UDP-GlcNAc complexes
Alternate Journal: Genes & development
ISSN: 0890-9369 (Print)
0890-9369
DOI: 10.1101/gad.275925.115
PMCID: PMC4840301
Accession Number: 27056667
Keywords: Amino Acid Motifs
Animals
Catalytic Domain
Computer Simulation
Evolution, Molecular
Host Cell Factor C1/*metabolism
Humans
Invertebrates/enzymology
Models, Molecular
Mutation
N-Acetylglucosaminyltransferases/*metabolism
Protein Processing, Post-Translational
Protein Structure, Tertiary
*Proteolysis
O-GlcNAcylation
post-translation modifications
proteolysis
Abstract: In complex with the cosubstrate UDP-N-acetylglucosamine (UDP-GlcNAc),O-linked-GlcNAc transferase (OGT) catalyzes Ser/ThrO-GlcNAcylation of many cellular proteins and proteolysis of the transcriptional coregulator HCF-1. Such a dual glycosyltransferase-protease activity, which occurs in the same active site, is unprecedented and integrates both reversible and irreversible forms of protein post-translational modification within one enzyme. Although occurring within the same active site, we show here that glycosylation and proteolysis occur through separable mechanisms. OGT consists of tetratricopeptide repeat (TPR) and catalytic domains, which, together with UDP-GlcNAc, are required for both glycosylation and proteolysis. Nevertheless, a specific TPR domain contact with the HCF-1 substrate is critical for proteolysis but not Ser/Thr glycosylation. In contrast, key catalytic domain residues and even a UDP-GlcNAc oxygen important for Ser/Thr glycosylation are irrelevant for proteolysis. Thus, from a dual glycosyltransferase-protease, essentially single-activity enzymes can be engineered both in vitro and in vivo. Curiously, whereas OGT-mediated HCF-1 proteolysis is limited to vertebrate species, invertebrate OGTs can cleave human HCF-1. We present a model for the evolution of HCF-1 proteolysis by OGT.
Notes: 1549-5477
Kapuria, Vaibhav
Röhrig, Ute F
Bhuiyan, Tanja
Borodkin, Vladimir S
van Aalten, Daan M F
Zoete, Vincent
Herr, Winship
WT087590MA/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Genes Dev. 2016 Apr 15;30(8):960-72. doi: 10.1101/gad.275925.115. Epub 2016 Apr 7.
Author Address: Center for Integrative Genomics, University of Lausanne, Lausanne 1015, Switzerland;
Molecular Modeling Group, SIB Swiss Institute of Bioinformatics, Lausanne 1015, Switzerland;
MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1968
Author: Kapuria, V., Röhrig, U. F., Waridel, P., Lammers, F., Borodkin, V. S., van Aalten, D. M. F., Zoete, V. and Herr, W.
Year: 2018
Title: The conserved threonine-rich region of the HCF-1(PRO) repeat activates promiscuous OGT:UDP-GlcNAc glycosylation and proteolysis activities
Journal: J Biol Chem
Volume: 293
Issue: 46
Pages: 17754-17768
Epub Date: 2018/09/19
Date: Nov 16
Short Title: The conserved threonine-rich region of the HCF-1(PRO) repeat activates promiscuous OGT:UDP-GlcNAc glycosylation and proteolysis activities
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA118.004185
PMCID: PMC6240873
Accession Number: 30224358
Keywords: Amino Acid Motifs
Amino Acid Sequence
Endopeptidases/genetics/*metabolism
Glycosylation
Host Cell Factor C1/genetics/*metabolism
Humans
Molecular Dynamics Simulation
Multifunctional Enzymes/genetics/metabolism
Mutation
N-Acetylglucosaminyltransferases/genetics/*metabolism
Proteolysis
Stereoisomerism
Substrate Specificity
Uridine Diphosphate N-Acetylglucosamine/analogs & derivatives/metabolism
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
*enzyme mechanism
*glycobiology
*host-cell factor-1
*post-translational modification (PTM)
this article
Abstract: O-Linked GlcNAc transferase (OGT) possesses dual glycosyltransferase-protease activities. OGT thereby stably glycosylates serines and threonines of numerous proteins and, via a transient glutamate glycosylation, cleaves a single known substrate-the so-called HCF-1(PRO) repeat of the transcriptional co-regulator host-cell factor 1 (HCF-1). Here, we probed the relationship between these distinct glycosylation and proteolytic activities. For proteolysis, the HCF-1(PRO) repeat possesses an important extended threonine-rich region that is tightly bound by the OGT tetratricopeptide-repeat (TPR) region. We report that linkage of this HCF-1(PRO)-repeat, threonine-rich region to heterologous substrate sequences also potentiates robust serine glycosylation with the otherwise poor R (p)-αS-UDP-GlcNAc diastereomer phosphorothioate and UDP-5S-GlcNAc OGT co-substrates. Furthermore, it potentiated proteolysis of a non-HCF-1(PRO)-repeat cleavage sequence, provided it contained an appropriately positioned glutamate residue. Using serine- or glutamate-containing HCF-1(PRO)-repeat sequences, we show that proposed OGT-based or UDP-GlcNAc-based serine-acceptor residue activation mechanisms can be circumvented independently, but not when disrupted together. In contrast, disruption of both proposed activation mechanisms even in combination did not inhibit OGT-mediated proteolysis. These results reveal a multiplicity of OGT glycosylation strategies, some leading to proteolysis, which could be targets of alternative molecular regulatory strategies.
Notes: 1083-351x
Kapuria, Vaibhav
Röhrig, Ute F
Waridel, Patrice
Lammers, Fabienne
Borodkin, Vladimir S
van Aalten, Daan M F
Zoete, Vincent
Herr, Winship
110061/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2018 Nov 16;293(46):17754-17768. doi: 10.1074/jbc.RA118.004185. Epub 2018 Sep 17.
Author Address: From the Center for Integrative Genomics, University of Lausanne, 1015 Lausanne, Switzerland.
Molecular Modelling Group, SIB Swiss Institute of Bioinformatics, Lausanne 1015, Switzerland.
Protein Analysis Facility, Center for Integrative Genomics, Faculty of Biology and Medicine, University of Lausanne, 1015 Lausanne, Switzerland.
Centre for Gene Regulation and Expression, School of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, United Kingdom.
Molecular Modelling Group, SIB Swiss Institute of Bioinformatics, Lausanne 1015, Switzerland; Department of Fundamental Oncology, Ludwig Lausanne Branch, Faculty of Biology and Medicine, University of Lausanne, 1066 Epalinges, Switzerland.
From the Center for Integrative Genomics, University of Lausanne, 1015 Lausanne, Switzerland. Electronic address: winship.herr@unil.ch.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1646
Author: Karakas, E. and Furukawa, H.
Year: 2014
Title: Crystal structure of a heterotetrameric NMDA receptor ion channel
Journal: Science
Volume: 344
Issue: 6187
Pages: 992-7
Epub Date: 2014/05/31
Date: May 30
Short Title: Crystal structure of a heterotetrameric NMDA receptor ion channel
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1251915
PMCID: PMC4113085
NIHMSID: NIHMS605196
Accession Number: 24876489
Keywords: Animals
Binding Sites
Calcium/chemistry/metabolism
Crystallography, X-Ray
Protein Multimerization
Protein Structure, Secondary
Protein Structure, Tertiary
Rats
Receptors, N-Methyl-D-Aspartate/*chemistry/metabolism
Abstract: N-Methyl-D-aspartate (NMDA) receptors belong to the family of ionotropic glutamate receptors, which mediate most excitatory synaptic transmission in mammalian brains. Calcium permeation triggered by activation of NMDA receptors is the pivotal event for initiation of neuronal plasticity. Here, we show the crystal structure of the intact heterotetrameric GluN1-GluN2B NMDA receptor ion channel at 4 angstroms. The NMDA receptors are arranged as a dimer of GluN1-GluN2B heterodimers with the twofold symmetry axis running through the entire molecule composed of an amino terminal domain (ATD), a ligand-binding domain (LBD), and a transmembrane domain (TMD). The ATD and LBD are much more highly packed in the NMDA receptors than non-NMDA receptors, which may explain why ATD regulates ion channel activity in NMDA receptors but not in non-NMDA receptors.
Notes: 1095-9203
Karakas, Erkan
Furukawa, Hiro
R01 GM105730/GM/NIGMS NIH HHS/United States
R01 MH085926/MH/NIMH NIH HHS/United States
MH085926/MH/NIMH NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2014 May 30;344(6187):992-7. doi: 10.1126/science.1251915.
Author Address: Cold Spring Harbor Laboratory, W. M. Keck Structural Biology Laboratory, One Bungtown Road, Cold Spring Harbor, NY 11724, USA.
Cold Spring Harbor Laboratory, W. M. Keck Structural Biology Laboratory, One Bungtown Road, Cold Spring Harbor, NY 11724, USA. furukawa@cshl.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1277
Author: Karakas, E., Simorowski, N. and Furukawa, H.
Year: 2009
Title: Structure of the zinc-bound amino-terminal domain of the NMDA receptor NR2B subunit
Journal: Embo j
Volume: 28
Issue: 24
Pages: 3910-20
Epub Date: 2009/11/17
Date: Dec 16
Short Title: Structure of the zinc-bound amino-terminal domain of the NMDA receptor NR2B subunit
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2009.338
PMCID: PMC2797058
Accession Number: 19910922
Keywords: Allosteric Site
Animals
Brain/embryology
Crystallography, X-Ray/methods
Electrophysiology/methods
Female
Glutamic Acid/chemistry
Molecular Conformation
Oocytes/metabolism
Protein Binding
Protein Structure, Tertiary
Rats
Receptors, N-Methyl-D-Aspartate/*chemistry
Xenopus laevis/metabolism
Zinc/chemistry
Abstract: N-methyl-D-aspartate (NMDA) receptors belong to the family of ionotropic glutamate receptors (iGluRs) that mediate the majority of fast excitatory synaptic transmission in the mammalian brain. One of the hallmarks for the function of NMDA receptors is that their ion channel activity is allosterically regulated by binding of modulator compounds to the extracellular amino-terminal domain (ATD) distinct from the L-glutamate-binding domain. The molecular basis for the ATD-mediated allosteric regulation has been enigmatic because of a complete lack of structural information on NMDA receptor ATDs. Here, we report the crystal structures of ATD from the NR2B NMDA receptor subunit in the zinc-free and zinc-bound states. The structures reveal the overall clamshell-like architecture distinct from the non-NMDA receptor ATDs and molecular determinants for the zinc-binding site, ion-binding sites, and the architecture of the putative phenylethanolamine-binding site.
Notes: 1460-2075
Karakas, Erkan
Simorowski, Noriko
Furukawa, Hiro
P30 EB009998/EB/NIBIB NIH HHS/United States
R01 MH085926/MH/NIMH NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2009 Dec 16;28(24):3910-20. doi: 10.1038/emboj.2009.338.
Author Address: WM Keck Structural Biology Laboratory, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 651
Author: Karlsson, N. G., Herrmann, A., Karlsson, H., Johansson, M. E., Carlstedt, I. and Hansson, G. C.
Year: 1997
Title: The glycosylation of rat intestinal Muc2 mucin varies between rat strains and the small and large intestine. A study of O-linked oligosaccharides by a mass spectrometric approach
Journal: J Biol Chem
Volume: 272
Issue: 43
Pages: 27025-34
Epub Date: 1997/10/27
Date: Oct 24
Short Title: The glycosylation of rat intestinal Muc2 mucin varies between rat strains and the small and large intestine. A study of O-linked oligosaccharides by a mass spectrometric approach
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.43.27025
Accession Number: 9341141
Keywords: ABO Blood-Group System/chemistry
Animals
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Gel
Epitopes/analysis
Gas Chromatography-Mass Spectrometry/methods
Glycopeptides/chemistry/isolation & purification
Glycosylation
Intestinal Mucosa/chemistry/*metabolism
Intestine, Large/*metabolism
Intestine, Small/*metabolism
Molecular Sequence Data
Mucin-2
Mucins/biosynthesis/*chemistry/*metabolism
Neoplasm Proteins/chemistry/metabolism
Oligosaccharides/*chemistry/isolation & purification
Organ Specificity
Rats
Sialic Acids/analysis
Abstract: The large glycosylated domains obtained from the rat intestinal mucin Muc2 were isolated from the large and small intestine of the inbred rat strains GOT-W and GOT-BW. The expression of the rat Muc2 in the large intestine was confirmed immunochemically and by Northern blotting. Released oligosaccharides were structurally characterized by gas chromatography-mass spectrometry (neutral and sialylated species) or by tandem mass spectrometry (sulfated species), and a total of 63 structures was assigned. The large intestinal oligosaccharides were found to be identical between the strains, while the small intestinal glycosylation differed. Until now, detailed structural analysis of oligosaccharides isolated from a single mucin core or mucin domain with different origin have not been performed, and the information of different mucin glycoforms has been limited to immunochemistry. Blood group A-determinants (GalNAcalpha1-3(Fucalpha1-2)Galbeta1-, and structures related to the blood group Sda/Cad-related epitope NeuAc/NeuGcalpha1-3(GalNAcbeta1-4)Galbeta1-, were found in GOT-BW small intestine, and also in both large intestines. Blood group H-determinants and NeuAc/NeuGcalpha1-3Galbeta1- were found in all samples. Core 1 (Galbeta1-3GalNAcalpha1-), core 2 (Galbeta1-3(GlcNAcbeta1-6)GalNAcalpha1-), core 3 (GlcNAcbeta1-3GalNAcalpha1-), and core 4 (GlcNAcbeta1-3(GlcNAcbeta1-6)GalNAcalpha1- were also found in all the samples. The large intestine were enriched in sulfated oligosaccharides and the small intestine contained higher amounts of sialylated species. Sulfation were found exclusively on C-6 of GlcNAc.
Notes: Karlsson, N G
Herrmann, A
Karlsson, H
Johansson, M E
Carlstedt, I
Hansson, G C
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1997 Oct 24;272(43):27025-34. doi: 10.1074/jbc.272.43.27025.
Author Address: Department of Medical Biochemistry, Göteborg University, Medicinaregatan 9A, 413 90 Gothenburg, Sweden. Niclas.Karlsson@medkem.gu.se
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 622
Author: Karlsson, N. G., Johansson, M. E., Asker, N., Karlsson, H., Gendler, S. J., Carlstedt, I. and Hansson, G. C.
Year: 1996
Title: Molecular characterization of the large heavily glycosylated domain glycopeptide from the rat small intestinal Muc2 mucin
Journal: Glycoconj J
Volume: 13
Issue: 5
Pages: 823-31
Epub Date: 1996/10/01
Date: Oct
Short Title: Molecular characterization of the large heavily glycosylated domain glycopeptide from the rat small intestinal Muc2 mucin
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/bf00702346
Accession Number: 8910009
Keywords: Animals
Blotting, Northern
Blotting, Southern
Carbohydrate Sequence
Electrophoresis, Agar Gel
Glycopeptides/*chemistry/immunology
Immunohistochemistry
Intestinal Mucosa/*chemistry
Intestine, Small/metabolism
Mass Spectrometry
Molecular Sequence Data
Mucin-2
Mucins/*chemistry/immunology/isolation & purification
Oligosaccharides/chemistry
Polymerase Chain Reaction
RNA, Messenger/metabolism
Rats
Rats, Inbred Strains
Sulfuric Acid Esters/chemistry
Abstract: The largest high-glycosylated domain, glycopeptide A, of the "insoluble' mucin complex of the rat small intestine has earlier been purified and characterized (Carlstedt et al., 1993, J Biol Chem 268: 18771-81). A rabbit antiserum raised against deglycosylated glycopeptide A was used to clone part of a mucin showing homology to the human MUC2 mucin (Hansson et al., 1994, Biochem Biophys Res Commun 198. 181-90). This serum specifically stained goblet cells (paranuclear) in the mouse small intestine. The size of the coding sequence of glycopeptide A was estimated by using reversed transcriptase PCR of mRNA from an inbred rat strain (GOT-W) using primers in the unique central and C-terminal parts of the proposed rat Muc2 sequences. The PCR and Southern blot of the PCR products showed a fragment of about 5.5 kb corresponding to about 1700 amino acids when the known Cys-rich sequences used for the primers were subtracted. This is slightly larger than the size estimated earlier by biochemical studies. The mRNA encoding the rat Muc2 was slightly smaller than the mRNA encoding the human MUC2 in a colorectal cell line. Although the size of glycopeptide A estimated from biochemical results and by PCR is not identical, the results obtained here further support that the "insoluble' mucin of the rat small intestine is encoded by the Muc2 gene. Most of the oligosaccharides in glycopeptide A were either neutral (40%) or sialylated (40%). The remaining ones were sulfated with the sulfate group attached to C-6 of N-acetylglucosamine linked to C-6 of the N-acetylgalactosaminitol as revealed by tandem mass spectrometry of the perdeuteroacetylated oligosaccharides. Eighteen oligosaccharides were found of which fourteen were characterized and found to be mostly novel. Our findings thus expand the current knowledge of the core peptide of the rat intestinal goblet cell mucin and provide a relatively complete picture of the glycosylation of a defined mucin domain.
Notes: Karlsson, N G
Johansson, M E
Asker, N
Karlsson, H
Gendler, S J
Carlstedt, I
Hansson, G C
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 1996 Oct;13(5):823-31. doi: 10.1007/BF00702346.
Author Address: Department of Medical Biochemistry, Göteborg University, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 728
Author: Karpa, K. D., Lidow, M. S., Pickering, M. T., Levenson, R. and Bergson, C.
Year: 1999
Title: N-linked glycosylation is required for plasma membrane localization of D5, but not D1, dopamine receptors in transfected mammalian cells
Journal: Mol Pharmacol
Volume: 56
Issue: 5
Pages: 1071-8
Epub Date: 1999/10/26
Date: Nov
Short Title: N-linked glycosylation is required for plasma membrane localization of D5, but not D1, dopamine receptors in transfected mammalian cells
Alternate Journal: Molecular pharmacology
ISSN: 0026-895X (Print)
0026-895x
DOI: 10.1124/mol.56.5.1071
Accession Number: 10531415
Keywords: Cell Membrane/drug effects/*metabolism
Cells, Cultured
Glycosylation
Humans
Ligands
Mutation
Oligosaccharides/pharmacology
Receptors, Dopamine D1/genetics/*metabolism
Receptors, Dopamine D5
Transfection
Tunicamycin/pharmacology
Abstract: We have analyzed the role of N-linked glycosylation in functional cell surface expression of the D1 and D5 dopamine receptor subtypes. Treatment of transfected HEK 293 cells with tunicamycin, an inhibitor of N-linked oligosaccharide addition, was found to prevent localization of D5 receptors in the plasma membrane. In contrast, tunicamycin treatment had no effect on the plasma membrane localization of the D1 receptor. Polymerase chain reaction mutagenesis was used to generate a panel of D5 receptors containing mutations in the three predicted sites of N-linked glycosylation. Expression of mutant receptors indicated that glycosylation of residue N7 was the major determinant of D5 receptor plasma membrane localization. Mutation of a comparable site in the D1 receptor at position N5 had no effect on the delivery of the D1 receptor to the cell surface. Tunicamycin treatment during receptor biosynthesis, but not N-glycosidase F digestion of mature receptors, abrogated binding of the D5 receptor antagonist [(3)H]SCH23390, suggesting that while oligosaccharide moieties play a key role in the cell surface expression of D5 receptors, they do not appear to contribute to the receptor's ligand binding properties. Together, our data indicate a differential requirement for N-linked glycosylation in functional cell surface expression of D1 and D5 dopamine receptors.
Notes: Karpa, K D
Lidow, M S
Pickering, M T
Levenson, R
Bergson, C
MH56608/MH/NIMH NIH HHS/United States
P50-MH44866/MH/NIMH NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Mol Pharmacol. 1999 Nov;56(5):1071-8. doi: 10.1124/mol.56.5.1071.
Author Address: Department of Pharmacology, Penn State College of Medicine, Milton S. Hershey Medical Center, Hershey, Pennsylvania 17033, USA. kjd136@psu.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 773
Author: Kastrup, I. B., Stevanovic, S., Arsequell, G., Valencia, G., Zeuthen, J., Rammensee, H. G., Elliott, T. and Haurum, J. S.
Year: 2000
Title: Lectin purified human class I MHC-derived peptides: evidence for presentation of glycopeptides in vivo
Journal: Tissue Antigens
Volume: 56
Issue: 2
Pages: 129-35
Epub Date: 2000/10/06
Date: Aug
Short Title: Lectin purified human class I MHC-derived peptides: evidence for presentation of glycopeptides in vivo
Alternate Journal: Tissue antigens
ISSN: 0001-2815 (Print)
0001-2815
DOI: 10.1034/j.1399-0039.2000.560203.x
Accession Number: 11019912
Keywords: Acetylglucosamine/immunology
Antigen Presentation/*immunology
Chromatography, Affinity
Chromatography, High Pressure Liquid
Galactosyltransferases
Glycopeptides/*immunology/metabolism
Glycosylation
Histocompatibility Antigens Class I/*immunology/*isolation & purification
Humans
Protein Binding/immunology
Protein Processing, Post-Translational/immunology
Sequence Analysis, Protein
*Wheat Germ Agglutinins
Abstract: Previously, using synthetic glycopeptides carrying a natural cytosolic type of monosaccharide O-beta-linked N-acetylglucosamine (GlcNAc) glycosylation of serine residues, we have shown that glycopeptides act as suitable substrates for TAP-mediated transport into the endoplasmic reticulum (ER), and that they bind efficiently to class I major histocompatibility complex (MHC) molecules and can elicit glycopeptide-specific cytotoxic T-lymphocyte (CTL) responses in mice. Recently, we have reported that peptides presented by human class I MHC molecules in vivo encompass a small but significant amount of peptides which seem to be carrying O-beta-linked monosaccharide GlcNAc. In the present report we provide further evidence that glycosylated peptides are indeed presented by class I MHC molecules in vivo. Thus, peptides derived from HLA-A*0201 were purified by wheat germ agglutinin (WGA) lectin affinity chromatography as previously described. Subsequently, the peptides contained in the WGA-eluate were subjected to sequence analysis by Edman degradation. It was found that the peptides derived from HLA-A*0201 which had been retained by the O-GlcNAc-binding lectin WGA did indeed carry a HLA-A*0201 binding motif. Furthermore, using an enzymatic labeling procedure we present evidence that the HLA-A*0201-derived peptides which bind to the WGA lectin are glycosylated with terminal GlcNAc residues. Together, these data provide further evidence for the natural presentation by human class I MHC of glycopeptides carrying terminal O-GlcNAc residues in vivo.
Notes: Kastrup, I B
Stevanovic, S
Arsequell, G
Valencia, G
Zeuthen, J
Rammensee, H G
Elliott, T
Haurum, J S
Journal Article
Research Support, Non-U.S. Gov't
England
Tissue Antigens. 2000 Aug;56(2):129-35. doi: 10.1034/j.1399-0039.2000.560203.x.
Author Address: Institute of Cancer Biology, Danish Cancer Society, Copenhagen.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1041
Author: Kato, K., Jeanneau, C., Tarp, M. A., Benet-Pagès, A., Lorenz-Depiereux, B., Bennett, E. P., Mandel, U., Strom, T. M. and Clausen, H.
Year: 2006
Title: Polypeptide GalNAc-transferase T3 and familial tumoral calcinosis. Secretion of fibroblast growth factor 23 requires O-glycosylation
Journal: J Biol Chem
Volume: 281
Issue: 27
Pages: 18370-7
Epub Date: 2006/04/28
Date: Jul 7
Short Title: Polypeptide GalNAc-transferase T3 and familial tumoral calcinosis. Secretion of fibroblast growth factor 23 requires O-glycosylation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M602469200
Accession Number: 16638743
Keywords: Amino Acid Motifs
Animals
Binding, Competitive
CHO Cells
Calcinosis/*genetics/metabolism
Cricetinae
Fibroblast Growth Factors/*metabolism
Glycosylation
Humans
Metabolic Diseases/genetics
Molecular Sequence Data
N-Acetylgalactosaminyltransferases/genetics/*metabolism
Neoplasm Proteins/genetics/*metabolism
Proprotein Convertases/genetics/metabolism
*Protein Processing, Post-Translational
Substrate Specificity
Abstract: Mutations in the gene encoding the glycosyltransferase polypeptide GalNAc-T3, which is involved in initiation of O-glycosylation, were recently identified as a cause of the rare autosomal recessive metabolic disorder familial tumoral calcinosis (OMIM 211900). Familial tumoral calcinosis is associated with hyperphosphatemia and massive ectopic calcifications. Here, we demonstrate that the secretion of the phosphaturic factor fibroblast growth factor 23 (FGF23) requires O-glycosylation, and that GalNAc-T3 selectively directs O-glycosylation in a subtilisin-like proprotein convertase recognition sequence motif, which blocks processing of FGF23. The study suggests a novel posttranslational regulatory model of FGF23 involving competing O-glycosylation and protease processing to produce intact FGF23.
Notes: Kato, Kentaro
Jeanneau, Charlotte
Tarp, Mads Agervig
Benet-Pagès, Anna
Lorenz-Depiereux, Bettina
Bennett, Eric Paul
Mandel, Ulla
Strom, Tim M
Clausen, Henrik
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Jul 7;281(27):18370-7. doi: 10.1074/jbc.M602469200. Epub 2006 Apr 25.
Author Address: Department of Medical Biochemistry and Genetics, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1520
Author: Kato, K., Nishimasu, H., Okudaira, S., Mihara, E., Ishitani, R., Takagi, J., Aoki, J. and Nureki, O.
Year: 2012
Title: Crystal structure of Enpp1, an extracellular glycoprotein involved in bone mineralization and insulin signaling
Journal: Proc Natl Acad Sci U S A
Volume: 109
Issue: 42
Pages: 16876-81
Epub Date: 2012/10/03
Date: Oct 16
Short Title: Crystal structure of Enpp1, an extracellular glycoprotein involved in bone mineralization and insulin signaling
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1208017109
PMCID: PMC3479499
Accession Number: 23027977
Keywords: Animals
Calcification, Physiologic/*genetics
Crystallography, X-Ray
Glycoproteins/*chemistry/metabolism
Insulin/*metabolism
Mice
*Models, Molecular
Phosphoric Diester Hydrolases/*chemistry/metabolism
*Protein Conformation
Pyrophosphatases/*chemistry/metabolism
Ribonucleotides/metabolism
Signal Transduction/*genetics
X-Ray Diffraction
Abstract: Enpp1 is a membrane-bound glycoprotein that regulates bone mineralization by hydrolyzing extracellular nucleotide triphosphates to produce pyrophosphate. Enpp1 dysfunction causes human diseases characterized by ectopic calcification. Enpp1 also inhibits insulin signaling, and an Enpp1 polymorphism is associated with insulin resistance. However, the precise mechanism by which Enpp1 functions in these cellular processes remains elusive. Here, we report the crystal structures of the extracellular region of mouse Enpp1 in complex with four different nucleotide monophosphates, at resolutions of 2.7-3.2 Å. The nucleotides are accommodated in a pocket formed by an insertion loop in the catalytic domain, explaining the preference of Enpp1 for an ATP substrate. Structural mapping of disease-associated mutations indicated the functional importance of the interdomain interactions. A structural comparison of Enpp1 with Enpp2, a lysophospholipase D, revealed marked differences in the domain arrangements and active-site architectures. Notably, the Enpp1 mutant lacking the insertion loop lost the nucleotide-hydrolyzing activity but instead gained the lysophospholipid-hydrolyzing activity of Enpp2. Our findings provide structural insights into how the Enpp family proteins evolved to exert their diverse cellular functions.
Notes: 1091-6490
Kato, Kazuki
Nishimasu, Hiroshi
Okudaira, Shinichi
Mihara, Emiko
Ishitani, Ryuichiro
Takagi, Junichi
Aoki, Junken
Nureki, Osamu
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2012 Oct 16;109(42):16876-81. doi: 10.1073/pnas.1208017109. Epub 2012 Oct 1.
Author Address: Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Bunkyo-ku, Tokyo 113-0032, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1795
Author: Kato, K., Satouh, Y., Nishimasu, H., Kurabayashi, A., Morita, J., Fujihara, Y., Oji, A., Ishitani, R., Ikawa, M. and Nureki, O.
Year: 2016
Title: Structural and functional insights into IZUMO1 recognition by JUNO in mammalian fertilization
Journal: Nat Commun
Volume: 7
Pages: 12198
Epub Date: 2016/07/16
Date: Jul 15
Short Title: Structural and functional insights into IZUMO1 recognition by JUNO in mammalian fertilization
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms12198
PMCID: PMC4947182
Accession Number: 27416963
Keywords: Animals
Binding Sites
Carrier Proteins/genetics/metabolism
Cell Adhesion
Crystallography, X-Ray
Egg Proteins
Female
Fertilization/*physiology
Folate Receptor 1/chemistry/metabolism
HEK293 Cells
Humans
Immunoglobulins/chemistry/genetics/*metabolism
Male
Membrane Proteins/chemistry/genetics/*metabolism
Mice, Knockout
Mutation
Receptors, Cell Surface/*chemistry/genetics/*metabolism
Sperm-Ovum Interactions/physiology
Tryptophan/genetics/metabolism
Abstract: Sperm-egg fusion is the critical step in mammalian fertilization, and requires the interaction between IZUMO1 on the sperm surface and JUNO (also known as folate receptor (FR) 4 or IZUMO1R) on the egg surface. Whereas other FRs bind and uptake folates, JUNO binds IZUMO1 and establishes the cell-cell adhesion. However, the mechanism of IZUMO1 recognition by JUNO has remained elusive. Here we report the crystal structure of mouse JUNO, at 2.3 Å resolution. A structural comparison of JUNO with the FRs revealed that JUNO and the FRs have similar overall structures, but JUNO lacks the folate-binding pocket, thereby explaining the inability of JUNO to bind folate. Further complementation of Juno knockout eggs with mutant Juno messenger RNAs revealed that the conserved, surface-exposed tryptophan residue of JUNO is required for sperm binding and fertilization. Our structure-based in vivo functional analyses provide a framework towards a mechanistic understanding of mammalian gamete recognition.
Notes: 2041-1723
Kato, Kazuki
Satouh, Yuhkoh
Orcid: 0000-0001-9207-9374
Nishimasu, Hiroshi
Kurabayashi, Arisa
Morita, Junko
Fujihara, Yoshitaka
Oji, Asami
Ishitani, Ryuichiro
Ikawa, Masahito
Orcid: 0000-0001-9859-6217
Nureki, Osamu
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2016 Jul 15;7:12198. doi: 10.1038/ncomms12198.
Author Address: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan.
Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
PRESTO, JST, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 279
Author: Kaushansky, K., Lopez, J. A. and Brown, C. B.
Year: 1992
Title: Role of carbohydrate modification in the production and secretion of human granulocyte macrophage colony-stimulating factor in genetically engineered and normal mesenchymal cells
Journal: Biochemistry
Volume: 31
Issue: 6
Pages: 1881-6
Epub Date: 1992/02/18
Date: Feb 18
Short Title: Role of carbohydrate modification in the production and secretion of human granulocyte macrophage colony-stimulating factor in genetically engineered and normal mesenchymal cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00121a042
Accession Number: 1737041
Keywords: Animals
Carbohydrate Conformation
Carbohydrates/*chemistry
Cell Line
Cricetinae
DNA/genetics
Endothelium, Vascular/metabolism
Fibroblasts/metabolism
*Genetic Engineering
Granulocyte-Macrophage Colony-Stimulating Factor/biosynthesis/*chemistry/genetics
Humans
Kidney
Mesoderm/metabolism
Mutagenesis, Site-Directed
Structure-Activity Relationship
Transfection
Abstract: Colony-stimulating factors (CSFs) are a group of acidic glycoproteins which stimulate the proliferation and differentiation of hematopoietic progenitor cells in vitro and stimulate hemopoiesis in vivo. Human GM-CSF contains two N-linked carbohydrate side chains of the complex acidic type and several sites of O-linked carbohydrate clustered on serine and threonine residues near the N-terminus of the molecule. Previous studies have failed to detect a significant functional role for the carbohydrate modification characteristic of human GM-CSF. Using permanent cell lines and transient expression systems which produce moderate to high levels of native or carbohydrate-deficient forms of the growth factor, the role of carbohydrate modification in the biosynthesis and secretion of GM-CSF was studied. Unlike a number of other secreted glycoproteins, the transient time and secretory efficiency of several carbohydrate-deficient mutants of GM-CSF are indistinguishable from those of the native growth factor in BHK, 293, COS, and ldlD cells. Furthermore, normal human endothelial cells and fibroblasts, which normally produce the growth factor, can synthesize and secrete GM-CSF that lacks all forms of carbohydrate modification. These studies help to point out the range of roles played by carbohydrate modification in the biosynthesis, assembly, and secretion of glycoprotein hormones.
Notes: Kaushansky, K
Lopez, J A
Brown, C B
AI 22772/AI/NIAID NIH HHS/United States
CA 31615/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1992 Feb 18;31(6):1881-6. doi: 10.1021/bi00121a042.
Author Address: Division of Hematology, University of Washington School of Medicine, Seattle 98195.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1559
Author: Kawaguchi, M., Okabe, T., Okudaira, S., Nishimasu, H., Ishitani, R., Kojima, H., Nureki, O., Aoki, J. and Nagano, T.
Year: 2013
Title: Screening and X-ray crystal structure-based optimization of autotaxin (ENPP2) inhibitors, using a newly developed fluorescence probe
Journal: ACS Chem Biol
Volume: 8
Issue: 8
Pages: 1713-21
Epub Date: 2013/05/22
Date: Aug 16
Short Title: Screening and X-ray crystal structure-based optimization of autotaxin (ENPP2) inhibitors, using a newly developed fluorescence probe
Alternate Journal: ACS chemical biology
ISSN: 1554-8929
DOI: 10.1021/cb400150c
Accession Number: 23688339
Keywords: Crystallography, X-Ray
Drug Evaluation, Preclinical/methods
Enzyme Activation/drug effects
Enzyme Inhibitors/*chemistry/pharmacology
Fluorescent Dyes/*chemistry
Inhibitory Concentration 50
Limit of Detection
Models, Molecular
Molecular Structure
Phosphoric Diester Hydrolases/*chemistry
Small Molecule Libraries/chemistry/pharmacology
Structure-Activity Relationship
Abstract: Autotaxin (ATX), also known as ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2), was originally identified as a tumor cell autocrine motility factor and was found to be identical to plasma lysophospholipase D, which is the predominant contributor to lysophosphatidic acid (LPA) production from lysophospholipids. ATX is therefore considered to regulate the physiological and pathological roles of LPA, including angiogenesis, lymphocyte trafficking, tissue fibrosis, and cancer cell invasion and metastasis. Thus, it is a potential therapeutic target. Here, we first developed a sensitive and specific ATX fluorescence probe, TG-mTMP, and used it to screen ATX inhibitors in a large chemical library. This probe, which is superior to previously available probes FS-3 and CPF4 in terms of sensitivity or specificity, enabled us to identify several novel ATX inhibitor scaffolds. We solved the crystal structures of ATX complexes with the hit compounds at high resolution (1.75-1.95 Å) and used this information to guide optimization of the structure of a selected inhibitor. The optimized compounds, 3BoA and its derivatives, exhibited potent ATX-inhibitory activity both in vitro and in vivo. These inhibitors are expected to be useful tools to understand the roles of ATX in vitro and in vivo and may also be candidate anti-ATX therapeutic agents.
Notes: 1554-8937
Kawaguchi, Mitsuyasu
Okabe, Takayoshi
Okudaira, Shinichi
Nishimasu, Hiroshi
Ishitani, Ryuichiro
Kojima, Hirotatsu
Nureki, Osamu
Aoki, Junken
Nagano, Tetsuo
Journal Article
Research Support, Non-U.S. Gov't
United States
ACS Chem Biol. 2013 Aug 16;8(8):1713-21. doi: 10.1021/cb400150c. Epub 2013 Jun 3.
Author Address: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1970
Author: Kawahara, R., Ortega, F., Rosa-Fernandes, L., Guimarães, V., Quina, D., Nahas, W., Schwämmle, V., Srougi, M., Leite, K. R. M., Thaysen-Andersen, M., Larsen, M. R. and Palmisano, G.
Year: 2018
Title: Distinct urinary glycoprotein signatures in prostate cancer patients
Journal: Oncotarget
Volume: 9
Issue: 69
Pages: 33077-33097
Epub Date: 2018/09/22
Date: Sep 4
Short Title: Distinct urinary glycoprotein signatures in prostate cancer patients
Alternate Journal: Oncotarget
ISSN: 1949-2553
DOI: 10.18632/oncotarget.26005
PMCID: PMC6145689
Accession Number: 30237853
Keywords: TMT-labeling
glycopeptide
glycoproteomics
prostate cancer
urine
Abstract: Novel biomarkers are needed to complement prostate specific antigen (PSA) in prostate cancer (PCa) diagnostic screening programs. Glycoproteins represent a hitherto largely untapped resource with a great potential as specific and sensitive tumor biomarkers due to their abundance in bodily fluids and their dynamic and cancer-associated glycosylation. However, quantitative glycoproteomics strategies to detect potential glycoprotein cancer markers from complex biospecimen are only just emerging. Here, we describe a glycoproteomics strategy for deep quantitative mapping of N- and O-glycoproteins in urine with a view to investigate the diagnostic value of the glycoproteome to discriminate PCa from benign prostatic hyperplasia (BPH), two conditions that remain difficult to clinically stratify. Total protein extracts were obtained, concentrated and digested from urine of six PCa patients (Gleason score 7) and six BPH patients. The resulting peptide mixtures were TMT-labeled and mixed prior to a multi-faceted sample processing including hydrophilic interaction liquid chromatography (HILIC) and titanium dioxide SPE based enrichment, endo-/exoglycosidase treatment and HILIC-HPLC pre-fractionation. The isolated N- and O-glycopeptides were detected and quantified using high resolution mass spectrometry. We accurately quantified 729 N-glycoproteins spanning 1,310 unique N-glycosylation sites and observed 954 and 965 unique intact N- and O-glycopeptides, respectively, across the two disease conditions. Importantly, a panel of 56 intact N-glycopeptides perfectly discriminated PCa and BPH (ROC: AUC = 1). This study has generated a panel of intact glycopeptides that has a potential for PCa detection.
Notes: 1949-2553
Kawahara, Rebeca
Ortega, Fabio
Rosa-Fernandes, Livia
Guimarães, Vanessa
Quina, Daniel
Nahas, Willian
Schwämmle, Veit
Srougi, Miguel
Leite, Katia R M
Thaysen-Andersen, Morten
Larsen, Martin R
Palmisano, Giuseppe
Journal Article
Oncotarget. 2018 Sep 4;9(69):33077-33097. doi: 10.18632/oncotarget.26005. eCollection 2018 Sep 4.
Author Address: Instituto de Ciências Biomédicas, Departamento de Parasitologia, Universidade de São Paulo, USP, São Paulo, Brazil.
Laboratório de Investigação Médica da Disciplina de Urologia da Faculdade de Medicina da USP, LIM55, São Paulo, Brazil.
Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.
Instituto do Câncer do Estado de São Paulo, ICESP, São Paulo, Brazil.
Department of Molecular Sciences, Macquarie University, Sydney, NSW, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 831
Author: Kawasaki, N., Haishima, Y., Ohta, M., Itoh, S., Hyuga, M., Hyuga, S. and Hayakawa, T.
Year: 2001
Title: Structural analysis of sulfated N-linked oligosaccharides in erythropoietin
Journal: Glycobiology
Volume: 11
Issue: 12
Pages: 1043-9
Epub Date: 2002/01/24
Date: Dec
Short Title: Structural analysis of sulfated N-linked oligosaccharides in erythropoietin
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/11.12.1043
Accession Number: 11805077
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Chromatography, Liquid
Cricetinae
Erythropoietin/*chemistry
Glycoside Hydrolases/metabolism
Humans
Magnetic Resonance Spectroscopy
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Recombinant Proteins
Spectrometry, Mass, Electrospray Ionization
Sugar Alcohols/chemistry
Sulfur/*chemistry
Abstract: We previously demonstrated that high-performance liquid chromatography with electrospray ionization mass spectrometry (LC/MS) equipped with a graphitized carbon column (GCC) is useful for the structural analysis of carbohydrates in glycoproteins. Using LC/MS with GCC, sulfated N-linked oligosaccharides were found in erythropoietin (EPO) expressed in baby hamster kidney cells. Sulfation occurs in a part of the N-linked oligosaccharides in the EPO. Sulfated monosaccharide residue in the sulfated N-linked oligosaccharide was determined by exoglycosidase digestion followed by sugar mapping by LC/MS. The linkage position and branch-location of the sulfate group in the tetraantennary oligosaccharide were analyzed by (1)H-nuclear magnetic resonance. It was suggested that sulfation occurs on the C-6 position of GlcNAc located in the GlcNAcbeta1-4Manalpha1-3 branch.
Notes: Kawasaki, N
Haishima, Y
Ohta, M
Itoh, S
Hyuga, M
Hyuga, S
Hayakawa, T
Journal Article
England
Glycobiology. 2001 Dec;11(12):1043-9. doi: 10.1093/glycob/11.12.1043.
Author Address: Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, 1-18-1, Kamiyoga, Setagaya-ku, Tokyo 158-8501 Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 268
Author: Kawashima, H., Murata, T., Yamamoto, K., Tateishi, A., Irimura, T. and Osawa, T.
Year: 1992
Title: A simple method for the release of asparagine-linked oligosaccharides from a glycoprotein purified by SDS-polyacrylamide gel electrophoresis
Journal: J Biochem
Volume: 111
Issue: 5
Pages: 620-2
Epub Date: 1992/05/01
Date: May
Short Title: A simple method for the release of asparagine-linked oligosaccharides from a glycoprotein purified by SDS-polyacrylamide gel electrophoresis
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a123807
Accession Number: 1639758
Keywords: Animals
Asialoglycoproteins/chemistry/isolation & purification
Asparagine/chemistry
Carbohydrate Sequence
Electrophoresis, Polyacrylamide Gel
Glycoproteins/chemistry/*isolation & purification
Humans
Hydrazines/chemistry
Immunoglobulin G/chemistry/isolation & purification
Molecular Sequence Data
Oligosaccharides/chemistry/*isolation & purification
Orosomucoid/analogs & derivatives/chemistry/isolation & purification
Sodium Dodecyl Sulfate
Abstract: A simple method for the release of oligosaccharides from glycoproteins separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) has been developed. Asialo-alpha 1-acid glycoprotein, which was tritiated at the nonreducing terminal D-galactopyranosyl residue by reduction with sodium borotritide after incubation with D-galactose oxidase, was used as a model compound. After electrophoretic separation of the glycoprotein, oligosaccharides were released by the use of a gas-phase hydrazinolysis apparatus. In the first method, the gel was stained with Coomassie Blue and the glycoprotein together with the gel was directly subjected to gas-phase hydrazinolysis after removal of water in a P2O5 desiccator. The recovery of released oligosaccharides was 25.9 +/- 2.4%, based on the amount of the glycoprotein loaded on the gel within the range of 3.5-28.5 micrograms. In the second method, the glycoprotein was electroblotted onto an Immobilon transfer membrane and was visualized by staining with Coomassie Blue. A small piece of the membrane with the corresponding band was cut out, dried in a desiccator and subjected to gas-phase hydrazinolysis. In this case, the recovery of released oligosaccharides was 15.2 +/- 1.0%. These procedures, particularly the first one, should be widely applicable for the isolation of oligosaccharides from glycoproteins separated by SDS-PAGE.
Notes: Kawashima, H
Murata, T
Yamamoto, K
Tateishi, A
Irimura, T
Osawa, T
Journal Article
England
J Biochem. 1992 May;111(5):620-2. doi: 10.1093/oxfordjournals.jbchem.a123807.
Author Address: Division of Chemical Toxicology and Immunochemistry, Faculty of Pharmaceutical Sciences, University of Tokyo.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1225
Author: Kawauchi, K., Araki, K., Tobiume, K. and Tanaka, N.
Year: 2009
Title: Loss of p53 enhances catalytic activity of IKKbeta through O-linked beta-N-acetyl glucosamine modification
Journal: Proc Natl Acad Sci U S A
Volume: 106
Issue: 9
Pages: 3431-6
Epub Date: 2009/02/10
Date: Mar 3
Short Title: Loss of p53 enhances catalytic activity of IKKbeta through O-linked beta-N-acetyl glucosamine modification
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0813210106
PMCID: PMC2651314
Accession Number: 19202066
Keywords: Acetylation/drug effects
Acetylglucosamine/*metabolism
Animals
*Biocatalysis
Cell Line
Glucose/pharmacology
Glycolysis
Humans
I-kappa B Kinase/genetics/*metabolism
Mice
Mice, Knockout
Tumor Suppressor Protein p53/*deficiency/genetics/*metabolism
Abstract: The IkappaB kinase (IKK)-NF-kappaB pathway plays a critical role in oncogenesis. Recently, we have shown that p53 regulates glucose metabolism through the IKK-NF-kappaB pathway and that, in the absence of p53, the positive feedback loop between IKK-NF-kappaB and glycolysis has an integral role in oncogene-induced cell transformation. Here, we demonstrate that IKKbeta, a component of the IKK complex, was constitutively modified with O-linked beta-N-acetyl glucosamine (O-GlcNAc) in both p53-deficient mouse embryonic fibroblasts (MEFs) and transformed human fibroblasts. In p53-deficient cells, the O-GlcNAcylated IKKbeta and the activating phosphorylation of IKK were decreased by p65/NF-kappaB knockdown or glucose depletion. We also found that high glucose induced the O-GlcNAcylation of IKKbeta and sustained the TNFalpha-dependent IKKbeta activity. Moreover, the O-GlcNAcase inhibitor streptozotocin intensified O-GlcNAcylation and concomitant activating phosphorylation of IKKbeta. Mutational analysis revealed that O-GlcNAcylation of IKKbeta occurred at Ser 733 in the C-terminal domain, which was identified as an inactivating phosphorylation site, suggesting that IKKbeta O-GlcNAcylation regulates its catalytic activity. Taken together, we propose a novel mechanism for the enhancement of NF-kappaB activity by loss of p53, which evokes positive feedback regulation from enhanced glucose metabolism to IKK in oncogenesis.
Notes: 1091-6490
Kawauchi, Keiko
Araki, Keigo
Tobiume, Kei
Tanaka, Nobuyuki
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2009 Mar 3;106(9):3431-6. doi: 10.1073/pnas.0813210106. Epub 2009 Feb 6.
Author Address: Department of Molecular Oncology, Institute of Gerontology, Nippon Medical School, Kawasaki, Kanagawa 211-8533, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 478
Author: Kehry, M. R., Fuhrman, J. S., Schilling, J. W., Rogers, J., Sibley, C. H. and Hood, L. E.
Year: 1982
Title: Complete amino acid sequence of a mouse mu chain: homology among heavy chain constant region domains
Journal: Biochemistry
Volume: 21
Issue: 22
Pages: 5415-24
Epub Date: 1982/10/26
Date: Oct 26
Short Title: Complete amino acid sequence of a mouse mu chain: homology among heavy chain constant region domains
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00265a006
Accession Number: 6816276
Keywords: Amino Acid Sequence
Animals
Base Sequence
Genes
Immunoglobulin Constant Regions/*analysis/genetics
Immunoglobulin Heavy Chains/*analysis/genetics
Immunoglobulin M/analysis
Immunoglobulin mu-Chains/*analysis/genetics
Immunoglobulins/*analysis
Mice
Neoplasms, Experimental/metabolism
Plasmacytoma/metabolism
Abstract: The complete amino acid sequence of the mouse mu chain secreted by the MOPC 104E myeloma tumor has been determined. There are four constant region domains in the mu chain and a 20-residue COOH-terminal segment that plays a role in the polymerization of pentameric immunoglobulin M molecules. There are six sites of carbohydrate attachment in the MOPC 104E mu chain. Three complex-type and two high-mannose oligosaccharides are located in the mu chain constant region. The general type and location of carbohydrate moieties in the mu chain constant region are completely conserved between mouse and human mu chains. Homology in the location of carbohydrate structures on different classes of heavy chains is discussed.
Notes: Kehry, M R
Fuhrman, J S
Schilling, J W
Rogers, J
Sibley, C H
Hood, L E
AI 13410/AI/NIAID NIH HHS/United States
AI 16950/AI/NIAID NIH HHS/United States
CA 20314/CA/NCI NIH HHS/United States
etc.
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1982 Oct 26;21(22):5415-24. doi: 10.1021/bi00265a006.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1086
Author: Keil, C., Maskos, K., Than, M., Hoopes, J. T., Huber, R., Tan, F., Deddish, P. A., Erdös, E. G., Skidgel, R. A. and Bode, W.
Year: 2007
Title: Crystal structure of the human carboxypeptidase N (kininase I) catalytic domain
Journal: J Mol Biol
Volume: 366
Issue: 2
Pages: 504-16
Epub Date: 2006/12/13
Date: Feb 16
Short Title: Crystal structure of the human carboxypeptidase N (kininase I) catalytic domain
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2006.11.025
Accession Number: 17157876
Keywords: Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Bradykinin/chemistry
*Catalytic Domain
Crystallography, X-Ray
Humans
Lysine Carboxypeptidase/*chemistry/genetics/isolation & purification
Models, Molecular
Molecular Sequence Data
Prealbumin/chemistry
*Protein Structure, Tertiary
Sequence Analysis, Protein
Sequence Homology, Amino Acid
Structure-Activity Relationship
Abstract: Human carboxypeptidase N (CPN), a member of the CPN/E subfamily of "regulatory" metallo-carboxypeptidases, is an extracellular glycoprotein synthesized in the liver and secreted into the blood, where it controls the activity of vasoactive peptide hormones, growth factors and cytokines by specifically removing C-terminal basic residues. Normally, CPN circulates in blood plasma as a hetero-tetramer consisting of two 83 kDa (CPN2) domains each flanked by a 48 to 55 kDa catalytic (CPN1) domain. We have prepared and crystallized the recombinant C-terminally truncated catalytic domain of human CPN1, and have determined and refined its 2.1 A crystal structure. The structural analysis reveals that CPN1 has a pear-like shape, consisting of a 319 residue N-terminal catalytic domain and an abutting, cylindrically shaped 79 residue C-terminal beta-sandwich transthyretin (TT) domain, more resembling CPD-2 than CPM. Like these other CPN/E members, two surface loops surrounding the active-site groove restrict access to the catalytic center, offering an explanation for why some larger protein carboxypeptidase inhibitors do not inhibit CPN. Modeling of the Pro-Phe-Arg C-terminal end of the natural substrate bradykinin into the active site shows that the S1' pocket of CPN1 might better accommodate P1'-Lys than Arg residues, in agreement with CPN's preference for cleaving off C-terminal Lys residues. Three Thr residues at the distal TT edge of CPN1 are O-linked to N-acetyl glucosamine sugars; equivalent sites in the membrane-anchored CPM are occupied by basic residues probably involved in membrane interaction. In tetrameric CPN, each CPN1 subunit might interact with the central leucine-rich repeat tandem of the cognate CPN2 subunit via a unique hydrophobic surface patch wrapping around the catalytic domain-TT interface, exposing the two active centers.
Notes: Keil, Cora
Maskos, Klaus
Than, Manuel
Hoopes, J Todd
Huber, Robert
Tan, Fulong
Deddish, Peter A
Erdös, Ervin G
Skidgel, Randal A
Bode, Wolfram
DK41431/DK/NIDDK NIH HHS/United States
HL36473/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2007 Feb 16;366(2):504-16. doi: 10.1016/j.jmb.2006.11.025. Epub 2006 Nov 11.
Author Address: Arbeitsgruppe Proteinaseforschung, Max-Planck-Institut für Biochemie, Am Klopferspitz 18, D-82152 Planegg-Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 377
Author: Kellermann, J., Lottspeich, F., Geiger, R. and Deutzmann, R.
Year: 1988
Title: Human urinary kallikrein--amino acid sequence and carbohydrate attachment sites
Journal: Protein Seq Data Anal
Volume: 1
Issue: 3
Pages: 177-82
Epub Date: 1988/02/01
Date: Feb
Short Title: Human urinary kallikrein--amino acid sequence and carbohydrate attachment sites
Alternate Journal: Protein sequences & data analysis
ISSN: 0931-9506 (Print)
0931-9506
Accession Number: 3163150
Keywords: Amino Acid Sequence
Amino Acids/analysis
Animals
Binding Sites
Carbohydrates
Humans
Kallikreins/*urine
Molecular Sequence Data
Serine Endopeptidases
Abstract: The amino acid sequence of human urinary kallikrein was determined by protein-chemical methods. The peptide chain contains 238 amino acid residues. Ten half-cystine residues form five disulfide bridges. Six carbohydrate side chains could be identified, three of them are N-glycosidically linked to asparagine which is present in the acceptor sequon Asn-Xaa-Thr (or -Ser) and three are O-glycosidically linked to two serine residues and one threonine residue, respectively. Comparison of the primary structure of human urinary kallikrein with that of other serine proteinases especially porcine kallikrein and rat kallikrein shows a high degree of homology. IVGGWECEQHSQPWQAALYHFSTFQCGGILVHRQWVLTA AHCISDNYQLWLGRHNLFDDENTAQFVHVSESFPHPGFN MSLLENHTRQADEDYSHDLMLLRLTEPADTITDAVKVVE LPTQEPEVGSTCLASGWGSIEPENFSFPDDLQCVDLKIL PNDECEKAHVQKVTDFMLCVGHLEGGKDTCVGDSGGPLM CDGVLQGVTSWGYVPCGTPNKPSVAVRVLSYVKWIEDTIAENS.
Notes: Kellermann, J
Lottspeich, F
Geiger, R
Deutzmann, R
Comparative Study
Journal Article
Germany
Protein Seq Data Anal. 1988 Feb;1(3):177-82.
Author Address: Max-Planck-Institut für Biochemie, Genzentrum, Martinsried, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 402
Author: Kellermann, J., Lottspeich, F., Henschen, A. and Müller-Esterl, W.
Year: 1986
Title: Completion of the primary structure of human high-molecular-mass kininogen. The amino acid sequence of the entire heavy chain and evidence for its evolution by gene triplication
Journal: Eur J Biochem
Volume: 154
Issue: 2
Pages: 471-8
Epub Date: 1986/01/15
Date: Jan 15
Short Title: Completion of the primary structure of human high-molecular-mass kininogen. The amino acid sequence of the entire heavy chain and evidence for its evolution by gene triplication
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1986.tb09421.x
Accession Number: 3484703
Keywords: Amino Acid Sequence
Biological Evolution
Carbohydrates/analysis
Chemical Phenomena
Chemistry
Chromatography, High Pressure Liquid
Computers
Cysteine Proteinase Inhibitors
Gene Amplification
Humans
Hydrolysis
Kininogens/*genetics
Molecular Weight
Protein Conformation
Proteins
Repetitive Sequences, Nucleic Acid
Trypsin
Abstract: The amino acid sequence of the heavy chain of human high-molecular-mass kininogen has been determined. It completes the primary structure of the high-Mr kininogen molecule. The heavy chain contains 362, the total kininogen molecule 626 amino acid residues. Three carbohydrate side chains were found in the heavy chain, all of them N-glycosidically linked to asparagine, which is present in the acceptor sequon Asn-Xaa-Thr (or -Ser); one additional potential glycosylation site devoid of a sugar side chain is found at position 30. There is a high degree of homology between the heavy chains of human high-Mr kininogen and bovine high-Mr kininogen (74% identity), or rat T-kininogen (61%). Comparison of the primary structure of human high-Mr kininogen with that of human low-Mr kininogen predicted from its cDNA sequence, reveals that the heavy chains of the two human kininogens are completely identical. Two heavy chain segments believed to contain the reactive sites for cysteine proteinase inhibition show an extensive sequence homology with other mammalian cysteine proteinase inhibitors. Within the heavy chain of human high-Mr kininogen are repetitive units strongly suggesting that the heavy chain of human kininogens has evolved from at least two ancestral units by a series of gene duplication and fusion events.
Notes: Kellermann, J
Lottspeich, F
Henschen, A
Müller-Esterl, W
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1986 Jan 15;154(2):471-8. doi: 10.1111/j.1432-1033.1986.tb09421.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 337
Author: Kentzer, E. J., Buko, A., Menon, G. and Sarin, V. K.
Year: 1990
Title: Carbohydrate composition and presence of a fucose-protein linkage in recombinant human pro-urokinase
Journal: Biochem Biophys Res Commun
Volume: 171
Issue: 1
Pages: 401-6
Epub Date: 1990/08/31
Date: Aug 31
Short Title: Carbohydrate composition and presence of a fucose-protein linkage in recombinant human pro-urokinase
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(90)91407-j
Accession Number: 2393398
Keywords: Amino Acid Sequence
Fucose/analysis/*metabolism
Glycosylation
Hexoses/analysis
Humans
Molecular Sequence Data
Peptide Fragments/analysis
Peptide Mapping
Protein Processing, Post-Translational
Recombinant Proteins
Trypsin
Urokinase-Type Plasminogen Activator/*metabolism
Abstract: A new post-translational modification site in the growth factor domain of urinary type plasminogen activator has been identified. A glycopeptide containing the monosaccharide, fucose, covalently linked directly to the peptide backbone has been isolated from the tryptic digest of pro-urokinase expressed in a mouse hybridoma cell line Sp 2/0 Ag 14. The glycopeptide was isolated by semi-preparative reversed phase high performance liquid chromatography. The identity of a fucose containing peptide was confirmed by carbohydrate analysis, amino acid analysis and plasma desorption mass spectrometry (PDMS). A combination of these methodologies showed an equimolar ratio of peptide and fucose in the glycopeptide. This modification is not detected without mass spectrometry because the fucose residue is hydrolyzed under standard acidic conditions of amino acid composition and N-terminal sequence analysis. The site of attachment of fucose to the peptide has been localized towards the N-terminus (within first 23 amino acids) of the protein. Also, the carbohydrate composition of recombinant pro-urokinase is reported.
Notes: Kentzer, E J
Buko, A
Menon, G
Sarin, V K
Journal Article
United States
Biochem Biophys Res Commun. 1990 Aug 31;171(1):401-6. doi: 10.1016/0006-291x(90)91407-j.
Author Address: PPD Analytical Research Department, Abbott Laboratories, North Chicago, Illinois 60064.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 221
Author: Kessler, M. J., Mise, T., Ghai, R. D. and Bahl, O. P.
Year: 1979
Title: Structure and location of the O-glycosidic carbohydrate units of human chorionic gonadotropin
Journal: J Biol Chem
Volume: 254
Issue: 16
Pages: 7909-14
Epub Date: 1979/08/25
Date: Aug 25
Short Title: Structure and location of the O-glycosidic carbohydrate units of human chorionic gonadotropin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 468798
Keywords: Amino Acid Sequence
Amino Acids/analysis
Carbohydrates/analysis
*Chorionic Gonadotropin
Glycopeptides/analysis
Glycosides/analysis
Humans
Peptide Fragments/analysis
Notes: Kessler, M J
Mise, T
Ghai, R D
Bahl, O P
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1979 Aug 25;254(16):7909-14.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 821
Author: Khandekar, S. S., Silverman, C., Wells-Marani, J., Bacon, A. M., Birrell, H., Brigham-Burke, M., DeMarini, D. J., Jonak, Z. L., Camilleri, P. and Fishman-Lobell, J.
Year: 2001
Title: Determination of carbohydrate structures N-linked to soluble CD154 and characterization of the interactions of CD40 with CD154 expressed in Pichia pastoris and Chinese hamster ovary cells
Journal: Protein Expr Purif
Volume: 23
Issue: 2
Pages: 301-10
Epub Date: 2001/10/26
Date: Nov
Short Title: Determination of carbohydrate structures N-linked to soluble CD154 and characterization of the interactions of CD40 with CD154 expressed in Pichia pastoris and Chinese hamster ovary cells
Alternate Journal: Protein expression and purification
ISSN: 1046-5928 (Print)
1046-5928
DOI: 10.1006/prep.2001.1501
Accession Number: 11676606
Keywords: Animals
Asparagine/chemistry
B-Lymphocytes/immunology
CD40 Antigens/*chemistry/genetics/immunology/metabolism
CD40 Ligand/*chemistry/genetics/immunology/metabolism
CHO Cells
Carbohydrate Conformation
Carbohydrate Metabolism
Carbohydrates/*chemistry
Cells, Cultured
Cloning, Molecular
Cricetinae
Humans
Mannose/chemistry/metabolism
Pichia/genetics
Protein Binding
Recombinant Proteins/chemistry/genetics/metabolism
Solubility
Abstract: CD40-CD154 (CD40 ligand) interactions are essential for the development of protective immunity. Previous studies have described the CD40 binding site as a shallow groove formed between two monomers of CD154. However, these studies have not examined the structure or biological function of the carbohydrate on CD154. Human CD154 contains a single N-linked glycosylation site at asparagine 240. We have characterized the interactions between CD40 and soluble (s) CD154 in which sCD154 contains different types of carbohydrates. Detailed carbohydrate analysis revealed high-mannose structures on sCD154 purified from Pichia pastoris, whereas CD154 purified from Chinese hamster ovary E1A contained heterogeneous populations of complex carbohydrates. sCD154 purified from either system was trimeric, it bound to CD40 with similar affinities of 10-30 nM, and it functionally induced CD69 and CD95 expression on primary B cells. Together, these results indicate that the presence of varied types of N-linked glycans on asparagine 240 of CD154 does not play a significant role in the CD40-CD154 interactions.
Notes: Khandekar, S S
Silverman, C
Wells-Marani, J
Bacon, A M
Birrell, H
Brigham-Burke, M
DeMarini, D J
Jonak, Z L
Camilleri, P
Fishman-Lobell, J
Journal Article
United States
Protein Expr Purif. 2001 Nov;23(2):301-10. doi: 10.1006/prep.2001.1501.
Author Address: Department of Protein Biochemistry, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1660
Author: Khatri, K., Staples, G. O., Leymarie, N., Leon, D. R., Turiák, L., Huang, Y., Yip, S., Hu, H., Heckendorf, C. F. and Zaia, J.
Year: 2014
Title: Confident assignment of site-specific glycosylation in complex glycoproteins in a single step
Journal: J Proteome Res
Volume: 13
Issue: 10
Pages: 4347-55
Epub Date: 2014/08/26
Date: Oct 3
Short Title: Confident assignment of site-specific glycosylation in complex glycoproteins in a single step
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr500506z
PMCID: PMC4184449
Accession Number: 25153361
Keywords: Amino Acid Sequence
Chromatography, Liquid
Glycoproteins/chemistry/*metabolism
Glycosylation
Hemagglutinin Glycoproteins, Influenza Virus/chemistry/metabolism
Mass Spectrometry
Orosomucoid/chemistry/metabolism
Proteomics
Cad
Glycopeptides
Hilic
tandem MS
Abstract: A glycoprotein may contain several sites of glycosylation, each of which is heterogeneous. As a consequence of glycoform diversity and signal suppression from nonglycosylated peptides that ionize more efficiently, typical reversed-phase LC-MS and bottom-up proteomics database searching workflows do not perform well for identification of site-specific glycosylation for complex glycoproteins. We present an LC-MS system for enrichment, separation, and analysis of glycopeptides from complex glycoproteins (>4 N-glycosylation sequons) in a single step. This system uses an online HILIC enrichment trap prior to reversed-phase C18-MS analysis. We demonstrated the effectiveness of the system using a set of glycoproteins including human transferrin (2 sequons), human alpha-1-acid glycoprotein (5 sequons), and influenza A virus hemagglutinin (9 sequons). The online enrichment renders glycopeptides the most abundant ions detected, thereby facilitating the generation of high-quality data-dependent tandem mass spectra. The tandem mass spectra exhibited product ions from both glycan and peptide backbone dissociation for a majority of the glycopeptides tested using collisionally activated dissociation that served to confidently assign site-specific glycosylation. We demonstrated the value of our system to define site-specific glycosylation using a hemagglutinin containing 9 N-glycosylation sequons from a single HILIC-C18-MS acquisition.
Notes: 1535-3907
Khatri, Kshitij
Staples, Gregory O
Leymarie, Nancy
Leon, Deborah R
Turiák, Lilla
Huang, Yu
Yip, Shun
Hu, Han
Heckendorf, Christian F
Zaia, Joseph
P41 GM104603/GM/NIGMS NIH HHS/United States
P41GM104603/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Proteome Res. 2014 Oct 3;13(10):4347-55. doi: 10.1021/pr500506z. Epub 2014 Sep 9.
Author Address: Center for Biomedical Mass Spectrometry, Department of Biochemistry, Boston University School of Medicine , Boston, Massachusetts 02118, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1975
Author: Khayat, W., Hackett, A., Shaw, M., Ilie, A., Dudding-Byth, T., Kalscheuer, V. M., Christie, L., Corbett, M. A., Juusola, J., Friend, K. L., Kirmse, B. M., Gecz, J., Field, M. and Orlowski, J.
Year: 2019
Title: A recurrent missense variant in SLC9A7 causes nonsyndromic X-linked intellectual disability with alteration of Golgi acidification and aberrant glycosylation
Journal: Hum Mol Genet
Volume: 28
Issue: 4
Pages: 598-614
Epub Date: 2018/10/20
Date: Feb 15
Short Title: A recurrent missense variant in SLC9A7 causes nonsyndromic X-linked intellectual disability with alteration of Golgi acidification and aberrant glycosylation
Alternate Journal: Human molecular genetics
ISSN: 0964-6906 (Print)
0964-6906
DOI: 10.1093/hmg/ddy371
PMCID: PMC6360272
Accession Number: 30335141
Keywords: Acids/metabolism
Animals
CHO Cells
Cell Membrane/genetics
Cricetinae
Cricetulus
Gene Expression Regulation/genetics
Genetic Diseases, X-Linked/*genetics/metabolism/pathology
Glycosylation
Golgi Apparatus/*genetics/metabolism
Humans
Intellectual Disability/*genetics/metabolism/pathology
Membrane Glycoproteins/genetics
Mutation, Missense/genetics
Protein Transport/genetics
Sodium-Hydrogen Exchangers/*genetics
Transfection
Viral Envelope Proteins/genetics
trans-Golgi Network/genetics
Abstract: We report two unrelated families with multigenerational nonsyndromic intellectual disability (ID) segregating with a recurrent de novo missense variant (c.1543C>T:p.Leu515Phe) in the alkali cation/proton exchanger gene SLC9A7 (also commonly referred to as NHE7). SLC9A7 is located on human X chromosome at Xp11.3 and has not yet been associated with a human phenotype. The gene is widely transcribed, but especially abundant in brain, skeletal muscle and various secretory tissues. Within cells, SLC9A7 resides in the Golgi apparatus, with prominent enrichment in the trans-Golgi network (TGN) and post-Golgi vesicles. In transfected Chinese hamster ovary AP-1 cells, the Leu515Phe mutant protein was correctly targeted to the TGN/post-Golgi vesicles, but its N-linked oligosaccharide maturation as well as that of a co-transfected secretory membrane glycoprotein, vesicular stomatitis virus G (VSVG) glycoprotein, was reduced compared to cells co-expressing SLC9A7 wild-type and VSVG. This correlated with alkalinization of the TGN/post-Golgi compartments, suggestive of a gain-of-function. Membrane trafficking of glycosylation-deficient Leu515Phe and co-transfected VSVG to the cell surface, however, was relatively unaffected. Mass spectrometry analysis of patient sera also revealed an abnormal N-glycosylation profile for transferrin, a clinical diagnostic marker for congenital disorders of glycosylation. These data implicate a crucial role for SLC9A7 in the regulation of TGN/post-Golgi pH homeostasis and glycosylation of exported cargo, which may underlie the cellular pathophysiology and neurodevelopmental deficits associated with this particular nonsyndromic form of X-linked ID.
Notes: 1460-2083
Khayat, Wujood
Hackett, Anna
Shaw, Marie
Ilie, Alina
Dudding-Byth, Tracy
Kalscheuer, Vera M
Christie, Louise
Corbett, Mark A
Juusola, Jane
Friend, Kathryn L
Kirmse, Brian M
Gecz, Jozef
Field, Michael
Orlowski, John
MOP-111191/CIHR/Canada
PJT-155976/CIHR/Canada
Journal Article
Research Support, Non-U.S. Gov't
Hum Mol Genet. 2019 Feb 15;28(4):598-614. doi: 10.1093/hmg/ddy371.
Author Address: Department of Physiology, McGill University, Montreal, Quebec, Canada.
Genetics of Learning Disability Service, Hunter Genetics, Waratah, NSW, Australia.
Adelaide Medical School and Robinson Research Institute, The University of Adelaide, Adelaide, SA, Australia.
Research Group Development and Disease, Max Planck Institute for Molecular Genetics, Berlin, Germany.
GeneDx, Gaithersburg, MD, USA.
Genetics and Molecular Pathology, SA Pathology, Adelaide, SA, Australia.
Department of Pediatrics, Division of Medical Genetics, University of Mississippi Medical Center, Jackson, MS, USA.
South Australian Health and Medical Research Institute, Adelaide, SA, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 951
Author: Khidekel, N., Ficarro, S. B., Peters, E. C. and Hsieh-Wilson, L. C.
Year: 2004
Title: Exploring the O-GlcNAc proteome: direct identification of O-GlcNAc-modified proteins from the brain
Journal: Proc Natl Acad Sci U S A
Volume: 101
Issue: 36
Pages: 13132-7
Epub Date: 2004/09/02
Date: Sep 7
Short Title: Exploring the O-GlcNAc proteome: direct identification of O-GlcNAc-modified proteins from the brain
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0403471101
PMCID: PMC516536
Accession Number: 15340146
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
*Brain Chemistry
Cattle
Glycosylation
Molecular Sequence Data
*Protein Processing, Post-Translational
*Proteome
Rats
Rats, Sprague-Dawley
alpha-Crystallin A Chain/chemistry
Abstract: The covalent modification of intracellular proteins by O-linked beta-N-acetylglucosamine (O-GlcNAc) is emerging as a crucial regulatory posttranslational modification akin to phosphorylation. Numerous studies point to the significance of O-GlcNAc in cellular processes such as nutrient sensing, protein degradation, and gene expression. Despite its importance, the breadth and functional roles of O-GlcNAc are only beginning to be elucidated. Advances in our understanding will require the development of new strategies for the detection and study of O-GlcNAc-modified proteins in vivo. Herein we report the direct, high-throughput analysis of O-GlcNAc-glycosylated proteins from the mammalian brain. The proteins were identified by using a chemoenzymatic approach that exploits an engineered galactosyltransferase enzyme to selectively label O-GlcNAc proteins with a ketone-biotin tag. The tag permits enrichment of low-abundance O-GlcNAc species from complex mixtures and localization of the modification to short amino acid sequences. Using this approach, we discovered 25 O-GlcNAc-glycosylated proteins from the brain, including regulatory proteins associated with gene expression, neuronal signaling, and synaptic plasticity. The functional diversity represented by this set of proteins suggests an expanded role for O-GlcNAc in regulating neuronal function. Moreover, the chemoenzymatic strategy described here should prove valuable for identifying O-GlcNAc-modified proteins in various tissues and facilitate studies of the physiological significance of O-GlcNAc across the proteome.
Notes: 1091-6490
Khidekel, Nelly
Ficarro, Scott B
Peters, Eric C
Hsieh-Wilson, Linda C
T32 GM007616/GM/NIGMS NIH HHS/United States
T32GM07616/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2004 Sep 7;101(36):13132-7. doi: 10.1073/pnas.0403471101. Epub 2004 Aug 30.
Author Address: Division of Chemistry and Chemical Engineering, California Institute of Technology, 1200 East California Boulevard, Pasadena, CA 91125, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 748
Author: Kiefer, J. R., Pawlitz, J. L., Moreland, K. T., Stegeman, R. A., Hood, W. F., Gierse, J. K., Stevens, A. M., Goodwin, D. C., Rowlinson, S. W., Marnett, L. J., Stallings, W. C. and Kurumbail, R. G.
Year: 2000
Title: Structural insights into the stereochemistry of the cyclooxygenase reaction
Journal: Nature
Volume: 405
Issue: 6782
Pages: 97-101
Epub Date: 2000/05/16
Date: May 4
Short Title: Structural insights into the stereochemistry of the cyclooxygenase reaction
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/35011103
Accession Number: 10811226
Keywords: Animals
Apoenzymes/chemistry/metabolism
Arachidonic Acid/*chemistry/metabolism
Binding Sites
Crystallography, X-Ray
Cyclooxygenase 2
Isoenzymes/*chemistry/metabolism
Mice
Models, Molecular
Prostaglandin H2
Prostaglandin-Endoperoxide Synthases/*chemistry/metabolism
Prostaglandins H/*chemistry/metabolism
Protein Conformation
Recombinant Proteins/chemistry/metabolism
Stereoisomerism
Abstract: Cyclooxygenases are bifunctional enzymes that catalyse the first committed step in the synthesis of prostaglandins, thromboxanes and other eicosanoids. The two known cyclooxygenases isoforms share a high degree of amino-acid sequence similarity, structural topology and an identical catalytic mechanism. Cyclooxygenase enzymes catalyse two sequential reactions in spatially distinct, but mechanistically coupled active sites. The initial cyclooxygenase reaction converts arachidonic acid (which is achiral) to prostaglandin G2 (which has five chiral centres). The subsequent peroxidase reaction reduces prostaglandin G2 to prostaglandin H2. Here we report the co-crystal structures of murine apo-cyclooxygenase-2 in complex with arachidonic acid and prostaglandin. These structures suggest the molecular basis for the stereospecificity of prostaglandin G2 synthesis.
Notes: Kiefer, J R
Pawlitz, J L
Moreland, K T
Stegeman, R A
Hood, W F
Gierse, J K
Stevens, A M
Goodwin, D C
Rowlinson, S W
Marnett, L J
Stallings, W C
Kurumbail, R G
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2000 May 4;405(6782):97-101. doi: 10.1038/35011103.
Author Address: Searle Discovery Research, Monsanto Company, St Louis, Missouri 63198, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 275
Author: Kiefer, M. C., Masiarz, F. R., Bauer, D. M. and Zapf, J.
Year: 1991
Title: Identification and molecular cloning of two new 30-kDa insulin-like growth factor binding proteins isolated from adult human serum
Journal: J Biol Chem
Volume: 266
Issue: 14
Pages: 9043-9
Epub Date: 1991/05/15
Date: May 15
Short Title: Identification and molecular cloning of two new 30-kDa insulin-like growth factor binding proteins isolated from adult human serum
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1709161
Keywords: Amino Acid Sequence
Base Sequence
Carrier Proteins/*genetics/immunology/metabolism
Cloning, Molecular
DNA/genetics
Humans
Insulin-Like Growth Factor Binding Protein 5
Molecular Sequence Data
Molecular Weight
Oligonucleotides/chemistry
Polymerase Chain Reaction
Somatomedins/*metabolism
Abstract: Three insulin-like growth factor binding proteins (IGFBP) with apparent molecular masses of 24, 28-30, and 30 kDa, nonreduced, have been isolated from human serum. The 15 NH2-terminal amino acids of the 24-kDa binding protein are identical with those of the 30-kDa BP. The apparent molecular mass of the latter is reduced to 24 kDa by N-glycanase, suggesting that the 30-kDa BP is the glycosylated form of the isolated 24-kDa BP. The complete amino acid sequences derived from the cloned cDNAs represent two new IGFBPs. They are tentatively termed IGFBP-4 and -5. The prepeptide sequences of BP-4 and -5 contain 27 and 21, the mature proteins 213 and 237 amino acids, respectively (Mr = 22,610 and 25,980). The NH2- and COOH-terminal thirds of BP-4 and -5 display pronounced homology to the other three human BPs. 16 of the 16-20 cysteines and 37 of the 213-289 amino acids (12.8-17.1%) are conserved in all five mature BPs. 10 amino acid positions located in the NH2-terminal region and shared by BP-1, -2, -3, and -5 are different in BP-4. These differences may account for the preferential affinity of BP-4 for IGF II. A most intriguing homology exists between the COOH-terminal quarter of the five IGFBPs, 10 repetitive domains of human thyroglobulin, a gastrointestinal tumor-associated antigen, and the invariant chain of the class II histocompatibility antigen. The cDNAs of five human IGFBPs are now available. They will allow their expression and production in sufficient quantities for in vivo studies to unravel their role in growth and metabolism.
Notes: Kiefer, M C
Masiarz, F R
Bauer, D M
Zapf, J
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1991 May 15;266(14):9043-9.
Author Address: Chiron Corporation, Emeryville, California 94608.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 379
Author: Killeen, N., Moessner, R., Arvieux, J., Willis, A. and Williams, A. F.
Year: 1988
Title: The MRC OX-45 antigen of rat leukocytes and endothelium is in a subset of the immunoglobulin superfamily with CD2, LFA-3 and carcinoembryonic antigens
Journal: Embo j
Volume: 7
Issue: 10
Pages: 3087-91
Epub Date: 1988/10/01
Date: Oct
Short Title: The MRC OX-45 antigen of rat leukocytes and endothelium is in a subset of the immunoglobulin superfamily with CD2, LFA-3 and carcinoembryonic antigens
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
PMCID: PMC454697
Accession Number: 3181129
Keywords: Animals
Antigens, CD
Antigens, Surface/*genetics/metabolism
Base Sequence
CD48 Antigen
Circular Dichroism
Cloning, Molecular
DNA/genetics
Disulfides
Endothelium, Vascular/immunology
Immunoglobulins/genetics
Leukocytes/*immunology
Membrane Glycoproteins/*genetics/metabolism/ultrastructure
Molecular Sequence Data
*Multigene Family
Phosphatidylinositols/metabolism
Rats
Abstract: The MRC OX-45 cell surface antigen is a glycoprotein of 45,000 apparent mol. wt of rat leukocytes and endothelium. Antibodies against the antigen inhibit T lymphocyte responses by stimulation of suppression by accessory cells. We now report the immunochemical characterization of this antigen and its cDNA sequence. The predicted protein sequence contains 240 amino acids including a leader sequence of 22 residues and a carboxy-terminal sequence of 23 residues that is replaced in the processed molecule by a glycosyl-phosphatidylinositol anchor attached at serine 195. Two Ig-related domains are predicted to account for all of the processed sequence and the circular dichroism spectrum shows pure beta-structure. The amino-terminal domain is V-like, but without a disulphide bond, while the second domain is C-like (C2-SET) with two disulphide bonds. The sequence matches particularly well with the extracellular parts of LFA-3 and CD2 antigens and the first two domains of carcinoembryonic antigen and non-specific, cross-reacting antigen.
Notes: 1460-2075
Killeen, N
Moessner, R
Arvieux, J
Willis, A
Williams, A F
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1988 Oct;7(10):3087-91.
Author Address: MRC Cellular Immunology Unit, Sir William Dunn School of Pathology, Oxford, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1474
Author: Kim, C., Nam, D. W., Park, S. Y., Song, H., Hong, H. S., Boo, J. H., Jung, E. S., Kim, Y., Baek, J. Y., Kim, K. S., Cho, J. W. and Mook-Jung, I.
Year: 2013
Title: O-linked β-N-acetylglucosaminidase inhibitor attenuates β-amyloid plaque and rescues memory impairment
Journal: Neurobiol Aging
Volume: 34
Issue: 1
Pages: 275-85
Epub Date: 2012/04/17
Date: Jan
Short Title: O-linked β-N-acetylglucosaminidase inhibitor attenuates β-amyloid plaque and rescues memory impairment
Alternate Journal: Neurobiology of aging
ISSN: 0197-4580
DOI: 10.1016/j.neurobiolaging.2012.03.001
Accession Number: 22503002
Keywords: Alzheimer Disease/complications/drug therapy/genetics
Amyloid Precursor Protein Secretases/metabolism
Amyloid beta-Peptides/metabolism
Amyloid beta-Protein Precursor/genetics
Analysis of Variance
Animals
Bridged Bicyclo Compounds, Heterocyclic/*therapeutic use
Conditioning, Psychological/drug effects
Disease Models, Animal
Dose-Response Relationship, Drug
Enzyme Inhibitors/*therapeutic use
Enzyme-Linked Immunosorbent Assay
Fear/drug effects
Gene Expression Regulation, Enzymologic/drug effects
HEK293 Cells
Humans
Maze Learning/drug effects
Memory Disorders/*drug therapy/*enzymology/etiology
Mice
Mice, Transgenic
Mutation/genetics
Peptide Fragments/metabolism
Plaque, Amyloid/drug therapy
Presenilin-1/genetics
Presenilin-2/genetics
Transfection
beta-N-Acetylhexosaminidases/*metabolism
Abstract: Deposition of β-amyloid (Aβ) as senile plaques and disrupted glucose metabolism are two main characteristics of Alzheimer's disease (AD). It is unknown, however, how these two processes are related in AD. Here we examined the relationship between O-GlcNAcylation, which is a glucose level-dependent post-translational modification that adds O-linked β-N-acetylglucosamine (O-GlcNAc) to proteins, and Aβ production in a mouse model of AD carrying 5XFAD genes. We found that 1,2-dideoxy-2'-propyl-α-d-glucopyranoso-[2,1-D]-Δ2'-thiazoline (NButGT), a specific inhibitor of O-GlcNAcase, reduces Aβ production by lowering γ-secretase activity both in vitro and in vivo. We also found that O-GlcNAcylation takes place at the S708 residue of nicastrin, which is a component of γ-secretase. Moreover, NButGT attenuated the accumulation of Aβ, neuroinflammation, and memory impairment in the 5XFAD mice. This is the first study to show the relationship between Aβ generation and O-GlcNAcylation in vivo. These results suggest that O-GlcNAcylation may be a suitable therapeutic target for the treatment of AD.
Notes: 1558-1497
Kim, Chaeyoung
Nam, Dong Woo
Park, Sang Yoon
Song, Hyundong
Hong, Hyun Seok
Boo, Jung Hyun
Jung, Eun Sun
Kim, Yoonhee
Baek, Ju Yuel
Kim, Kwan Soo
Cho, Jin Won
Mook-Jung, Inhee
Journal Article
Research Support, Non-U.S. Gov't
United States
Neurobiol Aging. 2013 Jan;34(1):275-85. doi: 10.1016/j.neurobiolaging.2012.03.001. Epub 2012 Apr 11.
Author Address: Department of Biochemistry and Biomedical Sciences, Seoul National University College of Medicine, Seoul, 110-799, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1903
Author: Kim, D. I., Cutler, J. A., Na, C. H., Reckel, S., Renuse, S., Madugundu, A. K., Tahir, R., Goldschmidt, H. L., Reddy, K. L., Huganir, R. L., Wu, X., Zachara, N. E., Hantschel, O. and Pandey, A.
Year: 2018
Title: BioSITe: A Method for Direct Detection and Quantitation of Site-Specific Biotinylation
Journal: J Proteome Res
Volume: 17
Issue: 2
Pages: 759-769
Epub Date: 2017/12/19
Date: Feb 2
Short Title: BioSITe: A Method for Direct Detection and Quantitation of Site-Specific Biotinylation
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/acs.jproteome.7b00775
PMCID: PMC6092923
NIHMSID: NIHMS933244
Accession Number: 29249144
Keywords: Acetylglucosamine/chemistry/*metabolism
Amino Acid Sequence
Animals
Antibodies, Immobilized/chemistry
B-Lymphocytes/chemistry
Biotin/*chemistry
Biotinylation
Cell Line
Chromatography, Liquid
Click Chemistry/*methods
HEK293 Cells
Humans
Mice
Peptides/chemistry/*isolation & purification
*Protein Processing, Post-Translational
Proteolysis
Streptavidin/*chemistry
Tandem Mass Spectrometry
*Apex
*BioID
*biotinylation
*peptide
*protein−protein interactions
*proximity-dependent biotinylation
*subcellular proteome
Abstract: Biotin-based labeling strategies are widely employed to study protein-protein interactions, subcellular proteomes and post-translational modifications, as well as, used in drug discovery. While the high affinity of streptavidin for biotin greatly facilitates the capture of biotinylated proteins, it still presents a challenge, as currently employed, for the recovery of biotinylated peptides. Here we describe a strategy designated Biotinylation Site Identification Technology (BioSITe) for the capture of biotinylated peptides for LC-MS/MS analyses. We demonstrate the utility of BioSITe when applied to proximity-dependent labeling methods, APEX and BioID, as well as biotin-based click chemistry strategies for identifying O-GlcNAc-modified sites. We demonstrate the use of isotopically labeled biotin for quantitative BioSITe experiments that simplify differential interactome analysis and obviate the need for metabolic labeling strategies such as SILAC. Our data also highlight the potential value of site-specific biotinylation in providing spatial and topological information about proteins and protein complexes. Overall, we anticipate that BioSITe will replace the conventional methods in studies where detection of biotinylation sites is important.
Notes: 1535-3907
Kim, Dae In
Cutler, Jevon A
Na, Chan Hyun
Reckel, Sina
Renuse, Santosh
Madugundu, Anil K
Tahir, Raiha
Goldschmidt, Hana L
Reddy, Karen L
Huganir, Richard L
Wu, Xinyan
Zachara, Natasha E
Hantschel, Oliver
Pandey, Akhilesh
Orcid: 0000-0001-9943-6127
F32 NS098946/NS/NINDS NIH HHS/United States
U24 CA210985/CA/NCI NIH HHS/United States
S10 OD021844/OD/NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Proteome Res. 2018 Feb 2;17(2):759-769. doi: 10.1021/acs.jproteome.7b00775. Epub 2017 Dec 28.
Author Address: McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Pre-Doctoral Training Program in Human Genetics, McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Department of Biological Chemistry, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Center for Proteomics Discovery, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Swiss Institute for Experimental Cancer Research (ISREC), School of Life Sciences, École polytechnique fédérale de Lausanne (EPFL) , CH-1015 Lausanne, Switzerland.
Institute of Bioinformatics , International Technology Park, Bangalore 560 066, India.
Biochemistry, Cellular and Molecular Biology Graduate Program, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Solomon H. Snyder Department of Neuroscience, Kavli Neuroscience Discovery Institute, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Center for Epigenetics, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Departments of Pathology and Oncology, Johns Hopkins University School of Medicine , Baltimore, Maryland 21205, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1565
Author: Kim, E. J. and Hanover, J. A.
Year: 2013
Title: Enzymatic characterization of recombinant enzymes of O-GlcNAc cycling
Journal: Methods Mol Biol
Volume: 1022
Pages: 129-45
Epub Date: 2013/06/15
Short Title: Enzymatic characterization of recombinant enzymes of O-GlcNAc cycling
Alternate Journal: Methods in molecular biology (Clifton, N.J.)
ISSN: 1064-3745
DOI: 10.1007/978-1-62703-465-4_11
Accession Number: 23765659
Keywords: Animals
Cloning, Molecular/methods
Enzyme Assays/*methods
Escherichia coli/genetics
Humans
N-Acetylglucosaminyltransferases/genetics/isolation & purification/*metabolism
Recombinant Proteins/genetics/isolation & purification/metabolism
Abstract: The dynamic addition of O-GlcNAc to target proteins is now recognized as a major signaling paradigm impacting phosphorylation, protein turnover, gene expression, and other posttranslational modifications influencing epigenetics. Here we describe the production of and methods for assay of the recombinant enzymes of O-GlcNAc cycling: O-linked GlcNAc Transferase (OGT) and O-GlcNAcase (OGA).
Notes: 1940-6029
Kim, Eun Ju
Hanover, John A
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
United States
Methods Mol Biol. 2013;1022:129-45. doi: 10.1007/978-1-62703-465-4_11.
Author Address: Laboratory Cell Biochemistry and Biology, NIDDK, National Institutes of Health, Bethesda, MD, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 505
Author: Kim, H., Yamaguchi, Y., Masuda, K., Matsunaga, C., Yamamoto, K., Irimura, T., Takahashi, N., Kato, K. and Arata, Y.
Year: 1994
Title: O-glycosylation in hinge region of mouse immunoglobulin G2b
Journal: J Biol Chem
Volume: 269
Issue: 16
Pages: 12345-50
Epub Date: 1994/04/22
Date: Apr 22
Short Title: O-glycosylation in hinge region of mouse immunoglobulin G2b
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7512967
Keywords: Amino Acid Sequence
Animals
Antibodies, Monoclonal/*chemistry/classification/isolation & purification
Cell Line
Chromatography, Affinity
Electrophoresis, Polyacrylamide Gel
Glycosylation
Hybridomas/immunology
Immunoglobulin Fragments/chemistry/isolation & purification
Immunoglobulin G/*chemistry/classification/isolation & purification
Magnetic Resonance Spectroscopy
Mice
Molecular Sequence Data
Neuraminidase
Oligosaccharides/analysis/chemistry
Papain
Peptide Fragments/chemistry/isolation & purification
Peptide Mapping
Phytohemagglutinins
*Protein Conformation
Spectrometry, Mass, Fast Atom Bombardment
alpha-Fetoproteins
Abstract: Mouse monoclonal immunoglobulin G2b (IgG2b) antibodies are known to contain two forms of the heavy chain that are different in susceptibility to the protease attack. In the present study, by use of an affinity column containing sialic acid-binding lectins from Maackia amurensis seeds, a mouse monoclonal IgG2b was successfully separated into three phenotypes, which are different in the degree of sialylation in the heavy chain. In the N-linked oligosaccharides from all of the IgG2b phenotypes, virtually no sialylation was detected. Elution profiles of the lysyl endopeptidase digestion products were compared for the three phenotypes. The peptides eluted at different retention times were subjected to fast atom bombardment-mass spectrometry and amino acid sequence analyses. It was revealed that approximately 40% of the heavy chain of the mouse IgG2b are O-glycosylated at Thr-221A in the hinge region, predominantly with a tetrasaccharide composed of GalNAc, Gal, and two N-glycolylneuraminic acid residues. We suggest that the O-glycosylation renders the hinge region resistant against the proteolyses of the heavy chain. A therapeutic significance of the O-glycosylation of IgG2b is briefly discussed.
Notes: Kim, H
Yamaguchi, Y
Masuda, K
Matsunaga, C
Yamamoto, K
Irimura, T
Takahashi, N
Kato, K
Arata, Y
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1994 Apr 22;269(16):12345-50.
Author Address: Faculty of Pharmaceutical Sciences, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1130
Author: Kim, H. M., Park, B. S., Kim, J. I., Kim, S. E., Lee, J., Oh, S. C., Enkhbayar, P., Matsushima, N., Lee, H., Yoo, O. J. and Lee, J. O.
Year: 2007
Title: Crystal structure of the TLR4-MD-2 complex with bound endotoxin antagonist Eritoran
Journal: Cell
Volume: 130
Issue: 5
Pages: 906-17
Epub Date: 2007/09/07
Date: Sep 7
Short Title: Crystal structure of the TLR4-MD-2 complex with bound endotoxin antagonist Eritoran
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2007.08.002
Accession Number: 17803912
Keywords: Amino Acid Sequence
Animals
Binding Sites
Cloning, Molecular
Crystallography, X-Ray
Dimerization
Disaccharides/*chemistry/metabolism/pharmacology
Fish Proteins/chemistry
Hagfishes
Humans
Hydrophobic and Hydrophilic Interactions
Immunoglobulin Variable Region/chemistry
Lipopolysaccharides/antagonists & inhibitors/*chemistry/metabolism
Lymphocyte Antigen 96/*chemistry/genetics/metabolism
Mice
Models, Molecular
Molecular Sequence Data
Molecular Structure
Mutation
Protein Binding
Protein Conformation
Protein Engineering
Protein Structure, Tertiary
Recombinant Fusion Proteins/chemistry
Structural Homology, Protein
Sugar Phosphates/*chemistry/metabolism/pharmacology
Toll-Like Receptor 4/antagonists & inhibitors/*chemistry/genetics/metabolism
Abstract: TLR4 and MD-2 form a heterodimer that recognizes LPS (lipopolysaccharide) from Gram-negative bacteria. Eritoran is an analog of LPS that antagonizes its activity by binding to the TLR4-MD-2 complex. We determined the structure of the full-length ectodomain of the mouse TLR4 and MD-2 complex. We also produced a series of hybrids of human TLR4 and hagfish VLR and determined their structures with and without bound MD-2 and Eritoran. TLR4 is an atypical member of the LRR family and is composed of N-terminal, central, and C-terminal domains. The beta sheet of the central domain shows unusually small radii and large twist angles. MD-2 binds to the concave surface of the N-terminal and central domains. The interaction with Eritoran is mediated by a hydrophobic internal pocket in MD-2. Based on structural analysis and mutagenesis experiments on MD-2 and TLR4, we propose a model of TLR4-MD-2 dimerization induced by LPS.
Notes: Kim, Ho Min
Park, Beom Seok
Kim, Jung-In
Kim, Sung Eun
Lee, Judong
Oh, Se Cheol
Enkhbayar, Purevjav
Matsushima, Norio
Lee, Hayyoung
Yoo, Ook Joon
Lee, Jie-Oh
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell. 2007 Sep 7;130(5):906-17. doi: 10.1016/j.cell.2007.08.002.
Author Address: Department of Chemistry, Korea Advanced Institute of Science and Technology, Daejon, Korea 305-701.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1914
Author: Kim, H. T., Na, B. K., Chung, J., Kim, S., Kwon, S. K., Cha, H., Son, J., Cho, J. M. and Hwang, K. Y.
Year: 2018
Title: Structural Basis for Inhibitor-Induced Hydrogen Peroxide Production by Kynurenine 3-Monooxygenase
Journal: Cell Chem Biol
Volume: 25
Issue: 4
Pages: 426-438.e4
Epub Date: 2018/02/13
Date: Apr 19
Short Title: Structural Basis for Inhibitor-Induced Hydrogen Peroxide Production by Kynurenine 3-Monooxygenase
Alternate Journal: Cell chemical biology
ISSN: 2451-9448
DOI: 10.1016/j.chembiol.2018.01.008
Accession Number: 29429898
Keywords: Amino Acid Sequence
Animals
Enzyme Inhibitors/*pharmacology
Flavins/metabolism
Humans
Hydrogen Peroxide/*metabolism
Kynurenine 3-Monooxygenase/*antagonists & inhibitors/chemistry/*metabolism
Molecular Docking Simulation
Oxidation-Reduction/drug effects
Protein Conformation/drug effects
Pseudomonas fluorescens/chemistry/*enzymology
Saccharomyces cerevisiae/chemistry/*enzymology
Sequence Alignment
Sulfonamides/*pharmacology
Thiazoles/*pharmacology
*KMO inhibitor
*drug design
*flavin reduction
*hydrogen peroxide
*kynurenine 3-monooxygenase
Abstract: Kynurenine 3-monooxygenase (KMO) inhibitors have been developed for the treatment of neurodegenerative disorders. The mechanisms of flavin reduction and hydrogen peroxide production by KMO inhibitors are unknown. Herein, we report the structure of human KMO and crystal structures of Saccharomyces cerevisiae (sc) and Pseudomonas fluorescens (pf) KMO with Ro 61-8048. Proton transfer in the hydrogen bond network triggers flavin reduction in p-hydroxybenzoate hydroxylase, but the mechanism triggering flavin reduction in KMO is different. Conformational changes via π-π interactions between the loop above the flavin and substrate or non-substrate effectors lead to disorder of the C-terminal α helix in scKMO and shifts of domain III in pfKMO, stimulating flavin reduction. Interestingly, Ro 61-8048 has two different binding modes. It acts as a competitive inhibitor in scKMO and as a non-substrate effector in pfKMO. These findings provide understanding of the catalytic cycle of KMO and insight for structure-based drug design of KMO inhibitors.
Notes: 2451-9448
Kim, Hyun Tae
Na, Byeong Kwan
Chung, Jiwoung
Kim, Sulhee
Kwon, Sool Ki
Cha, Hyunju
Son, Jonghyeon
Cho, Joong Myung
Hwang, Kwang Yeon
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell Chem Biol. 2018 Apr 19;25(4):426-438.e4. doi: 10.1016/j.chembiol.2018.01.008. Epub 2018 Feb 8.
Author Address: Crystalgenomics, Inc., 5F, Tower A, Korea Bio Park 700, Daewangpangyo-ro, Bundang-gu, Seongnam-si, Gyeonggi-do 13524, Korea; Division of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul 02841, Korea.
Crystalgenomics, Inc., 5F, Tower A, Korea Bio Park 700, Daewangpangyo-ro, Bundang-gu, Seongnam-si, Gyeonggi-do 13524, Korea.
Division of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul 02841, Korea.
Crystalgenomics, Inc., 5F, Tower A, Korea Bio Park 700, Daewangpangyo-ro, Bundang-gu, Seongnam-si, Gyeonggi-do 13524, Korea. Electronic address: jmcho@cgxinc.com.
Division of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul 02841, Korea. Electronic address: chahong@korea.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 968
Author: Kim, J. I., Lee, C. J., Jin, M. S., Lee, C. H., Paik, S. G., Lee, H. and Lee, J. O.
Year: 2005
Title: Crystal structure of CD14 and its implications for lipopolysaccharide signaling
Journal: J Biol Chem
Volume: 280
Issue: 12
Pages: 11347-51
Epub Date: 2005/01/13
Date: Mar 25
Short Title: Crystal structure of CD14 and its implications for lipopolysaccharide signaling
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M414607200
Accession Number: 15644310
Keywords: Amino Acid Sequence
Animals
Antigens, Ly/physiology
Hydrophobic and Hydrophilic Interactions
Lipopolysaccharide Receptors/*chemistry/metabolism
Lipopolysaccharides/*metabolism
Lymphocyte Antigen 96
Mice
Molecular Sequence Data
Protein Structure, Secondary
Receptors, Cell Surface/physiology
Signal Transduction
Toll-Like Receptor 4
Abstract: Lipopolysaccharide, the endotoxin of Gram-negative bacteria, induces extensive immune responses that can lead to fatal septic shock syndrome. The core receptors recognizing lipopolysaccharide are CD14, TLR4, and MD-2. CD14 binds to lipopolysaccharide and presents it to the TLR4/MD-2 complex, which initiates intracellular signaling. In addition to lipopolysaccharide, CD14 is capable of recognizing a few other microbial and cellular products. Here, we present the first crystal structure of CD14 to 2.5 angstroms resolution. A large hydrophobic pocket was found on the NH2-terminal side of the horseshoe-like structure. Previously identified regions involved in lipopolysaccharide binding map to the rim and bottom of the pocket indicating that the pocket is the main component of the lipopolysaccharide-binding site. Mutations that interfere with lipopolysaccharide signaling but not with lipopolysaccharide binding are also clustered in a separate area near the pocket. Ligand diversity of CD14 could be explained by the generous size of the pocket, the considerable flexibility of the rim of the pocket, and the multiplicity of grooves available for ligand binding.
Notes: Kim, Jung-In
Lee, Chang Jun
Jin, Mi Sun
Lee, Cherl-Ho
Paik, Sang-Gi
Lee, Hayyoung
Lee, Jie-Oh
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2005 Mar 25;280(12):11347-51. doi: 10.1074/jbc.M414607200. Epub 2005 Jan 10.
Author Address: Department of Chemistry, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1915
Author: Kim, M. J., Choi, M. Y., Lee, D. H., Roh, G. S., Kim, H. J., Kang, S. S., Cho, G. J., Kim, Y. S. and Choi, W. S.
Year: 2018
Title: O-linked N-acetylglucosamine transferase enhances secretory clusterin expression via liver X receptors and sterol response element binding protein regulation in cervical cancer
Journal: Oncotarget
Volume: 9
Issue: 4
Pages: 4625-4636
Epub Date: 2018/02/13
Date: Jan 12
Short Title: O-linked N-acetylglucosamine transferase enhances secretory clusterin expression via liver X receptors and sterol response element binding protein regulation in cervical cancer
Alternate Journal: Oncotarget
ISSN: 1949-2553
DOI: 10.18632/oncotarget.23588
PMCID: PMC5797001
Accession Number: 29435130
Keywords: LXRs
O-GlcNAcylation
Ogt
Srebp-1
cisplatin
Abstract: O-linked N-acetylglucosamine transferase (OGT) expression is increased in various cancer types, indicating the potential importance of O-GlcNAcylation in tumorigenesis. Secretory clusterin (sCLU) is involved in cancer cell proliferation and drug resistance, and recently, liver X receptors (LXRs) and sterol response element binding protein-1 (SREBP-1) were reported to regulate sCLU transcription. Here, we found that sCLU is significantly increased in cervical cancer cell lines, which have higher expression levels of O-GlcNAc and OGT than keratinocytes. OGT knockdown decreased expression of LXRs, SREBP-1 and sCLU through hypo-O-GlcNAcylation of LXRs. Additionally, treatment with Thiamet G, O-GlcNAcase OGA inhibitor, increased expression of O-GlcNAcylation and sCLU, and high glucose increased levels of LXRs, SREBP-1 and sCLU in HeLa cells. Moreover, OGT knockdown induced G(0)/G(1) phase cell cycle arrest and late apoptosis in cisplatin-treated HeLa cells, and decreased viability compared to OGT intact HeLa cells. Taken together, these findings suggest that OGT, O-GlcNAcylated LXRs, and SREBP-1 increase sCLU expression in cervical cancer cells, which contributes to drug resistance.
Notes: 1949-2553
Kim, Min Jun
Choi, Mee Young
Lee, Dong Hoon
Roh, Gu Seob
Kim, Hyun Joon
Kang, Sang Soo
Cho, Gyeong Jae
Kim, Yoon Sook
Choi, Wan Sung
Journal Article
Oncotarget. 2017 Dec 21;9(4):4625-4636. doi: 10.18632/oncotarget.23588. eCollection 2018 Jan 12.
Author Address: Department of Anatomy and Convergence Medical Science, Institute of Health Sciences, College of Medicine, Gyeongsang National University, Jinju, Gyeongnam, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1196
Author: Kim, M. L., Chandrasekharan, K., Glass, M., Shi, S., Stahl, M. C., Kaspar, B., Stanley, P. and Martin, P. T.
Year: 2008
Title: O-fucosylation of muscle agrin determines its ability to cluster acetylcholine receptors
Journal: Mol Cell Neurosci
Volume: 39
Issue: 3
Pages: 452-64
Epub Date: 2008/09/09
Date: Nov
Short Title: O-fucosylation of muscle agrin determines its ability to cluster acetylcholine receptors
Alternate Journal: Molecular and cellular neurosciences
ISSN: 1044-7431 (Print)
1044-7431
DOI: 10.1016/j.mcn.2008.07.026
PMCID: PMC2646263
NIHMSID: NIHMS77248
Accession Number: 18775496
Keywords: Agrin/genetics/*metabolism
Animals
CHO Cells
Cricetinae
Cricetulus
Fucose/chemistry/*metabolism
Fucosyltransferases/genetics/*metabolism
Humans
Mice
Muscle, Skeletal/cytology/*physiology
RNA, Small Interfering/genetics/metabolism
Receptor Aggregation/*physiology
Receptor Protein-Tyrosine Kinases/genetics/metabolism
Receptors, Cholinergic/*metabolism
Receptors, Notch/metabolism
Recombinant Proteins/genetics/metabolism
Signal Transduction/physiology
Synapses/physiology
Abstract: Protein O-fucosyltransferase 1 (Pofut1) transfers fucose to serine or threonine on proteins, including Notch receptors, that contain EGF repeats with a particular consensus sequence. Here we demonstrate that agrin is O-fucosylated in a Pofut1-dependent manner, and that this glycosylation can regulate agrin function. Fucosylation of recombinant C45 agrin, both active (neural, z8) and inactive (muscle, z0) splice forms, was eliminated when agrin was overexpressed in Pofut1-deficient cells or by mutation of a consensus site for Pofut1 fucosylation (serine 1726 in the EGF4 domain). Loss of O-fucosylation caused a gain of function for muscle agrin such that it stimulated AChR clustering and MuSK phosphorylation in cultured myotubes at levels normally only found with the neural splice form. Deletion of Pofut1 in cultured primary myotubes and in adult skeletal muscle increased AChR aggregation. In addition, Pofut1 gene and protein expression and Pofut1 activity of the EGF4 domain of agrin were modulated during neuromuscular development. These data are consistent with a role for Pofut1 in AChR aggregation during synaptogenesis via the regulation of the synaptogenic activity of muscle agrin.
Notes: 1095-9327
Kim, Mi-Lyang
Chandrasekharan, Kumaran
Glass, Matthew
Shi, Shaolin
Stahl, Mark C
Kaspar, Brian
Stanley, Pamela
Martin, Paul T
R01 AR050202/AR/NIAMS NIH HHS/United States
R01 95022/PHS HHS/United States
R01 AR050202-05/AR/NIAMS NIH HHS/United States
R01 AR047922/AR/NIAMS NIH HHS/United States
R01 AR049722-03/AR/NIAMS NIH HHS/United States
R01 AR049722/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Mol Cell Neurosci. 2008 Nov;39(3):452-64. doi: 10.1016/j.mcn.2008.07.026. Epub 2008 Aug 15.
Author Address: Center for Gene Therapy, The Research Institute at Nationwide Children's Hospital, Columbus, OH 43205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2036
Author: Kim, M. Y., Kim, Y. S., Kim, M., Choi, M. Y., Roh, G. S., Lee, D. H., Kim, H. J., Kang, S. S., Cho, G. J., Shin, J. K. and Choi, W. S.
Year: 2019
Title: Metformin inhibits cervical cancer cell proliferation via decreased AMPK O-GlcNAcylation
Journal: Anim Cells Syst (Seoul)
Volume: 23
Issue: 4
Pages: 302-309
Epub Date: 2019/09/07
Short Title: Metformin inhibits cervical cancer cell proliferation via decreased AMPK O-GlcNAcylation
Alternate Journal: Animal cells and systems
ISSN: 1976-8354 (Print)
1976-8354
DOI: 10.1080/19768354.2019.1614092
PMCID: PMC6711131
Accession Number: 31489252
Keywords: Ampk
O-GlcNAcylation
cervical cancer cells
p21
p27
Abstract: Metformin is a widely used drug for the treatment of type 2 diabetes. Antidiabetic drugs are also known to influence cancer progression, as high glucose levels affect both cancer and diabetes. Metformin induces cell cycle arrest in cancer cells, but the underlying mechanism remains unclear in cervical cancer system. Here, we examined how metformin affects cell cycle arrest and apoptosis in cervical cancer cells. Western blot analysis showed that levels of O-linked N-acetylglucosamine (O-GlcNAc) and O-GlcNAc transferase (OGT) were increased in cervical cancer cells; these effects were reversed by metformin treatment. Immunoprecipitation analysis was used to examine the interplay between O-GlcNAcylation and phosphorylation in HeLa cells, revealing that metformin decreased O-GlcNAcylated AMP-activated protein kinase (AMPK) and increased levels of phospho-AMPK compared to untreated cells. These results were associated with decreased cell cycle arrest and apoptotic cell death in HeLa cells, as shown by flow cytometry. Moreover, 6-diazo-5-oxo-L-norleucine (a glutamine fructose-6-phosphate aminotransferase inhibitor) or thiamet G (an O-GlcNAcase inhibitor) decreased or increased levels of O-GlcNAcylated AMPK, and increased or decreased levels of phosphorylated AMPK, respectively, suggesting that O-GlcNAc modification affects AMPK activation. Of note, we found that metformin treatment of HeLa cells increased the levels of p21 and p27 (which are AMPK-dependent cell cycle inhibitors), leading to increased cell cycle arrest and apoptosis in HeLa cells compared to untreated cells. These findings suggest that metformin may serve as a useful antiproliferative drug in cervical cancer cells, with potential therapeutic benefit.
Notes: 2151-2485
Kim, Min Young
Kim, Yoon Sook
Kim, Minjun
Choi, Mee Young
Roh, Gu Seob
Lee, Dong Hoon
Kim, Hyun Joon
Kang, Sang Soo
Cho, Gyeong Jae
Shin, Jeong Kyu
Choi, Wan Sung
Journal Article
Anim Cells Syst (Seoul). 2019 May 14;23(4):302-309. doi: 10.1080/19768354.2019.1614092. eCollection 2019.
Author Address: Gyeongsang National University, Jinju, The Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1518
Author: Kim, S. Y., Kang, H. T., Han, J. A. and Park, S. C.
Year: 2012
Title: The transcription factor Sp1 is responsible for aging-dependent altered nucleocytoplasmic trafficking
Journal: Aging Cell
Volume: 11
Issue: 6
Pages: 1102-9
Epub Date: 2012/09/28
Date: Dec
Short Title: The transcription factor Sp1 is responsible for aging-dependent altered nucleocytoplasmic trafficking
Alternate Journal: Aging cell
ISSN: 1474-9718
DOI: 10.1111/acel.12012
Accession Number: 23013401
Keywords: Animals
Cell Nucleus/*drug effects/metabolism
Cells, Cultured
Cellular Senescence/drug effects/*genetics
Child
Cytosol/*drug effects/metabolism
Diploidy
Epidermal Growth Factor/pharmacology
Fibroblasts/cytology/*drug effects/metabolism
*Gene Expression Regulation/drug effects
Gene Knockdown Techniques
Humans
Hydrogen Peroxide/pharmacology
Male
Mice
Mice, Inbred C57BL
Mitogen-Activated Protein Kinase 1/genetics/metabolism
Mitogen-Activated Protein Kinase 3/genetics/metabolism
Protein Transport/drug effects
Reactive Oxygen Species/metabolism
Signal Transduction/drug effects
Sp1 Transcription Factor/*genetics/metabolism
Abstract: Hyporesponsiveness to external signals, such as growth factors and apoptotic stimuli, is a cardinal feature of cellular senescence. We previously reported that an aging-dependent marked reduction in nucleocytoplasmic trafficking (NCT)-related genes could be responsible for this phenomenon. In searching for the mechanism, we identified the transcription factor, Sp1, as a common regulator of NCT genes, including various nucleoporins, importins, exportins, and Ran GTPase cycle-related genes. Sp1 knockdown led to a reduction of those genes in young human diploid fibroblast cells (HDF); Sp1 overexpression induced those genes in senescent cells. In addition, epidermal growth factor stimulation-induced p-ERK1/2 nuclear translocation and Elk-1 phosphorylation were severely impaired by Sp1 depletion in young HDFs; Sp1 overexpression restored the nuclear translocation of p-ERK1/2 in senescent HDFs. Furthermore, we observed that Sp1 protein levels were decreased in senescent cells, and H(2) O(2) treatment decreased Sp1 levels in a proteasome-dependent manner. In addition, O-GlcNAcylation of Sp1 was decreased in senescent cells as well as in H(2) O(2) -treated cells. Taken together, these results suggest that Sp1 could be a key regulator in the control of NCT genes and that reactive oxygen species-mediated alteration in Sp1 stability may be responsible for the generalized repression of those genes, leading to formation of the senescence-dependent functional nuclear barrier, resulting in subsequent hyporesponsiveness to external signals.
Notes: 1474-9726
Kim, Sung Y
Kang, Hyun T
Han, Jeong A
Park, Sang C
Journal Article
Research Support, Non-U.S. Gov't
England
Aging Cell. 2012 Dec;11(6):1102-9. doi: 10.1111/acel.12012. Epub 2012 Oct 19.
Author Address: Department of Biochemistry, Gachon University of Medicine and Science, Incheon, 406-799, South Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 340
Author: King, I. A. and Hounsell, E. F.
Year: 1989
Title: Cytokeratin 13 contains O-glycosidically linked N-acetylglucosamine residues
Journal: J Biol Chem
Volume: 264
Issue: 24
Pages: 14022-8
Epub Date: 1989/08/25
Date: Aug 25
Short Title: Cytokeratin 13 contains O-glycosidically linked N-acetylglucosamine residues
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2474541
Keywords: Acetylglucosamine/*isolation & purification/metabolism
Animals
Carbohydrate Conformation
Carcinoma, Squamous Cell/analysis
Cell Line
Cells, Cultured
Glucosamine/*analogs & derivatives
Glycosides/*metabolism
Humans
KB Cells/analysis
Keratins/*isolation & purification/metabolism
Mice
Skin
Abstract: The glycosylation of human cytokeratins was investigated in cultured human keratinocytes and A431 cells by metabolic labeling with [3H]glucosamine. In the presence of tunicamycin, keratinocytes incorporated [3H]glucosamine into a vitamin A-regulated acidic 53-kDa component of the cytoskeleton which was identified as cytokeratin 13 by one- and two-dimensional immunoblotting with specific monoclonal antibodies. This cytoskeletal component was also labeled with [3H]glucosamine in A431 cells but not in KB cells, which do not express cytokeratin 13. Its labeling was resistant to tunicamycin, suggesting that [3H]glucosamine had not been incorporated into N-linked oligosaccharides. Acid hydrolysis followed by paper and ion-exchange chromatography showed that the radioactivity in electrophoretically purified cytokeratin 13 was still present as glucosamine. Radioactivity was completely removed by treatment with beta-N-acetylglucosaminidase, suggesting that it was present in terminal N-acetylglucosamine residues. The labeled carbohydrate was released by alkaline borohydride treatment and was bound by a phenylboronic acid column, indicating an O-glycosidic linkage. On Bio-Gel P-2 columns, the beta-eliminated carbohydrate co-eluted with authentic N-acetylglucosaminitol. The results indicate that cytokeratin 13 contains single residues of N-acetylglucosamine O-glycosidically linked to the polypeptide chain.
Notes: King, I A
Hounsell, E F
Journal Article
United States
J Biol Chem. 1989 Aug 25;264(24):14022-8.
Author Address: Dermatology Research Group, Medical Research Council Clinical Research Center, Harrow, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 868
Author: Kinjo, T. G., Szerencsei, R. T., Winkfein, R. J., Kang, K. and Schnetkamp, P. P.
Year: 2003
Title: Topology of the retinal cone NCKX2 Na/Ca-K exchanger
Journal: Biochemistry
Volume: 42
Issue: 8
Pages: 2485-91
Epub Date: 2003/02/26
Date: Mar 4
Short Title: Topology of the retinal cone NCKX2 Na/Ca-K exchanger
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi0270788
Accession Number: 12600216
Keywords: Amino Acid Sequence
Amino Acid Substitution/genetics
Animals
Cell Line
Cell Membrane/chemistry/genetics/metabolism
Cysteine/genetics
Extracellular Space/chemistry/genetics/metabolism
Glycosylation
Humans
Models, Molecular
Molecular Sequence Data
Moths
Mutagenesis, Site-Directed
Protein Structure, Secondary/genetics
Retinal Cone Photoreceptor Cells/chemistry
Rod Cell Outer Segment/*chemistry/metabolism
Sodium-Calcium Exchanger/*chemistry/genetics/metabolism
Transfection
Abstract: The Na/Ca-K exchanger (NCKX) is a polytopic membrane protein that plays a critical role in Ca(2+) homeostasis in retinal rod and cone photoreceptors. The NCKX1 isoform is found in rods, while the NCKX2 isoform is found in cones, in retinal ganglion cells, and in various parts of the brain. The topology of the Na/Ca-K exchanger is thought to consist of two large hydrophilic loops and two sets of transmembrane spanning segments (TMs). The first large hydrophilic loop is located extracellularly at the N-terminus; the other is cytoplasmic and separates the two sets of TMs. The TMs consist of either five and five membrane spanning helices or five and six membrane spanning helices, depending upon the predictive algorithm used. Little specific information is yet available on the orientation of the various membrane spanning helices and the localization of the short loops connecting these helices. In this study, we have determined which of the connecting loops are exposed to the extracellular milieu using two different methods: accessibility of substituted cysteine residues and insertion of N-glycosylation sites. The two methods resulted in a consistent NCKX topology in which the two sets of TMs each contain five membrane spanning helices. Our new model places what was previously membrane spanning helix six in the cytoplasm, which places the C-terminus on the extracellular surface. Surprisingly, this NCKX topology model is different from the current NCX topology model with respect to the C-terminal three membrane helices.
Notes: Kinjo, Tashi G
Szerencsei, Robert T
Winkfein, Robert J
Kang, KyeongJin
Schnetkamp, Paul P M
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2003 Mar 4;42(8):2485-91. doi: 10.1021/bi0270788.
Author Address: Department of Physiology & Biophysics, Faculty of Medicine, University of Calgary, 3330 Hospital Drive, N.W. Calgary, Alberta, Canada T2N 4N1.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 257
Author: Kirszbaum, L., Bozas, S. E. and Walker, I. D.
Year: 1992
Title: SP-40,40, a protein involved in the control of the complement pathway, possesses a unique array of disulphide bridges
Journal: FEBS Lett
Volume: 297
Issue: 1-2
Pages: 70-6
Epub Date: 1992/02/03
Date: Feb 3
Short Title: SP-40,40, a protein involved in the control of the complement pathway, possesses a unique array of disulphide bridges
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(92)80330-j
Accession Number: 1551440
Keywords: Amino Acid Sequence
Blood Proteins/*metabolism
Chromatography, High Pressure Liquid
Clusterin
Complement System Proteins/*metabolism
Cyanogen Bromide/chemistry
Disulfides/*metabolism
Electrophoresis, Polyacrylamide Gel
*Glycoproteins
*Molecular Chaperones
Molecular Sequence Data
Trypsin/chemistry
Abstract: SP-40,40 is a two-chain serum protein which acts in vitro as a potent inhibitor of the assembly of the membrane attack complex of human complement. It contains 10 cysteine residues, the numbers and locations of which are conserved in several mammalian species. Evidence is presented that all the cysteine residues are involved in interchain (alpha-beta) disulphide bonds. There are no free cysteine residues. The disulphide bond motif established in this study for SP-40,40 is unique and bears no obvious homology to those complement components whose disulphide bonds have been assigned, nor is there any homology apparent between SP-40,40 and other multi-chain proteins containing disulphide bonds.
Notes: Kirszbaum, L
Bozas, S E
Walker, I D
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1992 Feb 3;297(1-2):70-6. doi: 10.1016/0014-5793(92)80330-j.
Author Address: Department of Veterinary Sciences, University of Melbourne, Parkville, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 357
Author: Kirszbaum, L., Sharpe, J. A., Murphy, B., d'Apice, A. J., Classon, B., Hudson, P. and Walker, I. D.
Year: 1989
Title: Molecular cloning and characterization of the novel, human complement-associated protein, SP-40,40: a link between the complement and reproductive systems
Journal: Embo j
Volume: 8
Issue: 3
Pages: 711-8
Epub Date: 1989/03/01
Date: Mar
Short Title: Molecular cloning and characterization of the novel, human complement-associated protein, SP-40,40: a link between the complement and reproductive systems
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
PMCID: PMC400866
Accession Number: 2721499
Keywords: Amino Acid Sequence
Animals
Base Sequence
Blood Proteins/*genetics/immunology/physiology
Cloning, Molecular
Clusterin
Complement System Proteins/*genetics
DNA/genetics
Glycoproteins/genetics
Humans
Male
*Molecular Chaperones
Molecular Sequence Data
Protein Precursors/genetics
Rats
*Reproduction
Semen/metabolism
Sequence Homology, Nucleic Acid
Abstract: The cDNA sequence encoding the human complement-associated protein, SP-40,40, is reported. The two chains of SP-40,40 are coded in a single open reading frame on the same mRNA molecule, indicating the existence of a biosynthetic precursor protein which matures post-synthetically by the proteolysis of at least one peptide bond. The precursor is preceded by a signal sequence for vectorial export and contains six N-linked glycosylation sites distributed equally between the two chains of the structure. The sequence of the SP-40,40 precursor bears a 77% identity to a rat sulphated glycoprotein-2 (SGP-2) which is the major secreted product of Sertoli cells. The presence of SP-40,40 within human seminal plasma at levels comparable to those in serum was demonstrated, indicating that SP-40,40 and SGP-2 are serum and seminal forms of the same protein. A sequence of 23 amino acids within the beta-chain of SP-40,40 exhibited significant homology to corresponding segments located within complement components C7, C8 and C9. The short cysteine-containing motif represented the only evidence of a possible vestigial relationship between SP-40,40 and other complement components. The precise role of SP-40,40 is not known in either blood or semen but the present findings document an intriguing link between the immune and the reproductive systems.
Notes: 1460-2075
Kirszbaum, L
Sharpe, J A
Murphy, B
d'Apice, A J
Classon, B
Hudson, P
Walker, I D
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1989 Mar;8(3):711-8.
Author Address: Department of Veterinary Preclinical Sciences, University of Melbourne, Victoria, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 611
Author: Kise, H., Nishioka, J., Kawamura, J. and Suzuki, K.
Year: 1996
Title: Characterization of semenogelin II and its molecular interaction with prostate-specific antigen and protein C inhibitor
Journal: Eur J Biochem
Volume: 238
Issue: 1
Pages: 88-96
Epub Date: 1996/05/15
Date: May 15
Short Title: Characterization of semenogelin II and its molecular interaction with prostate-specific antigen and protein C inhibitor
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1996.0088q.x
Accession Number: 8665956
Keywords: Amino Acid Sequence
Cations
Dextran Sulfate/chemistry/pharmacology
Gonadal Steroid Hormones/*chemistry/*metabolism
Heparin/metabolism/pharmacology
Humans
Hydrogen-Ion Concentration
Male
Molecular Sequence Data
Organophosphorus Compounds/metabolism
Osmolar Concentration
Prostate-Specific Antigen/antagonists & inhibitors/*metabolism
Protein C Inhibitor/*metabolism/pharmacology
Protein Precursors/*chemistry/*metabolism
Semen/chemistry/drug effects
*Seminal Plasma Proteins
*Seminal Vesicle Secretory Proteins
Solubility
Abstract: The serine protease, prostate-specific antigen (PSA), its protein substrates, semenogelin (Sg) I and II, and protein C inhibitor (PCI) have been described as components of human seminal plasma. PCI was found to inhibit the PSA-catalyzed degradation of insoluble coagula Sg I + II by forming a PSA-PCI complex. Digestion of seminal coagula with PSA released PCI and PSA-PCI complex from the coagula into a soluble phase, suggesting the presence of active PCI binding to the coagula. To investigate the molecular interaction of Sg with PSA and PCI, we purified Sg II from seminal coagula as a soluble form and found that Sg II is glycosylated with heterogeneous carbohydrate moieties. Sg II bound to the solid-phase complex of diisopropylfluorophosphate (iPr2FP) and PSA with an apparent dissociation constant (kd) of 41 nM and to PCI with a Kd of 28 nM. The binding of Sg II to iPr2P-PSA was not affected by PCI and that of Sg II to PCI was not affected by iPr2P-PSA, suggesting that Sg II forms a ternary complex with PSA and PCI. The bindings of Sg II to both iPr2P-PSA and PCI were influenced by pH, ionic strength, heparin, dextran sulfate, and divalent cations, particularly by Zn2+. Treatment of Sg II with heparinase, heparitinase, N-glycanase, or with O-glycanase following sialidase did not affect the binding of Sg II to iPr2P-PSA and PCI. These findings suggested that PCI bound to Sg in seminal vesicles regulates the PSA-catalyzed degradation of Sg in seminal plasma, and that the binding of PCI and PSA to Sg is modulated by several factors such as pH, ionic strength, divalent cations, and heparin-like substances in seminal plasma.
Notes: Kise, H
Nishioka, J
Kawamura, J
Suzuki, K
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1996 May 15;238(1):88-96. doi: 10.1111/j.1432-1033.1996.0088q.x.
Author Address: Department of Molecular Pathobiology, Mie University School of Medicine, Tsu-city, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1695
Author: Kita, S., Matsubara, H., Kasai, Y., Tamaoki, T., Okabe, Y., Fukuhara, H., Kamishikiryo, J., Krayukhina, E., Uchiyama, S., Ose, T., Kuroki, K. and Maenaka, K.
Year: 2015
Title: Crystal structure of extracellular domain of human lectin-like transcript 1 (LLT1), the ligand for natural killer receptor-P1A
Journal: Eur J Immunol
Volume: 45
Issue: 6
Pages: 1605-13
Epub Date: 2015/04/01
Date: Jun
Short Title: Crystal structure of extracellular domain of human lectin-like transcript 1 (LLT1), the ligand for natural killer receptor-P1A
Alternate Journal: European journal of immunology
ISSN: 0014-2980
DOI: 10.1002/eji.201545509
Accession Number: 25826155
Keywords: Amino Acid Sequence
Binding Sites
Crystallization
Humans
Lectins, C-Type/*chemistry/genetics/metabolism
Ligands
*Models, Molecular
Molecular Sequence Data
Mutation
NK Cell Lectin-Like Receptor Subfamily B/*chemistry/metabolism
Protein Binding
*Protein Conformation
*Protein Interaction Domains and Motifs
Protein Multimerization
Receptors, Cell Surface/*chemistry/genetics/metabolism
Sequence Alignment
C-type lectin-like receptor
Cell surface receptor
Crystal structure
Natural killer cell
Protein-protein interactions
Abstract: Emerging evidence has revealed the pivotal roles of C-type lectin-like receptors (CTLRs) in the regulation of a wide range of immune responses. Human natural killer cell receptor-P1A (NKRP1A) is one of the CTLRs and recognizes another CTLR, lectin-like transcript 1 (LLT1) on target cells to control NK, NKT and Th17 cells. The structural basis for the NKRP1A-LLT1 interaction was limitedly understood. Here, we report the crystal structure of the ectodomain of LLT1. The plausible receptor-binding face of the C-type lectin-like domain is flat, and forms an extended β-sheet. The residues of this face are relatively conserved with another CTLR, keratinocyte-associated C-type lectin, which binds to the CTLR member, NKp65. A LLT1-NKRP1A complex model, prepared using the crystal structures of LLT1 and the keratinocyte-associated C-type lectin-NKp65 complex, reasonably satisfies the charge consistency and the conformational complementarity to explain a previous mutagenesis study. Furthermore, crystal packing and analytical ultracentrifugation revealed dimer formation, which supports a complex model. Our results provide structural insights for understanding the binding modes and signal transduction mechanisms, which are likely to be conserved in the CTLR family, and for further rational drug design towards regulating the LLT1 function.
Notes: 1521-4141
Kita, Shunsuke
Matsubara, Haruki
Kasai, Yoshiyuki
Tamaoki, Takaharu
Okabe, Yuki
Fukuhara, Hideo
Kamishikiryo, Jun
Krayukhina, Elena
Uchiyama, Susumu
Ose, Toyoyuki
Kuroki, Kimiko
Maenaka, Katsumi
Journal Article
Research Support, Non-U.S. Gov't
Germany
Eur J Immunol. 2015 Jun;45(6):1605-13. doi: 10.1002/eji.201545509. Epub 2015 Apr 28.
Author Address: Laboratory of Biomolecular Science, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
Department of Materials Structure Science, Structural Biology Research Center, Photon Factory, Institute of Materials Structure Science, High Energy Accelerator Research Organization (KEK), The Graduate University for Advanced Studies, Tsukuba, Ibaraki, Japan.
Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, 1-7-1 Kohnodai, Ichikawa, Chiba, Japan.
Faculty of Pharmacy and Pharmaceutical Sciences, Fukuyama University, Fukuyama, Japan.
Department of Biotechnology, Graduate School of Engineering, Osaka University, Osaka, Japan.
U-Medico Corporation, Osaka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1740
Author: Kizuka, Y., Nakano, M., Kitazume, S., Saito, T., Saido, T. C. and Taniguchi, N.
Year: 2016
Title: Bisecting GlcNAc modification stabilizes BACE1 protein under oxidative stress conditions
Journal: Biochem J
Volume: 473
Issue: 1
Pages: 21-30
Epub Date: 2015/10/16
Date: Jan 1
Short Title: Bisecting GlcNAc modification stabilizes BACE1 protein under oxidative stress conditions
Alternate Journal: The Biochemical journal
ISSN: 0264-6021
DOI: 10.1042/bj20150607
Accession Number: 26467158
Keywords: Acetylglucosamine/*biosynthesis/genetics
Amyloid Precursor Protein Secretases/*biosynthesis
Animals
Aspartic Acid Endopeptidases/*biosynthesis
Brain/metabolism/pathology
Brain Injuries/genetics/metabolism/pathology
Cell Line, Transformed
Cell Line, Tumor
Male
Mice
Mice, Inbred C57BL
Mice, Knockout
Oxidative Stress/*physiology
Alzheimer’s disease
Bace1
GnT-III
bisecting GlcNAc
glycobiology
oxidative stress
Abstract: β-Site amyloid precursor protein-cleaving enzyme-1 (BACE1) is a protease essential for amyloid-β (Aβ) production in Alzheimer's disease (AD). BACE1 protein is known to be up-regulated by oxidative stress-inducing stimuli but the mechanism for this up-regulation still needs to be clarified. We have recently found that BACE1 is modified with bisecting N-acetylglucosamine (GlcNAc) by N-acetylglucosaminyltransferase-III (GnT-III, encoded by the Mgat3 gene) and that GnT-III deficiency reduces Aβ-plaque formation in the brain by accelerating lysosomal degradation of BACE1. Therefore, we hypothesized that bisecting GlcNAc would stabilize BACE1 protein on oxidative stress. In the present study, we first show that Aβ deposition in the mouse brain induces oxidative stress, together with an increase in levels of BACE1 and bisecting GlcNAc. Furthermore, prooxidant treatment induces expression of BACE1 protein in wild-type mouse embryonic fibroblasts (MEFs), whereas it reduces BACE1 protein in GnT-III (Mgat3) knock-out MEFs by accelerating lysosomal degradation of BACE1. We purified BACE1 from Neuro2A cells and performed LC/ESI/MS analysis for BACE1-derived glycopeptides and mapped bisecting GlcNAc-modified sites on BACE1. Point mutations at two N-glycosylation sites (Asn(153) and Asn(223)) abolish the bisecting GlcNAc modification on BACE1. These mutations almost cancelled the enhanced BACE1 degradation seen in Mgat3(-/-) MEFs, indicating that bisecting GlcNAc on BACE1 indeed regulates its degradation. Finally, we show that traumatic brain injury-induced BACE1 up-regulation is significantly suppressed in the Mgat3(-/-) brain. These results highlight the role of bisecting GlcNAc in oxidative stress-induced BACE1 expression and offer a novel glycan-targeted strategy for suppressing Aβ generation.
Notes: 1470-8728
Kizuka, Yasuhiko
Nakano, Miyako
Kitazume, Shinobu
Saito, Takashi
Saido, Takaomi C
Taniguchi, Naoyuki
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem J. 2016 Jan 1;473(1):21-30. doi: 10.1042/BJ20150607. Epub 2015 Oct 14.
Author Address: Disease Glycomics Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Centre for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Graduate School of Advanced Sciences of Matter, Hiroshima University, 1-3-1 Kagamiyama, Higashihiroshima, Hiroshima 739-8530, Japan.
Disease Glycomics Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Centre for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan shinobuk@riken.jp dglycotani@riken.jp rvw.tani@sanken.osaka-u.ac.jp.
Laboratory for Proteolytic Neuroscience, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1313
Author: Kjaer, S., Hanrahan, S., Totty, N. and McDonald, N. Q.
Year: 2010
Title: Mammal-restricted elements predispose human RET to folding impairment by HSCR mutations
Journal: Nat Struct Mol Biol
Volume: 17
Issue: 6
Pages: 726-31
Epub Date: 2010/05/18
Date: Jun
Short Title: Mammal-restricted elements predispose human RET to folding impairment by HSCR mutations
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.1808
Accession Number: 20473317
Keywords: Amino Acid Sequence
Animals
Cysteine/chemistry
Endoplasmic Reticulum/metabolism
Genetic Association Studies
Hirschsprung Disease/*genetics/*metabolism
Humans
Mammals
Models, Molecular
Molecular Sequence Data
*Mutation, Missense
Phylogeny
Protein Folding
Protein Interaction Domains and Motifs
Protein Multimerization
Protein Processing, Post-Translational
Proto-Oncogene Proteins c-ret/*chemistry/*genetics/metabolism
Sequence Homology, Amino Acid
Static Electricity
Abstract: The maturation of human RET is adversely affected by a range of missense mutations found in patients with Hirschsprung's disease (HSCR), a complex multigenic disease. Here we show that two N-terminal cadherin-like domains, CLD1 and CLD2 (CLD(1-2)), from human RET adopt a clam-shell arrangement distinct from that of classical cadherins. CLD1 structural elements and disulfide composition are unique to mammals, indicating an unexpected structural diversity within higher and lower vertebrate RET CLD regions. We identify two unpaired cysteines that predispose human RET to maturation impediments in the endoplasmic reticulum and establish a quantitative cell-based RET maturation assay that offers a biochemical correlate of HSCR disease severity. Our findings provide a key conceptual framework and means of testing and predicting genotype-phenotype correlations in HSCR.
Notes: 1545-9985
Kjaer, Svend
Hanrahan, Sarah
Totty, Nick
McDonald, Neil Q
Cancer Research UK/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2010 Jun;17(6):726-31. doi: 10.1038/nsmb.1808. Epub 2010 May 16.
Author Address: Structural Biology Laboratory, the London Research Institute, Cancer Research UK, London, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 526
Author: Kjeldsen, L., Johnsen, A. H., Sengeløv, H. and Borregaard, N.
Year: 1993
Title: Isolation and primary structure of NGAL, a novel protein associated with human neutrophil gelatinase
Journal: J Biol Chem
Volume: 268
Issue: 14
Pages: 10425-32
Epub Date: 1993/05/15
Date: May 15
Short Title: Isolation and primary structure of NGAL, a novel protein associated with human neutrophil gelatinase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7683678
Keywords: *Acute-Phase Proteins
Alpha-Globulins/chemistry
Amino Acid Sequence
Animals
Antibodies
Blotting, Western
Carrier Proteins/*blood/chemistry/*isolation & purification
Chromatography, Liquid
Collagenases/chemistry/*isolation & purification
Electrophoresis, Polyacrylamide Gel
Glycoproteins/*blood/*isolation & purification
Glycoside Hydrolases
Humans
Lipocalin-2
Lipocalins
Macromolecular Substances
Matrix Metalloproteinase 9
Mice
Molecular Sequence Data
Molecular Weight
Neutrophils/drug effects/*enzymology
*Oncogene Proteins
Proto-Oncogene Proteins
Rats
Sequence Homology, Amino Acid
Tetradecanoylphorbol Acetate/pharmacology
Abstract: A 25-kDa protein was found to be associated with purified human neutrophil gelatinase. Polyclonal antibodies raised against gelatinase not only recognized gelatinase but also this 25-kDa protein. Specific antibodies against the 25-kDa protein were obtained by affinity purification of the gelatinase antibodies. Immunoblotting and immunoprecipitation studies demonstrated the 135-kDa form of gelatinase to be a complex of 92-kDa gelatinase and the 25-kDa protein, and the 220-kDa form was demonstrated to be a homodimer of the 92-kDa protein, thus explaining the 220-, 135-, and 92-kDa forms characteristic of neutrophil gelatinase. The 25-kDa protein was purified to apparent homogeneity from exocytosed material from phorbol myristate acetate-stimulated neutrophils. The primary structure of the 25-kDa protein was determined as a 178-residue protein. It was susceptible to treatment with N-glycanase, and one N-glycosylation site was identified. The sequence did not match any known human protein, but showed a high degree of similarity with the deduced sequences of rat alpha 2-microglobulin-related protein and the mouse protein 24p3. It is thus a new member of the lipocalin family. The function of the 25-kDa protein, named neutrophil gelatinase-associated lipocalin (NGAL), remains to be determined.
Notes: Kjeldsen, L
Johnsen, A H
Sengeløv, H
Borregaard, N
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1993 May 15;268(14):10425-32.
Author Address: Departments of Hematology, University Hospital, Rigshospitalet, Copenhagen, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1261
Author: Klein, A. L., Berkaw, M. N., Buse, M. G. and Ball, L. E.
Year: 2009
Title: O-linked N-acetylglucosamine modification of insulin receptor substrate-1 occurs in close proximity to multiple SH2 domain binding motifs
Journal: Mol Cell Proteomics
Volume: 8
Issue: 12
Pages: 2733-45
Epub Date: 2009/08/13
Date: Dec
Short Title: O-linked N-acetylglucosamine modification of insulin receptor substrate-1 occurs in close proximity to multiple SH2 domain binding motifs
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M900207-MCP200
PMCID: PMC2816021
Accession Number: 19671924
Keywords: Acetylglucosamine/*analogs & derivatives/*metabolism
Amino Acid Motifs
Amino Acid Sequence
Animals
Binding Sites
Cell Line
Humans
Insulin Receptor Substrate Proteins/chemistry/genetics/*metabolism
Mass Spectrometry
Molecular Sequence Data
Phosphatidylinositol 3-Kinases/chemistry/metabolism
Phosphorylation
Polymorphism, Genetic
Protein Binding
Protein Processing, Post-Translational
Protein Subunits/chemistry/metabolism
Protein Tyrosine Phosphatase, Non-Receptor Type 11/chemistry/metabolism
Rats
Serine/metabolism
*src Homology Domains
Abstract: Insulin receptor substrate-1 (IRS-1) is a highly phosphorylated adaptor protein critical to insulin and IGF-1 receptor signaling. Ser/Thr kinases impact the metabolic and mitogenic effects elicited by insulin and IGF-1 through feedback and feed forward regulation at the level of IRS-1. Ser/Thr residues of IRS-1 are also O-GlcNAc-modified, which may influence the phosphorylation status of the protein. To facilitate the understanding of the functional effects of O-GlcNAc modification on IRS-1-mediated signaling, we identified the sites of O-GlcNAc modification of rat and human IRS-1. Tandem mass spectrometric analysis of IRS-1, exogenously expressed in HEK293 cells, revealed that the C terminus, which is rich in docking sites for SH2 domain-containing proteins, was O-GlcNAc-modified at multiple residues. Rat IRS-1 was O-GlcNAc-modified at Ser(914), Ser(1009), Ser(1036), and Ser(1041). Human IRS-1 was O-GlcNAc-modified at Ser(984) or Ser(985), at Ser(1011), and possibly at multiple sites within residues 1025-1045. O-GlcNAc modification at a conserved residue in rat (Ser(1009)) and human (Ser(1011)) IRS-1 is adjacent to a putative binding motif for the N-terminal SH2 domains of p85alpha and p85beta regulatory subunits of phosphatidylinositol 3-kinase and the tyrosine phosphatase SHP2 (PTPN11). Immunoblot analysis using an antibody generated against human IRS-1 Ser(1011) GlcNAc further confirmed the site of attachment and the identity of the +203.2-Da mass shift as beta-N-acetylglucosamine. The accumulation of IRS-1 Ser(1011) GlcNAc in HEPG2 liver cells and MC3T3-E1 preosteoblasts upon inhibition of O-GlcNAcase indicates that O-GlcNAcylation of endogenously expressed IRS-1 is a dynamic process that occurs at normal glucose concentrations (5 mm). O-GlcNAc modification did not occur at any known or newly identified Ser/Thr phosphorylation sites and in most cases occurred simultaneously with phosphorylation of nearby residues. These findings suggest that O-GlcNAc modification represents an additional layer of posttranslational regulation that may impact the specificity of effects elicited by insulin and IGF-1.
Notes: 1535-9484
Klein, Amanda L
Berkaw, Mary N
Buse, Maria G
Ball, Lauren E
R01 DK002001/DK/NIDDK NIH HHS/United States
R56 DK002001/DK/NIDDK NIH HHS/United States
P20RR017696/RR/NCRR NIH HHS/United States
U24 DE016508/DE/NIDCR NIH HHS/United States
P20 RR017696/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2009 Dec;8(12):2733-45. doi: 10.1074/mcp.M900207-MCP200. Epub 2009 Aug 11.
Author Address: Department of Molecular and Cellular Pharmacology, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1257
Author: Klíma, M., Zájedová, J., Doubravská, L. and Andera, L.
Year: 2009
Title: Functional analysis of the posttranslational modifications of the death receptor 6
Journal: Biochim Biophys Acta
Volume: 1793
Issue: 10
Pages: 1579-87
Epub Date: 2009/08/06
Date: Oct
Short Title: Functional analysis of the posttranslational modifications of the death receptor 6
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbamcr.2009.07.008
Accession Number: 19654028
Keywords: Cell Line
Cell Line, Tumor
Glycosylation
HL-60 Cells
HeLa Cells
Humans
Jurkat Cells
Lipoylation
Male
Membrane Microdomains/metabolism
Molecular Weight
Mutagenesis, Site-Directed
Protein Isoforms/chemistry/genetics/physiology
Protein Processing, Post-Translational
Protein Structure, Tertiary
Receptors, Tumor Necrosis Factor/*chemistry/genetics/*metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Deletion
Abstract: Death receptor 6 (DR6/TNFRSF21) is a death domain-containing receptor of the TNFR superfamily with an apparent regulatory function in hematopoietic and neuronal cells. In this study we document that DR6 is an extensively posttranslationally modified transmembrane protein and that N- and O-glycosylations of amino acids in its extracellular part are mainly responsible for its approximately 40 kDa mobility shift in SDS polyacrylamide gels. Site-directed mutagenesis confirmed that all six extracellular asparagines are N-glycosylated and that the Ser/Thr/Pro cluster in the "stalk" domain juxtaposed to the cysteine-rich domains (CRDs) is a major site for the likely mucine-type of O-glycosylation. Deletion of the entire linker region between CRDs and the transmembrane domain, spanning over 130 amino acids, severely compromises the plasma membrane localization of DR6 and leads to its intracellular retention. Biosynthetic labeling with radiolabeled palmitate and side-directed mutagenesis also revealed that the membrane-proximal Cys368 in the intracellular part of DR6 is, similarly as cysteines in Fas/CD95 or DR4 ICPs, S-palmitoylated. However, palmitoylation of Cys368 is apparently not required for DR6 targeting into Brij-98 insoluble lipid rafts. In contrast, we show that N-glycosylation of the extracellular part might participate in directing DR6 into these membrane microdomains.
Notes: Klíma, Martin
Zájedová, Jitka
Doubravská, Lenka
Andera, Ladislav
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2009 Oct;1793(10):1579-87. doi: 10.1016/j.bbamcr.2009.07.008. Epub 2009 Aug 3.
Author Address: Laboratory of Cell Signaling and Apoptosis, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1232
Author: Klusák, V., Barinka, C., Plechanovová, A., Mlcochová, P., Konvalinka, J., Rulísek, L. and Lubkowski, J.
Year: 2009
Title: Reaction mechanism of glutamate carboxypeptidase II revealed by mutagenesis, X-ray crystallography, and computational methods
Journal: Biochemistry
Volume: 48
Issue: 19
Pages: 4126-38
Epub Date: 2009/03/24
Date: May 19
Short Title: Reaction mechanism of glutamate carboxypeptidase II revealed by mutagenesis, X-ray crystallography, and computational methods
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi900220s
PMCID: PMC7289149
NIHMSID: NIHMS1589328
Accession Number: 19301871
Keywords: Alanine/metabolism
Amino Acid Substitution
Binding Sites/genetics
Computational Biology/*methods
*Crystallography, X-Ray
Dipeptides/genetics/metabolism
Glutamate Carboxypeptidase II/chemistry/*genetics/*metabolism
Humans
Hydrogen Bonding
Hydrolysis
Kinetics
Models, Biological
Models, Molecular
Molecular Sequence Data
*Mutagenesis
Protein Binding/genetics
Quantum Theory
Substrate Specificity/genetics
Thermodynamics
Zinc/chemistry
Abstract: Glutamate carboxypeptidase II (GCPII, EC 3.4.17.21) is a zinc-dependent exopeptidase and an important therapeutic target for neurodegeneration and prostate cancer. The hydrolysis of N-acetyl-l-aspartyl-l-glutamate (N-Ac-Asp-Glu), the natural dipeptidic substrate of the GCPII, is intimately involved in cellular signaling within the mammalian nervous system, but the exact mechanism of this reaction has not yet been determined. To investigate peptide hydrolysis by GCPII in detail, we constructed a mutant of human GCPII [GCPII(E424A)], in which Glu424, a putative proton shuttle residue, is substituted with alanine. Kinetic analysis of GCPII(E424A) using N-Ac-Asp-Glu as substrate revealed a complete loss of catalytic activity, suggesting the direct involvement of Glu424 in peptide hydrolysis. Additionally, we determined the crystal structure of GCPII(E424A) in complex with N-Ac-Asp-Glu at 1.70 A resolution. The presence of the intact substrate in the GCPII(E424A) binding cavity substantiates our kinetic data and allows a detailed analysis of GCPII/N-Ac-Asp-Glu interactions. The experimental data are complemented by the combined quantum mechanics/molecular mechanics calculations (QM/MM) which enabled us to characterize the transition states, including the associated reaction barriers, and provided detailed information concerning the GCPII reaction mechanism. The best estimate of the reaction barrier was calculated to be DeltaG(++) approximately 22(+/-5) kcal x mol(-1), which is in a good agreement with the experimentally observed reaction rate constant (k(cat) approximately 1 s(-1)). Combined together, our results provide a detailed and consistent picture of the reaction mechanism of this highly interesting enzyme at the atomic level.
Notes: 1520-4995
Klusák, Vojtech
Barinka, Cyril
Plechanovová, Anna
Mlcochová, Petra
Konvalinka, Jan
Rulísek, Lubomír
Lubkowski, Jacek
Z01 BC010761/Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Biochemistry. 2009 May 19;48(19):4126-38. doi: 10.1021/bi900220s.
Author Address: Institute of Organic Chemistry and Biochemistry, Gilead Sciences Research Center and IOCB, Academy of Sciences of the Czech Republic, Flemingovo nam. 2, 166 10 Praha 6, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1528
Author: Knapp, K., Zebisch, M., Pippel, J., El-Tayeb, A., Müller, C. E. and Sträter, N.
Year: 2012
Title: Crystal structure of the human ecto-5'-nucleotidase (CD73): insights into the regulation of purinergic signaling
Journal: Structure
Volume: 20
Issue: 12
Pages: 2161-73
Epub Date: 2012/11/13
Date: Dec 5
Short Title: Crystal structure of the human ecto-5'-nucleotidase (CD73): insights into the regulation of purinergic signaling
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126
DOI: 10.1016/j.str.2012.10.001
Accession Number: 23142347
Keywords: 5'-Nucleotidase/antagonists & inhibitors/*chemistry
Adenosine/chemistry
Adenosine Diphosphate/analogs & derivatives/chemistry
Amino Acid Sequence
Catalytic Domain
Coordination Complexes/chemistry
Crystallography, X-Ray
Enzyme Inhibitors/chemistry
Flavonoids/chemistry
GPI-Linked Proteins/antagonists & inhibitors/chemistry
Humans
Hydrogen Bonding
Hydrophobic and Hydrophilic Interactions
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Interaction Domains and Motifs
Protein Structure, Quaternary
Protein Structure, Secondary
Protein Subunits/antagonists & inhibitors/chemistry
*Signal Transduction
Structural Homology, Protein
Zinc/chemistry
Abstract: In vertebrates ecto-5'-nucleotidase (e5NT) catalyzes the hydrolysis of extracellular AMP to adenosine and represents the major control point for extracellular adenosine levels. Due to its pivotal role for activation of P1 adenosine receptors, e5NT has emerged as an appealing drug target for treatment of inflammation, chronic pain, hypoxia, and cancer. Crystal structures of the dimeric human e5NT reveal an extensive 114° conformational switch between the open and closed forms of the enzyme. The dimerization interface is formed by the C-terminal domains and exhibits interchain motions of up to 13°. Complex structures with adenosine and AMPCP indicate that structural control of the domain movement determines the selectivity for monophosphate nucleotides. Binding modes of nucleotide-derived and flavonoid-based compounds complexed to the C-terminal domain in the open form reveal an additional binding pocket of ∼210 Å(3) that might be explored to design more potent inhibitors.
Notes: 1878-4186
Knapp, Karen
Zebisch, Matthias
Pippel, Jan
El-Tayeb, Ali
Müller, Christa E
Sträter, Norbert
Journal Article
Research Support, Non-U.S. Gov't
United States
Structure. 2012 Dec 5;20(12):2161-73. doi: 10.1016/j.str.2012.10.001. Epub 2012 Nov 8.
Author Address: Institute of Bioanalytical Chemistry, Center for Biotechnology and Biomedicine, University of Leipzig, Deutscher Platz 5, 04103 Leipzig, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 601
Author: Knäuper, V., López-Otin, C., Smith, B., Knight, G. and Murphy, G.
Year: 1996
Title: Biochemical characterization of human collagenase-3
Journal: J Biol Chem
Volume: 271
Issue: 3
Pages: 1544-50
Epub Date: 1996/01/19
Date: Jan 19
Short Title: Biochemical characterization of human collagenase-3
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.3.1544
Accession Number: 8576151
Keywords: Amino Acid Sequence
Animals
Binding Sites
Collagenases/chemistry/*isolation & purification/*metabolism
DNA, Complementary
Electrophoresis, Polyacrylamide Gel
Enzyme Activation
Enzyme Precursors/chemistry/isolation & purification/*metabolism
Glycosylation
Humans
Kinetics
Matrix Metalloproteinase 13
Models, Structural
Molecular Sequence Data
Molecular Weight
Peptide Fragments/chemistry
*Protein Structure, Secondary
Recombinant Proteins/chemistry/isolation & purification/metabolism
Sequence Homology, Amino Acid
Substrate Specificity
Abstract: The cDNA of a novel matrix metalloproteinase, collagenase-3 (MMP-13) has been isolated from a breast tumor library (Freije, J. M. P., Dicz-Itza, I., Balbin, M., Sanchez, L. M., Blasco, R., Tolivia, J., and López-Otin, C. (1994) J. Biol. Chem. 269, 16766-16773), and a potential role in tumor progression has been proposed for this enzyme. In order to establish the possible role of collagenase-3 in connective tissue turnover, we have expressed and purified recombinant human procollagenase-3 and characterized the enzyme biochemically. The purified procollagenase-3 was shown to be glycosylated and displayed a M(r) of 60,000, the N-terminal sequence being LPLPSGGD, which is consistent with the cDNA-predicted sequence. The proenzyme was activated by p-aminophenylmercuric acetate or stromelysin, yielding an intermediate form of M(r) 50,000, which displayed the N-terminal sequence L58EVTGK. Further processing resulted in cleavage of the Glu84-Tyr85 peptide bond to the final active enzyme (M(r) 48,000). Trypsin activation of procollagenase-3 also generated a Tyr85 N terminus, but it was evident that the C-terminal domain was rapidly lost, and hence the collagenolytic activity diminished. Analysis of the substrate specificity of collagenase-3 revealed that soluble type II collagen was preferentially hydrolyzed, while the enzyme was 5 or 6 times less efficient at cleaving type I or III collagen. Fibrillar type I collagen was cleaved with comparable efficiency to the fibroblast and neutrophil collagenases (MMP-1 and MMP-8), respectively. Unlike these collagenases, gelatin and the peptide substrates Mea-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 and Mca-Pro-Cha-Gly-Nva-His-Ala-Dpa-NH2 were efficiently hydrolyzed as well, as would be predicted from the similarities between the active site sequence of collagenase-3 (MMP-13) and the gelatinases A and B. Active collagenase-3 was inhibited in a 1:1 stoichiometric fashion by the tissue inhibitors of metalloproteinases, TIMP-1, TIMP-2, and TIMP-3. These results suggest that in vivo collagenase-3 could play a significant role in the turnover of connective tissue matrix constituents.
Notes: Knäuper, V
López-Otin, C
Smith, B
Knight, G
Murphy, G
Wellcome Trust/United Kingdom
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1996 Jan 19;271(3):1544-50. doi: 10.1074/jbc.271.3.1544.
Author Address: Strangeways Research Laboratory, Department of Cell and Molecular Biology, Worts' Causeway, Cambridge, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 242
Author: Knepper, T. P., Arbogast, B., Schreurs, J. and Deinzer, M. L.
Year: 1992
Title: Determination of the glycosylation patterns, disulfide linkages, and protein heterogeneities of baculovirus-expressed mouse interleukin-3 by mass spectrometry
Journal: Biochemistry
Volume: 31
Issue: 46
Pages: 11651-9
Epub Date: 1992/11/24
Date: Nov 24
Short Title: Determination of the glycosylation patterns, disulfide linkages, and protein heterogeneities of baculovirus-expressed mouse interleukin-3 by mass spectrometry
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00161a053
Accession Number: 1445902
Keywords: Amino Acid Sequence
Animals
Baculoviridae/*genetics
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Disulfides/*chemistry
Glycosylation
Interleukin-3/*chemistry
Mass Spectrometry/methods
Mice
Molecular Sequence Data
Molecular Weight
Proteins/*chemistry
Abstract: The primary structure of mouse interleukin-3 (IL-3) expressed by recombinant baculovirus-infected silkworm (Bombyx mori) larvae was analyzed by subjecting isolated IL-3 derived peptides to liquid secondary ion mass spectrometry. Two species of IL-3 were isolated from the silkworm hemolymph by reverse-phase high-pressure liquid chromatography. The major component has M(r)20-22 x 10(3) as determined by SDS-PAGE. Liquid secondary ion mass spectrometric analysis was carried out on the reduced tryptic and endopeptidase lysyl-C peptides of glycosylated and deglycosylated IL-3. These studies provided evidence that (1) Asn-16 is heterogeneously glycosylated with four different oligosaccharides, (2) Asn-86 is either nonglycosylated or has attached to it one oligosaccharide, (3) the N-glycosylation sites Asn-44 and Asn-51 are not glycosylated, and (4) there is no O-glycosylation. Liquid secondary ion mass spectrometric analysis of the unreduced tryptic peptides provided evidence for disulfide linkages between Cys-140 and Cys-79 or Cys-80 and between Cys-17 and Cys-79 or Cys-80. In comparison to the major component, a minor IL-3 species (M(r) 17-19 x 10(3) by SDS-PAGE) isolated from the hemolymph showed no difference with respect to the glycosylation pattern or the disulfide linkages, but it was cleaved between Ala-127 and Ser-128, and only a disulfide linkage between Cys-140 and Cys-79 or Cys-80 held the molecule together.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Knepper, T P
Arbogast, B
Schreurs, J
Deinzer, M L
ES00210/ES/NIEHS NIH HHS/United States
ES0040/ES/NIEHS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1992 Nov 24;31(46):11651-9. doi: 10.1021/bi00161a053.
Author Address: Department of Agricultural Chemistry, Oregon State University, Corvallis 97331-7301.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1290
Author: Kobayashi, S., Fukuhara, A., Taguchi, T., Matsuda, M., Tochino, Y., Otsuki, M. and Shimomura, I.
Year: 2010
Title: Identification of a new secretory factor, CCDC3/Favine, in adipocytes and endothelial cells
Journal: Biochem Biophys Res Commun
Volume: 392
Issue: 1
Pages: 29-35
Epub Date: 2010/01/02
Date: Jan 29
Short Title: Identification of a new secretory factor, CCDC3/Favine, in adipocytes and endothelial cells
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2009.12.142
Accession Number: 20043878
Keywords: Adipocytes/*metabolism
Amino Acid Sequence
Animals
Blood Vessels/cytology/metabolism
COS Cells
Chlorocebus aethiops
Endothelium, Vascular/*metabolism
Glycosylation
Golgi Apparatus/metabolism
Humans
Male
Mice
Mice, Inbred C57BL
Molecular Sequence Data
Protein Multimerization
Proteins/genetics/*metabolism
Abstract: The vascular system secretes many bioactive factors. In a gene chip database, we searched for novel genes with signal sequences that are specifically expressed in murine aorta, and focused on one gene previously named CCDC3 (NCBI nucleotide entry NM_028804), and we designated as Favine (fat/vessel-derived secretory protein). Northern blot analysis revealed that CCDC3 was expressed abundantly in the aorta and adipose tissues. The mRNA levels of CCDC3 were higher in adipose tissues of obese db/db mice than control mice, and induced during differentiation of rat primary adipocytes. In differentiated adipocytes, CCDC3 mRNA expression was enhanced by insulin and pioglitazone, a PPARgamma agonist, and suppressed by TNF-alpha, isoproterenol and norepinephrine. Transient expression experiments followed by N-terminal amino acid sequence analysis revealed secretion of CCDC3 protein into the culture medium, which was dose-dependently reduced by brefeldin A, an inhibitor of Golgi-mediated secretory pathway. When expressed in COS-7 cells, CCDC3 protein was post-transcriptionally modified with N-glycosylation, and formed a dimer complex. These results indicate that CCDC3 is a protein secreted by adipocytes and endothelial cells, and that its level is regulated both hormonally and nutritionally.
Notes: 1090-2104
Kobayashi, Sachiko
Fukuhara, Atsunori
Taguchi, Takashi
Matsuda, Morihiro
Tochino, Yoshihiro
Otsuki, Michio
Shimomura, Iichiro
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2010 Jan 29;392(1):29-35. doi: 10.1016/j.bbrc.2009.12.142. Epub 2009 Dec 30.
Author Address: Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1823
Author: Kober, D. L., Alexander-Brett, J. M., Karch, C. M., Cruchaga, C., Colonna, M., Holtzman, M. J. and Brett, T. J.
Year: 2016
Title: Neurodegenerative disease mutations in TREM2 reveal a functional surface and distinct loss-of-function mechanisms
Journal: Elife
Volume: 5
Epub Date: 2016/12/21
Date: Dec 20
Short Title: Neurodegenerative disease mutations in TREM2 reveal a functional surface and distinct loss-of-function mechanisms
Alternate Journal: eLife
ISSN: 2050-084x
DOI: 10.7554/eLife.20391
PMCID: PMC5173322
Accession Number: 27995897
Keywords: Crystallography, X-Ray
Humans
Membrane Glycoproteins/*chemistry/genetics/*metabolism
Models, Molecular
Mutant Proteins/*chemistry/genetics/*metabolism
*Mutation
Neurodegenerative Diseases/*genetics
Protein Conformation
Receptors, Immunologic/*chemistry/genetics/*metabolism
*Alzheimer's disease
*biophysics
*crystallography
*human
*inflammation
*microglia
*mouse
*neuroscience
*structural biology
Abstract: Genetic variations in the myeloid immune receptor TREM2 are linked to several neurodegenerative diseases. To determine how TREM2 variants contribute to these diseases, we performed structural and functional studies of wild-type and variant proteins. Our 3.1 Å TREM2 crystal structure revealed that mutations found in Nasu-Hakola disease are buried whereas Alzheimer's disease risk variants are found on the surface, suggesting that these mutations have distinct effects on TREM2 function. Biophysical and cellular methods indicate that Nasu-Hakola mutations impact protein stability and decrease folded TREM2 surface expression, whereas Alzheimer's risk variants impact binding to a TREM2 ligand. Additionally, the Alzheimer's risk variants appear to epitope map a functional surface on TREM2 that is unique within the larger TREM family. These findings provide a guide to structural and functional differences among genetic variants of TREM2, indicating that therapies targeting the TREM2 pathway should be tailored to these genetic and functional differences with patient-specific medicine approaches for neurodegenerative disorders.
Notes: 2050-084x
Kober, Daniel L
Alexander-Brett, Jennifer M
Karch, Celeste M
Orcid: 0000-0002-6854-5547
Cruchaga, Carlos
Colonna, Marco
Holtzman, Michael J
Brett, Thomas J
Orcid: 0000-0002-6871-6676
K08 HL121168/HL/NHLBI NIH HHS/United States
P50 AG005681/AG/NIA NIH HHS/United States
R01 HL119813/HL/NHLBI NIH HHS/United States
R01 HL120153/HL/NHLBI NIH HHS/United States
K01 AG046374/AG/NIA NIH HHS/United States
RF1 AG044546/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Elife. 2016 Dec 20;5:e20391. doi: 10.7554/eLife.20391.
Author Address: Molecular Microbiology and Microbial Pathogenesis Program, Washington University School of Medicine, St. Louis, United States.
Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Washington University School of Medicine, St. Louis, United States.
Department of Psychiatry, Washington University School of Medicine, St. Louis, United States.
Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, United States.
Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, United States.
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1289
Author: Kodiha, M., Tran, D., Morogan, A., Qian, C. and Stochaj, U.
Year: 2009
Title: Dissecting the signaling events that impact classical nuclear import and target nuclear transport factors
Journal: PLoS One
Volume: 4
Issue: 12
Pages: e8420
Epub Date: 2009/12/31
Date: Dec 24
Short Title: Dissecting the signaling events that impact classical nuclear import and target nuclear transport factors
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0008420
PMCID: PMC2793512
Accession Number: 20041180
Keywords: Active Transport, Cell Nucleus/drug effects
Cell Compartmentation/drug effects
Cell Nucleus/*metabolism
Cellular Apoptosis Susceptibility Protein/metabolism
Genes, Reporter
HeLa Cells
Humans
MAP Kinase Signaling System/drug effects
Mitogen-Activated Protein Kinase Kinases/antagonists & inhibitors
Nuclear Envelope/metabolism
Nuclear Localization Signals/metabolism
Nuclear Pore Complex Proteins/metabolism
Nuclear Proteins/*metabolism
Oxidative Stress/drug effects
Phosphoinositide-3 Kinase Inhibitors
Protein Kinase Inhibitors/pharmacology
Protein Processing, Post-Translational/drug effects
*Signal Transduction/drug effects
Solubility/drug effects
Stress, Physiological/drug effects
Up-Regulation/drug effects
alpha Karyopherins/metabolism
Abstract: BACKGROUND: Signaling through MEK-->ERK1/2 and PI3 kinases is implicated in many aspects of cell physiology, including the survival of oxidant exposure. Oxidants play a role in numerous physiological and pathophysiological processes, many of which rely on transport in and out of the nucleus. However, how oxidative stress impacts nuclear trafficking is not well defined. METHODOLOGY/PRINCIPAL FINDINGS: To better understand the effect of stress on nucleocytoplasmic trafficking, we exposed cells to the oxidant diethyl maleate. This treatment activated MEK-->ERK1/2 as well as PI3 kinase-->Akt cascades and triggered the inhibition of classical nuclear import. To define the molecular mechanisms that regulate nuclear transport, we examined whether MEK and PI3 kinase signaling affected the localization of key transport factors. Using recently developed tools for image acquisition and analysis, the subcellular distributions of importin-alpha, CAS, and nucleoporins Nup153 and Nup88 were quantified in different cellular compartments. These studies identified specific profiles for the localization of transport factors in the nucleus and cytoplasm, and at the nuclear envelope. Our results demonstrate that MEK and PI3 kinase signaling as well as oxidative stress control nuclear trafficking and the localization of transport components. Furthermore, stress not only induced changes in transport factor distribution, but also upregulated post-translational modification of transport factors. Our results are consistent with the idea that the phosphorylation of importin-alpha, CAS, Nup153, and Nup88, and the O-GlcNAc modification of Nup153 increase when cells are exposed to oxidant. CONCLUSIONS/SIGNIFICANCE: Our studies defined the complex regulation of classical nuclear import and identified key transport factors that are targeted by stress, MEK, and PI3 kinase signaling.
Notes: 1932-6203
Kodiha, Mohamed
Tran, Dan
Morogan, Andreea
Qian, Cynthia
Stochaj, Ursula
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2009 Dec 24;4(12):e8420. doi: 10.1371/journal.pone.0008420.
Author Address: Department of Physiology, McGill University, Montreal, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1157
Author: Koehnke, J., Jin, X., Budreck, E. C., Posy, S., Scheiffele, P., Honig, B. and Shapiro, L.
Year: 2008
Title: Crystal structure of the extracellular cholinesterase-like domain from neuroligin-2
Journal: Proc Natl Acad Sci U S A
Volume: 105
Issue: 6
Pages: 1873-8
Epub Date: 2008/02/06
Date: Feb 12
Short Title: Crystal structure of the extracellular cholinesterase-like domain from neuroligin-2
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0711701105
PMCID: PMC2538853
Accession Number: 18250328
Keywords: Alternative Splicing
Animals
Binding Sites
Cell Adhesion Molecules, Neuronal
Cell Line
Cholinesterases/*chemistry
Crystallography, X-Ray
Dimerization
Humans
Membrane Proteins/*chemistry
Mice
Models, Molecular
Nerve Tissue Proteins/*chemistry
Protein Conformation
Abstract: Neuroligins (NLs) are catalytically inactive members of a family of cholinesterase-like transmembrane proteins that mediate cell adhesion at neuronal synapses. Postsynaptic neuroligins engage in Ca2+-dependent transsynaptic interactions via their extracellular cholinesterase domain with presynaptic neurexins (NRXs). These interactions may be regulated by two short splice insertions (termed A and B) in the NL cholinesterase domain. Here, we present the 3.3-A crystal structure of the ectodomain from NL2 containing splice insertion A (NL2A). The overall structure of NL2A resembles that of cholinesterases, but several structural features are unique to the NL proteins. First, structural elements surrounding the esterase active-site region differ significantly between active esterases and NL2A. On the opposite surface of the NL2A molecule, the positions of the A and B splice insertions identify a candidate NRX interaction site of the NL protein. Finally, sequence comparisons of NL isoforms allow for mapping the location of residues of previously identified mutations in NL3 and NL4 found in patients with autism spectrum disorders. Overall, the NL2 structure promises to provide a valuable model for dissecting NL isoform- and synapse-specific functions.
Notes: 1091-6490
Koehnke, Jesko
Jin, Xiangshu
Budreck, Elaine C
Posy, Shoshana
Scheiffele, Peter
Honig, Barry
Shapiro, Lawrence
F30 MH083473/MH/NIMH NIH HHS/United States
R01 NS045014/NS/NINDS NIH HHS/United States
U54 CA121852/CA/NCI NIH HHS/United States
F30MH083473/MH/NIMH NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2008 Feb 12;105(6):1873-8. doi: 10.1073/pnas.0711701105. Epub 2008 Feb 4.
Author Address: Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1371
Author: Kogure, A., Shiratori, I., Wang, J., Lanier, L. L. and Arase, H.
Year: 2011
Title: PANP is a novel O-glycosylated PILRα ligand expressed in neural tissues
Journal: Biochem Biophys Res Commun
Volume: 405
Issue: 3
Pages: 428-33
Epub Date: 2011/01/19
Date: Feb 18
Short Title: PANP is a novel O-glycosylated PILRα ligand expressed in neural tissues
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/j.bbrc.2011.01.047
PMCID: PMC4089865
NIHMSID: NIHMS588942
Accession Number: 21241660
Keywords: Amino Acid Sequence
Animals
Cloning, Molecular
Humans
Ligands
Melanoma, Experimental
Membrane Glycoproteins/*metabolism
Mice
Molecular Sequence Data
Nerve Tissue Proteins/chemistry/genetics/*metabolism
Rats
Rats, Wistar
Receptors, Immunologic/*metabolism
Abstract: PILRα is an immune inhibitory receptor possessing an immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic domain enabling it to deliver inhibitory signals. Binding of PILRα to its ligand CD99 is involved in immune regulation; however, whether there are other PILRα ligands in addition to CD99 is not known. Here, we report that a novel molecule, PILR-associating neural protein (PANP), acts as an additional ligand for PILRα. Transcription of PANP was mainly observed in neural tissues. PILRα-Ig fusion protein bound cells transfected with PANP and the transfectants stimulated PILRα reporter cells. Specific O-glycan structures on PANP were found to be required for PILR recognition of this ligand. These results suggest that PANP is involved in immune regulation as a ligand of the PILRα.
Notes: 1090-2104
Kogure, Amane
Shiratori, Ikuo
Wang, Jing
Lanier, Lewis L
Arase, Hisashi
R01 AI068129/AI/NIAID NIH HHS/United States
AI068129/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochem Biophys Res Commun. 2011 Feb 18;405(3):428-33. doi: 10.1016/j.bbrc.2011.01.047. Epub 2011 Jan 15.
Author Address: Department of Immunochemistry, Research Institute for Microbial Diseases, Osaka University, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 980
Author: Kohno, T. and Igarashi, Y.
Year: 2004
Title: Roles for N-glycosylation in the dynamics of Edg-1/S1P1 in sphingosine 1-phosphate-stimulated cells
Journal: Glycoconj J
Volume: 21
Issue: 8-9
Pages: 497-501
Epub Date: 2005/03/08
Short Title: Roles for N-glycosylation in the dynamics of Edg-1/S1P1 in sphingosine 1-phosphate-stimulated cells
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/s10719-004-5540-8
Accession Number: 15750791
Keywords: Amino Acid Sequence
Animals
Cell Line
Glycosylation
Lysophospholipids/*metabolism/pharmacology
Molecular Sequence Data
Protein Transport
Receptors, Lysosphingolipid/*metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Sphingosine/*analogs & derivatives/*metabolism/pharmacology
Abstract: Sphingosine 1-phosphate (Sph-1-P) is a bioactive lipid mediator released from activated platelets. To date, 5 seven-transmembrane-spanning receptors, Edg-1/S1P1, Edg-3/S1P3, Edg-5/S1P2, Edg-6/S1P4 and Edg-8/S1P5, have been identified as specific Sph-1-P receptors. Our recent novel studies established that Edg-1/S1P1 is glycosylated in its N-terminal extracellular portion and further identified the specific glycosylation site as asparagine 30. We also demonstrated that the structure of the N-terminal ectodomain of Edg-1/S1P1 affects both its transport to the cell surface and the N-glycosylation process. These studies revealed a possible regulatory role for the N-glycan on Edg-1/S1P1 in the dynamics of the receptor, such as its lateral and internal movements within the membrane, in ligand-stimulated mammalian cells.
Notes: Kohno, Takayuki
Igarashi, Yasuyuki
Journal Article
Research Support, Non-U.S. Gov't
Review
United States
Glycoconj J. 2004;21(8-9):497-501. doi: 10.1007/s10719-004-5540-8.
Author Address: Department of Biomembrane and Biofunctional Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 562
Author: Kokenyesi, R. and Bernfield, M.
Year: 1994
Title: Core protein structure and sequence determine the site and presence of heparan sulfate and chondroitin sulfate on syndecan-1
Journal: J Biol Chem
Volume: 269
Issue: 16
Pages: 12304-9
Epub Date: 1994/04/22
Date: Apr 22
Short Title: Core protein structure and sequence determine the site and presence of heparan sulfate and chondroitin sulfate on syndecan-1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8163535
Keywords: Amino Acid Sequence
Animals
Antibodies
Blotting, Western
CHO Cells
Cells, Cultured
Chondroitin Sulfates/*analysis/chemistry
Cricetinae
Culture Media, Conditioned
Cyanogen Bromide
Epithelium/metabolism
Female
Heparitin Sulfate/*analysis/chemistry
Mammary Glands, Animal/*metabolism
Membrane Glycoproteins/biosynthesis/*chemistry/isolation & purification
Mice
Molecular Sequence Data
Oligopeptides/chemical synthesis/immunology
Peptide Fragments/chemistry/isolation & purification
Proteoglycans/biosynthesis/*chemistry/isolation & purification
Syndecan-1
Syndecans
Transfection
Abstract: Most proteoglycans bear either chondroitin sulfate or heparan sulfate chains linked to serine residues at Ser-Gly attachment sites on the core protein. However, only a fraction of proteins with Ser-Gly sites exhibit glycosaminoglycan chains. A variable proportion of these sites may be glycanated, and an unknown mechanism distinguishes whether these sites are for chondroitin sulfate or heparan sulfate. To evaluate the core protein features that determine whether and where chondroitin sulfate or heparan sulfate will be linked, we have studied mouse syndecan-1, a transmembrane proteoglycan that is invariably glycanated and can contain both chondroitin sulfate and heparan sulfate chains. The extracellular domain of the syndecan-1 core protein contains five Ser-Gly sites, three clustered near its N terminus and two adjacent to the transmembrane domain near its C terminus. We have established the distribution of glycosaminoglycans on these attachment clusters. In contrast to the C-terminal cluster, the N-terminal cluster was always glycanated, suggesting that this domain of the core protein contains sequences responsible for the invariable attachment of glycosaminoglycan chains. Solely chondroitin sulfate was found on the C-terminal cluster. This cluster contains the sequences EGSGE and ETSGE, both estimated to be on the protein surface in a hydrophilic environment. Heparan sulfate was found solely on the N-terminal cluster, which also bears some chondroitin sulfate. This cluster contains the sequences FSGSGTG and DGSGD, the former estimated to be in a hydrophobic pocket and the latter, similar to the sequence on the C-terminal cluster, in an exposed hydrophilic region. This glycosaminoglycan distribution was identical on mouse syndecan-1 produced by either mouse epithelial (NMuMG) or hamster mesenchymal (CHO) cells, suggesting that site-specific attachment of glycosaminoglycans is independent of cell type. These results implicate a cellular mechanism that distinguishes among the potential sites and attaches the correct glycosaminoglycan type unambiguously. Thus, structural elements of the core protein other than the Ser-Gly attachment sites determine if a site will be glycanated and, if so, whether with chondroitin sulfate or heparan sulfate.
Notes: Kokenyesi, R
Bernfield, M
CA28735/CA/NCI NIH HHS/United States
HD06763/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1994 Apr 22;269(16):12304-9.
Author Address: Joint Program in Neonatology, Harvard Medical School, Boston, Massachusetts 02115.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1491
Author: Kolarich, D., Jensen, P. H., Altmann, F. and Packer, N. H.
Year: 2012
Title: Determination of site-specific glycan heterogeneity on glycoproteins
Journal: Nat Protoc
Volume: 7
Issue: 7
Pages: 1285-98
Epub Date: 2012/06/09
Date: Jun 7
Short Title: Determination of site-specific glycan heterogeneity on glycoproteins
Alternate Journal: Nature protocols
ISSN: 1750-2799
DOI: 10.1038/nprot.2012.062
Accession Number: 22678432
Keywords: Chromatography, Liquid/methods
Electrophoresis, Polyacrylamide Gel
Erythropoietin/chemistry/metabolism
Glycoproteins/*chemistry/metabolism
Glycosylation
Humans
Polysaccharides/*chemistry/metabolism
Proteomics/*methods
Software
Tandem Mass Spectrometry/methods
Abstract: The comprehensive analysis of protein glycosylation is a major requirement for understanding glycoprotein function in biological systems, and is a prerequisite for producing recombinant glycoprotein therapeutics. This protocol describes workflows for the characterization of glycopeptides and their site-specific heterogeneity, showing examples of the analysis of recombinant human erythropoietin (rHuEPO), α1-proteinase inhibitor (A1PI) and immunoglobulin (IgG). Glycoproteins of interest can be proteolytically digested either in solution or in-gel after electrophoretic separation, and the (glyco)peptides are analyzed by capillary/nano-liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). If required, specific glycopeptide enrichment steps, such as hydrophilic interaction liquid chromatography (HILIC), can also be performed. Particular emphasis is placed on data interpretation and the determination of site-specific glycan heterogeneity. The described workflow takes approximately 3-5 d, including sample preparation and data analysis. The data obtained from analyzing released glycans of rHuEPO and IgG, described in the second protocol of this series (10.1038/nprot.2012.063), provide complementary detailed glycan structural information that facilitates characterization of the glycopeptides.
Notes: 1750-2799
Kolarich, Daniel
Jensen, Pia H
Altmann, Friedrich
Packer, Nicolle H
Journal Article
Research Support, Non-U.S. Gov't
England
Nat Protoc. 2012 Jun 7;7(7):1285-98. doi: 10.1038/nprot.2012.062.
Author Address: Biomolecular Frontiers Research Centre, Faculty of Science, Macquarie University, Sydney, New South Wales, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1153
Author: Kolarich, D., Weber, A., Pabst, M., Stadlmann, J., Teschner, W., Ehrlich, H., Schwarz, H. P. and Altmann, F.
Year: 2008
Title: Glycoproteomic characterization of butyrylcholinesterase from human plasma
Journal: Proteomics
Volume: 8
Issue: 2
Pages: 254-63
Epub Date: 2008/01/19
Date: Jan
Short Title: Glycoproteomic characterization of butyrylcholinesterase from human plasma
Alternate Journal: Proteomics
ISSN: 1615-9853 (Print)
1615-9853
DOI: 10.1002/pmic.200700720
Accession Number: 18203274
Keywords: Amino Acid Sequence
Butyrylcholinesterase/*blood
Carbohydrate Sequence
Chromatography, Liquid
Glycoproteins/*chemistry
Glycoside Hydrolases/metabolism
Humans
Molecular Sequence Data
Oligosaccharides, Branched-Chain
Polysaccharides/chemistry
Proteomics/*methods
Abstract: Human butyrylcholinesterase (hBChE) is a highly glycosylated protein present in human plasma. The enzyme hydrolyses choline esters, for example benzoylcholine, butyrylthiocholine and acetylthiocholine as well as noncholine esters like heroin and aspirin. hBChE is primarily involved in neuronal transmission and is a potential bioscavenger of toxic organophosphates to protect acetylcholinesterase. A prerequisite for the therapeutic use of hBChE is a detailed characterization of this glycoprotein purified from human plasma. In this study, MS/MS could confirm most of the protein backbone, including the N- and the C-terminus. Site-specific analysis of all nine potential N-glycosylation sites revealed mainly mono- and disialylated N-glycans to be present on this glycoprotein. Sialic acids (Neu5Ac) are mainly alpha2,6-linked, however a fraction of the N-glycans contained Neu5Ac also in alpha2,3 linkage. On monosialylated N-glycans, sialic acid is exclusively located on the 3-arm and in alpha2,6 linkage, as verified by 2D-HPLC and exoglycosidase digests of 2-aminopyridine (PA)-labelled N-glycans. This first comprehensive glycoproteomic analysis of the important human plasma glycoprotein BChE did not give any indication of O-glycosylation or any other kind of PTMs as previously postulated.
Notes: Kolarich, Daniel
Weber, Alfred
Pabst, Martin
Stadlmann, Johannes
Teschner, Wolfgang
Ehrlich, Hartmut
Schwarz, Hans-Peter
Altmann, Friedrich
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2008 Jan;8(2):254-63. doi: 10.1002/pmic.200700720.
Author Address: Biochemistry Division, Department of Chemistry, University of Natural Resources and Applied Life Sciences, Vienna, Austria. daniel.kolarich@mq.edu.au
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1039
Author: Kolarich, D., Weber, A., Turecek, P. L., Schwarz, H. P. and Altmann, F.
Year: 2006
Title: Comprehensive glyco-proteomic analysis of human alpha1-antitrypsin and its charge isoforms
Journal: Proteomics
Volume: 6
Issue: 11
Pages: 3369-80
Epub Date: 2006/04/20
Date: Jun
Short Title: Comprehensive glyco-proteomic analysis of human alpha1-antitrypsin and its charge isoforms
Alternate Journal: Proteomics
ISSN: 1615-9853 (Print)
1615-9853
DOI: 10.1002/pmic.200500751
Accession Number: 16622833
Keywords: Carbohydrate Conformation
Cysteine/*chemistry
Fucose/chemistry
Glycosylation
Humans
Molecular Sequence Data
Peptide Fragments/chemistry
Polysaccharides/chemistry
Protein Isoforms/chemistry
Proteomics
Spectrometry, Mass, Electrospray Ionization
alpha 1-Antitrypsin/*chemistry
Abstract: Human alpha1-antitrypsin (A1PI) is a well-known glycoprotein in human plasma important for the protection of tissues from proteolytic enzymes. The three N-glycosylation sites of A1PI contain diantennary N-glycans but also triantennary and even traces of tetraantennary structures leading to the typical IEF pattern observed for A1PI. Here we present an approach to characterize A1PI isoforms from human plasma and its PTMs by LC-ESI-MS and LC-ESI-MS/MS of peptides obtained by proteolytic digestion. The single cysteine residue of A1PI formed a disulfide bridge with free cysteine. The variability of the number of antennae and hence sialic acids on glycosylation site N107, which even contained minute amounts of tetraantennary structures, emerged as a major cause for the IEF pattern of A1PI. Only negligible amounts of triantennary structures were identified attached to N70, and exclusively diantennary structures were present on site N271 in each of the isoforms analyzed. Exoglycosidase digests revealed alpha2,6-linked neuraminic acids on diantennary N-glycans, and triantennary contained additionally one single alpha2,3-neuraminic acid per N-glycan, which, together with a fucose, formed a sialyl Lewis X determinant on the beta1,4-linked N-acetylglucosamine, as shown by 2-D-HPLC of pyridylaminated asialoglycans. Fucosylation of diantennary structures was marginal and of the core alpha1,6 type.
Notes: Kolarich, Daniel
Weber, Alfred
Turecek, Peter L
Schwarz, Hans-Peter
Altmann, Friedrich
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2006 Jun;6(11):3369-80. doi: 10.1002/pmic.200500751.
Author Address: Department of Chemistry, Biochemistry Division, University of Natural Resources and Applied Life Sciences (BOKU), Vienna, Austria. daniel.kolarich@boku.ac.at
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1358
Author: Kommaddi, R. P., Dickson, K. M. and Barker, P. A.
Year: 2011
Title: Stress-induced expression of the p75 neurotrophin receptor is regulated by O-GlcNAcylation of the Sp1 transcription factor
Journal: J Neurochem
Volume: 116
Issue: 3
Pages: 396-405
Epub Date: 2010/11/26
Date: Feb
Short Title: Stress-induced expression of the p75 neurotrophin receptor is regulated by O-GlcNAcylation of the Sp1 transcription factor
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042
DOI: 10.1111/j.1471-4159.2010.07120.x
Accession Number: 21105874
Keywords: Acetylglucosamine/*metabolism
Acylation/physiology
Animals
Apoptosis/physiology
Cells, Cultured
Down-Regulation/genetics
Gene Expression Regulation, Enzymologic
HEK293 Cells
Humans
Mice
Mutation/genetics
N-Acetylglucosaminyltransferases/*metabolism
Osmolar Concentration
Osmotic Pressure/physiology
Receptors, Nerve Growth Factor/biosynthesis/*genetics/*metabolism
Sp1 Transcription Factor/genetics/*metabolism/physiology
Stress, Physiological/*physiology
Abstract: Injury-induced expression of p75 neurotrophin receptor (p75NTR) in the CNS induces neuronal apoptosis and prevents neuronal regrowth. The mechanisms regulating injury-induced p75NTR expression are poorly characterized but previous studies have established that reductions in extracellular osmolarity which mimic cytotoxic edema induce p75NTR gene expression through pathways that activate the Sp1 transcription factor. In this report, we examined how extracellular osmolarity converges on Sp1 to regulate p75NTR expression. We report that levels of O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT), the enzyme that mediates O-linked attachment of GlcNAc, are reduced by extracellular hypo-osmolarity and that global levels of protein O-GlcNAcylation and of Sp1 show a corresponding decline. We demonstrate that chemical and RNAi-based treatments that reduce cellular O-GlcNAcylation facilitate p75NTR induction by hypo-osmolarity, directly linking protein O-GlcNAcylation to p75NTR induction. To determine if Sp1 O-GlcNAc content regulates p75NTR expression, we replaced endogenous Sp1 with a Sp1 mutated at O-GlcNAc target residues. This O-GlcNAc-deficient form of Sp1-enhanced p75NTR expression, demonstrating that O-GlcNAcylation of Sp1 negatively regulates p75NTR expression. We conclude that a stress-induced decline in the O-GlcNAc content of Sp1 drives expression of p75NTR.
Notes: 1471-4159
Kommaddi, Reddy P
Dickson, Kathleen M
Barker, Philip A
MOP37850/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
England
J Neurochem. 2011 Feb;116(3):396-405. doi: 10.1111/j.1471-4159.2010.07120.x. Epub 2010 Dec 13.
Author Address: Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1611
Author: Kong, L., Giang, E., Nieusma, T., Kadam, R. U., Cogburn, K. E., Hua, Y., Dai, X., Stanfield, R. L., Burton, D. R., Ward, A. B., Wilson, I. A. and Law, M.
Year: 2013
Title: Hepatitis C virus E2 envelope glycoprotein core structure
Journal: Science
Volume: 342
Issue: 6162
Pages: 1090-4
Epub Date: 2013/11/30
Date: Nov 29
Short Title: Hepatitis C virus E2 envelope glycoprotein core structure
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1243876
PMCID: PMC3954638
NIHMSID: NIHMS562050
Accession Number: 24288331
Keywords: Antibodies, Neutralizing/chemistry
Antiviral Agents/chemistry
Binding Sites
Crystallography, X-Ray
Drug Design
Epitopes/chemistry/genetics
Humans
Immunoglobulin Fab Fragments/chemistry
Mutagenesis, Site-Directed
Protein Folding
Protein Structure, Tertiary
Tetraspanin 28/chemistry
Viral Envelope Proteins/*chemistry/immunology
Viral Hepatitis Vaccines/chemistry/immunology
Abstract: Hepatitis C virus (HCV), a Hepacivirus, is a major cause of viral hepatitis, liver cirrhosis, and hepatocellular carcinoma. HCV envelope glycoproteins E1 and E2 mediate fusion and entry into host cells and are the primary targets of the humoral immune response. The crystal structure of the E2 core bound to broadly neutralizing antibody AR3C at 2.65 angstroms reveals a compact architecture composed of a central immunoglobulin-fold β sandwich flanked by two additional protein layers. The CD81 receptor binding site was identified by electron microscopy and site-directed mutagenesis and overlaps with the AR3C epitope. The x-ray and electron microscopy E2 structures differ markedly from predictions of an extended, three-domain, class II fusion protein fold and therefore provide valuable information for HCV drug and vaccine design.
Notes: 1095-9203
Kong, Leopold
Giang, Erick
Nieusma, Travis
Kadam, Rameshwar U
Cogburn, Kristin E
Hua, Yuanzi
Dai, Xiaoping
Stanfield, Robyn L
Burton, Dennis R
Ward, Andrew B
Wilson, Ian A
Law, Mansun
AI079031/AI/NIAID NIH HHS/United States
R21 AI080916/AI/NIAID NIH HHS/United States
RR017573/RR/NCRR NIH HHS/United States
P41 GM103310/GM/NIGMS NIH HHS/United States
P41RR001209/RR/NCRR NIH HHS/United States
R01 AI084817/AI/NIAID NIH HHS/United States
R01 AI071084/AI/NIAID NIH HHS/United States
P41 RR017573/RR/NCRR NIH HHS/United States
U54 GM094586/GM/NIGMS NIH HHS/United States
R56 AI084817/AI/NIAID NIH HHS/United States
R01 AI079031/AI/NIAID NIH HHS/United States
P41 RR001209/RR/NCRR NIH HHS/United States
AI071084/AI/NIAID NIH HHS/United States
AI080916/AI/NIAID NIH HHS/United States
AI084817/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Science. 2013 Nov 29;342(6162):1090-4. doi: 10.1126/science.1243876.
Author Address: Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1644
Author: Kontro, H., Joenväärä, S., Haglund, C. and Renkonen, R.
Year: 2014
Title: Comparison of sialylated N-glycopeptide levels in serum of pancreatic cancer patients, acute pancreatitis patients, and healthy controls
Journal: Proteomics
Volume: 14
Issue: 15
Pages: 1713-23
Epub Date: 2014/05/21
Date: Aug
Short Title: Comparison of sialylated N-glycopeptide levels in serum of pancreatic cancer patients, acute pancreatitis patients, and healthy controls
Alternate Journal: Proteomics
ISSN: 1615-9853
DOI: 10.1002/pmic.201300270
Accession Number: 24841998
Keywords: Case-Control Studies
Glycopeptides/*blood/*chemistry
Humans
N-Acetylneuraminic Acid
Pancreatic Neoplasms/*blood
Pancreatitis/*blood
Acute pancreatitis
Glycoproteomics
Ms
Pancreatic cancer
Sialylation
Technology
Abstract: Serum protein glycosylation is known to be affected by pathological conditions, including cancer and inflammatory diseases. Pancreatic cancer patients would benefit from early diagnosis, as the disease is often detected in an advanced stage and has poor prognosis. Searching for changes in serum protein site-specific glycosylation could reveal novel glycoprotein biomarkers. We used Sambucus nigra lectin affinity chromatography to enrich α-2,6 sialylated tryptic N-glycopeptides from albumin-depleted sera of pancreatic cancer patients, acute pancreatitis patients, and healthy individuals, and compared their relative abundance using ultra performance LC-MS. Relative quantitation was done using the spectrum processing software MZmine. Identification was performed on the web-based tool GlycopeptideID, developed for in silico analysis of intact N-glycopeptides. Seventeen high-abundance serum proteins, mainly acute-phase proteins, and immunoglobulins, with total 27 N-glycosylation sites, and 62 glycoforms, were identified. Pancreatitis patient sera contained 38, and pancreatic cancer patients sera contained 13 glycoform changes with statistical significance (p < 0.05). In pancreatitis, up to tenfold changes were found in some glycoforms, and in pancreatic cancer, threefold. Analysis showed that the changes often concerned one or two, but not all, N-glycosylation sites in a specific glycoprotein. In conclusion, the analysis shows that pancreatic cancer, and acute pancreatitis are associated with changes in concentrations of intact sialylated N-glycopeptides derived from acute-phase proteins, and immunoglobulins, and that changes are site specific.
Notes: 1615-9861
Kontro, Hilkka
Joenväärä, Sakari
Haglund, Caj
Renkonen, Risto
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2014 Aug;14(15):1713-23. doi: 10.1002/pmic.201300270.
Author Address: Transplantation Laboratory, Haartman Institute, University of Helsinki, Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1654
Author: Koo, J. and Bahk, Y. Y.
Year: 2014
Title: In vivo putative O-GlcNAcylation of human SCP1 and evidence for possible role of its N-terminal disordered structure
Journal: BMB Rep
Volume: 47
Issue: 10
Pages: 593-8
Epub Date: 2014/08/02
Date: Oct
Short Title: In vivo putative O-GlcNAcylation of human SCP1 and evidence for possible role of its N-terminal disordered structure
Alternate Journal: BMB reports
ISSN: 1976-6696 (Print)
1976-6696
DOI: 10.5483/bmbrep.2014.47.10.144
PMCID: PMC4261519
Accession Number: 25081999
Keywords: Animals
Glycosylation
Green Fluorescent Proteins/metabolism
Humans
Intrinsically Disordered Proteins/chemistry/metabolism
Mice
NIH 3T3 Cells
Nuclear Proteins/*chemistry/*metabolism
Phosphoprotein Phosphatases/*chemistry/*metabolism
Protein Processing, Post-Translational
Serine/metabolism
Substrate Specificity
Abstract: RNA polymerase II carboxyl-terminal domain (RNAPII CTD) phosphatases are responsible for the dephosphorylation of the C-terminal domain of the small subunit of RNAPII in eukaryotes. Recently, we demonstrated the identification of several interacting partners with human small CTD phosphatase1 (hSCP1) and the substrate specificity to delineate an appearance of the dephosphorylation catalyzed by SCP1. In this study, using the established cells for inducibly expressing hSCP1 proteins, we monitored the modification of β-O-linked N-acetylglucosamine (O-GlcNAc). O-GlcNAcylation is one of the most common post-translational modifications (PTMs). To gain insight into the PTM of hSCP1, we used the Western blot, immunoprecipitation, succinylayed wheat germ agglutininprecipitation, liquid chromatography-mass spectrometry analyses, and site-directed mutagenesis and identified the Ser41 residue of hSCP1 as the O-GlcNAc modification site. These results suggest that hSCP1 may be an O-GlcNAcylated protein in vivo, and its N-terminus may function a possible role in the PTM, providing a scaffold for binding the protein(s).
Notes: 1976-670x
Koo, JaeHyung
Bahk, Young Yil
Journal Article
Research Support, Non-U.S. Gov't
BMB Rep. 2014 Oct;47(10):593-8. doi: 10.5483/bmbrep.2014.47.10.144.
Author Address: Department of Brain Science, Daegu-Gyeongbuk Institute of Science and Technology (DGIST), Daegu 711-873, Korea.
Department of Biotechnology, Konkuk University, Chungju 380-701, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1397
Author: Korekane, H., Korekane, A., Yamaguchi, Y., Kato, M., Miyamoto, Y., Matsumoto, A., Hasegawa, T., Suzuki, K., Taniguchi, N. and Ookawara, T.
Year: 2011
Title: N-Glycosylation profiling of recombinant mouse extracellular superoxide dismutase produced in Chinese hamster ovary cells
Journal: Glycoconj J
Volume: 28
Issue: 3-4
Pages: 183-96
Epub Date: 2011/05/17
Date: May
Short Title: N-Glycosylation profiling of recombinant mouse extracellular superoxide dismutase produced in Chinese hamster ovary cells
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080
DOI: 10.1007/s10719-011-9333-6
Accession Number: 21573946
Keywords: Animals
CHO Cells
Chromatography, High Pressure Liquid
Cricetinae
Cricetulus
Glycosylation
Mass Spectrometry
Mice
Recombinant Proteins/chemistry/genetics/metabolism
Superoxide Dismutase/chemistry/*genetics/metabolism
Abstract: Extracellular superoxide dismutase (EC-SOD), the major SOD isoenzyme in biological fluids, is known to be N-glycosylated and heterogeneous as was detected in most glycoproteins. However, only one N-glycan structure has been reported in recombinant human EC-SOD produced in Chinese hamster ovary (CHO) cells. Thus, a precise N-glycan profile of the recombinant EC-SOD is not available. In this study, we report profiling of the N-glycan in the recombinant mouse EC-SOD produced in CHO cells using high-resolution techniques, including the liberation of N-glycans by treatment with PNGase F, fluorescence labeling by pyridylamination, characterization by anion-exchange, normal and reversed phase-HPLC separation, and mass spectrometry. We succeeded in identifying 26 different types of N-glycans in the recombinant enzyme. The EC-SOD N-glycans were basically core-fucosylated (98.3% of the total N-glycan content), and were high mannose sugar chain, and mono-, bi-, tri-, and tetra-antennary complex sugar chains exhibiting varying degrees of sialylation. Four of the identified N-glycans were uniquely modified with a sulfate group, a Lewis(x) structure, or an α-Gal epitope. The findings will shed new light on the structure-function relationships of EC-SOD N-glycans.
Notes: 1573-4986
Korekane, Hiroaki
Korekane, Atsuko
Yamaguchi, Yoshiki
Kato, Masaki
Miyamoto, Yasuhide
Matsumoto, Akio
Hasegawa, Tomoko
Suzuki, Keiichiro
Taniguchi, Naoyuki
Ookawara, Tomomi
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 2011 May;28(3-4):183-96. doi: 10.1007/s10719-011-9333-6. Epub 2011 May 15.
Author Address: Department of Disease Glycomics (Seikagaku Corporation), The Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka, 567-0047, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 460
Author: Korrel, S. A., Clemetson, K. J., Van Halbeek, H., Kamerling, J. P., Sixma, J. J. and Vliegenthart, J. F.
Year: 1984
Title: Structural studies on the O-linked carbohydrate chains of human platelet glycocalicin
Journal: Eur J Biochem
Volume: 140
Issue: 3
Pages: 571-6
Epub Date: 1984/05/02
Date: May 2
Short Title: Structural studies on the O-linked carbohydrate chains of human platelet glycocalicin
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1984.tb08140.x
Accession Number: 6327299
Keywords: Blood Platelets/*analysis
Carbohydrates/*analysis
Chemical Phenomena
Chemistry
Glycoproteins/*blood
Humans
Magnetic Resonance Spectroscopy
Membrane Proteins/*blood
*Platelet Glycoprotein GPIb-IX Complex
Protons
Abstract: Glycocalicin (140 kDa), constituting the main part of glycoprotein Ib (160 kDa), was released from the human platelet membrane by the action of a Ca2+-dependent protease, present in the platelet cytoplasm and liberated during sonication of the platelet suspension. After activation of the protease by Ca2+, the sonicated platelet suspension was subjected to differential centrifugation. The supernatant was applied to a column of wheat germ agglutinin linked to Sepharose 4B; glycocalicin was eluted from the column with 2.5% (w/v) N-acetylglucosamine. Glycocalicin was found to contain 40% carbohydrate by weight, representing N- as well as O-glycosidically linked carbohydrate chains. The O-glycosidic chains were split off by alkaline cleavage in the presence of 3H-labelled NaBH4. The liberated 3H-labelled oligosaccharide-alditols were fractionated on a DEAE-Sephadex A-25 column. The structures of the oligosaccharide-alditols were investigated by 500-MHz 1H-NMR spectroscopy. The major compound was identified as NeuAc alpha(2----3)Ga1 beta(1----3)[NeuAc alpha(2----3)Ga1 beta(1----4)GlcNAc beta(1----6)]GalNAc-ol. Two minor compounds were found to be NeuAc alpha(2----3)Gal beta(1----3)[NeuAc alpha(2----6)]GalNAc-ol and NeuAc alpha(2----3)Gal beta(1----3)GalNAc-ol.
Notes: Korrel, S A
Clemetson, K J
Van Halbeek, H
Kamerling, J P
Sixma, J J
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1984 May 2;140(3):571-6. doi: 10.1111/j.1432-1033.1984.tb08140.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 389
Author: Korrel, S. A., Clemetson, K. J., van Halbeek, H., Kamerling, J. P., Sixma, J. J. and Vliegenthart, J. F.
Year: 1988
Title: Identification of a tetrasialylated monofucosylated tetraantennary N-linked carbohydrate chain in human platelet glycocalicin
Journal: FEBS Lett
Volume: 228
Issue: 2
Pages: 321-6
Epub Date: 1988/02/15
Date: Feb 15
Short Title: Identification of a tetrasialylated monofucosylated tetraantennary N-linked carbohydrate chain in human platelet glycocalicin
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(88)80024-3
Accession Number: 3342888
Keywords: Amino Sugars/analysis
Blood Platelets/*analysis
Carbohydrate Conformation
Carbohydrates/*analysis
Cell Membrane/analysis
Chromatography, Ion Exchange
Fucose/*analysis
Glycosylation
Humans
Hydrazines
Magnetic Resonance Spectroscopy
N-Acetylneuraminic Acid
*Platelet Glycoprotein GPIb-IX Complex
Platelet Membrane Glycoproteins/*analysis
Sialic Acids/*analysis
Abstract: Glycocalicin (140 kDa), the main constituent of the glycoprotein Ib alpha-chain (150 kDa) of the human platelet membrane, contains 4 putative N-glycosylation sites. For the structural analysis of the N-glycosidic carbohydrate chains of glycocalicin, the glycoprotein has been subjected to the hydrazinolysis procedure. The acidic carbohydrate chains obtained were fractionated by ion-exchange chromatography on DEAE-Sephadex A-25, and subsequently analyzed by sugar analysis, anion-exchange chromatography on Mono Q HR 5/5 and 500 MHz 1H-NMR spectroscopy. A novel tetrasialylated monofucosylated tetraantennary chain was identified in the glycoprotein. It could also be deduced that in all structures the alpha 2----6-linked NeuAc is attached exclusively at the Gal beta 1----2Man alpha 1----3 antenna, whereas the other antennae can be terminated with alpha 2----3-linked NeuAc. As minor constituents sialylated N-linked carbohydrate chains with a terminal Fuc alpha 1----2Gal beta 1----sequence were detected.
Notes: Korrel, S A
Clemetson, K J
van Halbeek, H
Kamerling, J P
Sixma, J J
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1988 Feb 15;228(2):321-6. doi: 10.1016/0014-5793(88)80024-3.
Author Address: Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1508
Author: Koth, C. M., Murray, J. M., Mukund, S., Madjidi, A., Minn, A., Clarke, H. J., Wong, T., Chiang, V., Luis, E., Estevez, A., Rondon, J., Zhang, Y., Hötzel, I. and Allan, B. B.
Year: 2012
Title: Molecular basis for negative regulation of the glucagon receptor
Journal: Proc Natl Acad Sci U S A
Volume: 109
Issue: 36
Pages: 14393-8
Epub Date: 2012/08/22
Date: Sep 4
Short Title: Molecular basis for negative regulation of the glucagon receptor
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1206734109
PMCID: PMC3437825
Accession Number: 22908259
Keywords: Amino Acid Sequence
Antibodies, Monoclonal/metabolism/pharmacology
Blotting, Western
Cell Line
Chromatography, Affinity
Crystallography
Enzyme-Linked Immunosorbent Assay
Humans
Immunoglobulin Fab Fragments/metabolism
*Models, Molecular
Molecular Sequence Data
Mutagenesis
*Protein Conformation
Protein Structure, Tertiary/genetics
Receptors, Glucagon/antagonists & inhibitors/*chemistry/*metabolism
Abstract: Members of the class B family of G protein-coupled receptors (GPCRs) bind peptide hormones and have causal roles in many diseases, ranging from diabetes and osteoporosis to anxiety. Although peptide, small-molecule, and antibody inhibitors of these GPCRs have been identified, structure-based descriptions of receptor antagonism are scarce. Here we report the mechanisms of glucagon receptor inhibition by blocking antibodies targeting the receptor's extracellular domain (ECD). These studies uncovered a role for the ECD as an intrinsic negative regulator of receptor activity. The crystal structure of the ECD in complex with the Fab fragment of one antibody, mAb1, reveals that this antibody inhibits glucagon receptor by occluding a surface extending across the entire hormone-binding cleft. A second antibody, mAb23, blocks glucagon binding and inhibits basal receptor activity, indicating that it is an inverse agonist and that the ECD can negatively regulate receptor activity independent of ligand binding. Biochemical analyses of receptor mutants in the context of a high-resolution ECD structure show that this previously unrecognized inhibitory activity of the ECD involves an interaction with the third extracellular loop of the receptor and suggest that glucagon-mediated structural changes in the ECD accompany receptor activation. These studies have implications for the design of drugs to treat class B GPCR-related diseases, including the potential for developing novel allosteric regulators that target the ECDs of these receptors.
Notes: 1091-6490
Koth, Christopher M
Murray, Jeremy M
Mukund, Susmith
Madjidi, Azadeh
Minn, Alexandra
Clarke, Holly J
Wong, Terence
Chiang, Vicki
Luis, Elizabeth
Estevez, Alberto
Rondon, Jesus
Zhang, Yingnan
Hötzel, Isidro
Allan, Bernard B
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2012 Sep 4;109(36):14393-8. doi: 10.1073/pnas.1206734109. Epub 2012 Aug 20.
Author Address: Department of Structural Biology, Molecular Biology, Antibody Engineering, Protein Chemistry, and Early Discovery Biochemistry, Genentech Inc, South San Francisco, CA 94080, USA. koth.christopher@gene.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1659
Author: Kowalewski, B., Lübke, T., Kollmann, K., Braulke, T., Reinheckel, T., Dierks, T. and Damme, M.
Year: 2014
Title: Molecular characterization of arylsulfatase G: expression, processing, glycosylation, transport, and activity
Journal: J Biol Chem
Volume: 289
Issue: 40
Pages: 27992-8005
Epub Date: 2014/08/20
Date: Oct 3
Short Title: Molecular characterization of arylsulfatase G: expression, processing, glycosylation, transport, and activity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M114.584144
PMCID: PMC4183830
Accession Number: 25135642
Keywords: Amino Acid Motifs
Animals
Arylsulfatases/chemistry/*genetics/*metabolism
Glycosylation
Humans
Lysosomes/chemistry/*enzymology/genetics
Mice
Mice, Knockout
Peptide Hydrolases/metabolism
Protein Precursors/chemistry/genetics/metabolism
Protein Processing, Post-Translational
Protein Transport
Arylsulfatase G
Enzyme Processing
Heparan Sulfate
Lysosomal Glycoprotein
Lysosomal Storage Disease
Lysosome
Mannose 6-Phosphate
Mucopolysaccharidosis
Proteolytic Processing
Sulfatase
Abstract: Arylsulfatase G (ARSG) is a recently identified lysosomal sulfatase that was shown to be responsible for the degradation of 3-O-sulfated N-sulfoglucosamine residues of heparan sulfate glycosaminoglycans. Deficiency of ARSG leads to a new type of mucopolysaccharidosis, as described in a mouse model. Here, we provide a detailed molecular characterization of the endogenous murine enzyme. ARSG is expressed and proteolytically processed in a tissue-specific manner. The 63-kDa single-chain precursor protein localizes to pre-lysosomal compartments and tightly associates with organelle membranes, most likely the endoplasmic reticulum. In contrast, proteolytically processed ARSG fragments of 34-, 18-, and 10-kDa were found in lysosomal fractions and lost their membrane association. The processing sites and a disulfide bridge between the 18- and 10-kDa chains could be roughly mapped. Proteases participating in the processing were identified as cathepsins B and L. Proteolytic processing is dispensable for hydrolytic sulfatase activity in vitro. Lysosomal transport of ARSG in the liver is independent of mannose 6-phosphate, sortilin, and Limp2. However, mutation of glycosylation site N-497 abrogates transport of ARSG to lysosomes in human fibrosarcoma cells, due to impaired mannose 6-phosphate modification.
Notes: 1083-351x
Kowalewski, Björn
Lübke, Torben
Kollmann, Katrin
Braulke, Thomas
Reinheckel, Thomas
Dierks, Thomas
Damme, Markus
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2014 Oct 3;289(40):27992-8005. doi: 10.1074/jbc.M114.584144. Epub 2014 Aug 18.
Author Address: From the Department of Chemistry, Biochemistry I, Bielefeld University, 33615 Bielefeld.
the Department of Biochemistry, Children's Hospital, University Medical Center Hamburg-Eppendorf, Hamburg 20246, and.
the Institute of Molecular Medicine and Cell Research and BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg 79104, Germany.
From the Department of Chemistry, Biochemistry I, Bielefeld University, 33615 Bielefeld, thomas.dierks@uni-bielefeld.de.
From the Department of Chemistry, Biochemistry I, Bielefeld University, 33615 Bielefeld, mdamme@biochem.uni-kiel.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1725
Author: Koymans, K. J., Feitsma, L. J., Brondijk, T. H., Aerts, P. C., Lukkien, E., Lössl, P., van Kessel, K. P., de Haas, C. J., van Strijp, J. A. and Huizinga, E. G.
Year: 2015
Title: Structural basis for inhibition of TLR2 by staphylococcal superantigen-like protein 3 (SSL3)
Journal: Proc Natl Acad Sci U S A
Volume: 112
Issue: 35
Pages: 11018-23
Epub Date: 2015/08/19
Date: Sep 1
Short Title: Structural basis for inhibition of TLR2 by staphylococcal superantigen-like protein 3 (SSL3)
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1502026112
PMCID: PMC4568226
Accession Number: 26283364
Keywords: Dimerization
Endotoxins/chemistry/genetics/*physiology
Molecular Structure
Mutagenesis
Protein Binding
Staphylococcus aureus/*physiology
Toll-Like Receptor 2/*antagonists & inhibitors/chemistry
S. aureus
Toll-like receptor
crystal structure
immune evasion
innate immunity
Abstract: Toll-like receptors (TLRs) are crucial in innate recognition of invading micro-organisms and their subsequent clearance. Bacteria are not passive bystanders and have evolved complex evasion mechanisms. Staphylococcus aureus secretes a potent TLR2 antagonist, staphylococcal superantigen-like protein 3 (SSL3), which prevents receptor stimulation by pathogen-associated lipopeptides. Here, we present crystal structures of SSL3 and its complex with TLR2. The structure reveals that formation of the specific inhibitory complex is predominantly mediated by hydrophobic contacts between SSL3 and TLR2 and does not involve interaction of TLR2-glycans with the conserved Lewis(X) binding site of SSL3. In the complex, SSL3 partially covers the entrance to the lipopeptide binding pocket in TLR2, reducing its size by ∼50%. We show that this is sufficient to inhibit binding of agonist Pam2CSK4 effectively, yet allows SSL3 to bind to an already formed TLR2-Pam2CSK4 complex. The binding site of SSL3 overlaps those of TLR2 dimerization partners TLR1 and TLR6 extensively. Combined, our data reveal a robust dual mechanism in which SSL3 interferes with TLR2 activation at two stages: by binding to TLR2, it blocks ligand binding and thus inhibits activation. Second, by interacting with an already formed TLR2-lipopeptide complex, it prevents TLR heterodimerization and downstream signaling.
Notes: 1091-6490
Koymans, Kirsten J
Feitsma, Louris J
Brondijk, T Harma C
Aerts, Piet C
Lukkien, Eddie
Lössl, Philip
van Kessel, Kok P M
de Haas, Carla J C
van Strijp, Jos A G
Huizinga, Eric G
Orcid: 0000-0001-6928-8426
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2015 Sep 1;112(35):11018-23. doi: 10.1073/pnas.1502026112. Epub 2015 Aug 17.
Author Address: Department of Medical Microbiology, University Medical Center Utrecht, NL-3584 CX, Utrecht, The Netherlands;
Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, NL-3584 CH, Utrecht, The Netherlands;
Biomolecular Mass Spectrometry and Proteomics, Bijvoet Centre for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Netherlands Proteomics Center, Utrecht University, NL-3584 CH, Utrecht, The Netherlands.
Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, NL-3584 CH, Utrecht, The Netherlands; e.g.huizinga@uu.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1900
Author: Kozielewicz, P., Alomar, H., Yusof, S., Grafton, G., Cooper, A. J., Curnow, S. J., Ironside, J. W., Pall, H. and Barnes, N. M.
Year: 2017
Title: N-glycosylation and expression in human tissues of the orphan GPR61 receptor
Journal: FEBS Open Bio
Volume: 7
Issue: 12
Pages: 1982-1993
Epub Date: 2017/12/12
Date: Dec
Short Title: N-glycosylation and expression in human tissues of the orphan GPR61 receptor
Alternate Journal: FEBS open bio
ISSN: 2211-5463 (Print)
2211-5463
DOI: 10.1002/2211-5463.12339
PMCID: PMC5715243
Accession Number: 29226084
Keywords: *Gpcr
*N‐linked glycosylation
*hippocampus
*lymphocyte
*membrane trafficking
Abstract: A number of members of the G protein-coupled receptor class of cell surface receptors are 'orphans' with no known endogenous ligand. One of these orphan receptors is GPR61; there are little data about its expression in human cells and tissues. In this study, we investigated the post-translational modification of GPR61 by N-glycosylation at an identified consensus N-glycosylation site (N12) and the impact of this modification upon the subcellular expression of the protein. The N-glycosylation inhibitor tunicamycin reduced the apparent molecular weight of immunoreactivity associated with myc-tagged GPR61 by 1-2 kDa, which was comparable to the evident molecular weight of the myc-tagged N12S GPR61 mutant with disrupted consensus N-glycosylation site. Analysis of GPR61 expression demonstrated that tunicamycin treatment reduced considerably heterologous expression of GPR61 in the cell membrane despite the N12S GPR61 mutant being readily expressed at the cell surface. These results demonstrate that GPR61 is subject to N-glycosylation but suggest this is not a prerequisite for cell surface expression, although N-glycosylation of other proteins may be important for cell membrane expression of GPR61. Expression of GPR61 protein was demonstrated at the cellular level in human hippocampus and human peripheral blood mononuclear cells. In the latter, there was a significantly higher expression of GPR61 in the Th17 cell subset in comparison with resting CD4+ cells, which may point toward a potential role for the GPR61 receptor in autoimmune diseases. This is the first report that GPR61 protein is subject to post-translational modification and is expressed in immune cell subsets and the hippocampus. These findings will help guide studies to investigate the function of GPR61.
Notes: 2211-5463
Kozielewicz, Paweł
Alomar, Hatun
Yusof, Syaratul
Grafton, Gillian
Cooper, Alison J
Curnow, S John
Ironside, James W
Pall, Hardev
Barnes, Nicholas M
G0600953/MRC_/Medical Research Council/United Kingdom
G0900580/MRC_/Medical Research Council/United Kingdom
MR/L016400/1/MRC_/Medical Research Council/United Kingdom
Journal Article
FEBS Open Bio. 2017 Nov 20;7(12):1982-1993. doi: 10.1002/2211-5463.12339. eCollection 2017 Dec.
Author Address: Institute of Clinical Sciences College of Medical and Dental Sciences University of Birmingham UK.
Present address: Department of Physiology and Pharmacology Karolinska Institutet Nanna Svartz väg 217 177 Stockholm Sweden.
Present address: Pharmacology and Toxicology Department College of Pharmacy King Saud University Riyadh 12372 Saudi Arabia.
Present address: Faculty of Pharmacy Universiti Kebangsaan Malaysia 50300 Kuala Lumpur Malaysia.
Institute of Inflammation and Ageing College of Medical and Dental Sciences University of Birmingham UK.
National CJD Research and Surveillance Unit Centre for Clinical Brain Sciences University of Edinburgh UK.
Neurology Department Queen Elizabeth Hospital Birmingham UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 828
Author: Kragh-Hansen, U., Donaldson, D. and Jensen, P. H.
Year: 2001
Title: The glycan structure of albumin Redhill, a glycosylated variant of human serum albumin
Journal: Biochim Biophys Acta
Volume: 1550
Issue: 1
Pages: 20-6
Epub Date: 2001/12/12
Date: Nov 26
Short Title: The glycan structure of albumin Redhill, a glycosylated variant of human serum albumin
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/s0167-4838(01)00264-3
Accession Number: 11738084
Keywords: Carbohydrate Sequence
Cyanogen Bromide
Glycoside Hydrolases
Glycosylation
Heterozygote
Humans
Hydrazines
Ligands
Molecular Conformation
Molecular Sequence Data
N-Acetylneuraminic Acid/chemistry
Neuraminidase
Oligosaccharides/chemistry/isolation & purification
Oxidation-Reduction
Peptide Fragments/isolation & purification
Polysaccharides/*chemistry
Serum Albumin/*chemistry/genetics
Serum Albumin, Human
Abstract: Although human serum albumin is synthesized without carbohydrate, glycosylated variants of the protein can be found. We have determined the structure of the glycan bound to the double-mutant albumin Redhill (-1 Arg, 320 Ala-->Thr). The oligosaccharide was released from the protein using anhydrous hydrazine, and its structure was investigated using neuraminidase and a reagent array analysis method, which is based on the use of specific exoglycosidases. The glycan was shown to be a disialylated biantennary complex type oligosaccharide N-linked to 318 Asn. However, a minor part (11 mol%) of the glycan was without sialic acid. The structure is principally the same as that of glycans bound to two other types of glycosylated albumin variants. Glycosylation can affect, for example, the fatty acid binding properties of albumin. Taking the present information into account, it is apparent that different effects on binding are caused not by different glycan structures but by different locations of attachment, with the possible addition of local conformational changes in the protein molecule.
Notes: Kragh-Hansen, U
Donaldson, D
Jensen, P H
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2001 Nov 26;1550(1):20-6. doi: 10.1016/s0167-4838(01)00264-3.
Author Address: Department of Medical Biochemistry, The University of Aarhus, Denmark. ukh@biokemi.au.dk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 649
Author: Krieg, J., Gläsner, W., Vicentini, A., Doucey, M. A., Löffler, A., Hess, D. and Hofsteenge, J.
Year: 1997
Title: C-Mannosylation of human RNase 2 is an intracellular process performed by a variety of cultured cells
Journal: J Biol Chem
Volume: 272
Issue: 42
Pages: 26687-92
Epub Date: 1997/10/23
Date: Oct 17
Short Title: C-Mannosylation of human RNase 2 is an intracellular process performed by a variety of cultured cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.42.26687
Accession Number: 9334252
Keywords: Animals
Antibody Specificity
Cell Line
Cloning, Molecular
Endoribonucleases/genetics/*metabolism
Humans
Mass Spectrometry
Peptide Mapping
Tryptophan/analogs & derivatives/metabolism
Abstract: C2-alpha-Mannosyltryptophan was discovered in RNase 2 from human urine, representing a novel way of attaching carbohydrate to a protein. Here, we have addressed two questions related to the biosynthesis of this modification: (i) is C-mannosylation part of the normal intracellular biosynthetic route, and (ii) how general is it, i.e. which organisms perform this kind of glycosylation? To answer the first question, RNase 2, which is identical to the eosinophil-derived neurotoxin, was isolated from intracellular stores of cultured human HL-60 cells. The enzyme was C-mannosylated at Trp-7, showing that the modification occurs intracellularly, before secretion of the protein. The second question was investigated by immunological and chemical analysis of RNase 2 purified from the supernatant of transiently transformed cells from different organisms. This revealed that C-mannosylation occurs in cells from man, green monkey, pig, mouse, and hamster. The observation that pig kidney cells contain the machinery for C-mannosylation of Trp-7 of human RNase 2 but that the homologous RNase from porcine kidney is not a substrate, since it does not contain a tryptophan at position 7, strongly suggests that C-mannosylated proteins other than RNase 2 exist. Recombinant RNase 2 isolated from insect cells, plant protoplasts, and Escherichia coli was not C-mannosylated. These results not only form the basis for further studies on the biochemical aspects of C-mannosylation but also have implications for the choice of cells for production of recombinant glycoproteins.
Notes: Krieg, J
Gläsner, W
Vicentini, A
Doucey, M A
Löffler, A
Hess, D
Hofsteenge, J
Journal Article
United States
J Biol Chem. 1997 Oct 17;272(42):26687-92. doi: 10.1074/jbc.272.42.26687.
Author Address: Friedrich Miescher-Institut, P. O. Box 2543, CH-4002 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 659
Author: Krieg, J., Hartmann, S., Vicentini, A., Gläsner, W., Hess, D. and Hofsteenge, J.
Year: 1998
Title: Recognition signal for C-mannosylation of Trp-7 in RNase 2 consists of sequence Trp-x-x-Trp
Journal: Mol Biol Cell
Volume: 9
Issue: 2
Pages: 301-9
Epub Date: 1998/04/04
Date: Feb
Short Title: Recognition signal for C-mannosylation of Trp-7 in RNase 2 consists of sequence Trp-x-x-Trp
Alternate Journal: Molecular biology of the cell
ISSN: 1059-1524 (Print)
1059-1524
DOI: 10.1091/mbc.9.2.301
PMCID: PMC25254
Accession Number: 9450956
Keywords: Amino Acid Sequence
Amino Acid Substitution
Animals
Carbon/chemistry
Cell Line
Endoribonucleases/*chemistry/genetics
Glycosylation
Humans
Kidney
Mannosyltransferases/*metabolism
Models, Molecular
Molecular Sequence Data
Protein Folding
Recombinant Fusion Proteins
Swine
Tryptophan/*chemistry
Abstract: C2-alpha-Mannosyltryptophan was discovered in human RNase 2, an enzyme that occurs in eosinophils and is involved in host defense. It represents a novel way of attaching carbohydrate to a protein in addition to the well-known N- and O-glycosylations. The reaction is specific, as in RNase 2 Trp-7, but never Trp-10, which is modified. In this article, we address which structural features provide the specificity of the reaction. Expression of chimeras of RNase 2 and nonglycosylated RNase 4 and deletion mutants in HEK293 cells identified residues 1-13 to be sufficient for C-mannosylation. Site-directed mutagenesis revealed the sequence Trp-x-x-Trp, in which the first Trp becomes mannosylated, as the specificity determinant. The Trp residue at position +3 can be replaced by Phe, which reduces the efficiency of the reaction threefold. Interpretation of the data in the context of the three-dimensional structure of RNase 2 strongly suggests that the primary, rather than the tertiary, structure forms the determinant. The sequence motif occurs in 336 mammalian proteins currently present in protein databases. Two of these proteins were analyzed protein chemically, which showed partial C-glycosylation of recombinant human interleukin 12. The frequent occurrence of the protein recognition motif suggests that C-glycosides could be part of the structure of more proteins than assumed so far.
Notes: Krieg, J
Hartmann, S
Vicentini, A
Gläsner, W
Hess, D
Hofsteenge, J
Journal Article
Mol Biol Cell. 1998 Feb;9(2):301-9. doi: 10.1091/mbc.9.2.301.
Author Address: Friedrich Miescher-Institut, CH-4002 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1946
Author: Kristensen, K. K., Midtgaard, S. R., Mysling, S., Kovrov, O., Hansen, L. B., Skar-Gislinge, N., Beigneux, A. P., Kragelund, B. B., Olivecrona, G., Young, S. G., Jørgensen, T. J. D., Fong, L. G. and Ploug, M.
Year: 2018
Title: A disordered acidic domain in GPIHBP1 harboring a sulfated tyrosine regulates lipoprotein lipase
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 26
Pages: E6020-e6029
Epub Date: 2018/06/15
Date: Jun 26
Short Title: A disordered acidic domain in GPIHBP1 harboring a sulfated tyrosine regulates lipoprotein lipase
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1806774115
PMCID: PMC6042107
Accession Number: 29899144
Keywords: Angiopoietin-like 4 Protein/chemistry/genetics/metabolism
Animals
Endothelial Cells/*metabolism/pathology
Humans
Hyperlipoproteinemia Type I/genetics/metabolism/pathology
Lipoprotein Lipase/chemistry/genetics/*metabolism
Mice
Protein Binding
Protein Domains
Receptors, Lipoprotein/chemistry/genetics/*metabolism
Tyrosine/chemistry/genetics/metabolism
*autoimmune disease
*electrostatic steering
*hypertriglyceridemia
*intravascular lipolysis
*intrinsically disordered region
investigators on a consortium grant. They did not collaborate directly.
Abstract: The intravascular processing of triglyceride-rich lipoproteins depends on lipoprotein lipase (LPL) and GPIHBP1, a membrane protein of endothelial cells that binds LPL within the subendothelial spaces and shuttles it to the capillary lumen. In the absence of GPIHBP1, LPL remains mislocalized within the subendothelial spaces, causing severe hypertriglyceridemia (chylomicronemia). The N-terminal domain of GPIHBP1, an intrinsically disordered region (IDR) rich in acidic residues, is important for stabilizing LPL's catalytic domain against spontaneous and ANGPTL4-catalyzed unfolding. Here, we define several important properties of GPIHBP1's IDR. First, a conserved tyrosine in the middle of the IDR is posttranslationally modified by O-sulfation; this modification increases both the affinity of GPIHBP1-LPL interactions and the ability of GPIHBP1 to protect LPL against ANGPTL4-catalyzed unfolding. Second, the acidic IDR of GPIHBP1 increases the probability of a GPIHBP1-LPL encounter via electrostatic steering, increasing the association rate constant (k(on)) for LPL binding by >250-fold. Third, we show that LPL accumulates near capillary endothelial cells even in the absence of GPIHBP1. In wild-type mice, we expect that the accumulation of LPL in close proximity to capillaries would increase interactions with GPIHBP1. Fourth, we found that GPIHBP1's IDR is not a key factor in the pathogenicity of chylomicronemia in patients with the GPIHBP1 autoimmune syndrome. Finally, based on biophysical studies, we propose that the negatively charged IDR of GPIHBP1 traverses a vast space, facilitating capture of LPL by capillary endothelial cells and simultaneously contributing to GPIHBP1's ability to preserve LPL structure and activity.
Notes: 1091-6490
Kristensen, Kristian K
Midtgaard, Søren Roi
Mysling, Simon
Kovrov, Oleg
Hansen, Lars Bo
Skar-Gislinge, Nicholas
Beigneux, Anne P
Kragelund, Birthe B
Olivecrona, Gunilla
Young, Stephen G
Jørgensen, Thomas J D
Fong, Loren G
Ploug, Michael
Orcid: 0000-0003-2215-4265
R01 HL087228/HL/NHLBI NIH HHS/United States
P01 HL146358/HL/NHLBI NIH HHS/United States
R01 HL125335/HL/NHLBI NIH HHS/United States
P01 HL090553/HL/NHLBI NIH HHS/United States
R35 HL139725/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2018 Jun 26;115(26):E6020-E6029. doi: 10.1073/pnas.1806774115. Epub 2018 Jun 13.
Author Address: Finsen Laboratory, Rigshospitalet, DK-2200 Copenhagen N, Denmark.
Biotech Research and Innovation Centre, University of Copenhagen, DK-2200 Copenhagen N, Denmark.
Structural Biophysics, Niels Bohr Institute, University of Copenhagen, DK-2100 Copenhagen Ø, Denmark.
Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense M, Denmark.
Department of Medical Biosciences, Umeå University, SE-901 87 Umeå, Sweden.
Zealand Pharma, DK-2600 Glostrup, Denmark.
Department of Medicine, University of California, Los Angeles, CA 90095.
Department of Biology, University of Copenhagen, DK-2200 Copenhagen N, Denmark.
Department of Medicine, University of California, Los Angeles, CA 90095; sgyoung@mednet.ucla.edu m-ploug@finsenlab.dk.
Department of Human Genetics, University of California, Los Angeles, CA 90095.
Finsen Laboratory, Rigshospitalet, DK-2200 Copenhagen N, Denmark; sgyoung@mednet.ucla.edu m-ploug@finsenlab.dk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 933
Author: Kristiansen, T. Z., Bunkenborg, J., Gronborg, M., Molina, H., Thuluvath, P. J., Argani, P., Goggins, M. G., Maitra, A. and Pandey, A.
Year: 2004
Title: A proteomic analysis of human bile
Journal: Mol Cell Proteomics
Volume: 3
Issue: 7
Pages: 715-28
Epub Date: 2004/04/16
Date: Jul
Short Title: A proteomic analysis of human bile
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M400015-MCP200
Accession Number: 15084671
Keywords: Amino Acid Sequence
Bile/*metabolism
Chromatography, Liquid
Electrophoresis, Polyacrylamide Gel
Humans
Isotope Labeling
Lectins/*chemistry
Mass Spectrometry
Molecular Sequence Data
*Proteome
Abstract: We have carried out a comprehensive characterization of human bile to define the bile proteome. Our approach involved fractionation of bile by one-dimensional gel electrophoresis and lectin affinity chromatography followed by liquid chromatography tandem mass spectrometry. Overall, we identified 87 unique proteins, including several novel proteins as well as known proteins whose functions are unknown. A large majority of the identified proteins have not been previously described in bile. Using lectin affinity chromatography and enzymatically labeling of asparagine residues carrying glycan moieties by (18)O, we have identified a total of 33 glycosylation sites. The strategy described in this study should be generally applicable for a detailed proteomic analysis of most body fluids. In combination with "tagging" approaches for differential proteomics, our method could be used for identification of cancer biomarkers from any body fluid.
Notes: Kristiansen, Troels Zakarias
Bunkenborg, Jakob
Gronborg, Mads
Molina, Henrik
Thuluvath, Paul J
Argani, Pedram
Goggins, Michael G
Maitra, Anirban
Pandey, Akhilesh
CA62924/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Mol Cell Proteomics. 2004 Jul;3(7):715-28. doi: 10.1074/mcp.M400015-MCP200. Epub 2004 Apr 14.
Author Address: McKusick-Nathans Institute of Genetic Medicine and Department of Biological Chemistry, Johns Hopkins University, Baltimore, MD 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 641
Author: Krogh, T. N., Bachmann, E., Teisner, B., Skjødt, K. and Højrup, P.
Year: 1997
Title: Glycosylation analysis and protein structure determination of murine fetal antigen 1 (mFA1)--the circulating gene product of the delta-like protein (dlk), preadipocyte factor 1 (Pref-1) and stromal-cell-derived protein 1 (SCP-1) cDNAs
Journal: Eur J Biochem
Volume: 244
Issue: 2
Pages: 334-42
Epub Date: 1997/03/01
Date: Mar 1
Short Title: Glycosylation analysis and protein structure determination of murine fetal antigen 1 (mFA1)--the circulating gene product of the delta-like protein (dlk), preadipocyte factor 1 (Pref-1) and stromal-cell-derived protein 1 (SCP-1) cDNAs
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1997.00334.x
Accession Number: 9118998
Keywords: Amino Acid Sequence
Animals
Calcium-Binding Proteins
Carbohydrate Sequence
Chemokine CXCL12
Chemokines/*genetics
*Chemokines, CXC
DNA, Complementary/genetics
Female
Glycoproteins/blood/*chemistry/*genetics
Glycosylation
Intercellular Signaling Peptides and Proteins
Intracellular Signaling Peptides and Proteins
Membrane Proteins/*genetics
Mice
Molecular Sequence Data
Molecular Structure
Neoplasm Proteins/*genetics
Peptide Fragments/chemistry/genetics/isolation & purification
Pregnancy
Repressor Proteins/*genetics
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: By means of sequence analysis, murine fetal antigen 1 (mFA1) isolated from Mus musculus amniotic fluid was shown to be the circulating protein of the delta-like protein, stromal-cell-derived protein 1 (SCP-1) and preadipocyte factor 1 (Pref-1) gene products. The protein contains 36 cysteine residues arranged in six epidermal-growth-factor-like domains. The purification of several C-terminal peptides of varying lengths showed mFA1 to be C-terminal heterogeneous. O-linked glycosylations of the NeuNAc alpha2-3Gal beta1-3(NeuNAc alpha2-6)GalNAc type were present on all C-terminal peptides at residues Thr235, Thr244 and Thr248, although glycosylation on Thr244 was only partial. Three N-linked glycosylations were localized in mFA1 (Asn77, Asn142 and Asn151), two of which (Asn142 and Asn151) were in the unusual Asn-Xaa-Cys motif. Fucosylated biantennary complex-type and small amounts (less than 5%) of triantennary complex-type structures were identified on the glycosylated asparagine residues using sequential exoglycosidase and endoglycosidase digestions combined with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The presence of O-linked monosaccharides (glucose attached to Ser71, Ser193 and fucose at Thr201) was tentatively ascertained by combining Edman degradation and MALDI-MS. The results presented shows mFA1 to be the circulating heterogeneous cleavage products of the membrane-bound protein encoded by the murine cDNAs dlk, pref-1 and SCP-1.
Notes: Krogh, T N
Bachmann, E
Teisner, B
Skjødt, K
Højrup, P
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1997 Mar 1;244(2):334-42. doi: 10.1111/j.1432-1033.1997.00334.x.
Author Address: Department of Molecular Biology, Odense University, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2027
Author: Kronlage, M., Dewenter, M., Grosso, J., Fleming, T., Oehl, U., Lehmann, L. H., Falcão-Pires, I., Leite-Moreira, A. F., Volk, N., Gröne, H. J., Müller, O. J., Sickmann, A., Katus, H. A. and Backs, J.
Year: 2019
Title: O-GlcNAcylation of Histone Deacetylase 4 Protects the Diabetic Heart From Failure
Journal: Circulation
Volume: 140
Issue: 7
Pages: 580-594
Epub Date: 2019/06/15
Date: Aug 13
Short Title: O-GlcNAcylation of Histone Deacetylase 4 Protects the Diabetic Heart From Failure
Alternate Journal: Circulation
ISSN: 0009-7322
DOI: 10.1161/circulationaha.117.031942
Accession Number: 31195810
Keywords: Animals
Animals, Newborn
Cells, Cultured
Diabetes Mellitus, Experimental/*metabolism/pathology
Diabetes Mellitus, Type 1/*metabolism/pathology
Heart Failure/*metabolism/pathology/*prevention & control
Histone Deacetylases/*metabolism
Humans
Male
Mice
Mice, 129 Strain
Mice, Inbred C57BL
Mice, Knockout
Rats
Rats, Wistar
Repressor Proteins/*metabolism
Serine/metabolism
*Hdac4
*calcium-calmodulin-dependent protein kinase type 2
*diabetes mellitus
*heart failure
Abstract: BACKGROUND: Worldwide, diabetes mellitus and heart failure represent frequent comorbidities with high socioeconomic impact and steadily growing incidence, calling for a better understanding of how diabetic metabolism promotes cardiac dysfunction. Paradoxically, some glucose-lowering drugs have been shown to worsen heart failure, raising the question of how glucose mediates protective versus detrimental cardiac signaling. Here, we identified a histone deacetylase 4 (HDAC4) subdomain as a molecular checkpoint of adaptive and maladaptive signaling in the diabetic heart. METHODS: A conditional HDAC4 allele was used to delete HDAC4 specifically in cardiomyocytes (HDAC4-knockout). Mice were subjected to diabetes mellitus either by streptozotocin injections (type 1 diabetes mellitus model) or by crossing into mice carrying a leptin receptor mutation (db/db; type 2 diabetes mellitus model) and monitored for remodeling and cardiac function. Effects of glucose and the posttranslational modification by β-linked N-acetylglucosamine (O-GlcNAc) on HDAC4 were investigated in vivo and in vitro by biochemical and cellular assays. RESULTS: We show that the cardio-protective N-terminal proteolytic fragment of HDAC4 is enhanced in vivo in patients with diabetes mellitus and mouse models, as well as in vitro under high-glucose and high-O-GlcNAc conditions. HDAC4-knockout mice develop heart failure in models of type 1 and type 2 diabetes mellitus, whereas wild-type mice do not develop clear signs of heart failure, indicating that HDAC4 protects the diabetic heart. Reexpression of the N-terminal fragment of HDAC4 prevents HDAC4-dependent diabetic cardiomyopathy. Mechanistically, the posttranslational modification of HDAC4 at serine (Ser)-642 by O-GlcNAcylation is an essential step for production of the N-terminal fragment of HDAC4, which was attenuated by Ca(2+)/calmodulin-dependent protein kinase II-mediated phosphorylation at Ser-632. Preventing O-GlcNAcylation at Ser-642 not only entirely precluded production of the N-terminal fragment of HDAC4 but also promoted Ca(2+)/calmodulin-dependent protein kinase II-mediated phosphorylation at Ser-632, pointing to a mutual posttranslational modification cross talk of (cardio-detrimental) phosphorylation at Ser-632 and (cardio-protective) O-GlcNAcylation at Ser-642. CONCLUSIONS: In this study, we found that O-GlcNAcylation of HDAC4 at Ser-642 is cardio-protective in diabetes mellitus and counteracts pathological Ca(2+)/calmodulin-dependent protein kinase II signaling. We introduce a molecular model explaining how diabetic metabolism possesses important cardio-protective features besides its known detrimental effects. A deeper understanding of the here-described posttranslational modification cross talk may lay the groundwork for the development of specific therapeutic concepts to treat heart failure in the context of diabetes mellitus.
Notes: 1524-4539
Kronlage, Mariya
Dewenter, Matthias
Grosso, Johannes
Fleming, Thomas
Oehl, Ulrike
Lehmann, Lorenz H
Falcão-Pires, Inês
Leite-Moreira, Adelino F
Volk, Nadine
Gröne, Hermann-Josef
Müller, Oliver J
Sickmann, Albert
Katus, Hugo A
Backs, Johannes
Journal Article
Research Support, Non-U.S. Gov't
United States
Circulation. 2019 Aug 13;140(7):580-594. doi: 10.1161/CIRCULATIONAHA.117.031942. Epub 2019 Jun 14.
Author Address: Institute of Experimental Cardiology (M.K., M.D., J.G., U.O., L.H.L., J.B.), Heidelberg University, Germany.
Department of Cardiology (M.K., L.H.L., O.J.M., H.A.K.), Heidelberg University, Germany.
German Centre for Cardiovascular Research (DZHK), partner site Heidelberg/Mannheim (M.K., M.D., J.G., U.O., L.H.L., J.B., L.H.L., O.J.M., H.A.K.).
Department of Internal Medicine I (T.F.), Heidelberg University, Germany.
Unidade de Investigação Cardiovascular, Departamento de Cirurgia e Fisiologia, Faculdade de Medicina, Universidade do Porto, Portugal (I.F.-P., A.F.L.-M.).
Tissue Bank of the National Center for Tumor Diseases, Heidelberg, Germany (N.V.).
Department of Cellular and Molecular Pathology, German Cancer Research Center, Heidelberg (H.-J.G.).
Institute of Pathology, University of Marburg, Germany (H.-J.G.).
Leibniz Institute for Analysical Sciences (ISAS), Dortmund, Germany (A.S.).
Medical Faculty, Medical Proteomics Center, Ruhr-University Bochum, Germany (A.S.).
Department of Chemistry, College of Physical Sciences, University of Aberdeen, United Kingdom (A.S.). Dr Müller is currently at the Department of Internal Medicine III, University of Kiel, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 383
Author: Krotkiewski, H., Lisowska, E., Angel, A. S. and Nilsson, B.
Year: 1988
Title: Structural analysis by fast-atom-bombardment mass spectrometry of the mixture of alditols derived from the O-linked oligosaccharides of murine glycophorins
Journal: Carbohydr Res
Volume: 184
Pages: 27-38
Epub Date: 1988/12/31
Date: Dec 31
Short Title: Structural analysis by fast-atom-bombardment mass spectrometry of the mixture of alditols derived from the O-linked oligosaccharides of murine glycophorins
Alternate Journal: Carbohydrate research
ISSN: 0008-6215 (Print)
0008-6215
DOI: 10.1016/0008-6215(88)80003-x
Accession Number: 3242812
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Erythrocytes/analysis
Glycopeptides/isolation & purification
*Glycophorins/isolation & purification
Indicators and Reagents
Mass Spectrometry/methods
Mice
Mice, Inbred BALB C
Molecular Sequence Data
*Oligosaccharides/isolation & purification
*Sialoglycoproteins/isolation & purification
Sugar Alcohols/*analysis
Abstract: The O-glycosylically linked oligosaccharides from glycophorins of BALB/c mouse erythrocytes were released as a mixture of alditol derivatives on reductive beta-elimination. A new approach, based on periodate oxidation in combination with f.a.b.-m.s., was used to elucidate the structure of one of the branched derivatives in the mixture. Evidence for the anomeric configuration was obtained by 500-MHz 1H-n.m.r. spectroscopy. The following structures were found: (Formula: see text).
Notes: Krotkiewski, H
Lisowska, E
Angel, A S
Nilsson, B
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Carbohydr Res. 1988 Dec 31;184:27-38. doi: 10.1016/0008-6215(88)80003-x.
Author Address: Department of Immunochemistry, Polish Academy of Sciences, Wrocław.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 584
Author: Krotkiewski, H., Lisowska, E., Nilsson, G., Grönberg, G. and Nilsson, B.
Year: 1993
Title: An improved approach to the analysis of the structure of small oligosaccharides of glycoproteins: application to the O-linked oligosaccharides from human glycophorin A
Journal: Carbohydr Res
Volume: 239
Pages: 35-50
Epub Date: 1993/02/01
Date: Feb 1
Short Title: An improved approach to the analysis of the structure of small oligosaccharides of glycoproteins: application to the O-linked oligosaccharides from human glycophorin A
Alternate Journal: Carbohydrate research
ISSN: 0008-6215 (Print)
0008-6215
DOI: 10.1016/0008-6215(93)84201-g
Accession Number: 8384526
Keywords: Acetylation
Blood Group Antigens/chemistry
Borohydrides
Carbohydrate Sequence
Erythrocytes/chemistry
Glycophorins/*chemistry
Humans
Magnetic Resonance Spectroscopy
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Oxidation-Reduction
Periodic Acid
Spectrometry, Mass, Fast Atom Bombardment
Stereoisomerism
Abstract: Treatment of purified human glycophorin A with alkaline borohydride cleaved the oligosaccharide side chains to yield alditol derivatives that were separated by gel filtration into three mixtures of low molecular weight compounds. Each mixture was oxidised with periodate, and the products were reduced with borohydride and analysed after acetylation or methylation by GLC-MS and FABMS. The resulting data allowed the monosaccharide sequence and linkage positions to be assigned to each component of the mixtures. The anomeric configuration was determined by 1H NMR spectroscopy of the intact fractions. The structures of a desialylated tetrasaccharide, two monosialylated trisaccharides, and five other minor products were defined.
Notes: Krotkiewski, H
Lisowska, E
Nilsson, G
Grönberg, G
Nilsson, B
Journal Article
Netherlands
Carbohydr Res. 1993 Feb 1;239:35-50. doi: 10.1016/0008-6215(93)84201-g.
Author Address: Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wroclaw.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 401
Author: Krusius, T. and Ruoslahti, E.
Year: 1986
Title: Primary structure of an extracellular matrix proteoglycan core protein deduced from cloned cDNA
Journal: Proc Natl Acad Sci U S A
Volume: 83
Issue: 20
Pages: 7683-7
Epub Date: 1986/10/01
Date: Oct
Short Title: Primary structure of an extracellular matrix proteoglycan core protein deduced from cloned cDNA
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.83.20.7683
PMCID: PMC386785
Accession Number: 3484330
Keywords: Amino Acid Sequence
Base Sequence
DNA/*analysis/isolation & purification
Decorin
Extracellular Matrix/*analysis
Extracellular Matrix Proteins
Humans
Nucleic Acid Hybridization
Proteoglycans/analysis/*genetics
Abstract: The core protein of a small chondroitin/dermatan sulfate proteoglycan expressed by human fibroblasts and present in extracellular matrices in association with collagen has been cloned from a lambda gt11 fibroblast cDNA library. cDNA clones were isolated by use of antibodies specific for the intact proteoglycan and antibodies against a peptide synthesized on the basis of the amino-terminal sequence of the core protein. A 1.8-kilobase cDNA was found to code for a prepro core protein composed of a signal peptide, a propeptide, and a mature core protein of 329 amino acids. The amino-terminal amino acid sequence deduced from the cDNA sequence was identical to that previously obtained by protein sequencing. The core protein contains three Ser-Gly dipeptide sequences, of which one is substituted with glycosaminoglycan. A protein data base homology search established the core protein sequence is a unique sequence distinct from published amino acid sequences. RNA blot hybridizations, performed using the cloned cDNA as a probe, revealed two related transcripts of 1.6 and 1.9 kilobases in RNA from both human fibroblast and placental tissue. Hybridization of genomic DNA restriction fragments suggested that there is one gene for the core protein of this proteoglycan and possibly one other closely related gene. Availability of the cloned cDNA for the proteoglycan now makes it possible to apply methods of molecular biology to study the collagen-binding and cell attachment-inhibiting properties of this proteoglycan.
Notes: 1091-6490
Krusius, T
Ruoslahti, E
CA28896/CA/NCI NIH HHS/United States
CA30199/CA/NCI NIH HHS/United States
F05 TW0 3477-01/TW/FIC NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1986 Oct;83(20):7683-7. doi: 10.1073/pnas.83.20.7683.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 774
Author: Kryger, G., Harel, M., Giles, K., Toker, L., Velan, B., Lazar, A., Kronman, C., Barak, D., Ariel, N., Shafferman, A., Silman, I. and Sussman, J. L.
Year: 2000
Title: Structures of recombinant native and E202Q mutant human acetylcholinesterase complexed with the snake-venom toxin fasciculin-II
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 56
Issue: Pt 11
Pages: 1385-94
Epub Date: 2000/10/29
Date: Nov
Short Title: Structures of recombinant native and E202Q mutant human acetylcholinesterase complexed with the snake-venom toxin fasciculin-II
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s0907444900010659
Accession Number: 11053835
Keywords: Acetylcholinesterase/*chemistry/genetics/isolation & purification
Amino Acid Sequence
Animals
Cholinesterase Inhibitors/*chemistry
Crystallography, X-Ray
Elapid Venoms/*chemistry
Elapidae
Humans
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Conformation
Recombinant Proteins/chemistry/genetics/isolation & purification
Sequence Homology, Amino Acid
Abstract: Structures of recombinant wild-type human acetylcholinesterase and of its E202Q mutant as complexes with fasciculin-II, a 'three-finger' polypeptide toxin purified from the venom of the eastern green mamba (Dendroaspis angusticeps), are reported. The structure of the complex of the wild-type enzyme was solved to 2.8 A resolution by molecular replacement starting from the structure of the complex of Torpedo californica acetylcholinesterase with fasciculin-II and verified by starting from a similar complex with mouse acetylcholinesterase. The overall structure is surprisingly similar to that of the T. californica enzyme with fasciculin-II and, as expected, to that of the mouse acetylcholinesterase complex. The structure of the E202Q mutant complex was refined starting from the corresponding wild-type human acetylcholinesterase structure, using the 2.7 A resolution data set collected. Comparison of the two structures shows that removal of the charged group from the protein core and its substitution by a neutral isosteric moiety does not disrupt the functional architecture of the active centre. One of the elements of this architecture is thought to be a hydrogen-bond network including residues Glu202, Glu450, Tyr133 and two bridging molecules of water, which is conserved in other vertebrate acetylcholinesterases as well as in the human enzyme. The present findings are consistent with the notion that the main role of this network is the proper positioning of the Glu202 carboxylate relative to the catalytic triad, thus defining its functional role in the interaction of acetylcholinesterase with substrates and inhibitors.
Notes: Kryger, G
Harel, M
Giles, K
Toker, L
Velan, B
Lazar, A
Kronman, C
Barak, D
Ariel, N
Shafferman, A
Silman, I
Sussman, J L
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
United States
Acta Crystallogr D Biol Crystallogr. 2000 Nov;56(Pt 11):1385-94. doi: 10.1107/s0907444900010659.
Author Address: Department of Structural Biology, Weizmann Institute of Science, Rehovot 76100, Israel.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 511
Author: Ku, N. O. and Omary, M. B.
Year: 1995
Title: Identification and mutational analysis of the glycosylation sites of human keratin 18
Journal: J Biol Chem
Volume: 270
Issue: 20
Pages: 11820-7
Epub Date: 1995/05/19
Date: May 19
Short Title: Identification and mutational analysis of the glycosylation sites of human keratin 18
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.270.20.11820
Accession Number: 7538124
Keywords: Amino Acid Sequence
Animals
Cell Line
Cricetinae
DNA, Complementary/genetics
Glycosylation
Humans
Keratins/genetics/*metabolism
Mice
Molecular Sequence Data
Mutagenesis, Site-Directed
Nucleopolyhedroviruses
*Protein Processing, Post-Translational
Recombinant Fusion Proteins/metabolism
Sequence Homology, Amino Acid
Spodoptera
Transfection
Abstract: Keratin polypeptides 8 and 18 (K8/18) are intermediate filament phosphoglycoproteins that are expressed preferentially in glandular epithelia. We previously showed that K8/18 phosphorylation occurs on serine residues and that K8/18 glycosylation consists of O-linked single N-acetylglucosamines (O-GlcNAc) that are linked to Ser/Thr. Since the function of these modifications is unknown, we sought as a first step to identify the precise modification sites and asked if they play a role in keratin filament assembly. For this, we generated a panel of K18 Ser and Thr-->Ala mutants at potential glycosylation sites followed by expression in a baculovirus-insect cell system. We identified the major glycosylation sites of K18 by comparing the tryptic 3H-glycopeptide pattern of the panel of mutant and wild type K18 expressed in the insect cells with the glycopeptides of K18 in human colonic cells. The identified sites occur on three serines in the head domain of K18. The precise modified residues in human cells were verified using Edman degradation and confirmed further by the lack of glycosylation of a K18 construct that was mutated at the molecularly identified sites then transfected into NIH-3T3 cells. Partial or total K18 glycosylation mutants transfected into mammalian cells manifested nondistinguishable filament assembly to cells transfected with wild type K8/18. Our results show that K18 glycosylation sites share some features with other already identified O-GlcNAc sites and may together help predict glycosylation sites of other intermediate filament proteins.
Notes: Ku, N O
Omary, M B
AA0947/AA/NIAAA NIH HHS/United States
DK38707/DK/NIDDK NIH HHS/United States
DK47918/DK/NIDDK NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1995 May 19;270(20):11820-7. doi: 10.1074/jbc.270.20.11820.
Author Address: Palo Alto Veterans Administration Medical Center, California 94304, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1042
Author: Kubota, T., Shiba, T., Sugioka, S., Furukawa, S., Sawaki, H., Kato, R., Wakatsuki, S. and Narimatsu, H.
Year: 2006
Title: Structural basis of carbohydrate transfer activity by human UDP-GalNAc: polypeptide alpha-N-acetylgalactosaminyltransferase (pp-GalNAc-T10)
Journal: J Mol Biol
Volume: 359
Issue: 3
Pages: 708-27
Epub Date: 2006/05/03
Date: Jun 9
Short Title: Structural basis of carbohydrate transfer activity by human UDP-GalNAc: polypeptide alpha-N-acetylgalactosaminyltransferase (pp-GalNAc-T10)
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2006.03.061
Accession Number: 16650853
Keywords: Amino Acid Sequence
Carbohydrates/*chemistry
Catalytic Domain
Crystallography, X-Ray
Glycosylation
Humans
Isoenzymes/chemistry/genetics
Manganese/chemistry
*Models, Molecular
Molecular Sequence Data
Mucins/chemistry
Mutation
N-Acetylgalactosaminyltransferases/*chemistry/genetics
Protein Binding
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics
Substrate Specificity
Uridine Diphosphate/chemistry
Abstract: Mucin-type O-glycans are important carbohydrate chains involved in differentiation and malignant transformation. Biosynthesis of the O-glycan is initiated by the transfer of N-acetylgalactosamine (GalNAc) which is catalyzed by UDP-GalNAc:polypeptide alpha-N-acetylgalactosaminyltransferases (pp-GalNAc-Ts). Here we present crystal structures of the pp-GalNAc-T10 isozyme, which has specificity for glycosylated peptides, in complex with the hydrolyzed donor substrate UDP-GalNAc and in complex with GalNAc-serine. A structural comparison with uncomplexed pp-GalNAc-T1 suggests that substantial conformational changes occur in two loops near the catalytic center upon donor substrate binding, and that a distinct interdomain arrangement between the catalytic and lectin domains forms a narrow cleft for acceptor substrates. The distance between the catalytic center and the carbohydrate-binding site on the lectin beta sub-domain influences the position of GalNAc glycosylation on GalNAc-glycosylated peptide substrates. A chimeric enzyme in which the two domains of pp-GalNAc-T10 are connected by a linker from pp-GalNAc-T1 acquires activity toward non-glycosylated acceptors, identifying a potential mechanism for generating the various acceptor specificities in different isozymes to produce a wide range of O-glycans.
Notes: Kubota, Tomomi
Shiba, Tomoo
Sugioka, Shigemi
Furukawa, Sanae
Sawaki, Hiromichi
Kato, Ryuich
Wakatsuki, Soichi
Narimatsu, Hisashi
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2006 Jun 9;359(3):708-27. doi: 10.1016/j.jmb.2006.03.061. Epub 2006 Apr 19.
Author Address: Glycogene Function Team of Research Center for Glycoscience (RCG), National Institute of Advanced Industrial Science and Technology (AIST), Central 2, 1-1-1 Umezono, Tsukuba, Ibaraki 305-8568, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1871
Author: Kubota, Y., Fujioka, K. and Takekawa, M.
Year: 2017
Title: WGA-based lectin affinity gel electrophoresis: A novel method for the detection of O-GlcNAc-modified proteins
Journal: PLoS One
Volume: 12
Issue: 7
Pages: e0180714
Epub Date: 2017/07/08
Short Title: WGA-based lectin affinity gel electrophoresis: A novel method for the detection of O-GlcNAc-modified proteins
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0180714
PMCID: PMC5501588
Accession Number: 28686627
Keywords: Acetylglucosamine/*chemistry/metabolism
Electrophoresis
Glycosylation
*Protein Processing, Post-Translational
Proteins/chemistry/isolation & purification/*metabolism
Wheat Germ Agglutinins/chemistry
Abstract: Post-translational modification with O-linked β-N-acetylglucosamine (O-GlcNAc) occurs selectively on serine and/or threonine residues of cytoplasmic and nuclear proteins, and dynamically regulates their molecular functions. Since conventional strategies to evaluate the O-GlcNAcylation level of a specific protein require time-consuming steps, the development of a rapid and easy method for the detection and quantification of an O-GlcNAcylated protein has been a challenging issue. Here, we describe a novel method in which O-GlcNAcylated and non-O-GlcNAcylated forms of proteins are separated by lectin affinity gel electrophoresis using wheat germ agglutinin (WGA), which primarily binds to N-acetylglucosamine residues. Electrophoresis of cell lysates through a gel containing copolymerized WGA selectively induced retardation of the mobility of O-GlcNAcylated proteins, thereby allowing the simultaneous visualization of both the O-GlcNAcylated and the unmodified forms of proteins. This method is therefore useful for the quantitative detection of O-GlcNAcylated proteins.
Notes: 1932-6203
Kubota, Yuji
Fujioka, Ko
Takekawa, Mutsuhiro
Journal Article
PLoS One. 2017 Jul 7;12(7):e0180714. doi: 10.1371/journal.pone.0180714. eCollection 2017.
Author Address: Division of Cell Signaling and Molecular Medicine, Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1580
Author: Kuhn, B., Benz, J., Greif, M., Engel, A. M., Sobek, H. and Rudolph, M. G.
Year: 2013
Title: The structure of human α-2,6-sialyltransferase reveals the binding mode of complex glycans
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 69
Issue: Pt 9
Pages: 1826-38
Epub Date: 2013/09/04
Date: Sep
Short Title: The structure of human α-2,6-sialyltransferase reveals the binding mode of complex glycans
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449
DOI: 10.1107/s0907444913015412
Accession Number: 23999306
Keywords: Antigens, CD/*chemistry/metabolism
Catalytic Domain
Crystallography, X-Ray
Humans
Polysaccharides/*chemistry/*metabolism
Protein Binding
Sialyltransferases/*chemistry/metabolism
glycosylation
sialyltransferases
β-galactoside α-2,6-sialyltransferase I
Abstract: Human β-galactoside α-2,6-sialyltransferase I (ST6Gal-I) establishes the final glycosylation pattern of many glycoproteins by transferring a sialyl moiety to a terminal galactose. Complete sialylation of therapeutic immunoglobulins is essential for their anti-inflammatory activity and protein stability, but is difficult to achieve in vitro owing to the limited activity of ST6Gal-I towards some galactose acceptors. No structural information on ST6Gal-I that could help to improve the enzymatic properties of ST6Gal-I for biotechnological purposes is currently available. Here, the crystal structures of human ST6Gal-I in complex with the product cytidine 5'-monophosphate and in complex with cytidine and phosphate are described. These complexes allow the rationalization of the inhibitory activity of cytosine-based nucleotides. ST6Gal-I adopts a variant of the canonical glycosyltransferase A fold and differs from related sialyltransferases by several large insertions and deletions that determine its regiospecificity and substrate specificity. A large glycan from a symmetry mate localizes to the active site of ST6Gal-I in an orientation compatible with catalysis. The glycan binding mode can be generalized to any glycoprotein that is a substrate of ST6Gal-I. Comparison with a bacterial sialyltransferase in complex with a modified sialyl donor lends insight into the Michaelis complex. The results support an SN2 mechanism with inversion of configuration at the sialyl residue and suggest substrate-assisted catalysis with a charge-relay mechanism that bears a conceptual similarity to serine proteases.
Notes: 1399-0047
Kuhn, Bernd
Benz, Jörg
Greif, Michael
Engel, Alfred M
Sobek, Harald
Rudolph, Markus G
Journal Article
United States
Acta Crystallogr D Biol Crystallogr. 2013 Sep;69(Pt 9):1826-38. doi: 10.1107/S0907444913015412. Epub 2013 Aug 17.
Author Address: pRED Pharma Research and Early Development, Discovery Technologies, F. Hoffmann-La Roche AG, Grenzacher Strasse 124, 4070 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 631
Author: Kumar, H. P., Hague, C., Haley, T., Starr, C. M., Besman, M. J., Lundblad, R. L. and Baker, D.
Year: 1996
Title: Elucidation of N-linked oligosaccharide structures of recombinant human factor VIII using fluorophore-assisted carbohydrate electrophoresis
Journal: Biotechnol Appl Biochem
Volume: 24
Issue: 3
Pages: 207-16
Epub Date: 1996/12/01
Date: Dec
Short Title: Elucidation of N-linked oligosaccharide structures of recombinant human factor VIII using fluorophore-assisted carbohydrate electrophoresis
Alternate Journal: Biotechnology and applied biochemistry
ISSN: 0885-4513 (Print)
0885-4513
Accession Number: 8969451
Keywords: Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cricetinae
Electrophoresis/*methods
Factor VIII/biosynthesis/*chemistry/genetics
*Fluorescent Dyes
Hexosaminidases
Humans
Molecular Sequence Data
Oligosaccharides/*chemistry
Recombinant Proteins/biosynthesis/chemistry
Abstract: Characterization of the carbohydrate moiety is a critical measure of manufacturing process consistency of recombinant human Factor VIII (rFVIII) in Chinese-hamster ovary (CHO) cells. FVIII, a large (300 kDa) glycoprotein, is employed therapeutically for the correction of haemophilia A. While N-linked and O-linked oligosaccharides are found in this protein, the current study focuses on the N-linked oligosaccharides. The N-linked oligosaccharides from rFVIII were released using either peptide N-glycosidase F or endoglycosidase H, derivatized with the fluorophore 8-aminonaphthalene-1,3,6-trisulphonate, and analysed by fluorophore-assisted carbohydrate electrophoresis (FACE). The electrophoretically resolved oligosaccharide bands were isolated and individual bands subjected to digestion with defined pools of exoglycosidases and re-electrophoresed on FACE sequencing gels. The resulting gel patterns were interpreted, based on band mobility shifts, to obtain the sequence structure of the oligosaccharides. A total of eight acidic and 12 neutral structures were identified, and the majority of the oligosaccharides (approximately 92%) were found to be sialylated. All of the major oligosaccharide structures found in CHO-cell-derived rFVIII have also been reported to be present in plasma-derived FVIII. Among the most abundant are disialylated, biantennary, core-fucosylated (approximately 40%), followed by trisialylated, triantennary, core-fucosylated and monosialo, biantennary, core-fucosylated structures (each approximately 18%). The Gal alpha 1-3Gal structures reported to be present in baby-hamster-kidney-cell-derived rFVIII were not found in the CHO-cell-derived protein. The glycosylation patterns were consistent in six random lots of rFVIII [coefficient of variation (%) 3-14] based on percentage lane luminance data of bands that represent approximately 98% of all asparagine-linked oligosaccharides.
Notes: Kumar, H P
Hague, C
Haley, T
Starr, C M
Besman, M J
Lundblad, R L
Baker, D
Journal Article
United States
Biotechnol Appl Biochem. 1996 Dec;24(3):207-16.
Author Address: Baxter Biotech, Hyland R&D Division, Duarte, CA 91010, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1351
Author: Kumar, J. and Mayer, M. L.
Year: 2010
Title: Crystal structures of the glutamate receptor ion channel GluK3 and GluK5 amino-terminal domains
Journal: J Mol Biol
Volume: 404
Issue: 4
Pages: 680-96
Epub Date: 2010/10/19
Date: Dec 10
Short Title: Crystal structures of the glutamate receptor ion channel GluK3 and GluK5 amino-terminal domains
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2010.10.006
PMCID: PMC2991425
NIHMSID: NIHMS246654
Accession Number: 20951142
Keywords: Animals
Crystallography, X-Ray
Ion Channels/*chemistry
Models, Molecular
Protein Multimerization
Protein Structure, Quaternary
Rats
Receptors, Kainic Acid/*chemistry
Abstract: Ionotropic glutamate receptors (iGluRs) mediate the majority of fast excitatory synaptic neurotransmission in the central nervous system. The selective assembly of iGluRs into AMPA, kainate, and N-methyl-d-aspartic acid (NMDA) receptor subtypes is regulated by their extracellular amino-terminal domains (ATDs). Kainate receptors are further classified into low-affinity receptor families (GluK1-GluK3) and high-affinity receptor families (GluK4-GluK5) based on their affinity for the neurotoxin kainic acid. These two families share a 42% sequence identity for the intact receptor but only a 27% sequence identity at the level of ATD. We have determined for the first time the high-resolution crystal structures of GluK3 and GluK5 ATDs, both of which crystallize as dimers but with a strikingly different dimer assembly at the R1 interface. By contrast, for both GluK3 and GluK5, the R2 domain dimer assembly is similar to those reported previously for other non-NMDA iGluRs. This observation is consistent with the reports that GluK4-GluK5 cannot form functional homomeric ion channels and require obligate coassembly with GluK1-GluK3. Our analysis also reveals that the relative orientation of domains R1 and R2 in individual non-NMDA receptor ATDs varies by up to 10°, in contrast to the 50° difference reported for the NMDA receptor GluN2B subunit. This restricted domain movement in non-NMDA receptor ATDs seems to result both from extensive intramolecular contacts between domain R1 and domain R2 and from their assembly as dimers, which interact at both R1 and R2 domains. Our results provide the first insights into the structure and function of GluK4-GluK5, the least understood family of iGluRs.
Notes: 1089-8638
Kumar, Janesh
Mayer, Mark L
ZIA HD000707-25/Intramural NIH HHS/United States
Y1-GM-1104/GM/NIGMS NIH HHS/United States
Y1-CO-1020/CO/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, U.S. Gov't, Non-P.H.S.
J Mol Biol. 2010 Dec 10;404(4):680-96. doi: 10.1016/j.jmb.2010.10.006. Epub 2010 Oct 14.
Author Address: Laboratory of Cellular and Molecular Neurophysiology, Porter Neuroscience Research Center, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1416
Author: Kumar, J., Schuck, P. and Mayer, M. L.
Year: 2011
Title: Structure and assembly mechanism for heteromeric kainate receptors
Journal: Neuron
Volume: 71
Issue: 2
Pages: 319-31
Epub Date: 2011/07/28
Date: Jul 28
Short Title: Structure and assembly mechanism for heteromeric kainate receptors
Alternate Journal: Neuron
ISSN: 0896-6273 (Print)
0896-6273
DOI: 10.1016/j.neuron.2011.05.038
PMCID: PMC3145919
NIHMSID: NIHMS304430
Accession Number: 21791290
Keywords: Alanine/analogs & derivatives/pharmacology
Animals
Cell Line, Transformed
Chromatography, Gel/methods
Crystallography/methods
Crystallography, X-Ray
Excitatory Amino Acid Agonists/pharmacology
Glutamic Acid/pharmacology
Humans
Membrane Potentials/drug effects/genetics
Mice
Models, Molecular
Mutagenesis/genetics
Oocytes
Protein Structure, Tertiary/genetics/*physiology
Pyrimidines/pharmacology
Receptors, AMPA/genetics/metabolism
Receptors, Kainic Acid/*chemistry/classification/genetics/*physiology
Transfection
Xenopus
alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid/pharmacology
Abstract: Native glutamate receptor ion channels are tetrameric assemblies containing two or more different subunits. NMDA receptors are obligate heteromers formed by coassembly of two or three divergent gene families. While some AMPA and kainate receptors can form functional homomeric ion channels, the KA1 and KA2 subunits are obligate heteromers which function only in combination with GluR5-7. The mechanisms controlling glutamate receptor assembly involve an initial step in which the amino terminal domains (ATD) assemble as dimers. Here, we establish by sedimentation velocity that the ATDs of GluR6 and KA2 coassemble as a heterodimer of K(d) 11 nM, 32,000-fold lower than the K(d) for homodimer formation by KA2; we solve crystal structures for the GluR6/KA2 ATD heterodimer and heterotetramer assemblies. Using these structures as a guide, we perform a mutant cycle analysis to probe the energetics of assembly and show that high-affinity ATD interactions are required for biosynthesis of functional heteromeric receptors.
Notes: 1097-4199
Kumar, Janesh
Schuck, Peter
Mayer, Mark L
Z99 HD999999/Intramural NIH HHS/United States
ZIA HD000707-26/Intramural NIH HHS/United States
Y1-GM-1104/GM/NIGMS NIH HHS/United States
Y1-CO-1020/CO/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, U.S. Gov't, Non-P.H.S.
Neuron. 2011 Jul 28;71(2):319-31. doi: 10.1016/j.neuron.2011.05.038.
Author Address: Laboratory of Cellular and Molecular Neurophysiology, Porter Neuroscience Research Center, NICHD, NIH, DHHS, Bethesda, MD 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 998
Author: Kunicki, T. J., Cheli, Y., Moroi, M. and Furihata, K.
Year: 2005
Title: The influence of N-linked glycosylation on the function of platelet glycoprotein VI
Journal: Blood
Volume: 106
Issue: 8
Pages: 2744-9
Epub Date: 2005/07/15
Date: Oct 15
Short Title: The influence of N-linked glycosylation on the function of platelet glycoprotein VI
Alternate Journal: Blood
ISSN: 0006-4971 (Print)
0006-4971
DOI: 10.1182/blood-2005-04-1454
PMCID: PMC1895313
Accession Number: 16014566
Keywords: Animals
Blood Platelets/drug effects/*metabolism
Cell Adhesion/drug effects
Cell Line
Chlorocebus aethiops
Endopeptidases/metabolism
Glycosylation/drug effects
Humans
Models, Molecular
Mutation/genetics
Platelet Membrane Glycoproteins/chemistry/genetics/*metabolism
Protein Structure, Tertiary
Structural Homology, Protein
Transfection
Tunicamycin/pharmacology
Abstract: Using recombinant human glycoprotein VI (GPVI), we evaluated the effect of N-linked glycosylation at the consensus site Asparagine92-Glycine-Serine94 (N92GS94) on binding of this platelet-specific receptor to its ligands, human type I collagen, collagen-related peptide (CRP), and the snake venom C-type lectin convulxin (CVX). In COS-7 cells transiently transfected with GPVI, deglycosylation with peptide-N-glycosidase F (PNGase F; specific for complex N-linked glycans) or tunicamycin decreases the molecular weight of GPVI and reduces transfected COS-7 cell binding to both CRP and CVX. In stably transfected Dami cells, the substitutions N92A or S94A, but not L95H, resulted in a 30% to 40% decrease in adhesion to CVX, but a 90% or greater decrease in adhesion to CRP and a 65% to 70% decrease in adhesion to type I collagen. Treatment with PNGase F, but not Endoglycosidase H (Endo H) (specific for high-mannose N-linked glycans), produced an equivalent decrease in molecular weight. Neither N92A nor S94A affected the expression of GPVI, based on the direct binding of murine anti-human GPVI monoclonal antibody 204-11 to transfected Dami cells. These findings indicate that N-linked glycosylation at N92 in human GPVI is not required for surface expression, but contributes to maximal adhesion to type I collagen, CRP and, to a lesser extent, CVX.
Notes: 1528-0020
Kunicki, Thomas J
Cheli, Yann
Moroi, Masaaki
Furihata, Kenichi
R01 HL46979/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, P.H.S.
Blood. 2005 Oct 15;106(8):2744-9. doi: 10.1182/blood-2005-04-1454. Epub 2005 Jul 12.
Author Address: Division of Experimental Hemostasis and Thrombosis, Department of Molecular and Experimental Medicine, The Scripps Research Institute, 10550 North Torrey Pines Road, MEM-150, La Jolla, CA 92037, USA. tomk@scripps.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 776
Author: Kunishima, N., Shimada, Y., Tsuji, Y., Sato, T., Yamamoto, M., Kumasaka, T., Nakanishi, S., Jingami, H. and Morikawa, K.
Year: 2000
Title: Structural basis of glutamate recognition by a dimeric metabotropic glutamate receptor
Journal: Nature
Volume: 407
Issue: 6807
Pages: 971-7
Epub Date: 2000/11/09
Date: Oct 26
Short Title: Structural basis of glutamate recognition by a dimeric metabotropic glutamate receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/35039564
Accession Number: 11069170
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Dimerization
Glutamic Acid/chemistry/*metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Protein Structure, Tertiary
Rats
Receptors, Glutamate/chemistry/*metabolism
Abstract: The metabotropic glutamate receptors (mGluRs) are key receptors in the modulation of excitatory synaptic transmission in the central nervous system. Here we have determined three different crystal structures of the extracellular ligand-binding region of mGluR1--in a complex with glutamate and in two unliganded forms. They all showed disulphide-linked homodimers, whose 'active' and 'resting' conformations are modulated through the dimeric interface by a packed alpha-helical structure. The bi-lobed protomer architectures flexibly change their domain arrangements to form an 'open' or 'closed' conformation. The structures imply that glutamate binding stabilizes both the 'active' dimer and the 'closed' protomer in dynamic equilibrium. Movements of the four domains in the dimer are likely to affect the separation of the transmembrane and intracellular regions, and thereby activate the receptor. This scheme in the initial receptor activation could be applied generally to G-protein-coupled neurotransmitter receptors that possess extracellular ligand-binding sites.
Notes: Kunishima, N
Shimada, Y
Tsuji, Y
Sato, T
Yamamoto, M
Kumasaka, T
Nakanishi, S
Jingami, H
Morikawa, K
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2000 Oct 26;407(6807):971-7. doi: 10.1038/35039564.
Author Address: Department of Structural Biology, Biomolecular Engineering Research Institute, Suita, Osaka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 856
Author: Kunita, R., Otomo, A. and Ikeda, J. E.
Year: 2002
Title: Identification and characterization of novel members of the CREG family, putative secreted glycoproteins expressed specifically in brain
Journal: Genomics
Volume: 80
Issue: 5
Pages: 456-60
Epub Date: 2002/11/01
Date: Nov
Short Title: Identification and characterization of novel members of the CREG family, putative secreted glycoproteins expressed specifically in brain
Alternate Journal: Genomics
ISSN: 0888-7543 (Print)
0888-7543
Accession Number: 12408961
Keywords: Amino Acid Sequence
Animals
Base Sequence
Blotting, Northern
Brain/*metabolism
Cell Culture Techniques
Cell Differentiation/drug effects/*physiology
Expressed Sequence Tags
Glycoproteins/*genetics/isolation & purification/metabolism
Glycosylation
HeLa Cells
Humans
Mice
Molecular Sequence Data
*Multigene Family
Repressor Proteins/*genetics/isolation & purification
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: The cellular repressor of E1A-stimulated genes, CREG, is a secreted glycoprotein that enhances differentiation of pluripotent stem cells. Here we report two novel members of the CREG family, human CREG2 and mouse Creg2 cDNAs. The predicted human and mouse protein sequences exhibit 35% identity with CREG protein. Northern blot analyses demonstrate specific CREG2 and Creg2 transcription in brain-in the case of CREG2, mainly in the limbic system of the brain. Both mouse and human CREG2 fused to the carboxy terminus of EGFP in NIH3T3 cells localize to the perinuclear region, which demonstrates implicit endoplasmic reticulum and Golgi localization. Human and mouse CREG2 are N-glycosylated in HeLa cells and deletion of amino-terminal sequences completely abolishes N-glycosylation, indicating that the N termini of both proteins may function as signal sequences. Thus, human and mouse CREG2 are putative secreted glycoproteins and may be novel neuronal extracellular molecules.
Notes: Kunita, Ryota
Otomo, Asako
Ikeda, Joh-E
Journal Article
Research Support, Non-U.S. Gov't
United States
Genomics. 2002 Nov;80(5):456-60.
Author Address: SORST, Japan Science and Technology Corporation (JST), Tokai University School of Medicine, Isehara, Kanagawa, 259-1193, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1084
Author: Kurayoshi, M., Yamamoto, H., Izumi, S. and Kikuchi, A.
Year: 2007
Title: Post-translational palmitoylation and glycosylation of Wnt-5a are necessary for its signalling
Journal: Biochem J
Volume: 402
Issue: 3
Pages: 515-23
Epub Date: 2006/11/23
Date: Mar 15
Short Title: Post-translational palmitoylation and glycosylation of Wnt-5a are necessary for its signalling
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20061476
PMCID: PMC1863570
Accession Number: 17117926
Keywords: Amino Acid Sequence
Animals
Cell Line
Cell Movement
Fatty Acids, Monounsaturated/*metabolism
Glycosylation
Humans
Mice
Molecular Sequence Data
Protein Binding
*Protein Processing, Post-Translational
Proto-Oncogene Proteins/chemistry/genetics/isolation & purification/*metabolism
Sequence Alignment
*Signal Transduction
TCF Transcription Factors/metabolism
Transcription Factor 7-Like 2 Protein
Wnt Proteins/chemistry/genetics/isolation & purification/*metabolism
Wnt-5a Protein
Abstract: Wnt-5a is a representative ligand that activates a beta-catenin-independent pathway in Wnt signalling. In the present paper, the roles of the post-translational modifications in the actions of Wnt-5a were investigated. We found that Wnt-5a is modified with palmitate at Cys104 and glycans at Asn114, Asn120, Asn311 and Asn325. The palmitoylation was not essential for the secretion of Wnt-5a, but was necessary for its ability to suppress Wnt-3a-dependent T-cell factor transcriptional activity and to stimulate cell migration. Wnt-5a activated focal adhesion kinase and this activation also required palmitoylation. Wild-type Wnt-5a induced the internalization of Fz (Frizzled) 5, but a Wnt-5a mutant that lacks the palmitoylation site did not. Furthermore, the binding of Wnt-5a to the extracellular domain of Fz5 required palmitoylation of Wnt-5a. These results indicate that palmitoylation of Wnt-5a is important for the triggering of signalling at the cell surface level and, therefore, that the lipid-unmodified form of Wnt-5a cannot activate intracellular signal cascades. In contrast, glycosylation was necessary for the secretion of Wnt-5a, but not essential for the actions of Wnt-5a. Thus the post-translational palmitoylation and glycosylation of Wnt-5a are important for the actions and secretion of Wnt-5a.
Notes: 1470-8728
Kurayoshi, Manabu
Yamamoto, Hideki
Izumi, Shunsuke
Kikuchi, Akira
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2007 Mar 15;402(3):515-23. doi: 10.1042/BJ20061476.
Author Address: Department of Biochemistry, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima 734-8551, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 874
Author: Kuroda, Y., Nakata, M., Makino, A., Matsumoto, A., Ohashi, K., Itahashi, K., Takeuchi, F., Goto, M., Kojima, N. and Mizuochi, T.
Year: 2002
Title: Structural studies on IgG oligosaccharides of patients with primary Sjögren's syndrome
Journal: Glycoconj J
Volume: 19
Issue: 1
Pages: 23-31
Epub Date: 2003/03/26
Date: Jan
Short Title: Structural studies on IgG oligosaccharides of patients with primary Sjögren's syndrome
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1023/a:1022528829799
Accession Number: 12652077
Keywords: Adult
Aged
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Female
Galactose/metabolism
Glycosylation
Humans
Immunoglobulin G/*chemistry/isolation & purification
Middle Aged
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Sjogren's Syndrome/*metabolism
Abstract: Sjögren's syndrome (SS) is an autoimmune disease, and some patients have been found to have SS complicated with rheumatoid arthritis (RA), in which IgG is known to carry abnormal N-linked oligosaccharides. In order to investigate the relationship between SS and RA, the structures of N-linked oligosaccharides of IgG from 12 primary SS patients without RA, 9 RA patients, and 8 healthy individuals were analyzed using reversed-phase high-performance liquid chromatography, in combination with sequential exoglycosidase treatment and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. All of the IgG samples obtained from primary SS patients, RA patients, and healthy individuals contained the same series of biantennary complex-type oligosaccharides, but the ratio of each oligosaccharide differed among these 3 groups. The incidence of galactose-lacking N-linked oligosaccharides obtained from the IgG of RA patients was significantly higher than that from healthy individuals, but that from the serum IgG of primary SS patients varied among individuals. The patients with primary SS were classified into two groups based on the galactosylation levels of IgG oligosaccharides; one group exhibits galactosylation levels as low as those of RA patients and another exhibits levels similar to those of healthy individuals. Measurement of levels of rheumatoid factor (RF) revealed that primary SS patients with a high incidence of RF belonged to the low galactosylation group, as did RA patients. These results suggest that appearance of IgG carrying abnormal N-linked oligosaccharides in primary SS may be related to future complication with RA.
Notes: Kuroda, Yasuhiro
Nakata, Munehiro
Makino, Ayako
Matsumoto, Ayano
Ohashi, Kazuhiro
Itahashi, Koju
Takeuchi, Fujio
Goto, Makoto
Kojima, Naoya
Mizuochi, Tsuguo
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 2002 Jan;19(1):23-31. doi: 10.1023/a:1022528829799.
Author Address: Department of Applied Biochemistry, Institute of Glycotechnology, Tokai University, Hiratsuka, Kanagawa, 259-1292, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1321
Author: Kurogochi, M., Matsushista, T., Amano, M., Furukawa, J., Shinohara, Y., Aoshima, M. and Nishimura, S.
Year: 2010
Title: Sialic acid-focused quantitative mouse serum glycoproteomics by multiple reaction monitoring assay
Journal: Mol Cell Proteomics
Volume: 9
Issue: 11
Pages: 2354-68
Epub Date: 2010/06/24
Date: Nov
Short Title: Sialic acid-focused quantitative mouse serum glycoproteomics by multiple reaction monitoring assay
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M110.000430
PMCID: PMC2984228
Accession Number: 20571061
Keywords: Amino Acid Sequence
Animals
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Liquid/methods
Diabetes Mellitus/blood
Disease Models, Animal
Female
Glycopeptides/*chemistry
Glycosylation
Male
Mass Spectrometry/methods
Mice
Mice, Inbred C57BL
Molecular Sequence Data
N-Acetylneuraminic Acid/*analysis
Proteomics/*methods
Abstract: Despite increasing importance of protein glycosylation, most of the large-scale glycoproteomics have been limited to profiling the sites of N-glycosylation. However, in-depth knowledge of protein glycosylation to uncover functions and their clinical applications requires quantitative glycoproteomics eliciting both peptide and glycan sequences concurrently. Here we describe a novel strategy for the multiplexed quantitative mouse serum glycoproteomics based on a specific chemical ligation, namely, reverse glycoblotting technique, focusing sialic acids and multiple reaction monitoring (MRM). LC-MS/MS analysis of de-glycosylated peptides identified 270 mouse serum peptides (95 glycoproteins) as sialylated glycopeptides, of which 67 glycopeptides were fully characterized by MS/MS analyses in a straightforward manner. We revealed the importance of a fragment ion containing innermost N-acetylglucosamine (GlcNAc) residue as MRM transitions regardless the sequence of the peptides. Versatility of the reverse glycoblotting-assisted MRM assays was demonstrated by quantitative comparison of 25 targeted glycopeptides from 16 proteins between mice with homo and hetero types of diabetes disease model.
Notes: 1535-9484
Kurogochi, Masaki
Matsushista, Takahiko
Amano, Maho
Furukawa, Jun-ichi
Shinohara, Yasuro
Aoshima, Masato
Nishimura, Shin-Ichiro
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2010 Nov;9(11):2354-68. doi: 10.1074/mcp.M110.000430. Epub 2010 Jun 22.
Author Address: Graduate School of Life Science, Frontier Research Center for the Post-Genomic Science and Technology, Hokkaido University, Kita-ku, Sapporo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 629
Author: Kurumbail, R. G., Stevens, A. M., Gierse, J. K., McDonald, J. J., Stegeman, R. A., Pak, J. Y., Gildehaus, D., Miyashiro, J. M., Penning, T. D., Seibert, K., Isakson, P. C. and Stallings, W. C.
Year: 1996
Title: Structural basis for selective inhibition of cyclooxygenase-2 by anti-inflammatory agents
Journal: Nature
Volume: 384
Issue: 6610
Pages: 644-8
Epub Date: 1996/12/19
Date: Dec 19-26
Short Title: Structural basis for selective inhibition of cyclooxygenase-2 by anti-inflammatory agents
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/384644a0
Accession Number: 8967954
Keywords: Animals
Anti-Inflammatory Agents, Non-Steroidal/*pharmacology
Cell Line
Cloning, Molecular
Crystallography, X-Ray
Cyclooxygenase 2
Cyclooxygenase 2 Inhibitors
Cyclooxygenase Inhibitors/*pharmacology
Flurbiprofen/pharmacology
Indomethacin/pharmacology
Isoenzymes/*chemistry
Mice
Models, Molecular
Prostaglandin-Endoperoxide Synthases/*chemistry
Protein Conformation
Pyrazoles/pharmacology
Structure-Activity Relationship
Abstract: Prostaglandins and glucocorticoids are potent mediators of inflammation. Non-steroidal anti-inflammatory drugs (NSAIDs) exert their effects by inhibition of prostaglandin production. The pharmacological target of NSAIDs is cyclooxygenase (COX, also known as PGH synthase), which catalyses the first committed step in arachidonic-acid metabolism. Two isoforms of the membrane protein COX are known: COX-1, which is constitutively expressed in most tissues, is responsible for the physiological production of prostaglandins; and COX-2, which is induced by cytokines, mitogens and endotoxins in inflammatory cells, is responsible for the elevated production of prostaglandins during inflammation. The structure of ovine COX-1 complexed with several NSAIDs has been determined. Here we report the structures of unliganded murine COX-2 and complexes with flurbiprofen, indomethacin and SC-558, a selective COX-2 inhibitor, determined at 3.0 to 2.5 A resolution. These structures explain the structural basis for the selective inhibition of COX-2, and demonstrate some of the conformational changes associated with time-dependent inhibition.
Notes: Kurumbail, R G
Stevens, A M
Gierse, J K
McDonald, J J
Stegeman, R A
Pak, J Y
Gildehaus, D
Miyashiro, J M
Penning, T D
Seibert, K
Isakson, P C
Stallings, W C
Journal Article
England
Nature. 1996 Dec 19-26;384(6610):644-8. doi: 10.1038/384644a0.
Author Address: G.D. Searle, St Louis, Missouri 63198, USA. rgkuru@punchiri.monsanto.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 757
Author: Kustedjo, K., Bracey, M. H. and Cravatt, B. F.
Year: 2000
Title: Torsin A and its torsion dystonia-associated mutant forms are lumenal glycoproteins that exhibit distinct subcellular localizations
Journal: J Biol Chem
Volume: 275
Issue: 36
Pages: 27933-9
Epub Date: 2000/06/29
Date: Sep 8
Short Title: Torsin A and its torsion dystonia-associated mutant forms are lumenal glycoproteins that exhibit distinct subcellular localizations
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M910025199
Accession Number: 10871631
Keywords: Animals
COS Cells
Carrier Proteins/chemistry/genetics/*physiology
Cell Line
Cell Nucleus/*physiology/ultrastructure
Dystonia Musculorum Deformans/*genetics
Humans
*Molecular Chaperones
Mutagenesis, Site-Directed
Recombinant Proteins/analysis/chemistry/metabolism
Sequence Deletion
Transfection
Abstract: Early-onset torsion dystonia is an autosomal dominant hyperkinetic movement disorder that has recently been linked to a 3-base pair deletion in the DYT1 gene. The DYT1 gene encodes a 332-amino acid protein, torsin A, that bears low but significant homology to the Hsp100/Clp family of ATPase chaperones. The deletion in DYT1 associated with torsion dystonia results in the loss of one of a pair of glutamic acid residues residing near the C terminus of torsin A (DeltaE-torsin A). At present, little is known about the expression, subcellular distribution, and/or function of either the torsin A or DeltaE-torsin A protein. When transfected into mammalian cells, both torsin A and DeltaE-torsin A were found to behave as lumenally oriented glycoproteins. Immunofluorescence studies revealed that torsin A localized to a diffuse network of intracellular membranes displaying significant co-immunoreactivity for the endoplasmic reticulum resident protein BiP, whereas DeltaE-torsin A resided in large spheroid intracellular structures exclusive of BiP immunoreactivity. These results initially suggested that DeltaE-torsin A might exist as insoluble aggregates. However, both torsin A and DeltaE-torsin A were readily solubilized by nonionic detergents, were similarly accessible to proteases, and displayed equivalent migration patterns on sucrose gradients. Collectively, these data support that both the wild type and torsion dystonia-associated forms of torsin A are properly folded, lumenal proteins of similar oligomeric states. The potential relationship between the altered subcellular distribution of DeltaE-torsin A and the disease-inducing phenotype of the protein is discussed.
Notes: Kustedjo, K
Bracey, M H
Cravatt, B F
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2000 Sep 8;275(36):27933-9. doi: 10.1074/jbc.M910025199.
Author Address: Skaggs Institute for Chemical Biology and Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 685
Author: Küster, B., Hunter, A. P., Wheeler, S. F., Dwek, R. A. and Harvey, D. J.
Year: 1998
Title: Structural determination of N-linked carbohydrates by matrix-assisted laser desorption/ionization-mass spectrometry following enzymatic release within sodium dodecyl sulphate-polyacrylamide electrophoresis gels: application to species-specific glycosylation of alpha1-acid glycoprotein
Journal: Electrophoresis
Volume: 19
Issue: 11
Pages: 1950-9
Epub Date: 1998/09/18
Date: Aug
Short Title: Structural determination of N-linked carbohydrates by matrix-assisted laser desorption/ionization-mass spectrometry following enzymatic release within sodium dodecyl sulphate-polyacrylamide electrophoresis gels: application to species-specific glycosylation of alpha1-acid glycoprotein
Alternate Journal: Electrophoresis
ISSN: 0173-0835 (Print)
0173-0835
DOI: 10.1002/elps.1150191113
Accession Number: 9740055
Keywords: Amidohydrolases
Animals
Carbohydrate Conformation
Carbohydrates/*chemistry
Cattle
Dogs
*Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Orosomucoid/*chemistry
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Polysaccharides/*chemistry
Sheep
Sodium Dodecyl Sulfate
Species Specificity
*Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
Abstract: This paper describes a sensitive method for analysis of N-linked carbohydrates released enzymatically from within the gel following separation of glycoproteins (50-100 pmols) by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The separated bands containing the glycoproteins were cut from the gel, destained, reduced and alkylated. N-linked glycans were then released by in-gel incubation with peptide N-glycosidase-F (PNGase-F) and extracted with water and acetonitrile. Sialic acid-containing glycans were converted into methyl esters by reaction with methyl iodide, salts and reagents were removed by passage through a mixed-bed column of ion-exchange resins and the glycans were examined by matrix-assisted laser desorption/ionization (MALDI)-mass spectrometry. Structural determination of the released glycans was performed by exoglycosidase digestion. Following glycan release and extraction, the protein could be digested within the gel with trypsin, and the masses of the tryptic peptides could be compared with those generated from a sequence database for protein identification. The method is applied to the analysis of N-linked glycans from alpha1-acid glycoprotein from man, cow, sheep and dog. Major species-specific differences in glycosylation were found. Thus, although all four species used N-acetyl-neuraminic acid, only cow and sheep additionally used N-glycolyl-neuraminic acid. Biantennary glycans were the predominant carbohydrates in cow, sheep and dog but man produced more triantennary glycans and a substantial amount of tetraantennary sugars. Fucosylation was only found in glycans from man and cow and both cow and sheep glycans were found to have beta1-3- and well as beta1-4-linked galactose residues in the antennae.
Notes: Küster, B
Hunter, A P
Wheeler, S F
Dwek, R A
Harvey, D J
Journal Article
Research Support, Non-U.S. Gov't
Germany
Electrophoresis. 1998 Aug;19(11):1950-9. doi: 10.1002/elps.1150191113.
Author Address: Oxford Glycobiology Institute, Department of Biochemistry, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1214
Author: Kuzmanov, U., Jiang, N., Smith, C. R., Soosaipillai, A. and Diamandis, E. P.
Year: 2009
Title: Differential N-glycosylation of kallikrein 6 derived from ovarian cancer cells or the central nervous system
Journal: Mol Cell Proteomics
Volume: 8
Issue: 4
Pages: 791-8
Epub Date: 2008/12/18
Date: Apr
Short Title: Differential N-glycosylation of kallikrein 6 derived from ovarian cancer cells or the central nervous system
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M800516-MCP200
PMCID: PMC2667357
Accession Number: 19088065
Keywords: Amino Acid Sequence
Anions
Antibodies
Blotting, Western
Body Fluids/enzymology
Central Nervous System/*enzymology
Chromatography, Ion Exchange
Enzyme-Linked Immunosorbent Assay
Female
Glycopeptides/chemistry
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Kallikreins/chemistry/*metabolism
Molecular Sequence Data
Mutagenesis, Site-Directed
Ovarian Neoplasms/*enzymology/*pathology
Plant Lectins/metabolism
Ribosome Inactivating Proteins/metabolism
Tandem Mass Spectrometry
Abstract: Ovarian cancer causes more deaths than any other gynecological disorder. Perturbed glycosylation is one of the hallmarks of this malignancy. Kallikrein 6 (KLK6) elevation in serum is a diagnostic and prognostic indicator in ovarian cancer. The majority of ovarian carcinomas express high levels of KLK6, which diffuses into the circulation. Under physiological conditions, KLK6 is expressed highly in the central nervous system and found at high levels in cerebrospinal fluid from where it enters the circulation. Our aim was to characterize and compare the N-glycosylation status of this protein in ovarian cancer ascites fluid and cerebrospinal fluid. Anion-exchange chromatography was used to reveal different post-translational modifications on the two isoforms. Mobility gel shift Western blot analysis coupled with glycosidase digestion showed that the molecular weight difference between the two isoforms was because of differential glycosylation patterns. The presence of a single N-glycosylation site on KLK6 was confirmed by site-directed mutagenesis. Using a Sambucus nigra agglutinin-monoclonal antibody sandwich enzyme-linked immunosorbent assay approach, it was shown that ovarian cancer-derived KLK6 was modified with alpha2-6-linked sialic acid. The structure and composition of glycans of both KLK6 isoforms was elucidated by glycopeptide monitoring with electrospray ionization-Orbitrap tandem mass spectrometry. Therefore, the extensive and almost exclusive sialylation of KLK6 from ovarian cancer cells could lead to the development of an improved biomarker for the early diagnosis of ovarian carcinoma.
Notes: 1535-9484
Kuzmanov, Uros
Jiang, Nianxin
Smith, Christopher R
Soosaipillai, Antoninus
Diamandis, Eleftherios P
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2009 Apr;8(4):791-8. doi: 10.1074/mcp.M800516-MCP200. Epub 2008 Dec 16.
Author Address: Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto M5G 1X5, Ontario, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 929
Author: Kvansakul, M., Adams, J. C. and Hohenester, E.
Year: 2004
Title: Structure of a thrombospondin C-terminal fragment reveals a novel calcium core in the type 3 repeats
Journal: Embo j
Volume: 23
Issue: 6
Pages: 1223-33
Epub Date: 2004/03/12
Date: Mar 24
Short Title: Structure of a thrombospondin C-terminal fragment reveals a novel calcium core in the type 3 repeats
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7600166
PMCID: PMC381422
Accession Number: 15014436
Keywords: Amino Acid Sequence
Animals
Binding Sites
Calcium/*metabolism
Cell Adhesion
Cell Line
Crystallography, X-Ray
Humans
Mice
Models, Molecular
Molecular Sequence Data
Mutation/genetics
Oligopeptides/chemistry/metabolism
Peptide Fragments/chemistry/genetics/metabolism
Protein Structure, Tertiary
Rats
*Repetitive Sequences, Amino Acid
Sequence Alignment
Thrombospondin 1/*chemistry/genetics/*metabolism
Abstract: Thrombospondins (TSPs) are extracellular regulators of cell-matrix interactions and cell phenotype. The most highly conserved region of all TSPs are the calcium-binding type 3 (T3) repeats and the C-terminal globular domain (CTD). The crystal structure of a cell-binding TSP-1 fragment, spanning three T3 repeats and the CTD, reveals a compact assembly. The T3 repeats lack secondary structure and are organised around a core of calcium ions; two DxDxDGxxDxxD motifs per repeat each encapsulate two calcium ions in a novel arrangement. The CTD forms a lectin-like beta-sandwich and contains four strictly conserved calcium-binding sites. Disruption of the hairpin structure of T3 repeats 6 and 7 decreases protein secretion and stability. The availability for cell attachment of an RGD motif in T3 repeat 7 is modulated by calcium loading. The central architectural role of calcium explains how it is critical for the functions of the TSP C-terminal region. Mutations in the T3 repeats of TSP-5/COMP, which cause two human skeletal disorders, are predicted to disrupt the tertiary structure of the T3-CTD assembly.
Notes: 1460-2075
Kvansakul, Marc
Adams, Josephine C
Hohenester, Erhard
054334/Wellcome Trust/United Kingdom
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
EMBO J. 2004 Mar 24;23(6):1223-33. doi: 10.1038/sj.emboj.7600166. Epub 2004 Mar 11.
Author Address: Department of Biological Sciences, Imperial College London, South Kensington Campus, London, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1252
Author: Kwon, H. J., Abi-Mosleh, L., Wang, M. L., Deisenhofer, J., Goldstein, J. L., Brown, M. S. and Infante, R. E.
Year: 2009
Title: Structure of N-terminal domain of NPC1 reveals distinct subdomains for binding and transfer of cholesterol
Journal: Cell
Volume: 137
Issue: 7
Pages: 1213-24
Epub Date: 2009/07/01
Date: Jun 26
Short Title: Structure of N-terminal domain of NPC1 reveals distinct subdomains for binding and transfer of cholesterol
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2009.03.049
PMCID: PMC2739658
NIHMSID: NIHMS130089
Accession Number: 19563754
Keywords: Binding Sites
Carrier Proteins/*chemistry/*metabolism
Cholesterol/*metabolism
Crystallography, X-Ray
Humans
Intracellular Signaling Peptides and Proteins
Lysosomes/metabolism
Membrane Glycoproteins/*chemistry/*metabolism
Models, Molecular
Mutagenesis
Protein Structure, Tertiary
Abstract: LDL delivers cholesterol to lysosomes by receptor-mediated endocytosis. Exit of cholesterol from lysosomes requires two proteins, membrane-bound Niemann-Pick C1 (NPC1) and soluble NPC2. NPC2 binds cholesterol with its isooctyl side chain buried and its 3beta-hydroxyl exposed. Here, we describe high-resolution structures of the N-terminal domain (NTD) of NPC1 and complexes with cholesterol and 25-hydroxycholesterol. NPC1(NTD) binds cholesterol in an orientation opposite to NPC2: 3beta-hydroxyl buried and isooctyl side chain exposed. Cholesterol transfer from NPC2 to NPC1(NTD) requires reorientation of a helical subdomain in NPC1(NTD), enlarging the opening for cholesterol entry. NPC1 with point mutations in this subdomain (distinct from the binding subdomain) cannot accept cholesterol from NPC2 and cannot restore cholesterol exit from lysosomes in NPC1-deficient cells. We propose a working model wherein after lysosomal hydrolysis of LDL-cholesteryl esters, cholesterol binds NPC2, which transfers it to NPC1(NTD), reversing its orientation and allowing insertion of its isooctyl side chain into the outer lysosomal membranes.
Notes: 1097-4172
Kwon, Hyock Joo
Abi-Mosleh, Lina
Wang, Michael L
Deisenhofer, Johann
Goldstein, Joseph L
Brown, Michael S
Infante, Rodney E
P01 HL020948-32/HL/NHLBI NIH HHS/United States
P01 HL020948/HL/NHLBI NIH HHS/United States
T32 GM008014/GM/NIGMS NIH HHS/United States
HL20948/HL/NHLBI NIH HHS/United States
Howard Hughes Medical Institute/United States
GM08014/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Cell. 2009 Jun 26;137(7):1213-24. doi: 10.1016/j.cell.2009.03.049.
Author Address: Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX 75390-9046, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1391
Author: Kwon, H. J., Palnitkar, M. and Deisenhofer, J.
Year: 2011
Title: The structure of the NPC1L1 N-terminal domain in a closed conformation
Journal: PLoS One
Volume: 6
Issue: 4
Pages: e18722
Epub Date: 2011/04/29
Date: Apr 15
Short Title: The structure of the NPC1L1 N-terminal domain in a closed conformation
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0018722
PMCID: PMC3078110
Accession Number: 21525977
Keywords: Amino Acid Sequence
Cholesterol/metabolism
Crystallography, X-Ray
Humans
Membrane Glycoproteins/chemistry
Membrane Proteins/*chemistry/metabolism
Membrane Transport Proteins
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Tertiary
Structural Homology, Protein
Abstract: BACKGROUND: NPC1L1 is the molecular target of the cholesterol lowering drug Ezetimibe and mediates the intestinal absorption of cholesterol. Inhibition or deletion of NPC1L1 reduces intestinal cholesterol absorption, resulting in reduction of plasma cholesterol levels. PRINCIPAL FINDINGS: Here we present the 2.8 Å crystal structure of the N-terminal domain (NTD) of NPC1L1 in the absence of cholesterol. The structure, combined with biochemical data, reveals the mechanism of cholesterol selectivity of NPC1L1. Comparison to the cholesterol free and bound structures of NPC1(NTD) reveals that NPC1L1(NTD) is in a closed conformation and the sterol binding pocket is occluded from solvent. CONCLUSION: The structure of NPC1L1(NTD) reveals a degree of flexibility surrounding the entrance to the sterol binding pocket, suggesting a gating mechanism that relies on multiple movements around the entrance to the sterol binding pocket.
Notes: 1932-6203
Kwon, Hyock Joo
Palnitkar, Maya
Deisenhofer, Johann
Howard Hughes Medical Institute/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
PLoS One. 2011 Apr 15;6(4):e18722. doi: 10.1371/journal.pone.0018722.
Author Address: Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas, United States of America. hyockjoo.kwon@gmail.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2025
Author: Kwon, O. H., Cho, Y. Y., Kim, T. W. and Chung, S.
Year: 2019
Title: O-GlcNAcylation of Amyloid-β Protein Precursor by Insulin Signaling Reduces Amyloid-β Production
Journal: J Alzheimers Dis
Volume: 69
Issue: 4
Pages: 1195-1211
Epub Date: 2019/06/04
Short Title: O-GlcNAcylation of Amyloid-β Protein Precursor by Insulin Signaling Reduces Amyloid-β Production
Alternate Journal: Journal of Alzheimer's disease : JAD
ISSN: 1387-2877
DOI: 10.3233/jad-190060
Accession Number: 31156159
Keywords: Alzheimer Disease/etiology/metabolism
Amyloid beta-Peptides/*metabolism
Amyloid beta-Protein Precursor/*metabolism
Animals
Blotting, Western
Cell Line, Tumor
Diabetes Mellitus/metabolism
Humans
Immunoprecipitation
Insulin/*metabolism
Rats
Rats, Sprague-Dawley
Signal Transduction
beta-N-Acetylhexosaminidases/*metabolism
*Alzheimer’s disease
*O-GlcNAcylation
*amyloid-β
*amyloid-β protein precursor
*insulin
Abstract: Alzheimer's disease (AD) is caused by the accumulation of neurotoxic amyloid-β (Aβ) peptides. Aβ is derived from amyloid-β protein precursor (AβPP). In the non-amyloidogenic pathway, AβPP is cleaved by α-secretase and γ-secretase at the plasma membrane, excluding Aβ production. Alternatively, AβPP in the plasma membrane is internalized via endocytosis, and delivered to early endosomes and lysosomes, where it is cleaved by β-secretase and γ-secretase. Recent studies have shown that insulin in the periphery crosses the blood-brain barrier, and plays important roles in the brain. Furthermore, impaired insulin signaling has been linked to the progression of AD, and intranasal insulin administration improves memory impairments and cognition. However, the underlying molecular mechanisms of insulin treatment remain largely unknown. To investigate the effects of insulin on AβPP processing, we tested the effects of insulin on neuroblastoma SH-SY5Y cells overexpressing AβPP, and cultured rat cortical neurons. We found that insulin increased the level of cell surface AβPP, decreasing the endocytosis rate of AβPP. Insulin reduced Aβ generation through upregulation of AβPP O-GlcNAcylation via Akt insulin signaling. Our present data suggest that insulin affects Aβ production by regulating AβPP processing through AβPP O-GlcNAcylation. These results provide mechanistic insight into the beneficial effects of insulin, and a possible link between insulin deficient diabetes and cerebral amyloidosis in the pathogenesis of AD.
Notes: 1875-8908
Kwon, Oh Hoon
Cho, Yoon Young
Kim, Tae-Wan
Chung, Sungkwon
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Alzheimers Dis. 2019;69(4):1195-1211. doi: 10.3233/JAD-190060.
Author Address: Department of Physiology, Sungkyunkwan University School of Medicine, Suwon, South Korea.
Department of Pathology and Cell Biology, and Taub Institute for Research on Alzheimer's Disease and the Aging Brain, Columbia University Medical Center, New York, NY, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 671
Author: Lacko, A. G., Reason, A. J., Nuckolls, C., Kudchodkar, B. J., Nair, M. P., Sundarrajan, G., Pritchard, P. H., Morris, H. R. and Dell, A.
Year: 1998
Title: Characterization of recombinant human plasma lecithin: cholesterol acyltransferase (LCAT): N-linked carbohydrate structures and catalytic properties
Journal: J Lipid Res
Volume: 39
Issue: 4
Pages: 807-20
Epub Date: 1998/04/29
Date: Apr
Short Title: Characterization of recombinant human plasma lecithin: cholesterol acyltransferase (LCAT): N-linked carbohydrate structures and catalytic properties
Alternate Journal: Journal of lipid research
ISSN: 0022-2275 (Print)
0022-2275
Accession Number: 9555945
Keywords: Apolipoprotein A-I/metabolism
Carbohydrate Sequence
Glycoproteins/*chemistry/genetics
Humans
Molecular Sequence Data
Phosphatidylcholine-Sterol O-Acyltransferase/*chemistry/genetics
Phospholipases/metabolism
Recombinant Proteins/chemistry
Sequence Analysis
Spectrometry, Mass, Fast Atom Bombardment
Abstract: The major N-linked carbohydrate structures were determined for recombinant human plasma lecithin:cholesterol acyltransferase (LCAT). The analysis of the structure of oligosaccharides by fast atom bombardment mass spectrometry (FAB-MS) and linkage analysis was preceded by reduction and carboxymethylation of the intact glycoproteins and digestion with trypsin and proline specific endopeptidase. The N-glycans were subsequently released from the glycopeptides by PNGase F digestion and the oligosaccharides were separated using a C18 Sep-pak cartridge. The data from the combination of FAB spectrometry and linkage analysis show that the N-linked glycans present on recombinant LCAT (rLCAT) were composed primarily of triantennary and tetraantennary structures with and without core fucosylation. A minor population of glycans (less than 5%) contained up to three repeats of N-acetyllactosamine in one or more antennae. The LCAT activities of both recombinant and circulating forms of plasma LCAT were determined using low molecular weight and lipoprotein substrates. The catalytic behavior of these two enzyme forms were found to be very similar if not identical. These findings validate the concept that the recombinant enzyme can serve as an appropriate model for structure/function studies of LCAT and provide the foundation for subsequent structural studies.
Notes: Lacko, A G
Reason, A J
Nuckolls, C
Kudchodkar, B J
Nair, M P
Sundarrajan, G
Pritchard, P H
Morris, H R
Dell, A
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
J Lipid Res. 1998 Apr;39(4):807-20.
Author Address: Department of Biochemistry and Molecular Biology, University of North Texas Health Science Center, Fort Worth 76107, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 428
Author: Lai, P. H., Everett, R., Wang, F. F., Arakawa, T. and Goldwasser, E.
Year: 1986
Title: Structural characterization of human erythropoietin
Journal: J Biol Chem
Volume: 261
Issue: 7
Pages: 3116-21
Epub Date: 1986/03/05
Date: Mar 5
Short Title: Structural characterization of human erythropoietin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3949763
Keywords: Amino Acid Sequence
Binding Sites
Carbohydrates/analysis
Circular Dichroism
Disulfides/analysis
Erythropoietin/*analysis
Humans
Protein Conformation
Spectrophotometry, Ultraviolet
Abstract: Erythropoietin is the primary regulator of red blood cell formation in mammals. Because of its extreme scarcity, very little information is available regarding structural features of this important glycoprotein. We report here the primary structure of human urinary erythropoietin, determined by protein sequencing. In addition, the sites of glycosylation, assignment of disulfide bonds, and the circular dichroism of the hormone analyzed for secondary structure in comparison with the prediction from the sequence are presented.
Notes: Lai, P H
Everett, R
Wang, F F
Arakawa, T
Goldwasser, E
HL 21676/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1986 Mar 5;261(7):3116-21.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1866
Author: Lai, X., Wichers, H. J., Soler-Lopez, M. and Dijkstra, B. W.
Year: 2017
Title: Structure of Human Tyrosinase Related Protein 1 Reveals a Binuclear Zinc Active Site Important for Melanogenesis
Journal: Angew Chem Int Ed Engl
Volume: 56
Issue: 33
Pages: 9812-9815
Epub Date: 2017/07/01
Date: Aug 7
Short Title: Structure of Human Tyrosinase Related Protein 1 Reveals a Binuclear Zinc Active Site Important for Melanogenesis
Alternate Journal: Angewandte Chemie (International ed. in English)
ISSN: 1433-7851 (Print)
1433-7851
DOI: 10.1002/anie.201704616
PMCID: PMC5601231
Accession Number: 28661582
Keywords: Binding Sites
Humans
Membrane Glycoproteins/chemistry/genetics/*metabolism
Models, Molecular
Mutation
Oxidoreductases/chemistry/genetics/*metabolism
Protein Conformation
Zinc/chemistry/*metabolism
*albinism
*human tyrosinase
*melanogenesis
*structure elucidation
*zinc-copper enzymes
Abstract: Tyrosinase-related protein 1 (TYRP1) is one of three tyrosinase-like glycoenzymes in human melanocytes that are key to the production of melanin, the compound responsible for the pigmentation of skin, eye, and hair. Difficulties with producing these enzymes in pure form have hampered the understanding of their activity and the effect of mutations that cause albinism and pigmentation disorders. Herein we show that the typical tyrosinase-like subdomain of TYRP1 contains two zinc ions in the active site instead of copper ions as found in tyrosinases, which explains why TYRP1 does not exhibit tyrosinase redox activity. In addition, the structures reveal for the first time that the Cys-rich subdomain, which is unique to vertebrate melanogenic proteins, has an epidermal growth factor-like fold and is tightly associated with the tyrosinase subdomain. Our structures suggest that most albinism-related mutations of TYRP1 affect its stability or activity.
Notes: 1521-3773
Lai, Xuelei
Orcid: 0000-0003-4989-2444
Wichers, Harry J
Soler-Lopez, Montserrat
Orcid: 0000-0002-5924-4734
Dijkstra, Bauke W
Orcid: 0000-0001-9731-6586
Journal Article
Research Support, Non-U.S. Gov't
Angew Chem Int Ed Engl. 2017 Aug 7;56(33):9812-9815. doi: 10.1002/anie.201704616. Epub 2017 Jul 17.
Author Address: Laboratory of Biophysical Chemistry, University of Groningen, Nijenborgh 7, 9747 AG, Groningen, The Netherlands.
Structural Biology Group, European Synchrotron Radiation Facility, 71 Avenue des Martyrs, 38000, Grenoble, France.
Wageningen Food & Biobased Research, Bornse Weilanden 9, 6708 WG, Wageningen, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 303
Author: Laine, A., Hachulla, E., Strecker, G., Michalski, J. C. and Wieruszeski, J. M.
Year: 1991
Title: Structure determination of the glycans of human-serum alpha 1-antichymotrypsin using 1H-NMR spectroscopy and deglycosylation by N-glycanase
Journal: Eur J Biochem
Volume: 197
Issue: 1
Pages: 209-15
Epub Date: 1991/04/10
Date: Apr 10
Short Title: Structure determination of the glycans of human-serum alpha 1-antichymotrypsin using 1H-NMR spectroscopy and deglycosylation by N-glycanase
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1991.tb15901.x
Accession Number: 2015821
Keywords: Amidohydrolases
Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Humans
Hydrogen
Immunoelectrophoresis, Two-Dimensional
Magnetic Resonance Spectroscopy/methods
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Polysaccharides/*chemistry/isolation & purification
alpha 1-Antichymotrypsin/*chemistry/isolation & purification
alpha 1-Antitrypsin/chemistry/isolation & purification
Abstract: alpha 1-Antichymotrypsin purified from normal human serum was separated by affinity chromatography into th ree microheterogeneous forms on a concanavalin-A-Sepharose column: a pass-through (peak 1), a retarded (peak 2) and a bound form (peaks 3 + 4). For each form the asparagine-linked carbohydrate chains were liberated as oligosaccharides by hydrazinolysis, submitted to reduction with NaBH4 after re-N-acetylation and further separated by affinity chromatography on a concanavalin-A-Sepharose column. The complete primary structure of the glycans was determined by high-resolution 1H-NMR spectroscopy. The results indicated the presence of disialyl diantennary and of trisialyl triantennary type glycanic structures, the latter being accompanied by traces of disialylated triantennary oligosaccharide. The N-glycanase was used for the deglycosylation of the unfractionated alpha 1-antichymotrypsin; the successive removal of the N-linked complex-type oligosaccharide side chains of alpha 1-antichymotrypsin was studied in the presence of detergents. From these experiments it is concluded that alpha 1-antichymotrypsin carries four oligosaccharide side chains. Moreover our results show that the peak 1 contains four triantennary glycans, the peak 2 three triantennary and one diantennary glycans while the bound peaks 3 + 4 possess, on average, about one triantennary and three diantennary glycans per molecule. Since we showed that the peak 4 contains mostly diantennary glycans, it can be deduced that in peak 3 there are molecules carrying two triantennary and two diantennary glycans and others carrying one triantennary and three diantennary glycans.
Notes: Laine, A
Hachulla, E
Strecker, G
Michalski, J C
Wieruszeski, J M
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1991 Apr 10;197(1):209-15. doi: 10.1111/j.1432-1033.1991.tb15901.x.
Author Address: Unité 16 INSERM, Lille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1400
Author: Lakner, J., Seyer, C., Hermsdorf, T. and Schöneberg, T.
Year: 2011
Title: Characterization of the expression, promoter activity and molecular architecture of fibin
Journal: BMC Biochem
Volume: 12
Pages: 26
Epub Date: 2011/05/28
Date: May 26
Short Title: Characterization of the expression, promoter activity and molecular architecture of fibin
Alternate Journal: BMC biochemistry
ISSN: 1471-2091
DOI: 10.1186/1471-2091-12-26
PMCID: PMC3115872
Accession Number: 21615908
Keywords: Amino Acid Sequence
Animals
Conserved Sequence
Evolution, Molecular
*Gene Expression Regulation
Glycoproteins/chemistry/*genetics/*metabolism
HEK293 Cells
Humans
Intracellular Space/metabolism
Mice
Molecular Sequence Data
Promoter Regions, Genetic/*genetics
Protein Processing, Post-Translational
Protein Refolding
Protein Transport
Abstract: BACKGROUND: Fibin was initially discovered as a secreted signal molecule essential for pectoral fin bud initiation in zebrafish. Currently, there is little information about the molecular architecture and biological relevance of fibin in humans and other mammals. RESULTS: Fibin is expressed in cerebellum, skeletal muscle and many other embryonic and adult mouse tissues suggesting not only a role during embryonic development but also in adult functions. A 2.5-kbp genomic sequence fragment upstream of the coding sequence is sufficient to drive and regulate fibin expression through stimulation by glucocorticoids, activators of the protein kinase C signalling pathways and manganese ions. Fibin is an evolutionarily conserved protein, carries a cleavable signal peptide (amino acids 1-18) and is glycosylated at Asn30. The two conserved cysteines participate in intermolecular disulfide bond and multimer formation. Although fibin displays all features of a secretory protein, it is mostly retained in the endoplasmic reticulum when heterologously expressed. CONCLUSION: Fibin is functionally relevant during embryogenesis and adult life. Its expression is regulated by a number of cellular signalling pathways and the protein is routed via the secretory pathway. However, proper secretion presumably requires an unknown covalently-linked or associated co-factor.
Notes: 1471-2091
Lakner, Johannes
Seyer, Christian
Hermsdorf, Thomas
Schöneberg, Torsten
Journal Article
Research Support, Non-U.S. Gov't
BMC Biochem. 2011 May 26;12:26. doi: 10.1186/1471-2091-12-26.
Author Address: Molecular Biochemistry, Institute of Biochemistry, Medical Faculty, University of Leipzig, Johannisallee 30, Leipzig, 04103, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1226
Author: Lakomek, K., Dickmanns, A., Mueller, U., Kollmann, K., Deuschl, F., Berndt, A., Lübke, T. and Ficner, R.
Year: 2009
Title: De novo sulfur SAD phasing of the lysosomal 66.3 kDa protein from mouse
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 65
Issue: Pt 3
Pages: 220-8
Epub Date: 2009/02/25
Date: Mar
Short Title: De novo sulfur SAD phasing of the lysosomal 66.3 kDa protein from mouse
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449
DOI: 10.1107/s0907444908041814
Accession Number: 19237744
Keywords: Animals
Crystallization
Crystallography, X-Ray/*methods/statistics & numerical data
Glycoproteins/*chemistry
Glycosylation
Lysosomes/*enzymology
Mannosephosphates/chemistry
Mice
Models, Molecular
Protein Conformation
Protein Processing, Post-Translational
Scattering, Radiation
Sulfur/chemistry
Xenon/chemistry
Abstract: The 66.3 kDa protein from mouse is a soluble protein of the lysosomal matrix. It is synthesized as a glycosylated 75 kDa preproprotein which is further processed into 28 and 40 kDa fragments. Despite bioinformatics approaches and molecular characterization of the 66.3 kDa protein, the mode of its maturation as well as its physiological function remained unknown. Therefore, it was decided to tackle this question by means of X-ray crystallography. After expression in a human fibrosarcoma cell line, the C-terminally His-tagged single-chain 66.3 kDa variant and the double-chain form consisting of a 28 kDa fragment and a 40 kDa fragment were purified to homogeneity but could not be separated during the purification procedure. This mixture was therefore used for crystallization. Single crystals were obtained and the structure of the 66.3 kDa protein was solved by means of sulfur SAD phasing using data collected at a wavelength of 1.9 A on the BESSY beamline BL14.2 of Freie Universität Berlin. Based on the anomalous signal, a 22-atom substructure comprising 21 intrinsic S atoms and one Xe atom with very low occupancy was found and refined at a resolution of 2.4 A using the programs SHELXC/D and SHARP. Density modification using SOLOMON and DM resulted in a high-quality electron-density map, enabling automatic model building with ARP/wARP. The initial model contained 85% of the amino-acid residues expected to be present in the asymmetric unit of the crystal. Subsequently, the model was completed and refined to an R(free) factor of 19.8%. The contribution of the single Xe atom to the anomalous signal was analyzed in comparison to that of the S atoms and was found to be negligible. This work should encourage the use of the weak anomalous scattering of intrinsic S atoms in SAD phasing, especially for proteins, which require both expensive and time-consuming expression and purification procedures, preventing extensive screening of heavy-atom crystal soaks.
Notes: 1399-0047
Lakomek, Kristina
Dickmanns, Achim
Mueller, Uwe
Kollmann, Katrin
Deuschl, Florian
Berndt, Annette
Lübke, Torben
Ficner, Ralf
Journal Article
Research Support, Non-U.S. Gov't
United States
Acta Crystallogr D Biol Crystallogr. 2009 Mar;65(Pt 3):220-8. doi: 10.1107/S0907444908041814. Epub 2009 Feb 20.
Author Address: Department of Molecular Structural Biology, Institute of Microbiology and Genetics, Georg-August University Göttingen, Göttingen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 457
Author: Lalanne, J. L., Delarbre, C., Gachelin, G. and Kourilsky, P.
Year: 1983
Title: A cDNA clone containing the entire coding sequence of a mouse H-2Kd histocompatibility antigen
Journal: Nucleic Acids Res
Volume: 11
Issue: 5
Pages: 1567-77
Epub Date: 1983/03/11
Date: Mar 11
Short Title: A cDNA clone containing the entire coding sequence of a mouse H-2Kd histocompatibility antigen
Alternate Journal: Nucleic acids research
ISSN: 0305-1048 (Print)
0305-1048
DOI: 10.1093/nar/11.5.1567
PMCID: PMC325816
Accession Number: 6298749
Keywords: Animals
Base Composition
*Cloning, Molecular
DNA/*metabolism
DNA Restriction Enzymes
DNA Transposable Elements
H-2 Antigens/*genetics
*Major Histocompatibility Complex
Mice
Plasmids
Abstract: We have isolated a cDNA clone carrying a 1560 bp long insert which contains the entire coding and 3' untranslated regions of an H-2K(d) mouse histocompatibility antigen. Its sequence and overal features are described. They point to the existence of unique properties of DNA sequences associated with the H-2K(d) antigen.
Notes: 1362-4962
Lalanne, J L
Delarbre, C
Gachelin, G
Kourilsky, P
Journal Article
Research Support, Non-U.S. Gov't
Nucleic Acids Res. 1983 Mar 11;11(5):1567-77. doi: 10.1093/nar/11.5.1567.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1475
Author: Lang, C. M., Fellerer, K., Schwenk, B. M., Kuhn, P. H., Kremmer, E., Edbauer, D., Capell, A. and Haass, C.
Year: 2012
Title: Membrane orientation and subcellular localization of transmembrane protein 106B (TMEM106B), a major risk factor for frontotemporal lobar degeneration
Journal: J Biol Chem
Volume: 287
Issue: 23
Pages: 19355-65
Epub Date: 2012/04/19
Date: Jun 1
Short Title: Membrane orientation and subcellular localization of transmembrane protein 106B (TMEM106B), a major risk factor for frontotemporal lobar degeneration
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.365098
PMCID: PMC3365973
Accession Number: 22511793
Keywords: Cell Membrane/genetics/*metabolism/pathology
Endosomes/genetics/*metabolism/pathology
Frontotemporal Lobar Degeneration/genetics/*metabolism/pathology
Genome-Wide Association Study
Glycosylation
HEK293 Cells
HeLa Cells
Humans
Lysosomes/genetics/*metabolism/pathology
Membrane Proteins/genetics/*metabolism
Nerve Tissue Proteins/genetics/*metabolism
*Polymorphism, Single Nucleotide
Protein Transport/genetics
Risk Factors
Vacuolar Proton-Translocating ATPases/genetics/metabolism
Abstract: TMEM106B was identified as a major risk factor in a genome-wide association study for frontotemporal lobar degeneration (FTLD) with TAR DNA-binding protein (TDP)-43 pathology. The most significant association of TMEM106B single nucleotide polymorphisms with risk of FTLD-TDP was observed in patients with progranulin (GRN) mutations. Subsequent studies suggested an inverse correlation between TMEM106B expression and GRN levels in patient serum. However, in this study, this was not confirmed as we failed to detect a significant alteration of GRN levels upon knockdown or exogenous expression of TMEM106B in heterologous cells. To provide a basis for understanding TMEM106B function in health and disease, we investigated the membrane orientation and subcellular localization of this completely uncharacterized protein. By differential membrane extraction and sequential mutagenesis of potential N-glycosylation sites, we identified TMEM106B as a type 2 integral membrane protein with a highly glycosylated luminal domain. Glycosylation is partially required for the transport of TMEM106B beyond the endoplasmic reticulum to late cellular compartments. Endogenous as well as overexpressed TMEM106B localizes to late endosomes and lysosomes. Interestingly, the inhibition of vacuolar H(+)-ATPases significantly increased the levels of TMEM106B, a finding that may provide an unexpected biochemical link to GRN, because this protein is also strongly increased under the same conditions. Our findings provide a biochemical and cell biological basis for the understanding of the pathological role of TMEM106B in FTLD, an incurable neurodegenerative disorder.
Notes: 1083-351x
Lang, Christina M
Fellerer, Katrin
Schwenk, Benjamin M
Kuhn, Peer-Hendrik
Kremmer, Elisabeth
Edbauer, Dieter
Capell, Anja
Haass, Christian
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Jun 1;287(23):19355-65. doi: 10.1074/jbc.M112.365098. Epub 2012 Apr 17.
Author Address: Adolf-Butenandt-Institute, Biochemistry, Ludwig-Maximilians-University, 80336 Munich, Germany. christian.haass@dzne.lmu.de
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1885
Author: Larsen, I. S. B., Narimatsu, Y., Joshi, H. J., Siukstaite, L., Harrison, O. J., Brasch, J., Goodman, K. M., Hansen, L., Shapiro, L., Honig, B., Vakhrushev, S. Y., Clausen, H. and Halim, A.
Year: 2017
Title: Discovery of an O-mannosylation pathway selectively serving cadherins and protocadherins
Journal: Proc Natl Acad Sci U S A
Volume: 114
Issue: 42
Pages: 11163-11168
Epub Date: 2017/10/05
Date: Oct 17
Short Title: Discovery of an O-mannosylation pathway selectively serving cadherins and protocadherins
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1708319114
PMCID: PMC5651762
Accession Number: 28973932
Keywords: Cadherins/*metabolism
Carrier Proteins/*genetics/metabolism
Glycosylation
Glycosyltransferases/*genetics/metabolism
HEK293 Cells
Humans
Mannose/*metabolism
Membrane Proteins/*genetics/metabolism
Multigene Family
*O-glycosylation
*gene editing
*glycoproteomics
*glycosyltransferase
*mass spectrometry
Abstract: The cadherin (cdh) superfamily of adhesion molecules carry O-linked mannose (O-Man) glycans at highly conserved sites localized to specific β-strands of their extracellular cdh (EC) domains. These O-Man glycans do not appear to be elongated like O-Man glycans found on α-dystroglycan (α-DG), and we recently demonstrated that initiation of cdh/protocadherin (pcdh) O-Man glycosylation is not dependent on the evolutionary conserved POMT1/POMT2 enzymes that initiate O-Man glycosylation on α-DG. Here, we used a CRISPR/Cas9 genetic dissection strategy combined with sensitive and quantitative O-Man glycoproteomics to identify a homologous family of four putative protein O-mannosyltransferases encoded by the TMTC1-4 genes, which were found to be imperative for cdh and pcdh O-Man glycosylation. KO of all four TMTC genes in HEK293 cells resulted in specific loss of cdh and pcdh O-Man glycosylation, whereas combined KO of TMTC1 and TMTC3 resulted in selective loss of O-Man glycans on specific β-strands of EC domains, suggesting that each isoenzyme serves a different function. In addition, O-Man glycosylation of IPT/TIG domains of plexins and hepatocyte growth factor receptor was not affected in TMTC KO cells, suggesting the existence of yet another O-Man glycosylation machinery. Our study demonstrates that regulation of O-mannosylation in higher eukaryotes is more complex than envisioned, and the discovery of the functions of TMTCs provide insight into cobblestone lissencephaly caused by deficiency in TMTC3.
Notes: 1091-6490
Larsen, Ida Signe Bohse
Narimatsu, Yoshiki
Joshi, Hiren Jitendra
Orcid: 0000-0002-8192-2829
Siukstaite, Lina
Harrison, Oliver J
Brasch, Julia
Goodman, Kerry M
Hansen, Lars
Shapiro, Lawrence
Honig, Barry
Vakhrushev, Sergey Y
Clausen, Henrik
Halim, Adnan
Orcid: 0000-0003-1629-187x
R01 GM107571/GM/NIGMS NIH HHS/United States
R01 GM118584/GM/NIGMS NIH HHS/United States
R01 MH114817/MH/NIMH NIH HHS/United States
HHMI/Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2017 Oct 17;114(42):11163-11168. doi: 10.1073/pnas.1708319114. Epub 2017 Oct 2.
Author Address: Department of Cellular and Molecular Medicine, Faculty of Health Sciences, Copenhagen Center for Glycomics, University of Copenhagen, DK-2200 Copenhagen, Denmark.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032.
Zuckerman Mind Brain Behavior Institute, Columbia University, New York, NY 10032.
Department of Systems Biology, Columbia University, New York, NY 10032.
Howard Hughes Medical Institute, Columbia University, New York, NY 10032.
Department of Cellular and Molecular Medicine, Faculty of Health Sciences, Copenhagen Center for Glycomics, University of Copenhagen, DK-2200 Copenhagen, Denmark; halim@sund.ku.dk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1367
Author: Laursen, N. S., Andersen, K. R., Braren, I., Spillner, E., Sottrup-Jensen, L. and Andersen, G. R.
Year: 2011
Title: Substrate recognition by complement convertases revealed in the C5-cobra venom factor complex
Journal: Embo j
Volume: 30
Issue: 3
Pages: 606-16
Epub Date: 2011/01/11
Date: Feb 2
Short Title: Substrate recognition by complement convertases revealed in the C5-cobra venom factor complex
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2010.341
PMCID: PMC3034014
Accession Number: 21217642
Keywords: Complement C3-C5 Convertases/chemistry/*metabolism
Complement C5/chemistry/*metabolism
Crystallography
Elapid Venoms/chemistry/*metabolism
Exotoxins/chemistry/*metabolism
Humans
*Models, Molecular
Multiprotein Complexes/chemistry/*metabolism
*Protein Conformation
Abstract: Complement acts as a danger-sensing system in the innate immune system, and its activation initiates a strong inflammatory response and cleavage of the proteins C3 and C5 by proteolytic enzymes, the convertases. These contain a non-catalytic substrate contacting subunit (C3b or C4b) in complex with a protease subunit (Bb or C2a). We determined the crystal structures of the C3b homologue cobra venom factor (CVF) in complex with C5, and in complex with C5 and the inhibitor SSL7 at 4.3 Å resolution. The structures reveal a parallel two-point attachment between C5 and CVF, where the presence of SSL7 only slightly affects the C5-CVF interface, explaining the IgA dependence for SSL7-mediated inhibition of C5 cleavage. CVF functions as a relatively rigid binding scaffold inducing a conformational change in C5, which positions its cleavage site in proximity to the serine protease Bb. A general model for substrate recognition by the convertases is presented based on the C5-CVF and C3b-Bb-SCIN structures. Prior knowledge concerning interactions between the endogenous convertases and their substrates is rationalized by this model.
Notes: 1460-2075
Laursen, Nick S
Andersen, Kasper R
Braren, Ingke
Spillner, Edzard
Sottrup-Jensen, Lars
Andersen, Gregers R
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2011 Feb 2;30(3):606-16. doi: 10.1038/emboj.2010.341. Epub 2011 Jan 7.
Author Address: Department of Molecular Biology, Aarhus University, Aarhus, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1353
Author: Law, R. H., Lukoyanova, N., Voskoboinik, I., Caradoc-Davies, T. T., Baran, K., Dunstone, M. A., D'Angelo, M. E., Orlova, E. V., Coulibaly, F., Verschoor, S., Browne, K. A., Ciccone, A., Kuiper, M. J., Bird, P. I., Trapani, J. A., Saibil, H. R. and Whisstock, J. C.
Year: 2010
Title: The structural basis for membrane binding and pore formation by lymphocyte perforin
Journal: Nature
Volume: 468
Issue: 7322
Pages: 447-51
Epub Date: 2010/11/03
Date: Nov 18
Short Title: The structural basis for membrane binding and pore formation by lymphocyte perforin
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature09518
Accession Number: 21037563
Keywords: Animals
Cell Membrane/*metabolism
Cholesterol/metabolism
Cryoelectron Microscopy
Crystallography, X-Ray
Epidermal Growth Factor/chemistry
Granzymes/metabolism
Humans
Lymphocytes/*metabolism
Mice
Models, Molecular
Pore Forming Cytotoxic Proteins/*chemistry/genetics/*metabolism/ultrastructure
Protein Structure, Tertiary
Abstract: Natural killer cells and cytotoxic T lymphocytes accomplish the critically important function of killing virus-infected and neoplastic cells. They do this by releasing the pore-forming protein perforin and granzyme proteases from cytoplasmic granules into the cleft formed between the abutting killer and target cell membranes. Perforin, a 67-kilodalton multidomain protein, oligomerizes to form pores that deliver the pro-apoptopic granzymes into the cytosol of the target cell. The importance of perforin is highlighted by the fatal consequences of congenital perforin deficiency, with more than 50 different perforin mutations linked to familial haemophagocytic lymphohistiocytosis (type 2 FHL). Here we elucidate the mechanism of perforin pore formation by determining the X-ray crystal structure of monomeric murine perforin, together with a cryo-electron microscopy reconstruction of the entire perforin pore. Perforin is a thin 'key-shaped' molecule, comprising an amino-terminal membrane attack complex perforin-like (MACPF)/cholesterol dependent cytolysin (CDC) domain followed by an epidermal growth factor (EGF) domain that, together with the extreme carboxy-terminal sequence, forms a central shelf-like structure. A C-terminal C2 domain mediates initial, Ca(2+)-dependent membrane binding. Most unexpectedly, however, electron microscopy reveals that the orientation of the perforin MACPF domain in the pore is inside-out relative to the subunit arrangement in CDCs. These data reveal remarkable flexibility in the mechanism of action of the conserved MACPF/CDC fold and provide new insights into how related immune defence molecules such as complement proteins assemble into pores.
Notes: 1476-4687
Law, Ruby H P
Lukoyanova, Natalya
Voskoboinik, Ilia
Caradoc-Davies, Tom T
Baran, Katherine
Dunstone, Michelle A
D'Angelo, Michael E
Orlova, Elena V
Coulibaly, Fasséli
Verschoor, Sandra
Browne, Kylie A
Ciccone, Annette
Kuiper, Michael J
Bird, Phillip I
Trapani, Joseph A
Saibil, Helen R
Whisstock, James C
079605/Wellcome Trust/United Kingdom
BB/D008573/1/Biotechnology and Biological Sciences Research Council/United Kingdom
Arthritis Research UK/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2010 Nov 18;468(7322):447-51. doi: 10.1038/nature09518. Epub 2010 Oct 31.
Author Address: Department of Biochemistry and Molecular Biology, Monash University, Clayton, Melbourne, Victoria 3800, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 727
Author: Lawrence, C. M., Ray, S., Babyonyshev, M., Galluser, R., Borhani, D. W. and Harrison, S. C.
Year: 1999
Title: Crystal structure of the ectodomain of human transferrin receptor
Journal: Science
Volume: 286
Issue: 5440
Pages: 779-82
Epub Date: 1999/10/26
Date: Oct 22
Short Title: Crystal structure of the ectodomain of human transferrin receptor
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.286.5440.779
Accession Number: 10531064
Keywords: Amino Acid Sequence
Animals
CHO Cells
Carboxypeptidases/chemistry
Cell Membrane/chemistry
Conserved Sequence
Cricetinae
Crystallography, X-Ray
Dimerization
Ferric Compounds/metabolism
Glycosylation
Humans
Hydrogen-Ion Concentration
Models, Molecular
Molecular Sequence Data
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Transferrin/*chemistry/metabolism
Transferrin/metabolism
Abstract: The transferrin receptor (TfR) undergoes multiple rounds of clathrin-mediated endocytosis and reemergence at the cell surface, importing iron-loaded transferrin (Tf) and recycling apotransferrin after discharge of iron in the endosome. The crystal structure of the dimeric ectodomain of the human TfR, determined here to 3.2 angstroms resolution, reveals a three-domain subunit. One domain closely resembles carboxy- and aminopeptidases, and features of membrane glutamate carboxypeptidase can be deduced from the TfR structure. A model is proposed for Tf binding to the receptor.
Notes: Lawrence, C M
Ray, S
Babyonyshev, M
Galluser, R
Borhani, D W
Harrison, S C
Journal Article
Research Support, Non-U.S. Gov't
United States
Science. 1999 Oct 22;286(5440):779-82. doi: 10.1126/science.286.5440.779.
Author Address: Howard Hughes Medical Institute and Children's Hospital Laboratory of Molecular Medicine, 320 Longwood Avenue, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 582
Author: Lawrence, C. W., Little, P. A., Little, B. W., Glushka, J., van Halbeek, H. and Alhadeff, J. A.
Year: 1993
Title: Human non-secretory ribonucleases. II. Structural characterization of the N-glycans of the kidney, liver and spleen enzymes by NMR spectroscopy and electrospray mass spectrometry
Journal: Glycobiology
Volume: 3
Issue: 3
Pages: 249-59
Epub Date: 1993/06/01
Date: Jun
Short Title: Human non-secretory ribonucleases. II. Structural characterization of the N-glycans of the kidney, liver and spleen enzymes by NMR spectroscopy and electrospray mass spectrometry
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/3.3.249
Accession Number: 8358150
Keywords: Base Sequence
Carbohydrate Sequence
Carbohydrates/analysis
Electrophoresis, Polyacrylamide Gel
Glycoproteins/*chemistry/isolation & purification/metabolism
Humans
Kidney/*enzymology
Liver/*enzymology
Magnetic Resonance Spectroscopy/methods
Mass Spectrometry
Molecular Sequence Data
Molecular Weight
Oligosaccharides/*chemistry/isolation & purification
Polysaccharides/*chemistry/isolation & purification
Ribonucleases/*chemistry/isolation & purification/metabolism
Spleen/*enzymology
Abstract: The N-glycans have been removed by peptide-N-glycosidase F (PNGase F) from purified human non-secretory RNases derived from kidney, liver and spleen. The spleen RNase was purified by two procedures, one of which did not include the usual acid treatment step (0.25 M H2SO4, 45 min, 4 degrees C), to determine if acid treatment alters the carbohydrate moieties. The N-glycans of the RNases were fractionated by Bio-Gel P-4 chromatography and analysed by 600 MHz 1H-NMR spectroscopy and electrospray mass spectrometry. All four non-secretory RNase preparations contained the following structures: [formula: see text] The relative amounts of the trisaccharide, pentasaccharide and hexasaccharide appeared to vary slightly in the different tissue RNases. The overall results indicate: (i) that acid treatment during purification does not alter the N-glycans of non-secretory RNases; (ii) that the N-glycans from kidney, liver and spleen non-secretory RNases are very similar, if not identical, to one another, but different from the N-glycan structures reported for secretory RNase.
Notes: Lawrence, C W
Little, P A
Little, B W
Glushka, J
van Halbeek, H
Alhadeff, J A
GM 08351/GM/NIGMS NIH HHS/United States
P41-RR-05351/RR/NCRR NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1993 Jun;3(3):249-59. doi: 10.1093/glycob/3.3.249.
Author Address: Department of Chemistry, Lehigh University, Bethlehem, PA 18015.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 969
Author: Laxmikanthan, G., Blaber, S. I., Bernett, M. J., Scarisbrick, I. A., Juliano, M. A. and Blaber, M.
Year: 2005
Title: 1.70 A X-ray structure of human apo kallikrein 1: structural changes upon peptide inhibitor/substrate binding
Journal: Proteins
Volume: 58
Issue: 4
Pages: 802-14
Epub Date: 2005/01/15
Date: Mar 1
Short Title: 1.70 A X-ray structure of human apo kallikrein 1: structural changes upon peptide inhibitor/substrate binding
Alternate Journal: Proteins
ISSN: 0887-3585
DOI: 10.1002/prot.20368
Accession Number: 15651049
Keywords: Animals
Aprotinin/chemistry
Binding Sites
Biomarkers
Blood Pressure
Cattle
Cloning, Molecular
Crystallography, X-Ray/*methods
DNA Primers/chemistry
Electrophoresis, Polyacrylamide Gel
Histidine/chemistry
Humans
Hydrolysis
Inflammation
Ligands
Models, Molecular
Peptides/*chemistry
Protein Binding
Protein Conformation
Recombinant Proteins/chemistry
Serine/chemistry
Serine Endopeptidases/chemistry
Solvents/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Substrate Specificity
Swine
Tissue Kallikreins/antagonists & inhibitors/*chemistry
Abstract: Human kallikreins are serine proteases that comprise a recently identified large and closely related 15-member family. The kallikreins include both regulatory- and degradative-type proteases, impacting a variety of physiological processes including regulation of blood pressure, neuronal health, and the inflammatory response. While the function of the majority of the kallikreins remains to be elucidated, two members are useful biomarkers for prostate cancer and several others are potentially useful biomarkers for breast cancer, Alzheimer's, and Parkinson's disease. Human tissue kallikrein (human K1) is the best functionally characterized member of this family, and is known to play an important role in blood pressure regulation. As part of this function, human K1 exhibits unique dual-substrate specificity in hydrolyzing low molecular weight kininogen between both Arg-Ser and Met-Lys sequences. We report the X-ray crystal structure of mature, active recombinant human apo K1 at 1.70 A resolution. The active site exhibits structural features intermediate between that of apo and pro forms of known kallikrein structures. The S2 to S2' pockets demonstrate a variety of conformational changes in comparison to the porcine homolog of K1 in complex with peptide inhibitors, including the displacement of an extensive solvent network. These results indicate that the binding of a peptide substrate contributes to a structural rearrangement of the active-site Ser 195 resulting in a catalytically competent juxtaposition with the active-site His 57. The solvent networks within the S1 and S1' pockets suggest how the Arg-Ser and Met-Lys dual substrate specificity of human K1 is accommodated.
Notes: 1097-0134
Laxmikanthan, Gurunathan
Blaber, Sachiko I
Bernett, Matthew J
Scarisbrick, Isobel A
Juliano, Maria Aparecida
Blaber, Michael
Journal Article
Research Support, Non-U.S. Gov't
United States
Proteins. 2005 Mar 1;58(4):802-14. doi: 10.1002/prot.20368.
Author Address: Institute of Molecular Biophysics Florida State University, Tallahassee, Florida 32306-3015, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1026
Author: Lazarus, B. D., Love, D. C. and Hanover, J. A.
Year: 2006
Title: Recombinant O-GlcNAc transferase isoforms: identification of O-GlcNAcase, yes tyrosine kinase, and tau as isoform-specific substrates
Journal: Glycobiology
Volume: 16
Issue: 5
Pages: 415-21
Epub Date: 2006/01/26
Date: May
Short Title: Recombinant O-GlcNAc transferase isoforms: identification of O-GlcNAcase, yes tyrosine kinase, and tau as isoform-specific substrates
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwj078
Accession Number: 16434389
Keywords: Acetylglucosaminidase/*metabolism
Amino Acid Motifs
Amino Acid Sequence
Catalytic Domain
Escherichia coli/genetics
Glycogen Synthase Kinase 3
Glycosylation
Histone Acetyltransferases/*metabolism
Humans
Isoenzymes/chemistry/genetics/metabolism
Multienzyme Complexes/*metabolism
N-Acetylglucosaminyltransferases/*chemistry/genetics/*metabolism
Protein Structure, Tertiary
Protein-Serine-Threonine Kinases/*metabolism
Proto-Oncogene Proteins c-yes/*metabolism
Recombinant Proteins/chemistry/metabolism
Substrate Specificity
beta-N-Acetylhexosaminidases
Abstract: O-linked N-acetylglucosaminyltransferase (OGT) catalyzes the transfer of O-linked GlcNAc to serine or threonine residues of a variety of substrate proteins, including nuclear pore proteins, transcription factors, and proteins implicated in diabetes and neurodegenerative disorders. We have identified two nucleocytoplasmic isoforms of OGT (ncOGT and sOGT) and one isoform that localizes to the mitochondria (mOGT). These three isoforms contain identical catalytic regions but differ in the number of tetratricopeptide repeat motifs found at the N-terminus of each enzyme. We expressed each of these OGT isoforms in a soluble form in Escherichia coli and have used them to identify novel targets including the Src-family tyrosine kinase yes and O-GlcNAc-ase. We demonstrate that some substrate proteins, such as Nup62 and casein kinase II, are glycosylated by both ncOGT and mOGT, while others such as O-GlcNAcase and tau are specifically modified by ncOGT. The yes kinase was specifically modified by mOGT. The short isoform of OGT (sOGT) did not glycosylate any of the substrates tested, although it retains a potentially active catalytic domain. Our findings demonstrate the potential utility of recombinant OGT in identifying new targets and illustrate the necessity to examine all active isoforms of the enzyme. The identification of a tyrosine kinase and O-GlcNAcase as OGT targets suggests the potential for OGT participation in numerous signal transduction cascades.
Notes: Lazarus, Brooke D
Love, Dona C
Hanover, John A
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
England
Glycobiology. 2006 May;16(5):415-21. doi: 10.1093/glycob/cwj078. Epub 2006 Jan 23.
Author Address: Laboratory of Cell Biology and Biochemistry, NIDDK, National Institutes of Health, Bethesda, MD 20897-0851, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1615
Author: Lazarus, M. B., Jiang, J., Kapuria, V., Bhuiyan, T., Janetzko, J., Zandberg, W. F., Vocadlo, D. J., Herr, W. and Walker, S.
Year: 2013
Title: HCF-1 is cleaved in the active site of O-GlcNAc transferase
Journal: Science
Volume: 342
Issue: 6163
Pages: 1235-9
Epub Date: 2013/12/07
Date: Dec 6
Short Title: HCF-1 is cleaved in the active site of O-GlcNAc transferase
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1243990
PMCID: PMC3930058
NIHMSID: NIHMS542151
Accession Number: 24311690
Keywords: Amino Acid Motifs
Amino Acid Substitution
Catalytic Domain
Crystallography, X-Ray
Glycosylation
Host Cell Factor C1/*chemistry/*metabolism
Humans
Hydrogen Bonding
Models, Molecular
N-Acetylglucosaminyltransferases/*chemistry/*metabolism
Protein Conformation
Protein Structure, Tertiary
Proteolysis
Pyrrolidonecarboxylic Acid/metabolism
Recombinant Fusion Proteins/chemistry/metabolism
Uridine Diphosphate N-Acetylglucosamine/chemistry/metabolism
Abstract: Host cell factor-1 (HCF-1), a transcriptional co-regulator of human cell-cycle progression, undergoes proteolytic maturation in which any of six repeated sequences is cleaved by the nutrient-responsive glycosyltransferase, O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT). We report that the tetratricopeptide-repeat domain of O-GlcNAc transferase binds the carboxyl-terminal portion of an HCF-1 proteolytic repeat such that the cleavage region lies in the glycosyltransferase active site above uridine diphosphate-GlcNAc. The conformation is similar to that of a glycosylation-competent peptide substrate. Cleavage occurs between cysteine and glutamate residues and results in a pyroglutamate product. Conversion of the cleavage site glutamate into serine converts an HCF-1 proteolytic repeat into a glycosylation substrate. Thus, protein glycosylation and HCF-1 cleavage occur in the same active site.
Notes: 1095-9203
Lazarus, Michael B
Jiang, Jiaoyang
Kapuria, Vaibhav
Bhuiyan, Tanja
Janetzko, John
Zandberg, Wesley F
Vocadlo, David J
Herr, Winship
Walker, Suzanne
R01 GM094263/GM/NIGMS NIH HHS/United States
R01GM094263/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2013 Dec 6;342(6163):1235-9. doi: 10.1126/science.1243990.
Author Address: Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 371
Author: Lazure, C., Leduc, R., Seidah, N. G., Thibault, G., Genest, J. and Chrétien, M.
Year: 1987
Title: The complete amino acid sequence of rat submaxillary gland tonin does contain the aspartic acid at the active site: confirmation by protein sequence analysis
Journal: Biochem Cell Biol
Volume: 65
Issue: 4
Pages: 321-37
Epub Date: 1987/04/01
Date: Apr
Short Title: The complete amino acid sequence of rat submaxillary gland tonin does contain the aspartic acid at the active site: confirmation by protein sequence analysis
Alternate Journal: Biochemistry and cell biology = Biochimie et biologie cellulaire
ISSN: 0829-8211 (Print)
0829-8211
DOI: 10.1139/o87-042
Accession Number: 3038148
Keywords: Amino Acid Sequence
Animals
*Aspartic Acid
Binding Sites
Cyanogen Bromide
Indicators and Reagents
Peptide Fragments/analysis
*Peptidyl-Dipeptidase A/metabolism
Rats
Submandibular Gland/*enzymology
Abstract: The revised amino acid sequence of rat submaxillary gland tonin, a serine protease, does contain the active site Asp residue. The active site of this kallikrein-related enzyme is thus made up of the same catalytic triad (Asp, Ser, and His) found in all known serine proteases. The important Asp residue has now been localized in a 16 amino acid peptide previously reported as missing in the tonin sequence. The complete amino acid sequence thus contains 235 residues corresponding to a molecular weight of 25,658, more in agreement with previously reported molecular weights. Moreover, the revised structure led (a) to the assignment of Arg, Asn, and Val residues instead of His, Asp, and Gly at positions 63, 165, and 169, respectively; (b) to the assignment of residues occupying an overlapping sequence at positions 165-171, and finally (c) to the localization of two N-glycosylation sites at positions 82 and 165. These results further document the close relationship of tonin to the ever expanding kallikrein family.
Notes: Lazure, C
Leduc, R
Seidah, N G
Thibault, G
Genest, J
Chrétien, M
Journal Article
Research Support, Non-U.S. Gov't
Canada
Biochem Cell Biol. 1987 Apr;65(4):321-37. doi: 10.1139/o87-042.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2076
Author: Lee, H., Beilhartz, G. L., Kucharska, I., Raman, S., Cui, H., Lam, M. H. Y., Liang, H., Rubinstein, J. L., Schramek, D., Julien, J. P., Melnyk, R. A. and Taipale, M.
Year: 2020
Title: Recognition of Semaphorin Proteins by P. sordellii Lethal Toxin Reveals Principles of Receptor Specificity in Clostridial Toxins
Journal: Cell
Volume: 182
Issue: 2
Pages: 345-356.e16
Epub Date: 2020/06/27
Date: Jul 23
Short Title: Recognition of Semaphorin Proteins by P. sordellii Lethal Toxin Reveals Principles of Receptor Specificity in Clostridial Toxins
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2020.06.005
PMCID: PMC7316060
Accession Number: 32589945
Keywords: Animals
Bacterial Toxins/chemistry/*metabolism/toxicity
Binding Sites
CRISPR-Cas Systems/genetics
Cell Line
Clostridium sordellii/*metabolism
Cryoelectron Microscopy
Edema/pathology/prevention & control
Female
Humans
Lung/drug effects/pathology
Mice
Mice, Inbred C57BL
Molecular Dynamics Simulation
Mutagenesis, Site-Directed
Protein Binding
Protein Structure, Tertiary
Recombinant Proteins/biosynthesis/isolation & purification/therapeutic use
Semaphorins/chemistry/genetics/*metabolism
*CRISPR/Cas9 screening
*Clostridial toxins
*P. sordellii
*Sema6a
*Sema6b
*TcsL
*bacterial exotoxins
*cryo-EM
*semaphorin
Abstract: Pathogenic clostridial species secrete potent toxins that induce severe host tissue damage. Paeniclostridium sordellii lethal toxin (TcsL) causes an almost invariably lethal toxic shock syndrome associated with gynecological infections. TcsL is 87% similar to C. difficile TcdB, which enters host cells via Frizzled receptors in colon epithelium. However, P. sordellii infections target vascular endothelium, suggesting that TcsL exploits another receptor. Here, using CRISPR/Cas9 screening, we establish semaphorins SEMA6A and SEMA6B as TcsL receptors. We demonstrate that recombinant SEMA6A can protect mice from TcsL-induced edema. A 3.3 Å cryo-EM structure shows that TcsL binds SEMA6A with the same region that in TcdB binds structurally unrelated Frizzled. Remarkably, 15 mutations in this evolutionarily divergent surface are sufficient to switch binding specificity of TcsL to that of TcdB. Our findings establish semaphorins as physiologically relevant receptors for TcsL and reveal the molecular basis for the difference in tissue targeting and disease pathogenesis between highly related toxins.
Notes: 1097-4172
Lee, Hunsang
Beilhartz, Greg L
Kucharska, Iga
Raman, Swetha
Cui, Hong
Lam, Mandy Hiu Yi
Liang, Huazhu
Rubinstein, John L
Schramek, Daniel
Julien, Jean-Philippe
Melnyk, Roman A
Taipale, Mikko
RN292258 – 366017/CIHR/Canada
Journal Article
Research Support, Non-U.S. Gov't
Cell. 2020 Jul 23;182(2):345-356.e16. doi: 10.1016/j.cell.2020.06.005. Epub 2020 Jun 25.
Author Address: Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON M5S 3E1, Canada.
Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON M5G 0A4, Canada.
Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON M5G 0A4, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada.
Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON M5G 0A4, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada.
Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada.
Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON M5G 0A4, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada; Department of Immunology, University of Toronto, Toronto, ON M5S 1A8, Canada; Molecular Architecture of Life Program, Canadian Institute for Advanced Research (CIFAR), Toronto, ON M5G 1M1, Canada. Electronic address: jean-philippe.julien@sickkids.ca.
Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON M5G 0A4, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada. Electronic address: roman.melnyk@sickkids.ca.
Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada; Molecular Architecture of Life Program, Canadian Institute for Advanced Research (CIFAR), Toronto, ON M5G 1M1, Canada. Electronic address: mikko.taipale@utoronto.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2029
Author: Lee, H., Li, C., Zhang, Y., Zhang, D., Otterbein, L. E. and Jin, Y.
Year: 2019
Title: Caveolin-1 selectively regulates microRNA sorting into microvesicles after noxious stimuli
Journal: J Exp Med
Volume: 216
Issue: 9
Pages: 2202-2220
Epub Date: 2019/06/27
Date: Sep 2
Short Title: Caveolin-1 selectively regulates microRNA sorting into microvesicles after noxious stimuli
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.20182313
PMCID: PMC6719430
Accession Number: 31235510
Keywords: Acetylglucosamine/metabolism
Animals
Caveolin 1/chemistry/*metabolism
Cell Line
Cell-Derived Microparticles/*metabolism/ultrastructure
Epithelial Cells/metabolism
Gene Expression Regulation
Glycosylation
Heterogeneous-Nuclear Ribonucleoprotein Group A-B/metabolism
Humans
Inflammation/pathology
Macrophage Activation
Macrophages/metabolism
Mice, Inbred C57BL
MicroRNAs/genetics/*metabolism
Models, Biological
Oxidative Stress
Phosphorylation
Phosphotyrosine/metabolism
Protein Binding
Protein Domains
Abstract: Emerging evidence suggests that extracellular vesicle (EV)-containing miRNAs mediate intercellular communications in response to noxious stimuli. It remains unclear how a cell selectively sorts the cellular miRNAs into EVs. We report that caveolin-1 (cav-1) is essential for sorting of selected miRNAs into microvesicles (MVs), a main type of EVs generated by outward budding of the plasma membrane. We found that cav-1 tyrosine 14 (Y14)-phosphorylation leads to interactions between cav-1 and hnRNPA2B1, an RNA-binding protein. The cav-1/hnRNPA2B1 complex subsequently traffics together into MVs. Oxidative stress induces O-GlcNAcylation of hnRNPA2B1, resulting in a robustly altered hnRNPA2B1-bound miRNA repertoire. Notably, cav-1 pY14 also promotes hnRNPA2B1 O-GlcNAcylation. Functionally, macrophages serve as the principal recipient of epithelial MVs in the lung. MV-containing cav-1/hnRNPA2B1 complex-bound miR-17/93 activate tissue macrophages. Collectively, cav-1 is the first identified membranous protein that directly guides RNA-binding protein into EVs. Our work delineates a novel mechanism by which oxidative stress compels epithelial cells to package and secrete specific miRNAs and elicits an innate immune response.
Notes: 1540-9538
Lee, Heedoo
Li, Chunhua
Zhang, Yang
Zhang, Duo
Otterbein, Leo E
Orcid: 0000-0002-7900-0825
Jin, Yang
Orcid: 0000-0002-4779-531x
R01 GM083107/GM/NIGMS NIH HHS/United States
R01 GM116960/GM/NIGMS NIH HHS/United States
R01 GM127596/GM/NIGMS NIH HHS/United States
R33 AI121644/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Exp Med. 2019 Sep 2;216(9):2202-2220. doi: 10.1084/jem.20182313. Epub 2019 Jun 24.
Author Address: Division of Pulmonary and Critical Care Medicine, Department of Medicine, Boston University Medical Campus, Boston, MA.
Department of Computational Medicine and Bioinformatics Department of Biological Chemistry, The University of Michigan, Ann Arbor, MI.
Department of Surgery, Harvard Medical School, Beth Israel Deaconess Medical Center, Boston, MA.
Division of Pulmonary and Critical Care Medicine, Department of Medicine, Boston University Medical Campus, Boston, MA yjin1@bu.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2022
Author: Lee, H., Oh, Y., Jeon, Y. J., Lee, S. Y., Kim, H., Lee, H. J. and Jung, Y. K.
Year: 2019
Title: DR4-Ser424 O-GlcNAcylation Promotes Sensitization of TRAIL-Tolerant Persisters and TRAIL-Resistant Cancer Cells to Death
Journal: Cancer Res
Volume: 79
Issue: 11
Pages: 2839-2852
Epub Date: 2019/04/17
Date: Jun 1
Short Title: DR4-Ser424 O-GlcNAcylation Promotes Sensitization of TRAIL-Tolerant Persisters and TRAIL-Resistant Cancer Cells to Death
Alternate Journal: Cancer research
ISSN: 0008-5472
DOI: 10.1158/0008-5472.can-18-1991
Accession Number: 30987996
Keywords: Acetylglucosamine/metabolism
Cell Death/genetics
Cell Line, Tumor
*Drug Resistance, Neoplasm/drug effects/physiology
Endometrial Neoplasms/genetics
Female
Glucose/metabolism
Humans
Membrane Microdomains/metabolism
Mutation
N-Acetylglucosaminyltransferases/genetics
Receptors, TNF-Related Apoptosis-Inducing Ligand/genetics/*metabolism
Serine/*metabolism
Stomach Neoplasms/drug therapy/metabolism/pathology
TNF-Related Apoptosis-Inducing Ligand/metabolism/*pharmacology
Abstract: TNF-related apoptosis-inducing ligand (TRAIL) resistance, including nongenetically acquired tolerance in cancer persister cells, is a major obstacle to translating TRAIL therapy into patients with cancer. However, the underlying mechanisms remain to be elucidated. Here, we show that DR4/TRAIL-R1 is O-GlcNAcylated at Ser424 in its death domain to mediate both apoptosis and necrosis upon TRAIL ligation. We found that DR4-Ser424 mutations, identified from our cell-based functional screen using a cancer patient-derived cDNA expression library and from The Cancer Genome Atlas, caused TRAIL resistance in various human cancer cell lines. Using O-GlcNAc transferase knockdown cells, DR4-preferred versus DR5-preferred cancer cells, and a DR5-neutralizing antibody, we evaluated the essential role of DR4-specific O-GlcNAc modification in TRAIL cytotoxicity. In contrast to DR4, DR5 was not O-GlcNAcylated by TRAIL treatment, discriminating DR4 from DR5-mediated signaling. Apart from genetic changes in DR4-Ser424, we further classified various cancer cell lines originated from stomach, colon, lung, and glioblastoma according to their sensitivity to and receptor preference upon TRAIL death signaling and generated TRAIL-tolerant persister-derived DLD-1(PER) cells. Among these, we discovered that DR4 was not modified by O-GlcNAc in most of the TRAIL-resistant cancer cells and DLD-1(PER) cells. Interestingly, promoting DR4 O-GlcNAcylation intentionally using 2-deoxy-d-glucose or a high concentration of glucose sensitized those resistant cancer cells to TRAIL. The O-GlcNAcylation-defective DR4 failed to form DISC/necrosome and could not translocate to aggregated platforms for receptor clustering. Our findings demonstrate that DR4 O-GlcNAcylation is crucial for TRAIL death signaling, providing new opportunities for TRAIL therapy overcoming TRAIL resistance in cancers. SIGNIFICANCE: This study reports that a novel posttranslational modification by O-GlcNAcylation of one of the two human TRAIL receptors with a death domain, TRAIL-R1 (DR4), plays a crucial role in enabling both apoptotic and necroptotic cell death induction by TRAIL.
Notes: 1538-7445
Lee, Hyeonjeong
Orcid: 0000-0002-6134-5569
Oh, Yumin
Jeon, Young-Jun
Lee, Song-Yi
Kim, Hyunjoo
Lee, Ho-June
Jung, Yong-Keun
Journal Article
Research Support, Non-U.S. Gov't
United States
Cancer Res. 2019 Jun 1;79(11):2839-2852. doi: 10.1158/0008-5472.CAN-18-1991. Epub 2019 Apr 15.
Author Address: School of Biological Science, Seoul National University, Gwanak-gu, Seoul, Republic of Korea.
The Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University School of Medicine, Baltimore, Maryland.
Stanford Cancer Institute, Stanford University, Stanford, California.
Department of Discovery Oncology, Genentech, Inc., South San Francisco, California.
School of Biological Science, Seoul National University, Gwanak-gu, Seoul, Republic of Korea. ykjung@snu.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1781
Author: Lee, J. S. and Zhang, Z.
Year: 2016
Title: O-linked N-acetylglucosamine transferase (OGT) interacts with the histone chaperone HIRA complex and regulates nucleosome assembly and cellular senescence
Journal: Proc Natl Acad Sci U S A
Volume: 113
Issue: 23
Pages: E3213-20
Epub Date: 2016/05/25
Date: Jun 7
Short Title: O-linked N-acetylglucosamine transferase (OGT) interacts with the histone chaperone HIRA complex and regulates nucleosome assembly and cellular senescence
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1600509113
PMCID: PMC4988580
Accession Number: 27217568
Keywords: Adaptor Proteins, Signal Transducing/metabolism
Cell Cycle Proteins/genetics/*metabolism
Cellular Senescence/*physiology
HEK293 Cells
Histone Chaperones/genetics/*metabolism
Histones/*metabolism
Humans
N-Acetylglucosaminyltransferases/genetics/*metabolism
Nuclear Proteins/metabolism
Nucleosomes/metabolism
Protein Binding
Protein Domains
Transcription Factors/genetics/*metabolism
Hira
Ogt
cellular senescence
histone H3.3
nucleosome assembly
Abstract: The histone chaperone HIRA complex, consisting of histone cell cycle regulator (HIRA), Ubinuclein1 (UBN1), and calcineurin binding protein 1 (CABIN1), deposits histone variant H3.3 to genic regions and regulates gene expression in various cellular processes, including cellular senescence. How HIRA-mediated nucleosome assembly of H3.3-H4 is regulated remains not well understood. Here, we show that O-linked N-acetylglucosamine (GlcNAc) transferase (OGT), an enzyme that catalyzes O-GlcNAcylation of serine or threonine residues, interacts with UBN1, modifies HIRA, and promotes nucleosome assembly of H3.3. Depletion of OGT or expression of the HIRA S231A O-GlcNAcylation-deficient mutant compromises formation of the HIRA-H3.3 complex and H3.3 nucleosome assembly. Importantly, OGT depletion or expression of the HIRA S231A mutant delays premature cellular senescence in primary human fibroblasts, whereas overexpression of OGT accelerates senescence. Taken together, these results support a model in which OGT modifies HIRA to regulate HIRA-H3.3 complex formation and H3.3 nucleosome assembly and reveal the mechanism by which OGT functions in cellular senescence.
Notes: 1091-6490
Lee, Jong-Sun
Zhang, Zhiguo
R01 CA157489/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2016 Jun 7;113(23):E3213-20. doi: 10.1073/pnas.1600509113. Epub 2016 May 23.
Author Address: Mayo Graduate School, Biochemistry and Molecular Biology Track, Mayo Clinic College of Medicine, Rochester, MN 55905;
Mayo Graduate School, Biochemistry and Molecular Biology Track, Mayo Clinic College of Medicine, Rochester, MN 55905; Epigenomics Developmental Program, Center for Individualized Medicine, Mayo Clinic College of Medicine, Rochester, MN 55905 zhang.zhiguo@mayo.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 311
Author: Lee, S. O., Connolly, J. M., Ramirez-Soto, D. and Poretz, R. D.
Year: 1990
Title: The polypeptide of immunoglobulin G influences its galactosylation in vivo
Journal: J Biol Chem
Volume: 265
Issue: 10
Pages: 5833-9
Epub Date: 1990/04/05
Date: Apr 5
Short Title: The polypeptide of immunoglobulin G influences its galactosylation in vivo
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2108149
Keywords: Acetylglucosaminidase/metabolism
Animals
Carbohydrate Sequence
Cloning, Molecular
Galactose/*metabolism
Glycopeptides/isolation & purification/metabolism
Glycoside Hydrolases/metabolism
Glycosylation
Hybridomas/immunology
Immunoglobulin G/*metabolism
Immunoglobulin Heavy Chains/*metabolism
Immunoglobulin gamma-Chains/analysis/metabolism
Macromolecular Substances
Methylation
Mice
Molecular Sequence Data
Pronase/metabolism
Tumor Cells, Cultured
Abstract: To examine the nature of the factors influencing the galactosylation pattern of the heavy chain of murine immunoglobulin G (IgG), cell fusion was performed between a myeloma (P3x63Ag8) and a hybridoma (Sp2HL/Bu) cell line which secrete different IgGs possessing structurally distinct CH2-linked oligosaccharide moieties. The glycosylation patterns of the IgGs of the parental and fused cells were studied. Pronase digestion of the purified heavy chains and subsequent end labeling with fluorescein isothiocyanate produced fluoresceinated glycopeptides which were detected and purified by polyacrylamide gel electrophoresis. Structural information was obtained by enzymatic digestion, lectin affinity chromatography, and methylation analysis. IgGs from both parental lines possessed oligosaccharide units displaying microheterogeneity based upon a common symmetrical biantennary structure terminating in beta-GlcNAc. The structures of both IgGs, however, differed in the pattern of the mono- and digalactosylated components. Clones, selected following the fusion of the parental cells, were expanded; and the individual IgGs were purified. All clones produced homodimeric IgG1 and IgG2b as well as heterodimeric IgG possessing both the gamma 1 and gamma 2b heavy chains. Analysis of the carbohydrate moieties of the gamma 1 chain from the homodimeric and heterodimeric IgGs and of the gamma 2b chain from the heterodimeric molecule demonstrates that the polypeptide structure of the heavy chain influences the terminal galactosylation of the glycan unit at the conserved site of glycosylation of IgGs.
Notes: Lee, S O
Connolly, J M
Ramirez-Soto, D
Poretz, R D
AI 06471/AI/NIAID NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1990 Apr 5;265(10):5833-9.
Author Address: Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New Jersey 08855-1059.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1589
Author: Lee, T. H., Mitchell, A., Liu Lau, S., An, H., Rajeaskariah, P., Wasinger, V., Raftery, M., Bryant, K. and Tedla, N.
Year: 2013
Title: Glycosylation in a mammalian expression system is critical for the production of functionally active leukocyte immunoglobulin-like receptor A3 protein
Journal: J Biol Chem
Volume: 288
Issue: 46
Pages: 32873-85
Epub Date: 2013/10/03
Date: Nov 15
Short Title: Glycosylation in a mammalian expression system is critical for the production of functionally active leukocyte immunoglobulin-like receptor A3 protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.478578
PMCID: PMC3829139
Accession Number: 24085305
Keywords: Cell Line
Female
*Gene Expression
Glycosylation
Humans
Macrophages/cytology/metabolism
Male
Pichia/genetics/metabolism
Receptors, Immunologic/*biosynthesis/chemistry/genetics
Recombinant Proteins/biosynthesis/chemistry/genetics
Glycoprotein
Leukocyte
Mass Spectrometry (MS)
Monocytes
Protein Deamidation
Protein Expression
Protein Purification
Recombinant Protein Expression
Tumor Necrosis Factor (TNF)
Abstract: The leukocyte immunoglobulin-like receptor (LILR) A3 is a member of the highly homologous activating and inhibitory receptors expressed on leukocytes. LILRA3 is a soluble receptor of unknown functions but is predicted to act as a broad antagonist to other membrane-bound LILRs. Functions of LILRA3 are unclear primarily because of the lack of high quality functional recombinant protein and insufficient knowledge regarding its ligand(s). Here, we expressed and characterized recombinant LILRA3 (rLILRA3) proteins produced in 293T cells, Escherichia coli, and Pichia pastoris. We found that the purified rLILRA3 produced in the mammalian system was the same size as a 70-kDa native macrophage LILRA3. This is 20 kDa larger than the calculated size, suggesting significant post-translational modifications. In contrast, rLILRA3 produced in E. coli was similar in size to the unprocessed protein, but yeast-produced protein was 2-4 times larger than the unprocessed protein. Treatment with peptide-N-glycosidase F reduced the size of the mammalian cell- and yeast-produced rLILRA3 to 50 kDa, suggesting that most modifications are due to glycosylation. Consistent with this, mass spectrometric analysis of the mammalian rLILRA3 revealed canonical N-glycosylation at the predicted Asn(140), Asn(281), Asn(302), Asn(341), and Asn(431) sites. Functionally, only mammalian cell-expressed rLILRA3 bound onto the surface of monocytes with high affinity, and importantly, only this significantly abrogated LPS-induced TNFα production by monocytes. Binding to monocytes was partially blocked by β-lactose, indicating that optimally glycosylated LILRA3 might be critical for ligand binding and function. Overall, our data demonstrated for the first time that LILRA3 is a potential new anti-inflammatory protein, and optimal glycosylation is required for its functions.
Notes: 1083-351x
Lee, Terry H Y
Mitchell, Ainslie
Liu Lau, Sydney
An, Hongyan
Rajeaskariah, Poornima
Wasinger, Valerie
Raftery, Mark
Bryant, Katherine
Tedla, Nicodemus
Journal Article
J Biol Chem. 2013 Nov 15;288(46):32873-85. doi: 10.1074/jbc.M113.478578. Epub 2013 Sep 30.
Author Address: From the Inflammation and Infection Research Centre, School of Medical Sciences.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1317
Author: Lee, Y., Kockx, M., Raftery, M. J., Jessup, W., Griffith, R. and Kritharides, L.
Year: 2010
Title: Glycosylation and sialylation of macrophage-derived human apolipoprotein E analyzed by SDS-PAGE and mass spectrometry: evidence for a novel site of glycosylation on Ser290
Journal: Mol Cell Proteomics
Volume: 9
Issue: 9
Pages: 1968-81
Epub Date: 2010/06/01
Date: Sep
Short Title: Glycosylation and sialylation of macrophage-derived human apolipoprotein E analyzed by SDS-PAGE and mass spectrometry: evidence for a novel site of glycosylation on Ser290
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M900430-MCP200
PMCID: PMC2938105
Accession Number: 20511397
Keywords: Amino Acid Sequence
Apolipoproteins E/*metabolism
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Immunoprecipitation
Macrophages/*metabolism
N-Acetylneuraminic Acid/*metabolism
Serine/*metabolism
Tandem Mass Spectrometry
Abstract: Apolipoprotein E (apoE) is a 34-kDa glycoprotein secreted from various cells including hepatocytes and macrophages and plays an important role in remnant lipoprotein clearance, immune responses, Alzheimer disease, and atherosclerosis. Cellular apoE and plasma apoE exist as multiple glycosylated and sialylated glycoforms with plasma apoE being less glycosylated/sialylated than cell-derived apoE. Some of the glycan structures on plasma apoE are characterized; however, the more complicated structures on plasma and cellular/secreted apoE remain unidentified. We investigated glycosylation and sialylation of cellular and secreted apoE from primary human macrophages by one- and two-dimensional gel electrophoresis and mass spectrometry. Our results identify eight different glycoforms with (HexNAc)(2)-Hex(2)-(NeuAc)(2) being the most complex glycan detected on Thr(194) in both cellular and secreted apoE. Four additional glycans were identified on apoE(283-299), and using beta-elimination/alkylation by methylamine in vitro, we identified Ser(290) as a novel site of glycan attachment. Comparison of plasma and cellular/secreted apoE from the same donor confirmed that cell-derived apoE is more extensively sialylated than plasma apoE. Given the importance of the C terminus of apoE in regulating apoE solubility, stability, and lipid binding, these results may have important implications for our understanding of apoE biochemistry.
Notes: 1535-9484
Lee, Youra
Kockx, Maaike
Raftery, Mark J
Jessup, Wendy
Griffith, Renate
Kritharides, Leonard
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2010 Sep;9(9):1968-81. doi: 10.1074/mcp.M900430-MCP200. Epub 2010 May 28.
Author Address: Centre for Vascular Research, School of Medical Sciences, University of New South Wales, Sydney, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 559
Author: Lefebvre, J. C., Giordanengo, V., Doglio, A., Cagnon, L., Breittmayer, J. P., Peyron, J. F. and Lesimple, J.
Year: 1994
Title: Altered sialylation of CD45 in HIV-1-infected T lymphocytes
Journal: Virology
Volume: 199
Issue: 2
Pages: 265-74
Epub Date: 1994/03/01
Date: Mar
Short Title: Altered sialylation of CD45 in HIV-1-infected T lymphocytes
Alternate Journal: Virology
ISSN: 0042-6822 (Print)
0042-6822
DOI: 10.1006/viro.1994.1124
Accession Number: 8122360
Keywords: Carbohydrate Sequence
Cells, Cultured
Glycosylation
*Hiv-1
Leukocyte Common Antigens/*biosynthesis/metabolism
Molecular Sequence Data
Protein Tyrosine Phosphatases/metabolism
Sialoglycoproteins/*biosynthesis/metabolism
T-Lymphocytes/*metabolism/*microbiology
Abstract: Immunodeficiency caused by HIV infection probably results from profound dysregulation of normal T lymphocyte properties by the virus. Despite description of the virus cytopathicity and numerous modifications in T cell functions, such as perturbation of antigen receptor signaling, CD4 downregulation, and induction of apoptosis, the precise mechanisms underlying the disruption of normal immune responses have not yet been elucidated. In the present study, we show that HIV-1-infected lymphocytes of the CEM cell line (either latent or virus-producing) and HIV-1-infected CD4+ lymphocytes have several membrane proteins with altered glycosylation patterns. Using lectins with specificity for different carbohydrate moieties, we could demonstrate the presence of two exposed nonsialylated disaccharides: a terminal Gal beta 1-->3GalNAc and a terminal Gal beta 1-->4GlcNAc. In particular, CD45, one of the major T cell glycoproteins, appeared to be partially sialylated on N- and O-linked carbohydrate moieties. Concerning the latter, PNA lectin which recognizes nonsialylated terminal Gal beta 1-->3GalNAc might precipitate up to 75% of the total tyrosine phosphatase activity displayed by CD45 molecules from one latently HIV-1-infected CEM cell line. Since CD45 glycoproteins are thought to play an important regulatory role in cell-to-cell interactions owing to their variable extracellular region and because they may regulate membrane signaling through their intracellular phosphatase domains, we suggest that these altered CD45 molecules may present an abnormal signal for natural ligands such as the B-cell-specific surface receptor CD22, thus perturbing the normal immune response in HIV-1-infected individuals.
Notes: Lefebvre, J C
Giordanengo, V
Doglio, A
Cagnon, L
Breittmayer, J P
Peyron, J F
Lesimple, J
Journal Article
Research Support, Non-U.S. Gov't
United States
Virology. 1994 Mar;199(2):265-74. doi: 10.1006/viro.1994.1124.
Author Address: Laboratoire de Virologie, U.E.R. de Médecine, Nice, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 862
Author: Lefebvre, T., Ferreira, S., Dupont-Wallois, L., Bussière, T., Dupire, M. J., Delacourte, A., Michalski, J. C. and Caillet-Boudin, M. L.
Year: 2003
Title: Evidence of a balance between phosphorylation and O-GlcNAc glycosylation of Tau proteins--a role in nuclear localization
Journal: Biochim Biophys Acta
Volume: 1619
Issue: 2
Pages: 167-76
Epub Date: 2003/01/16
Date: Jan 20
Short Title: Evidence of a balance between phosphorylation and O-GlcNAc glycosylation of Tau proteins--a role in nuclear localization
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/s0304-4165(02)00477-4
Accession Number: 12527113
Keywords: Blotting, Western
Cell Nucleus/*metabolism
Cytosol/metabolism
Electrophoresis, Gel, Two-Dimensional
Enzyme Inhibitors/pharmacology
Glycosylation/drug effects
Humans
N-Acetylglucosaminyltransferases/antagonists & inhibitors/metabolism
Okadaic Acid/pharmacology
Phosphorylation
Protein Isoforms/metabolism
Tumor Cells, Cultured
tau Proteins/biosynthesis/genetics/*metabolism
Abstract: Both phosphorylation and O-GlcNAc glycosylation posttranslationally modify microtubule-associated Tau proteins. Whereas the hyperphosphorylation of these proteins that occurs in Alzheimer's disease is well characterized, little is known about the O-GlcNAc glycosylation. The present study demonstrates that a balance exists between phosphorylation and O-GlcNAc glycosylation of Tau proteins, and furthermore that a dysfunction of this balance correlates with reduced nuclear localization. The affinity of Tau proteins for WGA lectin, together with evidence from [3H]-galactose transfer and analysis of beta-eliminated products, demonstrated the presence of O-GlcNAc residues on both cytosolic and nuclear Tau proteins. In addition, our data indicated the existence of a balance between phosphorylation and O-GlcNAc glycosylation events. Indeed, as demonstrated by 2D-electrophoresis and Western blotting, O-GlcNAc residues were mainly located on the less phosphorylated Tau 441 variants, whereas the more phosphorylated forms were devoid of O-GlcNAc residues. Furthermore, the Tau protein hyperphosphorylation induced by cellular okadaic acid treatment was correlated with reduced incorporation of O-GlcNAc residues into Tau proteins and with diminished Tau transfer into the nucleus. Hence, this paper establishes a direct relationship between O-GlcNAc glycosylation, phosphorylation and cellular localization of Tau proteins.
Notes: Lefebvre, Tony
Ferreira, Stéphanie
Dupont-Wallois, Laetitia
Bussière, Thierry
Dupire, Marie-Joëlle
Delacourte, André
Michalski, Jean-Claude
Caillet-Boudin, Marie-Laure
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2003 Jan 20;1619(2):167-76. doi: 10.1016/s0304-4165(02)00477-4.
Author Address: Laboratoire de Chimie Biologique, Unité Mixte de Recherches 8576 du CNRS, Université des Sciences et Technologies de Lille I, F-59655 Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 953
Author: Lefebvre, T., Pinte, S., Guérardel, C., Deltour, S., Martin-Soudant, N., Slomianny, M. C., Michalski, J. C. and Leprince, D.
Year: 2004
Title: The tumor suppressor HIC1 (hypermethylated in cancer 1) is O-GlcNAc glycosylated
Journal: Eur J Biochem
Volume: 271
Issue: 19
Pages: 3843-54
Epub Date: 2004/09/18
Date: Oct
Short Title: The tumor suppressor HIC1 (hypermethylated in cancer 1) is O-GlcNAc glycosylated
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.2004.04316.x
Accession Number: 15373830
Keywords: Acetylglucosamine/analogs & derivatives/chemistry/*metabolism
Animals
CHO Cells
COS Cells
Cells, Cultured
Chlorocebus aethiops
Chromatography, Affinity
Cricetinae
DNA Methylation
Electrophoretic Mobility Shift Assay
*Genes, Tumor Suppressor
Glycoconjugates/chemistry/metabolism
Glycosylation
Kruppel-Like Transcription Factors
Neoplasms/*metabolism/pathology
Peptide Fragments/chemistry/*metabolism
Protein Conformation
Transcription Factors/*metabolism
Trypsin/pharmacology
Wheat Germ Agglutinins/metabolism
Abstract: HIC1 (hypermethylated in cancer 1) is a transcriptional repressor containing five Krüppel-like C(2)H(2) zinc fingers and an N-terminal dimerization and autonomous repression domain called BTB/POZ. Here, we demonstrate that full-length HIC1 proteins are modified both in vivo and in vitro with O-linked N-acetylglucosamine (O-GlcNAc). This is a highly dynamic glycosylation found within the cytosolic and the nuclear compartments of eukaryotes. Analysis of [(3)H]Gal-labeled tryptic peptides indicates that HIC1 has three major sites for O-GlcNAc glycosylation. Using C-terminal deletion mutants, we have shown that O-GlcNAc modification of HIC1 proteins occurred preferentially in the DNA-binding domain. Nonglycosylated and glycosylated forms of full-length HIC1 proteins separated by wheat germ agglutinin affinity purification, displayed the same specific DNA-binding activity in electrophoretic mobility shift assays proving that the O-GlcNAc modification is not directly implicated in the specific DNA recognition of HIC1. Intriguingly, N-terminal truncated forms corresponding to BTB-POZ-deleted proteins exhibited a strikingly differential activity, as the glycosylated truncated forms are unable to bind DNA whereas the unglycosylated ones do. Electrophoretic mobility shift assays performed with separated pools of glycosylated and unglycosylated forms of a construct exhibiting only the DNA-binding domain and the C-terminal tail of HIC1 (residues 399-714) and supershift experiments with wheat germ agglutinin or RL-2, an antibody raised against O-GlcNAc residues, fully corroborated these results. Interestingly, these truncated proteins are O-GlcNAc modified in their C-terminal tail (residues 670-711) and not in the DNA-binding domain, as for the full-length proteins. Thus, the O-GlcNAc modification of HIC1 does not affect its specific DNA-binding activity and is highly sensitive to conformational effects, notably its dimerization through the BTB/POZ domain.
Notes: Lefebvre, Tony
Pinte, Sébastien
Guérardel, Cateline
Deltour, Sophie
Martin-Soudant, Nathalie
Slomianny, Marie-Christine
Michalski, Jean-Claude
Leprince, Dominique
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 2004 Oct;271(19):3843-54. doi: 10.1111/j.1432-1033.2004.04316.x.
Author Address: UMR 8526 du CNRS, Institut de Biologie de Lille, Institut Pasteur de Lille, Lille Cédex BP447, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 535
Author: Legrand, D., Salmon, V., Coddeville, B., Benaissa, M., Plancke, Y. and Spik, G.
Year: 1995
Title: Structural determination of two N-linked glycans isolated from recombinant human lactoferrin expressed in BHK cells
Journal: FEBS Lett
Volume: 365
Issue: 1
Pages: 57-60
Epub Date: 1995/05/22
Date: May 22
Short Title: Structural determination of two N-linked glycans isolated from recombinant human lactoferrin expressed in BHK cells
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(95)00441-b
Accession Number: 7774715
Keywords: Animals
Carbohydrate Sequence
Cells, Cultured
Cricetinae
Glycosylation
Humans
Lactoferrin/*chemistry/genetics
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Monosaccharides/analysis
Polysaccharides/*chemistry/genetics
Recombinant Proteins/chemistry
Sequence Analysis
Abstract: A full-length cDNA coding for human lactoferrin was isolated from a mammary gland library and the recombinant protein was expressed in BHK cells as described by Stowell K. M. et al. [1991, Biochem. J. 276, 349-355]. Two N-linked glycans from purified recombinant lactoferrin were released by hydrazinolysis and analyzed by 400-MHz 1H-NMR spectroscopy. The identified structures corresponded to N-acetyllactosaminic biantennary glycans and were alpha-2,3-disialylated forms (80%) or alpha-2,3-monosialylated (20%) forms. Moreover, 70% of total glycans were alpha-1,6-fucosylated at the GlcNAc residue linked to asparagine. In regard to its glycan moiety, the recombinant glycoprotein is close to native lactoferrins from milk or leucocytes but shows specific structural features which should be taken into account prior to in vivo and in vitro biological studies.
Notes: Legrand, D
Salmon, V
Coddeville, B
Benaissa, M
Plancke, Y
Spik, G
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1995 May 22;365(1):57-60. doi: 10.1016/0014-5793(95)00441-b.
Author Address: Laboratoire de Chimie Biologique, UMR CNRS n.111, Université des Sciences et Technologies de Lille, Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1957
Author: Lei, L., Xie, J., Yu, J., Li, Y. and Liu, Y.
Year: 2018
Title: Parallel study on protein O-GlcNAcylation in prostate cancer cell with a sensitive microarray biochip
Journal: Anal Biochem
Volume: 558
Pages: 53-59
Epub Date: 2018/08/08
Date: Oct 1
Short Title: Parallel study on protein O-GlcNAcylation in prostate cancer cell with a sensitive microarray biochip
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697
DOI: 10.1016/j.ab.2018.08.004
Accession Number: 30086259
Keywords: Acylation
Cell Line, Tumor
Cell Movement
Cell Proliferation
Humans
*Lab-On-A-Chip Devices
Male
N-Acetylglucosaminyltransferases/*metabolism
NF-kappa B/metabolism
*Oligonucleotide Array Sequence Analysis
Prostatic Neoplasms/*metabolism/pathology
Proto-Oncogene Proteins c-myc/metabolism
Tumor Suppressor Protein p53/metabolism
*Cell migration
*Prostate cancer
*Protein O-GlcNAcylation
*Protein microarray
Abstract: Although a variety of approaches have been developed to analyze protein O-GlcNAcylation, efficient investigations on O-GlcNAcylation of proteins of interest in high-throughput manner are still in high demand to further explore its functionality. In this work, we first develop a powerful microarray platform for a sensitive, specific and high-throughput analysis of protein O-GlcNAcylation. The developed array biochip is then utilized to parallelly analyze the O-GlcNAcylation of three oncogenic transcription factors C-Myc, NF-κB and p53 in normal prostate epithelial cell (RWPE-1) and prostate cancer cell line (PC-3). The levels of O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) are also monitored by the microarray platform. The experimental results show that the overall O-GlcNAcylation and OGT expression level are obviously elevated in PC-3 as compared to RWPE-1. The protein expression-normalized O-GlcNAcylation of C-Myc and NF-κB in PC-3 is significantly higher than that in RWPE-1, while opposite result is observed from p53. In addition, the biological behaviors including proliferation and migration of PC-3 cells are also studied when OGA inhibitor Thiamet G is applied to elevate the total O-GlcNAcylation level.
Notes: 1096-0309
Lei, Lingli
Xie, Jin
Yu, Jie
Li, Yuan
Liu, Yingshuai
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Biochem. 2018 Oct 1;558:53-59. doi: 10.1016/j.ab.2018.08.004. Epub 2018 Aug 4.
Author Address: Southwest University, Institute for Clean Energy and Advanced Materials, Chongqing, 400715, China; Chongqing Key Laboratory for Advanced Materials and Technologies of Clean Energies, Southwest University, Chongqing, 400715, China.
Central Laboratory of Yongchuan Hospital, Chongqing Medical University, Chongqing, 402160, China. Electronic address: liyuan1985999@163.com.
Southwest University, Institute for Clean Energy and Advanced Materials, Chongqing, 400715, China; Chongqing Key Laboratory for Advanced Materials and Technologies of Clean Energies, Southwest University, Chongqing, 400715, China. Electronic address: ysliu@swu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2059
Author: Lei, Y., Chen, T., Li, Y., Shang, M., Zhang, Y., Jin, Y., Yu, Q., Guo, F. and Wang, T.
Year: 2020
Title: O-GlcNAcylation of PFKFB3 is required for tumor cell proliferation under hypoxia
Journal: Oncogenesis
Volume: 9
Issue: 2
Pages: 21
Epub Date: 2020/02/16
Date: Feb 14
Short Title: O-GlcNAcylation of PFKFB3 is required for tumor cell proliferation under hypoxia
Alternate Journal: Oncogenesis
ISSN: 2157-9024 (Print)
2157-9024
DOI: 10.1038/s41389-020-0208-1
PMCID: PMC7021673
Accession Number: 32060258
Abstract: The protein O-GlcNAcylation catalysed by O-GlcNAc transferase (OGT) is tightly regulated by glucose availability. It is upregulated and essential for tumor cell proliferation under hypoxic conditions. However, the mechanism behind is still unclear. Here, we showed that the glycolytic regulator 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB3), which also promotes cell cycle progression in the nucleus, was O-GlcNAcylated in response to hypoxia. The O-GlcNAcylation of PFKFB3 could compete phosphorylation by hypoxia-activated ERK at the same modification site Ser172. Phosphorylated PFKFB3 could interact with the protein G3BP2 and retain in the cytosol; this in turn led to the accumulation of hypoxia-induced-P27 in the nucleus resulting in the cell cycle arrest. Such a pathway was compromised by high level of PFKFB3 O-GlcNAcylation in tumor cells contributing to cell cycle progression. Consistently, the PFKFB3-Ser172 phosphorylation level inversely correlated with the OGT level in pancreatic cancer patients. Our findings uncovered an O-GlcNAcylation mediated mechanism to promote tumor cell proliferation under metabolic stress, linking the aberrant OGT activity to tumorigenesis in pancreatic cancer.
Notes: 2157-9024
Lei, Yinrui
Chen, Tao
Li, Yeyi
Shang, Man
Zhang, Yan
Jin, Yuepeng
Yu, Qiujing
Guo, Fang
Wang, Ting
Orcid: 0000-0003-2257-0699
Journal Article
Oncogenesis. 2020 Feb 14;9(2):21. doi: 10.1038/s41389-020-0208-1.
Author Address: Department of Pharmacology And Tianjin Key Laboratory of Inflammation Biology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin, 300070, China.
Key Laboratory of Systems Biomedicine (Ministry of Education), Shanghai Center for Systems Biomedicine, Shanghai Jiao Tong University, 800 Dongchuan Road, Shanghai, 200240, China.
Endoscopy Center, Shanghai East Hospital, Tongji University, 150Jimo Road, Shanghai, 200120, China.
Department of General Surgery, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou, 325000, China.
Key Laboratory of Immune Microenvironment and Disease (Ministry of Education), Department of Immunology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin, 300070, China.
Department of Pharmacology And Tianjin Key Laboratory of Inflammation Biology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin, 300070, China. twang1@tmu.edu.cn.
The Institute of Cell Metabolism and Disease, Shanghai Key Laboratory of Pancreatic Cancer, Shanghai General Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, 201620, China. twang1@tmu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1954
Author: Leloup, N., Chataigner, L. M. P. and Janssen, B. J. C.
Year: 2018
Title: Structural insights into SorCS2-Nerve Growth Factor complex formation
Journal: Nat Commun
Volume: 9
Issue: 1
Pages: 2979
Epub Date: 2018/08/01
Date: Jul 30
Short Title: Structural insights into SorCS2-Nerve Growth Factor complex formation
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-018-05405-z
PMCID: PMC6065357
Accession Number: 30061605
Keywords: Amino Acid Motifs
Animals
Biophysics
Brain-Derived Neurotrophic Factor/*chemistry
Crystallography, X-Ray
HEK293 Cells
Humans
Ligands
Mice
Mutation
Nerve Growth Factor/*chemistry
Nerve Tissue Proteins/*chemistry
Neuronal Plasticity
Protein Binding
Protein Domains
Protein Folding
Protein Multimerization
Protein Structure, Secondary
Receptors, Cell Surface/*chemistry
Recombinant Proteins/chemistry
Signal Transduction
Surface Plasmon Resonance
Abstract: Signaling of SorCS receptors by proneurotrophin ligands regulates neuronal plasticity, induces apoptosis and is associated with mental disorders. The detailed structure of SorCS2 and its extracellular specificity are unresolved. Here we report crystal structures of the SorCS2-NGF complex and unliganded SorCS2 ectodomain, revealing cross-braced SorCS2 homodimers with two NGF dimers bound in a 2:4 stoichiometry. Five out of six SorCS2 domains directly contribute to dimer formation and a C-terminal membrane proximal unreported domain, with an RNA recognition motif fold, locks the dimer in an intermolecular head-to-tail interaction. The complex structure shows an altered SorCS2 conformation indicating substantial structural plasticity. Both NGF dimer chains interact exclusively with the top face of a SorCS2 β-propeller. Biophysical experiments reveal that NGF, proNGF, and proBDNF bind at this site on SorCS2. Taken together, our data reveal a structurally flexible SorCS2 receptor that employs the large β-propeller as a ligand binding platform.
Notes: 2041-1723
Leloup, Nadia
Orcid: 0000-0002-1504-3690
Chataigner, Lucas M P
Janssen, Bert J C
Orcid: 0000-0002-8101-8370
317371/EC | Seventh Framework Programme (European Union Seventh Framework Programme)/International
723.012.002/Nederlandse Organisatie voor Wetenschappelijk Onderzoek (Netherlands Organisation for Scientific Research)/International
721.012.004/Nederlandse Organisatie voor Wetenschappelijk Onderzoek (Netherlands Organisation for Scientific Research)/International
677500/EC | European Research Council (ERC)/International
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2018 Jul 30;9(1):2979. doi: 10.1038/s41467-018-05405-z.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Faculty of Science, Utrecht University, 3584 CH, Utrecht, The Netherlands.
Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Faculty of Science, Utrecht University, 3584 CH, Utrecht, The Netherlands. b.j.c.janssen@uu.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1045
Author: Lemieux, M. J., Mark, B. L., Cherney, M. M., Withers, S. G., Mahuran, D. J. and James, M. N.
Year: 2006
Title: Crystallographic structure of human beta-hexosaminidase A: interpretation of Tay-Sachs mutations and loss of GM2 ganglioside hydrolysis
Journal: J Mol Biol
Volume: 359
Issue: 4
Pages: 913-29
Epub Date: 2006/05/16
Date: Jun 16
Short Title: Crystallographic structure of human beta-hexosaminidase A: interpretation of Tay-Sachs mutations and loss of GM2 ganglioside hydrolysis
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2006.04.004
PMCID: PMC2910082
NIHMSID: CAMS1392
Accession Number: 16698036
Keywords: Acetylglucosamine/analogs & derivatives/chemistry/metabolism
Amino Acid Substitution
Arginine/genetics/metabolism
Aspartic Acid/genetics/metabolism
Binding Sites
Crystallography, X-Ray
Dimerization
Gangliosidoses, GM2/*metabolism
Glycine/genetics/metabolism
Glycosylation
Hexosaminidase A
Humans
Hydrolysis
Models, Molecular
*Mutation
Protein Conformation
Protein Subunits
Tay-Sachs Disease/*genetics
Thiazoles/chemistry/metabolism
beta-N-Acetylhexosaminidases/*chemistry/genetics/*metabolism
Abstract: Lysosomal beta-hexosaminidase A (Hex A) is essential for the degradation of GM2 gangliosides in the central and peripheral nervous system. Accumulation of GM2 leads to severely debilitating neurodegeneration associated with Tay-Sachs disease (TSD), Sandoff disease (SD) and AB variant. Here, we present the X-ray crystallographic structure of Hex A to 2.8 A resolution and the structure of Hex A in complex with NAG-thiazoline, (NGT) to 3.25 A resolution. NGT, a mechanism-based inhibitor, has been shown to act as a chemical chaperone that, to some extent, prevents misfolding of a Hex A mutant associated with adult onset Tay Sachs disease and, as a result, increases the residual activity of Hex A to a level above the critical threshold for disease. The crystal structure of Hex A reveals an alphabeta heterodimer, with each subunit having a functional active site. Only the alpha-subunit active site can hydrolyze GM2 gangliosides due to a flexible loop structure that is removed post-translationally from beta, and to the presence of alphaAsn423 and alphaArg424. The loop structure is involved in binding the GM2 activator protein, while alphaArg424 is critical for binding the carboxylate group of the N-acetyl-neuraminic acid residue of GM2. The beta-subunit lacks these key residues and has betaAsp452 and betaLeu453 in their place; the beta-subunit therefore cleaves only neutral substrates efficiently. Mutations in the alpha-subunit, associated with TSD, and those in the beta-subunit, associated with SD are discussed. The effect of NGT binding in the active site of a mutant Hex A and its effect on protein function is discussed.
Notes: 1089-8638
Lemieux, M Joanne
Mark, Brian L
Cherney, Maia M
Withers, Stephen G
Mahuran, Don J
James, Michael N G
82944/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
J Mol Biol. 2006 Jun 16;359(4):913-29. doi: 10.1016/j.jmb.2006.04.004. Epub 2006 Apr 27.
Author Address: CIHR Group in Protein Structure and Function, Department of Biochemistry, University of Alberta, Edmonton, Alta., Canada T6G 2H7.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1310
Author: Lenertz, L. Y., Wang, Z., Guadarrama, A., Hill, L. M., Gavala, M. L. and Bertics, P. J.
Year: 2010
Title: Mutation of putative N-linked glycosylation sites on the human nucleotide receptor P2X7 reveals a key residue important for receptor function
Journal: Biochemistry
Volume: 49
Issue: 22
Pages: 4611-9
Epub Date: 2010/05/11
Date: Jun 8
Short Title: Mutation of putative N-linked glycosylation sites on the human nucleotide receptor P2X7 reveals a key residue important for receptor function
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi902083n
PMCID: PMC2895974
NIHMSID: NIHMS205418
Accession Number: 20450227
Keywords: Amino Acid Sequence
Animals
Asparagine/*genetics/metabolism
COS Cells
Cell Line
Chlorocebus aethiops
Conserved Sequence
Down-Regulation/genetics
Extracellular Space/genetics
Glycosylation
Humans
Mice
Molecular Sequence Data
*Point Mutation
Polymorphism, Single Nucleotide
Protein Processing, Post-Translational/genetics
Protein Transport/genetics
Purinergic P2 Receptor Agonists
Receptors, Purinergic P2/*genetics/*metabolism/physiology
Receptors, Purinergic P2X7
Signal Transduction/genetics
Abstract: The nucleotide receptor P2X(7) is an immunomodulatory cation channel and a potential therapeutic target. P2X(7) is expressed in immune cells such as monocytes and macrophages and is activated by extracellular ATP following tissue injury or infection. Ligand binding to P2X(7) can stimulate ERK1/2, the transcription factor CREB, enzymes linked to the production of reactive oxygen species and interleukin-1 isoforms, and the formation of a nonspecific pore. However, little is known about the biochemistry of P2X(7), including whether the receptor is N-linked glycosylated and if this modification affects receptor function. Here we provide evidence that P2X(7) is sensitive to the glycosidases EndoH and PNGase F and that the human receptor appears glycosylated at N187, N202, N213, N241, and N284. Mutation of N187 results in weakened P2X(7) agonist-induced phosphorylation of ERK1/2, CREB, and p90 ribosomal S6 kinase, as well as a decreased level of pore formation. In further support of a role for glycosylation in receptor function, treatment of RAW 264.7 macrophages with the N-linked glycosylation synthesis inhibitor tunicamycin attenuates P2X(7) agonist-induced, but not phorbol ester-induced, ERK1/2 phosphorylation. Interestingly, residue N187 belongs to an N-linked glycosylation consensus sequence found in six of the seven P2X family members, suggesting this site is fundamentally important to P2X receptor function. To address the mechanism whereby N187 mutation attenuates receptor activity, we developed a live cell proteinase K digestion assay that demonstrated altered cell surface expression of P2X(7) N187A. This is the first report to map human P2X(7) glycosylation sites and reveal residue N187 is critical for receptor trafficking and function.
Notes: 1520-4995
Lenertz, Lisa Y
Wang, Ziyi
Guadarrama, Arturo
Hill, Lindsay M
Gavala, Monica L
Bertics, Paul J
U19 AI070503-040002/AI/NIAID NIH HHS/United States
2 R01 HL069116/HL/NHLBI NIH HHS/United States
U19 AI070503/AI/NIAID NIH HHS/United States
R01 HL069116/HL/NHLBI NIH HHS/United States
1 U19 AI070503/AI/NIAID NIH HHS/United States
1 P01 HL0885940/HL/NHLBI NIH HHS/United States
U19 AI070503-030002/AI/NIAID NIH HHS/United States
P01 HL088594-020003/HL/NHLBI NIH HHS/United States
P01 HL088594-030003/HL/NHLBI NIH HHS/United States
P01 HL088594/HL/NHLBI NIH HHS/United States
P01 HL088594-03/HL/NHLBI NIH HHS/United States
R01 HL069116-09/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochemistry. 2010 Jun 8;49(22):4611-9. doi: 10.1021/bi902083n.
Author Address: Department of Biomolecular Chemistry, The University of Wisconsin, Madison, Wisconsin 53706, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1861
Author: Leney, A. C., Rafie, K., van Aalten, D. M. F. and Heck, A. J. R.
Year: 2017
Title: Direct Monitoring of Protein O-GlcNAcylation by High-Resolution Native Mass Spectrometry
Journal: ACS Chem Biol
Volume: 12
Issue: 8
Pages: 2078-2084
Epub Date: 2017/06/14
Date: Aug 18
Short Title: Direct Monitoring of Protein O-GlcNAcylation by High-Resolution Native Mass Spectrometry
Alternate Journal: ACS chemical biology
ISSN: 1554-8929 (Print)
1554-8929
DOI: 10.1021/acschembio.7b00371
PMCID: PMC5565903
Accession Number: 28609614
Keywords: Acetylglucosamine/*chemistry
Acylation
Glycosylation
Kinetics
*Mass Spectrometry
Protein Processing, Post-Translational
Proteins/chemistry/*metabolism
Abstract: O-GlcNAcylation is one of the most abundant metazoan nuclear-cytoplasmic post-translational modifications. Proteins modified by O-GlcNAc play key cellular roles in signaling, transcription, metabolism, and cell division. Mechanistic studies on protein O-GlcNAcylation are hampered by the lack of methods that can simultaneously quantify O-GlcNAcylation, determine its stoichiometry, and monitor O-GlcNAcylation kinetics. Here, we demonstrate that high-resolution native mass spectrometry can be employed to monitor the small mass shifts induced by modification by O-GlcNAc on two known protein substrates, CK2α and TAB1, without the need for radioactive labeling or chemoenzymatic tagging using large mass tags. Limited proteolysis enabled further localization of the O-GlcNAc sites. In peptide-centric MS analysis, the O-GlcNAc moiety is known to be easily lost. In contrast, we demonstrate that the O-GlcNAc is retained under native MS conditions, enabling precise quantitative analysis of stoichiometry and O-GlcNAcylation kinetics. Together, the data highlight that high resolution native MS may provide an alternative tool to monitor kinetics on one of the most labile of protein post-translational modifications, in an efficient, reliable, and quantitative manner.
Notes: 1554-8937
Leney, Aneika C
Rafie, Karim
van Aalten, Daan M F
Heck, Albert J R
Orcid: 0000-0002-2405-4404
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
ACS Chem Biol. 2017 Aug 18;12(8):2078-2084. doi: 10.1021/acschembio.7b00371. Epub 2017 Jun 28.
Author Address: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences and Netherlands Proteomics Centre, Utrecht University , Padualaan 8, 3584 CH, Utrecht, The Netherlands.
School of Life Sciences, University of Dundee , Dow Street, DD1 5EH, Dundee, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2031
Author: Lenin, R., Nagy, P. G., Jha, K. A. and Gangaraju, R.
Year: 2019
Title: GRP78 translocation to the cell surface and O-GlcNAcylation of VE-Cadherin contribute to ER stress-mediated endothelial permeability
Journal: Sci Rep
Volume: 9
Issue: 1
Pages: 10783
Epub Date: 2019/07/28
Date: Jul 25
Short Title: GRP78 translocation to the cell surface and O-GlcNAcylation of VE-Cadherin contribute to ER stress-mediated endothelial permeability
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/s41598-019-47246-w
PMCID: PMC6658495
Accession Number: 31346222
Keywords: Antigens, CD/*metabolism
Blood-Retinal Barrier/cytology/*metabolism
Cadherins/*metabolism
*Capillary Permeability
Cell Membrane/metabolism
Cell Movement
Cells, Cultured
Endoplasmic Reticulum/drug effects/metabolism
*Endoplasmic Reticulum Stress
Endothelial Cells/*metabolism
Glycosylation
Heat-Shock Proteins/*metabolism
Humans
Monocytes/physiology
Protein Transport
Taurochenodeoxycholic Acid/pharmacology
Abstract: Increased O-GlcNAcylation, a well-known post-translational modification of proteins causally linked to various detrimental cellular functions in pathological conditions including diabetic retinopathy (DR). Previously we have shown that endothelial activation induced by inflammation and hyperglycemia results in the endoplasmic reticulum (ER) stress-mediated intercellular junction alterations accompanied by visual deficits in a tie2-TNF-α transgenic mouse model. In this study, we tested the hypothesis that increased ER stress via O-GlcNAcylation of VE-Cadherin likely contribute to endothelial permeability. We show that ER stress leads to GRP78 translocation to the plasma membrane, increased O-GlcNAcylation of proteins, particularly VE-Cadherin resulting in a defective complex partnering leading to the loss of retinal endothelial barrier integrity and increased transendothelial migration of monocytes. We further show an association of GRP78 with the VE-Cadherin under these conditions. Interestingly, cells exposed to ER stress inhibitor, tauroursodeoxycholic acid partially mitigated all these effects. Our findings suggest an essential role for ER stress and O-GlcNAcylation in altering the endothelial barrier function and reveal a potential therapeutic target in the treatment of DR.
Notes: 2045-2322
Lenin, Raji
Nagy, Peter G
Jha, Kumar Abhiram
Gangaraju, Rajashekhar
Orcid: 0000-0002-6664-8286
R01 EY023427/EY/NEI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Sci Rep. 2019 Jul 25;9(1):10783. doi: 10.1038/s41598-019-47246-w.
Author Address: Department of Ophthalmology, University of Tennessee Health Science Center, Memphis, TN, USA.
Department of Ophthalmology, University of Tennessee Health Science Center, Memphis, TN, USA. sgangara@uthsc.edu.
Department of Anatomy & Neurobiology, University of Tennessee Health Science Center, Memphis, TN, USA. sgangara@uthsc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1360
Author: Leppänen, V. M., Jeltsch, M., Anisimov, A., Tvorogov, D., Aho, K., Kalkkinen, N., Toivanen, P., Ylä-Herttuala, S., Ballmer-Hofer, K. and Alitalo, K.
Year: 2011
Title: Structural determinants of vascular endothelial growth factor-D receptor binding and specificity
Journal: Blood
Volume: 117
Issue: 5
Pages: 1507-15
Epub Date: 2010/12/15
Date: Feb 3
Short Title: Structural determinants of vascular endothelial growth factor-D receptor binding and specificity
Alternate Journal: Blood
ISSN: 0006-4971
DOI: 10.1182/blood-2010-08-301549
Accession Number: 21148085
Keywords: Amino Acid Sequence
Animals
Cell Proliferation
Cells, Cultured
Crystallography, X-Ray
Humans
Hydrogen Bonding
Immunoenzyme Techniques
Immunoprecipitation
Mice
Models, Molecular
Molecular Sequence Data
Muscle, Skeletal/cytology/*metabolism
Mutagenesis, Site-Directed
Mutation/genetics
Protein Binding
Protein Conformation
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Homology, Amino Acid
Vascular Endothelial Growth Factor C/chemistry/genetics/metabolism
Vascular Endothelial Growth Factor D/*chemistry/genetics/*metabolism
Vascular Endothelial Growth Factor Receptor-2/*metabolism
Vascular Endothelial Growth Factor Receptor-3/*metabolism
Abstract: Vascular endothelial growth factors (VEGFs) and their tyrosine kinase receptors (VEGFR-1-3) are central mediators of angiogenesis and lymphangiogenesis. VEGFR-3 ligands VEGF-C and VEGF-D are produced as precursor proteins with long N- and C-terminal propeptides and show enhanced VEGFR-2 and VEGFR-3 binding on proteolytic removal of the propeptides. Two different proteolytic cleavage sites have been reported in the VEGF-D N-terminus. We report here the crystal structure of the human VEGF-D Cys117Ala mutant at 2.9 Å resolution. Comparison of the VEGF-D and VEGF-C structures shows similar extended N-terminal helices, conserved overall folds, and VEGFR-2 interacting residues. Consistent with this, the affinity and the thermodynamic parameters for VEGFR-2 binding are very similar. In comparison with VEGF-C structures, however, the VEGF-D N-terminal helix was extended by 2 more turns because of a better resolution. Both receptor binding and functional assays of N-terminally truncated VEGF-D polypeptides indicated that the residues between the reported proteolytic cleavage sites are important for VEGF-D binding and activation of VEGFR-3, but not of VEGFR-2. Thus, we define here a VEGFR-2-specific form of VEGF-D that is angiogenic but not lymphangiogenic. These results provide important new insights into VEGF-D structure and function.
Notes: 1528-0020
Leppänen, Veli-Matti
Jeltsch, Michael
Anisimov, Andrey
Tvorogov, Denis
Aho, Kukka
Kalkkinen, Nisse
Toivanen, Pyry
Ylä-Herttuala, Seppo
Ballmer-Hofer, Kurt
Alitalo, Kari
Journal Article
Research Support, Non-U.S. Gov't
United States
Blood. 2011 Feb 3;117(5):1507-15. doi: 10.1182/blood-2010-08-301549. Epub 2010 Dec 8.
Author Address: Molecular Cancer Biology Program, Research Programs Unit, Haartman Institute, Institute for Molecular Medicine Finland and Helsinki University Central Hospital, Biomedicum Helsinki, University of Helsinki, Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1295
Author: Leppänen, V. M., Prota, A. E., Jeltsch, M., Anisimov, A., Kalkkinen, N., Strandin, T., Lankinen, H., Goldman, A., Ballmer-Hofer, K. and Alitalo, K.
Year: 2010
Title: Structural determinants of growth factor binding and specificity by VEGF receptor 2
Journal: Proc Natl Acad Sci U S A
Volume: 107
Issue: 6
Pages: 2425-30
Epub Date: 2010/02/11
Date: Feb 9
Short Title: Structural determinants of growth factor binding and specificity by VEGF receptor 2
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0914318107
PMCID: PMC2823880
Accession Number: 20145116
Keywords: Animals
Binding Sites/genetics
Cell Line
Cell Survival
Crystallography, X-Ray
Humans
Hydrogen Bonding
Kinetics
Models, Molecular
Mutation
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Recombinant Fusion Proteins/chemistry/genetics/metabolism
Spodoptera
Vascular Endothelial Growth Factor A/*chemistry/genetics/metabolism
Vascular Endothelial Growth Factor C/*chemistry/genetics/metabolism
Vascular Endothelial Growth Factor Receptor-1/chemistry/genetics/metabolism
Vascular Endothelial Growth Factor Receptor-2/*chemistry/genetics/metabolism
Vascular Endothelial Growth Factor Receptor-3/chemistry/genetics/metabolism
Abstract: Vascular endothelial growth factors (VEGFs) regulate blood and lymph vessel formation through activation of three receptor tyrosine kinases, VEGFR-1, -2, and -3. The extracellular domain of VEGF receptors consists of seven immunoglobulin homology domains, which, upon ligand binding, promote receptor dimerization. Dimerization initiates transmembrane signaling, which activates the intracellular tyrosine kinase domain of the receptor. VEGF-C stimulates lymphangiogenesis and contributes to pathological angiogenesis via VEGFR-3. However, proteolytically processed VEGF-C also stimulates VEGFR-2, the predominant transducer of signals required for physiological and pathological angiogenesis. Here we present the crystal structure of VEGF-C bound to the VEGFR-2 high-affinity-binding site, which consists of immunoglobulin homology domains D2 and D3. This structure reveals a symmetrical 22 complex, in which left-handed twisted receptor domains wrap around the 2-fold axis of VEGF-C. In the VEGFs, receptor specificity is determined by an N-terminal alpha helix and three peptide loops. Our structure shows that two of these loops in VEGF-C bind to VEGFR-2 subdomains D2 and D3, while one interacts primarily with D3. Additionally, the N-terminal helix of VEGF-C interacts with D2, and the groove separating the two VEGF-C monomers binds to the D2/D3 linker. VEGF-C, unlike VEGF-A, does not bind VEGFR-1. We therefore created VEGFR-1/VEGFR-2 chimeric proteins to further study receptor specificity. This biochemical analysis, together with our structural data, defined VEGFR-2 residues critical for the binding of VEGF-A and VEGF-C. Our results provide significant insights into the structural features that determine the high affinity and specificity of VEGF/VEGFR interactions.
Notes: 1091-6490
Leppänen, Veli-Matti
Prota, Andrea E
Jeltsch, Michael
Anisimov, Andrey
Kalkkinen, Nisse
Strandin, Tomas
Lankinen, Hilkka
Goldman, Adrian
Ballmer-Hofer, Kurt
Alitalo, Kari
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2010 Feb 9;107(6):2425-30. doi: 10.1073/pnas.0914318107.
Author Address: Molecular Cancer Biology Program, Biomedicum Helsinki, Department of Pathology, Haartman Institute and Helsinki University Central Hospital, PO Box 63, University of Helsinki, Haartmaninkatu 8, FI-00014 Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1573
Author: Leppänen, V. M., Tvorogov, D., Kisko, K., Prota, A. E., Jeltsch, M., Anisimov, A., Markovic-Mueller, S., Stuttfeld, E., Goldie, K. N., Ballmer-Hofer, K. and Alitalo, K.
Year: 2013
Title: Structural and mechanistic insights into VEGF receptor 3 ligand binding and activation
Journal: Proc Natl Acad Sci U S A
Volume: 110
Issue: 32
Pages: 12960-5
Epub Date: 2013/07/24
Date: Aug 6
Short Title: Structural and mechanistic insights into VEGF receptor 3 ligand binding and activation
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1301415110
PMCID: PMC3740881
Accession Number: 23878260
Keywords: Amino Acid Sequence
Binding Sites/genetics
Binding, Competitive
Crystallography, X-Ray
Electrophoresis, Polyacrylamide Gel
Humans
Ligands
Microscopy, Electron
Models, Molecular
Molecular Sequence Data
Multiprotein Complexes/chemistry/metabolism/ultrastructure
Mutation
Protein Binding
*Protein Multimerization
*Protein Structure, Tertiary
Scattering, Small Angle
Sequence Homology, Amino Acid
Thermodynamics
Vascular Endothelial Growth Factor C/*chemistry/genetics/metabolism
Vascular Endothelial Growth Factor Receptor-3/*chemistry/genetics/metabolism
X-Ray Diffraction
receptor tyrosine kinase
signal transduction
Abstract: Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are key drivers of blood and lymph vessel formation in development, but also in several pathological processes. VEGF-C signaling through VEGFR-3 promotes lymphangiogenesis, which is a clinically relevant target for treating lymphatic insufficiency and for blocking tumor angiogenesis and metastasis. The extracellular domain of VEGFRs consists of seven Ig homology domains; domains 1-3 (D1-3) are responsible for ligand binding, and the membrane-proximal domains 4-7 (D4-7) are involved in structural rearrangements essential for receptor dimerization and activation. Here we analyzed the crystal structures of VEGF-C in complex with VEGFR-3 domains D1-2 and of the VEGFR-3 D4-5 homodimer. The structures revealed a conserved ligand-binding interface in D2 and a unique mechanism for VEGFR dimerization and activation, with homotypic interactions in D5. Mutation of the conserved residues mediating the D5 interaction (Thr446 and Lys516) and the D7 interaction (Arg737) compromised VEGF-C induced VEGFR-3 activation. A thermodynamic analysis of VEGFR-3 deletion mutants showed that D3, D4-5, and D6-7 all contribute to ligand binding. A structural model of the VEGF-C/VEGFR-3 D1-7 complex derived from small-angle X-ray scattering data is consistent with the homotypic interactions in D5 and D7. Taken together, our data show that ligand-dependent homotypic interactions in D5 and D7 are essential for VEGFR activation, opening promising possibilities for the design of VEGFR-specific drugs.
Notes: 1091-6490
Leppänen, Veli-Matti
Tvorogov, Denis
Kisko, Kaisa
Prota, Andrea E
Jeltsch, Michael
Anisimov, Andrey
Markovic-Mueller, Sandra
Stuttfeld, Edward
Goldie, Kenneth N
Ballmer-Hofer, Kurt
Alitalo, Kari
268804/European Research Council/International
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2013 Aug 6;110(32):12960-5. doi: 10.1073/pnas.1301415110. Epub 2013 Jul 22.
Author Address: Translational Cancer Biology Program, Institute for Molecular Medicine Finland and Helsinki University Central Hospital, Biomedicum Helsinki, University of Helsinki, 00014 Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 632
Author: Lesage, F., Reyes, R., Fink, M., Duprat, F., Guillemare, E. and Lazdunski, M.
Year: 1996
Title: Dimerization of TWIK-1 K+ channel subunits via a disulfide bridge
Journal: Embo j
Volume: 15
Issue: 23
Pages: 6400-7
Epub Date: 1996/12/02
Date: Dec 2
Short Title: Dimerization of TWIK-1 K+ channel subunits via a disulfide bridge
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
PMCID: PMC452465
Accession Number: 8978667
Keywords: Amino Acid Sequence
Animals
Brain/*metabolism
Cell Line
Cell Membrane/physiology/ultrastructure
Diacetyl/analogs & derivatives/pharmacology
Dimerization
Dithiothreitol/pharmacology
Female
Glycosylation
Humans
Macromolecular Substances
Membrane Potentials/drug effects
Molecular Sequence Data
Myocardium/*metabolism
Oocytes/physiology
Potassium Channels/*biosynthesis/*chemistry/physiology
*Potassium Channels, Tandem Pore Domain
Protein Structure, Secondary
Recombinant Proteins/chemistry/metabolism
Spodoptera
Transfection
Xenopus
Abstract: TWIK-1 is a new type of K+ channel with two P domains and is abundantly expressed in human heart and brain. Here we show that TWIK-1 subunits can self-associate to give dimers containing an interchain disulfide bridge. This assembly involves a 34 amino acid domain that is localized to the extracellular M1P1 linker loop. Cysteine 69 which is part of this interacting domain is implicated in the formation of the disulfide bond. Replacing this cysteine with a serine residue results in the loss of functional K+ channel expression. This is the first example of a covalent association of functional subunits in voltage-sensitive channels via a disulfide bridge.
Notes: 1460-2075
Lesage, F
Reyes, R
Fink, M
Duprat, F
Guillemare, E
Lazdunski, M
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1996 Dec 2;15(23):6400-7.
Author Address: Institut de Pharmacologie Moléculaire et Cellulaire, CNRS, 660, Valbonne, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1955
Author: Leturcq, M., Mortuaire, M., Hardivillé, S., Schulz, C., Lefebvre, T. and Vercoutter-Edouart, A. S.
Year: 2018
Title: O-GlcNAc transferase associates with the MCM2-7 complex and its silencing destabilizes MCM-MCM interactions
Journal: Cell Mol Life Sci
Volume: 75
Issue: 23
Pages: 4321-4339
Epub Date: 2018/08/03
Date: Dec
Short Title: O-GlcNAc transferase associates with the MCM2-7 complex and its silencing destabilizes MCM-MCM interactions
Alternate Journal: Cellular and molecular life sciences : CMLS
ISSN: 1420-682X (Print)
1420-682x
DOI: 10.1007/s00018-018-2874-0
PMCID: PMC6208770
Accession Number: 30069701
Keywords: Blotting, Western
Cell Line, Tumor
Chromatin/*genetics/metabolism
*Gene Silencing
Glycosylation
HEK293 Cells
Humans
MCF-7 Cells
Minichromosome Maintenance Proteins/*genetics/metabolism
N-Acetylglucosaminyltransferases/*genetics/metabolism
Protein Binding
Protein Isoforms/genetics/metabolism
Protein Subunits/genetics/metabolism
beta-N-Acetylhexosaminidases/genetics/metabolism
Cell cycle
Chromatin
DNA replication
O-GlcNAcylation
Post-translational modification
Pre-replicative complex
Abstract: O-GlcNAcylation of proteins is governed by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). The homeostasis of O-GlcNAc cycling is regulated during cell cycle progression and is essential for proper cellular division. We previously reported the O-GlcNAcylation of the minichromosome maintenance proteins MCM2, MCM3, MCM6 and MCM7. These proteins belong to the MCM2-7 complex which is crucial for the initiation of DNA replication through its DNA helicase activity. Here we show that the six subunits of MCM2-7 are O-GlcNAcylated and that O-GlcNAcylation of MCM proteins mainly occurs in the chromatin-bound fraction of synchronized human cells. Moreover, we identify stable interaction between OGT and several MCM subunits. We also show that down-regulation of OGT decreases the chromatin binding of MCM2, MCM6 and MCM7 without affecting their steady-state level. Finally, OGT silencing or OGA inhibition destabilizes MCM2/6 and MCM4/7 interactions in the chromatin-enriched fraction. In conclusion, OGT is a new partner of the MCM2-7 complex and O-GlcNAcylation homeostasis might regulate MCM2-7 complex by regulating the chromatin loading of MCM6 and MCM7 and stabilizing MCM/MCM interactions.
Notes: 1420-9071
Leturcq, Maïté
Mortuaire, Marlène
Hardivillé, Stéphan
Schulz, Céline
Lefebvre, Tony
Vercoutter-Edouart, Anne-Sophie
Orcid: 0000-0002-3243-7122
Journal Article
Cell Mol Life Sci. 2018 Dec;75(23):4321-4339. doi: 10.1007/s00018-018-2874-0. Epub 2018 Aug 1.
Author Address: Univ. Lille, CNRS, UMR 8576, UGSF, Unité de Glycobiologie Structurale et Fonctionnelle, 59000, Lille, France.
Univ. Lille, CNRS, UMR 8576, UGSF, Unité de Glycobiologie Structurale et Fonctionnelle, 59000, Lille, France. anne-sophie.vercoutter@univ-lille.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1535
Author: Leung, M. C., Hitchen, P. G., Ward, D. G., Messer, A. E. and Marston, S. B.
Year: 2013
Title: Z-band alternatively spliced PDZ motif protein (ZASP) is the major O-linked β-N-acetylglucosamine-substituted protein in human heart myofibrils
Journal: J Biol Chem
Volume: 288
Issue: 7
Pages: 4891-8
Epub Date: 2012/12/29
Date: Feb 15
Short Title: Z-band alternatively spliced PDZ motif protein (ZASP) is the major O-linked β-N-acetylglucosamine-substituted protein in human heart myofibrils
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.410316
PMCID: PMC3576093
Accession Number: 23271734
Keywords: Acetylglucosamine/*chemistry
Adaptor Proteins, Signal Transducing/metabolism/*physiology
Alternative Splicing
Amino Acid Sequence
Antibodies, Monoclonal/chemistry
Fluorescent Antibody Technique, Indirect/methods
*Gene Expression Regulation
Heart/*physiology
Humans
LIM Domain Proteins/metabolism/*physiology
Microscopy, Fluorescence/methods
Molecular Sequence Data
Myofibrils/*metabolism
Peptides/chemistry
Signal Transduction
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: We studied O-linked β-N-acetylglucosamine (O-GlcNAc) modification of contractile proteins in human heart using SDS-PAGE and three detection methods: specific enzymatic conjugation of O-GlcNAc with UDP-N-azidoacetylgalactosamine (UDP-GalNAz) that is then linked to a tetramethylrhodamine fluorescent tag and CTD110.6 and RL2 monoclonal antibodies to O-GlcNAc. All three methods showed that O-GlcNAc modification was predominantly in a group of bands ~90 kDa that did not correspond to any of the major myofibrillar proteins. MALDI-MS/MS identified the 90-kDa band as the protein ZASP (Z-band alternatively spliced PDZ motif protein), a minor component of the Z-disc (about 1 per 400 α-actinin) important for myofibrillar development and mechanotransduction. This was confirmed by the co-localization of O-GlcNAc and ZASP in Western blotting and by immunofluorescence microscopy. O-GlcNAcylation of ZASP increased in diseased heart, being 49 ± 5% of all O-GlcNAc in donor, 68 ± 9% in end-stage failing heart, and 76 ± 6% in myectomy muscle samples (donor versus myectomy p < 0.05). ZASP is only 22% of all O-GlcNAcylated proteins in mouse heart myofibrils.
Notes: 1083-351x
Leung, Man-Ching
Hitchen, Paul G
Ward, Douglas G
Messer, Andrew E
Marston, Steven B
PG/08/077/25587/British Heart Foundation/United Kingdom
RG/11/20/29266/British Heart Foundation/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2013 Feb 15;288(7):4891-8. doi: 10.1074/jbc.M112.410316. Epub 2012 Dec 27.
Author Address: Myocardial Function Section, National Heart and Lung Institute, Imperial College London, London W12 0NN, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1918
Author: Levine, Z. G., Fan, C., Melicher, M. S., Orman, M., Benjamin, T. and Walker, S.
Year: 2018
Title: O-GlcNAc Transferase Recognizes Protein Substrates Using an Asparagine Ladder in the Tetratricopeptide Repeat (TPR) Superhelix
Journal: J Am Chem Soc
Volume: 140
Issue: 10
Pages: 3510-3513
Epub Date: 2018/02/28
Date: Mar 14
Short Title: O-GlcNAc Transferase Recognizes Protein Substrates Using an Asparagine Ladder in the Tetratricopeptide Repeat (TPR) Superhelix
Alternate Journal: Journal of the American Chemical Society
ISSN: 0002-7863 (Print)
0002-7863
DOI: 10.1021/jacs.7b13546
PMCID: PMC5937710
NIHMSID: NIHMS951734
Accession Number: 29485866
Keywords: Asparagine/*metabolism
Humans
N-Acetylglucosaminyltransferases/*metabolism
*Protein Array Analysis
Proteins/*chemistry/*metabolism
Substrate Specificity
*Tetratricopeptide Repeat
Abstract: The essential mammalian enzyme O-GlcNAc Transferase (OGT) is uniquely responsible for transferring N-acetylglucosamine to over a thousand nuclear and cytoplasmic proteins, yet there is no known consensus sequence and it remains unclear how OGT recognizes its substrates. To address this question, we developed a protein microarray assay that chemoenzymatically labels de novo sites of glycosylation with biotin, allowing us to simultaneously assess OGT activity across >6000 human proteins. With this assay we examined the contribution to substrate selection of a conserved asparagine ladder within the lumen of OGT's superhelical tetratricopeptide repeat (TPR) domain. When five asparagines were mutated, OGT retained significant activity against short peptides, but showed limited limited glycosylation of protein substrates on the microarray. O-GlcNAcylation of protein substrates in cell extracts was also greatly attenuated. We conclude that OGT recognizes the majority of its substrates by binding them to the asparagine ladder in the TPR lumen proximal to the catalytic domain.
Notes: 1520-5126
Levine, Zebulon G
Fan, Chenguang
Melicher, Michael S
Orman, Marina
Benjamin, Tania
Walker, Suzanne
Orcid: 0000-0002-0545-914x
R01 GM094263/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Am Chem Soc. 2018 Mar 14;140(10):3510-3513. doi: 10.1021/jacs.7b13546. Epub 2018 Mar 5.
Author Address: Department of Microbiology and Immunobiology , Harvard Medical School , Boston , Massachusetts 02115 , United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1015
Author: Lewandrowski, U., Moebius, J., Walter, U. and Sickmann, A.
Year: 2006
Title: Elucidation of N-glycosylation sites on human platelet proteins: a glycoproteomic approach
Journal: Mol Cell Proteomics
Volume: 5
Issue: 2
Pages: 226-33
Epub Date: 2005/11/03
Date: Feb
Short Title: Elucidation of N-glycosylation sites on human platelet proteins: a glycoproteomic approach
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M500324-MCP200
Accession Number: 16263699
Keywords: Amino Acid Sequence
Epidermal Growth Factor
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
P-Selectin/chemistry
Platelet Membrane Glycoproteins/*chemistry/*metabolism
*Proteomics
Thrombospondin 1/chemistry/metabolism
Abstract: Among known platelet proteins, a prominent and functionally important group is represented by glycoprotein isoforms. They account e.g. for secretory proteins and plasma membrane receptors including integrins and glycoprotein VI as well as intracellular components of cytosol and organelles including storage proteins (multimerin 1 etc.). Although many of those proteins have been studied for some time with regard to their function, little attention has been paid with respect to their glycosylation sites. Here we report the analysis of N-glycosylation sites of human platelet proteins. For the enrichment of glycopeptides, lectin affinity chromatography as well as chemical trapping of protein bound oligosaccharides was used. Therefore, concanavalin A was used for specific interaction with carbohydrate species along with periodic acid oxidation and hydrazide bead trapping of glycosylated proteins. Derivatization by peptide:N-glycosidase F yielded deglycosylated peptides, which provided the basis for the elucidation of proteins and their sites of modification. Using both methods in combination with nano-LC-ESI-MS/MS analysis 70 different glycosylation sites within 41 different proteins were identified. Comparison with the Swiss-Prot database established that the majority of these 70 sites have not been specifically determined by previous research projects. With this approach including hydrazide bead affinity trapping, the immunoglobulin receptor G6f, which is known to couple to the Ras-mitogen-activated protein kinase pathway in the immune system, was shown here for the first time to be present in human platelets.
Notes: Lewandrowski, Urs
Moebius, Jan
Walter, Ulrich
Sickmann, Albert
Journal Article
United States
Mol Cell Proteomics. 2006 Feb;5(2):226-33. doi: 10.1074/mcp.M500324-MCP200. Epub 2005 Oct 31.
Author Address: Protein Mass Spectrometry and Functional Proteomics Group, Rudolf Virchow Center for Experimental Biomedicine, Versbacher Strasse 9, 97078 Wuerzburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1777
Author: Lewis, B. A., Burlingame, A. L. and Myers, S. A.
Year: 2016
Title: Human RNA Polymerase II Promoter Recruitment in Vitro Is Regulated by O-Linked N-Acetylglucosaminyltransferase (OGT)
Journal: J Biol Chem
Volume: 291
Issue: 27
Pages: 14056-14061
Epub Date: 2016/04/30
Date: Jul 1
Short Title: Human RNA Polymerase II Promoter Recruitment in Vitro Is Regulated by O-Linked N-Acetylglucosaminyltransferase (OGT)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M115.684365
PMCID: PMC4933165
Accession Number: 27129214
Keywords: Humans
In Vitro Techniques
N-Acetylglucosaminyltransferases/*metabolism
*Promoter Regions, Genetic
RNA Polymerase II/*genetics
O-GlcNAcylation
O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
RNA polymerase II
gene regulation
transcription
Abstract: Although the O-linked N-acetylglucosamine (O-GlcNAc) modification of the RNA polymerase II C-terminal domain was described 20 years ago, the function of this RNA polymerase II (pol II) species is not known. We show here that an O-GlcNAcylated pol II species (pol IIγ) exists on promoters in vitro Inhibition of O-GlcNAc-transferase activity and O-GlcNAcylation prevents pol II entry into the promoter, and O-GlcNAc removal from pol II is an ATP-dependent step during initiation. These data indicate that O-GlcNAc-transferase activity is essential for RNA pol II promoter recruitment and that pol II goes through a cycling of O-GlcNAcylation at the promoter. Mass spectrometry shows that serine residues 2 and 5 of the pol II C-terminal domain are O-GlcNAcylated, suggesting an overlap with the transcription factor IIH (TFIIH)-dependent serine 5 phosphorylation events during initiation and P-TEFb (positive transcriptional elongation factor b) events during elongation. These data provide unexpected and important insights into the role of a previously ill-defined species of RNA polymerase II in regulating transcription.
Notes: 1083-351x
Lewis, Brian A
Burlingame, Alma L
Myers, Samuel A
P41 GM103481/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2016 Jul 1;291(27):14056-14061. doi: 10.1074/jbc.M115.684365. Epub 2016 Apr 27.
Author Address: Transcriptional Regulation and Biochemistry Unit, Metabolism Branch, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892. Electronic address: lewisbri@mail.nih.gov.
Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94158.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 834
Author: Li, H., Ryan, T. J., Chave, K. J. and Van Roey, P.
Year: 2002
Title: Three-dimensional structure of human gamma -glutamyl hydrolase. A class I glatamine amidotransferase adapted for a complex substate
Journal: J Biol Chem
Volume: 277
Issue: 27
Pages: 24522-9
Epub Date: 2002/04/16
Date: Jul 5
Short Title: Three-dimensional structure of human gamma -glutamyl hydrolase. A class I glatamine amidotransferase adapted for a complex substate
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M202020200
Accession Number: 11953431
Keywords: Amino Acid Sequence
Catalysis
Crystallography, X-Ray
Humans
Models, Molecular
Protein Conformation
gamma-Glutamyl Hydrolase/*chemistry
Abstract: gamma-Glutamyl hydrolase catalyzes the cleavage of the gamma-glutamyl chain of folylpoly-gamma-glutamyl substrates and is a central enzyme in folyl and antifolyl poly-gamma-glutamate metabolism. The crystal structure of human gamma-glutamyl hydrolase, determined at 1.6-A resolution, reveals that the protein is a homodimer. The overall structure of human gamma-glutamyl hydrolase contains 11 alpha-helices and 14 beta-strands, with a fold in which a central eight-stranded beta-sheet is sandwiched by three and five alpha-helices on each side. The topology is very similar to that of the class I glutamine amidotransferase domains, with the only major differences consisting of extensions in four loops and at the C terminus. These insertions are important for defining the substrate binding cleft and/or the dimer interface. Two sequence motifs are found in common between human gamma-glutamyl hydrolase and the class I glutamine amidotransferase family and include the catalytically essential residues, Cys-110 and His-220. These residues are located in the center of a large l-shaped cleft that is closed at one end and open at the other. Several conserved residues, including Glu-114, His-171, Gln-218, and Lys-223, may be important for substrate binding. Modeling of a methotrexate thioester intermediate, based on the corresponding complex of the glutamate thioester intermediate of Escherichia coli carbamoyl-phosphate synthetase, indicates that the substrate binds in an orientation with the pteroyl group toward the open end of the cleft.
Notes: Li, Hongmin
Ryan, Thomas J
Chave, Karen J
Van Roey, Patrick
CA82425/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2002 Jul 5;277(27):24522-9. doi: 10.1074/jbc.M202020200. Epub 2002 Apr 12.
Author Address: Division of Molecular Medicine, Wadsworth Center, Albany, New York 12201-0509, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 754
Author: Li, J., Quilty, J., Popov, M. and Reithmeier, R. A.
Year: 2000
Title: Processing of N-linked oligosaccharide depends on its location in the anion exchanger, AE1, membrane glycoprotein
Journal: Biochem J
Volume: 349
Issue: Pt 1
Pages: 51-7
Epub Date: 2000/06/22
Date: Jul 1
Short Title: Processing of N-linked oligosaccharide depends on its location in the anion exchanger, AE1, membrane glycoprotein
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/0264-6021:3490051
PMCID: PMC1221119
Accession Number: 10861210
Keywords: Animals
Anions
Antiporters/*metabolism
Binding Sites
Biotinylation
COS Cells
Cell Line
Cell Membrane/metabolism
Chloride-Bicarbonate Antiporters
Electrophoresis, Polyacrylamide Gel
Endoplasmic Reticulum/metabolism
Glycosylation
Humans
Immunoblotting
Lectins/metabolism
Mutagenesis
Oligosaccharides/*metabolism
Plasmids/metabolism
Protein Binding
Protein Isoforms
Protein Structure, Tertiary
Sequence Analysis, DNA
Time Factors
Transfection
Abstract: The human erythrocyte anion exchanger (AE)1 (Band 3) contains a single complex N-linked oligosaccharide that is attached to Asn(642) in the fourth extracellular loop of this polytopic membrane protein, while other isoforms (AE2, AE3 and trout AE1) are N-glycosylated on the preceding extracellular loop. Human AE1 expressed in transfected human embryonic kidney (HEK)-293 or COS-7 cells contained a high-mannose oligosaccharide. The lack of oligosaccharide processing was not due to retention of AE1 in the endoplasmic reticulum since biotinylation assays showed that approx. 30% of the protein was expressed at the cell surface. Moving the N-glycosylation site to the preceding extracellular loop in an AE1 glycosylation mutant (N555) resulted in processing of the oligosaccharide and production of a complex form of AE1. A double N-glycosylation mutant (N555/N642) contained both a high-mannose and a complex oligosaccharide chain. The complex form of the N555 mutant could be biotinylated showing that this form of the glycoprotein was at the cell surface. Pulse-chase experiments showed that the N555 mutant was efficiently converted from a high-mannose to a complex oligosaccharide with a half-time of approx. 4 h, which reflected the time course of trafficking of AE1 from the endoplasmic reticulum to the plasma membrane. The turnover of the complex form of the N555 mutant occurred with a half-life of approx. 15 h. The results show that the oligosaccharide attached to the endogenous site in extracellular loop 4 in human AE1 is not processed in HEK-293 or COS-7 cells, while the oligosaccharide attached to the preceding loop is converted into the complex form.
Notes: 1470-8728
Li, J
Quilty, J
Popov, M
Reithmeier, R A
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2000 Jul 1;349(Pt 1):51-7. doi: 10.1042/0264-6021:3490051.
Author Address: Medical Research Council Group in Membrane Biology, Department of Medicine, Room 7344, Medical Sciences Building, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1127
Author: Li, J. G., Chen, C. and Liu-Chen, L. Y.
Year: 2007
Title: N-Glycosylation of the human kappa opioid receptor enhances its stability but slows its trafficking along the biosynthesis pathway
Journal: Biochemistry
Volume: 46
Issue: 38
Pages: 10960-70
Epub Date: 2007/08/23
Date: Sep 25
Short Title: N-Glycosylation of the human kappa opioid receptor enhances its stability but slows its trafficking along the biosynthesis pathway
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi700443j
Accession Number: 17711303
Keywords: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide,
(trans)-Isomer/pharmacology
Amino Acid Substitution
Analgesics, Non-Narcotic/pharmacology
Animals
CHO Cells
Cricetinae
Cricetulus
Diprenorphine/pharmacology
Down-Regulation
Electrophoresis, Polyacrylamide Gel
*Gene Expression Regulation/physiology
Glycoside Hydrolases/antagonists & inhibitors/metabolism
Glycosylation
Guanosine 5'-O-(3-Thiotriphosphate)/metabolism
Humans
Immunoprecipitation
Kinetics
Mutagenesis, Site-Directed
*Protein Processing, Post-Translational/physiology
Protein Structure, Tertiary
Protein Transport/physiology
Receptors, Cell Surface/biosynthesis/*metabolism
Receptors, Opioid, kappa/biosynthesis/*metabolism
Recombinant Fusion Proteins/drug effects/metabolism
Signal Transduction
Sulfur Radioisotopes
Tritium
Tunicamycin/pharmacology
Abstract: We examined glycosylation of FLAG-hKOR expressed in CHO cells and determined its functional significance. FLAG-hKOR was resolved as a broad and diffuse 55-kDa band and a less diffuse 45-kDa band by immunoblotting, indicating that the receptor is glycosylated. Endoglycosidase H cleaved the 45-kDa band to approximately 38 kDa but did not change the 55-kDa band, demonstrating that the 45-kDa band is N-glycosylated with high-mannose or hybrid-type glycan. Peptide-N-glycosidase F digestion of solubilized hKOR or incubation of cells with tunicamycin resulted in two species of 43 and 38 kDa, suggesting that the 43-kDa band is O-glycosylated. FLAG-hKOR was reduced to lower Mr bands by neuraminidase and O-glycosidase, indicating that the hKOR contains O-linked glycan. Mutation of Asn25 or Asn39 to Gln in the N-terminal domain reduced the Mr by approximately 5 kDa, indicating that both residues were glycosylated. The double mutant hKOR-N25/39Q was resolved as a 43-kDa (mature form) and a 38-kDa (intermediate form) band. When transiently expressed, hKOR-N25/39Q had a lower expression level than the wild type. In CHO cells stably expressing the hKOR-N25/39Q, pulse-chase experiments revealed that the turnover rate constants (ke) of the intermediate and mature forms were approximately 3 times those of the wild type. In addition, the maturation rate constant (ka) of the 43-kDa form of hKOR-N25/39Q was 6 times that of the mature form of the wild type. The hKOR-N25/39Q mutant showed increased agonist-induced receptor phosphorylation, desensitization, internalization, and downregulation, without changing ligand binding affinity or receptor-G protein coupling. Thus, N-glycosylation of the hKOR plays important roles in stability and trafficking along the biosynthesis pathway of the receptor protein as well as agonist-induced receptor regulation.
Notes: Li, Jian-Guo
Chen, Chongguang
Liu-Chen, Lee-Yuan
DA04745/DA/NIDA NIH HHS/United States
DA17302/DA/NIDA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
Biochemistry. 2007 Sep 25;46(38):10960-70. doi: 10.1021/bi700443j. Epub 2007 Aug 21.
Author Address: Department of Pharmacology and Center for Substance Abuse Research, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1545
Author: Li, M. D., Ruan, H. B., Hughes, M. E., Lee, J. S., Singh, J. P., Jones, S. P., Nitabach, M. N. and Yang, X.
Year: 2013
Title: O-GlcNAc signaling entrains the circadian clock by inhibiting BMAL1/CLOCK ubiquitination
Journal: Cell Metab
Volume: 17
Issue: 2
Pages: 303-10
Epub Date: 2013/02/12
Date: Feb 5
Short Title: O-GlcNAc signaling entrains the circadian clock by inhibiting BMAL1/CLOCK ubiquitination
Alternate Journal: Cell metabolism
ISSN: 1550-4131 (Print)
1550-4131
DOI: 10.1016/j.cmet.2012.12.015
PMCID: PMC3647362
NIHMSID: NIHMS434108
Accession Number: 23395176
Keywords: ARNTL Transcription Factors/genetics/*metabolism
Acetylglucosamine/*metabolism
Animals
CLOCK Proteins/genetics/*metabolism
Cell Line, Tumor
*Circadian Clocks/genetics
Gene Expression Regulation
Glucose/metabolism
Homeostasis
Humans
Liver/enzymology
Mice
N-Acetylglucosaminyltransferases/metabolism
Protein Stability
*Signal Transduction/genetics
*Ubiquitination/genetics
Abstract: Circadian clocks are coupled to metabolic oscillations through nutrient-sensing pathways. Nutrient flux into the hexosamine biosynthesis pathway triggers covalent protein modification by O-linked β-D-N-acetylglucosamine (O-GlcNAc). Here we show that the hexosamine/O-GlcNAc pathway modulates peripheral clock oscillation. O-GlcNAc transferase (OGT) promotes expression of BMAL1/CLOCK target genes and affects circadian oscillation of clock genes in vitro and in vivo. Both BMAL1 and CLOCK are rhythmically O-GlcNAcylated, and this protein modification stabilizes BMAL1 and CLOCK by inhibiting their ubiquitination. In vivo analysis of genetically modified mice with perturbed hepatic OGT expression shows aberrant circadian rhythms of glucose homeostasis. These results establish the counteraction between O-GlcNAcylation and ubiquitination as a key mechanism that regulates the circadian clock and suggest a crucial role for O-GlcNAc signaling in transducing nutritional signals to the core circadian timing machinery.
Notes: 1932-7420
Li, Min-Dian
Ruan, Hai-Bin
Hughes, Michael E
Lee, Jeong-Sang
Singh, Jay P
Jones, Steven P
Nitabach, Michael N
Yang, Xiaoyong
R01 DK089098/DK/NIDDK NIH HHS/United States
P20 RR024489/RR/NCRR NIH HHS/United States
R01 NS056443/NS/NINDS NIH HHS/United States
R01 HL094419/HL/NHLBI NIH HHS/United States
R21 NS058330/NS/NINDS NIH HHS/United States
R01 NS083875/NS/NINDS NIH HHS/United States
P01 DK057751/DK/NIDDK NIH HHS/United States
F32GM096577/GM/NIGMS NIH HHS/United States
R01 HL083320/HL/NHLBI NIH HHS/United States
P01 HL078825/HL/NHLBI NIH HHS/United States
R01 GM098931/GM/NIGMS NIH HHS/United States
F32 GM096577/GM/NIGMS NIH HHS/United States
R01 NS055035/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell Metab. 2013 Feb 5;17(2):303-10. doi: 10.1016/j.cmet.2012.12.015.
Author Address: Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, CT 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1465
Author: Li, M. D., Ruan, H. B., Singh, J. P., Zhao, L., Zhao, T., Azarhoush, S., Wu, J., Evans, R. M. and Yang, X.
Year: 2012
Title: O-GlcNAc transferase is involved in glucocorticoid receptor-mediated transrepression
Journal: J Biol Chem
Volume: 287
Issue: 16
Pages: 12904-12
Epub Date: 2012/03/01
Date: Apr 13
Short Title: O-GlcNAc transferase is involved in glucocorticoid receptor-mediated transrepression
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.303792
PMCID: PMC3339970
Accession Number: 22371499
Keywords: Acetylglucosamine/metabolism
Anti-Inflammatory Agents/metabolism/pharmacology
Apoptosis/physiology
Cell Line, Tumor
Drug Design
Gene Expression Regulation, Enzymologic/drug effects/*physiology
HEK293 Cells
Humans
Hydrocortisone/metabolism/pharmacology
Lung Neoplasms
N-Acetylglucosaminyltransferases/*genetics/*metabolism
NF-kappa B/metabolism
Nuclear Receptor Coactivator 2/metabolism
RNA Polymerase II/metabolism
Receptors, Cytoplasmic and Nuclear/metabolism
Receptors, Glucocorticoid/*genetics/*metabolism
Signal Transduction/physiology
Abstract: Recruitment of O-GlcNAc transferase (OGT) to promoters plays an important role in gene repression. Glucocorticoid signaling represses the transcriptional activities of NF-κB and AP-1 through direct binding, yet the molecular mechanisms remain to be elucidated. Here we report that OGT is an important component of GR-mediated transrepression. OGT associates with ligand-bound GR in a multi-protein repression complex. Overexpression of OGT potentiates the GR transrepression pathway, whereas depletion of endogenous OGT by RNA interference abolishes the repression. The recruitment of OGT by GR leads to increased O-GlcNAcylation and decreased phosphorylation of RNA polymerase II on target genes. Functionally, overexpression of OGT enhances glucocorticoid-induced apoptosis in resistant cell lines while knockdown of OGT prevents sensitive cell lines from apoptosis. These studies identify a molecular mechanism of GR transrepression, and highlight the function of O-GlcNAc in hormone signaling.
Notes: 1083-351x
Li, Min-Dian
Ruan, Hai-Bin
Singh, Jay P
Zhao, Lin
Zhao, Tingting
Azarhoush, Sascha
Wu, Jing
Evans, Ronald M
Yang, Xiaoyong
R01 DK089098/DK/NIDDK NIH HHS/United States
R01 HL105278/HL/NHLBI NIH HHS/United States
DK062434/DK/NIDDK NIH HHS/United States
DK089098/DK/NIDDK NIH HHS/United States
U19 DK062434/DK/NIDDK NIH HHS/United States
Howard Hughes Medical Institute/United States
CA014195-38/CA/NCI NIH HHS/United States
F32 DK066971/DK/NIDDK NIH HHS/United States
HL105278/HL/NHLBI NIH HHS/United States
DK066971/DK/NIDDK NIH HHS/United States
P30 CA014195/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Apr 13;287(16):12904-12. doi: 10.1074/jbc.M111.303792. Epub 2012 Feb 27.
Author Address: Department of Cellular and Molecular Physiology, Section of Comparative Medicine and Program in Integrative Cell Signaling and Neurobiology of Metabolism, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1839
Author: Li, Q. Q., Hao, J. J., Zhang, Z., Krane, L. S., Hammerich, K. H., Sanford, T., Trepel, J. B., Neckers, L. and Agarwal, P. K.
Year: 2017
Title: Proteomic analysis of proteome and histone post-translational modifications in heat shock protein 90 inhibition-mediated bladder cancer therapeutics
Journal: Sci Rep
Volume: 7
Issue: 1
Pages: 201
Epub Date: 2017/03/17
Date: Mar 15
Short Title: Proteomic analysis of proteome and histone post-translational modifications in heat shock protein 90 inhibition-mediated bladder cancer therapeutics
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/s41598-017-00143-6
PMCID: PMC5427839
Accession Number: 28298630
Keywords: Antineoplastic Agents/*pharmacology
Benzamides/pharmacology
Benzodioxoles/pharmacology
Cell Line, Tumor
Cell Proliferation/drug effects
Cell Survival/drug effects
Dose-Response Relationship, Drug
Gene Expression Regulation, Neoplastic/drug effects
HSP90 Heat-Shock Proteins/*antagonists & inhibitors
Heterocyclic Compounds, 4 or More Rings/pharmacology
Histones/*metabolism
Humans
Imidazoles/pharmacology
Isoindoles/pharmacology
Isoxazoles/pharmacology
Protein Processing, Post-Translational
Proteomics/*methods
Resorcinols/pharmacology
Triazoles/pharmacology
Urinary Bladder Neoplasms/drug therapy/*metabolism
Abstract: Heat shock protein 90 (HSP90) inhibition is an attractive strategy for cancer treatment. Several HSP90 inhibitors have shown promising effects in clinical oncology trials. However, little is known about HSP90 inhibition-mediated bladder cancer therapy. Here, we report a quantitative proteomic study that evaluates alterations in protein expression and histone post-translational modifications (PTMs) in bladder carcinoma in response to HSP90 inhibition. We show that 5 HSP90 inhibitors (AUY922, ganetespib, SNX2112, AT13387, and CUDC305) potently inhibited the proliferation of bladder cancer 5637 cells in a dose- and time-dependent manner. Our proteomic study quantified 518 twofold up-regulated and 811 twofold down-regulated proteins common to both AUY922 and ganetespib treatment. Bioinformatic analyses revealed that those differentially expressed proteins were involved in multiple cellular processes and enzyme-regulated signaling pathways, including chromatin modifications and cell death-associated pathways. Furthermore, quantitative proteome studies identified 14 types of PTMs with 93 marks on the core histones, including 34 novel histone marks of butyrylation, citrullination, 2-hydroxyisobutyrylation, methylation, O-GlcNAcylation, propionylation, and succinylation in AUY922- and ganetespib-treated 5637 cells. Together, this study outlines the association between proteomic changes and histone PTMs in response to HSP90 inhibitor treatment in bladder carcinoma cells, and thus intensifies the understanding of HSP90 inhibition-mediated bladder cancer therapeutics.
Notes: 2045-2322
Li, Qingdi Quentin
Hao, Jian-Jiang
Zhang, Zheng
Krane, L Spencer
Hammerich, Kai H
Sanford, Thomas
Trepel, Jane B
Neckers, Len
Agarwal, Piyush K
Journal Article
Research Support, N.I.H., Intramural
Sci Rep. 2017 Mar 15;7(1):201. doi: 10.1038/s41598-017-00143-6.
Author Address: Urologic Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, 20892, USA. liquenti@mail.nih.gov.
Poochon Scientific, Frederick, Maryland, 21701, USA.
Urologic Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, 20892, USA.
Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, 20892, USA. trepelj@mail.nih.gov.
Urologic Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, 20892, USA. agarwalpk2@mail.nih.gov.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1758
Author: Li, S., Zhu, H., Wang, J., Wang, X., Li, X., Ma, C., Wen, L., Yu, B., Wang, Y., Li, J. and Wang, P. G.
Year: 2016
Title: Comparative analysis of Cu (I)-catalyzed alkyne-azide cycloaddition (CuAAC) and strain-promoted alkyne-azide cycloaddition (SPAAC) in O-GlcNAc proteomics
Journal: Electrophoresis
Volume: 37
Issue: 11
Pages: 1431-6
Epub Date: 2016/02/09
Date: Jun
Short Title: Comparative analysis of Cu (I)-catalyzed alkyne-azide cycloaddition (CuAAC) and strain-promoted alkyne-azide cycloaddition (SPAAC) in O-GlcNAc proteomics
Alternate Journal: Electrophoresis
ISSN: 0173-0835 (Print)
0173-0835
DOI: 10.1002/elps.201500491
PMCID: PMC5967854
NIHMSID: NIHMS968488
Accession Number: 26853435
Keywords: A549 Cells
Acetylglucosamine/*chemistry
Alkynes/chemistry
Azides/chemistry
Catalysis
Click Chemistry
Copper/chemistry
Cycloaddition Reaction
Electrophoresis, Polyacrylamide Gel
Humans
Mass Spectrometry
*Protein Processing, Post-Translational
Proteomics/*methods
*Bioorthogonal chemistry
*Biotin-DIBO-Alkyne
*Biotin-Diazo-Alkyne
*O-GlcNAc
*Proteomics
Abstract: O-linked β-N-acetylglucosamine (O-GlcNAc) is emerging as an essential protein post-translational modification in a range of organisms. It is involved in various cellular processes such as nutrient sensing, protein degradation, gene expression, and is associated with many human diseases. Despite its importance, identifying O-GlcNAcylated proteins is a major challenge in proteomics. Here, using peracetylated N-azidoacetylglucosamine (Ac4 GlcNAz) as a bioorthogonal chemical handle, we described a gel-based mass spectrometry method for the identification of proteins with O-GlcNAc modification in A549 cells. In addition, we made a labeling efficiency comparison between two modes of azide-alkyne bioorthogonal reactions in click chemistry: copper-catalyzed azide-alkyne cycloaddition (CuAAC) with Biotin-Diazo-Alkyne and stain-promoted azide-alkyne cycloaddition (SPAAC) with Biotin-DIBO-Alkyne. After conjugation with click chemistry in vitro and enrichment via streptavidin resin, proteins with O-GlcNAc modification were separated by SDS-PAGE and identified with mass spectrometry. Proteomics data analysis revealed that 229 putative O-GlcNAc modified proteins were identified with Biotin-Diazo-Alkyne conjugated sample and 188 proteins with Biotin-DIBO-Alkyne conjugated sample, among which 114 proteins were overlapping. Interestingly, 74 proteins identified from Biotin-Diazo-Alkyne conjugates and 46 verified proteins from Biotin-DIBO-Alkyne conjugates could be found in the O-GlcNAc modified proteins database dbOGAP (http://cbsb.lombardi.georgetown.edu/hulab/OGAP.html). These results suggested that CuAAC with Biotin-Diazo-Alkyne represented a more powerful method in proteomics with higher protein identification and better accuracy compared to SPAAC. The proteomics credibility was also confirmed by the molecular function and cell component gene ontology (GO). Together, the method we reported here combining metabolic labeling, click chemistry, affinity-based enrichment, SDS-PAGE separation, and mass spectrometry, would be adaptable for other post-translationally modified proteins in proteomics.
Notes: 1522-2683
Li, Shanshan
Zhu, He
Wang, Jiajia
Wang, Xiaomin
Li, Xu
Ma, Cheng
Wen, Liuqing
Yu, Bingchen
Wang, Yuehua
Li, Jing
Wang, Peng George
R01 GM085267/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Electrophoresis. 2016 Jun;37(11):1431-6. doi: 10.1002/elps.201500491. Epub 2016 Mar 1.
Author Address: Department of Chemistry and Center of Diagnostics & Therapeutics, Georgia State University, Atlanta, GA, USA.
State Key Laboratory of Medicinal Chemical Biology, College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin, P. R. China.
Department of Pathology, Shanxi Tumor Hospital, Taiyuan, P. R. China.
State Key Laboratory of Medicinal Chemical Biology, College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin, P. R. China. pwang11@gsu.edu.
Department of Chemistry and Center of Diagnostics & Therapeutics, Georgia State University, Atlanta, GA, USA. pwang11@gsu.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 987
Author: Li, S. H., Lee, R. K., Hsiao, Y. L. and Chen, Y. H.
Year: 2005
Title: Demonstration of a glycoprotein derived from the Ceacam10 gene in mouse seminal vesicle secretions
Journal: Biol Reprod
Volume: 73
Issue: 3
Pages: 546-53
Epub Date: 2005/05/20
Date: Sep
Short Title: Demonstration of a glycoprotein derived from the Ceacam10 gene in mouse seminal vesicle secretions
Alternate Journal: Biology of reproduction
ISSN: 0006-3363 (Print)
0006-3363
DOI: 10.1095/biolreprod.105.039651
PMCID: PMC7109865
Accession Number: 15901639
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cell Adhesion Molecules/*chemistry/genetics/isolation & purification/*physiology
Epithelial Cells/metabolism
Gene Expression Regulation, Developmental
Male
Mice
Molecular Sequence Data
RNA, Messenger
Seminal Vesicles/cytology/*metabolism
Sperm Motility/physiology
Spermatozoa/physiology
Abstract: CEACAM10 was purified from mouse seminal vesicle secretions by a series of purification steps that included ion exchange chromatography on a DEAE-Sephacel column and ion exchange high-performance liquid chromatography on a sulfopropyl column. It was shown to be a 36-kDa glycoprotein with an N-linked carbohydrate moiety. The circular dichromoism spectrum of CEACAM10 in 50 mM phosphate buffer at pH 7.4 appeared as one negative band arising from the beta form at 217 nm. CEACAM10 was expressed predominantly in seminal vesicles of adult mice. Both CEACAM10 and its mRNA were demonstrated on the luminal epithelium of the mucosal folds in the seminal vesicle. The amount of Ceacam10 mRNA in the seminal vesicle was correlated with the stage of animal maturation. Castration of adult mice resulted in cessation of Ceacam10 expression, while treatment of castrated mice with testosterone propionate in corn oil restored Ceacam10 expression in the seminal vesicle. During the entire course of pregnancy, Ceacam10 might be silent in the embryo. A cytochemical study illustrated the presence of the CEACAM10 binding region on the entire surface of mouse sperm. CEACAM10-sperm binding greatly enhanced sperm motility in vitro.
Notes: 1529-7268
Li, Sheng-Hsiang
Lee, Robert Kuo-Kuang
Hsiao, Ya-Ling
Chen, Yee-Hsiung
Journal Article
Research Support, Non-U.S. Gov't
Biol Reprod. 2005 Sep;73(3):546-53. doi: 10.1095/biolreprod.105.039651. Epub 2005 May 18.
Author Address: Department of Medical Research, Mackay Memorial Hospital, Tamshui 251, Taiwan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1989
Author: Li, T., Li, X., Attri, K. S., Liu, C., Li, L., Herring, L. E., Asara, J. M., Lei, Y. L., Singh, P. K., Gao, C. and Wen, H.
Year: 2018
Title: O-GlcNAc Transferase Links Glucose Metabolism to MAVS-Mediated Antiviral Innate Immunity
Journal: Cell Host Microbe
Volume: 24
Issue: 6
Pages: 791-803.e6
Epub Date: 2018/12/14
Date: Dec 12
Short Title: O-GlcNAc Transferase Links Glucose Metabolism to MAVS-Mediated Antiviral Innate Immunity
Alternate Journal: Cell host & microbe
ISSN: 1931-3128 (Print)
1931-3128
DOI: 10.1016/j.chom.2018.11.001
PMCID: PMC6296827
NIHMSID: NIHMS1511884
Accession Number: 30543776
Keywords: Adaptor Proteins, Signal Transducing/genetics/*metabolism
Animals
Chlorocebus aethiops
Glucose/metabolism
HEK293 Cells
HT29 Cells
Hexosamines/metabolism
Humans
Immunity, Innate/*immunology
Mice
Mice, Inbred C57BL
N-Acetylglucosaminyltransferases/genetics/*metabolism
Rhabdoviridae Infections/*immunology/virology
THP-1 Cells
Vero Cells
Vesicular stomatitis Indiana virus/*immunology
*Mavs
*O-GlcNAc transferase
*antiviral immunity
*glucose metabolism
*hexosamine biosynthesis pathway (HBP)
Abstract: Increased glucose metabolism in immune cells not only serves as a hallmark feature of acute inflammation but also profoundly affects disease outcome following bacterial infection and tissue damage. However, the role of individual glucose metabolic pathways during viral infection remains largely unknown. Here we demonstrate an essential function of the hexosamine biosynthesis pathway (HBP)-associated O-linked β-N-acetylglucosamine (O-GlcNAc) signaling in promoting antiviral innate immunity. Challenge of macrophages with vesicular stomatitis viruses (VSVs) enhances HBP activity and downstream protein O-GlcNAcylation. Human and murine cells deficient of O-GlcNAc transferase, a key enzyme for protein O-GlcNAcylation, show defective antiviral immune responses upon VSV challenge. Mechanistically, O-GlcNAc transferase-mediated O-GlcNAcylation of the signaling adaptor MAVS on serine 366 is required for K63-linked ubiquitination of MAVS and subsequent downstream retinoic-acid inducible gene-like receptor -antiviral signaling activation. Thus, our study identifies a molecular mechanism by which HBP-mediated O-GlcNAcylation regulates MAVS function and highlights the importance of glucose metabolism in antiviral innate immunity.
Notes: 1934-6069
Li, Tianliang
Li, Xinghui
Attri, Kuldeep S
Liu, Changhong
Li, Lupeng
Herring, Laura E
Asara, John M
Lei, Yu L
Singh, Pankaj K
Gao, Chengjiang
Wen, Haitao
R01 CA216853/CA/NCI NIH HHS/United States
U54 CA163120/CA/NCI NIH HHS/United States
P30 CA016086/CA/NCI NIH HHS/United States
P01 CA120964/CA/NCI NIH HHS/United States
P50 CA127297/CA/NCI NIH HHS/United States
R01 GM120496/GM/NIGMS NIH HHS/United States
K99 DE024173/DE/NIDCR NIH HHS/United States
R01 CA163649/CA/NCI NIH HHS/United States
R01 CA210439/CA/NCI NIH HHS/United States
R01 DE026728/DE/NIDCR NIH HHS/United States
R00 DE024173/DE/NIDCR NIH HHS/United States
P30 CA006516/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell Host Microbe. 2018 Dec 12;24(6):791-803.e6. doi: 10.1016/j.chom.2018.11.001.
Author Address: Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE 68198, USA; Holland Regenerative Medicine Program, University of Nebraska Medical Center, Omaha, NE 68198, USA.
Eppley Institute for Research in Cancer and Applied Diseases, University of Nebraska Medical Center, Omaha, NE 68198, USA.
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE 68198, USA; Holland Regenerative Medicine Program, University of Nebraska Medical Center, Omaha, NE 68198, USA; Department of Gastroenterology, Shandong Provincial Qianfoshan Hospital, Jinan 250014, China.
Proteomics Core Facility, Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Division of Signal Transduction, Beth Israel Deaconess Medical Center and Department of Medicine, Harvard Medical School, Boston, MA 02215, USA.
Department of Periodontics and Oral Medicine, University of Michigan School of Dentistry, University of Michigan Comprehensive Cancer Center, University of Michigan, Ann Arbor, MI 48109, USA.
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE 68198, USA; Eppley Institute for Research in Cancer and Applied Diseases, University of Nebraska Medical Center, Omaha, NE 68198, USA.
Department of Immunology and Key Laboratory of Infection and Immunity of Shandong Province, Shandong University School of Basic Medical Sciences, Jinan 250012, China.
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE 68198, USA; Holland Regenerative Medicine Program, University of Nebraska Medical Center, Omaha, NE 68198, USA. Electronic address: haiwen75@gmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2009
Author: Li, X., Gong, W., Wang, H., Li, T., Attri, K. S., Lewis, R. E., Kalil, A. C., Bhinderwala, F., Powers, R., Yin, G., Herring, L. E., Asara, J. M., Lei, Y. L., Yang, X., Rodriguez, D. A., Yang, M., Green, D. R., Singh, P. K. and Wen, H.
Year: 2019
Title: O-GlcNAc Transferase Suppresses Inflammation and Necroptosis by Targeting Receptor-Interacting Serine/Threonine-Protein Kinase 3
Journal: Immunity
Volume: 50
Issue: 3
Pages: 576-590.e6
Epub Date: 2019/02/17
Date: Mar 19
Short Title: O-GlcNAc Transferase Suppresses Inflammation and Necroptosis by Targeting Receptor-Interacting Serine/Threonine-Protein Kinase 3
Alternate Journal: Immunity
ISSN: 1074-7613 (Print)
1074-7613
DOI: 10.1016/j.immuni.2019.01.007
PMCID: PMC6426684
NIHMSID: NIHMS1519255
Accession Number: 30770249
Keywords: Animals
Apoptosis/*physiology
Cell Line
Glucose/metabolism
Humans
Immunity, Innate/physiology
Inflammation/*metabolism
Mice
Mice, Inbred C57BL
N-Acetylglucosaminyltransferases/*metabolism
Necrosis/*metabolism
Receptor-Interacting Protein Serine-Threonine Kinases/*metabolism
Serine/metabolism
Signal Transduction/physiology
Threonine/metabolism
*Hbp
*O-GlcNAc
*Ogt
*Ripk3
*inflammation
*necroptosis
Abstract: Elevated glucose metabolism in immune cells represents a hallmark feature of many inflammatory diseases, such as sepsis. However, the role of individual glucose metabolic pathways during immune cell activation and inflammation remains incompletely understood. Here, we demonstrate a previously unrecognized anti-inflammatory function of the O-linked β-N-acetylglucosamine (O-GlcNAc) signaling associated with the hexosamine biosynthesis pathway (HBP). Despite elevated activities of glycolysis and the pentose phosphate pathway, activation of macrophages with lipopolysaccharide (LPS) resulted in attenuated HBP activity and protein O-GlcNAcylation. Deletion of O-GlcNAc transferase (OGT), a key enzyme for protein O-GlcNAcylation, led to enhanced innate immune activation and exacerbated septic inflammation. Mechanistically, OGT-mediated O-GlcNAcylation of the serine-threonine kinase RIPK3 on threonine 467 (T467) prevented RIPK3-RIPK1 hetero- and RIPK3-RIPK3 homo-interaction and inhibited downstream innate immunity and necroptosis signaling. Thus, our study identifies an immuno-metabolic crosstalk essential for fine-tuning innate immune cell activation and highlights the importance of glucose metabolism in septic inflammation.
Notes: 1097-4180
Li, Xinghui
Gong, Wei
Wang, Hao
Li, Tianliang
Attri, Kuldeep S
Lewis, Robert E
Kalil, Andre C
Bhinderwala, Fatema
Powers, Robert
Yin, Guowei
Herring, Laura E
Asara, John M
Lei, Yu L
Yang, Xiaoyong
Rodriguez, Diego A
Yang, Mao
Green, Douglas R
Singh, Pankaj K
Wen, Haitao
R01 CA216853/CA/NCI NIH HHS/United States
R35 CA231620/CA/NCI NIH HHS/United States
R01 CA210439/CA/NCI NIH HHS/United States
R01 DE026728/DE/NIDCR NIH HHS/United States
R00 DE024173/DE/NIDCR NIH HHS/United States
R01 AI044828/AI/NIAID NIH HHS/United States
R37 AI044828/AI/NIAID NIH HHS/United States
R01 CA169291/CA/NCI NIH HHS/United States
R01 GM120496/GM/NIGMS NIH HHS/United States
R01 CA163649/CA/NCI NIH HHS/United States
R01 DK102648/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Immunity. 2019 Mar 19;50(3):576-590.e6. doi: 10.1016/j.immuni.2019.01.007. Epub 2019 Feb 12.
Author Address: Department of Microbial Infection and Immunity, Infectious Disease Institute, The Ohio State University Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210, USA.
Department of Hepatobiliary Surgery and Liver Transplantation, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong, China.
Department of Critical Care Medicine, Qilu Hospital of Shandong University, Jinan, Shandong, China.
Eppley Institute for Research in Cancer and Allied Diseases, Nebraska Medical Center, Omaha, NE 68198, USA.
Department of Internal Medicine, Division of infectious Diseases, University of Nebraska Medical Center, Omaha, NE 68198, USA.
Department of Chemistry, Nebraska Center for Integrated Biomolecular Communication, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
Department of Psychiatry, Vagelos College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA.
Proteomics Core Facility, Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Division of Signal Transduction, Beth Israel Deaconess Medical Center and Department of Medicine, Harvard Medical School, Boston, MA 02215, USA.
Department of Periodontics and Oral Medicine, University of Michigan School of Dentistry, University of Michigan Rogel Cancer Center, University of Michigan, Ann Arbor, MI 48105, USA.
Program in Integrative Cell Signaling and Neurobiology of Metabolism, Department of Comparative Medicine, Yale University School of Medicine, New Haven, CT 06520, USA.
Department of Immunology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
Department of Microbial Infection and Immunity, Infectious Disease Institute, The Ohio State University Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210, USA. Electronic address: haitao.wen@osumc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1878
Author: Li, X., Lu, F., Trinh, M. N., Schmiege, P., Seemann, J., Wang, J. and Blobel, G.
Year: 2017
Title: 3.3 Å structure of Niemann-Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport
Journal: Proc Natl Acad Sci U S A
Volume: 114
Issue: 34
Pages: 9116-9121
Epub Date: 2017/08/09
Date: Aug 22
Short Title: 3.3 Å structure of Niemann-Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1711716114
PMCID: PMC5576846
Accession Number: 28784760
Keywords: Binding Sites/genetics
Biological Transport/genetics
Carrier Proteins/chemistry/genetics/*metabolism
Cholesterol, LDL/*metabolism
Crystallography, X-Ray
Cysteine/chemistry/genetics/metabolism
Endosomes/*metabolism
Humans
Intracellular Signaling Peptides and Proteins
Membrane Glycoproteins/chemistry/genetics/*metabolism
Models, Molecular
Mutation
Niemann-Pick Disease, Type C/genetics/metabolism
Protein Domains
*Niemann–Pick type C disease
*cholesterol transport
*crystal structure
*cysteine-rich domain
*sterol-sensing domain
Abstract: Niemann-Pick C1 (NPC1) and NPC2 proteins are indispensable for the export of LDL-derived cholesterol from late endosomes. Mutations in these proteins result in Niemann-Pick type C disease, a lysosomal storage disease. Despite recent reports of the NPC1 structure depicting its overall architecture, the function of its C-terminal luminal domain (CTD) remains poorly understood even though 45% of NPC disease-causing mutations are in this domain. Here, we report a crystal structure at 3.3 Å resolution of NPC1* (residues 314-1,278), which-in contrast to previous lower resolution structures-features the entire CTD well resolved. Notably, all eight cysteines of the CTD form four disulfide bonds, one of which (C909-C914) enforces a specific loop that in turn mediates an interaction with a loop of the N-terminal domain (NTD). Importantly, this loop and its interaction with the NTD were not observed in any previous structures due to the lower resolution. Our mutagenesis experiments highlight the physiological relevance of the CTD-NTD interaction, which might function to keep the NTD in the proper orientation for receiving cholesterol from NPC2. Additionally, this structure allows us to more precisely map all of the disease-causing mutations, allowing future molecular insights into the pathogenesis of NPC disease.
Notes: 1091-6490
Li, Xiaochun
Lu, Feiran
Orcid: 0000-0002-1757-2895
Trinh, Michael N
Orcid: 0000-0003-0600-1349
Schmiege, Philip
Seemann, Joachim
Wang, Jiawei
Blobel, Günter
P01 HL020948/HL/NHLBI NIH HHS/United States
R01 GM096070/GM/NIGMS NIH HHS/United States
HHMI/Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2017 Aug 22;114(34):9116-9121. doi: 10.1073/pnas.1711716114. Epub 2017 Aug 7.
Author Address: Laboratory of Cell Biology, The Rockefeller University, New York, NY 10065; xli05@rockefeller.edu blobel@rockefeller.edu.
Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065.
Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, TX 75390.
Laboratory of Cell Biology, The Rockefeller University, New York, NY 10065.
Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390.
State Key Laboratory of Membrane Biology, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1244
Author: Li, X., Molina, H., Huang, H., Zhang, Y. Y., Liu, M., Qian, S. W., Slawson, C., Dias, W. B., Pandey, A., Hart, G. W., Lane, M. D. and Tang, Q. Q.
Year: 2009
Title: O-linked N-acetylglucosamine modification on CCAAT enhancer-binding protein beta: role during adipocyte differentiation
Journal: J Biol Chem
Volume: 284
Issue: 29
Pages: 19248-54
Epub Date: 2009/05/30
Date: Jul 17
Short Title: O-linked N-acetylglucosamine modification on CCAAT enhancer-binding protein beta: role during adipocyte differentiation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.005678
PMCID: PMC2740549
Accession Number: 19478079
Keywords: 3T3-L1 Cells
Acetylglucosamine/*metabolism
Adipocytes/cytology/*metabolism
Amino Acid Sequence
Animals
Blotting, Western
CCAAT-Enhancer-Binding Protein-beta/chemistry/genetics/*metabolism
*Cell Differentiation
Electrophoresis, Polyacrylamide Gel
Electrophoretic Mobility Shift Assay
Glycogen Synthase Kinase 3/metabolism
Glycogen Synthase Kinase 3 beta
Glycosylation
Luciferases/genetics/metabolism
Mass Spectrometry/methods
Mice
Mitogen-Activated Protein Kinases/metabolism
Molecular Sequence Data
Oligonucleotides/genetics/metabolism
Peptide Fragments/chemistry
Phosphorylation
Promoter Regions, Genetic/genetics
Protein Binding
Recombinant Fusion Proteins/genetics/metabolism
Trypsin/metabolism
Abstract: CCAAT enhancer-binding protein (C/EBP)beta is a basic leucine zipper transcription factor family member, and can be phosphorylated, acetylated, and sumoylated. C/EBPbeta undergoes sequential phosphorylation during 3T3-L1 adipocyte differentiation. Phosphorylation on Thr(188) by MAPK or cyclin A/cdk2 primes the phosphorylations on Ser(184)/Thr(179) by GSK3beta, and these phosphorylations are required for the acquisition of DNA binding activity of C/EBPbeta. Here we show that C/EBPbeta is modified by O-GlcNAc, a dynamic single sugar modification found on nucleocytoplasmic proteins. The GlcNAcylation sites are Ser(180) and Ser(181), which are in the regulation domain and are very close to the phosphorylation sites (Thr(188), Ser(184), and Thr(179)) required for the gain of DNA binding activity. Both in vitro and ex vivo experiments demonstrate that GlcNAcylation on Ser(180) and Ser(181) prevents phosphorylation on Thr(188), Ser(184), and Thr(179), as indicated by the decreased relative phosphorylation and DNA binding activity of C/EBPbeta delayed the adipocyte differentiation program. Mutation of both Ser(180) and Ser(181) to Ala significantly increase the transcriptional activity of C/EBPbeta. These data suggest that GlcNAcylation regulates both the phosphorylation and DNA binding activity of C/EBPbeta.
Notes: 1083-351x
Li, Xi
Molina, Henrik
Huang, Haiyan
Zhang, You-You
Liu, Mei
Qian, Shu-Wen
Slawson, Chad
Dias, Wagner B
Pandey, Akhilesh
Hart, Gerald W
Lane, M Daniel
Tang, Qi-Qun
R01 DK061671/DK/NIDDK NIH HHS/United States
R37 HD013563/HD/NICHD NIH HHS/United States
DK61671/DK/NIDDK NIH HHS/United States
HD13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2009 Jul 17;284(29):19248-54. doi: 10.1074/jbc.M109.005678. Epub 2009 May 28.
Author Address: Key Laboratory of Molecular Medicine, Ministry of Education, Fudan University Shanghai Medical College, Shanghai 200032, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1806
Author: Li, X., Saha, P., Li, J., Blobel, G. and Pfeffer, S. R.
Year: 2016
Title: Clues to the mechanism of cholesterol transfer from the structure of NPC1 middle lumenal domain bound to NPC2
Journal: Proc Natl Acad Sci U S A
Volume: 113
Issue: 36
Pages: 10079-84
Epub Date: 2016/08/24
Date: Sep 6
Short Title: Clues to the mechanism of cholesterol transfer from the structure of NPC1 middle lumenal domain bound to NPC2
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1611956113
PMCID: PMC5018801
Accession Number: 27551080
Keywords: Amino Acid Motifs
Binding Sites
Biological Transport
Carrier Proteins/*chemistry/genetics/metabolism
Cholesterol Esters/*chemistry/metabolism
Cloning, Molecular
Crystallography, X-Ray
Escherichia coli/genetics/metabolism
Gene Expression
Genetic Vectors/chemistry/metabolism
Glycoproteins/*chemistry/genetics/metabolism
Humans
Hydrophobic and Hydrophilic Interactions
Intracellular Signaling Peptides and Proteins
Kinetics
Lysosomes/*metabolism
Membrane Glycoproteins/*chemistry/genetics/metabolism
Molecular Docking Simulation
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Recombinant Proteins/chemistry/genetics/metabolism
Vesicular Transport Proteins
*Ebola virus glycoprotein
*Niemann–Pick type C disease
*cholesterol trafficking
*crystal structure
Abstract: Export of LDL-derived cholesterol from lysosomes requires the cooperation of the integral membrane protein Niemann-Pick C1 (NPC1) and a soluble protein, Niemann-Pick C2 (NPC2). Mutations in the genes encoding these proteins lead to Niemann-Pick disease type C (NPC). NPC2 binds to NPC1's second (middle), lumenally oriented domain (MLD) and transfers cholesterol to NPC1's N-terminal domain (NTD). Here, we report the 2.4-Å resolution crystal structure of a complex of human NPC1-MLD and NPC2 bearing bound cholesterol-3-O-sulfate. NPC1-MLD uses two protruding loops to bind NPC2, analogous to its interaction with the primed Ebola virus glycoprotein. Docking of the NPC1-NPC2 complex onto the full-length NPC1 structure reveals a direct cholesterol transfer tunnel between NPC2 and NTD cholesterol binding pockets, supporting the "hydrophobic hand-off" cholesterol transfer model.
Notes: 1091-6490
Li, Xiaochun
Saha, Piyali
Li, Jian
Blobel, Günter
Pfeffer, Suzanne R
R01 DK037332/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2016 Sep 6;113(36):10079-84. doi: 10.1073/pnas.1611956113. Epub 2016 Aug 22.
Author Address: Laboratory of Cell Biology, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065; xli05@rockefeller.edu blobel@rockefeller.edu pfeffer@stanford.edu.
Department of Biochemistry, Stanford University School of Medicine, Stanford, CA 94305.
Department of Biochemistry, Stanford University School of Medicine, Stanford, CA 94305 xli05@rockefeller.edu blobel@rockefeller.edu pfeffer@stanford.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2019
Author: Li, X., Zhu, Q., Shi, X., Cheng, Y., Li, X., Xu, H., Duan, X., Hsieh-Wilson, L. C., Chu, J., Pelletier, J., Ni, M., Zheng, Z., Li, S. and Yi, W.
Year: 2019
Title: O-GlcNAcylation of core components of the translation initiation machinery regulates protein synthesis
Journal: Proc Natl Acad Sci U S A
Volume: 116
Issue: 16
Pages: 7857-7866
Epub Date: 2019/04/04
Date: Apr 16
Short Title: O-GlcNAcylation of core components of the translation initiation machinery regulates protein synthesis
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1813026116
PMCID: PMC6475381
Accession Number: 30940748
Keywords: Cell Line, Tumor
Eukaryotic Initiation Factor-4G/chemistry/metabolism
*Glycosylation
Humans
Models, Molecular
Neoplasms/chemistry/metabolism
*Peptide Chain Initiation, Translational
Poly(A)-Binding Proteins/chemistry/metabolism
RNA, Messenger/chemistry/metabolism
*protein synthesis
*translation initiation
Abstract: Protein synthesis is essential for cell growth, proliferation, and survival. Protein synthesis is a tightly regulated process that involves multiple mechanisms. Deregulation of protein synthesis is considered as a key factor in the development and progression of a number of diseases, such as cancer. Here we show that the dynamic modification of proteins by O-linked β-N-acetyl-glucosamine (O-GlcNAcylation) regulates translation initiation by modifying core initiation factors eIF4A and eIF4G, respectively. Mechanistically, site-specific O-GlcNAcylation of eIF4A on Ser322/323 disrupts the formation of the translation initiation complex by perturbing its interaction with eIF4G. In addition, O-GlcNAcylation inhibits the duplex unwinding activity of eIF4A, leading to impaired protein synthesis, and decreased cell proliferation. In contrast, site-specific O-GlcNAcylation of eIF4G on Ser61 promotes its interaction with poly(A)-binding protein (PABP) and poly(A) mRNA. Depletion of eIF4G O-GlcNAcylation results in inhibition of protein synthesis, cell proliferation, and soft agar colony formation. The differential glycosylation of eIF4A and eIF4G appears to be regulated in the initiation complex to fine-tune protein synthesis. Our study thus expands the current understanding of protein synthesis, and adds another dimension of complexity to translational control of cellular proteins.
Notes: 1091-6490
Li, Xuexia
Zhu, Qiang
Shi, Xiaoliu
Cheng, Yaxian
Li, Xueliu
Xu, Huan
Duan, Xiaotao
Hsieh-Wilson, Linda C
Chu, Jennifer
Pelletier, Jerry
Ni, Maowei
Zheng, Zhiguo
Li, Sihui
Yi, Wen
Orcid: 0000-0002-4257-3355
RF1 AG060540/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2019 Apr 16;116(16):7857-7866. doi: 10.1073/pnas.1813026116. Epub 2019 Apr 2.
Author Address: Ministry of Education Laboratory of Biosystems Homeostasis & Protection, College of Life Sciences, Zhejiang University, 310058 Hangzhou, China.
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, 100850 Beijing, China.
Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125.
Department of Biochemistry, McGill University, Montreal, QC H3G 1Y6, Canada.
Zhejiang Cancer Hospital, Zhejiang Cancer Research Institute, 310022 Hangzhou, China.
Ministry of Education Laboratory of Biosystems Homeostasis & Protection, College of Life Sciences, Zhejiang University, 310058 Hangzhou, China; lisihui@zju.edu.cn wyi@zju.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1220
Author: Li, Y., Cao, C., Jia, W., Yu, L., Mo, M., Wang, Q., Huang, Y., Lim, J. M., Ishihara, M., Wells, L., Azadi, P., Robinson, H., He, Y. W., Zhang, L. and Mariuzza, R. A.
Year: 2009
Title: Structure of the F-spondin domain of mindin, an integrin ligand and pattern recognition molecule
Journal: Embo j
Volume: 28
Issue: 3
Pages: 286-97
Epub Date: 2009/01/21
Date: Feb 4
Short Title: Structure of the F-spondin domain of mindin, an integrin ligand and pattern recognition molecule
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2008.288
PMCID: PMC2637340
Accession Number: 19153605
Keywords: Amino Acid Sequence
Animals
Binding Sites
Calcium/metabolism
Cell Adhesion
Cell Line
Crystallography, X-Ray
Extracellular Matrix Proteins/*chemistry/*metabolism
Humans
Integrins/*metabolism
Lipopolysaccharides/metabolism
Mannose/metabolism
Mass Spectrometry
Mice
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Pattern Recognition/*chemistry/*metabolism
Repetitive Sequences, Amino Acid
Static Electricity
Abstract: Mindin (spondin-2) is an extracellular matrix protein of unknown structure that is required for efficient T-cell priming by dendritic cells. Additionally, mindin functions as a pattern recognition molecule for initiating innate immune responses. These dual functions are mediated by interactions with integrins and microbial pathogens, respectively. Mindin comprises an N-terminal F-spondin (FS) domain and C-terminal thrombospondin type 1 repeat (TSR). We determined the structure of the FS domain at 1.8-A resolution. The structure revealed an eight-stranded antiparallel beta-sandwich motif resembling that of membrane-targeting C2 domains, including a bound calcium ion. We demonstrated that the FS domain mediates integrin binding and identified the binding site by mutagenesis. The mindin FS domain therefore represents a new integrin ligand. We further showed that mindin recognizes lipopolysaccharide (LPS) through its TSR domain, and obtained evidence that C-mannosylation of the TSR influences LPS binding. Through these dual interactions, the FS and TSR domains of mindin promote activation of both adaptive and innate immune responses.
Notes: 1460-2075
Li, Yili
Cao, Chunzhang
Jia, Wei
Yu, Lily
Mo, Min
Wang, Qian
Huang, Yuping
Lim, Jae-Min
Ishihara, Mayumi
Wells, Lance
Azadi, Parastoo
Robinson, Howard
He, You-Wen
Zhang, Li
Mariuzza, Roy A
P01 HL054710/HL/NHLBI NIH HHS/United States
U01 AI061364/AI/NIAID NIH HHS/United States
P01 HL54710/HL/NHLBI NIH HHS/United States
R01 AI065612/AI/NIAID NIH HHS/United States
AI061364/AI/NIAID NIH HHS/United States
P30 EB009998/EB/NIBIB NIH HHS/United States
AI054658/AI/NIAID NIH HHS/United States
R01 AI054658/AI/NIAID NIH HHS/United States
AI065612/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
EMBO J. 2009 Feb 4;28(3):286-97. doi: 10.1038/emboj.2008.288. Epub 2009 Jan 15.
Author Address: WM Keck Laboratory for Structural Biology, Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute, Rockville, MD 20850, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1873
Author: Li, Y., Wang, L., Liu, J., Zhang, P., An, M., Han, C., Li, Y., Guan, X. and Zhang, K.
Year: 2017
Title: O-GlcNAcylation modulates Bmi-1 protein stability and potential oncogenic function in prostate cancer
Journal: Oncogene
Volume: 36
Issue: 45
Pages: 6293-6305
Epub Date: 2017/07/18
Date: Nov 9
Short Title: O-GlcNAcylation modulates Bmi-1 protein stability and potential oncogenic function in prostate cancer
Alternate Journal: Oncogene
ISSN: 0950-9232
DOI: 10.1038/onc.2017.223
Accession Number: 28714959
Keywords: Acylation
Case-Control Studies
Cell Line, Tumor
Humans
Immunoprecipitation
Male
Mass Spectrometry
Mitogen-Activated Protein Kinase 7/genetics/*metabolism
N-Acetylglucosaminyltransferases/metabolism
Oncogenes
Polycomb Repressive Complex 1/genetics/*metabolism
Prostatic Neoplasms/enzymology/genetics/*metabolism
Protein Stability
Transfection
Abstract: The Polycomb group transcriptional repressor Bmi-1 often overexpressed and participated in stem cells self-renewal and tumorigenesis initiating of prostate cancer. In this progression, Bmi-1 protein was regulated by transcription and post-translational modifications (PTMs). Nobly, the underlying PTMs regulation of Bmi-1 is poorly known. Here we use co-immunoprecipitation show that in C4-2 cell line, Bmi-1 directly interacted with OGT which is the only known enzyme catalyzed the O-GlcNAcylation in human. Furthermore, we identified that Ser255 is the site for Bmi-1 O-GlcNAcylation, and O-GlcNAcylation promoted Bmi-1 protein stability and its oncogenic activity. Finally, microarray analysis has characterized potential oncogenes associated pathway subject to repression via the OGT-Bmi-1 axis. Taken together, these results indicate that OGT-mediated O-GlcNAcylation at Ser255 stabilizes Bmi-1 and hence inhibits the TP53, PTEN and CDKN1A/CDKN2A pathway. The study not only uncovers a novel functional PTMs of Bmi-1 but also reveals a unique oncogenic role of O-GlcNAcylation in prostate cancer.
Notes: 1476-5594
Li, Y
Wang, L
Liu, J
Zhang, P
An, M
Han, C
Guan, X
Zhang, K
Journal Article
England
Oncogene. 2017 Nov 9;36(45):6293-6305. doi: 10.1038/onc.2017.223. Epub 2017 Jul 17.
Author Address: Sino-UK Regenerative Medicine Center, the First Affiliated Hospital of Dalian Medical University, Dalian, People's Republic of China.
Department of Pathology, the First Affiliated Hospital of Dalian Medical University, Dalian, People's Republic of China.
Department of Bioengineering, University of California, San Diego, La Jolla, CA, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1568
Author: Li, Y., Wang, Q., Chen, S., Brown, P. H. and Mariuzza, R. A.
Year: 2013
Title: Structure of NKp65 bound to its keratinocyte ligand reveals basis for genetically linked recognition in natural killer gene complex
Journal: Proc Natl Acad Sci U S A
Volume: 110
Issue: 28
Pages: 11505-10
Epub Date: 2013/06/28
Date: Jul 9
Short Title: Structure of NKp65 bound to its keratinocyte ligand reveals basis for genetically linked recognition in natural killer gene complex
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1303300110
PMCID: PMC3710857
Accession Number: 23803857
Keywords: Amino Acid Sequence
Humans
Keratinocytes/*metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Protein Conformation
Receptors, NK Cell Lectin-Like/chemistry/genetics/*metabolism
Sequence Homology, Amino Acid
Abstract: The natural killer (NK) gene complex (NKC) encodes numerous C-type lectin-like receptors that govern the activity of NK cells. Although some of these receptors (Ly49s, NKG2D, CD94/NKG2A) recognize MHC or MHC-like molecules, others (Nkrp1, NKRP1A, NKp80, NKp65) instead bind C-type lectin-like ligands to which they are genetically linked in the NKC. To understand the basis for this recognition, we determined the structure of human NKp65, an activating receptor implicated in the immunosurveillance of skin, bound to its NKC-encoded ligand keratinocyte-associated C-type lectin (KACL). Whereas KACL forms a homodimer resembling other C-type lectin-like dimers, NKp65 is monomeric. The binding mode in the NKp65-KACL complex, in which a monomeric receptor engages a dimeric ligand, is completely distinct from those used by Ly49s, NKG2D, or CD94/NKG2A. The structure explains the exceptionally high affinity of the NKp65-KACL interaction compared with other cell-cell interaction pairs (KD = 6.7 × 10(-10) M), which may compensate for the monomeric nature of NKp65 to achieve cell activation. This previously unreported structure of an NKC-encoded receptor-ligand complex, coupled with mutational analysis of the interface, establishes a docking template that is directly applicable to other genetically linked pairs in the NKC, including Nkrp1-Clr, NKRP1A-LLT1, and NKp80-AICL.
Notes: 1091-6490
Li, Yili
Wang, Qian
Chen, Sharon
Brown, Patrick H
Mariuzza, Roy A
R01 AI047990/AI/NIAID NIH HHS/United States
AI047990/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2013 Jul 9;110(28):11505-10. doi: 10.1073/pnas.1303300110. Epub 2013 Jun 26.
Author Address: W. M. Keck Laboratory for Structural Biology, University of Maryland Institute for Bioscience and Biotechnology Research, Rockville, MD 20850, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1385
Author: Li, Y., Wang, Q. and Mariuzza, R. A.
Year: 2011
Title: Structure of the human activating natural cytotoxicity receptor NKp30 bound to its tumor cell ligand B7-H6
Journal: J Exp Med
Volume: 208
Issue: 4
Pages: 703-14
Epub Date: 2011/03/23
Date: Apr 11
Short Title: Structure of the human activating natural cytotoxicity receptor NKp30 bound to its tumor cell ligand B7-H6
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.20102548
PMCID: PMC3135353
Accession Number: 21422170
Keywords: Amino Acid Sequence
Antigens, CD/chemistry
Apoptosis Regulatory Proteins/chemistry
B7 Antigens
B7-1 Antigen/*chemistry/metabolism
CTLA-4 Antigen
Humans
Models, Molecular
Molecular Sequence Data
Natural Cytotoxicity Triggering Receptor 3/*chemistry/metabolism
Programmed Cell Death 1 Receptor
Protein Binding
Protein Structure, Quaternary
Protein Structure, Secondary
Sequence Alignment
Surface Plasmon Resonance
Abstract: Natural killer (NK) cells are lymphocytes of the innate immune system that participate in the elimination of tumor cells. In humans, the activating natural cytotoxicity receptors (NCRs) NKp30, NKp44, and NKp46 play a major role in NK cell-mediated tumor cell lysis. NKp30 recognizes B7-H6, a member of the B7 family which is expressed on tumor, but not healthy, cells. To understand the basis for tumor surveillance by NCRs, we determined the structure of NKp30, a member of the CD28 family which includes CTLA-4 and PD-1, in complex with B7-H6. The overall organization of the NKp30-B7-H6-activating complex differs considerably from those of the CTLA-4-B7 and PD-1-PD-L T cell inhibitory complexes. Whereas CTLA-4 and PD-1 use only the front β-sheet of their Ig-like domain to bind ligands, NKp30 uses both front and back β-sheets, resulting in engagement of B7-H6 via the side, as well as face, of the β-sandwich. Moreover, B7-H6 contacts NKp30 through the complementarity-determining region (CDR)-like loops of its V-like domain in an antibody-like interaction that is not observed for B7 or PD-L. This first structure of an NCR bound to ligand provides a template for designing molecules to stimulate NKp30-mediated cytolytic activity for tumor immunotherapy.
Notes: 1540-9538
Li, Yili
Wang, Qian
Mariuzza, Roy A
P30 EB009998/EB/NIBIB NIH HHS/United States
R01 AI047990/AI/NIAID NIH HHS/United States
AI047990/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Exp Med. 2011 Apr 11;208(4):703-14. doi: 10.1084/jem.20102548. Epub 2011 Mar 21.
Author Address: WM Keck Laboratory for Structural Biology, University of Maryland Institute for Bioscience and Biotechnology Research, Rockville, MD 20850, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1877
Author: Li, Z., Fischer, M., Satkunarajah, M., Zhou, D., Withers, S. G. and Rini, J. M.
Year: 2017
Title: Structural basis of Notch O-glucosylation and O-xylosylation by mammalian protein-O-glucosyltransferase 1 (POGLUT1)
Journal: Nat Commun
Volume: 8
Issue: 1
Pages: 185
Epub Date: 2017/08/05
Date: Aug 4
Short Title: Structural basis of Notch O-glucosylation and O-xylosylation by mammalian protein-O-glucosyltransferase 1 (POGLUT1)
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-017-00255-7
PMCID: PMC5543122
Accession Number: 28775322
Keywords: Catalytic Domain
Gene Expression Regulation, Enzymologic/*physiology
Glucosyltransferases/genetics/*metabolism
HEK293 Cells
Humans
Models, Molecular
Protein Conformation
Receptors, Notch/genetics/*metabolism
Abstract: Protein O-glucosyltransferase 1/Rumi-mediated glucosylation of Notch epidermal growth factor-like (EGF-like) domains plays an important role in Notch signaling. Protein O-glucosyltransferase 1 shows specificity for folded EGF-like domains, it can only glycosylate serine residues in the C(1)XSXPC(2) motif, and it possesses an uncommon dual donor substrate specificity. Using several EGF-like domains and donor substrate analogs, we have determined the structures of human Protein O-glucosyltransferase 1 substrate/product complexes that provide mechanistic insight into the basis for these properties. Notably, we show that Protein O-glucosyltransferase 1's requirement for folded EGF-like domains also leads to its serine specificity and that two distinct local conformational states are likely responsible for its ability to transfer both glucose and xylose. We also show that Protein O-glucosyltransferase 1 possesses the potential to xylosylate a much broader range of EGF-like domain substrates than was previously thought. Finally, we show that Protein O-glucosyltransferase 1 has co-evolved with EGF-like domains of the type found in Notch.POGLUT1 is a protein-O-glucosyltransferase that transfers glucose and xylose to the EGF-like domains of Notch and other signaling receptors. Here the authors report the structure of human POGLUT1 in complexes with 3 different EGF-like domains and donor substrates and shed light on the enzyme's substrate specificity and catalytic mechanism.
Notes: 2041-1723
Li, Zhijie
Orcid: 0000-0001-9283-6072
Fischer, Michael
Satkunarajah, Malathy
Zhou, Dongxia
Withers, Stephen G
Rini, James M
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2017 Aug 4;8(1):185. doi: 10.1038/s41467-017-00255-7.
Author Address: Department of Biochemistry, University of Toronto, Toronto, ON, Canada, M5S 1A8.
Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada, M5S 1A8.
Department of Chemistry, University of British Columbia, Vancouver, BC, Canada, V6T 1Z1.
Department of Biochemistry, University of Toronto, Toronto, ON, Canada, M5S 1A8. james.rini@utoronto.ca.
Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada, M5S 1A8. james.rini@utoronto.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1233
Author: Lim, K. and Chang, H. I.
Year: 2009
Title: O-GlcNAcylation of Sp1 interrupts Sp1 interaction with NF-Y
Journal: Biochem Biophys Res Commun
Volume: 382
Issue: 3
Pages: 593-7
Epub Date: 2009/03/24
Date: May 8
Short Title: O-GlcNAcylation of Sp1 interrupts Sp1 interaction with NF-Y
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2009.03.075
Accession Number: 19302979
Keywords: Acetylglucosamine/*metabolism
Acylation
CCAAT-Binding Factor/*metabolism
Cell Line
Humans
Immunoprecipitation
Promoter Regions, Genetic
Protein Structure, Tertiary
Sp1 Transcription Factor/*metabolism
Transcription, Genetic
*Transcriptional Activation
Abstract: O-linked N-acetylglucosamine (O-GlcNAc), a monosaccharide N-acetylglucosamine addition on nucleocytoplasmic proteins, is abundant in transcription regulators and has been implicated in gene regulation. Sp1 transcription factor is multiply modified by O-GlcNAc within its serine/threonine-rich region and glutamine-rich transactivation domain. In the present study, we show that O-GlcNAc of Sp1 serine/threonine-rich region interrupts a physical interaction between Sp1 and NF-YA, thus inhibiting Sp1-NF-Y cooperative activation of gene transcription. Our results strengthen the notion that O-GlcNAc regulates gene transcription by modulating the protein-protein interaction network among transcription regulatory proteins.
Notes: 1090-2104
Lim, Kihong
Chang, Hyo-Ihl
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2009 May 8;382(3):593-7. doi: 10.1016/j.bbrc.2009.03.075. Epub 2009 Mar 18.
Author Address: School of Life Sciences and Biotechnology, Korea University, 5-1 Anam-dong, Seongbuk-gu, Seoul 136-701, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1331
Author: Lim, K. and Chang, H. I.
Year: 2010
Title: Elevated O-linked N-acetylglucosamine correlated with reduced Sp1 cooperative DNA binding with its collaborating factors in vivo
Journal: Biosci Biotechnol Biochem
Volume: 74
Issue: 8
Pages: 1668-72
Epub Date: 2010/08/12
Short Title: Elevated O-linked N-acetylglucosamine correlated with reduced Sp1 cooperative DNA binding with its collaborating factors in vivo
Alternate Journal: Bioscience, biotechnology, and biochemistry
ISSN: 0916-8451
DOI: 10.1271/bbb.100289
Accession Number: 20699577
Keywords: Acetylglucosamine/*metabolism
Animals
Base Sequence
Cell Line, Tumor
DNA/*metabolism
Humans
Molecular Sequence Data
Oxygen/*metabolism
Protein Binding/drug effects
RNA, Small Nuclear/genetics
Rats
Sp1 Transcription Factor/*metabolism
Streptozocin/pharmacology
Transcription, Genetic
Abstract: O-Linked N-acetylglucosamine (O-GlcNAc), a single GlcNAc modification of proteins, is abundant in nucleocytoplasmic proteins of eukaryotes. Most nuclear transcriptional regulator proteins carry O-GlcNAc, implicating O-GlcNAc in gene regulation. This study suggested the possibility that O-GlcNAc regulates cooperative binding of Sp1 and its collaborating transcription factors, Oct1 and Elf-1, onto DNA templates in vivo. Chromatin immunoprecipitation assays on cells in which O-GlcNAc was modulated pharmacologically revealed that Sp1-Oct1- and Sp1-Elf-1-paired occupancies of previously known target promoter regions were suppressed by elevated O-GlcNAc modification. Since these pairs of transcription factors bind the target promoters cooperatively and DNA binding of Sp1 alone is not affected by O-GlcNAc, our results imply that O-GlcNAc weakens the DNA binding of Sp1 and its cooperative binding partners by inhibiting stable interaction on DNA templates.
Notes: 1347-6947
Lim, Kihong
Chang, Hyo-Ihl
Journal Article
Research Support, Non-U.S. Gov't
England
Biosci Biotechnol Biochem. 2010;74(8):1668-72. doi: 10.1271/bbb.100289. Epub 2010 Aug 7.
Author Address: College of Life Sciences and Biotechnology, Korea University, Anam-dong, Seongbuk-gu, Seoul, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1743
Author: Lim, K., Yoon, B. H. and Ha, C. H.
Year: 2015
Title: O-Linked N-acetylglucosaminylation of Sp1 interferes with Sp1 activation of glycolytic genes
Journal: Biochem Biophys Res Commun
Volume: 468
Issue: 1-2
Pages: 349-53
Epub Date: 2015/10/27
Date: Dec 4-11
Short Title: O-Linked N-acetylglucosaminylation of Sp1 interferes with Sp1 activation of glycolytic genes
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2015.10.096
Accession Number: 26499076
Keywords: Base Sequence
Glyceraldehyde-3-Phosphate Dehydrogenases/genetics
*Glycolysis
HEK293 Cells
Humans
Molecular Sequence Data
Mutation
Phosphofructokinases/genetics
Promoter Regions, Genetic
Sp1 Transcription Factor/genetics/*metabolism
*Transcriptional Activation
Uridine Diphosphate N-Acetylglucosamine/genetics/*metabolism
Glycolysis
O-GlcNAc
Sp1
Transcription
Abstract: Glycolysis, the primary pathway metabolizing glucose for energy production, is connected to the hexosamine biosynthetic pathway (HBP) which produces UDP-N-acetylglucosamine (UDP-GlcNAc), a GlcNAc donor for O-linked GlcNAc modification (O-GlcNAc), as well as for traditional elongated glycosylation. Thus, glycolysis and O-GlcNAc are intimately associated. The present study reports the transcriptional activation of glycolytic genes by the transcription factor Sp1 and the O-GlcNAc-mediated suppression of Sp1-dependent activation of glycolytic genes. O-GlcNAc-deficient mutant Sp1 stimulated the transcription of nine glycolytic genes and cellular production of pyruvate, the final product of glycolysis, to a greater extent than wild-type Sp1. Consistently, this mutant Sp1 increased the protein levels of the two key glycolytic enzymes, phosphofructokinase (PFK) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), to a greater extent than wild-type Sp1. Finally, the mutant Sp1 occupied GC-rich elements on PFK and GAPDH promoters more efficiently than wild-type Sp1. These results suggest that O-GlcNAcylation of Sp1 suppresses Sp1-mediated activation of glycolytic gene transcription.
Notes: 1090-2104
Lim, Kihong
Yoon, Bo Hyun
Ha, Chang Hoon
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2015 Dec 4-11;468(1-2):349-53. doi: 10.1016/j.bbrc.2015.10.096. Epub 2015 Oct 22.
Author Address: David H. Smith Center for Vaccine Biology and Immunology, University of Rochester Medical Center, 601 Elmwood Avenue, Box 609, Rochester, NY 14642, United States.
Asan Institute for Life Sciences, Asan Medical Center, College of Medicine, University of Ulsan, 86 Asanbyeoungwon-gil, Songpa-gu, Seoul 138-736, Republic of Korea.
Asan Institute for Life Sciences, Asan Medical Center, College of Medicine, University of Ulsan, 86 Asanbyeoungwon-gil, Songpa-gu, Seoul 138-736, Republic of Korea. Electronic address: chhoonha@amc.seoul.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1728
Author: Lim, S., Haque, M. M., Nam, G., Ryoo, N., Rhim, H. and Kim, Y. K.
Year: 2015
Title: Monitoring of Intracellular Tau Aggregation Regulated by OGA/OGT Inhibitors
Journal: Int J Mol Sci
Volume: 16
Issue: 9
Pages: 20212-24
Epub Date: 2015/09/08
Date: Aug 26
Short Title: Monitoring of Intracellular Tau Aggregation Regulated by OGA/OGT Inhibitors
Alternate Journal: International journal of molecular sciences
ISSN: 1422-0067
DOI: 10.3390/ijms160920212
PMCID: PMC4613198
Accession Number: 26343633
Keywords: Cell Line
Enzyme Inhibitors/*pharmacology
Glycosylation
Humans
N-Acetylglucosaminyltransferases/*antagonists & inhibitors
Phosphorylation
Protein Aggregation, Pathological/*metabolism
Pyrans/pharmacology
Tauopathies/metabolism
Thiazoles/pharmacology
tau Proteins/*metabolism
O-GlcNAc transferase
O-GlcNAcase
O-GlcNAcylation
tau aggregation
tau phosphorylation
tau protein
Abstract: Abnormal phosphorylation of tau has been considered as a key pathogenic mechanism inducing tau aggregation in multiple neurodegenerative disorders, collectively called tauopathies. Recent evidence showed that tau phosphorylation sites are protected with O-linked β-N-acetylglucosamine (O-GlcNAc) in normal brain. In pathological condition, tau is de-glycosylated and becomes a substrate for kinases. Despite the importance of O-GlcNAcylation in tau pathology, O-GlcNAc transferase (OGT), and an enzyme catalyzing O-GlcNAc to tau, has not been carefully investigated in the context of tau aggregation. Here, we investigated intracellular tau aggregation regulated by BZX2, an inhibitor of OGT. Upon the inhibition of OGT, tau phosphorylation increased 2.0-fold at Ser199 and 1.5-fold at Ser396, resulting in increased tau aggregation. Moreover, the BZX2 induced tau aggregation was efficiently reduced by the treatment of Thiamet G, an inhibitor of O-GlcNAcase (OGA). Our results demonstrated the protective role of OGT in tau aggregation and also suggest the counter-regulatory mechanism of OGA and OGT in tau pathology.
Notes: 1422-0067
Lim, Sungsu
Haque, Md Mamunul
Nam, Ghilsoo
Ryoo, Nayeon
Rhim, Hyewhon
Kim, Yun Kyung
Journal Article
Research Support, Non-U.S. Gov't
Int J Mol Sci. 2015 Aug 26;16(9):20212-24. doi: 10.3390/ijms160920212.
Author Address: Center for Neuro-Medicine, Brain Science Institute, Korea Institute of Science and Technology (KIST), Seoul 136-791, South Korea. sungsulim85@gmail.com.
Center for Neuro-Medicine, Brain Science Institute, Korea Institute of Science and Technology (KIST), Seoul 136-791, South Korea. mamunbge@gmail.com.
Biological Chemistry, University of Science and Technology (UST), Daejeon 305-333, South Korea. mamunbge@gmail.com.
Center for Neuro-Medicine, Brain Science Institute, Korea Institute of Science and Technology (KIST), Seoul 136-791, South Korea. gsnam@kist.re.kr.
Center for Neuroscience, Brain Science Institute, Korea Institute of Science and Technology (KIST), Seoul 136-791, South Korea. ryoo8099@gmail.com.
Center for Neuroscience, Brain Science Institute, Korea Institute of Science and Technology (KIST), Seoul 136-791, South Korea. hrhim@kist.re.kr.
Department of Neuroscience, University of Science and Technology (UST), Daejeon 305-333, South Korea. hrhim@kist.re.kr.
Center for Neuro-Medicine, Brain Science Institute, Korea Institute of Science and Technology (KIST), Seoul 136-791, South Korea. yunkyungkim@kist.re.kr.
Biological Chemistry, University of Science and Technology (UST), Daejeon 305-333, South Korea. yunkyungkim@kist.re.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1749
Author: Lim, S. M., Yeung, K., Trésaugues, L., Ling, T. H. and Nordlund, P.
Year: 2016
Title: The structure and catalytic mechanism of human sphingomyelin phosphodiesterase like 3a--an acid sphingomyelinase homologue with a novel nucleotide hydrolase activity
Journal: Febs j
Volume: 283
Issue: 6
Pages: 1107-23
Epub Date: 2016/01/20
Date: Mar
Short Title: The structure and catalytic mechanism of human sphingomyelin phosphodiesterase like 3a--an acid sphingomyelinase homologue with a novel nucleotide hydrolase activity
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/febs.13655
Accession Number: 26783088
Keywords: Amino Acid Sequence
Catalytic Domain
Conserved Sequence
Crystallography, X-Ray
Cytidine Monophosphate/metabolism
Disulfides/chemistry
Glycosylation
Humans
Models, Molecular
Molecular Sequence Data
Mutant Proteins/chemistry/genetics/metabolism
Niemann-Pick Diseases/enzymology/genetics
Phylogeny
Point Mutation
Protein Conformation
Sequence Homology, Amino Acid
Sphingomyelin Phosphodiesterase/*chemistry/genetics/*metabolism
Static Electricity
Substrate Specificity
Zinc/metabolism
Niemann-Pick disease
acid sphingomyelinase like
binuclear metallophosphodiesterase
calcineurin-like phosphodiesterase
nucleotide hydrolase
Abstract: Human sphingomyelinase phosphodiesterase like 3a (SMPDL3a) is a secreted enzyme that shares a conserved catalytic domain with human acid sphingomyelinase (aSMase), the enzyme carrying mutations causative of Niemann-Pick disease. We have solved the structure of SMPDL3a revealing a calcineurin-like fold. A dimetal site, glycosylation pattern and a disulfide bond network are likely to be conserved also in human aSMase. We show that the binuclear site of SMPDL3a is occupied by two Zn(2+) ions and that excess Zn(2+) leads to inhibition of enzyme activity through binding to additional sites. As an extension of recent biochemical work we uncovered that SMPDL3a catalyses the hydrolysis of several modified nucleotides that include cytidine 5'-diphosphocholine, cytidine diphosphate ethanolamine and ADP-ribose, but not the aSMase substrate, sphingomyelin. We subsequently determined the structure of SMPDL3a in complex with the product 5'-cytidine monophosphate (CMP), a structure that is consistent with several distinct coordination modes of the substrate/product in the active site during the reaction cycle. Based on the structure of CMP complexes, we propose a phosphoryl transfer mechanism for SMPDL3a. Finally, a homology model of human aSMase was constructed to allow for the mapping of selected Niemann-Pick disease mutations on a three-dimensional framework to guide further characterization of their effects on aSMase function. DATABASE: Structural data are available in the PDB database under the accession numbers 5EBB and 5EBE.
Notes: 1742-4658
Lim, Sing Mei
Yeung, Kit
Trésaugues, Lionel
Ling, Teo Hsiang
Nordlund, Pär
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2016 Mar;283(6):1107-23. doi: 10.1111/febs.13655. Epub 2016 Feb 5.
Author Address: Division of Biomedical Structural Biology, School of Biological Sciences, Nanyang Technological University, Singapore.
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
Institute of Molecular and Cell Biology, A*STAR, Singapore city, Singapore.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1294
Author: Lin, L., Gårdsvoll, H., Huai, Q., Huang, M. and Ploug, M.
Year: 2010
Title: Structure-based engineering of species selectivity in the interaction between urokinase and its receptor: implication for preclinical cancer therapy
Journal: J Biol Chem
Volume: 285
Issue: 14
Pages: 10982-92
Epub Date: 2010/02/06
Date: Apr 2
Short Title: Structure-based engineering of species selectivity in the interaction between urokinase and its receptor: implication for preclinical cancer therapy
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.093492
PMCID: PMC2856303
Accession Number: 20133942
Keywords: Activating Transcription Factor 1/chemistry/genetics/metabolism
Amino Acid Sequence
Animals
Crystallization
Crystallography, X-Ray
*Drug Design
Humans
Mice
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Neoplasms/drug therapy
Protein Conformation
Receptors, Urokinase Plasminogen Activator/*chemistry/genetics/*metabolism
Sequence Homology, Amino Acid
Species Specificity
Surface Plasmon Resonance
Urokinase-Type Plasminogen Activator/*chemistry/genetics/*metabolism
Abstract: The high affinity interaction between the urokinase-type plasminogen activator (uPA) and its glycolipid-anchored receptor (uPAR) is decisive for cell surface-associated plasminogen activation. Because plasmin activity controls fibrinolysis in a variety of pathological conditions, including cancer and wound healing, several intervention studies have focused on targeting the uPA.uPAR interaction in vivo. Evaluations of such studies in xenotransplanted tumor models are, however, complicated by the pronounced species selectivity in this interaction. We now report the molecular basis underlying this difference by solving the crystal structure for the murine uPA.uPAR complex and demonstrate by extensive surface plasmon resonance studies that the kinetic rate constants for this interaction can be swapped completely between these orthologs by exchanging only two residues. This study not only discloses the structural basis required for a successful rational design of the species selectivity in the uPA.uPAR interaction, which is highly relevant for functional studies in mouse models, but it also suggests the possible development of general inhibitors that will target the uPA.uPAR interaction across species barriers.
Notes: 1083-351x
Lin, Lin
Gårdsvoll, Henrik
Huai, Qing
Huang, Mingdong
Ploug, Michael
F32 HL008658/HL/NHLBI NIH HHS/United States
R01 HL086584/HL/NHLBI NIH HHS/United States
P30 EB009998/EB/NIBIB NIH HHS/United States
R01 HL086584-04/HL/NHLBI NIH HHS/United States
HL08658/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Apr 2;285(14):10982-92. doi: 10.1074/jbc.M109.093492. Epub 2010 Feb 4.
Author Address: Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1189
Author: Lin, M. H., Lee, R. K., Hwu, Y. M., Lu, C. H., Chu, S. L., Chen, Y. J., Chang, W. C. and Li, S. H.
Year: 2008
Title: SPINKL, a Kazal-type serine protease inhibitor-like protein purified from mouse seminal vesicle fluid, is able to inhibit sperm capacitation
Journal: Reproduction
Volume: 136
Issue: 5
Pages: 559-71
Epub Date: 2008/08/22
Date: Nov
Short Title: SPINKL, a Kazal-type serine protease inhibitor-like protein purified from mouse seminal vesicle fluid, is able to inhibit sperm capacitation
Alternate Journal: Reproduction (Cambridge, England)
ISSN: 1470-1626
DOI: 10.1530/rep-07-0375
Accession Number: 18715980
Keywords: Acrosome Reaction/drug effects
Amino Acid Sequence
Animals
Blotting, Western/methods
Body Fluids/chemistry
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Female
Fertility/drug effects
Male
Mice
Molecular Sequence Data
Proteinase Inhibitory Proteins, Secretory/genetics/*isolation &
purification/pharmacology
RNA, Messenger/analysis
Seminal Vesicles/*chemistry
Sequence Alignment
Serine Proteinase Inhibitors/*pharmacology
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Sperm Capacitation/*drug effects
Spermatozoa/*drug effects
Abstract: We report a secreted serine protease inhibitor Kazal-type-like (SPINKL) protein. The SPINKL protein was purified from mouse seminal vesicle secretions through a series of steps, including ion-exchange chromatography on a diethylaminoethyl-Sephacel column, gel filtration on a Sephadex G-75 column, and ion-exchange HPLC on a Q strong anion exchange column. Further analysis identified several SPINKL proteins with various N-linked carbohydrates. The SPINKL protein has six conserved cysteine residues that are nearly identical to those of members of the SPINK protein family. It was noted that the SPINKL protein showed no inhibitory activities against common serine proteases such as trypsin, chymotrypsin, subtilisin, or elastase. Spinkl mRNA and SPINKL proteins were found to be primarily expressed in seminal vesicles. Immunohistochemistry revealed that the SPINKL protein occurred in the luminal fluid and mucosal epithelium of the seminal vesicles and was regulated by testosterone. The SPINKL protein was able to bind onto sperm and enhance sperm motility. Also, it was able to suppress BSA-stimulated sperm capacitation and block sperm-oocyte interactions in vitro, suggesting that SPINKL may be a decapacitation factor.
Notes: 1741-7899
Lin, Ming-Huei
Lee, Robert Kuo-Kuang
Hwu, Yuh-Ming
Lu, Chung-Hao
Chu, Shian-Ling
Chen, Ying-Jie
Chang, Wei-Chao
Li, Sheng-Hsiang
Journal Article
Research Support, Non-U.S. Gov't
England
Reproduction. 2008 Nov;136(5):559-71. doi: 10.1530/REP-07-0375. Epub 2008 Aug 20.
Author Address: Department of Obstetrics and Gynecology, Mackay Memorial Hospital, Mackay Medicine, Nursing and Management College, Taipei, Taiwan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1384
Author: Lin, S. J., Hu, Y., Zhu, J., Woodruff, T. K. and Jardetzky, T. S.
Year: 2011
Title: Structure of betaglycan zona pellucida (ZP)-C domain provides insights into ZP-mediated protein polymerization and TGF-beta binding
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 13
Pages: 5232-6
Epub Date: 2011/03/16
Date: Mar 29
Short Title: Structure of betaglycan zona pellucida (ZP)-C domain provides insights into ZP-mediated protein polymerization and TGF-beta binding
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1010689108
PMCID: PMC3069177
Accession Number: 21402931
Keywords: Animals
Crystallography, X-Ray
Mice
Models, Molecular
Molecular Sequence Data
Polymerization
Protein Binding
Protein Structure, Secondary
*Protein Structure, Tertiary
Proteoglycans/*chemistry/genetics/metabolism
Rats
Receptors, Transforming Growth Factor beta/*chemistry/genetics/metabolism
Transforming Growth Factor beta/*metabolism
Zona Pellucida/*metabolism
Abstract: The zona pellucida (ZP) domain is a bipartite protein structural element comprised of ZP-N and ZP-C regions. Most notable for its ability to mediate protein polymerization, many ZP proteins polymerize and assemble into long fibrils that form specialized extracellular matrices. Other ZP proteins (namely, betaglycan and endoglin) do not polymerize but serve as important membrane coreceptors for ligands in the transforming growth factor-β (TGF-β) superfamily. Here, we present the 2.0-Å resolution crystal structure of the betaglycan ZP-C region in combination with a downstream region known as the external hydrophobic patch (EHP). Similar to the ZP-N region, the ZP-C region also adopts an immunoglobulin-like fold, despite sharing no sequence homology and possessing different disulfide linkages. The EHP region, which was previously thought to be external to the ZP region, is integral to the ZP-C domain and corresponds to the ZP-C G strand. Our structure also indicates that the critical maturation cleavage of ZP proteins, a process that activates nascent ZP proteins for polymerization, occurs within the immunoglobulin domain at the FG loop. Nonpolymerizing ZP proteins such as betaglycan and endoglin do not contain this cleavage site. Finally, our structure suggests that the AB loop and the convex surface pocket are regions important for TGF-β ligand binding.
Notes: 1091-6490
Lin, S Jack
Hu, Yaoxiong
Zhu, Jie
Woodruff, Teresa K
Jardetzky, Theodore S
U54 HD041857/HD/NICHD NIH HHS/United States
R01 HD037096/HD/NICHD NIH HHS/United States
Canadian Institutes of Health Research/Canada
U54HD041857/HD/NICHD NIH HHS/United States
R01HD37096/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Mar 29;108(13):5232-6. doi: 10.1073/pnas.1010689108. Epub 2011 Mar 14.
Author Address: Department of Structural Biology, Stanford University School of Medicine, 371 Serra Mall, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1741
Author: Lin, W., Gao, L. and Chen, X.
Year: 2015
Title: Protein-Specific Imaging of O-GlcNAcylation in Single Cells
Journal: Chembiochem
Volume: 16
Issue: 18
Pages: 2571-5
Epub Date: 2015/10/22
Date: Dec
Short Title: Protein-Specific Imaging of O-GlcNAcylation in Single Cells
Alternate Journal: Chembiochem : a European journal of chemical biology
ISSN: 1439-4227
DOI: 10.1002/cbic.201500544
Accession Number: 26488919
Keywords: Acetylglucosamine/metabolism
Animals
CHO Cells
Cell Line, Tumor
Cricetinae
Cricetulus
Dogs
Fluorescence Recovery After Photobleaching
Glycosylation
Green Fluorescent Proteins/genetics/metabolism
Humans
Madin Darby Canine Kidney Cells
Microscopy, Fluorescence
Protein Processing, Post-Translational
Rhodamines/chemistry/metabolism
beta Catenin/genetics/*metabolism
beta-N-Acetylhexosaminidases/metabolism
tau Proteins/genetics/*metabolism
Flim
Fret
O-GlcNAc
glycan imaging
protein labeling
Abstract: Thousands of intracellular proteins are post-translationally modified with O-GlcNAc, and O-GlcNAcylation impacts the function of modified proteins and mediates diverse biological processes. However, the ubiquity of this important glycosylation makes it highly challenging to probe the O-GlcNAcylation state of a specific protein at the cellular level. Herein, we report the development of a FLIM-FRET-based strategy, which exploits the spatial proximity of the O-GlcNAc moiety and the attaching protein, for protein-specific imaging of O-GlcNAcylation in single cells. We demonstrated this strategy by imaging the O-GlcNAcylation state of tau and β-catenin inside the cells. Furthermore, the changes in tau O-GlcNAcylation were monitored when the overall cellular O-GlcNAc was pharmacologically altered by using the OGT and OGA inhibitors. We envision that the FLIM-FRET strategy will be broadly applicable to probe the O-GlcNAcylation state of various proteins in the cells.
Notes: 1439-7633
Lin, Wei
Gao, Ling
Chen, Xing
Journal Article
Research Support, Non-U.S. Gov't
Germany
Chembiochem. 2015 Dec;16(18):2571-5. doi: 10.1002/cbic.201500544. Epub 2015 Nov 13.
Author Address: Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, 100871, P. R. China.
Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, 100871, P. R. China. xingchen@pku.edu.cn.
Synthetic and Functional Biomolecules Center, Key Laboratory of Bioorganic Chemistry, and Molecular Engineering of Ministry of Education, Peking University, Beijing, 100871, P. R. China. xingchen@pku.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 590
Author: Lindqvist, Y., Schneider, G. and Vihko, P.
Year: 1993
Title: Three-dimensional structure of rat acid phosphatase in complex with L(+)-tartrate
Journal: J Biol Chem
Volume: 268
Issue: 28
Pages: 20744-6
Epub Date: 1993/10/05
Date: Oct 5
Short Title: Three-dimensional structure of rat acid phosphatase in complex with L(+)-tartrate
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8407898
Keywords: Acid Phosphatase/antagonists & inhibitors/*chemistry
Amino Acid Sequence
Animals
Hydrogen Bonding
Molecular Sequence Data
Protein Conformation
Rats
Recombinant Proteins/chemistry
Tartrates/*chemistry
Abstract: The crystal structure of recombinant rat prostatic acid phosphatase in complex with the inhibitor L(+)-tartrate was determined to 3-A resolution with protein crystallographic methods. The inhibitor binds at the carboxyl end of the parallel strands of the alpha/beta domain. One of the carboxyl groups of the tartrate molecule interacts with the conserved residues Arg-11, His-12, and Arg-15, which form part of the phosphate binding site. Furthermore, the C2 and C3 hydroxyl groups interact with His-257 and Arg-79. The second carboxyl group is close to Arg-79 but makes no direct hydrogen bonds to the protein. A sequence comparison between tartrate-sensitive and -resistant acid phosphatases suggests that these enzymes have different three-dimensional structures.
Notes: Lindqvist, Y
Schneider, G
Vihko, P
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1993 Oct 5;268(28):20744-6.
Author Address: Department of Molecular Biology, Swedish University of Agricultural Sciences, Uppsala Biomedical Center.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1075
Author: Liou, H. L., Dixit, S. S., Xu, S., Tint, G. S., Stock, A. M. and Lobel, P.
Year: 2006
Title: NPC2, the protein deficient in Niemann-Pick C2 disease, consists of multiple glycoforms that bind a variety of sterols
Journal: J Biol Chem
Volume: 281
Issue: 48
Pages: 36710-23
Epub Date: 2006/10/05
Date: Dec 1
Short Title: NPC2, the protein deficient in Niemann-Pick C2 disease, consists of multiple glycoforms that bind a variety of sterols
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M608743200
Accession Number: 17018531
Keywords: Animals
Biological Transport
Brain/metabolism
Carrier Proteins/chemistry/*physiology
Cholesterol/chemistry
Fibroblasts/metabolism
Glycoproteins/chemistry/*physiology
Glycosylation
Humans
Lipids/chemistry
Liver/metabolism
Mice
Niemann-Pick Diseases/*metabolism
Recombinant Proteins/chemistry
Sterols/*chemistry
Vesicular Transport Proteins
Abstract: Niemann-Pick C disease is a fatal neurodegenerative disorder characterized by an endolysosomal accumulation of cholesterol and other lipids. One form of the disease is caused by a deficiency in NPC2, a soluble lysosomal glycoprotein that binds cholesterol. To better understand the biological function of NPC2 and how its deficiency results in disease, we have characterized the structural and functional properties of recombinant human protein. Highly purified NPC2 consists of a complex mixture of glycosylated isoforms, similar to that observed in human brain autopsy specimens. Mass spectrometric analysis revealed that of the three potential N-linked glycosylation sites present in the mature protein, Asn-19 is not utilized; Asn-39 is linked to an endoglycosidase H (Endo H)-sensitive oligosaccharide, and Asn-116 is variably utilized, either being unmodified or linked to Endo H-sensitive or Endo H-resistant oligosaccharides. All glycoforms are endocytosed and ameliorate the cholesterol storage phenotype of NPC2-deficient fibroblasts. In addition, the purified preparation contains a mixture of both free and lipid-bound protein. All glycoforms bind cholesterol, and sterol binding to NPC2 significantly alters its behavior upon cation-exchange chromatography. Based on this observation, we developed chromatography-based binding assays and determined that NPC2 forms an equimolar complex with the fluorescent cholesterol analog dehydroergosterol. In addition, we find that NPC2 binds a range of cholesterol-related molecules (cholesterol precursors, plant sterols, some oxysterols, cholesterol sulfate, cholesterol acetate, and 5-alpha-cholestan-3-one) and that 27-hydroxysterol accumulates in NPC2-deficient mouse liver. Binding was not detected for various glycolipids, phospholipids, or fatty acids. These biochemical properties support a direct and specialized function of NPC2 in lysosomal sterol transport.
Notes: Liou, Heng-Ling
Dixit, Sayali S
Xu, Sujuan
Tint, G Stephen
Stock, Ann M
Lobel, Peter
Howard Hughes Medical Institute/United States
R01 DK054317/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Biol Chem. 2006 Dec 1;281(48):36710-23. doi: 10.1074/jbc.M608743200. Epub 2006 Oct 2.
Author Address: Center for Advanced Biotechnology and Medicine, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2075
Author: Lippold, S., Büttner, A., Choo, M. S. F., Hook, M., de Jong, C. J., Nguyen-Khuong, T., Haberger, M., Reusch, D., Wuhrer, M. and de Haan, N.
Year: 2020
Title: Cysteine Aminoethylation Enables the Site-Specific Glycosylation Analysis of Recombinant Human Erythropoietin using Trypsin
Journal: Anal Chem
Volume: 92
Issue: 14
Pages: 9476-9481
Epub Date: 2020/06/25
Date: Jul 21
Short Title: Cysteine Aminoethylation Enables the Site-Specific Glycosylation Analysis of Recombinant Human Erythropoietin using Trypsin
Alternate Journal: Analytical chemistry
ISSN: 0003-2700
DOI: 10.1021/acs.analchem.0c01794
Accession Number: 32578997
Keywords: Cysteine/*chemistry
Erythropoietin/*chemistry
Glycosylation
Humans
Recombinant Proteins/*chemistry
Trypsin/*chemistry
Abstract: Recombinant human erythropoietin (rhEPO) is an important biopharmaceutical for which glycosylation is a critical quality attribute. Therefore, robust analytical methods are needed for the in-depth characterization of rhEPO glycosylation. Currently, the protease GluC is widely established for the site-specific glycosylation analysis of rhEPO. However, this enzyme shows disadvantages, such as its specificity and the characteristics of the resulting (glyco)peptides. The use of trypsin, the gold standard protease in proteomics, as the sole protease for rhEPO is compromised, as no natural tryptic cleavage site is located between the glycosylation sites Asn24 and Asn38. Here, cysteine aminoethylation using 2-bromoethylamine was applied as an alternative alkylation strategy to introduce artificial tryptic cleavage sites at Cys29 and Cys33 in rhEPO. The (glyco)peptides resulting from a subsequent digestion using trypsin were analyzed by reverse-phase liquid chromatography-mass spectrometry. The new trypsin-based workflow was easily implemented by adapting the alkylation step in a conventional workflow and was directly compared to an established approach using GluC. The new method shows an improved specificity, a significantly reduced chromatogram complexity, allows for shorter analysis times, and simplifies data evaluation. Furthermore, the method allows for the monitoring of additional attributes, such as oxidation and deamidation at specific sites in parallel to the site-specific glycosylation analysis of rhEPO.
Notes: 1520-6882
Lippold, Steffen
Orcid: 0000-0002-1032-5808
Büttner, Alexander
Orcid: 0000-0003-3545-4208
Choo, Matthew S F
Orcid: 0000-0002-1376-3352
Hook, Michaela
de Jong, Coen J
Nguyen-Khuong, Terry
Orcid: 0000-0002-5852-542x
Haberger, Markus
Reusch, Dietmar
Wuhrer, Manfred
Orcid: 0000-0002-0814-4995
de Haan, Noortje
Orcid: 0000-0001-7026-6750
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Chem. 2020 Jul 21;92(14):9476-9481. doi: 10.1021/acs.analchem.0c01794. Epub 2020 Jul 9.
Author Address: Center for Proteomics and Metabolomics, Leiden University Medical Center, Albinusdreef 2, 2333 ZA Leiden, The Netherlands.
Pharma Technical Development, Roche Diagnostics GmbH, Nonnenwald 2, 82377 Penzberg, Germany.
Bioprocessing Technology Institute, Agency for Science Technology and Research, 20 Biopolis Way No. 06-01, Singapore 138668.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1701
Author: Lira-Navarrete, E., de Las Rivas, M., Compañón, I., Pallarés, M. C., Kong, Y., Iglesias-Fernández, J., Bernardes, G. J., Peregrina, J. M., Rovira, C., Bernadó, P., Bruscolini, P., Clausen, H., Lostao, A., Corzana, F. and Hurtado-Guerrero, R.
Year: 2015
Title: Dynamic interplay between catalytic and lectin domains of GalNAc-transferases modulates protein O-glycosylation
Journal: Nat Commun
Volume: 6
Pages: 6937
Epub Date: 2015/05/06
Date: May 5
Short Title: Dynamic interplay between catalytic and lectin domains of GalNAc-transferases modulates protein O-glycosylation
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms7937
PMCID: PMC4432651
Accession Number: 25939779
Keywords: *Catalytic Domain
Crystallography, X-Ray
Glycopeptides/chemistry/metabolism
Glycosylation
Lectins/*chemistry
Models, Molecular
Molecular Dynamics Simulation
N-Acetylgalactosaminyltransferases/*chemistry/*metabolism
Nucleotides/metabolism
Structure-Activity Relationship
Substrate Specificity
Abstract: Protein O-glycosylation is controlled by polypeptide GalNAc-transferases (GalNAc-Ts) that uniquely feature both a catalytic and lectin domain. The underlying molecular basis of how the lectin domains of GalNAc-Ts contribute to glycopeptide specificity and catalysis remains unclear. Here we present the first crystal structures of complexes of GalNAc-T2 with glycopeptides that together with enhanced sampling molecular dynamics simulations demonstrate a cooperative mechanism by which the lectin domain enables free acceptor sites binding of glycopeptides into the catalytic domain. Atomic force microscopy and small-angle X-ray scattering experiments further reveal a dynamic conformational landscape of GalNAc-T2 and a prominent role of compact structures that are both required for efficient catalysis. Our model indicates that the activity profile of GalNAc-T2 is dictated by conformational heterogeneity and relies on a flexible linker located between the catalytic and the lectin domains. Our results also shed light on how GalNAc-Ts generate dense decoration of proteins with O-glycans.
Notes: 2041-1723
Lira-Navarrete, Erandi
de Las Rivas, Matilde
Compañón, Ismael
Pallarés, María Carmen
Kong, Yun
Iglesias-Fernández, Javier
Bernardes, Gonçalo J L
Peregrina, Jesús M
Rovira, Carme
Bernadó, Pau
Bruscolini, Pierpaolo
Clausen, Henrik
Lostao, Anabel
Corzana, Francisco
Hurtado-Guerrero, Ramon
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2015 May 5;6:6937. doi: 10.1038/ncomms7937.
Author Address: BIFI, University of Zaragoza, BIFI-IQFR (CSIC) Joint Unit, Mariano Esquillor s/n, Campus Rio Ebro, Edificio I+D, Zaragoza 50018, Spain.
Departamento de Química, Universidad de La Rioja, Centro de Investigación en Síntesis Química, E-26006 Logroño, Spain.
LMA, INA, Universidad de Zaragoza, 50018 Zaragoza, Spain.
Copenhagen Center for Glycomics, Department of Cellular and Molecular Medicine, School of Dentistry, University of Copenhagen, Copenhagen DK-2200, Denmark.
Departament de Química Orgànica i IQTCUB, Universitat de Barcelona, Martí i Franquès 1, 08028 Barcelona, Spain.
1] Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, UK [2] Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Av Prof Egas Moniz, 1649-028 Lisboa, Portugal.
1] Departament de Química Orgànica i IQTCUB, Universitat de Barcelona, Martí i Franquès 1, 08028 Barcelona, Spain [2] ICREA, Passeig Lluís Companys 23, 08020 Barcelona, Spain.
Centre de Biochimie Structurale, INSERM U1054, CNRS UMR 5048, Université Montpellier 1 and 2, 29 rue de Navacelles, 34090 Montpellier, France.
1] BIFI, University of Zaragoza, BIFI-IQFR (CSIC) Joint Unit, Mariano Esquillor s/n, Campus Rio Ebro, Edificio I+D, Zaragoza 50018, Spain [2] Departamento de Física Teórica, Universidad de Zaragoza, Zaragoza 50009, Spain.
1] LMA, INA, Universidad de Zaragoza, 50018 Zaragoza, Spain [2] Fundación ARAID, 50018 Zaragoza, Spain.
1] BIFI, University of Zaragoza, BIFI-IQFR (CSIC) Joint Unit, Mariano Esquillor s/n, Campus Rio Ebro, Edificio I+D, Zaragoza 50018, Spain [2] Fundación ARAID, 50018 Zaragoza, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1825
Author: Liu, B., Wang, J., Li, M., Yuan, Q., Xue, M., Xu, F. and Chen, Y.
Year: 2017
Title: Inhibition of ALDH2 by O-GlcNAcylation contributes to the hyperglycemic exacerbation of myocardial ischemia/reperfusion injury
Journal: Oncotarget
Volume: 8
Issue: 12
Pages: 19413-19426
Epub Date: 2016/12/31
Date: Mar 21
Short Title: Inhibition of ALDH2 by O-GlcNAcylation contributes to the hyperglycemic exacerbation of myocardial ischemia/reperfusion injury
Alternate Journal: Oncotarget
ISSN: 1949-2553
DOI: 10.18632/oncotarget.14297
PMCID: PMC5386694
Accession Number: 28038474
Keywords: Acetylglucosamine/*metabolism
Aldehyde Dehydrogenase/chemistry/*metabolism
Animals
Apoptosis
Cell Proliferation
Cells, Cultured
Glycosylation
Humans
Hyperglycemia/enzymology/etiology/*pathology
Male
Myocardial Reperfusion Injury/*complications
Myocytes, Cardiac/drug effects/enzymology/*pathology
*Protein Processing, Post-Translational
Rats
Rats, Wistar
Aldh2
O-GlcNAcylation
hyperglycemia
myocardial ischemia/reperfusion
Abstract: Although hyperglycemia is causally related to adverse outcomes after myocardial ischemia/reperfusion (I/R), the underlying mechanisms are largely unknown. Here, we investigated whether excessive O-linked-N-acetylglucosamine (O-GlcNAc) modification of acetaldehyde dehydrogenase 2 (ALDH2), an important cardioprotective enzyme, was a mechanism for the hyperglycemic exacerbation of myocardial I/R injury. Both acute hyperglycemia (AHG) and diabetes (DM)-induced chronic hyperglycemia increased cardiac dysfunction, infarct size and apoptosis index compared with normal saline (NS)+I/R rats (P<0.05). ALDH2 O-GlcNAc modification was increased whereas its activity was decreased in AHG+I/R and DM+I/R rats. High glucose (HG, 30mmol/L) markedly increased ALDH2 O-GlcNAc modification compared with Con group (5mmol/L) (P<0.05). ALDH2 O-GlcNAc modification was increased by 62.9% in Con+PUGNAc group whereas it was decreased by 44.1% in Con+DON group compared with Con group (P<0.05). Accordingly, ALDH2 activity was decreased by 18.1% in Con+PUGNAc group whereas it was increased by 17.9% in Con+DON group. Moreover, DON decreased levels of 4-hydroxy-2-nonenal (4-HNE), aldehydes, protein carbonyl accumulation and apoptosis index compared with HG+H/R group (P<0.05). Alda-1, a specific activator of ALDH2, significantly decreased ALDH2 O-GlcNAc modification and improved infarct size, apoptosis index and cardiac dysfunction induced by I/R combined with hyperglycemia. These findings demonstrate that ALDH2 O-GlcNAc modification is a key mechanism for the hyperglycemic exacerbation of myocardial I/R injury and Alda-1 has therapeutic potential for inducing cardioprotection.
Notes: 1949-2553
Liu, Baoshan
Wang, Jiali
Li, Minghua
Yuan, Qiuhuan
Xue, Mengyang
Xu, Feng
Chen, Yuguo
Journal Article
Oncotarget. 2017 Mar 21;8(12):19413-19426. doi: 10.18632/oncotarget.14297.
Author Address: Department of Emergency, Qilu Hospital, Shandong University, Jinan, China.
Chest Pain Center, Qilu Hospital, Shandong University, Jinan, China.
Key Laboratory of Emergency and Critical Care Medicine of Shandong Province, Qilu Hospital, Shandong University, Jinan, China.
Key Laboratory of Cardiovascular Remodeling & Function Research, Chinese Ministry of Education & Chinese Ministry of Public Health, Qilu Hospital, Shandong University, Jinan, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2069
Author: Liu, C., Shi, Y., Li, J., Liu, X., Xiahou, Z., Tan, Z., Chen, X. and Li, J.
Year: 2020
Title: O-GlcNAcylation of myosin phosphatase targeting subunit 1 (MYPT1) dictates timely disjunction of centrosomes
Journal: J Biol Chem
Volume: 295
Issue: 21
Pages: 7341-7349
Epub Date: 2020/04/17
Date: May 22
Short Title: O-GlcNAcylation of myosin phosphatase targeting subunit 1 (MYPT1) dictates timely disjunction of centrosomes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA119.012401
PMCID: PMC7247298
Accession Number: 32295844
Keywords: CDC2 Protein Kinase/genetics/metabolism
Cell Cycle Proteins/genetics/metabolism
Cell Line
Centrosome/*metabolism
Glycosylation
Humans
*Mitosis
Myosin-Light-Chain Phosphatase/genetics/*metabolism
Protein-Serine-Threonine Kinases/genetics/metabolism
Proto-Oncogene Proteins/genetics/metabolism
*Cdk1
*Mypt1
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc)
*Plk1
*cell cycle
*centrosome
*protein phosphorylation
this article
Abstract: The role of O-linked N-acetylglucosamine (O-GlcNAc) modification in the cell cycle has been enigmatic. Previously, both O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) disruptions have been shown to derail the mitotic centrosome numbers, suggesting that mitotic O-GlcNAc oscillation needs to be in concert with mitotic progression to account for centrosome integrity. Here, using both chemical approaches and biological assays with HeLa cells, we attempted to address the underlying molecular mechanism and observed that incubation of the cells with the OGA inhibitor Thiamet-G strikingly elevates centrosomal distances, suggestive of premature centrosome disjunction. These aberrations could be overcome by inhibiting Polo-like kinase 1 (PLK1), a mitotic master kinase. PLK1 inactivation is modulated by the myosin phosphatase targeting subunit 1 (MYPT1)-protein phosphatase 1cβ (PP1cβ) complex. Interestingly, MYPT1 has been shown to be abundantly O-GlcNAcylated, and the modified residues have been detected in a recent O-GlcNAc-profiling screen utilizing chemoenzymatic labeling and bioorthogonal conjugation. We demonstrate here that MYPT1 is O-GlcNAcylated at Thr-577, Ser-585, Ser-589, and Ser-601, which antagonizes CDK1-dependent phosphorylation at Ser-473 and attenuates the association between MYPT1 and PLK1, thereby promoting PLK1 activity. We conclude that under high O-GlcNAc levels, PLK1 is untimely activated, conducive to inopportune centrosome separation and disruption of the cell cycle. We propose that too much O-GlcNAc is equally deleterious as too little O-GlcNAc, and a fine balance between the OGT/OGA duo is indispensable for successful mitotic divisions.
Notes: 1083-351x
Liu, Caifei
Shi, Yingxin
Li, Jie
Liu, Xuewen
Xiahou, Zhikai
Tan, Zhongping
Chen, Xing
Li, Jing
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2020 May 22;295(21):7341-7349. doi: 10.1074/jbc.RA119.012401. Epub 2020 Apr 15.
Author Address: Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China.
Department of Radiation Oncology, Hunan Cancer Hospital and the Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha 410006, China; Key Laboratory of Translational Radiation Oncology, Hunan 410006, China.
Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China.
College of Chemistry and Molecular Engineering, Beijing National Laboratory for Molecular Sciences, Peking-Tsinghua Center for Life Sciences, Synthetic and Functional Biomolecules Center, and Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University, Beijing 100871, China.
Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China. Electronic address: jing_li@mail.cnu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1002
Author: Liu, D., Cramer, C. C., Scafidi, J. and Davis, A. E., 3rd
Year: 2005
Title: N-linked glycosylation at Asn3 and the positively charged residues within the amino-terminal domain of the c1 inhibitor are required for interaction of the C1 Inhibitor with Salmonella enterica serovar typhimurium lipopolysaccharide and lipid A
Journal: Infect Immun
Volume: 73
Issue: 8
Pages: 4478-87
Epub Date: 2005/07/26
Date: Aug
Short Title: N-linked glycosylation at Asn3 and the positively charged residues within the amino-terminal domain of the c1 inhibitor are required for interaction of the C1 Inhibitor with Salmonella enterica serovar typhimurium lipopolysaccharide and lipid A
Alternate Journal: Infection and immunity
ISSN: 0019-9567 (Print)
0019-9567
DOI: 10.1128/iai.73.8.4478-4487.2005
PMCID: PMC1201182
Accession Number: 16040958
Keywords: Amino Acid Substitution
Animals
Asparagine/*metabolism
COS Cells
Chlorocebus aethiops
Complement C1 Inactivator Proteins
Complement C1 Inhibitor Protein
Glycosylation
Humans
Lipid A/*metabolism
Mice
Mutation
Protein Structure, Tertiary
Salmonella typhimurium/*metabolism
Serpins/genetics/*metabolism
Abstract: The C1 inhibitor (C1INH), a plasma complement regulatory protein, prevents endotoxin shock, at least partially via the direct interaction of its amino-terminal heavily glycosylated nonserpin region with gram-negative bacterial lipopolysaccharide (LPS). To further characterize the potential LPS-binding site(s) within the amino-terminal domain, mutations were introduced into C1INH at the three N-linked glycosylation sites and at the four positively charged amino acid residues. A mutant in which Asn(3) was replaced with Ala was markedly less effective in its binding to LPS, while substitution of Asn(47) or Asn(59) had little effect on binding. The mutation of C1INH at all four positively charged amino acid residues (Arg(18), Lys(22), Lys(30), and Lys(55)) resulted in near-complete failure to interact with LPS. The C1INH mutants that did not bind to LPS also did not suppress LPS binding or LPS-induced up-regulation of tumor necrosis factor alpha mRNA expression in RAW 264.7 macrophages. In addition, the binding of C1INH mutants to diphosphoryl lipid A was decreased in comparison with that of recombinant wild-type C1INH. Therefore, the interaction of C1INH with gram-negative bacterial LPS is dependent both on the N-linked carbohydrate at Asn(3) and on the positively charged residues within the amino-terminal domain.
Notes: 1098-5522
Liu, Dongxu
Cramer, Cort C
Scafidi, Jennifer
Davis, Alvin E 3rd
R01 HD033727/HD/NICHD NIH HHS/United States
R37 HD022082/HD/NICHD NIH HHS/United States
HD22082/HD/NICHD NIH HHS/United States
HD33727/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, P.H.S.
Infect Immun. 2005 Aug;73(8):4478-87. doi: 10.1128/IAI.73.8.4478-4487.2005.
Author Address: The CBR Institute for Biomedical Research, Harvard Medical School, 800 Huntington Avenue, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 942
Author: Liu, F., Iqbal, K., Grundke-Iqbal, I., Hart, G. W. and Gong, C. X.
Year: 2004
Title: O-GlcNAcylation regulates phosphorylation of tau: a mechanism involved in Alzheimer's disease
Journal: Proc Natl Acad Sci U S A
Volume: 101
Issue: 29
Pages: 10804-9
Epub Date: 2004/07/14
Date: Jul 20
Short Title: O-GlcNAcylation regulates phosphorylation of tau: a mechanism involved in Alzheimer's disease
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0400348101
PMCID: PMC490015
Accession Number: 15249677
Keywords: Acetylglucosamine/*metabolism
Aged
Aged, 80 and over
Alzheimer Disease/*metabolism/pathology
Animals
Brain/*metabolism
Female
Glycosylation
Humans
In Vitro Techniques
Male
Mice
Mice, Inbred C57BL
Middle Aged
PC12 Cells
Phosphorylation
Rats
Rats, Wistar
Starvation
tau Proteins/genetics/*metabolism
Abstract: Microtubule-associated protein tau is abnormally hyperphosphorylated and aggregated into neurofibrillary tangles in brains of individuals with Alzheimer's disease (AD) and other tauopathies. Tau pathology is critical to pathogenesis and correlates to the severity of dementia. However, the mechanisms leading to abnormal hyperphosphorylation are unknown. Here, we demonstrate that human brain tau was modified by O-GlcNAcylation, a type of protein O-glycosylation by which the monosaccharide beta-N-acetylglucosamine (GlcNAc) attaches to serine/threonine residues via an O-linked glycosidic bond. O-GlcNAcylation regulated phosphorylation of tau in a site-specific manner both in vitro and in vivo. At most of the phosphorylation sites, O-GlcNAcylation negatively regulated tau phosphorylation. In an animal model of starved mice, low glucose uptake/metabolism that mimicked those observed in AD brain produced a decrease in O-GlcNAcylation and consequent hyperphosphorylation of tau at the majority of the phosphorylation sites. The O-GlcNAcylation level in AD brain extracts was decreased as compared to that in controls. These results reveal a mechanism of regulation of tau phosphorylation and suggest that abnormal hyperphosphorylation of tau could result from decreased tau O-GlcNAcylation, which probably is induced by deficient brain glucose uptake/metabolism in AD and other tauopathies.
Notes: 1091-6490
Liu, Fei
Iqbal, Khalid
Grundke-Iqbal, Inge
Hart, Gerald W
Gong, Cheng-Xin
AG16760/AG/NIA NIH HHS/United States
AG19158/AG/NIA NIH HHS/United States
R01 AG016760/AG/NIA NIH HHS/United States
R01 AG019158/AG/NIA NIH HHS/United States
R37 HD013563/HD/NICHD NIH HHS/United States
HD13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2004 Jul 20;101(29):10804-9. doi: 10.1073/pnas.0400348101. Epub 2004 Jul 12.
Author Address: Department of Neurochemistry, New York State Institute for Basic Research in Developmental Disabilities, 1050 Forest Hill Road, Staten Island, NY 10314, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1242
Author: Liu, F., Shi, J., Tanimukai, H., Gu, J., Gu, J., Grundke-Iqbal, I., Iqbal, K. and Gong, C. X.
Year: 2009
Title: Reduced O-GlcNAcylation links lower brain glucose metabolism and tau pathology in Alzheimer's disease
Journal: Brain
Volume: 132
Issue: Pt 7
Pages: 1820-32
Epub Date: 2009/05/20
Date: Jul
Short Title: Reduced O-GlcNAcylation links lower brain glucose metabolism and tau pathology in Alzheimer's disease
Alternate Journal: Brain : a journal of neurology
ISSN: 0006-8950 (Print)
0006-8950
DOI: 10.1093/brain/awp099
PMCID: PMC2702834
Accession Number: 19451179
Keywords: Aged, 80 and over
Alzheimer Disease/*metabolism/pathology
Brain/*metabolism
Cells, Cultured
Down-Regulation
Female
Glucose/*metabolism
Hexosamines/biosynthesis
Humans
Male
N-Acetylglucosaminyltransferases/genetics/metabolism/*physiology
Phosphorylation
Protein Phosphatase 2/metabolism
tau Proteins/*metabolism
Abstract: It has been established for a long time that brain glucose metabolism is impaired in Alzheimer's disease. Recent studies have demonstrated that impaired brain glucose metabolism precedes the appearance of clinical symptoms, implying its active role in the development of Alzheimer's disease. However, the molecular mechanism by which this impairment contributes to the disease is not known. In this study, we demonstrated that protein O-GlcNAcylation, a common post-translational modification of nucleocytoplasmic proteins with beta-N-acetyl-glucosamine and a process regulated by glucose metabolism, was markedly decreased in Alzheimer's disease cerebrum. More importantly, the decrease in O-GlcNAc correlated negatively with phosphorylation at most phosphorylation sites of tau protein, which is known to play a crucial role in the neurofibrillary degeneration of Alzheimer's disease. We also found that hyperphosphorylated tau contained 4-fold less O-GlcNAc than non-hyperphosphorylated tau, demonstrating for the first time an inverse relationship between O-GlcNAcylation and phosphorylation of tau in the human brain. Downregulation of O-GlcNAcylation by knockdown of O-GlcNAc transferase with small hairpin RNA led to increased phosphorylation of tau in HEK-293 cells. Inhibition of the hexosamine biosynthesis pathway in rat brain resulted in decreased O-GlcNAcylation and increased phosphorylation of tau, which resembled changes of O-GlcNAcylation and phosphorylation of tau in rodent brains with decreased glucose metabolism induced by fasting, but not those in rat brains when protein phosphatase 2A was inhibited. Comparison of tau phosphorylation patterns under various conditions suggests that abnormal tau hyperphosphorylation in Alzheimer's disease brain may result from downregulation of both O-GlcNAcylation and protein phosphatase 2A. These findings suggest that impaired brain glucose metabolism leads to abnormal hyperphosphorylation of tau and neurofibrillary degeneration via downregulation of tau O-GlcNAcylation in Alzheimer's disease. Thus, restoration of brain tau O-GlcNAcylation and protein phosphatase 2A activity may offer promising therapeutic targets for treating Alzheimer's disease.
Notes: 1460-2156
Liu, Fei
Shi, Jianhua
Tanimukai, Hitoshi
Gu, Jinhua
Gu, Jianlan
Grundke-Iqbal, Inge
Iqbal, Khalid
Gong, Cheng-Xin
P30 AG019610/AG/NIA NIH HHS/United States
P30 AG19 610/AG/NIA NIH HHS/United States
R01 AG019 158/AG/NIA NIH HHS/United States
R01 AG027 429/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Brain. 2009 Jul;132(Pt 7):1820-32. doi: 10.1093/brain/awp099. Epub 2009 May 18.
Author Address: Department of Neurochemistry, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York 10314, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1090
Author: Liu, H., Chen, X., Focia, P. J. and He, X.
Year: 2007
Title: Structural basis for stem cell factor-KIT signaling and activation of class III receptor tyrosine kinases
Journal: Embo j
Volume: 26
Issue: 3
Pages: 891-901
Epub Date: 2007/01/27
Date: Feb 7
Short Title: Structural basis for stem cell factor-KIT signaling and activation of class III receptor tyrosine kinases
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7601545
PMCID: PMC1794399
Accession Number: 17255936
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cloning, Molecular
Crystallization
Dimerization
Mice/*genetics
*Models, Molecular
Molecular Conformation
Molecular Sequence Data
Protein Binding
Protein Structure, Tertiary
Protein-Tyrosine Kinases/*metabolism
Proto-Oncogene Proteins c-kit/*genetics/metabolism
Sequence Analysis, DNA
Signal Transduction/*genetics
Stem Cell Factor/*genetics/metabolism
Abstract: Stem cell factor (SCF) binds to and activates the KIT receptor, a class III receptor tyrosine kinase (RTK), to stimulate diverse processes including melanogenesis, gametogenesis and hematopoeisis. Dysregulation of KIT activation is associated with many cancers. We report a 2.5 A crystal structure of the functional core of SCF bound to the extracellular ligand-binding domains of KIT. The structure reveals a 'wrapping' SCF-recognition mode by KIT, in which KIT adopts a bent conformation to facilitate each of its first three immunoglobulin (Ig)-like domains to interact with SCF. Three surface epitopes on SCF, an extended loop, the B and C helices, and the N-terminal segment, contact distinct KIT domains, with two of the epitopes undergoing large conformational changes upon receptor binding. The SCF/KIT complex reveals a unique RTK dimerization assembly, and a novel recognition mode between four-helix bundle cytokines and Ig-family receptors. It serves as a framework for understanding the activation mechanisms of class III RTKs.
Notes: 1460-2075
Liu, Heli
Chen, Xiaoyan
Focia, Pamela J
He, Xiaolin
P41 RR015301/RR/NCRR NIH HHS/United States
R01 GM078055/GM/NIGMS NIH HHS/United States
RR-15301/RR/NCRR NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
EMBO J. 2007 Feb 7;26(3):891-901. doi: 10.1038/sj.emboj.7601545. Epub 2007 Jan 25.
Author Address: Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1355
Author: Liu, H., Focia, P. J. and He, X.
Year: 2011
Title: Homophilic adhesion mechanism of neurofascin, a member of the L1 family of neural cell adhesion molecules
Journal: J Biol Chem
Volume: 286
Issue: 1
Pages: 797-805
Epub Date: 2010/11/05
Date: Jan 7
Short Title: Homophilic adhesion mechanism of neurofascin, a member of the L1 family of neural cell adhesion molecules
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.180281
PMCID: PMC3013039
Accession Number: 21047790
Keywords: Animals
Cell Adhesion
Cell Adhesion Molecules/*chemistry/genetics/*metabolism
Cell Line
Crystallography, X-Ray
Humans
Immunoglobulins/chemistry
Models, Molecular
Mutagenesis
Nerve Growth Factors/*chemistry/genetics/*metabolism
Neurons/cytology/*metabolism
Protein Multimerization
Protein Structure, Quaternary
Protein Structure, Tertiary
Abstract: The L1 family neural cell adhesion molecules play key roles in specifying the formation and remodeling of the neural network, but their homophilic interaction that mediates adhesion is not well understood. We report two crystal structures of a dimeric form of the headpiece of neurofascin, an L1 family member. The four N-terminal Ig-like domains of neurofascin form a horseshoe shape, akin to several other immunoglobulin superfamily cell adhesion molecules such as hemolin, axonin, and Dscam. The neurofascin dimer, captured in two crystal forms with independent packing patterns, reveals a pair of horseshoes in trans-synaptic adhesion mode. The adhesion interaction is mediated mostly by the second Ig-like domain, which features an intermolecular β-sheet formed by the joining of two individual GFC β-sheets and a large but loosely packed hydrophobic cluster. Mutagenesis combined with gel filtration assays suggested that the side chain hydrogen bonds at the intermolecular β-sheet are essential for the homophilic interaction and that the residues at the hydrophobic cluster play supplementary roles. Our structures reveal a conserved homophilic adhesion mode for the L1 family and also shed light on how the pathological mutations of L1 affect its structure and function.
Notes: 1083-351x
Liu, Heli
Focia, Pamela J
He, Xiaolin
Journal Article
J Biol Chem. 2011 Jan 7;286(1):797-805. doi: 10.1074/jbc.M110.180281. Epub 2010 Nov 3.
Author Address: Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1333
Author: Liu, H., Juo, Z. S., Shim, A. H., Focia, P. J., Chen, X., Garcia, K. C. and He, X.
Year: 2010
Title: Structural basis of semaphorin-plexin recognition and viral mimicry from Sema7A and A39R complexes with PlexinC1
Journal: Cell
Volume: 142
Issue: 5
Pages: 749-61
Epub Date: 2010/08/24
Date: Sep 3
Short Title: Structural basis of semaphorin-plexin recognition and viral mimicry from Sema7A and A39R complexes with PlexinC1
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2010.07.040
PMCID: PMC2936782
NIHMSID: NIHMS227807
Accession Number: 20727575
Keywords: Amino Acid Sequence
Antigens, CD/*chemistry/metabolism
Crystallography, X-Ray
GPI-Linked Proteins
Humans
Models, Molecular
*Molecular Mimicry
Molecular Sequence Data
Protein Multimerization
Receptors, Virus/*chemistry/metabolism
Semaphorins/*chemistry/metabolism
Sequence Alignment
Vaccinia virus/*chemistry
Viral Proteins/*chemistry/metabolism
Abstract: Repulsive signaling by Semaphorins and Plexins is crucial for the development and homeostasis of the nervous, immune, and cardiovascular systems. Sema7A acts as both an immune and a neural Semaphorin through PlexinC1, and A39R is a Sema7A mimic secreted by smallpox virus. We report the structures of Sema7A and A39R complexed with the Semaphorin-binding module of PlexinC1. Both structures show two PlexinC1 molecules symmetrically bridged by Semaphorin dimers, in which the Semaphorin and PlexinC1 beta propellers interact in an edge-on, orthogonal orientation. Both binding interfaces are dominated by the insertion of the Semaphorin's 4c-4d loop into a deep groove in blade 3 of the PlexinC1 propeller. A39R appears to achieve Sema7A mimicry by preserving key Plexin-binding determinants seen in the mammalian Sema7A complex that have evolved to achieve higher affinity binding to the host-derived PlexinC1. The complex structures support a conserved Semaphorin-Plexin recognition mode and suggest that Plexins are activated by dimerization.
Notes: 1097-4172
Liu, Heli
Juo, Z Sean
Shim, Ann Hye-Ryong
Focia, Pamela J
Chen, Xiaoyan
Garcia, K Christopher
He, Xiaolin
1R01GM07805/GM/NIGMS NIH HHS/United States
R01 GM098259/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 GM078055/GM/NIGMS NIH HHS/United States
R01 AI051321/AI/NIAID NIH HHS/United States
R01-AI51321/AI/NIAID NIH HHS/United States
R01 AI051321-09/AI/NIAID NIH HHS/United States
R01 GM078055-05/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell. 2010 Sep 3;142(5):749-61. doi: 10.1016/j.cell.2010.07.040. Epub 2010 Aug 19.
Author Address: Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1264
Author: Liu, H., Shim, A. H. and He, X.
Year: 2009
Title: Structural characterization of the ectodomain of a disintegrin and metalloproteinase-22 (ADAM22), a neural adhesion receptor instead of metalloproteinase: insights on ADAM function
Journal: J Biol Chem
Volume: 284
Issue: 42
Pages: 29077-86
Epub Date: 2009/08/21
Date: Oct 16
Short Title: Structural characterization of the ectodomain of a disintegrin and metalloproteinase-22 (ADAM22), a neural adhesion receptor instead of metalloproteinase: insights on ADAM function
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.014258
PMCID: PMC2781453
Accession Number: 19692335
Keywords: ADAM Proteins/*chemistry
Amino Acid Sequence
Animals
Calorimetry/methods
Cell Adhesion Molecules, Neuronal/*metabolism
Cysteine/chemistry
Dimerization
Humans
Insecta
Metalloproteases/*chemistry
Models, Molecular
Molecular Conformation
Molecular Sequence Data
Nerve Tissue Proteins/*chemistry
Peptides/chemistry
Protein Structure, Tertiary
Abstract: ADAMs (a disintegrin and metalloproteinases) are a family of multidomain transmembrane glycoproteins with diverse roles in physiology and diseases, with several members being drug targets for cancer and inflammation therapies. The spatial organization of the ADAM extracellular segment and its influence on the function of ADAMs have been unclear. Although most members of the ADAM family are active zinc metalloproteinases, 8 of 21 ADAMs lack functional metalloproteinase domains and are implicated in protein-protein interactions instead of membrane protein ectodomain shedding. One of such non-proteinase ADAMs, ADAM22, acts as a receptor on the surface of the postsynaptic neuron to regulate synaptic signal transmission. The crystal structure of the full ectodomain of mature human ADAM22 shows that it is a compact four-leaf clover with the metalloproteinase-like domain held in the concave face of a rigid module formed by the disintegrin, cysteine-rich, and epidermal growth factor-like domains. The loss of metalloproteinase activity is ensured by the absence of critical catalytic residues, the filling of the substrate groove, and the steric hindrance by the cysteine-rich domain. The structure, combined with calorimetric experiments, suggests distinct roles of three putative calcium ions bound to ADAM22, with one in the metalloproteinase-like domain being regulatory and two in the disintegrin domain being structural. The metalloproteinase-like domain contacts the rest of ADAM22 with discontinuous, hydrophilic, and poorly complemented interactions, suggesting the possibility of modular movement of ADAM22 and other ADAMs. The ADAM22 structure provides a framework for understanding how different ADAMs exert their adhesive function and shedding activities.
Notes: 1083-351x
Liu, Heli
Shim, Ann H R
He, Xiaolin
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2009 Oct 16;284(42):29077-86. doi: 10.1074/jbc.M109.014258. Epub 2009 Aug 18.
Author Address: Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1645
Author: Liu, H., Yu, S., Zhang, H. and Xu, J.
Year: 2014
Title: Identification of nitric oxide as an endogenous inhibitor of 26S proteasomes in vascular endothelial cells
Journal: PLoS One
Volume: 9
Issue: 5
Pages: e98486
Epub Date: 2014/05/24
Short Title: Identification of nitric oxide as an endogenous inhibitor of 26S proteasomes in vascular endothelial cells
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0098486
PMCID: PMC4031199
Accession Number: 24853093
Keywords: Animals
Endothelium, Vascular/cytology/*enzymology
Human Umbilical Vein Endothelial Cells
Humans
Male
Mice
Mice, Inbred C57BL
Nitric Oxide/*physiology
Proteasome Endopeptidase Complex/*drug effects
Abstract: The 26S proteasome plays a fundamental role in almost all eukaryotic cells, including vascular endothelial cells. However, it remains largely unknown how proteasome functionality is regulated in the vasculature. Endothelial nitric oxide (NO) synthase (eNOS)-derived NO is known to be essential to maintain endothelial homeostasis. The aim of the present study was to establish the connection between endothelial NO and 26S proteasome functionality in vascular endothelial cells. The 26S proteasome reporter protein levels, 26S proteasome activity, and the O-GlcNAcylation of Rpt2, a key subunit of the proteasome regulatory complex, were assayed in 26S proteasome reporter cells, human umbilical vein endothelial cells (HUVEC), and mouse aortic tissues isolated from 26S proteasome reporter and eNOS knockout mice. Like the other selective NO donors, NO derived from activated eNOS (by pharmacological and genetic approach) increased O-GlcNAc modification of Rpt2, reduced proteasome chymotrypsin-like activity, and caused 26S proteasome reporter protein accumulation. Conversely, inactivation of eNOS reversed all the effects. SiRNA knockdown of O-GlcNAc transferase (OGT), the key enzyme that catalyzes protein O-GlcNAcylation, abolished NO-induced effects. Consistently, adenoviral overexpression of O-GlcNAcase (OGA), the enzyme catalyzing the removal of the O-GlcNAc group, mimicked the effects of OGT knockdown. Finally, compared to eNOS wild type aortic tissues, 26S proteasome reporter mice lacking eNOS exhibited elevated 26S proteasome functionality in parallel with decreased Rpt2 O-GlcNAcylation, without changing the levels of Rpt2 protein. In conclusion, the eNOS-derived NO functions as a physiological suppressor of the 26S proteasome in vascular endothelial cells.
Notes: 1932-6203
Liu, Hongtao
Yu, Shujie
Zhang, Hua
Xu, Jian
10SDG2600164/AHA/American Heart Association-American Stroke Association/United States
P20 RR 024215-05/RR/NCRR NIH HHS/United States
P20 GM104934/GM/NIGMS NIH HHS/United States
P20 RR024215/RR/NCRR NIH HHS/United States
14BGIA20030027/AHA/American Heart Association-American Stroke Association/United States
9P20GM104934-06/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
PLoS One. 2014 May 22;9(5):e98486. doi: 10.1371/journal.pone.0098486. eCollection 2014.
Author Address: Section of Endocrinology, Department of Medicine and Harold Hamm Oklahoma Diabetes Center, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, United States of America.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1164
Author: Liu, L., Botos, I., Wang, Y., Leonard, J. N., Shiloach, J., Segal, D. M. and Davies, D. R.
Year: 2008
Title: Structural basis of toll-like receptor 3 signaling with double-stranded RNA
Journal: Science
Volume: 320
Issue: 5874
Pages: 379-81
Epub Date: 2008/04/19
Date: Apr 18
Short Title: Structural basis of toll-like receptor 3 signaling with double-stranded RNA
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1155406
PMCID: PMC2761030
NIHMSID: NIHMS113699
Accession Number: 18420935
Keywords: Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray
Dimerization
Humans
Ligands
Mice
Models, Molecular
Molecular Sequence Data
Mutant Proteins/chemistry/genetics/metabolism
NF-kappa B/metabolism
Nucleic Acid Conformation
Protein Conformation
Protein Structure, Tertiary
RNA, Double-Stranded/*chemistry/*metabolism
*Signal Transduction
Toll-Like Receptor 3/*chemistry/genetics/*metabolism
Abstract: Toll-like receptor 3 (TLR3) recognizes double-stranded RNA (dsRNA), a molecular signature of most viruses, and triggers inflammatory responses that prevent viral spread. TLR3 ectodomains (ECDs) dimerize on oligonucleotides of at least 40 to 50 base pairs in length, the minimal length required for signal transduction. To establish the molecular basis for ligand binding and signaling, we determined the crystal structure of a complex between two mouse TLR3-ECDs and dsRNA at 3.4 angstrom resolution. Each TLR3-ECD binds dsRNA at two sites located at opposite ends of the TLR3 horseshoe, and an intermolecular contact between the two TLR3-ECD C-terminal domains coordinates and stabilizes the dimer. This juxtaposition could mediate downstream signaling by dimerizing the cytoplasmic Toll interleukin-1 receptor (TIR) domains. The overall shape of the TLR3-ECD does not change upon binding to dsRNA.
Notes: 1095-9203
Liu, Lin
Botos, Istvan
Wang, Yan
Leonard, Joshua N
Shiloach, Joseph
Segal, David M
Davies, David R
Z01 BC009254-33/Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, U.S. Gov't, Non-P.H.S.
Science. 2008 Apr 18;320(5874):379-81. doi: 10.1126/science.1155406.
Author Address: Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2040
Author: Liu, L., Li, L., Ma, C., Shi, Y., Liu, C., Xiao, Z., Zhang, Y., Tian, F., Gao, Y., Zhang, J., Ying, W., Wang, P. G. and Zhang, L.
Year: 2019
Title: O-GlcNAcylation of Thr(12)/Ser(56) in short-form O-GlcNAc transferase (sOGT) regulates its substrate selectivity
Journal: J Biol Chem
Volume: 294
Issue: 45
Pages: 16620-16633
Epub Date: 2019/09/19
Date: Nov 8
Short Title: O-GlcNAcylation of Thr(12)/Ser(56) in short-form O-GlcNAc transferase (sOGT) regulates its substrate selectivity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA119.009085
PMCID: PMC6851338
Accession Number: 31527085
Keywords: Amino Acid Sequence
Cell Cycle Checkpoints
Cell Nucleus/metabolism
Cell Proliferation
Chromatography, High Pressure Liquid
Glycopeptides/analysis/chemistry
Glycosylation
HEK293 Cells
HeLa Cells
Humans
Mutagenesis, Site-Directed
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/*metabolism
RNA Interference
RNA, Small Interfering/metabolism
Recombinant Proteins/biosynthesis/isolation & purification
Serine/*metabolism
Substrate Specificity
Tandem Mass Spectrometry
Threonine/*metabolism
*G2/M arrest
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
*cell cycle
*cell proliferation
*enzyme kinetics
*sOGT
*substrate selectivity
this article
Abstract: O-GlcNAcylation is a ubiquitous protein glycosylation playing different roles on variant proteins. O-GlcNAc transferase (OGT) is the unique enzyme responsible for the sugar addition to nucleocytoplasmic proteins. Recently, multiple O-GlcNAc sites have been observed on short-form OGT (sOGT) and nucleocytoplasmic OGT (ncOGT), both of which locate in the nucleus and cytoplasm in cell. Moreover, O-GlcNAcylation of Ser(389) in ncOGT (1036 amino acids) affects its nuclear translocation in HeLa cells. To date, the major O-GlcNAcylation sites and their roles in sOGT remain unknown. Here, we performed LC-MS/MS and mutational analyses to seek the major O-GlcNAcylation site on sOGT. We identified six O-GlcNAc sites in the tetratricopeptide repeat domain in sOGT, with Thr(12) and Ser(56) being two "key" sites. Thr(12) is a dominant O-GlcNAcylation site, whereas the modification of Ser(56) plays a role in regulating sOGT O-GlcNAcylation, partly through Thr(12) In vitro activity and pulldown assays demonstrated that O-GlcNAcylation does not affect sOGT activity but does affect sOGT-interacting proteins. In HEK293T cells, S56A bound to and hence glycosylated more proteins in contrast to T12A and WT sOGT. By proteomic and bioinformatics analyses, we found that T12A and S56A differed in substrate proteins (e.g. HNRNPU and PDCD6IP), which eventually affected cell cycle progression and/or cell proliferation. These findings demonstrate that O-GlcNAcylation modulates sOGT substrate selectivity and affects its role in the cell. The data also highlight the regulatory role of O-GlcNAcylation at Thr(12) and Ser(56).
Notes: 1083-351x
Liu, Li
Li, Ling
Ma, Cheng
Orcid: 0000-0002-0017-2182
Shi, Yangde
Liu, Congcong
Xiao, Zikang
Zhang, Yong
Orcid: 0000-0003-0513-8941
Tian, Fang
Gao, Yang
Zhang, Jie
Ying, Wantao
Wang, Peng George
Zhang, Lianwen
Orcid: 0000-0002-2817-3072
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2019 Nov 8;294(45):16620-16633. doi: 10.1074/jbc.RA119.009085. Epub 2019 Sep 16.
Author Address: College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300353, China.
Center for Diagnostics and Therapeutics, Department of Chemistry, Georgia State University, Atlanta, Georgia 30303.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, China.
West China-Washington Mitochondria and Metabolism Research Center, Key Laboratory of Transplant Engineering and Immunology, MOH, West China Hospital, Sichuan University, Chengdu 610041, China.
School of medicine, Nankai University, Tianjin 300071, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, China yingwantao@mail.ncpsb.org.cn.
College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300353, China lianwen@nankai.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1641
Author: Liu, M., Zhang, Y., Chen, Y., Yan, G., Shen, C., Cao, J., Zhou, X., Liu, X., Zhang, L., Shen, H., Lu, H., He, F. and Yang, P.
Year: 2014
Title: Efficient and accurate glycopeptide identification pipeline for high-throughput site-specific N-glycosylation analysis
Journal: J Proteome Res
Volume: 13
Issue: 6
Pages: 3121-9
Epub Date: 2014/04/29
Date: Jun 6
Short Title: Efficient and accurate glycopeptide identification pipeline for high-throughput site-specific N-glycosylation analysis
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr500238v
Accession Number: 24766575
Keywords: Amino Acid Motifs
Blood Proteins/chemistry/metabolism
Carbohydrate Conformation
Carbohydrate Sequence
Glycoproteins/blood/*chemistry
Glycosylation
Humans
Molecular Sequence Data
Peptide Mapping
*Protein Processing, Post-Translational
Tandem Mass Spectrometry
Abstract: Study of site-specific N-glycosylation in complex sample remains a huge analytical challenge because protein glycosylation is structurally diverse in post-translational modifications, resulting in an intricacy of N-glycopeptides. Here we have developed a novel approach for high-throughput N-glycopeptide profiling based on a network-centric algorithm for deciphering glycan fragmentation in mass spectrometry. We performed an extensive validation and a high-throughput N-glycosylation study on serum and identified thousands of N-glycopeptide spectra with high confidence. The results revealed a similar level of glycan microheterogeneity to that of conventional glycomics approach on individual proteins and provided the unique in-depth site-specific information that could only be studied through glycopeptide profiling.
Notes: 1535-3907
Liu, Mingqi
Zhang, Yang
Chen, Yaohan
Yan, Guoquan
Shen, Chengping
Cao, Jing
Zhou, Xinwen
Liu, Xiaohui
Zhang, Lei
Shen, Huali
Lu, Haojie
He, Fuchu
Yang, Pengyuan
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2014 Jun 6;13(6):3121-9. doi: 10.1021/pr500238v. Epub 2014 May 8.
Author Address: Department of Chemistry, Fudan University , 220 Han Dan Road, Shanghai 200433, P. R. China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1712
Author: Liu, N., Xiong, Y., Ren, Y., Zhang, L., He, X., Wang, X., Liu, M., Li, D., Shui, W. and Zhou, J.
Year: 2015
Title: Proteomic Profiling and Functional Characterization of Multiple Post-Translational Modifications of Tubulin
Journal: J Proteome Res
Volume: 14
Issue: 8
Pages: 3292-304
Epub Date: 2015/07/15
Date: Aug 7
Short Title: Proteomic Profiling and Functional Characterization of Multiple Post-Translational Modifications of Tubulin
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/acs.jproteome.5b00308
Accession Number: 26165356
Keywords: Acetylation
Amino Acid Sequence
Binding Sites/genetics
Chromatography, Liquid
Glycosylation
Green Fluorescent Proteins/genetics/metabolism
HeLa Cells
Humans
Immunoblotting
Mass Spectrometry
Microscopy, Fluorescence
Microtubules/genetics/*metabolism
Models, Molecular
Molecular Sequence Data
Phosphorylation
*Protein Processing, Post-Translational
Protein Structure, Tertiary
Proteomics/*methods
Sequence Homology, Amino Acid
Tubulin/chemistry/genetics/*metabolism
Ubiquitination
intracellular transport
microtubule assembly
microtubule stability
post-translational modification
tubulin
Abstract: Tubulin is known to undergo unique post-translational modifications (PTMs), such as detyrosination and polyglutamylation, particularly in the unstructured carboxy-terminal tails (CTTs). However, more conventional PTMs of tubulin and their roles in the regulation of microtubule properties and functions remain poorly defined. Here, we report the comprehensive profiling of tubulin phosphorylation, acetylation, ubiquitylation, and O-GlcNAcylation in HeLa cells with a proteomic approach. Our tubulin-targeted analysis has identified 80 residues bearing single or multiple conventional PTMs including 24 novel PTM sites not covered in previous global proteomic surveys. By using a series of PTM-deficient or PTM-mimicking mutants, we further find that tubulin phosphorylation and acetylation play important roles in the control of microtubule assembly and stability. In addition, these tubulin PTMs have distinct effects on the retrograde transport of adenoviruses along microtubules. These findings thus enlarge the repertoire of tubulin PTMs and foster our understanding of their versatile roles in the regulation of microtubule dynamics and cellular functions.
Notes: 1535-3907
Liu, Ningning
Xiong, Yun
Ren, Yiran
Zhang, Linlin
He, Xianfei
Wang, Xincheng
Liu, Min
Li, Dengwen
Shui, Wenqing
Zhou, Jun
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2015 Aug 7;14(8):3292-304. doi: 10.1021/acs.jproteome.5b00308. Epub 2015 Jul 17.
Author Address: †State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin 300071, China.
‡Key Laboratory of Systems Microbial Biotechnology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1814
Author: Liu, Q., Tao, T., Liu, F., Ni, R., Lu, C. and Shen, A.
Year: 2016
Title: Hyper-O-GlcNAcylation of YB-1 affects Ser102 phosphorylation and promotes cell proliferation in hepatocellular carcinoma
Journal: Exp Cell Res
Volume: 349
Issue: 2
Pages: 230-238
Epub Date: 2016/10/19
Date: Dec 10
Short Title: Hyper-O-GlcNAcylation of YB-1 affects Ser102 phosphorylation and promotes cell proliferation in hepatocellular carcinoma
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2016.10.011
Accession Number: 27751836
Keywords: Acetylglucosamine/metabolism
Adult
Aged
Carcinoma, Hepatocellular/*metabolism
Cell Movement/physiology
Cell Proliferation/*physiology
Female
Humans
Liver Neoplasms/*metabolism
Male
Middle Aged
Phosphorylation
Protein Processing, Post-Translational/*physiology
Serine/genetics/metabolism
Up-Regulation
Y-Box-Binding Protein 1/*metabolism
*Cell proliferation
*Hepatocellular carcinoma
*O-GlcNAc
*Ser102 phosphorylation
*Yb-1
Abstract: As an essential post-translational modification, O-GlcNAcylation has been thought to be able to modulate various nuclear and cytoplasmic proteins and is emerging as a key regulator of multiple biological processes, such as transcription, cell growth, signal transduction, and cell motility. Recently, authoritative glycomics analyses have reported extensive crosstalk between O-GlcNAcylation and phosphorylation, which always dynamically interplay with each other and regulate signaling, transcription, and other cellular processes. Also, plentiful studies have shown close correlation between YB-1 phosphorylation and tumorigenesis. Therefore, our study aimed to determine whether YB-1 was O-GlcNAc modified and whether such modification could interact with its phosphorylation during the process of HCC development. Western blot and immunohistochemistry were firstly conducted to reveal obvious up-regulation of YB-1, OGT and O-GlcNAc modification in HCC tissues. What is more, not only YB-1 was identified to be O-GlcNAcylated but hyper-O-GlcNAcylation was demonstrated to facilitate HCC cell proliferation in a YB-1 dependent manner. Moreover, we detected four specific O-GlcNAc sites and confirmed T126A to be the most effective mutant in HCC cell proliferation via close O-GlcNAcylation-phosphorylation interaction. Even more interestingly, we discovered that T126A-induced HCC cell retardation and subdued transcriptional activity of YB-1 could be partially reversed by T126A/S102E mutant. From all above, it is not difficult to find that glycosylated-YB-1 mainly enhanced cell proliferation through congenerous actions with YB-1 phosphorylation and thus played indispensable roles in fine-tuning cell proliferation and procession of HCC.
Notes: 1090-2422
Liu, Qingqing
Tao, Tao
Liu, Fang
Ni, Runzhou
Lu, Cuihua
Shen, Aiguo
Journal Article
United States
Exp Cell Res. 2016 Dec 10;349(2):230-238. doi: 10.1016/j.yexcr.2016.10.011. Epub 2016 Oct 14.
Author Address: Department of Gastroenterology, Affiliated Hospital of Nantong University, Nantong 226001, Jiangsu Province, People's Republic of China; Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qi-xiu Road, Nantong 226001, Jiangsu Province, People's Republic of China.
Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qi-xiu Road, Nantong 226001, Jiangsu Province, People's Republic of China.
Key Laboratory of Neuroregeneration, Nantong University, Nantong 226001, Jiangsu Province, People's Republic of China.
Department of Gastroenterology, Affiliated Hospital of Nantong University, Nantong 226001, Jiangsu Province, People's Republic of China.
Department of Gastroenterology, Affiliated Hospital of Nantong University, Nantong 226001, Jiangsu Province, People's Republic of China. Electronic address: lch1516@yeah.net.
Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qi-xiu Road, Nantong 226001, Jiangsu Province, People's Republic of China; Key Laboratory of Neuroregeneration, Nantong University, Nantong 226001, Jiangsu Province, People's Republic of China. Electronic address: shag@ntu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1835
Author: Liu, R., Ma, X., Chen, L., Yang, Y., Zeng, Y., Gao, J., Jiang, W., Zhang, F., Li, D., Han, B., Han, R., Qiu, R., Huang, W., Wang, Y. and Hao, J.
Year: 2017
Title: MicroRNA-15b Suppresses Th17 Differentiation and Is Associated with Pathogenesis of Multiple Sclerosis by Targeting O-GlcNAc Transferase
Journal: J Immunol
Volume: 198
Issue: 7
Pages: 2626-2639
Epub Date: 2017/02/24
Date: Apr 1
Short Title: MicroRNA-15b Suppresses Th17 Differentiation and Is Associated with Pathogenesis of Multiple Sclerosis by Targeting O-GlcNAc Transferase
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767
DOI: 10.4049/jimmunol.1601727
Accession Number: 28228555
Keywords: Animals
Blotting, Western
Cell Differentiation/genetics/immunology
Cell Separation
Down-Regulation
Encephalomyelitis, Autoimmune, Experimental/genetics/immunology
Flow Cytometry
Gene Expression Regulation/*immunology
Gene Knockdown Techniques
Humans
Immunoprecipitation
Mice
Mice, Inbred C57BL
MicroRNAs/genetics/*immunology
Multiple Sclerosis/genetics/*immunology
N-Acetylglucosaminyltransferases/*biosynthesis/immunology
Real-Time Polymerase Chain Reaction
Th17 Cells/*immunology
Abstract: IL-17-producing Th17 cells have gradually become considered as key factors in the pathogenesis of many autoimmune diseases, including multiple sclerosis (MS). Although the involvement of certain microRNAs in the development of MS has been reported, their role in Th17-driven autoimmunity is still poorly understood. In this study, we identified microRNA (miR)-15b as an important factor in Th17-associated effects and determined that the expression of miR-15b is significantly downregulated in MS patients and in mice with experimental autoimmune encephalomyelitis. Overexpression of miR-15b alleviated experimental autoimmune encephalomyelitis, whereas knockdown of miR-15b aggravated it. We demonstrated that miR-15b suppressed Th17 differentiation both in vivo and in vitro. We also found that O-linked N-acetylglucosamine transferase is a potential target of miR-15b, enabling it to affect the transcriptional regulation of retinoic acid-related orphan receptor γT through O-linked N-acetylglucosamine glycosylation of NF-κB. These results contribute to the importance of miR-15b in Th17 differentiation and the pathogenesis of MS.
Notes: 1550-6606
Liu, Ruiqiong
Ma, Xiaofeng
Chen, Li
Yang, Yang
Zeng, Yi
Gao, Jie
Jiang, Wei
Zhang, Fang
Li, Daojing
Han, Bin
Han, Ranran
Qiu, Rongfang
Huang, Wei
Wang, Yan
Hao, Junwei
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 2017 Apr 1;198(7):2626-2639. doi: 10.4049/jimmunol.1601727. Epub 2017 Feb 22.
Author Address: Department of Neurology, Tianjin Neurological Institute, Tianjin Medical University General Hospital, Tianjin 300052, China.
Tianjin Key Laboratory of Medical Epigenetics, Department of Biochemistry and Molecular Biology, Tianjin Medical University, Tianjin 300070, China.
Department of Neurology, Tianjin Huanhu Hospital, Tianjin 300350, China.
Tianjin Key Laboratory of Medical Epigenetics, Department of Biochemistry and Molecular Biology, Tianjin Medical University, Tianjin 300070, China; yanwang@tmu.edu.cn hjw@tmu.edu.cn.
Department of Neurology, Tianjin Neurological Institute, Tianjin Medical University General Hospital, Tianjin 300052, China; yanwang@tmu.edu.cn hjw@tmu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1560
Author: Liu, S., Song, X., Chrunyk, B. A., Shanker, S., Hoth, L. R., Marr, E. S. and Griffor, M. C.
Year: 2013
Title: Crystal structures of interleukin 17A and its complex with IL-17 receptor A
Journal: Nat Commun
Volume: 4
Pages: 1888
Epub Date: 2013/05/23
Short Title: Crystal structures of interleukin 17A and its complex with IL-17 receptor A
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms2880
Accession Number: 23695682
Keywords: Allosteric Regulation
Amino Acid Sequence
Conserved Sequence
Crystallization
Crystallography, X-Ray
HEK293 Cells
Humans
Interleukin-17/*chemistry/metabolism
Models, Molecular
Molecular Sequence Data
Mutagenesis
Protein Binding
Protein Multimerization
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Interleukin-17/*chemistry/metabolism
Surface Plasmon Resonance
Abstract: The constituent polypeptides of the interleukin-17 family form six different homodimeric cytokines (IL-17A-F) and the heterodimeric IL-17A/F. Their interactions with IL-17 receptors A-E (IL-17RA-E) mediate host defenses while also contributing to inflammatory and autoimmune responses. IL-17A and IL-17F both preferentially engage a receptor complex containing one molecule of IL-17RA and one molecule of IL-17RC. More generally, IL-17RA appears to be a shared receptor that pairs with other members of its family to allow signaling of different IL-17 cytokines. Here we report crystal structures of homodimeric IL-17A and its complex with IL-17RA. Binding to IL-17RA at one side of the IL-17A molecule induces a conformational change in the second, symmetry-related receptor site of IL-17A. This change favors, and is sufficient to account for, the selection of a different receptor polypeptide to complete the cytokine-receptor complex. The structural results are supported by biophysical studies with IL-17A variants produced by site-directed mutagenesis.
Notes: 2041-1723
Liu, Shenping
Song, Xi
Chrunyk, Boris A
Shanker, Suman
Hoth, Lise R
Marr, Eric S
Griffor, Matthew C
Journal Article
England
Nat Commun. 2013;4:1888. doi: 10.1038/ncomms2880.
Author Address: Structural Biology and Biophysics Group, Pfizer Groton Laboratories, Eastern Point Road, Groton, Connecticut 06340, USA. shenping.liu@pfizer.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1019
Author: Liu, T., Qian, W. J., Gritsenko, M. A., Camp, D. G., 2nd, Monroe, M. E., Moore, R. J. and Smith, R. D.
Year: 2005
Title: Human plasma N-glycoproteome analysis by immunoaffinity subtraction, hydrazide chemistry, and mass spectrometry
Journal: J Proteome Res
Volume: 4
Issue: 6
Pages: 2070-80
Epub Date: 2005/12/13
Date: Nov-Dec
Short Title: Human plasma N-glycoproteome analysis by immunoaffinity subtraction, hydrazide chemistry, and mass spectrometry
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr0502065
PMCID: PMC1850943
NIHMSID: NIHMS15603
Accession Number: 16335952
Keywords: Amino Acid Motifs
Amino Acid Sequence
Blood Proteins/*chemistry
Cations
Chromatography, Ion Exchange
Chromatography, Liquid
Glycopeptides/chemistry
Glycoproteins/*chemistry
Glycosylation
Humans
Hydrazines/*chemistry
Mass Spectrometry/*methods
Molecular Sequence Data
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/chemistry
Peptides/chemistry
Proteins/chemistry
Proteome
Proteomics/*methods
Spectroscopy, Fourier Transform Infrared
Subcellular Fractions
Abstract: The enormous complexity, wide dynamic range of relative protein abundances of interest (over 10 orders of magnitude), and tremendous heterogeneity (due to post-translational modifications, such as glycosylation) of the human blood plasma proteome severely challenge the capabilities of existing analytical methodologies. Here, we describe an approach for broad analysis of human plasma N-glycoproteins using a combination of immunoaffinity subtraction and glycoprotein capture to reduce both the protein concentration range and the overall sample complexity. Six high-abundance plasma proteins were simultaneously removed using a pre-packed, immobilized antibody column. N-linked glycoproteins were then captured from the depleted plasma using hydrazide resin and enzymatically digested, and the bound N-linked glycopeptides were released using peptide-N-glycosidase F (PNGase F). Following strong cation exchange (SCX) fractionation, the deglycosylated peptides were analyzed by reversed-phase capillary liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Using stringent criteria, a total of 2053 different N-glycopeptides were confidently identified, covering 303 nonredundant N-glycoproteins. This enrichment strategy significantly improved detection and enabled identification of a number of low-abundance proteins, exemplified by interleukin-1 receptor antagonist protein (approximately 200 pg/mL), cathepsin L (approximately 1 ng/mL), and transforming growth factor beta 1 (approximately 2 ng/mL). A total of 639 N-glycosylation sites were identified, and the overall high accuracy of these glycosylation site assignments as assessed by accurate mass measurement using high-resolution liquid chromatography coupled to Fourier transform ion cyclotron resonance mass spectrometry (LC-FTICR) is initially demonstrated.
Notes: Liu, Tao
Qian, Wei-Jun
Gritsenko, Marina A
Camp, David G 2nd
Monroe, Matthew E
Moore, Ronald J
Smith, Richard D
P41 RR018522/RR/NCRR NIH HHS/United States
U54 GM062119/GM/NIGMS NIH HHS/United States
RR18522/RR/NCRR NIH HHS/United States
U54 GM-62119-02/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Proteome Res. 2005 Nov-Dec;4(6):2070-80. doi: 10.1021/pr0502065.
Author Address: Biological Sciences Division and Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, Washington 99354, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1578
Author: Liu, X., Hammel, M., He, Y., Tainer, J. A., Jeng, U. S., Zhang, L., Wang, S. and Wang, X.
Year: 2013
Title: Structural insights into the interaction of IL-33 with its receptors
Journal: Proc Natl Acad Sci U S A
Volume: 110
Issue: 37
Pages: 14918-23
Epub Date: 2013/08/28
Date: Sep 10
Short Title: Structural insights into the interaction of IL-33 with its receptors
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1308651110
PMCID: PMC3773798
Accession Number: 23980170
Keywords: Amino Acid Sequence
Crystallography, X-Ray
Humans
Interleukin-1 Receptor Accessory Protein/*chemistry/genetics/*metabolism
Interleukin-1 Receptor-Like 1 Protein
Interleukin-33
Interleukins/*chemistry/genetics/*metabolism
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Conformation
Protein Interaction Domains and Motifs
Receptors, Cell Surface/*chemistry/genetics/*metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Scattering, Small Angle
Static Electricity
Surface Plasmon Resonance
X-Ray Diffraction
Saxs
X-ray crystallography
cytokine signaling
protein–protein interaction
Abstract: Interleukin (IL)-33 is an important member of the IL-1 family that has pleiotropic activities in innate and adaptive immune responses in host defense and disease. It signals through its ligand-binding primary receptor ST2 and IL-1 receptor accessory protein (IL-1RAcP), both of which are members of the IL-1 receptor family. To clarify the interaction of IL-33 with its receptors, we determined the crystal structure of IL-33 in complex with the ectodomain of ST2 at a resolution of 3.27 Å. Coupled with structure-based mutagenesis and binding assay, the structural results define the molecular mechanism by which ST2 specifically recognizes IL-33. Structural comparison with other ligand-receptor complexes in the IL-1 family indicates that surface-charge complementarity is critical in determining ligand-binding specificity of IL-1 primary receptors. Combined crystallography and small-angle X-ray-scattering studies reveal that ST2 possesses hinge flexibility between the D3 domain and D1D2 module, whereas IL-1RAcP exhibits a rigid conformation in the unbound state in solution. The molecular flexibility of ST2 provides structural insights into domain-level conformational change of IL-1 primary receptors upon ligand binding, and the rigidity of IL-1RAcP explains its inability to bind ligands directly. The solution architecture of IL-33-ST2-IL-1RAcP complex from small-angle X-ray-scattering analysis resembles IL-1β-IL-1RII-IL-1RAcP and IL-1β-IL-1RI-IL-1RAcP crystal structures. The collective results confer IL-33 structure-function relationships, supporting and extending a general model for ligand-receptor assembly and activation in the IL-1 family.
Notes: 1091-6490
Liu, Xi
Hammel, Michal
He, Yanfeng
Tainer, John A
Jeng, U-Ser
Zhang, Linqi
Wang, Shuying
Wang, Xinquan
R01 GM105404/GM/NIGMS NIH HHS/United States
GM105404/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2013 Sep 10;110(37):14918-23. doi: 10.1073/pnas.1308651110. Epub 2013 Aug 26.
Author Address: Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1640
Author: Liu, X., Li, L., Wang, Y., Yan, H., Ma, X., Wang, P. G. and Zhang, L.
Year: 2014
Title: A peptide panel investigation reveals the acceptor specificity of O-GlcNAc transferase
Journal: Faseb j
Volume: 28
Issue: 8
Pages: 3362-72
Epub Date: 2014/04/25
Date: Aug
Short Title: A peptide panel investigation reveals the acceptor specificity of O-GlcNAc transferase
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638
DOI: 10.1096/fj.13-246850
Accession Number: 24760753
Keywords: Amino Acids/chemistry
Animals
Cattle
Chromatography, High Pressure Liquid
Computer Simulation
Databases, Protein
Genes, Synthetic
Glycosylation
HeLa Cells
Humans
Mutagenesis, Site-Directed
N-Acetylglucosaminyltransferases/*metabolism
Peptide Fragments/chemistry
Peptide Library
Peptides/chemistry/metabolism
Protein Conformation
Protein Processing, Post-Translational
Protein Structure, Secondary
Recombinant Fusion Proteins/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Structure-Activity Relationship
Substrate Specificity
alpha-Crystallins/*chemistry/metabolism
Ogt
amino acid preference
Abstract: O-linked β-N-acetylglucosaminylation (O-GlcNAcylation) is widely distributed on nucleocytoplasmic proteins and participates in various physiological processes. But O-GlcNAc status on numerous proteins remains unknown. To better understand this modification, computational analysis combined with experimental study was performed in this work. Structural analysis of many O-GlcNAcylation sites indicated that the modification occurred predominantly in a random coil region. Frequency analysis on many O-GlcNAcylated peptides revealed a signature sequence, PPVS/TSATT, around the modification site (underlined, position 0). Based on the sequence, a peptide panel was designed to investigate key positions affecting O-GlcNAcylation of peptides and their amino acid preference. It was indicated that 3 positions (-2, -1, and +2) had an important role for this modification, where the presence of uncharged amino acids with small side chains could confer high reactivity. The amino acid preference at key positions was further investigated on bovine crystalline α via site-directed mutagenesis. The preferred amino acids were Pro > Ala > Gly at position -2, Ala > Thr > Val > Lys > Pro at position -1, and Ala > Gly > Arg > Glu at position +2. Altogether, these findings suggested that a substrate (peptide or protein) with Pro, Ala at position -2, and/or Val, Ala, Thr, Ser at position -1, and/or Ala, Ser, Pro, Thr, Gly at position +2 would have more chances for O-GlcNAcylation. To test the rule, 2 O-GlcNAcylation sites on sOGT (S52 and T449) were predicted and confirmed by Western blot. The present work systematically investigated the sequence signature for O-GlcNAcylation. The result will contribute to predicting the O-GlcNAc status of a protein and further functional studies.
Notes: 1530-6860
Liu, Xiaoyan
Li, Ling
Wang, Yuqiu
Yan, Hui
Ma, Xiaofeng
Wang, Peng George
Zhang, Lianwen
Journal Article
Research Support, Non-U.S. Gov't
United States
FASEB J. 2014 Aug;28(8):3362-72. doi: 10.1096/fj.13-246850. Epub 2014 Apr 23.
Author Address: State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy, Nankai University, Tianjin, China.
State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy, Nankai University, Tianjin, China lianwen@nankai.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1935
Author: Liu, Y., Cao, Y., Pan, X., Shi, M., Wu, Q., Huang, T., Jiang, H., Li, W. and Zhang, J.
Year: 2018
Title: O-GlcNAc elevation through activation of the hexosamine biosynthetic pathway enhances cancer cell chemoresistance
Journal: Cell Death Dis
Volume: 9
Issue: 5
Pages: 485
Epub Date: 2018/05/01
Date: May 1
Short Title: O-GlcNAc elevation through activation of the hexosamine biosynthetic pathway enhances cancer cell chemoresistance
Alternate Journal: Cell death & disease
DOI: 10.1038/s41419-018-0522-0
PMCID: PMC5924752
Accession Number: 29706631
Keywords: Antineoplastic Agents/*pharmacology
Apoptosis/drug effects
Camptothecin/*pharmacology
Doxorubicin/*pharmacology
*Drug Resistance, Neoplasm
*Energy Metabolism
Glycosylation
HL-60 Cells
HeLa Cells
Hexosamines/*biosynthesis
Humans
MCF-7 Cells
Neoplasms/*drug therapy/genetics/*metabolism/pathology
*Protein Processing, Post-Translational
Proto-Oncogene Proteins c-akt/metabolism
Signal Transduction
Tumor Cells, Cultured
X-Box Binding Protein 1/genetics/metabolism
Abstract: Chemoresistance has become a major obstacle to the success of cancer therapy, but the mechanisms underlying chemoresistance are not yet fully understood. O-GlcNAcylation is a post-translational modification that is regulated by the hexosamine biosynthetic pathway (HBP) and has an important role in a wide range of cellular functions. Here we assessed the role of O-GlcNAcylation in chemoresistance and investigated the underlying cellular mechanisms. The results showed that the HBP has an important role in cancer cell chemoresistance by regulating O-GlcNAcylation. An increase in the levels of O-GlcNAcylation indicates an increased resistance of cancer cells to chemotherapy. Acute treatment with doxorubicin (DOX) or camptothecin (CPT) induced O-GlcNAcylation through HBP activation. In fact, the chemotherapy agents activated the AKT/X-box-binding protein 1 (XBP1) axis and then induced the HBP. Furthermore, the observed elevation of cellular O-GlcNAcylation led to activation of survival signalling pathways and chemoresistance in cancer cells. Finally, suppression of O-GlcNAcylation reduced the resistance of both established and primary cancer cells to chemotherapy. These results provide significant novel insights regarding the important role of the HBP and O-GlcNAcylation in regulating cancer chemoresistance. Thus, O-GlcNAc inhibition might offer a new strategy for improving the efficacy of chemotherapy.
Notes: 2041-4889
Liu, Yubo
Cao, Yu
Pan, Xiaoqing
Shi, Meiyun
Wu, Qiong
Huang, Tianmiao
Jiang, Hui
Li, Wenli
Zhang, Jianing
Journal Article
Research Support, Non-U.S. Gov't
Cell Death Dis. 2018 May 1;9(5):485. doi: 10.1038/s41419-018-0522-0.
Author Address: School of Life Science and Medicine, Dalian University of Technology, Panjin, China.
School of Life Science and Biotechnology, Dalian University of Technology, Dalian, China.
School of Life Science and Medicine, Dalian University of Technology, Panjin, China. jnzhang@dlut.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1850
Author: Liu, Y., Huang, H., Cao, Y., Wu, Q., Li, W. and Zhang, J.
Year: 2017
Title: Suppression of OGT by microRNA24 reduces FOXA1 stability and prevents breast cancer cells invasion
Journal: Biochem Biophys Res Commun
Volume: 487
Issue: 3
Pages: 755-762
Epub Date: 2017/04/30
Date: Jun 3
Short Title: Suppression of OGT by microRNA24 reduces FOXA1 stability and prevents breast cancer cells invasion
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2017.04.135
Accession Number: 28455227
Keywords: Breast Neoplasms/*genetics/*metabolism
Cell Line, Tumor
Cell Movement
Gene Expression Regulation, Neoplastic
Gene Silencing
Hepatocyte Nuclear Factor 3-alpha/*metabolism
Humans
MicroRNAs/*metabolism
N-Acetylglucosaminyltransferases/*metabolism
Neoplasm Invasiveness
Breast cancer
Foxa1
Invasion
Ogt
miRNA24
Abstract: O-GlcNAc transferase (OGT) catalyzes the addition of O-GlcNAc to certain serine or threonine residue on a wide variety of cytosolic and nuclear proteins and regulates cellular activities such as signaling and transcription. Although there are emerging evidences that OGT plays important roles in breast cancer metastasis, the underlying mechanism is not fully understood. In this study, we demonstrated that up-regulation of OGT correlates with breast cancer cells invasion. Over-expression of OGT stimulates cells invasion, while OGT silence exhibits the opposite effects. OGT is further identified as a target of microRNA24 (miR24). miR24 down-regulates OGT expression and subsequently suppresses cells invasion. Re-expression of OGT significantly rescues miR24-mediated invasion repression. Furthermore, our data showed that FOXA1 is subjected to O-GlcNAcylation, which instabilizes FOXA1 protein and promotes breast cancer cells invasion. In conclusion, our results demonstrated that miR24 inhibits breast cancer cells invasion by targeting OGT and reducing FOXA1 stability. These results also indicated that OGT might be a potential target for the diagnosis and therapy of breast cancer metastasis.
Notes: 1090-2104
Liu, Yubo
Huang, Huang
Cao, Yu
Wu, Qiong
Li, Wenli
Zhang, Jianing
Journal Article
United States
Biochem Biophys Res Commun. 2017 Jun 3;487(3):755-762. doi: 10.1016/j.bbrc.2017.04.135. Epub 2017 Apr 26.
Author Address: School of Life Science and Medicine, Dalian University of Technology, Panjin, China.
School of Life Science and Medicine, Dalian University of Technology, Panjin, China. Electronic address: jnzhang@dlut.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1897
Author: Liu, Y., Lu, Z., Shi, Y. and Sun, F.
Year: 2018
Title: AMOT is required for YAP function in high glucose induced liver malignancy
Journal: Biochem Biophys Res Commun
Volume: 495
Issue: 1
Pages: 1555-1561
Epub Date: 2017/12/09
Date: Jan 1
Short Title: AMOT is required for YAP function in high glucose induced liver malignancy
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2017.12.010
Accession Number: 29217192
Keywords: Adaptor Proteins, Signal Transducing/*metabolism
Animals
Blood Glucose/*metabolism
Cell Cycle Proteins
Cell Nucleus/metabolism
Diabetes Complications/*metabolism/pathology
Intercellular Signaling Peptides and Proteins/*metabolism
Liver Neoplasms/*etiology/*metabolism/pathology
Mice
Microfilament Proteins/*metabolism
Phosphoproteins/*metabolism
*Hippo signaling
*Liver tumorigenesis
*O-GlcNAcylation
*Subcellular localization
Abstract: AMOT has been identified as a YAP interactor. However, how AMOT regulates YAP remains unclear and controversy. Here, we identified that besides YAP, AMOT was another Hippo signaling core factor which could be O-GlcNAcylated. Moreover, high glucose (HG) was able to enhance the expression and O-GlcNAcylation of AMOT. We also found that HG stimulated nuclear accumulation, transcription activity, interaction with transcription factor and transcription of target genes of YAP via AMOT, while AMOT acted as a suppressor of YAP in normal glucose level. Finally, we observed the upregulation and nuclear accumulation of AMOT and YAP in Streptozocin (STZ) induced high glucose mice. Collectively, we have uncovered that AMOT acts as a YAP stimulator in high glucose level. Targeting the aberrantly regulated core factors in Hippo pathway might be a more effective therapeutic approach for liver cancer associated with possibly diabetes.
Notes: 1090-2104
Liu, Ya
Lu, Zhicheng
Shi, Yi
Sun, Fenyong
Journal Article
United States
Biochem Biophys Res Commun. 2018 Jan 1;495(1):1555-1561. doi: 10.1016/j.bbrc.2017.12.010. Epub 2017 Dec 5.
Author Address: Department of Clinical Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai, 200072, China.
Department of Laboratory Medicine, Seventh People's Hospital of Shanghai University of TCM, Shanghai, 200137, China.
Department of Clinical Laboratory, Shanghai Fourth People's Hospital, Shanghai, 200081, China. Electronic address: yishishanghai@126.com.
Department of Clinical Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai, 200072, China. Electronic address: sunfenyong@126.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2010
Author: Liu, Y., Wang, X., Zhu, T., Zhang, N., Wang, L., Huang, T., Cao, Y., Li, W. and Zhang, J.
Year: 2019
Title: Resistance to bortezomib in breast cancer cells that downregulate Bim through FOXA1 O-GlcNAcylation
Journal: J Cell Physiol
Volume: 234
Issue: 10
Pages: 17527-17537
Epub Date: 2019/02/23
Date: Aug
Short Title: Resistance to bortezomib in breast cancer cells that downregulate Bim through FOXA1 O-GlcNAcylation
Alternate Journal: Journal of cellular physiology
ISSN: 0021-9541
DOI: 10.1002/jcp.28376
Accession Number: 30793308
Keywords: Antineoplastic Agents/*pharmacology
Apoptosis/drug effects
Bortezomib/*pharmacology
Breast Neoplasms/*drug therapy/*metabolism/pathology
Cell Line, Tumor
DNA-Binding Proteins/deficiency/*metabolism
Down-Regulation
Drug Resistance, Neoplasm
Female
Glycosylation/drug effects
Hepatocyte Nuclear Factor 3-alpha/chemistry/*metabolism
Humans
MCF-7 Cells
Proteasome Inhibitors/pharmacology
Protein Stability/drug effects
Transcription Factors/deficiency/*metabolism
*Foxa1
*O-GlcNAc
*apoptosis
*bortezomib
*breast cancer
Abstract: Bortezomib (BTZ), a well-established proteasome inhibitor used in the clinical therapy, leads the modulation of several biological alterations and in turn induces apoptosis. Although clinical trials with BTZ have shown promising results for some types of cancers, but not for some others, including those of the breast. The molecular basis of BTZ resistance in breast cancer remains elusive. In the present study, we found that cellular O-GlcNAc modification was dramatically elevated by BTZ treatment in intrinsic resistant MCF-7 and T47D cells, but not in sensitive MDA-MB-231 cells. A progressive increase in O-GlcNAcylation characterized the increased acquired resistance of MDA-MB-231-derived cells. We showed that elevated O-GlcNAc subsequently modified breast cancer related pioneer factor FOXA1 and reduced its protein stability. Further, we demonstrated that FOXA1 attenuation was involved in transcriptional downregulation of proapoptotic Bim and thus suppressed breast cancer cell apoptosis. Finally, the combination of O-GlcNAc inhibitor L01 to BTZ sensitized resistant cells. Our results have revealed a new regulatory mechanism that involves O-GlcNAc elevation mediated Bim deficiency, which plays a key role in the apoptotic dysregulation and BTZ resistance in breast cancer cells.
Notes: 1097-4652
Liu, Yubo
Wang, Xue
Zhu, Tong
Zhang, Nana
Wang, Lingyan
Huang, Tianmiao
Cao, Yu
Li, Wenli
Zhang, Jianing
Orcid: 0000-0002-4514-9197
Journal Article
Research Support, Non-U.S. Gov't
United States
J Cell Physiol. 2019 Aug;234(10):17527-17537. doi: 10.1002/jcp.28376. Epub 2019 Feb 21.
Author Address: Department of Biochemistry, School of Life Science & Medicine, Dalian University of Technology, Panjin, China.
Department of Biochemistry, School of Life Science & Biotechnology, Dalian University of Technology, Dalian, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 984
Author: Llinas, P., Le Du, M. H., Gårdsvoll, H., Danø, K., Ploug, M., Gilquin, B., Stura, E. A. and Ménez, A.
Year: 2005
Title: Crystal structure of the human urokinase plasminogen activator receptor bound to an antagonist peptide
Journal: Embo j
Volume: 24
Issue: 9
Pages: 1655-63
Epub Date: 2005/04/30
Date: May 4
Short Title: Crystal structure of the human urokinase plasminogen activator receptor bound to an antagonist peptide
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7600635
PMCID: PMC1142576
Accession Number: 15861141
Keywords: Crystallization/methods
Humans
Molecular Conformation
Peptides/*chemistry/metabolism
Protein Binding
Receptors, Cell Surface/antagonists & inhibitors/*chemistry/metabolism
Receptors, Urokinase Plasminogen Activator
Recombinant Proteins/chemistry
Urokinase-Type Plasminogen Activator/chemistry/metabolism
Abstract: We report the crystal structure of a soluble form of human urokinase-type plasminogen activator receptor (uPAR/CD87), which is expressed at the invasive areas of the tumor-stromal microenvironment in many human cancers. The structure was solved at 2.7 A in association with a competitive peptide inhibitor of the urokinase-type plasminogen activator (uPA)-uPAR interaction. uPAR is composed of three consecutive three-finger domains organized in an almost circular manner, which generates both a deep internal cavity where the peptide binds in a helical conformation, and a large external surface. This knowledge combined with the discovery of a convergent binding motif shared by the antagonist peptide and uPA allowed us to build a model of the human uPA-uPAR complex. This model reveals that the receptor-binding module of uPA engages the uPAR central cavity, thus leaving the external receptor surface accessible for other protein interactions (vitronectin and integrins). By this unique structural assembly, uPAR can orchestrate the fine interplay with the partners that are required to guide uPA-focalized proteolysis on the cell surface and control cell adhesion and migration.
Notes: 1460-2075
Llinas, Paola
Le Du, Marie Hélène
Gårdsvoll, Henrik
Danø, Keld
Ploug, Michael
Gilquin, Bernard
Stura, Enrico A
Ménez, André
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2005 May 4;24(9):1655-63. doi: 10.1038/sj.emboj.7600635. Epub 2005 Apr 7.
Author Address: CEA, Département d'Ingénierie et d'Etudes des Protéines, CE Saclay, Gif sur Yvette, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1057
Author: Llinas, P., Masella, M., Stigbrand, T., Ménez, A., Stura, E. A. and Le Du, M. H.
Year: 2006
Title: Structural studies of human alkaline phosphatase in complex with strontium: implication for its secondary effect in bones
Journal: Protein Sci
Volume: 15
Issue: 7
Pages: 1691-700
Epub Date: 2006/07/04
Date: Jul
Short Title: Structural studies of human alkaline phosphatase in complex with strontium: implication for its secondary effect in bones
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.062123806
PMCID: PMC2242561
Accession Number: 16815919
Keywords: Alkaline Phosphatase/*chemistry
*Calcification, Physiologic
Calcium
GPI-Linked Proteins
Humans
Isoenzymes
Osteomalacia/*etiology
Protein Conformation
Strontium/*chemistry
Abstract: Strontium is used in the treatment of osteoporosis as a ranelate compound, and in the treatment of painful scattered bone metastases as isotope. At very high doses and in certain conditions, it can lead to osteomalacia characterized by impairment of bone mineralization. The osteomalacia symptoms resemble those of hypophosphatasia, a rare inherited disorder associated with mutations in the gene encoding for tissue-nonspecific alkaline phosphatase (TNAP). Human alkaline phosphatases have four metal binding sites--two for zinc, one for magnesium, and one for calcium ion--that can be substituted by strontium. Here we present the crystal structure of strontium-substituted human placental alkaline phosphatase (PLAP), a related isozyme of TNAP, in which such replacement can have important physiological implications. The structure shows that strontium substitutes the calcium ion with concomitant modification of the metal coordination. The use of the flexible and polarizable force-field TCPEp (topological and classical polarization effects for proteins) predicts that calcium or strontium has similar interaction energies at the calcium-binding site of PLAP. Since calcium helps stabilize a large area that includes loops 210-228 and 250-297, its substitution by strontium could affect the stability of this region. Energy calculations suggest that only at high doses of strontium, comparable to those found for calcium, can strontium substitute for calcium. Since osteomalacia is observed after ingestion of high doses of strontium, alkaline phosphatase is likely to be one of the targets of strontium, and thus this enzyme might be involved in this disease.
Notes: 1469-896x
Llinas, Paola
Masella, Michel
Stigbrand, Torgny
Ménez, André
Stura, Enrico A
Le Du, Marie Hélène
Journal Article
Protein Sci. 2006 Jul;15(7):1691-700. doi: 10.1110/ps.062123806.
Author Address: Laboratoire de Structure des Protéines, Département d'Ingénierie et d'Etude des Protéines, Commissariat à l'Energie Atomique, Gif sur Yvette, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 990
Author: Llinas, P., Stura, E. A., Ménez, A., Kiss, Z., Stigbrand, T., Millán, J. L. and Le Du, M. H.
Year: 2005
Title: Structural studies of human placental alkaline phosphatase in complex with functional ligands
Journal: J Mol Biol
Volume: 350
Issue: 3
Pages: 441-51
Epub Date: 2005/06/11
Date: Jul 15
Short Title: Structural studies of human placental alkaline phosphatase in complex with functional ligands
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2005.04.068
Accession Number: 15946677
Keywords: Adenosine Monophosphate/chemistry
Alkaline Phosphatase/*chemistry
Binding Sites
Catalytic Domain
Cell Proliferation
Crystallography, X-Ray
Down-Regulation
Fibroblasts/metabolism
Humans
Hydrolysis
Ligands
Models, Molecular
Molecular Conformation
Nitrophenols
Organophosphonates/pharmacology
Phenylalanine/chemistry
Placenta/*enzymology
Protein Conformation
Protein Isoforms
Protein Structure, Tertiary
Abstract: The activity of human placental alkaline phosphatase (PLAP) is downregulated by a number of effectors such as l-phenylalanine, an uncompetitive inhibitor, 5'-AMP, an antagonist of the effects of PLAP on fibroblast proliferation and by p-nitrophenyl-phosphonate (PNPPate), a non-hydrolysable substrate analogue. For the first two, such regulation may be linked to its biological function that requires a reduced and better-regulated hydrolytic rate. To understand how such disparate ligands are able to inhibit the enzyme, we solved the structure of the complexes at 1.6A, 1.9A and 1.9A resolution, respectively. These crystal structures are the first of an alkaline phosphatase in complex with organic inhibitors. Of the three inhibitors, only l-Phe and PNPPate bind at the active site hydrophobic pocket, providing structural data on the uncompetitive inhibition process. In contrast, all three ligands interact at a remote peripheral site located 28A from the active site. In order to extend these observations to the other members of the human alkaline phosphatase family, we have modelled the structures of the other human isozymes and compared them to PLAP. This comparison highlights the crucial role played by position 429 at the active site in the modulation of the catalytic process, and suggests that the peripheral binding site may be involved in the functional specialization of the PLAP isozyme.
Notes: Llinas, Paola
Stura, Enrico A
Ménez, André
Kiss, Zoltan
Stigbrand, Torgny
Millán, José Luis
Le Du, Marie Hélène
Journal Article
England
J Mol Biol. 2005 Jul 15;350(3):441-51. doi: 10.1016/j.jmb.2005.04.068.
Author Address: Laboratoire de Structure des Protéines, Département d'Ingénierie et d'Etude des Protéines, Commissariat à l'Energie Atomique, 91191 Gif sur Yvette, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 630
Author: Lloyd, K. O., Burchell, J., Kudryashov, V., Yin, B. W. and Taylor-Papadimitriou, J.
Year: 1996
Title: Comparison of O-linked carbohydrate chains in MUC-1 mucin from normal breast epithelial cell lines and breast carcinoma cell lines. Demonstration of simpler and fewer glycan chains in tumor cells
Journal: J Biol Chem
Volume: 271
Issue: 52
Pages: 33325-34
Epub Date: 1996/12/27
Date: Dec 27
Short Title: Comparison of O-linked carbohydrate chains in MUC-1 mucin from normal breast epithelial cell lines and breast carcinoma cell lines. Demonstration of simpler and fewer glycan chains in tumor cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.52.33325
Accession Number: 8969192
Keywords: Breast/*chemistry
Breast Neoplasms/*chemistry
Chromatography, Gel
Epithelial Cells
Female
Glycosylation
Humans
Models, Chemical
Mucin-1/*chemistry
Periodic Acid/metabolism
Polysaccharides/*chemistry
Tumor Cells, Cultured
Abstract: MUC-1 mucin is considered to be aberrantly glycosylated in breast, ovary, and other carcinomas in comparison with mucin from corresponding normal tissues. In order to clarify these differences in glycosylation, we have compared the O-linked carbohydrate chains from MUC-1 immunoprecipitated from [3H]GlcN-labeled breast epithelial cell lines (MMSV1-1, MTSV1-7, and HB-2) derived from cells cultured from human milk, with three breast cancer cell lines (MCF-7, BT-20, and T47D). Analysis by high pH anion chromatography showed that the normal cell lines had a higher ratio of GlcN/GalN and more complex oligosaccharide profiles than the cancer cell lines. Structural analyses were carried out on the oligosaccharides from MTSV1-7 and T47D MUC-1, and the following structures were proposed. MUC-1 from T47D had rather a simple glycosylation pattern, with NeuAcalpha2-3Galbeta1-3GalNAc-ol, Galbeta1-3GalNAc-ol, and GalNAc-ol predominating; in contrast, MUC-1 from MTSV1-7 had more complex structures, including a number of disialo, core 2 species, i.e. NeuAcalpha2-3Galbeta1-4GlcNAcbeta1-6[NeuAcalpha2 -3Galbeta1-3]GalNAc- ol and NeuAcalpha2-3Galbeta1-4GlcNAcbeta1-6[NeuAcalpha2 -3Galbeta1-4GlcNAcbet a1-3Galbeta1-3]GalNAc-ol. Double-labeling experiments with [3H]GlcN and 14C-aminoacids and analysis of GalNAc or GalNAc-ol:protein ratios in MUC-1 showed that there was also a significant difference in the degree of glycosylation of the mucin between the two cell types. We conclude that MUC-1 from breast cancer cell lines has simpler, and fewer, carbohydrate chains than MUC-1 from normal breast epithelial cells, and that these differences, combined or separately, explain the differential tumor specificity of some MUC-1 antibodies and T cells.
Notes: Lloyd, K O
Burchell, J
Kudryashov, V
Yin, B W
Taylor-Papadimitriou, J
CA-08748/CA/NCI NIH HHS/United States
CA-52477/CA/NCI NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1996 Dec 27;271(52):33325-34. doi: 10.1074/jbc.271.52.33325.
Author Address: Immunology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 644
Author: Lloyd, K. O., Yin, B. W. and Kudryashov, V.
Year: 1997
Title: Isolation and characterization of ovarian cancer antigen CA 125 using a new monoclonal antibody (VK-8): identification as a mucin-type molecule
Journal: Int J Cancer
Volume: 71
Issue: 5
Pages: 842-50
Epub Date: 1997/05/29
Date: May 29
Short Title: Isolation and characterization of ovarian cancer antigen CA 125 using a new monoclonal antibody (VK-8): identification as a mucin-type molecule
Alternate Journal: International journal of cancer
ISSN: 0020-7136 (Print)
0020-7136
DOI: 10.1002/(sici)1097-0215(19970529)71:5<842::aid-ijc24>3.0.co;2-8
Accession Number: 9180155
Keywords: Acetylgalactosamine/analysis
Acetylglucosamine/analysis
Animals
*Antibodies, Monoclonal/immunology
Antibody Specificity
CA-125 Antigen/*chemistry/immunology/*isolation & purification
Carbohydrate Conformation
Electrophoresis, Polyacrylamide Gel
Enzyme-Linked Immunosorbent Assay
Female
Fucose/metabolism
Galactose/analysis
Glycosylation
Humans
Immunosorbent Techniques
Iodine Radioisotopes
Lewis Blood Group Antigens
Mice
Mucins/*chemistry/immunology
Ovarian Neoplasms/*immunology
Abstract: A new murine monoclonal antibody (MAb VK-8), detecting the CA 125 ovarian cancer antigen, was used to purify this antigen from OVCAR-3 ovarian cancer cells by affinity chromatography. The biochemical properties of the purified antigen are characteristic of a mucin-type glycoprotein: (1) the molecule is highly glycosylated (77% w/w), mainly with galactose, N-acetylglucosamine, and N-acetylgalactosamine, (2) the protein moiety is rich in serine, threonine and proline, (3) many of the serine and threonine residues are glycosylated, (4) the glycan chains are almost entirely O-linked, with core 2 [Galbeta1 --> 3(GlcNAcbeta1 --> 6)GalNAc] structures predominating and (5) these chains carry fucosylated Type 2 (Le(y) and Le(x) and H type 2) blood group structures. The antigen exhibited a very high m.w. (> 10(3) kDa) in aqueous buffer as well as in urea, but was degraded by proteolytic enzymes to smaller fragments that no longer reacted with the antibody. Although this result, and other immunochemical data, indicate that OC125, the original MAb to CA125, and VK-8 antibodies detect epitopes on the protein portion of the molecule, the involvement of carbohydrate cannot be ruled out. Further insight into the structure and function of the CA125 antigen will come from cloning the gene coding for the peptide backbone, and from more detailed carbohydrate structural analysis.
Notes: Lloyd, K O
Yin, B W
Kudryashov, V
CA 08748/CA/NCI NIH HHS/United States
CA 52477/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Int J Cancer. 1997 May 29;71(5):842-50. doi: 10.1002/(sici)1097-0215(19970529)71:5<842::aid-ijc24>3.0.co;2-8.
Author Address: Immunology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA. klloyd@ski.mskcc.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1949
Author: Lo, P. W., Shie, J. J., Chen, C. H., Wu, C. Y., Hsu, T. L. and Wong, C. H.
Year: 2018
Title: O-GlcNAcylation regulates the stability and enzymatic activity of the histone methyltransferase EZH2
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 28
Pages: 7302-7307
Epub Date: 2018/06/27
Date: Jul 10
Short Title: O-GlcNAcylation regulates the stability and enzymatic activity of the histone methyltransferase EZH2
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1801850115
PMCID: PMC6048490
Accession Number: 29941599
Keywords: Acetylglucosamine/chemistry/genetics/*metabolism
Amino Acid Substitution
Cell Line
Enhancer of Zeste Homolog 2 Protein/chemistry/genetics/*metabolism
Enzyme Stability
Glycosylation
Humans
*Mutation, Missense
Protein Domains
*H3K27me3
*O-GlcNAcylation
*cancer
*methyltransferase EZH2
Abstract: Protein O-glycosylation by attachment of β-N-acetylglucosamine (GlcNAc) to the Ser or Thr residue is a major posttranslational glycosylation event and is often associated with protein folding, stability, and activity. The methylation of histone H3 at Lys-27 catalyzed by the methyltransferase EZH2 was known to suppress gene expression and cancer development, and we previously reported that the O-GlcNAcylation of EZH2 at S76 stabilized EZH2 and facilitated the formation of H3K27me3 to inhibit tumor suppression. In this study, we employed a fluorescence-based method of sugar labeling combined with mass spectrometry to investigate EZH2 glycosylation and identified five O-GlcNAcylation sites. We also find that mutation of one or more of the O-GlcNAcylation sites S73A, S76A, S84A, and T313A in the N-terminal region decreases the stability of EZH2, but does not affect its association with the PRC2 components SUZ12 and EED. Mutation of the C-terminal O-GlcNAcylation site (S729A) in the catalytic domain of EZH2 abolishes the di- and trimethylation activities, but not the monomethylation of H3K27, nor the integrity of the PRC2/EZH2 core complex. Our results show the effect of individual O-GlcNAcylation sites on the function of EZH2 and suggest an alternative approach to tumor suppression through selective inhibition of EZH2 O-GlcNAcylation.
Notes: 1091-6490
Lo, Pei-Wen
Shie, Jiun-Jie
Chen, Chein-Hung
Wu, Chung-Yi
Hsu, Tsui-Ling
Wong, Chi-Huey
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2018 Jul 10;115(28):7302-7307. doi: 10.1073/pnas.1801850115. Epub 2018 Jun 25.
Author Address: Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Institute of Chemistry, Academia Sinica, Taipei 115, Taiwan.
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan; chwong@gate.sinica.edu.tw.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 408
Author: Lockridge, O., Bartels, C. F., Vaughan, T. A., Wong, C. K., Norton, S. E. and Johnson, L. L.
Year: 1987
Title: Complete amino acid sequence of human serum cholinesterase
Journal: J Biol Chem
Volume: 262
Issue: 2
Pages: 549-57
Epub Date: 1987/01/15
Date: Jan 15
Short Title: Complete amino acid sequence of human serum cholinesterase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3542989
Keywords: Amino Acid Sequence
Binding Sites
Cholinesterases/*blood
Cyanogen Bromide
Humans
Macromolecular Substances
Peptide Fragments/analysis
Peptide Hydrolases
Phenylthiohydantoin
Abstract: The complete amino acid sequence of human serum cholinesterase (choline esterase II (unspecific), EC 3.1.1.8) was determined by Edman degradation of purified peptides. The protein contains 574 amino acids per subunit and nine carbohydrate chains attached to 9 asparagines. The four subunits of cholinesterase appear to be identical. The active site serine is the 198th residue from the amino terminus. The sequence of human serum cholinesterase is 53.8% identical with the sequence of acetylcholinesterase from Torpedo californica and 28% identical with the carboxyl-terminal portion of bovine thyroglobulin.
Notes: Lockridge, O
Bartels, C F
Vaughan, T A
Wong, C K
Norton, S E
Johnson, L L
GM 27028/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1987 Jan 15;262(2):549-57.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 618
Author: Löffler, A., Doucey, M. A., Jansson, A. M., Müller, D. R., de Beer, T., Hess, D., Meldal, M., Richter, W. J., Vliegenthart, J. F. and Hofsteenge, J.
Year: 1996
Title: Spectroscopic and protein chemical analyses demonstrate the presence of C-mannosylated tryptophan in intact human RNase 2 and its isoforms
Journal: Biochemistry
Volume: 35
Issue: 37
Pages: 12005-14
Epub Date: 1996/09/17
Date: Sep 17
Short Title: Spectroscopic and protein chemical analyses demonstrate the presence of C-mannosylated tryptophan in intact human RNase 2 and its isoforms
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi9610515
Accession Number: 8810905
Keywords: Amino Acid Sequence
Blotting, Western
Endoribonucleases/*chemistry/isolation & purification/urine
Female
Glycosylation
Humans
Isoenzymes/*chemistry/isolation & purification/urine
Magnetic Resonance Spectroscopy
Mannose/*analysis
Mass Spectrometry
Menopause
Models, Molecular
Molecular Sequence Data
Molecular Weight
Peptide Fragments/chemistry/isolation & purification
Pregnancy
Protein Conformation
*Tryptophan/*analogs & derivatives/analysis
Abstract: Recently, the C-mannosylation of a specific tryptophan residue in RNase 2 from human urine has been reported [Hofsteenge, J., et al. (1994) Biochemistry 33, 13524-13530; de Beer, T., et al. (1995) Biochemistry 34, 11785-11789]. In those studies, identification of this unusual modification was accomplished by mass spectrometric and NMR spectroscopic analysis of peptide fragments. The evidence for the occurrence of C2-alpha-mannosyltryptophan [(C2-Man-)Trp] in the intact protein relied exclusively on the detection of the same phenylthiohydantoin derivatives during Edman degradation. In this paper, we have (1) excluded the possibility that (C2-Man-)Trp arose artificially under the acidic conditions previously employed for protein and peptide isolation and analysis, by maintaining the pH > 5 throughout these procedures, (2) demonstrated the occurrence of (C2-Man-)Trp in the intact protein, by NMR spectroscopy, (3) showed that (C2-Man-)Trp is not unique for RNase 2 from urine but that it is also present in the enzyme isolated from erythrocytes, and (4) found also that high-molecular mass isoforms of urinary RNase 2 are C-mannosylated. These observations firmly establish C-mannosylation as a novel way of post-translationally attaching carbohydrate to protein, in addition to the well-known N- and O-glycosylations. Furthermore, the NMR data, in combination with molecular dynamics calculations, indicate that in the native protein the mannopyranosyl residue is in a different conformation than in the glycopeptide or denatured protein, due to protein-carbohydrate interactions.
Notes: Löffler, A
Doucey, M A
Jansson, A M
Müller, D R
de Beer, T
Hess, D
Meldal, M
Richter, W J
Vliegenthart, J F
Hofsteenge, J
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1996 Sep 17;35(37):12005-14. doi: 10.1021/bi9610515.
Author Address: Friedrich Miescher-Institut, Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1054
Author: Longenecker, K. L., Stewart, K. D., Madar, D. J., Jakob, C. G., Fry, E. H., Wilk, S., Lin, C. W., Ballaron, S. J., Stashko, M. A., Lubben, T. H., Yong, H., Pireh, D., Pei, Z., Basha, F., Wiedeman, P. E., von Geldern, T. W., Trevillyan, J. M. and Stoll, V. S.
Year: 2006
Title: Crystal structures of DPP-IV (CD26) from rat kidney exhibit flexible accommodation of peptidase-selective inhibitors
Journal: Biochemistry
Volume: 45
Issue: 24
Pages: 7474-82
Epub Date: 2006/06/14
Date: Jun 20
Short Title: Crystal structures of DPP-IV (CD26) from rat kidney exhibit flexible accommodation of peptidase-selective inhibitors
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi060184f
Accession Number: 16768443
Keywords: Animals
Binding Sites
Crystallization
Dimerization
Dipeptidases/*antagonists & inhibitors/chemistry/metabolism
Dipeptidyl Peptidase 4/*chemistry/genetics/isolation & purification/*metabolism
Glucagon-Like Peptide 1/chemistry/metabolism
Humans
Kidney/*enzymology
Kinetics
Models, Molecular
Molecular Structure
Protein Conformation
Rats
Recombinant Proteins/chemistry/metabolism
Structure-Activity Relationship
Tyrosine/chemistry
X-Ray Diffraction
Abstract: Dipeptidyl peptidase IV (DPP-IV) belongs to a family of serine peptidases, and due to its indirect regulatory role in plasma glucose modulation, DPP-IV has become an attractive pharmaceutical target for diabetes therapy. DPP-IV inactivates the glucagon-like peptide (GLP-1) and several other naturally produced bioactive peptides that contain preferentially a proline or alanine residue in the second amino acid sequence position by cleaving the N-terminal dipeptide. To elucidate the details of the active site for structure-based drug design, we crystallized a natural source preparation of DPP-IV isolated from rat kidney and determined its three-dimensional structure using X-ray diffraction techniques. With a high degree of similarity to structures of human DPP-IV, the active site architecture provides important details for the design of inhibitory compounds, and structures of inhibitor-protein complexes offer detailed insight into three-dimensional structure-activity relationships that include a conformational change of Tyr548. Such accommodation is exemplified by the response to chemical substitution on 2-cyanopyrrolidine inhibitors at the 5 position, which conveys inhibitory selectivity for DPP-IV over closely related homologues. A similar conformational change is also observed in the complex with an unrelated synthetic inhibitor containing a xanthine core that is also selective for DPP-IV. These results suggest the conformational flexibility of Tyr548 is unique among protein family members and may be utilized in drug design to achieve peptidase selectivity.
Notes: Longenecker, Kenton L
Stewart, Kent D
Madar, David J
Jakob, Clarissa G
Fry, Elizabeth H
Wilk, Sherwin
Lin, Chun W
Ballaron, Stephen J
Stashko, Michael A
Lubben, Thomas H
Yong, Hong
Pireh, Daisy
Pei, Zhonghua
Basha, Fatima
Wiedeman, Paul E
von Geldern, Thomas W
Trevillyan, James M
Stoll, Vincent S
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2006 Jun 20;45(24):7474-82. doi: 10.1021/bi060184f.
Author Address: Department of Structural Biology, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, Illinois 60064-6098, USA. Kenton.Longenecker@Abbott.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1635
Author: Loos, A., Gruber, C., Altmann, F., Mehofer, U., Hensel, F., Grandits, M., Oostenbrink, C., Stadlmayr, G., Furtmüller, P. G. and Steinkellner, H.
Year: 2014
Title: Expression and glycoengineering of functionally active heteromultimeric IgM in plants
Journal: Proc Natl Acad Sci U S A
Volume: 111
Issue: 17
Pages: 6263-8
Epub Date: 2014/04/08
Date: Apr 29
Short Title: Expression and glycoengineering of functionally active heteromultimeric IgM in plants
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1320544111
PMCID: PMC4035941
Accession Number: 24706782
Keywords: Chromatography, Affinity
Glycomics/*methods
Glycosylation
Humans
Immunoglobulin M/chemistry/*metabolism
Models, Molecular
Plant Cells/metabolism
Plant Leaves/metabolism
Polysaccharides/chemistry/metabolism
*Protein Multimerization
Spectrometry, Mass, Electrospray Ionization
Tobacco/*metabolism
N-glycosylation
recombinant biopharmaceuticals
subsidiary of Patrys Limited, which holds the rights of the described IgM.
Abstract: IgM antibodies are an important player of the human's innate defense mechanisms and increasingly have gained interest as therapeutics. Although the expression of IgM antibodies in mammalian cell culture is established, this approach remains costly and alternative methods have not been developed yet. Plants have a proven record for the production of therapeutically relevant recombinant proteins. However, whether they are able to express proteins like IgM antibodies, which range among the most complex human proteins, remains unknown so far. Here we report the in planta generation of the functionally active monoclonal antitumor IgM PAT-SM6 (SM6). SM6 efficiently accumulates in plant leaves and assembles correctly into heterooligomers (pentamers and hexamers). Detailed glycosylation analysis exhibited complex and oligomannosidic N-glycans in a site-specific manner on human-serum IgM and on plant- and human-cell-line-produced SM6. Moreover, extensive in planta glycoengineering allowed the generation of SM6 decorated with sialylated human-type oligosaccharides, comparable to plasma-derived IgM. A glycosylated model of pentameric IgM exhibits different accessibility of the glycosylation sites, explaining site-specific glycosylation. Biochemical and biophysical properties and importantly biological activities of plant-derived SM6 glycoforms are comparable to the human-cell-derived counterparts. The in planta generation of one of the most complex human proteins opens new pathways toward the production of difficult-to-express proteins for pharmaceutical applications. Moreover, the generation of IgMs with a controlled glycosylation pattern allows the study of the so far unknown contribution of sugar moieties to the function of IgMs.
Notes: 1091-6490
Loos, Andreas
Gruber, Clemens
Altmann, Friedrich
Mehofer, Ulrich
Hensel, Frank
Grandits, Melanie
Oostenbrink, Chris
Stadlmayr, Gerhard
Furtmüller, Paul G
Steinkellner, Herta
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2014 Apr 29;111(17):6263-8. doi: 10.1073/pnas.1320544111. Epub 2014 Mar 31.
Author Address: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, 1190 Vienna, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 392
Author: Lopez, J. A., Chung, D. W., Fujikawa, K., Hagen, F. S., Davie, E. W. and Roth, G. J.
Year: 1988
Title: The alpha and beta chains of human platelet glycoprotein Ib are both transmembrane proteins containing a leucine-rich amino acid sequence
Journal: Proc Natl Acad Sci U S A
Volume: 85
Issue: 7
Pages: 2135-9
Epub Date: 1988/04/01
Date: Apr
Short Title: The alpha and beta chains of human platelet glycoprotein Ib are both transmembrane proteins containing a leucine-rich amino acid sequence
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.85.7.2135
PMCID: PMC279943
Accession Number: 3353370
Keywords: Amino Acid Sequence
Base Sequence
DNA/genetics
Humans
Leucine/analysis
Molecular Sequence Data
Platelet Membrane Glycoproteins/*genetics
Sequence Homology, Nucleic Acid
Abstract: The primary structure of the beta chain of human glycoprotein Ib (GPIb), the platelet receptor for von Willebrand factor, has been established by a combination of cDNA cloning and amino acid sequence analysis. A lambda phage cDNA expression library prepared from human erythroleukemia cells (HEL cells) was screened with a radiolabeled affinity-purified rabbit polyclonal antibody to the beta chain of GPIb. Eighteen positive clones were isolated and plaque-purified and the nucleotide sequences of three were determined. The composite sequence spanned 968 nucleotides and included a 5' untranslated region of 22 nucleotides, an open reading frame of 618 nucleotides encoding a signal peptide of 28 amino acids and a mature protein of 181 amino acids, a stop codon, and a 3' noncoding region of 307 nucleotides. The 3' noncoding sequence also contained a polyadenylylation signal (AATAAA) 14 nucleotides upstream from the poly(A) tail of 18 nucleotides. Edman degradation of the intact beta chain and of peptides produced by chemical cleavage yielded amino acid sequences spanning 76 residues that were identical to those predicted from the cDNA. The amino-terminal region of the beta chain contains a leucine-rich sequence of 24 amino acids that is similar to a sequence that occurs as seven tandem repeats in the alpha chain of GPIb and nine tandem repeats in leucine-rich alpha 2-glycoprotein. The leucine-rich sequence in the beta chain of GPIb is flanked on both sides by amino acid sequences that are similar to those flanking the leucine-rich tandem repeats of the alpha chain of GPIb and leucine-rich alpha 2-glycoprotein. The amino-terminal region of the beta chain of GPIb is followed by a transmembrane segment of 25 amino acids and an intracellular segment of 34 amino acids at the carboxyl terminus of the protein. The intracellular segment contains an unpaired cysteine and two potential sites for phosphorylation by cAMP-dependent protein kinase.
Notes: 1091-6490
Lopez, J A
Chung, D W
Fujikawa, K
Hagen, F S
Davie, E W
Roth, G J
HL 16919/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1988 Apr;85(7):2135-9. doi: 10.1073/pnas.85.7.2135.
Author Address: Department of Biochemistry, University of Washington, Seattle.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2061
Author: Loregger, A., Raaben, M., Nieuwenhuis, J., Tan, J. M. E., Jae, L. T., van den Hengel, L. G., Hendrix, S., van den Berg, M., Scheij, S., Song, J. Y., Huijbers, I. J., Kroese, L. J., Ottenhoff, R., van Weeghel, M., van de Sluis, B., Brummelkamp, T. and Zelcer, N.
Year: 2020
Title: Haploid genetic screens identify SPRING/C12ORF49 as a determinant of SREBP signaling and cholesterol metabolism
Journal: Nat Commun
Volume: 11
Issue: 1
Pages: 1128
Epub Date: 2020/03/01
Date: Feb 28
Short Title: Haploid genetic screens identify SPRING/C12ORF49 as a determinant of SREBP signaling and cholesterol metabolism
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-020-14811-1
PMCID: PMC7048761
Accession Number: 32111832
Keywords: Animals
Cell Line
Cholesterol/*metabolism
Embryonic Development/genetics
Endoplasmic Reticulum/metabolism
Gene Expression
Golgi Apparatus/metabolism
Haploidy
Hepatocytes/metabolism
Humans
Intracellular Signaling Peptides and Proteins/genetics/metabolism
Lipid Metabolism/genetics
Liver/metabolism
Membrane Glycoproteins/genetics/*metabolism
Membrane Proteins/genetics/*metabolism
Mice
Mice, Inbred C57BL
*Signal Transduction
Sterol Regulatory Element Binding Proteins/genetics/*metabolism
Abstract: The sterol-regulatory element binding proteins (SREBP) are central transcriptional regulators of lipid metabolism. Using haploid genetic screens we identify the SREBP Regulating Gene (SPRING/C12ORF49) as a determinant of the SREBP pathway. SPRING is a glycosylated Golgi-resident membrane protein and its ablation in Hap1 cells, Hepa1-6 hepatoma cells, and primary murine hepatocytes reduces SREBP signaling. In mice, Spring deletion is embryonic lethal yet silencing of hepatic Spring expression also attenuates the SREBP response. Mechanistically, attenuated SREBP signaling in SPRING(KO) cells results from reduced SREBP cleavage-activating protein (SCAP) and its mislocalization to the Golgi irrespective of the cellular sterol status. Consistent with limited functional SCAP in SPRING(KO) cells, reintroducing SCAP restores SREBP-dependent signaling and function. Moreover, in line with the role of SREBP in tumor growth, a wide range of tumor cell lines display dependency on SPRING expression. In conclusion, we identify SPRING as a previously unrecognized modulator of SREBP signaling.
Notes: 2041-1723
Loregger, Anke
Raaben, Matthijs
Nieuwenhuis, Joppe
Orcid: 0000-0003-4738-5895
Tan, Josephine M E
Jae, Lucas T
Orcid: 0000-0002-3531-1760
van den Hengel, Lisa G
Hendrix, Sebastian
Orcid: 0000-0003-2055-9883
van den Berg, Marlene
Scheij, Saskia
Song, Ji-Ying
Huijbers, Ivo J
Orcid: 0000-0002-7954-8604
Kroese, Lona J
Ottenhoff, Roelof
van Weeghel, Michel
Orcid: 0000-0002-4916-2866
van de Sluis, Bart
Orcid: 0000-0003-3039-4365
Brummelkamp, Thijn
Zelcer, Noam
Orcid: 0000-0001-6935-7532
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2020 Feb 28;11(1):1128. doi: 10.1038/s41467-020-14811-1.
Author Address: Department of Medical Biochemistry, Amsterdam UMC, Amsterdam Cardiovascular Sciences and Gastroenterology and Metabolism, University of Amsterdam, Meibergdreef 9, 1105AZ, Amsterdam, The Netherlands.
Oncode Institute, Division of Biochemistry, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX, Amsterdam, The Netherlands.
Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität München, Feodor Lynen-Str. 25, 81377, Munich, Germany.
Division of Experimental Animal Pathology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX, Amsterdam, The Netherlands.
Mouse Clinic for Cancer and Aging (MCCA) Transgenic Facility, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX, Amsterdam, The Netherlands.
Laboratory of Genetic and Metabolic Diseases and Core Facility Metabolomics, Academic Medical Center of the University of Amsterdam, Meibergdreef 9, 1105AZ, Amsterdam, The Netherlands.
Department of Pediatrics, University Medical Center Groningen, Antonius Deusinglaan 1, 9713AV, Groningen, The Netherlands.
iPSC/CRISPR Center Groningen, University Medical Center Groningen, Antonius Deusinglaan 1, 9713AV, Groningen, The Netherlands.
Oncode Institute, Division of Biochemistry, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX, Amsterdam, The Netherlands. t.brummelkamp@nki.nl.
CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Lazarettgasse 14, A-1090, Vienna, Austria. t.brummelkamp@nki.nl.
Cancer Genomics Center, Amsterdam, The Netherlands. t.brummelkamp@nki.nl.
Department of Medical Biochemistry, Amsterdam UMC, Amsterdam Cardiovascular Sciences and Gastroenterology and Metabolism, University of Amsterdam, Meibergdreef 9, 1105AZ, Amsterdam, The Netherlands. n.zelcer@amsterdamumc.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 783
Author: Lorenzo, P., Aspberg, A., Onnerfjord, P., Bayliss, M. T., Neame, P. J. and Heinegard, D.
Year: 2001
Title: Identification and characterization of asporin. a novel member of the leucine-rich repeat protein family closely related to decorin and biglycan
Journal: J Biol Chem
Volume: 276
Issue: 15
Pages: 12201-11
Epub Date: 2001/01/12
Date: Apr 13
Short Title: Identification and characterization of asporin. a novel member of the leucine-rich repeat protein family closely related to decorin and biglycan
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M010932200
Accession Number: 11152692
Keywords: Adult
Amino Acid Sequence
Animals
Base Sequence
Biglycan
Carrier Proteins
Cartilage, Articular/chemistry
Chromatography, Ion Exchange
Chromosome Mapping
Chromosomes, Human, Pair 9
Cloning, Molecular
DNA, Complementary
Decorin
Electrophoresis, Polyacrylamide Gel
Extracellular Matrix Proteins
Glycoproteins/*chemistry/genetics/isolation & purification
Humans
Mice
Middle Aged
Molecular Sequence Data
Proteins/*chemistry/genetics
Proteoglycans/*chemistry
Sequence Homology, Amino Acid
Abstract: Asporin, a novel member of the leucine-rich repeat family of proteins, was partially purified from human articular cartilage and meniscus. Cloning of human and mouse asporin cDNAs revealed that the protein is closely related to decorin and biglycan. It contains a putative propeptide, 4 amino-terminal cysteines, 10 leucine-rich repeats, and 2 C-terminal cysteines. In contrast to decorin and biglycan, asporin is not a proteoglycan. Instead, asporin contains a unique stretch of aspartic acid residues in its amino-terminal region. A polymorphism was identified in that the number of consecutive aspartate residues varied from 11 to 15. The 8 exons of the human asporin gene span 26 kilobases on chromosome 9q31.1-32, and the putative promoter region lacks TATA consensus sequences. The asporin mRNA is expressed in a variety of human tissues with higher levels in osteoarthritic articular cartilage, aorta, uterus, heart, and liver. The deduced amino acid sequence of asporin was confirmed by mass spectrometry of the isolated protein resulting in 84% sequence coverage. The protein contains an N-glycosylation site at Asn(281) with a heterogeneous oligosaccharide structure and a potential O-glycosylation site at Ser(54). The name asporin reflects the aspartate-rich amino terminus and the overall similarity to decorin.
Notes: Lorenzo, P
Aspberg, A
Onnerfjord, P
Bayliss, M T
Neame, P J
Heinegard, D
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2001 Apr 13;276(15):12201-11. doi: 10.1074/jbc.M010932200. Epub 2001 Jan 10.
Author Address: Department of Cell and Molecular Biology, Section for Connective Tissue Biology, Lund University, BMC plan C12, SE-221 84 Lund, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 684
Author: Lorenzo, P., Neame, P., Sommarin, Y. and Heinegård, D.
Year: 1998
Title: Cloning and deduced amino acid sequence of a novel cartilage protein (CILP) identifies a proform including a nucleotide pyrophosphohydrolase
Journal: J Biol Chem
Volume: 273
Issue: 36
Pages: 23469-75
Epub Date: 1998/08/29
Date: Sep 4
Short Title: Cloning and deduced amino acid sequence of a novel cartilage protein (CILP) identifies a proform including a nucleotide pyrophosphohydrolase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.36.23469
Accession Number: 9722584
Keywords: Amino Acid Sequence
Animals
Base Sequence
COS Cells
Cartilage/*chemistry
Chondrocytes/metabolism
Cloning, Molecular
DNA, Complementary/genetics
Extracellular Matrix Proteins/chemistry/*genetics/metabolism
Glycoproteins/chemistry/*genetics/metabolism
Humans
Molecular Sequence Data
Peptide Mapping
Polymerase Chain Reaction
Protein Precursors/*genetics
Protein Processing, Post-Translational
Pyrophosphatases/*genetics
Recombinant Proteins/biosynthesis
Restriction Mapping
Sequence Analysis
Sequence Homology, Amino Acid
Abstract: The cDNA cloning and expression in vitro and in eukaryotic cells of a novel protein isolated from human articular cartilage, cartilage intermediate layer protein (CILP) is described. A single 4. 2-kilobase mRNA detected in human articular cartilage encodes a polypeptide of 1184 amino acids with a calculated molecular mass of 132.5 kDa. The protein has a putative signal peptide of 21 amino acids, and is a proform of two polypeptides. The amino-terminal half corresponds to CILP (molecular mass of 78.5 kDa, not including post-translational modifications) and the carboxyl-terminal half corresponds to a protein homologous to a porcine nucleotide pyrophosphohydrolase, NTPPHase (molecular mass of 51.8 kDa, not including post-translational modifications). CILP has 30 cysteines and six putative N-glycosylation sites. The human homolog of porcine NTPPHase described here contains 10 cysteine residues and two putative N-glycosylation sites. In the precursor protein the NTPPHase region is immediately preceded by a tetrapeptide conforming to a furin proteinase cleavage consensus sequence. Expression of the full-length cDNA in a cell-free translation system and in COS-7 or EBNA cells indicates that the precursor protein is synthesized as a single polypeptide chain that is processed, possibly by a furin-like protease, into two polypeptides upon or preceding secretion.
Notes: Lorenzo, P
Neame, P
Sommarin, Y
Heinegård, D
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1998 Sep 4;273(36):23469-75. doi: 10.1074/jbc.273.36.23469.
Author Address: Department of Cell and Molecular Biology, Section for Connective Tissue Biology, University of Lund, P.O.Box 94, S-221 00 Lund, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 433
Author: Lottspeich, F., Kellermann, J., Henschen, A., Foertsch, B. and Müller-Esterl, W.
Year: 1985
Title: The amino acid sequence of the light chain of human high-molecular-mass kininogen
Journal: Eur J Biochem
Volume: 152
Issue: 2
Pages: 307-14
Epub Date: 1985/10/15
Date: Oct 15
Short Title: The amino acid sequence of the light chain of human high-molecular-mass kininogen
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1985.tb09199.x
Accession Number: 4054110
Keywords: Amino Acid Sequence
Animals
Cattle
Chemical Phenomena
Chemistry
Chromatography, High Pressure Liquid
Computers
Humans
Kininogens/*analysis
Molecular Weight
Peptide Fragments/analysis
Protein Conformation
Solubility
Species Specificity
Abstract: The complete amino acid sequence of the light chain of human high-molecular-mass kininogen has been determined. The peptide chain contains 255 amino acid residues. The half-cystine, which forms the disulfide bridge to the heavy chain, was identified in position 225. Nine carbohydrate attachment sites were found. All carbohydrate side chains are O-glycosidically linked. Alignment of the present sequence with the bovine kininogen light chain sequence shows a high degree of homology, except for an extension of 22 amino acids within the histidine-rich part of the sequence. The histidine-rich region may have arisen by gene multiplication during evolution.
Notes: Lottspeich, F
Kellermann, J
Henschen, A
Foertsch, B
Müller-Esterl, W
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1985 Oct 15;152(2):307-14. doi: 10.1111/j.1432-1033.1985.tb09199.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1065
Author: Lou, M., Garrett, T. P., McKern, N. M., Hoyne, P. A., Epa, V. C., Bentley, J. D., Lovrecz, G. O., Cosgrove, L. J., Frenkel, M. J. and Ward, C. W.
Year: 2006
Title: The first three domains of the insulin receptor differ structurally from the insulin-like growth factor 1 receptor in the regions governing ligand specificity
Journal: Proc Natl Acad Sci U S A
Volume: 103
Issue: 33
Pages: 12429-34
Epub Date: 2006/08/09
Date: Aug 15
Short Title: The first three domains of the insulin receptor differ structurally from the insulin-like growth factor 1 receptor in the regions governing ligand specificity
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0605395103
PMCID: PMC1533800
Accession Number: 16894147
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cricetinae
Crystallography, X-Ray
Humans
Insulin-Like Growth Factor I/*chemistry/genetics/metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Mutation
Protein Binding
*Protein Structure, Secondary
*Protein Structure, Tertiary
Receptor, IGF Type 1/chemistry/genetics/metabolism
Receptor, Insulin/*chemistry/genetics/metabolism
Sequence Alignment
Abstract: The insulin receptor (IR) and the type-1 insulin-like growth factor receptor (IGF1R) are homologous multidomain proteins that bind insulin and IGF with differing specificity. Here we report the crystal structure of the first three domains (L1-CR-L2) of human IR at 2.3 A resolution and compare it with the previously determined structure of the corresponding fragment of IGF1R. The most important differences seen between the two receptors are in the two regions governing ligand specificity. The first is at the corner of the ligand-binding surface of the L1 domain, where the side chain of F39 in IR forms part of the ligand binding surface involving the second (central) beta-sheet. This is very different to the location of its counterpart in IGF1R, S35, which is not involved in ligand binding. The second major difference is in the sixth module of the CR domain, where IR contains a larger loop that protrudes further into the ligand-binding pocket. This module, which governs IGF1-binding specificity, shows negligible sequence identity, significantly more alpha-helix, an additional disulfide bond, and opposite electrostatic potential compared to that of the IGF1R.
Notes: 1091-6490
Lou, Meizhen
Garrett, Thomas P J
McKern, Neil M
Hoyne, Peter A
Epa, V Chandana
Bentley, John D
Lovrecz, George O
Cosgrove, Leah J
Frenkel, Maurice J
Ward, Colin W
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2006 Aug 15;103(33):12429-34. doi: 10.1073/pnas.0605395103. Epub 2006 Aug 7.
Author Address: Division of Molecular and Health Technologies, Commonwealth Scientific and Industrial Research Organization, 343 Royal Parade, Parkville, Victoria 3052, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 469
Author: Lozier, J., Takahashi, N. and Putnam, F. W.
Year: 1984
Title: Complete amino acid sequence of human plasma beta 2-glycoprotein I
Journal: Proc Natl Acad Sci U S A
Volume: 81
Issue: 12
Pages: 3640-4
Epub Date: 1984/06/01
Date: Jun
Short Title: Complete amino acid sequence of human plasma beta 2-glycoprotein I
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.81.12.3640
PMCID: PMC345274
Accession Number: 6587378
Keywords: Amino Acid Sequence
Binding Sites
Biological Evolution
*Blood Proteins
Disulfides
Glucosamine
Glycoproteins/*blood
Humans
Macromolecular Substances
Molecular Weight
beta 2-Glycoprotein I
Abstract: We have determined the complete amino acid sequence of beta 2-glycoprotein I (Mr, congruent to 50,000), a human plasma protein that is associated with lipids and binds to platelets but whose function is not yet known. The protein consists of 326 amino acids and has five attached glucosamine-containing oligosaccharides. The protein is rich in cysteine and proline, and the sequence is notable for the frequent occurrence of Cys-Pro linkages at regular intervals. Computerized analysis of the sequence reveals five consecutive homologous segments in which cysteine, proline, and tryptophan appear to be highly conserved. This suggests that beta 2-glycoprotein I may have evolved by repeated duplications of a gene coding for a 60-amino acid segment of protein.
Notes: 1091-6490
Lozier, J
Takahashi, N
Putnam, F W
AM 19221/AM/NIADDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1984 Jun;81(12):3640-4. doi: 10.1073/pnas.81.12.3640.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1343
Author: Lu, C., Mi, L. Z., Grey, M. J., Zhu, J., Graef, E., Yokoyama, S. and Springer, T. A.
Year: 2010
Title: Structural evidence for loose linkage between ligand binding and kinase activation in the epidermal growth factor receptor
Journal: Mol Cell Biol
Volume: 30
Issue: 22
Pages: 5432-43
Epub Date: 2010/09/15
Date: Nov
Short Title: Structural evidence for loose linkage between ligand binding and kinase activation in the epidermal growth factor receptor
Alternate Journal: Molecular and cellular biology
ISSN: 0270-7306 (Print)
0270-7306
DOI: 10.1128/mcb.00742-10
PMCID: PMC2976375
Accession Number: 20837704
Keywords: Amino Acid Sequence
Cell Line
Cross-Linking Reagents/chemistry
Cysteine/chemistry/genetics
Dimerization
Disulfides/chemistry
Enzyme Activation
ErbB Receptors/*chemistry/genetics/*metabolism
Humans
*Ligands
Models, Molecular
Molecular Sequence Data
Protein Binding
*Protein Structure, Quaternary
*Protein Structure, Secondary
Sequence Alignment
Signal Transduction/*physiology
X-Ray Diffraction
Abstract: The mechanisms by which signals are transmitted across the plasma membrane to regulate signaling are largely unknown for receptors with single-pass transmembrane domains such as the epidermal growth factor receptor (EGFR). A crystal structure of the extracellular domain of EGFR dimerized by epidermal growth factor (EGF) reveals the extended, rod-like domain IV and a small, hydrophobic domain IV interface compatible with flexibility. The crystal structure and disulfide cross-linking suggest that the 7-residue linker between the extracellular and transmembrane domains is flexible. Disulfide cross-linking of the transmembrane domain shows that EGF stimulates only moderate association in the first two α-helical turns, in contrast to association throughout the membrane over five α-helical turns in glycophorin A and integrin. Furthermore, systematic mutagenesis to leucine and phenylalanine suggests that no specific transmembrane interfaces are required for EGFR kinase activation. These results suggest that linkage between ligand-induced dimerization and tyrosine kinase activation is much looser than was previously envisioned.
Notes: 1098-5549
Lu, Chafen
Mi, Li-Zhi
Grey, Michael J
Zhu, Jieqing
Graef, Elizabeth
Yokoyama, Shigeyuki
Springer, Timothy A
P01 HL048675/HL/NHLBI NIH HHS/United States
HL-48675/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cell Biol. 2010 Nov;30(22):5432-43. doi: 10.1128/MCB.00742-10. Epub 2010 Sep 13.
Author Address: Immune Disease Institute and Department of Pathology, Harvard Medical School, 3 Blackfan Circle, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 274
Author: Lu, H. S., Clogston, C. L., Wypych, J., Fausset, P. R., Lauren, S., Mendiaz, E. A., Zsebo, K. M. and Langley, K. E.
Year: 1991
Title: Amino acid sequence and post-translational modification of stem cell factor isolated from buffalo rat liver cell-conditioned medium
Journal: J Biol Chem
Volume: 266
Issue: 13
Pages: 8102-7
Epub Date: 1991/05/05
Date: May 5
Short Title: Amino acid sequence and post-translational modification of stem cell factor isolated from buffalo rat liver cell-conditioned medium
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1708771
Keywords: Amino Acid Sequence
Animals
Cells, Cultured
Disulfides/chemistry
Glycosylation
Hematopoietic Cell Growth Factors/chemistry/*metabolism
Liver/cytology/*metabolism
Mass Spectrometry
Molecular Sequence Data
*Protein Processing, Post-Translational
Rats
Rats, Inbred BUF
Stem Cell Factor
Abstract: Stem cell factor (SCF) isolated from culture medium conditioned by Buffalo rat liver cells was subjected to detailed structural analysis. Attempts at direct N-terminal sequencing of the factor indicated that its N terminus is blocked as pyroglutamic acid (Zsebo, K. M., Wypych, J., McNiece, I. K., Lu, H. S., Smith, K. A., Karkare, S. B., Sachdev, R. K., Yuschenkoff, V. N., Birkett, N. C., Williams, L. R., Satyagal, V. N., Bosselman, R. A., Mendiaz, E. A., and Langley, K. E. (1990) Cell 63, 195-201). The removal of the blocking pyroglutamate by pyroglutamate aminopeptidase allowed sequencing of the polypeptide chain to position 47. Stem cell factor was also digested with CNBr, trypsin, Staphylococcus aureus protease (strain V8), and AspN peptidase to generate different sets of peptides that were then separated by reverse-phase high-performance liquid chromatography and sequenced. Sequence of an internal peptide fragment obtained by cleavage of stem cell factor at a single tryptophanyl peptide bond was also obtained. From these analyses, the complete amino acid sequence could be constructed. The factor as isolated is a single polypeptide of 164 or 165 amino acids. The sequence is confirmatory to a sequence deduced from a cDNA sequence and provides important evidence for C-terminal processing of the polypeptide encoded by cDNA. There are four potential N-linked glycosylation sites. Asn65, Asn72, Asn109, and Asn120. Sequence determination of isolated peptides suggested that Asn120 is glycosylated, Asn65 and Asn109 glycosylated in some molecules but not in others, and Asn72 not glycosylated. Amino acids at three positions, i.e. 142, 143, and 155, could not be detected during sequence analysis. Since the gene sequence codes for Ser, Thr, and Thr at these positions (Martin, F. H., Suggs, S. V., Langley, K. E., Lu, H. S., Ting, J., Okino, K. H., Morris, C. F., McNiece, I. K., Jacobsen, F. W., Mendiaz, E. A., Birkett, N. C., Smith, K. C., Johnson, M. J., Parker, V. P., Flores, J. C., Patel, A. C., Fisher, E. F., Erjavec, H. O., Herrera, C. J., Wypych, J., Sachdev, R. K., Pope, J. A., Leslie, I., Wen, D., Lin, C. W., Cupples, R. L., and Zsebo, K. M. (1990) Cell 63, 203-211), they could be sites of O-linked carbohydrate attachment. The four cysteines form two intramolecular disulfide bonds, Cys4-Cys89 and Cys43-Cys138.
Notes: Lu, H S
Clogston, C L
Wypych, J
Fausset, P R
Lauren, S
Mendiaz, E A
Zsebo, K M
Langley, K E
Journal Article
United States
J Biol Chem. 1991 May 5;266(13):8102-7.
Author Address: Amgen Inc., Thousand Oaks, California 91320.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 234
Author: Lu, H. S., Clogston, C. L., Wypych, J., Parker, V. P., Lee, T. D., Swiderek, K., Baltera, R. F., Jr., Patel, A. C., Chang, D. C., Brankow, D. W. and et al.
Year: 1992
Title: Post-translational processing of membrane-associated recombinant human stem cell factor expressed in Chinese hamster ovary cells
Journal: Arch Biochem Biophys
Volume: 298
Issue: 1
Pages: 150-8
Epub Date: 1992/10/11
Date: Oct
Short Title: Post-translational processing of membrane-associated recombinant human stem cell factor expressed in Chinese hamster ovary cells
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1016/0003-9861(92)90106-7
Accession Number: 1381905
Keywords: Amino Acid Sequence
Animals
CHO Cells
Chromatography, High Pressure Liquid
Cricetinae
Disulfides/chemistry
Glycosylation
Hematopoietic Cell Growth Factors/chemistry/*metabolism
Humans
In Vitro Techniques
Mass Spectrometry
Molecular Sequence Data
Molecular Weight
Protein Processing, Post-Translational
Proto-Oncogene Proteins/metabolism
Proto-Oncogene Proteins c-kit
Receptors, Cell Surface/metabolism
Recombinant Proteins/metabolism
Stem Cell Factor
Structure-Activity Relationship
Abstract: This report describes the structure of soluble human stem cell factor isolated from the conditioned medium of Chinese hamster ovary (CHO) cells transfected with stem cell factor (SCF) cDNA, which encodes a leader sequence plus 248 additional amino acids. The 248 amino acids include a hydrophobic transmembrane region at positions 190-212. The isolated material is glycosylated and three bands (apparent M(r) 28,000, M(r) 35,000, and M(r) 40,000) are evident by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. After complete deglycosylation, the molecular weight by SDS-polyacrylamide gel electrophoresis is 18,000-19,000. Structural analyses of the intact SCF, the deglycosylated SCF, and a deglycosylated C-terminal peptide were performed by laser desorption, fast atom bombardment, or electrospray mass spectrometry. Pulse-labeling of cells with 35S-labeled Met and Cys resulted in cell-associated glycosylated SCF of M(r) 33,000-45,000 which was converted to M(r) 33,000 by in vitro treatment with glycosidases. During a chase with unlabeled Met and Cys, labeled SCF of M(r) 28,000, M(r) 35,000, and M(r) 40,000 appeared in the medium; it was converted to M(r) 18,000-19,000 by glycosidase treatment. SCF at the surface of the transfected CHO cells could be demonstrated by immunofluorescence. The data obtained indicate that the recombinant human stem cell factor, as isolated, represents proteolytically processed forms containing amino acids 1-165, derived from the initially synthesized membrane-bound form of 248 amino acids. Further characterization indicated that the M(r) 28,000 form is glycosylated at Asn120, the M(r) 35,000 form at Asn120 and Asn65, and the M(r) 40,000 form at Asn120, Asn93, and Asn65. Each form also contains O-linked carbohydrate. The N-linked glycosylation, particularly that at Asn93 and at Asn65, adversely affects in vitro biological activity and receptor binding.
Notes: Lu, H S
Clogston, C L
Wypych, J
Parker, V P
Lee, T D
Swiderek, K
Baltera, R F Jr
Patel, A C
Chang, D C
Brankow, D W
Journal Article
United States
Arch Biochem Biophys. 1992 Oct;298(1):150-8. doi: 10.1016/0003-9861(92)90106-7.
Author Address: Amgen, Inc., Amgen Center, Thousand Oaks, California 91320.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1431
Author: Lu, J., Ellsworth, J. L., Hamacher, N., Oak, S. W. and Sun, P. D.
Year: 2011
Title: Crystal structure of Fcγ receptor I and its implication in high affinity γ-immunoglobulin binding
Journal: J Biol Chem
Volume: 286
Issue: 47
Pages: 40608-13
Epub Date: 2011/10/04
Date: Nov 25
Short Title: Crystal structure of Fcγ receptor I and its implication in high affinity γ-immunoglobulin binding
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.257550
PMCID: PMC3220521
Accession Number: 21965667
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cricetinae
Cricetulus
Crystallography, X-Ray
Humans
Immunoglobulin Fc Fragments/metabolism
Immunoglobulin G/*metabolism
Models, Molecular
Molecular Sequence Data
N-Acetylneuraminic Acid/metabolism
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, IgG/*chemistry/*metabolism
Substrate Specificity
Abstract: Fcγ receptors (FcγRs) play critical roles in humoral and cellular immune responses through interactions with the Fc region of immunoglobulin G (IgG). Among them, FcγRI is the only high affinity receptor for IgG and thus is a potential target for immunotherapy. Here we report the first crystal structure of an FcγRI with all three extracellular Ig-like domains (designated as D1, D2, and D3). The structure shows that, first, FcγRI has an acute D1-D2 hinge angle similar to that of FcεRI but much smaller than those observed in the low affinity Fcγ receptors. Second, the D3 domain of FcγRI is positioned away from the putative IgG binding site on the receptor and is thus unlikely to make direct contacts with Fc. Third, the replacement of FcγRIII FG-loop ((171)LVGSKNV(177)) with that of FcγRI ((171)MGKHRY(176)) resulted in a 15-fold increase in IgG(1) binding affinity, whereas a valine insertion in the FcγRI FG-loop ((171)MVGKHRY(177)) abolished the affinity enhancement. Thus, the FcγRI FG-loop with its conserved one-residue deletion is critical to the high affinity IgG binding. The structural results support FcγRI binding to IgG in a similar mode as its low affinity counterparts. Taken together, our study suggests a molecular mechanism for the high affinity IgG recognition by FcγRI and provides a structural basis for understanding its physiological function and its therapeutic implication in treating autoimmune diseases.
Notes: 1083-351x
Lu, Jinghua
Ellsworth, Jeff L
Hamacher, Nels
Oak, Si Won
Sun, Peter D
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
J Biol Chem. 2011 Nov 25;286(47):40608-13. doi: 10.1074/jbc.M111.257550. Epub 2011 Sep 29.
Author Address: Structural Immunology Section, Laboratory of Immunogenetics, NIAID, National Institutes of Health, Rockville, Maryland, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1752
Author: Lu, L., Fan, D., Hu, C. W., Worth, M., Ma, Z. X. and Jiang, J.
Year: 2016
Title: Distributive O-GlcNAcylation on the Highly Repetitive C-Terminal Domain of RNA Polymerase II
Journal: Biochemistry
Volume: 55
Issue: 7
Pages: 1149-58
Epub Date: 2016/01/26
Date: Feb 23
Short Title: Distributive O-GlcNAcylation on the Highly Repetitive C-Terminal Domain of RNA Polymerase II
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/acs.biochem.5b01280
PMCID: PMC6314847
NIHMSID: NIHMS1001510
Accession Number: 26807597
Keywords: Acetylglucosamine/*metabolism
Gene Deletion
Glycosylation
HeLa Cells
Humans
Kinetics
Molecular Weight
N-Acetylglucosaminyltransferases/chemistry/genetics/*metabolism
Peptide Fragments/chemistry/genetics/metabolism
Peptide Mapping
Protein Interaction Domains and Motifs
*Protein Processing, Post-Translational
RNA Polymerase II/chemistry/genetics/*metabolism
Recombinant Fusion Proteins/chemistry/metabolism
Recombinant Proteins/chemistry/metabolism
Repetitive Sequences, Amino Acid
Reproducibility of Results
Spectrometry, Mass, Electrospray Ionization
Substrate Specificity
Tandem Mass Spectrometry
Abstract: O-GlcNAcylation is a nutrient-responsive glycosylation that plays a pivotal role in transcriptional regulation. Human RNA polymerase II (Pol II) is extensively modified by O-linked N-acetylglucosamine (O-GlcNAc) on its unique C-terminal domain (CTD), which consists of 52 heptad repeats. One approach to understanding the function of glycosylated Pol II is to determine the mechanism of dynamic O-GlcNAcylation on the CTD. Here, we discovered that the Pol II CTD can be extensively O-GlcNAcylated in vitro and in cells. Efficient glycosylation requires a minimum of 20 heptad repeats of the CTD and more than half of the N-terminal domain of O-GlcNAc transferase (OGT). Under conditions of saturated sugar donor, we monitored the attachment of more than 20 residues of O-GlcNAc to the full-length CTD. Surprisingly, glycosylation on the periodic CTD follows a distributive mechanism, resulting in highly heterogeneous glycoforms. Our data suggest that initial O-GlcNAcylation can take place either on the proximal or on the distal region of the CTD, and subsequent glycosylation occurs similarly over the entire CTD with nonuniform distributions. Moreover, removal of O-GlcNAc from glycosylated CTD is also distributive and is independent of O-GlcNAcylation level. Our results suggest that O-GlcNAc cycling enzymes can employ a similar mechanism to react with other protein substrates on multiple sites. Distributive O-GlcNAcylation on Pol II provides another regulatory mechanism of transcription in response to fluctuating cellular conditions.
Notes: 1520-4995
Lu, Lei
Fan, Dacheng
Hu, Chia-Wei
Worth, Matthew
Ma, Zhi-Xiong
Jiang, Jiaoyang
S10 RR029531/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochemistry. 2016 Feb 23;55(7):1149-58. doi: 10.1021/acs.biochem.5b01280. Epub 2016 Feb 9.
Author Address: Pharmaceutical Sciences Division, School of Pharmacy, and ‡Department of Chemistry, University of Wisconsin-Madison , Madison, Wisconsin 53705, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2070
Author: Lu, S., Yin, X., Wang, J., Gu, Q., Huang, Q., Jin, N., Chu, D., Xu, Z., Liu, F. and Qian, W.
Year: 2020
Title: SIRT1 regulates O-GlcNAcylation of tau through OGT
Journal: Aging (Albany NY)
Volume: 12
Issue: 8
Pages: 7042-7055
Epub Date: 2020/04/21
Date: Apr 20
Short Title: SIRT1 regulates O-GlcNAcylation of tau through OGT
Alternate Journal: Aging
ISSN: 1945-4589
DOI: 10.18632/aging.103062
PMCID: PMC7202539
Accession Number: 32310828
Keywords: Acetylglucosamine/*metabolism
Animals
COS Cells
Chlorocebus aethiops
Cyclic AMP Response Element-Binding Protein/physiology
Glycosylation
HEK293 Cells
Humans
N-Acetylglucosaminyltransferases/genetics/*physiology
Phosphorylation
Rats
Rats, Sprague-Dawley
Sirtuin 1/*physiology
tau Proteins/*metabolism
*Alzheimer’s disease
*O-GlcNAcylation of tau
*Ogt
*Sirt1
*phosphorylation of tau
Abstract: Tau is modified with O-GlcNAcylation extensively in human brain. The O-GlcNAcylation levels of tau are decreased in Alzheimer's disease (AD) brain. Sirtuin type 1 (SIRT1) is an enzyme that deacetylates proteins including transcriptional factors and associates with neurodegenerative diseases, such as AD. Aberrant SIRT1 expression levels in AD brain is in parallel with the accumulation of tau. cAMP response element binding protein (CREB), a cellular transcription factor, plays a critical role in learning and memory. In this present study, we found SIRT1 deacetylates CREB and inhibits phosphorylation of CREB at Ser133. The inactivated CREB suppresses OGT expression and therefore decreases the O-GlcNAcylation of tau and thus increases the phosphorylation of tau at specific sites. These findings suggest that SIRT1 may be a potential therapeutic target for treating tauopathies.
Notes: 1945-4589
Lu, Shu
Yin, Xiaomin
Wang, Jia
Gu, Qun
Huang, Qin
Jin, Nana
Chu, Dandan
Xu, Ziqi
Liu, Fei
Qian, Wei
Journal Article
Research Support, Non-U.S. Gov't
Aging (Albany NY). 2020 Apr 20;12(8):7042-7055. doi: 10.18632/aging.103062. Epub 2020 Apr 20.
Author Address: Department of Intensive Care Unit, The Affiliated Hospital of Nantong University, Nantong, Jiangsu, P. R. China.
Department of Biochemistry and Molecular Biology, Medical School, Nantong, Jiangsu, P. R. China.
Jiangsu Key Laboratory of Neuroregeneration of Jiangsu and Ministry of Education of China, Co-innovation Center of Neuroregeneration, Nantong University, Nantong, Jiangsu, P. R. China.
Department of Neurochemistry, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, NY 10314, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1719
Author: Lu, Y. C., Nazarko, O. V., Sando, R., 3rd, Salzman, G. S., Li, N. S., Südhof, T. C. and Araç, D.
Year: 2015
Title: Structural Basis of Latrophilin-FLRT-UNC5 Interaction in Cell Adhesion
Journal: Structure
Volume: 23
Issue: 9
Pages: 1678-1691
Epub Date: 2015/08/04
Date: Sep 1
Short Title: Structural Basis of Latrophilin-FLRT-UNC5 Interaction in Cell Adhesion
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2015.06.024
PMCID: PMC4851429
NIHMSID: NIHMS780319
Accession Number: 26235030
Keywords: Binding Sites
Cell Adhesion
Crystallography, X-Ray
HEK293 Cells
Humans
Membrane Glycoproteins
Membrane Proteins/*chemistry/genetics/*metabolism
Models, Molecular
Mutation
Netrin Receptors
Protein Multimerization
Receptors, Cell Surface/chemistry/*metabolism
Receptors, G-Protein-Coupled/*chemistry/genetics/*metabolism
Receptors, Peptide/*chemistry/genetics/*metabolism
Synapses/metabolism
adhesion-type GPCR
attention deficit hyperactivity disorder
neural development
neuronal communication
synaptic junction
Abstract: Fibronectin leucine-rich repeat transmembrane proteins (FLRTs) are cell-adhesion molecules with emerging functions in cortical development and synapse formation. Their extracellular regions interact with latrophilins (LPHNs) to mediate synapse development, and with Uncoordinated-5 (UNC5)/netrin receptors to control the migration of neurons in the developing cortex. Here, we present the crystal structures of FLRT3 in isolation and in complex with LPHN3. The LPHN3/FLRT3 structure reveals that LPHN3 binds to FLRT3 at a site distinct from UNC5. Structure-based mutations specifically disrupt LPHN3/FLRT3 binding, but do not disturb their interactions with other proteins or their cell-membrane localization. Thus, they can be used as molecular tools to dissect the functions of FLRTs and LPHNs in vivo. Our results suggest that UNC5 and LPHN3 can simultaneously bind to FLRT3, forming a trimeric complex, and that FLRT3 may form transsynaptic complexes with both LPHN3 and UNC5. These findings provide molecular insights for understanding the role of cell-adhesion proteins in synapse function.
Notes: 1878-4186
Lu, Yue C
Nazarko, Olha V
Sando, Richard 3rd
Salzman, Gabriel S
Li, Nan-Sheng
Südhof, Thomas C
Araç, Demet
T32 GM007281/GM/NIGMS NIH HHS/United States
Journal Article
Structure. 2015 Sep 1;23(9):1678-1691. doi: 10.1016/j.str.2015.06.024. Epub 2015 Jul 30.
Author Address: Department of Biochemistry and Molecular Biology, The University of Chicago, 929 East 57(th) Street, GCIS W219, Chicago, IL 60637, USA.
Department of Molecular and Cellular Physiology, Stanford University, Stanford, CA 94305, USA; Howard Hughes Medical Institute, Stanford University, Stanford, CA 94305, USA.
Department of Biochemistry and Molecular Biology, The University of Chicago, 929 East 57(th) Street, GCIS W219, Chicago, IL 60637, USA. Electronic address: arac@uchicago.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1490
Author: Luan, X., Lu, Q., Jiang, Y., Zhang, S., Wang, Q., Yuan, H., Zhao, W., Wang, J. and Wang, X.
Year: 2012
Title: Crystal structure of human RANKL complexed with its decoy receptor osteoprotegerin
Journal: J Immunol
Volume: 189
Issue: 1
Pages: 245-52
Epub Date: 2012/06/06
Date: Jul 1
Short Title: Crystal structure of human RANKL complexed with its decoy receptor osteoprotegerin
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767
DOI: 10.4049/jimmunol.1103387
Accession Number: 22664871
Keywords: Animals
Crystallography, X-Ray
Cysteine/chemistry/metabolism
Humans
Mice
Mutagenesis, Site-Directed
Osteoprotegerin/*chemistry/genetics/*physiology
Protein Binding/immunology
Protein Multimerization
Protein Structure, Tertiary
RANK Ligand/*chemistry/genetics/*physiology
Abstract: Receptor activator of NF-κB ligand (RANKL), its signaling receptor RANK, and its decoy receptor osteoprotegerin (OPG) constitute a molecular triad that is critical in regulating bone remodeling, and also plays multiple roles in the immune system. OPG binds RANKL directly to block its interaction with RANK. In this article, we report the 2.7-Å crystal structure of human RANKL trimer in complex with the N-terminal fragment of human OPG containing four cysteine-rich TNFR homologous domains (OPG-CRD). The structure shows that RANKL trimer uses three equivalent grooves between two neighboring monomers to interact with three OPG-CRD monomers symmetrically. A loop from the CRD3 domain of OPG-CRD inserts into the shallow groove of RANKL, providing the major binding determinant that is further confirmed by affinity measurement and osteoclast differentiation assay. These results, together with a previously reported mouse RANKL/RANK complex structure, reveal that OPG exerts its decoy receptor function by directly blocking the accessibilities of important interacting residues of RANKL for RANK recognition. Structural comparison with TRAIL/death receptor 5 complex also reveals structural basis for the cross-reactivity of OPG to TRAIL.
Notes: 1550-6606
Luan, Xudong
Lu, Qingyu
Jiang, Yinan
Zhang, Senyan
Wang, Qing
Yuan, Huihui
Zhao, Wenming
Wang, Jiawei
Wang, Xinquan
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 2012 Jul 1;189(1):245-52. doi: 10.4049/jimmunol.1103387. Epub 2012 Jun 4.
Author Address: Center for Structural Biology, School of Life Sciences, Ministry of Education Key Laboratory of Protein Science, Tsinghua University, Beijing 100084, People's Republic of China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1881
Author: Luanpitpong, S., Angsutararux, P., Samart, P., Chanthra, N., Chanvorachote, P. and Issaragrisil, S.
Year: 2017
Title: Hyper-O-GlcNAcylation induces cisplatin resistance via regulation of p53 and c-Myc in human lung carcinoma
Journal: Sci Rep
Volume: 7
Issue: 1
Pages: 10607
Epub Date: 2017/09/08
Date: Sep 6
Short Title: Hyper-O-GlcNAcylation induces cisplatin resistance via regulation of p53 and c-Myc in human lung carcinoma
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/s41598-017-10886-x
PMCID: PMC5587763
Accession Number: 28878262
Keywords: Antineoplastic Agents/*pharmacology
Apoptosis/drug effects
Caspases/metabolism
Cell Line, Tumor
Cisplatin/*pharmacology
Drug Resistance, Neoplasm/*genetics
Gene Expression Regulation, Neoplastic/*drug effects
*Genes, myc
Humans
Lung Neoplasms
Protein Processing, Post-Translational/drug effects
RNA, Messenger/genetics
Signal Transduction
Tumor Suppressor Protein p53/*genetics/*metabolism
beta-N-Acetylhexosaminidases/antagonists & inhibitors/genetics/metabolism
Abstract: Aberrant metabolism in hexosamine biosynthetic pathway (HBP) has been observed in several cancers, affecting cellular signaling and tumor progression. However, the role of O-GlcNAcylation, a post-translational modification through HBP flux, in apoptosis remains unclear. Here, we found that hyper-O-GlcNAcylation in lung carcinoma cells by O-GlcNAcase inhibition renders the cells to apoptosis resistance to cisplatin (CDDP). Profiling of various key regulatory proteins revealed an implication of either p53 or c-Myc in the apoptosis regulation by O-GlcNAcylation, independent of p53 status. Using co-immunoprecipitation and correlation analyses, we found that O-GlcNAcylation of p53 under certain cellular contexts, i.e. high p53 activation, promotes its ubiquitin-mediated proteasomal degradation, resulting in a gain of oncogenic and anti-apoptotic functions. By contrast, O-GlcNAcylation of c-Myc inhibits its ubiquitination and subsequent proteasomal degradation. Gene manipulation studies revealed that O-GlcNAcylation of p53/c-Myc is in part a regulator of CDDP-induced apoptosis. Accordingly, we classified CDDP resistance by hyper-O-GlcNAcylation in lung carcinoma cells as either p53 or c-Myc dependence based on their molecular targets. Together, our findings provide novel mechanisms for the regulation of lung cancer cell apoptosis that could be important in understanding clinical drug resistance and suggest O-GlcNAcylation as a potential target for cancer therapy.
Notes: 2045-2322
Luanpitpong, Sudjit
Orcid: 0000-0002-1639-4935
Angsutararux, Paweorn
Samart, Parinya
Chanthra, Nawin
Chanvorachote, Pithi
Issaragrisil, Surapol
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2017 Sep 6;7(1):10607. doi: 10.1038/s41598-017-10886-x.
Author Address: Siriraj Center of Excellence for Stem Cell Research, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand.
Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand.
Department of Pharmacology and Physiology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, 10330, Thailand.
Siriraj Center of Excellence for Stem Cell Research, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. surapol.iss@mahidol.ac.th.
Division of Hematology, Department of Medicine, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. surapol.iss@mahidol.ac.th.
Bangkok Hematology Center, Wattanosoth Hospital, BDMS Center of Excellence for Cancer, Bangkok, 10310, Thailand. surapol.iss@mahidol.ac.th.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1894
Author: Luanpitpong, S., Chanthra, N., Janan, M., Poohadsuan, J., Samart, P., Y, U. Pratya, Rojanasakul, Y. and Issaragrisil, S.
Year: 2018
Title: Inhibition of O-GlcNAcase Sensitizes Apoptosis and Reverses Bortezomib Resistance in Mantle Cell Lymphoma through Modification of Truncated Bid
Journal: Mol Cancer Ther
Volume: 17
Issue: 2
Pages: 484-496
Epub Date: 2017/11/24
Date: Feb
Short Title: Inhibition of O-GlcNAcase Sensitizes Apoptosis and Reverses Bortezomib Resistance in Mantle Cell Lymphoma through Modification of Truncated Bid
Alternate Journal: Molecular cancer therapeutics
ISSN: 1535-7163 (Print)
1535-7163
DOI: 10.1158/1535-7163.mct-17-0390
PMCID: PMC5805589
NIHMSID: NIHMS922304
Accession Number: 29167312
Keywords: Apoptosis
Bortezomib/pharmacology/*therapeutic use
Humans
Lymphoma, Mantle-Cell/*drug therapy/pathology
N-Acetylglucosaminyltransferases/*antagonists & inhibitors
Protein Processing, Post-Translational
Signal Transduction
Abstract: Aberrant energy metabolism represents a hallmark of cancer and contributes to numerous aggressive behaviors of cancer cells, including cell death and survival. Despite the poor prognosis of mantle cell lymphoma (MCL), due to the inevitable development of drug resistance, metabolic reprograming of MCL cells remains an unexplored area. Posttranslational modification of proteins via O-GlcNAcylation is an ideal sensor for nutritional changes mediated by O-GlcNAc transferase (OGT) and is removed by O-GlcNAcase (OGA). Using various small-molecule inhibitors of OGT and OGA, we found for the first time that O-GlcNAcylation potentiates MCL response to bortezomib. CRISPR interference of MGEA5 (encoding OGA) validated the apoptosis sensitization by O-GlcNAcylation and OGA inhibition. To identify the potential clinical candidates, we tested MCL response to drug-like OGA inhibitor, ketoconazole, and verified that it exerts similar sensitizing effect on bortezomib-induced apoptosis. Investigations into the underlying molecular mechanisms reveal that bortezomib and ketoconazole act in concert to cause the accumulation of truncated Bid (tBid). Not only does ketoconazole potentiate tBid induction, but also increases tBid stability through O-GlcNAcylation that interferes with tBid ubiquitination and proteasomal degradation. Remarkably, ketoconazole strongly enhances bortezomib-induced apoptosis in de novo bortezomib-resistant MCL cells and in patient-derived primary cells with minimal cytotoxic effect on normal peripheral blood mononuclear cells and hepatocytes, suggesting its potential utility as a safe and effective adjuvant for MCL. Together, our findings provide novel evidence that combination of bortezomib and ketoconazole or other OGA inhibitors may present a promising strategy for the treatment of drug-resistant MCL. Mol Cancer Ther; 17(2); 484-96. ©2017 AACR.
Notes: 1538-8514
Luanpitpong, Sudjit
Chanthra, Nawin
Janan, Montira
Poohadsuan, Jirarat
Samart, Parinya
U-Pratya, Yaowalak
Rojanasakul, Yon
Issaragrisil, Surapol
R01 ES022968/ES/NIEHS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cancer Ther. 2018 Feb;17(2):484-496. doi: 10.1158/1535-7163.MCT-17-0390. Epub 2017 Nov 22.
Author Address: Siriraj Center of Excellence for Stem Cell Research, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
Division of Hematology, Department of Medicine, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
WVU Cancer Institute, West Virginia University, Morgantown, West Virginia.
Siriraj Center of Excellence for Stem Cell Research, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand. surapolsi@gmail.com.
Bangkok Hematology Center, Wattanosoth Hospital, BDMS Center of Excellence for Cancer, Bangkok, Thailand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1684
Author: Luca, V. C., Jude, K. M., Pierce, N. W., Nachury, M. V., Fischer, S. and Garcia, K. C.
Year: 2015
Title: Structural biology. Structural basis for Notch1 engagement of Delta-like 4
Journal: Science
Volume: 347
Issue: 6224
Pages: 847-53
Epub Date: 2015/02/24
Date: Feb 20
Short Title: Structural biology. Structural basis for Notch1 engagement of Delta-like 4
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1261093
PMCID: PMC4445638
NIHMSID: NIHMS691940
Accession Number: 25700513
Keywords: Alagille Syndrome/genetics
Amino Acid Sequence
Amino Acid Substitution
Animals
Cell Line
Conserved Sequence
Crystallography, X-Ray
Fucose/chemistry
Glucose/chemistry
Glycosylation
Intracellular Signaling Peptides and Proteins/*chemistry/genetics
Ligands
Membrane Proteins/*chemistry/genetics/ultrastructure
Molecular Sequence Data
Molecular Targeted Therapy
Polysaccharides/chemistry
Precursor T-Cell Lymphoblastic Leukemia-Lymphoma/drug therapy/genetics
Protein Binding
Protein Structure, Tertiary
Rats
Receptor, Notch1/*chemistry/genetics/ultrastructure
Serine/chemistry/genetics
Threonine/chemistry/genetics
Abstract: Notch receptors guide mammalian cell fate decisions by engaging the proteins Jagged and Delta-like (DLL). The 2.3 angstrom resolution crystal structure of the interacting regions of the Notch1-DLL4 complex reveals a two-site, antiparallel binding orientation assisted by Notch1 O-linked glycosylation. Notch1 epidermal growth factor-like repeats 11 and 12 interact with the DLL4 Delta/Serrate/Lag-2 (DSL) domain and module at the N-terminus of Notch ligands (MNNL) domains, respectively. Threonine and serine residues on Notch1 are functionalized with O-fucose and O-glucose, which act as surrogate amino acids by making specific, and essential, contacts to residues on DLL4. The elucidation of a direct chemical role for O-glycans in Notch1 ligand engagement demonstrates how, by relying on posttranslational modifications of their ligand binding sites, Notch proteins have linked their functional capacity to developmentally regulated biosynthetic pathways.
Notes: 1095-9203
Luca, Vincent C
Jude, Kevin M
Pierce, Nathan W
Nachury, Maxence V
Fischer, Suzanne
Garcia, K Christopher
Howard Hughes Medical Institute/United States
R01 GM089933/GM/NIGMS NIH HHS/United States
R01 GM097015/GM/NIGMS NIH HHS/United States
1R01-GM097015/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Science. 2015 Feb 20;347(6224):847-53. doi: 10.1126/science.1261093.
Author Address: Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA 94305, USA. Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305, USA. Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA 94305, USA. Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305, USA. Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305, USA. kcgarcia@stanford.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1761
Author: Lucido, M. J., Orlando, B. J., Vecchio, A. J. and Malkowski, M. G.
Year: 2016
Title: Crystal Structure of Aspirin-Acetylated Human Cyclooxygenase-2: Insight into the Formation of Products with Reversed Stereochemistry
Journal: Biochemistry
Volume: 55
Issue: 8
Pages: 1226-38
Epub Date: 2016/02/10
Date: Mar 1
Short Title: Crystal Structure of Aspirin-Acetylated Human Cyclooxygenase-2: Insight into the Formation of Products with Reversed Stereochemistry
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/acs.biochem.5b01378
PMCID: PMC4775376
NIHMSID: NIHMS761856
Accession Number: 26859324
Keywords: Acetylation/*drug effects
Animals
Anti-Inflammatory Agents, Non-Steroidal/*chemistry/pharmacology
Aspirin/*chemistry/pharmacology
Crystallography, X-Ray
Cyclooxygenase 2/*chemistry/metabolism
Humans
Mice
Molecular Docking Simulation
Protein Conformation
Abstract: Aspirin and other nonsteroidal anti-inflammatory drugs target the cyclooxygenase enzymes (COX-1 and COX-2) to block the formation of prostaglandins. Aspirin is unique in that it covalently modifies each enzyme by acetylating Ser-530 within the cyclooxygenase active site. Acetylation of COX-1 leads to complete loss of activity, while acetylation of COX-2 results in the generation of the monooxygenated product 15(R)-hydroxyeicosatetraenoic acid (15R-HETE). Ser-530 has also been shown to influence the stereochemistry for the addition of oxygen to the prostaglandin product. We determined the crystal structures of S530T murine (mu) COX-2, aspirin-acetylated human (hu) COX-2, and huCOX-2 in complex with salicylate to 1.9, 2.0, and 2.4 Å, respectively. The structures reveal that (1) the acetylated Ser-530 completely blocks access to the hydrophobic groove, (2) the observed binding pose of salicylate is reflective of the enzyme-inhibitor complex prior to acetylation, and (3) the observed Thr-530 rotamer in the S530T muCOX-2 crystal structure does not impede access to the hydrophobic groove. On the basis of these structural observations, along with functional analysis of the S530T/G533V double mutant, we propose a working hypothesis for the generation of 15R-HETE by aspirin-acetylated COX-2. We also observe differential acetylation of COX-2 purified in various detergent systems and nanodiscs, indicating that detergent and lipid binding within the membrane-binding domain of the enzyme alters the rate of the acetylation reaction in vitro.
Notes: 1520-4995
Lucido, Michael J
Orlando, Benjamin J
Vecchio, Alex J
Malkowski, Michael G
R01 GM077176/GM/NIGMS NIH HHS/United States
P41 GM103485/GM/NIGMS NIH HHS/United States
P41 RR001646/RR/NCRR NIH HHS/United States
DMR0225180/GM/NIGMS NIH HHS/United States
R01 GM115386/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Biochemistry. 2016 Mar 1;55(8):1226-38. doi: 10.1021/acs.biochem.5b01378. Epub 2016 Feb 19.
Author Address: Department of Structural Biology, The State University of New York at Buffalo and Hauptman-Woodward Medical Research Institute , Buffalo, New York 14203, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 627
Author: Lund, J., Takahashi, N., Pound, J. D., Goodall, M. and Jefferis, R.
Year: 1996
Title: Multiple interactions of IgG with its core oligosaccharide can modulate recognition by complement and human Fc gamma receptor I and influence the synthesis of its oligosaccharide chains
Journal: J Immunol
Volume: 157
Issue: 11
Pages: 4963-9
Epub Date: 1996/12/01
Date: Dec 1
Short Title: Multiple interactions of IgG with its core oligosaccharide can modulate recognition by complement and human Fc gamma receptor I and influence the synthesis of its oligosaccharide chains
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
Accession Number: 8943402
Keywords: Animals
Binding Sites
Carbohydrate Sequence
Cell Line
Complement System Proteins/*metabolism
Cricetinae
Glycosylation
Humans
Immunoglobulin G/*chemistry/genetics/*metabolism
In Vitro Techniques
Mice
Models, Molecular
Molecular Sequence Data
Molecular Structure
Oligosaccharides/biosynthesis/chemistry/*metabolism
Protein Conformation
Protein Engineering
Receptors, IgG/*metabolism
Recombinant Fusion Proteins/chemistry/genetics/metabolism
Abstract: Glycosylation at Asn297 within the CH2 domains of IgG is important for recognition by the effector ligands Fc gammaR and C. Protein engineering has been used to replace amino acid residues within the extensive oligosaccharide interaction site that contact the core hexasaccharide (GlcNAc2Man3GlcNAc). Replacement of residues Phe241, Val264, or Asp265, in particular, results in reduced recognition of human chimeric anti-nitroiodophenacetyl IgG3 produced in Chinese hamster ovary cells, by guinea pig C and human C1q. Replacement of residues Val264 or Asp265, in particular, results in reduced superoxide production triggered through human Fc gammaRI expressed on U937 cells. These results suggest that noncovalent interactions of multiple amino acid residues of IgG with oligosaccharide residues that include the primary and secondary GlcNAc are necessary for optimal recognition of IgG by human Fc gammaRI and C1q. Replacement of residues 241, 243, 264, 265, or 301 with alanine in each case resulted in increased galactosylation and sialylation relative to the wild-type oligosaccharide chains. In particular, for the mutant FA243 there was much increased sialylation of its oligosaccharide chains (73%) relative to the wild-type (4%). Thus, even single residue replacements within the oligosaccharide interaction site of the C region can influence galactosylation and sialylation of its oligosaccharide chains. These data suggest a protein engineering route to the production of more homogeneously glycosylated IgG molecules with or without compromised biologic activities.
Notes: Lund, J
Takahashi, N
Pound, J D
Goodall, M
Jefferis, R
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 1996 Dec 1;157(11):4963-9.
Author Address: Department of Immunology, The Medical School, Birmingham, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1810
Author: Lund, P. J., Elias, J. E. and Davis, M. M.
Year: 2016
Title: Global Analysis of O-GlcNAc Glycoproteins in Activated Human T Cells
Journal: J Immunol
Volume: 197
Issue: 8
Pages: 3086-3098
Epub Date: 2016/09/23
Date: Oct 15
Short Title: Global Analysis of O-GlcNAc Glycoproteins in Activated Human T Cells
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
DOI: 10.4049/jimmunol.1502031
PMCID: PMC5055199
Accession Number: 27655845
Keywords: Acetylglucosamine/metabolism
Animals
Cell Line
Glycoproteins/metabolism
Glycosylation
Humans
Lymphocyte Activation
Mice
N-Acetylglucosaminyltransferases/*metabolism
Protein Isoforms/metabolism
Protein Processing, Post-Translational
Proteomics
Receptors, Antigen, T-Cell/metabolism
Signal Transduction
T-Lymphocytes/*metabolism
Abstract: T cell activation in response to Ag is largely regulated by protein posttranslational modifications. Although phosphorylation has been extensively characterized in T cells, much less is known about the glycosylation of serine/threonine residues by O-linked N-acetylglucosamine (O-GlcNAc). Given that O-GlcNAc appears to regulate cell signaling pathways and protein activity similarly to phosphorylation, we performed a comprehensive analysis of O-GlcNAc during T cell activation to address the functional importance of this modification and to identify the modified proteins. Activation of T cells through the TCR resulted in a global elevation of O-GlcNAc levels and in the absence of O-GlcNAc, IL-2 production and proliferation were compromised. T cell activation also led to changes in the relative expression of O-GlcNAc transferase (OGT) isoforms and accumulation of OGT at the immunological synapse of murine T cells. Using a glycoproteomics approach, we identified >200 O-GlcNAc proteins in human T cells. Many of the identified proteins had a functional relationship to RNA metabolism, and consistent with a connection between O-GlcNAc and RNA, inhibition of OGT impaired nascent RNA synthesis upon T cell activation. Overall, our studies provide a global analysis of O-GlcNAc dynamics during T cell activation and the first characterization, to our knowledge, of the O-GlcNAc glycoproteome in human T cells.
Notes: 1550-6606
Lund, Peder J
Orcid: 0000-0002-2146-8724
Elias, Joshua E
Orcid: 0000-0002-8063-3259
Davis, Mark M
P30 NS069375/NS/NINDS NIH HHS/United States
R01 AI022511/AI/NIAID NIH HHS/United States
U19 AI057229/AI/NIAID NIH HHS/United States
U19 AI090019/AI/NIAID NIH HHS/United States
HHMI/Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Immunol. 2016 Oct 15;197(8):3086-3098. doi: 10.4049/jimmunol.1502031. Epub 2016 Sep 21.
Author Address: Interdepartmental Program in Immunology, Stanford University, Stanford, CA 94305.
Department of Microbiology and Immunology, Stanford University, Stanford, CA 94305.
Department of Chemical and Systems Biology, Stanford University, Stanford, CA 94305.
Department of Microbiology and Immunology, Stanford University, Stanford, CA 94305; mmdavis@stanford.edu.
Stanford Institute for Immunity, Transplantation, and Infection, Stanford University, Stanford, CA 94305; and.
Howard Hughes Medical Institute, Stanford University, Stanford, CA 94305.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1480
Author: Luo, J., Obmolova, G., Malia, T. J., Wu, S. J., Duffy, K. E., Marion, J. D., Bell, J. K., Ge, P., Zhou, Z. H., Teplyakov, A., Zhao, Y., Lamb, R. J., Jordan, J. L., San Mateo, L. R., Sweet, R. W. and Gilliland, G. L.
Year: 2012
Title: Lateral clustering of TLR3:dsRNA signaling units revealed by TLR3ecd:3Fabs quaternary structure
Journal: J Mol Biol
Volume: 421
Issue: 1
Pages: 112-24
Epub Date: 2012/05/15
Date: Aug 3
Short Title: Lateral clustering of TLR3:dsRNA signaling units revealed by TLR3ecd:3Fabs quaternary structure
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2012.05.006
PMCID: PMC3920545
NIHMSID: NIHMS551209
Accession Number: 22579623
Keywords: Antibody Affinity
Binding Sites
Cell Line
Humans
Immunoglobulin Fab Fragments/chemistry/immunology/metabolism
Models, Molecular
Mutation
Protein Structure, Tertiary
RNA, Double-Stranded/*metabolism
Signal Transduction
Toll-Like Receptor 3/*chemistry/genetics/*metabolism
Abstract: Toll-like receptor 3 (TLR3) recognizes dsRNA and initiates an innate immune response through the formation of a signaling unit (SU) composed of one double-stranded RNA (dsRNA) and two TLR3 molecules. We report the crystal structure of human TLR3 ectodomain (TLR3ecd) in a quaternary complex with three neutralizing Fab fragments. Fab15 binds an epitope that overlaps the C-terminal dsRNA binding site and, in biochemical assays, blocks the interaction of TLR3ecd with dsRNA, thus directly antagonizing TLR3 signaling through inhibition of SU formation. In contrast, Fab12 and Fab1068 bind TLR3ecd at sites distinct from the N- and C-terminal regions that interact with dsRNA and do not inhibit minimal SU formation with short dsRNA. Molecular modeling based on the co-structure rationalizes these observations by showing that both Fab12 and Fab1068 prevent lateral clustering of SUs along the length of the dsRNA ligand. This model is further supported by cell-based assay results using dsRNA ligands of lengths that support single and multiple SUs. Thus, their antagonism of TLR3 signaling indicates that lateral clustering of SUs is required for TLR3 signal transduction.
Notes: 1089-8638
Luo, Jinquan
Obmolova, Galina
Malia, Thomas J
Wu, Sheng-Jiun
Duffy, Karen E
Marion, James D
Bell, Jessica K
Ge, Peng
Zhou, Z Hong
Teplyakov, Alexey
Zhao, Yonghong
Lamb, Roberta J
Jordan, Jarrat L
San Mateo, Lani R
Sweet, Raymond W
Gilliland, Gary L
K22 CA122828/CA/NCI NIH HHS/United States
P30 CA016059/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Mol Biol. 2012 Aug 3;421(1):112-24. doi: 10.1016/j.jmb.2012.05.006. Epub 2012 May 9.
Author Address: Biologics Research, Janssen Research and Development, L.L.C., 145 King of Prussia Road, Radnor, PA 19087, USA. jluo@its.jnj.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1298
Author: Lupardus, P. J., Birnbaum, M. E. and Garcia, K. C.
Year: 2010
Title: Molecular basis for shared cytokine recognition revealed in the structure of an unusually high affinity complex between IL-13 and IL-13Ralpha2
Journal: Structure
Volume: 18
Issue: 3
Pages: 332-42
Epub Date: 2010/03/13
Date: Mar 10
Short Title: Molecular basis for shared cytokine recognition revealed in the structure of an unusually high affinity complex between IL-13 and IL-13Ralpha2
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2010.01.003
PMCID: PMC2850121
NIHMSID: NIHMS188925
Accession Number: 20223216
Keywords: Alanine/genetics
Binding Sites
Crystallography, X-Ray
Cytokines/*metabolism
Humans
Interleukin-13/*chemistry/metabolism
Interleukin-13 Receptor alpha2 Subunit/*chemistry/metabolism
Kinetics
Models, Molecular
Protein Conformation
Signal Transduction
Abstract: Interleukin-13 is a cytokine important for development of T helper cell type 2 (Th2) responses and plays a critical role in asthma and allergy. The IL-13 Receptor alpha2 (IL-13Ralpha2) is a receptor for IL-13 lacking canonical Jak/STAT signaling functions. Here we present the crystal structure along with a mutational and biophysical analysis of the IL-13/IL-13Ralpha2 complex. While retaining a similar mode of IL-13 binding to its related signaling receptor, IL-13Ralpha1, IL-13Ralpha2 uses peripheral receptor residues unused in the IL-13/IL-13Ralpha1 complex to generate a larger and more complementary interface for IL-13. This results in a four orders of magnitude increase in affinity, to the femtomolar level, compared to IL-13Ralpha1. Alanine scanning mutagenesis of the IL-13 interface reveals several common "hotspot" residues important for binding to both receptors, but also identifies a prominent IL-13Ralpha2-specific contact. These results provide a framework for development of receptor subtype-selective IL-13 antagonists and indicate a decoy function for IL-13Ralpha2.
Notes: 1878-4186
Lupardus, Patrick J
Birnbaum, Michael E
Garcia, K Christopher
R01 AI051321-05/AI/NIAID NIH HHS/United States
R37 AI051321/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
AI51321/AI/NIAID NIH HHS/United States
R01 AI051321/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Structure. 2010 Mar 10;18(3):332-42. doi: 10.1016/j.str.2010.01.003.
Author Address: Howard Hughes Medical Institute, Department of Molecular and Cellular Physiology, Department of Structural Biology, and Program in Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1997
Author: Ma, J., Wang, W. H., Li, Z., Shabanowitz, J., Hunt, D. F. and Hart, G. W.
Year: 2019
Title: O-GlcNAc Site Mapping by Using a Combination of Chemoenzymatic Labeling, Copper-Free Click Chemistry, Reductive Cleavage, and Electron-Transfer Dissociation Mass Spectrometry
Journal: Anal Chem
Volume: 91
Issue: 4
Pages: 2620-2625
Epub Date: 2019/01/19
Date: Feb 19
Short Title: O-GlcNAc Site Mapping by Using a Combination of Chemoenzymatic Labeling, Copper-Free Click Chemistry, Reductive Cleavage, and Electron-Transfer Dissociation Mass Spectrometry
Alternate Journal: Analytical chemistry
ISSN: 0003-2700 (Print)
0003-2700
DOI: 10.1021/acs.analchem.8b05688
PMCID: PMC6756848
NIHMSID: NIHMS1051119 a share of royalty received by the university on sales of the CTD 110.6 antibody, which are managed by the Johns Hopkins University.
Accession Number: 30657688
Keywords: Acetylglucosamine/*analysis/chemistry
Adaptor Proteins, Signal Transducing/*chemistry/metabolism
Alkynes/chemistry
Azides/chemistry
Click Chemistry
Cycloaddition Reaction
Glycosylation
HEK293 Cells
Humans
Oxidation-Reduction
Peptides/*chemistry/metabolism
Protein Processing, Post-Translational
Tandem Mass Spectrometry/*methods
Uridine Diphosphate N-Acetylgalactosamine/analogs & derivatives/chemistry
alpha-Crystallins/*chemistry/metabolism
Abstract: As a dynamic post-translational modification, O-linked β- N-acetylglucosamine ( O-GlcNAc) modification (i.e., O-GlcNAcylation) of proteins regulates many biological processes involving cellular metabolism and signaling. However, O-GlcNAc site mapping, a prerequisite for site-specific functional characterization, has been a challenge since its discovery. Herein we present a novel method for O-GlcNAc enrichment and site mapping. In this method, the O-GlcNAc moiety on peptides was labeled with UDP-GalNAz followed by copper-free azide-alkyne cycloaddition with a multifunctional reagent bearing a terminal cyclooctyne, a disulfide bridge, and a biotin handle. The tagged peptides were then released from NeutrAvidin beads upon reductant treatment, alkylated with (3-acrylamidopropyl)trimethylammonium chloride, and subjected to electron-transfer dissociation mass spectrometry analysis. After validation by using standard synthetic peptide gCTD and model protein α-crystallin, such an approach was applied to the site mapping of overexpressed TGF-β-activated kinase 1/MAP3K7 binding protein 2 (TAB2), with four O-GlcNAc sites unambiguously identified. Our method provides a promising tool for the site-specific characterization of O-GlcNAcylation of important proteins.
Notes: 1520-6882
Ma, Junfeng
Orcid: 0000-0002-5183-5425
Wang, Wei-Han
Li, Zengxia
Shabanowitz, Jeffrey
Hunt, Donald F
Hart, Gerald W
P01 HL107153/HL/NHLBI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
R01 GM037537/GM/NIGMS NIH HHS/United States
R01 GM116891/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Anal Chem. 2019 Feb 19;91(4):2620-2625. doi: 10.1021/acs.analchem.8b05688. Epub 2019 Feb 4.
Author Address: Department of Biological Chemistry , The Johns Hopkins University School of Medicine , Baltimore , Maryland 21205 , United States.
Department of Chemistry , University of Virginia , Charlottesville , Virginia 22904 , United States.
Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences , Fudan University , Shanghai 200032 , China.
Department of Pathology, Health Sciences Center , University of Virginia , Charlottesville , Virginia 22908 , United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 902
Author: Ma, S., Hill, K. E., Burk, R. F. and Caprioli, R. M.
Year: 2003
Title: Mass spectrometric identification of N- and O-glycosylation sites of full-length rat selenoprotein P and determination of selenide-sulfide and disulfide linkages in the shortest isoform
Journal: Biochemistry
Volume: 42
Issue: 32
Pages: 9703-11
Epub Date: 2003/08/13
Date: Aug 19
Short Title: Mass spectrometric identification of N- and O-glycosylation sites of full-length rat selenoprotein P and determination of selenide-sulfide and disulfide linkages in the shortest isoform
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi0346300
Accession Number: 12911312
Keywords: Amino Acid Sequence
Animals
Binding Sites
Carbohydrate Sequence
Disulfides/*chemistry
Glycosylation
Molecular Sequence Data
Peptide Fragments/analysis
Peptide Mapping
Protein Isoforms
Proteins/*chemistry/genetics/metabolism
Rats
Selenium/*chemistry
Selenocysteine/chemistry
Selenoprotein P
Selenoproteins
Spectrometry, Mass, Electrospray Ionization/methods
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
Sulfides/*chemistry
Abstract: Rat selenoprotein P is an extracellular glycoprotein of 366 amino acid residues that is rich in cysteine and selenocysteine. Plasma contains four isoforms that differ principally by length at the C-terminal end. Mass spectrometry was used to identify sites of glycosylation on the full-length protein. Of the potential N-glycosylation sites, three located at residues 64, 155, and 169 were occupied, while the two at residues 351 and 356 were not occupied. Threonine 346 was variably O-glycosylated. Thus, full-length selenoprotein P is both N- and O-glycosylated. The shortest isoform of selenoprotein P, which terminates at residue 244, was analyzed for selenide-sulfide and disulfide linkages. In this isoform, a single selenocysteine and seven cysteines are present. Mass spectrometric analysis indicated that a selenide-sulfide bond exists between Sec40 and Cys43. Two disulfides were also detected as Cys149-Cys167 and Cys153-Cys156. The finding of a selenide-sulfide bond in the shortest isoform is compatible with a redox function of this pair that might be analogous to the selenol-thiol pair near the C terminus of animal thioredoxin reductase. The disulfide formed by Cys153-Cys156 also has some characteristics of a redox active pair.
Notes: Ma, Shuguang
Hill, Kristina E
Burk, Raymond F
Caprioli, Richard M
DK26657/DK/NIDDK NIH HHS/United States
ES00267/ES/NIEHS NIH HHS/United States
ES02497/ES/NIEHS NIH HHS/United States
GM58008/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2003 Aug 19;42(32):9703-11. doi: 10.1021/bi0346300.
Author Address: Mass Spectrometry Research Center, Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1896
Author: Ma, X., Liu, H., Li, J., Wang, Y., Ding, Y. H., Shen, H., Yang, Y., Sun, C., Huang, M., Tu, Y., Liu, Y., Zhao, Y., Dong, M. Q., Xu, P., Tang, T. S. and Guo, C.
Year: 2017
Title: Polη O-GlcNAcylation governs genome integrity during translesion DNA synthesis
Journal: Nat Commun
Volume: 8
Issue: 1
Pages: 1941
Epub Date: 2017/12/07
Date: Dec 5
Short Title: Polη O-GlcNAcylation governs genome integrity during translesion DNA synthesis
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-017-02164-1
PMCID: PMC5717138
Accession Number: 29208956
Keywords: Antineoplastic Agents/pharmacology
Cell Line, Tumor
Cisplatin/pharmacology
DNA/*biosynthesis
*DNA Damage
*DNA Repair
DNA Replication
DNA-Directed DNA Polymerase/*metabolism
HEK293 Cells
Humans
Mutagenesis
N-Acetylglucosaminyltransferases/*metabolism
Polyubiquitin
Protein Processing, Post-Translational
Pyrimidine Dimers
Ubiquitination
Ultraviolet Rays
Abstract: DNA polymerase η (Polη) facilitates translesion DNA synthesis (TLS) across ultraviolet (UV) irradiation- and cisplatin-induced DNA lesions implicated in skin carcinogenesis and chemoresistant phenotype formation, respectively. However, whether post-translational modifications of Polη are involved in these processes remains largely unknown. Here, we reported that human Polη undergoes O-GlcNAcylation at threonine 457 by O-GlcNAc transferase upon DNA damage. Abrogation of this modification results in a reduced level of CRL4(CDT2)-dependent Polη polyubiquitination at lysine 462, a delayed p97-dependent removal of Polη from replication forks, and significantly enhanced UV-induced mutagenesis even though Polη focus formation and its efficacy to bypass across cyclobutane pyrimidine dimers after UV irradiation are not affected. Furthermore, the O-GlcNAc-deficient T457A mutation impairs TLS to bypass across cisplatin-induced lesions, causing increased cellular sensitivity to cisplatin. Our findings demonstrate a novel role of Polη O-GlcNAcylation in TLS regulation and genome stability maintenance and establish a new rationale to improve chemotherapeutic treatment.
Notes: 2041-1723
Ma, Xiaolu
Liu, Hongmei
Li, Jing
Orcid: 0000-0002-3977-1641
Wang, Yihao
Ding, Yue-He
Shen, Hongyan
Orcid: 0000-0002-3592-6220
Yang, Yeran
Sun, Chenyi
Huang, Min
Tu, Yingfeng
Liu, Yang
Zhao, Yongliang
Dong, Meng-Qiu
Orcid: 0000-0002-6094-1182
Xu, Ping
Tang, Tie-Shan
Guo, Caixia
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2017 Dec 5;8(1):1941. doi: 10.1038/s41467-017-02164-1.
Author Address: CAS Key Laboratory of Genomics and Precision Medicine, Beijing Institute of Genomics, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Beijing, 100101, China.
State Key Laboratory of Membrane Biology, Institute of Zoology, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Beijing, 100101, China.
Beijing Key Laboratory of DNA Damage Response, College of Life Sciences, Capital Normal University, Beijing, 100048, China.
State Key Laboratory of Proteomics National Center for Protein Sciences Beijing, Beijing Proteome Research Center, National Engineering Research Center for Protein Drugs, Beijing Institute of Radiation Medicine, Beijing, 102206, China.
National Institute of Biological Sciences (Beijing), Beijing, 102206, China.
State Key Laboratory of Membrane Biology, Institute of Zoology, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Beijing, 100101, China. tangtsh@ioz.ac.cn.
CAS Key Laboratory of Genomics and Precision Medicine, Beijing Institute of Genomics, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Beijing, 100101, China. guocx@big.ac.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1847
Author: Ma, Z., Chalkley, R. J. and Vosseller, K.
Year: 2017
Title: Hyper-O-GlcNAcylation activates nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) signaling through interplay with phosphorylation and acetylation
Journal: J Biol Chem
Volume: 292
Issue: 22
Pages: 9150-9163
Epub Date: 2017/04/19
Date: Jun 2
Short Title: Hyper-O-GlcNAcylation activates nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) signaling through interplay with phosphorylation and acetylation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.766568
PMCID: PMC5454098
Accession Number: 28416608
Keywords: Acetylation
Animals
HEK293 Cells
Humans
I-kappa B Kinase/genetics/metabolism
Mice
Phosphorylation/physiology
Signal Transduction/*physiology
Transcription Factor RelA/genetics/*metabolism
p300-CBP Transcription Factors/genetics/metabolism
*E1A-binding protein p300 (p300)
*NF-kappaB
*O-GlcNAcylation
*acetylation
*phosphorylation
this article.
Abstract: O-GlcNAcylation is the covalent addition of an O-linked β-N-acetylglucosamine (O-GlcNAc) sugar moiety to hydroxyl groups of serine/threonine residues of cytosolic and nuclear proteins. O-GlcNAcylation, analogous to phosphorylation, plays critical roles in gene expression through direct modification of transcription factors, such as NF-κB. Aberrantly increased NF-κB O-GlcNAcylation has been linked to NF-κB constitutive activation and cancer development. Therefore, it is of a great biological and clinical significance to dissect the molecular mechanisms that tune NF-κB activity. Recently, we and others have shown that O-GlcNAcylation affects the phosphorylation and acetylation of NF-κB subunit p65/RelA. However, the mechanism of how O-GlcNAcylation activates NF-κB signaling through phosphorylation and acetylation is not fully understood. In this study, we mapped O-GlcNAcylation sites of p65 at Thr-305, Ser-319, Ser-337, Thr-352, and Ser-374. O-GlcNAcylation of p65 at Thr-305 and Ser-319 increased CREB-binding protein (CBP)/p300-dependent activating acetylation of p65 at Lys-310, contributing to NF-κB transcriptional activation. Moreover, elevation of O-GlcNAcylation by overexpression of OGT increased the expression of p300, IKKα, and IKKβ and promoted IKK-mediated activating phosphorylation of p65 at Ser-536, contributing to NF-κB activation. In addition, we also identified phosphorylation of p65 at Thr-308, which might impair the O-GlcNAcylation of p65 at Thr-305. These results indicate mechanisms through which both non-pathological and oncogenic O-GlcNAcylation regulate NF-κB signaling through interplay with phosphorylation and acetylation.
Notes: 1083-351x
Ma, Zhiyuan
Chalkley, Robert J
Vosseller, Keith
Journal Article
J Biol Chem. 2017 Jun 2;292(22):9150-9163. doi: 10.1074/jbc.M116.766568. Epub 2017 Apr 17.
Author Address: From the Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102 and zhiyuan.ma@dm.duke.edu.
the Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94158.
From the Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102 and kvosseller@gmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1553
Author: Ma, Z., Vocadlo, D. J. and Vosseller, K.
Year: 2013
Title: Hyper-O-GlcNAcylation is anti-apoptotic and maintains constitutive NF-κB activity in pancreatic cancer cells
Journal: J Biol Chem
Volume: 288
Issue: 21
Pages: 15121-30
Epub Date: 2013/04/18
Date: May 24
Short Title: Hyper-O-GlcNAcylation is anti-apoptotic and maintains constitutive NF-κB activity in pancreatic cancer cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.470047
PMCID: PMC3663532
Accession Number: 23592772
Keywords: Active Transport, Cell Nucleus/genetics
Acylation/genetics
Amino Acid Substitution
Animals
*Apoptosis
Carcinoma, Pancreatic Ductal/genetics/*metabolism/pathology
Cell Line, Transformed
Cell Nucleus/genetics/*metabolism/pathology
Cell Proliferation
*Gene Expression Regulation, Neoplastic
Humans
Mice
Mutation, Missense
Neoplasm Proteins/genetics/*metabolism
Pancreatic Neoplasms/genetics/*metabolism/pathology
Transcription Factor RelA/genetics/*metabolism
Transcription, Genetic/genetics
Uridine Diphosphate N-Acetylgalactosamine/genetics/*metabolism
Apoptosis
NF-kappa B (NF-κB)
O-GlcNAc
O-GlcNAcylation
Pancreatic Cancer
Warburg Effect
Abstract: Cancer cell metabolic reprogramming includes a shift in energy production from oxidative phosphorylation to less efficient glycolysis even in the presence of oxygen (Warburg effect) and use of glutamine for increased biosynthetic needs. This necessitates greatly increased glucose and glutamine uptake, both of which enter the hexosamine biosynthetic pathway (HBP). The HBP end product UDP-N-acetylglucosamine (UDP-GlcNAc) is used in enzymatic post-translational modification of many cytosolic and nuclear proteins by O-linked β-N-acetylglucosamine (O-GlcNAc). Here, we observed increased HBP flux and hyper-O-GlcNAcylation in human pancreatic ductal adenocarcinoma (PDAC). PDAC hyper-O-GlcNAcylation was associated with elevation of OGT and reduction of the enzyme that removes O-GlcNAc (OGA). Reducing hyper-O-GlcNAcylation had no effect on non-transformed pancreatic epithelial cell growth, but inhibited PDAC cell proliferation, anchorage-independent growth, orthotopic tumor growth, and triggered apoptosis. PDAC is supported by oncogenic NF-κB transcriptional activity. The NF-κB p65 subunit and upstream kinases IKKα/IKKβ were O-GlcNAcylated in PDAC. Reducing hyper-O-GlcNAcylation decreased PDAC cell p65 activating phosphorylation (S536), nuclear translocation, NF-κB transcriptional activity, and target gene expression. Conversely, mimicking PDAC hyper-O-GlcNAcylation through pharmacological inhibition of OGA suppressed suspension culture-induced apoptosis and increased IKKα and p65 O-GlcNAcylation, accompanied by activation of NF-κB signaling. Finally, reducing p65 O-GlcNAcylation specifically by mutating two p65 O-GlcNAc sites (T322A and T352A) attenuated the induction of PDAC cell anchorage-independent growth. Our data indicate that hyper-O-GlcNAcylation is anti-apoptotic and contributes to NF-κB oncogenic activation in PDAC.
Notes: 1083-351x
Ma, Zhiyuan
Vocadlo, David J
Vosseller, Keith
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2013 May 24;288(21):15121-30. doi: 10.1074/jbc.M113.470047. Epub 2013 Apr 16.
Author Address: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 374
Author: Maeda, H., Hashimoto, R. K., Ogura, T., Hiraga, S. and Uzawa, H.
Year: 1987
Title: Molecular cloning of a human apoC-III variant: Thr 74----Ala 74 mutation prevents O-glycosylation
Journal: J Lipid Res
Volume: 28
Issue: 12
Pages: 1405-9
Epub Date: 1987/12/01
Date: Dec
Short Title: Molecular cloning of a human apoC-III variant: Thr 74----Ala 74 mutation prevents O-glycosylation
Alternate Journal: Journal of lipid research
ISSN: 0022-2275 (Print)
0022-2275
Accession Number: 3123586
Keywords: Alanine
Amino Acid Sequence
Apolipoprotein C-III
Apolipoproteins C/*genetics
Base Sequence
*Cloning, Molecular
DNA/analysis
Glycosylation
Humans
Lipoproteins, VLDL/blood
Molecular Sequence Data
Mutation
Threonine
Abstract: Apolipoprotein C-III (apoC-III) is a major protein of very low density lipoprotein (VLDL). The apoC-III polypeptide contains a carbohydrate chain containing galactosamine, galactose, and sialic acid attached in O-linkage to a threonine residue at position 74. We have cloned the apoC-III gene from a subject whose serum contained unusually high amounts of apoC-III lacking the carbohydrate moiety (C-III-0). DNA sequence analysis of the cloned gene revealed a single nucleotide substitution (A----G) that encodes an alanine at position 74 instead of the normal threonine. As a result of this amino acid replacement, the mutant apoC-III polypeptide is not glycosylated. The mutation in the apoC-III gene creates a novel AluI site that permits diagnosis of the change by Southern blotting of genomic DNA.
Notes: Maeda, H
Hashimoto, R K
Ogura, T
Hiraga, S
Uzawa, H
Journal Article
Research Support, Non-U.S. Gov't
United States
J Lipid Res. 1987 Dec;28(12):1405-9.
Author Address: Department of Molecular Genetics, Kumamoto University Medical School, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1406
Author: Maher, G. J., Hilton, E. N., Urquhart, J. E., Davidson, A. E., Spencer, H. L., Black, G. C. and Manson, F. D.
Year: 2011
Title: The cataract-associated protein TMEM114, and TMEM235, are glycosylated transmembrane proteins that are distinct from claudin family members
Journal: FEBS Lett
Volume: 585
Issue: 14
Pages: 2187-92
Epub Date: 2011/06/22
Date: Jul 21
Short Title: The cataract-associated protein TMEM114, and TMEM235, are glycosylated transmembrane proteins that are distinct from claudin family members
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1016/j.febslet.2011.05.060
Accession Number: 21689651
Keywords: Amino Acid Sequence
Animals
Calcium Channels/genetics/metabolism
Cataract/genetics
Cell Line
Claudins/genetics/*metabolism
Embryo, Nonmammalian/anatomy & histology/physiology
Eye/embryology/growth & development/metabolism/pathology
Humans
Membrane Glycoproteins/classification/*genetics/*metabolism
Membrane Proteins/classification/*genetics/*metabolism
Molecular Sequence Data
Phylogeny
Sequence Alignment
Xenopus
Abstract: A novel gene, TMEM114, was annotated as a member of the claudin gene family and was subsequently associated as a cause of autosomal dominant cataract because of a translocation in its putative promoter. Our bioinformatic and molecular analyses of TMEM114, and the closely related TMEM235, demonstrate that these proteins are more closely related to members of the voltage dependent calcium channel gamma subunit family. TMEM114 and TMEM235 differed from claudins in terms of localisation in polarised epithelial cells and by the presence of N-linked glycans. By gene expression knockdown in Xenopus tropicalis we also demonstrate a role for Tmem114 in eye development.
Notes: 1873-3468
Maher, Geoffrey J
Hilton, Emma N
Urquhart, Jill E
Davidson, Alice E
Spencer, Helen L
Black, Graeme C
Manson, Forbes D
Biotechnology and Biological Sciences Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2011 Jul 21;585(14):2187-92. doi: 10.1016/j.febslet.2011.05.060. Epub 2011 Jun 16.
Author Address: School of Biomedicine, The University of Manchester, Central Manchester University Hospitals NHS Foundation Trust, Manchester, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1787
Author: Mahrhold, S., Bergström, T., Stern, D., Dorner, B. G., Åstot, C. and Rummel, A.
Year: 2016
Title: Only the complex N559-glycan in the synaptic vesicle glycoprotein 2C mediates high affinity binding to botulinum neurotoxin serotype A1
Journal: Biochem J
Volume: 473
Issue: 17
Pages: 2645-54
Epub Date: 2016/06/18
Date: Sep 1
Short Title: Only the complex N559-glycan in the synaptic vesicle glycoprotein 2C mediates high affinity binding to botulinum neurotoxin serotype A1
Alternate Journal: The Biochemical journal
ISSN: 0264-6021
DOI: 10.1042/bcj20160439
Accession Number: 27313224
Keywords: Azacitidine/*analogs & derivatives/pharmacology
Botulinum Toxins, Type A/*metabolism
Decitabine
Glycosylation
Membrane Glycoproteins/*metabolism
Protein Binding
Surface Plasmon Resonance
Synaptic Vesicles/*metabolism
*HEK cell expression
*Lc–ms/ms
*N-glycan
*botulinum neurotoxin A
*surface plasmon resonance
*synaptic vesicle glycoprotein 2C
Abstract: The extraordinary potency of botulinum neurotoxins (BoNTs) is mediated by their high neurospecificity, targeting peripheral cholinergic motoneurons leading to flaccid paralysis and successive respiratory failure. Complex polysialo gangliosides accumulate BoNTs on the plasma membrane and facilitate subsequent binding to synaptic vesicle membrane proteins which results in toxin endocytosis. The luminal domain 4 (LD4) of the three synaptic vesicle glycoprotein 2 (SV2) isoforms A-C mediates uptake of the clinically most relevant serotype BoNT/A1. SV2C-LD4 exhibits the strongest protein-protein interaction and comprises five putative N-glycosylation sites (PNG sites). Here, we expressed human SV2C-LD4 fused to human IgG-Fc in prokaryotic and eukaryotic expression systems to analyse the effect of N-glycosylation of SV2C on the interaction with BoNT/A1. Mass spectrometric analysis of gSV2CLD-Fc demonstrates glycosylation of N534, N559 and N565, the latter two residing at the BoNT/A interface. Mutational analysis demonstrates that only the N559-glycan, but not N565-glycan increases affinity of BoNT/A for human gSV2C-LD4. The N559-glycan was characterised as a complex core-fucosylated type with a heterogeneity ranging up to tetra-antennary structure with bisecting N-acetylglucosamine which can establish extensive interactions with BoNT/A. The mutant gSV2CLD-Fc N559A displayed a 50-fold increased dissociation rate kd resulting in an overall 12-fold decreased binding affinity in surface plasmon resonance (SPR) experiments. The delayed dissociation might provide BoNT/A more time for endocytosis into synaptic vesicles. In conclusion, we show the importance of the complex N559-glycan of SV2C-LD4, adding a third anchor point beside a ganglioside and the SV2C-LD4 peptide, for BoNT/A neuronal cell surface binding and uptake.
Notes: 1470-8728
Mahrhold, Stefan
Bergström, Tomas
Stern, Daniel
Dorner, Brigitte G
Åstot, Crister
Rummel, Andreas
Journal Article
England
Biochem J. 2016 Sep 1;473(17):2645-54. doi: 10.1042/BCJ20160439. Epub 2016 Jun 16.
Author Address: Institut für Toxikologie, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, 30625 Hannover, Germany.
Division for CBRN Defence and Security, Swedish Defence Research Agency, Cementvägen 20, 90182 Umeå, Sweden.
Biological Toxins, Centre for Biological Threats and Special Pathogens, Robert Koch Institute, Seestr. 10, 13353 Berlin, Germany.
Institut für Toxikologie, Medizinische Hochschule Hannover, Carl-Neuberg-Str. 1, 30625 Hannover, Germany rummel.andreas@mh-hannover.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1577
Author: Maita, N., Tsukimura, T., Taniguchi, T., Saito, S., Ohno, K., Taniguchi, H. and Sakuraba, H.
Year: 2013
Title: Human α-L-iduronidase uses its own N-glycan as a substrate-binding and catalytic module
Journal: Proc Natl Acad Sci U S A
Volume: 110
Issue: 36
Pages: 14628-33
Epub Date: 2013/08/21
Date: Sep 3
Short Title: Human α-L-iduronidase uses its own N-glycan as a substrate-binding and catalytic module
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1306939110
PMCID: PMC3767532
Accession Number: 23959878
Keywords: Amino Acid Sequence
Binding Sites
Biocatalysis
Circular Dichroism
Crystallography, X-Ray
Dermatan Sulfate/metabolism
Electrophoresis, Polyacrylamide Gel
Heparitin Sulfate/metabolism
Humans
Iduronidase/*chemistry/genetics/*metabolism
Kinetics
Mannose/chemistry/metabolism
*Models, Molecular
Molecular Sequence Data
Mucopolysaccharidosis I/enzymology/metabolism
Mutation
Polysaccharides/*chemistry/*metabolism
Protein Binding
Protein Structure, Tertiary
Sequence Homology, Amino Acid
Substrate Specificity
N-linked glycan
X-ray crystallography
glycoside hydrolase family 39
Abstract: N-glycosylation is a major posttranslational modification that endows proteins with various functions. It is established that N-glycans are essential for the correct folding and stability of some enzymes; however, the actual effects of N-glycans on their activities are poorly understood. Here, we show that human α-l-iduronidase (hIDUA), of which a dysfunction causes accumulation of dermatan/heparan sulfate leading to mucopolysaccharidosis type I, uses its own N-glycan as a substrate binding and catalytic module. Structural analysis revealed that the mannose residue of the N-glycan attached to N372 constituted a part of the substrate-binding pocket and interacted directly with a substrate. A deglycosylation study showed that enzyme activity was highly correlated with the N-glycan attached to N372. The kinetics of native and deglycosylated hIDUA suggested that the N-glycan is also involved in catalytic processes. Our study demonstrates a previously unrecognized function of N-glycans.
Notes: 1091-6490
Maita, Nobuo
Tsukimura, Takahiro
Taniguchi, Takako
Saito, Seiji
Ohno, Kazuki
Taniguchi, Hisaaki
Sakuraba, Hitoshi
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2013 Sep 3;110(36):14628-33. doi: 10.1073/pnas.1306939110. Epub 2013 Aug 19.
Author Address: Laboratory of X-Ray Crystallography, Institute for Enzyme Research, University of Tokushima, Tokushima 770-8503, Japan. nmaita@tokushima-u.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1664
Author: Májek, P., Pecankova, K., Maly, M., Oravec, M., Riedel, T. and Dyr, J. E.
Year: 2015
Title: N-Glycosylation of apolipoprotein A1 in cardiovascular diseases
Journal: Transl Res
Volume: 165
Issue: 2
Pages: 360-2
Epub Date: 2014/09/30
Date: Feb
Short Title: N-Glycosylation of apolipoprotein A1 in cardiovascular diseases
Alternate Journal: Translational research : the journal of laboratory and clinical medicine
ISSN: 1878-1810
DOI: 10.1016/j.trsl.2014.09.003
Accession Number: 25262938
Keywords: Apolipoprotein A-I/*blood
Carbohydrates/*chemistry
Humans
Myocardial Infarction/*blood
*Proteome
Notes: 1878-1810
Májek, Pavel
Pecankova, Klara
Maly, Martin
Oravec, Milan
Riedel, Tomas
Dyr, Jan E
Comment
Letter
Research Support, Non-U.S. Gov't
United States
Transl Res. 2015 Feb;165(2):360-2. doi: 10.1016/j.trsl.2014.09.003. Epub 2014 Sep 11.
Author Address: Institute of Hematology and Blood Transfusion, Prague, Czech Republic. Electronic address: pavel.majek@uhkt.cz.
Institute of Hematology and Blood Transfusion, Prague, Czech Republic.
Central Military Hospital, 1st Medical Faculty, Charles University, Prague, Czech Republic.
Complex Cardiovascular Center for Adult Patients, Cardiology Clinic, 2nd Faculty of Medicine, Charles University and University Hospital Motol, Prague, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1018
Author: Majumdar, G., Harrington, A., Hungerford, J., Martinez-Hernandez, A., Gerling, I. C., Raghow, R. and Solomon, S.
Year: 2006
Title: Insulin dynamically regulates calmodulin gene expression by sequential o-glycosylation and phosphorylation of sp1 and its subcellular compartmentalization in liver cells
Journal: J Biol Chem
Volume: 281
Issue: 6
Pages: 3642-50
Epub Date: 2005/12/08
Date: Feb 10
Short Title: Insulin dynamically regulates calmodulin gene expression by sequential o-glycosylation and phosphorylation of sp1 and its subcellular compartmentalization in liver cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M511223200
Accession Number: 16332679
Keywords: Animals
Antibodies, Monoclonal/chemistry
Blotting, Western
Calmodulin/*metabolism
Cell Line, Tumor
Cell Nucleus/metabolism
Cytoplasm/metabolism
*Gene Expression Regulation
Glycosylation
Immunohistochemistry
Immunoprecipitation
Insulin/*metabolism
Liver/*metabolism
Mass Spectrometry
Microscopy, Confocal
Microscopy, Fluorescence
Peptides/chemistry
Phosphorylation
Protein Processing, Post-Translational
RNA, Messenger/metabolism
Rats
Serine/chemistry
Sp1 Transcription Factor/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Time Factors
Transcription, Genetic
Abstract: O-glycosylation and phosphorylation of Sp1 are thought to modulate the expression of a number of genes in normal and diabetic state. Sp1 is an obligatory transcription factor for constitutive and insulin-responsive expression of the calmodulin gene (Majumdar, G., Harmon, A., Candelaria, R., Martinez-Hernandez, A., Raghow, R., and Solomon, S. S. (2003) Am. J. Physiol. 285, E584-E591). Here we report the temporal dynamics of accumulation of total, O-GlcNAc-modified, and phosphorylated Sp1 in H-411E hepatoma cells by immunohistochemistry with monospecific antibodies, confocal microscopy, and matrix-assisted laser desorption and ionization-time of flight mass spectrometry. Insulin elicited sequential and reciprocal post-translational modifications of Sp1. The O-glycosylation of Sp1 and its nuclear accumulation induced by insulin peaked early (approximately 30 min), followed by a steady decline of O-GlcNAc-modified Sp1 to negligible levels by 240 min. The accumulation of phosphorylated Sp1 in the nuclei of insulin-treated cells showed an opposite pattern, increasing steadily until reaching a maximum around 240 min after treatment. Analyses of the total, O-GlcNAc-modified, or phosphorylated Sp1 by Western blot and mass spectrometry corroborated the sequential and reciprocal control of post-translational modifications of Sp1 in response to insulin. Treatment of cells with streptozotocin (a potent inhibitor of O-GlcNAcase) led to hyperglycosylation of Sp1 that failed to be significantly phosphorylated. The mass spectrometry data indicated that a number of common serine residues of Sp1 undergo time-dependent, reciprocal O-glycosylation and phosphorylation, paralleling its rapid translocation from cytoplasm to the nucleus. Later, changes in the steady state levels of phosphorylated Sp1 mimicked the enhanced steady state levels of calmodulin mRNA seen after insulin treatment. Thus, O-glycosylation of Sp1 appears to be critical for its localization into the nucleus, where it undergoes obligatory phosphorylation that is needed for Sp1 to activate calmodulin gene expression.
Notes: Majumdar, Gipsy
Harrington, Adrienne
Hungerford, James
Martinez-Hernandez, Antonio
Gerling, Ivan C
Raghow, Rajendra
Solomon, Solomon
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Biol Chem. 2006 Feb 10;281(6):3642-50. doi: 10.1074/jbc.M511223200. Epub 2005 Dec 6.
Author Address: Research, Medical, and Pathology Services, Veterans Affairs Medical Center Memphis, Tennessee 38104, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 863
Author: Makarova, O., Roh, M. H., Liu, C. J., Laurinec, S. and Margolis, B.
Year: 2003
Title: Mammalian Crumbs3 is a small transmembrane protein linked to protein associated with Lin-7 (Pals1)
Journal: Gene
Volume: 302
Issue: 1-2
Pages: 21-9
Epub Date: 2003/01/16
Date: Jan 2
Short Title: Mammalian Crumbs3 is a small transmembrane protein linked to protein associated with Lin-7 (Pals1)
Alternate Journal: Gene
ISSN: 0378-1119 (Print)
0378-1119
DOI: 10.1016/s0378111902010843
Accession Number: 12527193
Keywords: Adaptor Proteins, Signal Transducing
Amino Acid Sequence
Animals
Base Sequence
Cloning, Molecular
DNA, Complementary/chemistry/genetics
Epithelial Cells/metabolism
Female
Gene Expression
Glycosylation
Humans
Immunohistochemistry
Male
Membrane Glycoproteins/*genetics/metabolism
Membrane Proteins/*genetics/*metabolism
Microscopy, Confocal
Molecular Sequence Data
Protein Binding
Rats
Sequence Alignment
Sequence Analysis, DNA
Sequence Homology, Amino Acid
Vesicular Transport Proteins
Abstract: Drosophila Crumbs is a transmembrane protein that plays an important role in epithelial cell polarity and photoreceptor development. Overexpression of Crumbs in Drosophila epithelia expands the apical surface and leads to disruption of cell polarity. Drosophila Crumbs also interacts with two other polarity genes, Stardust and Discs Lost. Recent work has identified a human orthologue of Drosophila Crumbs, known as CRB1, that is mutated in the eye disorders, retinitis pigmentosa and Leber congenital amaurosis. Our work has demonstrated that human CRB1 can form a complex with mammalian orthologues of Stardust and Discs Lost, known as protein associated with Lin-7 (Pals1) and Pals1 associated tight junction (PATJ), respectively. In the current report we have cloned a full length cDNA for a human paralogue of CRB1 called Crumbs3 (CRB3). In contrast to Drosophila Crumbs and CRB1, CRB3 has a very short extracellular domain but like these proteins it has a conserved intracellular domain that allows it to complex with Pals1 and PATJ. Mouse and human CRB3 have identical intracellular domains but divergent extracellular domains except for a conserved N-glycosylation site. CRB3 is localized to the apical surface and tight junctions but the conserved N linked glycosylation site does not appear to be necessary for CRB3 apical targeting. CRB3 is a specialized isoform of the Crumbs protein family that is expressed in epithelia and can tie the apical membrane to the tight junction.
Notes: Makarova, Olga
Roh, Michael H
Liu, Chia-Jen
Laurinec, Stephanie
Margolis, Ben
DK58208/DK/NIDDK NIH HHS/United States
T32 GM07544-24/GM/NIGMS NIH HHS/United States
T32 GM07863/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Netherlands
Gene. 2003 Jan 2;302(1-2):21-9. doi: 10.1016/s0378111902010843.
Author Address: Howard Hughes Medical Institute, University of Michigan Medical School, 4570 MSRB II, Box 0650, 1150 W. Medical Center Drive, Ann Arbor, MI 48109-0650, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 765
Author: Makino, Y., Omichi, K., Kuraya, N., Ogawa, H., Nishimura, H., Iwanaga, S. and Hase, S.
Year: 2000
Title: Structural analysis of N-linked sugar chains of human blood clotting factor IX
Journal: J Biochem
Volume: 128
Issue: 2
Pages: 175-80
Epub Date: 2000/08/02
Date: Aug
Short Title: Structural analysis of N-linked sugar chains of human blood clotting factor IX
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a022738
Accession Number: 10920251
Keywords: Carbohydrate Sequence
Carbohydrates/*chemistry
Chromatography, High Pressure Liquid
Factor IX/*chemistry
Humans
Molecular Sequence Data
N-Acetylneuraminic Acid/chemistry
Abstract: The structures of N-glycans of human blood clotting factor IX were studied. N-Glycans liberated by hydrazinolysis were N-acetylated and the reducing-end sugar residues were tagged with 2-aminopyridine. The pyridylamino (PA-) sugar chains thus obtained were purified by HPLC. Each PA-sugar chain was analyzed by two-dimensional sugar mapping combined with glycosidase digestion. The major structures of the N-linked sugar chains of human factor IX were found to be sialotetraantennary and sialotriantennary chains with or without fucose residues. These highly sialylated sugar chains are located on the activation peptide of the protein.
Notes: Makino, Y
Omichi, K
Kuraya, N
Ogawa, H
Nishimura, H
Iwanaga, S
Hase, S
Journal Article
Research Support, Non-U.S. Gov't
England
J Biochem. 2000 Aug;128(2):175-80. doi: 10.1093/oxfordjournals.jbchem.a022738.
Author Address: Department of Chemistry, Graduate School of Science, Osaka University, Machikaneyama-cho, Toyonaka, Osaka 560-0043, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1840
Author: Malaker, S. A., Penny, S. A., Steadman, L. G., Myers, P. T., Loke, J. C., Raghavan, M., Bai, D. L., Shabanowitz, J., Hunt, D. F. and Cobbold, M.
Year: 2017
Title: Identification of Glycopeptides as Posttranslationally Modified Neoantigens in Leukemia
Journal: Cancer Immunol Res
Volume: 5
Issue: 5
Pages: 376-384
Epub Date: 2017/03/21
Date: May
Short Title: Identification of Glycopeptides as Posttranslationally Modified Neoantigens in Leukemia
Alternate Journal: Cancer immunology research
ISSN: 2326-6066 (Print)
2326-6066
DOI: 10.1158/2326-6066.cir-16-0280
PMCID: PMC5508727
NIHMSID: NIHMS860644 Associated Glycopeptides as Targets for Cancer Immunotherapy,” has been filed U.S. Patent Application No. 62/202,359 which has been licensed to Agenus Inc. D.F.H and M.C. receive research support and consultancy from Agenus Inc. The other authors declare no competing interests.
Accession Number: 28314751
Keywords: Antigens, Neoplasm/*immunology/metabolism
Cell Line
Cell Line, Tumor
Glycopeptides/*immunology/metabolism
Glycosylation
HLA-B7 Antigen/*immunology/metabolism
Humans
Leukemia/*immunology
Methylation
Protein Processing, Post-Translational
T-Lymphocytes/immunology
Abstract: Leukemias are highly immunogenic, but they have a low mutational load, providing few mutated peptide targets. Thus, the identification of alternative neoantigens is a pressing need. Here, we identify 36 MHC class I-associated peptide antigens with O-linked β-N-acetylglucosamine (O-GlcNAc) modifications as candidate neoantigens, using three experimental approaches. Thirteen of these peptides were also detected with disaccharide units on the same residues and two contain either mono- and/or di-methylated arginine residues. A subset were linked with key cancer pathways, and these peptides were shared across all of the leukemia patient samples tested (5/5). Seven of the O-GlcNAc peptides were synthesized and five (71%) were shown to be associated with multifunctional memory T-cell responses in healthy donors. An O-GlcNAc-specific T-cell line specifically killed autologous cells pulsed with the modified peptide, but not the equivalent unmodified peptide. Therefore, these posttranslationally modified neoantigens provide logical targets for cancer immunotherapy. Cancer Immunol Res; 5(5); 376-84. ©2017 AACR.
Notes: 2326-6074
Malaker, Stacy A
Penny, Sarah A
Steadman, Lora G
Myers, Paisley T
Loke, Justin C
Raghavan, Manoj
Bai, Dina L
Shabanowitz, Jeffrey
Hunt, Donald F
Cobbold, Mark
R01 AI033993/AI/NIAID NIH HHS/United States
R01 GM037537/GM/NIGMS NIH HHS/United States
R37 AI033993/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cancer Immunol Res. 2017 May;5(5):376-384. doi: 10.1158/2326-6066.CIR-16-0280. Epub 2017 Mar 17.
Author Address: Department of Chemistry, University of Virginia, Charlottesville, Virginia.
Department of Clinical Immunology, University of Birmingham, Birmingham, United Kingdom.
Department of Pathology, University of Virginia, Charlottesville, Virginia.
Department of Clinical Immunology, University of Birmingham, Birmingham, United Kingdom. mcobbold@mgh.harvard.edu.
Massachusetts General Hospital Cancer Center and Department of Medicine, Harvard Medical School, Charlestown, Massachusetts.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 410
Author: Malfroy, B., Schofield, P. R., Kuang, W. J., Seeburg, P. H., Mason, A. J. and Henzel, W. J.
Year: 1987
Title: Molecular cloning and amino acid sequence of rat enkephalinase
Journal: Biochem Biophys Res Commun
Volume: 144
Issue: 1
Pages: 59-66
Epub Date: 1987/04/14
Date: Apr 14
Short Title: Molecular cloning and amino acid sequence of rat enkephalinase
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/s0006-291x(87)80475-8
Accession Number: 3555489
Keywords: Amino Acid Sequence
Animals
Base Sequence
Brain/enzymology
Carboxypeptidases/genetics
Cloning, Molecular
DNA/genetics
Endopeptidases/*genetics
Kidney/enzymology
Neprilysin
Rats
Rats, Inbred Strains
Abstract: cDNA clones encoding rat enkephalinase (neutral endopeptidase, EC 3.4.24.11) have been isolated in lambda gt10 libraries from both brain and kidney mRNAs and the complete 742 amino acid sequence of rat enkephalinase is presented. The enzyme possesses a single transmembrane spanning domain near the N-terminal of the molecule but lacks a signal sequence. Because enkephalinase has it active site located extracellularly and is thus an ectopeptidase, we suggest that the N-terminal transmembrane region of the enzyme anchors the protein in membranes and that the majority of the protein, including the carboxy terminus, is extracellular. Enkephalinase, a zinc-containing metallo enzyme, displays homology with other zinc metallo enzymes such as carboxypeptidase A, B and E, suggesting enzymatic similarities in these enzymes.
Notes: Malfroy, B
Schofield, P R
Kuang, W J
Seeburg, P H
Mason, A J
Henzel, W J
Comparative Study
Journal Article
United States
Biochem Biophys Res Commun. 1987 Apr 14;144(1):59-66. doi: 10.1016/s0006-291x(87)80475-8.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1243
Author: Mandel, A. L., Ozdener, H. and Utermohlen, V.
Year: 2009
Title: Identification of pro- and mature brain-derived neurotrophic factor in human saliva
Journal: Arch Oral Biol
Volume: 54
Issue: 7
Pages: 689-95
Epub Date: 2009/05/27
Date: Jul
Short Title: Identification of pro- and mature brain-derived neurotrophic factor in human saliva
Alternate Journal: Archives of oral biology
ISSN: 0003-9969 (Print)
0003-9969
DOI: 10.1016/j.archoralbio.2009.04.005
PMCID: PMC2716651
NIHMSID: NIHMS111536
Accession Number: 19467646
Keywords: Adult
Blotting, Western
Brain-Derived Neurotrophic Factor/*analysis/blood/genetics
Electrophoresis, Polyacrylamide Gel
Female
Fibrinolysin
Genotype
Glycoside Hydrolases
Humans
Male
Methionine/genetics
Molecular Weight
Polymorphism, Single Nucleotide/genetics
Protein Precursors/*analysis/blood/genetics
Saliva/*chemistry
Valine/genetics
Abstract: OBJECTIVE: Growth factors, including brain-derived neurotrophic factor (BDNF), are polypeptides that are involved in the maintenance, survival, and death of central and peripheral cells. Numerous growth factors have been identified in saliva and are thought to promote wound healing and maintenance of the oral epithelium. The aim of this study was to determine if BDNF is also found in human saliva. METHODS: Whole, unstimulated saliva samples (n = 30) were analyzed by SDS-PAGE and Western blot using an anti-human BDNF antibody. Proteolytic cleavage products were similarly assessed following the incubation of pooled saliva with N-glycanase F and plasmin. Subjects were also genotyped for the BDNF Val66Met single nucleotide polymorphism (SNP). RESULTS: These experiments revealed the presence of immunoreactive bands at 14, 32 and 34 kDa, corresponding to mature (mBDNF) and proBDNF, as well as a truncated pro-form at 24 kDa. Not every sample contained all forms of BDNF. Treatment with N-glycanase and plasmin reduced the size of the higher molecular weight bands, confirming the glycosylated pro-form of BDNF. mBDNF was detected significantly less often in subjects with the Val66Met SNP, compared to those without the polymorphism (chi(2) = 4.05; P < 0.05). CONCLUSIONS: While the function of salivary BDNF still requires elucidation, these findings suggest that it may be possible to use saliva in lieu of blood in future studies of BDNF and the Val66Met polymorphism.
Notes: 1879-1506
Mandel, A L
Ozdener, H
Utermohlen, V
T32 DK007158/DK/NIDDK NIH HHS/United States
T32 DK007158-31/DK/NIDDK NIH HHS/United States
5 T32 DK-007158 31/DK/NIDDK NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Arch Oral Biol. 2009 Jul;54(7):689-95. doi: 10.1016/j.archoralbio.2009.04.005. Epub 2009 May 20.
Author Address: Monell Chemical Senses Center, Philadelphia, PA 19104, USA. amandel@monell.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 918
Author: Mani, K., Cheng, F., Havsmark, B., David, S. and Fransson, L. A.
Year: 2004
Title: Involvement of glycosylphosphatidylinositol-linked ceruloplasmin in the copper/zinc-nitric oxide-dependent degradation of glypican-1 heparan sulfate in rat C6 glioma cells
Journal: J Biol Chem
Volume: 279
Issue: 13
Pages: 12918-23
Epub Date: 2004/01/07
Date: Mar 26
Short Title: Involvement of glycosylphosphatidylinositol-linked ceruloplasmin in the copper/zinc-nitric oxide-dependent degradation of glypican-1 heparan sulfate in rat C6 glioma cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M313678200
Accession Number: 14707133
Keywords: Animals
Ascorbic Acid/metabolism
Cell Line, Tumor
Cell-Free System
Ceruloplasmin/*chemistry/metabolism
Chromatography
Copper/chemistry
Glioma/metabolism
Glycosylphosphatidylinositols/*chemistry
Heparan Sulfate Proteoglycans/*chemistry
Heparitin Sulfate/*chemistry
Ions
Microscopy, Confocal
Models, Biological
Nitric Oxide/chemistry
Nitrogen/metabolism
Rats
Zinc/chemistry
Abstract: The core protein of glypican-1, a glycosylphosphatidylinositol-linked heparan sulfate proteoglycan, can bind Cu(II) or Zn(II) ions and undergo S-nitrosylation in the presence of nitric oxide. Cu(II)-to-Cu(I)-reduction supports extensive and permanent nitrosothiol formation, whereas Zn(II) ions appear to support a more limited, possibly transient one. Ascorbate induces release of nitric oxide, which catalyzes deaminative degradation of the heparan sulfate chains on the same core protein. Although free Zn(II) ions support a more limited degradation, Cu(II) ions support a more extensive self-pruning process. Here, we have investigated processing of glypican-1 in rat C6 glioma cells and the possible participation of the copper-containing glycosylphosphatidylinositol-linked splice variant of ceruloplasmin in nitrosothiol formation. Confocal microscopy demonstrated colocalization of glypican-1 and ceruloplasmin in endosomal compartments. Ascorbate induced extensive, Zn(II)-supported heparan sulfate degradation, which could be demonstrated using a specific zinc probe. RNA interference silencing of ceruloplasmin expression reduced the extent of Zn(II)-supported degradation. In cell-free experiments, the presence of free Zn(II) ions prevented free Cu(II) ion from binding to glypican-1 and precluded extensive heparan sulfate autodegradation. However, in the presence of Cu(II)-loaded ceruloplasmin, heparan sulfate in Zn(II)-loaded glypican-1 underwent extensive, ascorbate-induced degradation. We propose that the Cu(II)-to-Cu(I)-reduction that is required for S-nitrosylation of glypican-1 can take place on ceruloplasmin and thereby ensure extensive glypican-1 processing in the presence of free Zn(II) ions.
Notes: Mani, Katrin
Cheng, Fang
Havsmark, Birgitta
David, Samuel
Fransson, Lars-Ake
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2004 Mar 26;279(13):12918-23. doi: 10.1074/jbc.M313678200. Epub 2004 Jan 5.
Author Address: Department of Cell and Molecular Biology, Section for Cell and Matrix Biology, Lund University, Biomedical Center C13, SE-221 84 Lund, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 561
Author: Manneberg, M., Friedlein, A., Kurth, H., Lahm, H. W. and Fountoulakis, M.
Year: 1994
Title: Structural analysis and localization of the carbohydrate moieties of a soluble human interferon gamma receptor produced in baculovirus-infected insect cells
Journal: Protein Sci
Volume: 3
Issue: 1
Pages: 30-8
Epub Date: 1994/01/01
Date: Jan
Short Title: Structural analysis and localization of the carbohydrate moieties of a soluble human interferon gamma receptor produced in baculovirus-infected insect cells
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1002/pro.5560030105
PMCID: PMC2142480
Accession Number: 8142896
Keywords: Amino Acid Sequence
Animals
Baculoviridae/*genetics
Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/analysis/*chemistry
Chromatography, High Pressure Liquid
Genetic Vectors
Glycosylation
Humans
Molecular Sequence Data
*Moths
Receptors, Interferon/*chemistry/genetics
Recombinant Proteins/chemistry
Abstract: A soluble form of the human interferon gamma receptor that is required for the identification of interferon gamma antagonists was expressed in baculovirus-infected insect cells. The protein carried N-linked carbohydrate and showed a heterogeneity on denaturing polyacrylamide gels. We investigated the utilization of the potential sites for N-linked glycosylation and the structure of the carbohydrate moieties of this soluble receptor. Amino acid sequence analysis and ion spray mass spectrometry revealed that of the five potential sites for N-linked glycosylation, Asn17 and Asn69 were always utilized, whereas Asn62 and Asn162 were utilized in approximately one-third of the protein population. Asn223 was never found to be glycosylated. The soluble receptor was treated with N-glycosidase F and the oligosaccharides released were analyzed by matrix-assisted laser desorption mass spectrometry, which showed that the protein carried six types of short carbohydrate chains. The predominant species was a hexasaccharide of molecular mass 1,039, containing a fucose subunit linked to the proximal N-acetylglucosamine residue: [formula: see text]
Notes: 1469-896x
Manneberg, M
Friedlein, A
Kurth, H
Lahm, H W
Fountoulakis, M
Journal Article
Protein Sci. 1994 Jan;3(1):30-8. doi: 10.1002/pro.5560030105.
Author Address: Department of Biology, F. Hoffmann-La Roche Ltd., Pharmaceutical Research-New Technologies, Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1809
Author: Mansoor, S. E., Lü, W., Oosterheert, W., Shekhar, M., Tajkhorshid, E. and Gouaux, E.
Year: 2016
Title: X-ray structures define human P2X(3) receptor gating cycle and antagonist action
Journal: Nature
Volume: 538
Issue: 7623
Pages: 66-71
Epub Date: 2016/09/15
Date: Oct 6
Short Title: X-ray structures define human P2X(3) receptor gating cycle and antagonist action
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature19367
PMCID: PMC5161641
NIHMSID: NIHMS821331
Accession Number: 27626375
Keywords: Apoproteins/agonists/antagonists & inhibitors/chemistry/metabolism
Binding Sites/drug effects
Binding, Competitive/drug effects
Crystallization
Crystallography, X-Ray
Humans
Ion Channel Gating/*drug effects
Ion Transport
Ligands
Models, Molecular
Porosity
Protein Conformation
Purinergic Agonists/pharmacology
Purinergic P2X Receptor Antagonists/*pharmacology
Receptors, Purinergic P2X3/*chemistry/*metabolism
Abstract: P2X receptors are trimeric, non-selective cation channels activated by ATP that have important roles in the cardiovascular, neuronal and immune systems. Despite their central function in human physiology and although they are potential targets of therapeutic agents, there are no structures of human P2X receptors. The mechanisms of receptor desensitization and ion permeation, principles of antagonism, and complete structures of the pore-forming transmembrane domains of these receptors remain unclear. Here we report X-ray crystal structures of the human P2X(3) receptor in apo/resting, agonist-bound/open-pore, agonist-bound/closed-pore/desensitized and antagonist-bound/closed states. The open state structure harbours an intracellular motif we term the 'cytoplasmic cap', which stabilizes the open state of the ion channel pore and creates lateral, phospholipid-lined cytoplasmic fenestrations for water and ion egress. The competitive antagonists TNP-ATP and A-317491 stabilize the apo/resting state and reveal the interactions responsible for competitive inhibition. These structures illuminate the conformational rearrangements that underlie P2X receptor gating and provide a foundation for the development of new pharmacological agents.
Notes: 1476-4687
Mansoor, Steven E
Lü, Wei
Oosterheert, Wout
Shekhar, Mrinal
Tajkhorshid, Emad
Gouaux, Eric
F32 GM108391/GM/NIGMS NIH HHS/United States
R01 GM100400/GM/NIGMS NIH HHS/United States
U54 GM087519/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
P41 GM104601/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2016 Oct 6;538(7623):66-71. doi: 10.1038/nature19367. Epub 2016 Sep 14.
Author Address: Vollum Institute, Oregon Health &Science University, Portland, Oregon 97239, USA.
Knight Cardiovascular Institute, Oregon Health &Science University, Portland, Oregon 97239, USA.
Department of Biochemistry, Center for Biophysics and Quantitative Biology, and Beckman Institute for Advanced Science and Technology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Howard Hughes Medical Institute, Oregon Health &Science University, Portland, Oregon 97239, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1398
Author: Margraf-Schönfeld, S., Böhm, C. and Watzl, C.
Year: 2011
Title: Glycosylation affects ligand binding and function of the activating natural killer cell receptor 2B4 (CD244) protein
Journal: J Biol Chem
Volume: 286
Issue: 27
Pages: 24142-9
Epub Date: 2011/05/25
Date: Jul 8
Short Title: Glycosylation affects ligand binding and function of the activating natural killer cell receptor 2B4 (CD244) protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.225334
PMCID: PMC3129195
Accession Number: 21606496
Keywords: Antigens, CD/genetics/immunology/*metabolism
CD48 Antigen
Glycosylation
HEK293 Cells
HeLa Cells
Humans
Killer Cells, Natural/cytology/immunology/*metabolism
Ligands
Lymphocyte Activation/*physiology
Receptors, Immunologic/genetics/immunology/*metabolism
Signaling Lymphocytic Activation Molecule Family
Abstract: 2B4 (CD244) is an important activating receptor for the regulation of natural killer (NK) cell responses. Here we show that 2B4 is heavily and differentially glycosylated in primary human NK cells and NK cell lines. The differential glycosylation could be attributed to sialic acid residues on N- and O-linked carbohydrates. Using a recombinant fusion protein of the extracellular domain of 2B4, we demonstrate that N-linked glycosylation of 2B4 is essential for the binding to its ligand CD48. In contrast, sialylation of 2B4 has a negative impact on ligand binding, as the interaction between 2B4 and CD48 is increased after the removal of sialic acids. This was confirmed in a functional assay system, where the desialylation of NK cells or the inhibition of O-linked glycosylation resulted in increased 2B4-mediated lysis of CD48-expressing tumor target cells. These data demonstrate that glycosylation has an important impact on 2B4-mediated NK cell function and suggest that regulated changes in glycosylation during NK cell development and activation might be involved in the regulation of NK cell responses.
Notes: 1083-351x
Margraf-Schönfeld, Stefanie
Böhm, Carolin
Watzl, Carsten
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 Jul 8;286(27):24142-9. doi: 10.1074/jbc.M111.225334. Epub 2011 May 23.
Author Address: Institute for Immunology, University Heidelberg, 69120 Heidelberg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 715
Author: Marie-Cardine, A., Kirchgessner, H., Bruyns, E., Shevchenko, A., Mann, M., Autschbach, F., Ratnofsky, S., Meuer, S. and Schraven, B.
Year: 1999
Title: SHP2-interacting transmembrane adaptor protein (SIT), a novel disulfide-linked dimer regulating human T cell activation
Journal: J Exp Med
Volume: 189
Issue: 8
Pages: 1181-94
Epub Date: 1999/04/20
Date: Apr 19
Short Title: SHP2-interacting transmembrane adaptor protein (SIT), a novel disulfide-linked dimer regulating human T cell activation
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.189.8.1181
PMCID: PMC2193021
Accession Number: 10209036
Keywords: Adaptor Proteins, Signal Transducing
Amino Acid Sequence
Carrier Proteins/chemistry/*genetics
Cloning, Molecular
DNA-Binding Proteins/metabolism
Dimerization
Disulfides/chemistry
Gene Expression Regulation/genetics
Humans
Intracellular Signaling Peptides and Proteins
Jurkat Cells
Lymphocyte Activation
Membrane Glycoproteins/chemistry/*genetics
Membrane Proteins/chemistry/*genetics
Molecular Sequence Data
NFATC Transcription Factors
*Nuclear Proteins
Phorbol Esters/pharmacology
Phosphorylation
Protein Tyrosine Phosphatase, Non-Receptor Type 11
Protein Tyrosine Phosphatase, Non-Receptor Type 6
Protein Tyrosine Phosphatases/*metabolism
RNA, Messenger/metabolism
Receptors, Antigen, T-Cell/metabolism
SH2 Domain-Containing Protein Tyrosine Phosphatases
Sequence Alignment
Signal Transduction/genetics
T-Lymphocytes/*metabolism
Transcription Factors/metabolism
src Homology Domains/genetics
Abstract: T lymphocytes express several low molecular weight transmembrane adaptor proteins that recruit src homology (SH)2 domain-containing intracellular molecules to the cell membrane via tyrosine-based signaling motifs. We describe here a novel molecule of this group termed SIT (SHP2 interacting transmembrane adaptor protein). SIT is a disulfide-linked homodimeric glycoprotein that is expressed in lymphocytes. After tyrosine phosphorylation by src and possibly syk protein tyrosine kinases SIT recruits the SH2 domain-containing tyrosine phosphatase SHP2 via an immunoreceptor tyrosine-based inhibition motif. Overexpression of SIT in Jurkat cells downmodulates T cell receptor- and phytohemagglutinin-mediated activation of the nuclear factor of activated T cells (NF-AT) by interfering with signaling processes that are probably located upstream of activation of phospholipase C. However, binding of SHP2 to SIT is not required for inhibition of NF-AT induction, suggesting that SIT not only regulates NF-AT activity but also controls NF-AT unrelated pathways of T cell activation involving SHP2.
Notes: 1540-9538
Marie-Cardine, A
Kirchgessner, H
Bruyns, E
Shevchenko, A
Mann, M
Autschbach, F
Ratnofsky, S
Meuer, S
Schraven, B
Journal Article
Research Support, Non-U.S. Gov't
J Exp Med. 1999 Apr 19;189(8):1181-94. doi: 10.1084/jem.189.8.1181.
Author Address: Immunomodulation Laboratory of the Institute for Immunology, University of Heidelberg, 69120 Heidelberg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1722
Author: Marino, F., Bern, M., Mommen, G. P. M., Leney, A. C., van Gaans-van den Brink, J. A. M., Bonvin, Amjj, Becker, C., van Els, Cacm and Heck, A. J. R.
Year: 2015
Title: Extended O-GlcNAc on HLA Class-I-Bound Peptides
Journal: J Am Chem Soc
Volume: 137
Issue: 34
Pages: 10922-10925
Epub Date: 2015/08/19
Date: Sep 2
Short Title: Extended O-GlcNAc on HLA Class-I-Bound Peptides
Alternate Journal: Journal of the American Chemical Society
ISSN: 0002-7863 (Print)
0002-7863
DOI: 10.1021/jacs.5b06586
PMCID: PMC4603548
NIHMSID: NIHMS725706
Accession Number: 26280087
Keywords: Acetylglucosamine/*chemistry
Histocompatibility Antigens Class I/*chemistry/metabolism
Humans
Models, Molecular
Peptides/*chemistry/metabolism
Abstract: We report unexpected mass spectrometric observations of glycosylated human leukocyte antigen (HLA) class I-bound peptides. Complemented by molecular modeling, in vitro enzymatic assays, and oxonium ion patterns, we propose that the observed O-linked glycans carrying up to five monosaccharides are extended O-GlcNAc's rather than GalNAc-initiated O-glycans. A cytosolic O-GlcNAc modification is normally terminal and does not extend to produce a polysaccharide, but O-GlcNAc on an HLA peptide presents a special case because the loaded HLA class I complex traffics through the endoplasmic reticulum and Golgi apparatus on its way to the cell membrane and is hence exposed to glycosyltransferases. We also report for the first time natural HLA class I presentation of O- and N-linked glycopeptides derived from membrane proteins. HLA class I peptides with centrally located oligosaccharides have been shown to be immunogenic and may thus be important targets for immune surveillance.
Notes: 1520-5126
Marino, Fabio
Bern, Marshall
Mommen, Geert P M
Leney, Aneika C
van Gaans-van den Brink, Jacqueline A M
Bonvin, Alexandre M J J
Becker, Christopher
van Els, Cécile A C M
Heck, Albert J R
R42 GM103362/GM/NIGMS NIH HHS/United States
GM103362/GM/NIGMS NIH HHS/United States
GM100634/GM/NIGMS NIH HHS/United States
R43 GM100634/GM/NIGMS NIH HHS/United States
R41 GM103362/GM/NIGMS NIH HHS/United States
R44 GM100634/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Am Chem Soc. 2015 Sep 2;137(34):10922-10925. doi: 10.1021/jacs.5b06586. Epub 2015 Aug 19.
Author Address: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Netherlands Proteomics Centre, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Protein Metrics Inc., San Carlos, CA 94070.
Institute for Translational Vaccinology, Bilthoven, Netherlands.
Centre for Infectious Disease Control, National Institute for Public Health and the Environment, The Netherlands.
Computational Structural Biology, Bijvoet Center for Biomolecular Research, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 393
Author: Marti, T., Schaller, J., Rickli, E. E., Schmid, K., Kamerling, J. P., Gerwig, G. J., van Halbeek, H. and Vliegenthart, J. F.
Year: 1988
Title: The N- and O-linked carbohydrate chains of human, bovine and porcine plasminogen. Species specificity in relation to sialylation and fucosylation patterns
Journal: Eur J Biochem
Volume: 173
Issue: 1
Pages: 57-63
Epub Date: 1988/04/05
Date: Apr 5
Short Title: The N- and O-linked carbohydrate chains of human, bovine and porcine plasminogen. Species specificity in relation to sialylation and fucosylation patterns
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1988.tb13966.x
Accession Number: 3356193
Keywords: Animals
Carbohydrate Conformation
Cattle
Fucose/*metabolism
Humans
N-Acetylneuraminic Acid
Neuraminic Acids
*Plasminogen/metabolism
*Polysaccharides/isolation & purification
Sialic Acids/*metabolism
*Species Specificity
Structure-Activity Relationship
Swine
Abstract: The structures of the N- and O-glycans of human, bovine and porcine plasminogen were determined by 500-MHz 1H-NMR spectroscopy. The N-glycans of all three species proved to be of the N-acetyllactosamine type differing from one another with respect to the sialylation and fucosylation patterns. In the N-glycan of human plasminogen the two antennae are sialylated with N-acetylneuraminic acid (NeuAc), whereas in the bovine counterpart both branches carry significant amounts of N-glycolylneuraminic acid (NeuGc). In porcine plasminogen the sialic acid is mainly NeuAc; the Man alpha 1----6 branch, however, is only partially sialylated. In addition, the porcine N-glycan is fucosylated to about 80% in alpha 1----6 linkage to the GlcNAc-1 residue. The O-glycans of the three species possess an identical Gal beta 1----3GalNAc core which is alpha 2----3 sialylated with NeuAc at Gal. The disialylated form, which is also present in all three species, has an additional NeuAc residue in alpha 2----6 linkage to GalNAc. Mono- and disialylated forms occur in different molar ratios in the different plasminogens: 80:20 in human, 70:30 in bovine and 50:50 in porcine. This study on the carbohydrate moiety of these three plasminogens reveals species specificity in terms of various types of microheterogeneities.
Notes: Marti, T
Schaller, J
Rickli, E E
Schmid, K
Kamerling, J P
Gerwig, G J
van Halbeek, H
Vliegenthart, J F
GM-10374/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Eur J Biochem. 1988 Apr 5;173(1):57-63. doi: 10.1111/j.1432-1033.1988.tb13966.x.
Author Address: Institute of Biochemistry, University of Bern, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 424
Author: Martiniuk, F., Ellenbogen, A. and Hirschhorn, R.
Year: 1985
Title: Identity of neutral alpha-glucosidase AB and the glycoprotein processing enzyme glucosidase II. Biochemical and genetic studies
Journal: J Biol Chem
Volume: 260
Issue: 2
Pages: 1238-42
Epub Date: 1985/01/25
Date: Jan 25
Short Title: Identity of neutral alpha-glucosidase AB and the glycoprotein processing enzyme glucosidase II. Biochemical and genetic studies
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3881423
Keywords: Chromatography, Gel
Chromatography, Ion Exchange
*Chromosomes, Human, 6-12 and X
Electrophoresis, Polyacrylamide Gel
Electrophoresis, Starch Gel
Female
Glucosidases/*genetics
Humans
Isoelectric Focusing
Isoenzymes/*genetics
Placenta/enzymology
Pregnancy
alpha-Glucosidases/*genetics
Abstract: We have previously partially purified, characterized, and chromosomally mapped a human isozyme of alpha-glucosidase which is active at neutral pH. This isozyme appears as a doublet of enzyme activity on native gel electrophoresis and was termed neutral alpha-glucosidase AB. We now report genetic and biochemical evidence that neutral alpha-glucosidase AB is synonymous with the glycoprotein processing enzyme glucosidase II. We have found that a mutant mouse lymphoma line which is deficient in glucosidase II is also deficient in neutral alpha-glucosidase AB, as defined electrophoretically and quantitatively (less than 0.5% of parental). In contrast, both mutant and parental cell lines exhibited several lysosomal hydrolases which are processed by glucosidase II. We have also further purified the human neutral alpha-glucosidase A component of neutral alpha-glucosidase AB 740-fold from placenta in order to compare its biochemical properties with those described for rat liver and pig kidney glucosidase II. Both glucosidase II and neutral alpha-glucosidase AB are high-molecular mass (greater than 200,000 dalton) anionic glycoproteins which bind to concanavalin A, have a broad pH optima (5.5-8.5), and have a similar Km for maltose (4.8 versus 2.1 mM) and the artificial substrate 4-methylumbelliferyl-alpha-D-glucopyranoside (35 versus 19 microM). Similar to human neutral alpha-glucosidase AB, purified rat glucosidase II migrates as a doublet of enzyme activity on native gel electrophoresis. Although rat glucosidase II has been reported to have a subunit size of 67 kDa, pig glucosidase II has been found to have a subunit size of 100 kDa, like the 98-kDa major protein in purified human neutral alpha-glucosidase A. Although we have not demonstrated that neutral alpha-glucosidase AB is microsomal nor that it hydrolyzes the natural substrate of glucosidase II, we believe that the genetic evidence is compelling for and the biochemical data consistent with the hypothesis that neutral alpha-glucosidase AB and glucosidase II are synonymous. These and previous results would localize glucosidase II to the long arm of human chromosome II.
Notes: Martiniuk, F
Ellenbogen, A
Hirschhorn, R
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1985 Jan 25;260(2):1238-42.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1175
Author: Marx, P. F., Brondijk, T. H., Plug, T., Romijn, R. A., Hemrika, W., Meijers, J. C. and Huizinga, E. G.
Year: 2008
Title: Crystal structures of TAFI elucidate the inactivation mechanism of activated TAFI: a novel mechanism for enzyme autoregulation
Journal: Blood
Volume: 112
Issue: 7
Pages: 2803-9
Epub Date: 2008/06/19
Date: Oct 1
Short Title: Crystal structures of TAFI elucidate the inactivation mechanism of activated TAFI: a novel mechanism for enzyme autoregulation
Alternate Journal: Blood
ISSN: 0006-4971
DOI: 10.1182/blood-2008-03-146001
Accession Number: 18559974
Keywords: Carboxypeptidase B2/*chemistry
Carboxypeptidases/chemistry/metabolism
Cell Line
Crystallography, X-Ray
Enzyme Activation
Enzyme Inhibitors/metabolism
Humans
Models, Biological
Mutation/genetics
Protein Precursors
Protein Structure, Secondary
Abstract: Thrombin-activatable fibrinolysis inhibitor (TAFI) is a pro-metallocarboxypeptidase that can be proteolytically activated (TAFIa). TAFIa is unique among carboxypeptidases in that it spontaneously inactivates with a short half-life, a property that is crucial for its role in controlling blood clot lysis. We studied the intrinsic instability of TAFIa by solving crystal structures of TAFI, a TAFI inhibitor (GEMSA) complex and a quadruple TAFI mutant (70-fold more stable active enzyme). The crystal structures show that TAFIa stability is directly related to the dynamics of a 55-residue segment (residues 296-350) that includes residues of the active site wall. Dynamics of this flap are markedly reduced by the inhibitor GEMSA, a known stabilizer of TAFIa, and stabilizing mutations. Our data provide the structural basis for a model of TAFI auto-regulation: in zymogen TAFI the dynamic flap is stabilized by interactions with the activation peptide. Release of the activation peptide increases dynamic flap mobility and in time this leads to conformational changes that disrupt the catalytic site and expose a cryptic thrombin-cleavage site present at Arg302. This represents a novel mechanism of enzyme control that enables TAFI to regulate its activity in plasma in the absence of specific inhibitors.
Notes: 1528-0020
Marx, Pauline F
Brondijk, T Harma C
Plug, Tom
Romijn, Roland A
Hemrika, Wieger
Meijers, Joost C M
Huizinga, Eric G
Journal Article
Research Support, Non-U.S. Gov't
United States
Blood. 2008 Oct 1;112(7):2803-9. doi: 10.1182/blood-2008-03-146001. Epub 2008 Jun 17.
Author Address: Department of Experimental Vascular Medicine, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1107
Author: Maryon, E. B., Molloy, S. A. and Kaplan, J. H.
Year: 2007
Title: O-linked glycosylation at threonine 27 protects the copper transporter hCTR1 from proteolytic cleavage in mammalian cells
Journal: J Biol Chem
Volume: 282
Issue: 28
Pages: 20376-87
Epub Date: 2007/05/26
Date: Jul 13
Short Title: O-linked glycosylation at threonine 27 protects the copper transporter hCTR1 from proteolytic cleavage in mammalian cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M701806200
Accession Number: 17525160
Keywords: Amino Acid Substitution
Animals
CHO Cells
Caco-2 Cells
Cation Transport Proteins/genetics/*metabolism
Copper/*metabolism
Copper Transporter 1
Cricetinae
Cricetulus
Dogs
Gene Expression
Glycosylation
Humans
Ion Transport/physiology
Mutation, Missense
Protein Modification, Translational/*physiology
Protein Structure, Tertiary
Threonine/genetics/*metabolism
Abstract: The major human copper uptake protein, hCTR1, has 190 amino acids and a predicted mass of 21 kDa. hCTR1 antibodies recognize multiple bands in SDS-PAGE centered at 35 kDa. Part of this increased mass is due to N-linked glycosylation at Asn-15. We show that in mammalian cells the N15Q mutant protein trafficked to the plasma membrane and mediated copper uptake at 75% of the rate of wild-type hCTR1. We demonstrate that the extracellular amino terminus of hCTR1 also contains O-linked polysaccharides. Glycosidase treatment that removed O-linked sugars reduced the apparent mass of hCTR1 or N15Q mutant protein by 1-2 kDa. Expression of amino-terminal truncations and alanine substitution mutants of hCTR1 in HEK293 and MDCK cells localized the site of O-linked glycosylation to Thr-27. Expression of alanine substitutions at Thr-27 resulted in proteolytic cleavage of hCTR1 on the carboxyl side of the T27A mutations. This cleavage produced a 17-kDa polypeptide missing approximately the first 30 amino acids of hCTR1. Expression of wild-type hCTR1 in mutant Chinese hamster ovary cells that were unable to initiate O-glycosylation also resulted in hCTR1 cleavage to produce the 17-kDa polypeptide. The 17-kDa hCTR1 polypeptide was located in the plasma membrane and mediated copper uptake at about 50% that of the rate of wild-type hCTR1. Thus, O-linked glycosylation at Thr-27 is necessary to prevent proteolytic cleavage that removes half of the extracellular amino terminus of hCTR1 and significantly impairs transport activity of the remaining polypeptide.
Notes: Maryon, Edward B
Molloy, Shannon A
Kaplan, Jack H
Comparative Study
Journal Article
United States
J Biol Chem. 2007 Jul 13;282(28):20376-87. doi: 10.1074/jbc.M701806200. Epub 2007 May 24.
Author Address: Department of Biochemistry and Molecular Genetics, University of Illinois, 900 S. Ashland Avenue, Chicago, IL 60607, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 588
Author: Mas, E., Abouakil, N., Roudani, S., Franc, J. L., Montreuil, J. and Lombardo, D.
Year: 1993
Title: Variation of the glycosylation of human pancreatic bile-salt-dependent lipase
Journal: Eur J Biochem
Volume: 216
Issue: 3
Pages: 807-12
Epub Date: 1993/09/15
Date: Sep 15
Short Title: Variation of the glycosylation of human pancreatic bile-salt-dependent lipase
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb18201.x
Accession Number: 8404899
Keywords: Bile Acids and Salts/*metabolism
Blotting, Western
Carbohydrate Sequence
Carbohydrates/analysis
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Lectins
Lipase/chemistry/*metabolism
Molecular Sequence Data
Pancreatic Juice/*enzymology
Abstract: Glycoproteins of human pancreatic juice were characterized by means of lectins after electrophoresis and electrotransfer to nitrocellulose membranes. For the detected glycoproteins, only a 100-kDa glycoprotein varied in the pancreatic juice from a normal patient (i.e. without any pancreatic disorder) compared to the pancreatic juice from a patient suffering from chronic pancreatitis. This protein, which is the only protein in human pancreatic juice which is O-glycosylated and N-glycosylated, was identified as the bile-salt-dependent lipase. Among the glycosylated proteins present in human pancreatic juice, only the glycosylation of bile-salt-dependent lipase differs between individuals. The enzyme was isolated either from normal or pathological human pancreatic juices. The purified variants have an identical molecular mass and amino-acid composition. As suspected from lectin affinity studies, the oligosaccharide composition differs between the variants. The structure of the N-linked oligosaccharides of the variant from the pancreatic juice of a normal donor correlated with complete processing and maturation of a complex-type N-glycan. Alteration of the maturation process can be detected for a bile-salt-dependent-lipase variant from a patient suffering with chronic pancreatitis, since the carbohydrate composition is compatible with the predominance of hybrid or high-mannose-type structures. The amount of sugar involved in O-glycosylation associated with the peanut agglutinin reactivity suggests the presence of 12-14 minimal Gal beta 1-->3GalNac-->T/S O-glycan structures which are sialylated and fucosylated. The amount of sugar involved in the O-linked oligosaccharide structure appears to be unchanged in the variants isolated from the pathological pancreatic juice.
Notes: Mas, E
Abouakil, N
Roudani, S
Franc, J L
Montreuil, J
Lombardo, D
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1993 Sep 15;216(3):807-12. doi: 10.1111/j.1432-1033.1993.tb18201.x.
Author Address: INSERM U260, Faculté de Médecine, Marseille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2013
Author: Masclef, L., Dehennaut, V., Mortuaire, M., Schulz, C., Leturcq, M., Lefebvre, T. and Vercoutter-Edouart, A. S.
Year: 2019
Title: Cyclin D1 Stability Is Partly Controlled by O-GlcNAcylation
Journal: Front Endocrinol (Lausanne)
Volume: 10
Pages: 106
Epub Date: 2019/03/12
Short Title: Cyclin D1 Stability Is Partly Controlled by O-GlcNAcylation
Alternate Journal: Frontiers in endocrinology
ISSN: 1664-2392 (Print)
1664-2392
DOI: 10.3389/fendo.2019.00106
PMCID: PMC6395391
Accession Number: 30853938
Keywords: O-GlcNAc
cell cycle
cyclin D
stability
ubiquitination
Abstract: Cyclin D1 is the regulatory partner of the cyclin-dependent kinases (CDKs) CDK4 or CDK6. Once associated and activated, the cyclin D1/CDK complexes drive the cell cycle entry and G1 phase progression in response to extracellular signals. To ensure their timely and accurate activation during cell cycle progression, cyclin D1 turnover is finely controlled by phosphorylation and ubiquitination. Here we show that the dynamic and reversible O-linked β-N-Acetyl-glucosaminylation (O-GlcNAcylation) regulates also cyclin D1 half-life. High O-GlcNAc levels increase the stability of cyclin D1, while reduction of O-GlcNAcylation strongly decreases it. Moreover, elevation of O-GlcNAc levels through O-GlcNAcase (OGA) inhibition significantly slows down the ubiquitination of cyclin D1. Finally, biochemical and cell imaging experiments in human cancer cells reveal that the O-GlcNAc transferase (OGT) binds to and glycosylates cyclin D1. We conclude that O-GlcNAcylation promotes the stability of cyclin D1 through modulating its ubiquitination.
Notes: 1664-2392
Masclef, Louis
Dehennaut, Vanessa
Mortuaire, Marlène
Schulz, Céline
Leturcq, Maïté
Lefebvre, Tony
Vercoutter-Edouart, Anne-Sophie
Journal Article
Front Endocrinol (Lausanne). 2019 Feb 22;10:106. doi: 10.3389/fendo.2019.00106. eCollection 2019.
Author Address: Université de Lille, CNRS, UMR 8576, UGSF, Unité de Glycobiologie Structurale et Fonctionnelle, Lille, France.
Institut Pasteur de Lille, Université de Lille, CNRS, UMR 8161, M3T: Mechanisms of Tumorigenesis and Targeted Therapies, Lille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 621
Author: Mason, A. J., Farnworth, P. G. and Sullivan, J.
Year: 1996
Title: Characterization and determination of the biological activities of noncleavable high molecular weight forms of inhibin A and activin A
Journal: Mol Endocrinol
Volume: 10
Issue: 9
Pages: 1055-65
Epub Date: 1996/09/01
Date: Sep
Short Title: Characterization and determination of the biological activities of noncleavable high molecular weight forms of inhibin A and activin A
Alternate Journal: Molecular endocrinology (Baltimore, Md.)
ISSN: 0888-8809 (Print)
0888-8809
DOI: 10.1210/mend.10.9.8885240
Accession Number: 8885240
Keywords: Activins
Animals
Arginine/genetics/metabolism
Binding Sites
Cells, Cultured
DNA, Complementary/genetics
Follicle Stimulating Hormone/metabolism
Glycosylation
Humans
*Inhibin-beta Subunits
Inhibins/*chemistry/*genetics/*metabolism/pharmacology
Kidney/cytology/embryology/metabolism
Male
Molecular Weight
Mutagenesis, Site-Directed
Pituitary Gland/cytology/drug effects/metabolism
Protein Conformation
Protein Precursors/genetics/metabolism
Protein Processing, Post-Translational
Rats
Recombinant Proteins/genetics/metabolism/pharmacology
Transfection
Abstract: Recombinant expression of human alpha- and beta A-inhibin subunit cDNAs in mammalian 293 cells results in the secretion of 20-53K free alpha-subunit-derived products, 30-105K alpha beta A-inhibin dimers, and 24-110K beta A-activin dimers. The present study verifies that the wide variation in the size of these products is due to incomplete cleavage of the proteolytic processing sites and the differential glycosylation of the N-linked glycosylation site at amino acid number 302 in the alpha C-subunit. The identity of each of these products was established by mutagenesis of proteolytic processing sites and N-linked glycosylation sites, combined with the analysis of transfection products by immunoprecipitation and one- and two-dimensional SDS-PAGE (SDS/SDS-beta-ME). Transient expression of processing site mutants of the alpha- and beta A-subunits in 293 cells was used to generate microgram quantities of noncleavable 55K and 65K inhibin dimers, and noncleavable 110K activin A dimers. The 55K and 65K inhibin A forms were purified and found to be fully biologically active in a rat pituitary cell bioassay. The 110K high molecular weight (HMW) form of human activin A failed to show any FSH-releasing activity in the pituitary assay. Since radioactively labeled 55K and 65K inhibin A and 110K activin A remained intact after incubation with rat pituitary cells for 72 h, there appears to be no conversion of these dimers to lower molecular weight forms by proteolytic cleavage at additional sites. These results show for the first time that 55K and 65K inhibit A are intrinsically biologically active and do not require cleavage to the 32K form for activation. In contrast, cleavage of the 110K activin A precursor to the 24K form would appear to be necessary for activity.
Notes: Mason, A J
Farnworth, P G
Sullivan, J
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Mol Endocrinol. 1996 Sep;10(9):1055-65. doi: 10.1210/mend.10.9.8885240.
Author Address: Prince Henry's Institute of Medical Research, Monash Medical Centre, Clayton, Victoria, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1359
Author: Massoura, A. N., Dover, T. J., Newman, A. S. and Barnes, N. M.
Year: 2011
Title: The identification of N-glycosylated residues of the human 5-HT3B receptor subunit: importance for cell membrane expression
Journal: J Neurochem
Volume: 116
Issue: 6
Pages: 975-83
Epub Date: 2010/12/09
Date: Mar
Short Title: The identification of N-glycosylated residues of the human 5-HT3B receptor subunit: importance for cell membrane expression
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042
DOI: 10.1111/j.1471-4159.2010.07129.x
Accession Number: 21138434
Keywords: Cell Membrane/drug effects/*metabolism
*Gene Expression Regulation
Glycosylation/drug effects
HEK293 Cells/cytology
Humans
Mutagenesis, Site-Directed/methods
Mutation/genetics
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/pharmacology
Protein Subunits/genetics/*metabolism
Receptors, Serotonin, 5-HT3/genetics/*metabolism
Sequence Analysis, Protein/methods
Transfection/methods
Abstract: The 5-hydroxytryptamine 3 (5-HT(3)) receptor is a pentameric ligand-gated ion channel with potential molecular isoforms arising from different subunit combinations and/or different post-translational modifications of the individual subunits. Since N-glycosylation of the 5-HT3A subunit impacts cell surface trafficking, the presence of N-glycosylation of the human (h) 5-HT3B subunit and the influence upon cell membrane expression was investigated. Following transient expression of the h5-HT3B subunit by human embryonic kidney cells (HEK293 cells) stably expressing the h5-HT3A subunit, the N-glycosylation inhibitor tunicamycin reduced the size of the predominant h5-HT3B-immunoreactive protein (∼ 55 kDa reduced to ∼ 40 kDa). Disruption of each consensus N-glycosylation sequences in the h5-HT3B subunit (N31S, N75S, N117S, N147S and N182S) resulted in a reduced molecular weight (by ∼ 2-4 kDa) of each mutant when expressed by HEK293 cells stably expressing the h5-HT3A subunit. Immunocytochemical studies demonstrated that disruption of each of the N-glycosylation sequences (individually or combined) reduced the expression of the mutant h5-HT3B subunit protein in the cell membrane when co-expressed with the h5-HT3A subunit. The present study has identified utilised N-glycosylation sites of the h5-HT3B subunit and demonstrated that they promote subunit expression in the cell membrane; a prerequisite for 5-HT(3) receptor function.
Notes: 1471-4159
Massoura, Andrew N
Dover, Terri J
Newman, Amy S
Barnes, Nicholas M
Journal Article
England
J Neurochem. 2011 Mar;116(6):975-83. doi: 10.1111/j.1471-4159.2010.07129.x. Epub 2011 Jan 19.
Author Address: Neuropharmacology and Neurobiology Section, Clinical and Experimental Medicine, The Medical School, University of Birmingham, Birmingham, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 737
Author: Masuda, K., Takahashi, N., Tsukamoto, Y., Honma, H. and Kohri, K.
Year: 2000
Title: N-Glycan structures of an osteopontin from human bone
Journal: Biochem Biophys Res Commun
Volume: 268
Issue: 3
Pages: 814-7
Epub Date: 2000/02/19
Date: Feb 24
Short Title: N-Glycan structures of an osteopontin from human bone
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1006/bbrc.2000.2224
Accession Number: 10679288
Keywords: Aminopyridines
Bone and Bones/*chemistry
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Humans
Mass Spectrometry
Molecular Sequence Data
Osteopontin
Polysaccharides/*chemistry
Sialoglycoproteins/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tissue Distribution
Abstract: N-Glycan structures of osteopontin (a bone matrix protein) from human bone (lumbar vertabrate) are reported in detail. Asn-linked glycan portion was released from 100 microg of osteopontin by digestion with glycoamidase A (from sweet almond), and the reducing ends of the N-glycans were reductively aminated with 2-aminopyridine. The derivatized N-glycans were separated and structurally identified by a multidimensional mapping technique on HPLC columns. Two major N-glycan structures were also confirmed by mass spectrometry. The proposed structures are shown below. The result should permit future comparison with the N-glycan structures of osteopontins obtained from other sources (kidney tissues, macrophages, urinary stones, human milk, etc.).
Notes: Masuda, K
Takahashi, N
Tsukamoto, Y
Honma, H
Kohri, K
Journal Article
United States
Biochem Biophys Res Commun. 2000 Feb 24;268(3):814-7. doi: 10.1006/bbrc.2000.2224.
Author Address: Suntory Institute for Bioorganic Research, Wakayama-dai, Shimamoto-cho, Mishima-gun, Osaka, 618, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 899
Author: Mateo, J., Kreda, S., Henry, C. E., Harden, T. K. and Boyer, J. L.
Year: 2003
Title: Requirement of Cys399 for processing of the human ecto-ATPase (NTPDase2) and its implications for determination of the activities of splice variants of the enzyme
Journal: J Biol Chem
Volume: 278
Issue: 41
Pages: 39960-8
Epub Date: 2003/07/31
Date: Oct 10
Short Title: Requirement of Cys399 for processing of the human ecto-ATPase (NTPDase2) and its implications for determination of the activities of splice variants of the enzyme
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M307854200
Accession Number: 12888562
Keywords: 3T3 Cells
Adenosine Triphosphatases/*chemistry/genetics/*metabolism
Alternative Splicing
Amino Acid Sequence
Animals
Asparagine/chemistry
Base Sequence
CHO Cells
Cell Line
Conserved Sequence
Cricetinae
Cysteine/chemistry
DNA, Complementary/genetics
Glycosylation
Humans
In Vitro Techniques
Mice
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Folding
Protein Processing, Post-Translational
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Homology, Amino Acid
Solubility
Abstract: Ecto-ATPase (CD39L1) corresponds to the type 2 enzyme of the ecto-nucleoside triphosphate diphosphohydrolase family (E-NTPDase). We have isolated from human ECV304 cells three cDNAs with high homology with members of the E-NTPDase family that encode predicted proteins of 495, 472, and 450 amino acids. Sequencing of a genomic DNA clone confirmed that these three sequences correspond to splice variants of the human ecto-ATPase (NTPDase2 alpha,-2 beta, and -2 gamma). Although all three enzyme forms were expressed heterologously to similar levels in Chinese hamster ovary cells clone K-1 (CHO-K1) cells, only the 495-amino acid protein (NTPDase2 alpha exhibited ecto-ATPase activity. Immunolocalization studies demonstrated that NTPDase2 alpha is fully processed and trafficked to the plasma membrane, whereas the NTPDase2 beta and -2 gamma splice variants were retained in not fully glycosylated forms in the endoplasmic reticulum. The potential roles of two highly conserved residues, Cys399 and Asn443, in the activity and cellular trafficking of the ecto-ATPase were examined. Mutation of Cys399, which is absent in NTPDase2 beta and -2 gamma, produced a protein completely devoid of nucleotidase activity, while mutation of Asn443 to Asp resulted in substantial loss of activity. Neither the Cys399 nor Asn443 mutants were fully glycosylated, and both were retained in the endoplasmic reticulum. These results indicate that the lack of ecto-nucleotidase activity exhibited by NTPDase2 beta and -2 gamma and the C399S mutant, as well as the large reduction of activity in the N443D mutant are due to alterations in the folding/maturation of these proteins.
Notes: Mateo, Jesús
Kreda, Silvia
Henry, Christopher E
Harden, T Kendall
Boyer, José L
GM38213/GM/NIGMS NIH HHS/United States
HL34322/HL/NHLBI NIH HHS/United States
HL54889/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2003 Oct 10;278(41):39960-8. doi: 10.1074/jbc.M307854200. Epub 2003 Jul 29.
Author Address: Department of Pharmacology, School of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599-7365, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1136
Author: Mathews, F. S., Gordon, M. M., Chen, Z., Rajashankar, K. R., Ealick, S. E., Alpers, D. H. and Sukumar, N.
Year: 2007
Title: Crystal structure of human intrinsic factor: cobalamin complex at 2.6-A resolution
Journal: Proc Natl Acad Sci U S A
Volume: 104
Issue: 44
Pages: 17311-6
Epub Date: 2007/10/24
Date: Oct 30
Short Title: Crystal structure of human intrinsic factor: cobalamin complex at 2.6-A resolution
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0703228104
PMCID: PMC2077253
Accession Number: 17954916
Keywords: Crystallography, X-Ray
Humans
Intrinsic Factor/*chemistry/genetics/*metabolism
Models, Molecular
Oncogene Protein v-cbl/chemistry/metabolism
Protein Binding
Protein Structure, Quaternary
Protein Structure, Tertiary
Static Electricity
Structural Homology, Protein
Vitamin B 12/*chemistry/*metabolism
Abstract: The structure of intrinsic factor (IF) in complex with cobalamin (Cbl) was determined at 2.6-A resolution. The overall fold of the molecule is that of an alpha(6)/alpha(6) barrel. It is a two-domain protein, and the Cbl is bound at the interface of the domains in a base-on conformation. Surprisingly, two full-length molecules, each comprising an alpha- and a beta-domain and one Cbl, and two truncated molecules with only an alpha- domain are present in the same asymmetric unit. The environment around Cbl is dominated by uncharged residues, and the sixth coordinate position of Co(2+) is empty. A detailed comparison between the IF-B12 complex and another Cbl transport protein complex, trans-Cbl-B12, has been made. The pH effect on the binding of Cbl analogues in transport proteins is analyzed. A possible basis for the lack of interchangeability of human and rat IF receptors is presented.
Notes: 1091-6490
Mathews, F S
Gordon, M M
Chen, Z
Rajashankar, K R
Ealick, S E
Alpers, D H
Sukumar, N
P41 RR015301/RR/NCRR NIH HHS/United States
F32 GM020530/GM/NIGMS NIH HHS/United States
RR-15301/RR/NCRR NIH HHS/United States
P DK01 DK-33487/DK/NIDDK NIH HHS/United States
P01 DK033487/DK/NIDDK NIH HHS/United States
GM20530/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2007 Oct 30;104(44):17311-6. doi: 10.1073/pnas.0703228104. Epub 2007 Oct 22.
Author Address: Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO 63110, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 285
Author: Matsui, T., Kihara, C., Fujimura, Y., Mizuochi, T. and Titani, K.
Year: 1991
Title: Carbohydrate analysis of human von Willebrand factor with horseradish peroxidase-conjugated lectins
Journal: Biochem Biophys Res Commun
Volume: 178
Issue: 3
Pages: 1253-9
Epub Date: 1991/08/15
Date: Aug 15
Short Title: Carbohydrate analysis of human von Willebrand factor with horseradish peroxidase-conjugated lectins
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(91)91028-b
Accession Number: 1872845
Keywords: Carbohydrates/analysis
Electrophoresis, Polyacrylamide Gel
Glycoproteins/blood
Glycoside Hydrolases
Horseradish Peroxidase
Humans
*Lectins
Molecular Weight
Oligosaccharides/chemistry/isolation & purification
Protein Binding
von Willebrand Factor/*chemistry/isolation & purification
Abstract: Human von Willebrand factor (vWF) immobilized on a polyvinylidene difluoride membrane was subjected to binding assay with a series of horseradish peroxidase-conjugated lectins. The protein was reactive with concanavalin A, Ricinus communis agglutinin 120, wheat germ agglutinin and Ulex europaeus agglutinin I (UEA-I) but not with peanut agglutinin before sialidase treatment. These reactivities were consistent with the major oligosaccharide structure reported except for UEA-I. The reactivity with UEA-I was greatly decreased after digestion of the protein with either alpha-L-fucosidase or peptide-N-glycosidase F, but no significant decrease was observed after mild alkaline treatment or delipidation. vWF and UEA-I have been independently used as a good marker for human endothelial cells. Our results indicate that vWF itself contains UEA-I reactive sugar chains in its Asn-linked oligosaccharides.
Notes: Matsui, T
Kihara, C
Fujimura, Y
Mizuochi, T
Titani, K
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 1991 Aug 15;178(3):1253-9. doi: 10.1016/0006-291x(91)91028-b.
Author Address: Division of Biomedical Polymer Science, Fujita Health University School of Medicine, Aichi, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 264
Author: Matsui, T., Titani, K. and Mizuochi, T.
Year: 1992
Title: Structures of the asparagine-linked oligosaccharide chains of human von Willebrand factor. Occurrence of blood group A, B, and H(O) structures
Journal: J Biol Chem
Volume: 267
Issue: 13
Pages: 8723-31
Epub Date: 1992/05/05
Date: May 5
Short Title: Structures of the asparagine-linked oligosaccharide chains of human von Willebrand factor. Occurrence of blood group A, B, and H(O) structures
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1577715
Keywords: ABO Blood-Group System/*chemistry
Asparagine/*chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Electrophoresis, Paper
Humans
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry
von Willebrand Factor/*chemistry
Abstract: The asparagine-linked oligosaccharide chains of human von Willebrand factor (vWF) purified from pooled plasma were quantitatively liberated from the polypeptide moiety by hydrazinolysis. After N-acetylation, these were fractionated by paper electrophoresis and sequential chromatography on lectin-affinity columns of concanavalin A, Phaseolus vulgaris erythrophytohemagglutinin, Datura stramonium agglutinin, Ricinus communis agglutinin 120, and Ulex europaeus agglutinin I and on a Bio-Gel P-4 column. Their structures were investigated by sequential exoglycosidase digestion in conjunction with methylation analysis. The glycoprotein was shown to be unique in its great diversity of oligosaccharide structures. Another noteworthy finding which had not been reported previously was the occurrence of asparagine-linked oligosaccharide chains with blood group A, B, and H(O) structures. In the present study, this glycoprotein was shown to contain mono- (0.4% of the total oligosaccharides), bi-(78.2%), tri- (12.3%), and tetraantennary (2.3%) complex type oligosaccharides in addition to a series of high mannose type oligosaccharides, Man6-9GlcNAc2 (0.8%). Biantennary complex type oligosaccharide chains were those with (8.2%) and without (70.0%) a bisecting GlcNAc residue and approximately 13.2%, 2.2%, and 0.4% of these contained blood group H(O), A, and B structures, respectively. The tri- and tetraantennary complex type chains were those with and without N-acetyllactosamine repeats, and about 13.0% of the triantennary chains without the N-acetyllactosamine repeat contained the blood group H(O) structure. Occurrence of these asparagine-linked oligosaccharides with blood group A and B structures suggest that the repeated use of factor VIII/vWF pooled concentrate for the treatment of hemophiliacs could result in the production of antibodies against vWF with a different blood group from that of the patient, and this development may be pathogenic.
Notes: Matsui, T
Titani, K
Mizuochi, T
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1992 May 5;267(13):8723-31.
Author Address: Division of Biomedical Polymer Science, Fujita Health University School of Medicine, Aichi, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 485
Author: Matsumoto, A., Yoshima, H., Takasaki, S. and Kobata, A.
Year: 1982
Title: Structural study of the sugar chains of human lactoferrin: finding of four novel complex-type asparagine-linked sugar chains
Journal: J Biochem
Volume: 91
Issue: 1
Pages: 143-55
Epub Date: 1982/01/01
Date: Jan
Short Title: Structural study of the sugar chains of human lactoferrin: finding of four novel complex-type asparagine-linked sugar chains
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a133671
Accession Number: 7068558
Keywords: *Asian Continental Ancestry Group
Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Gel
Electrophoresis, Paper
*European Continental Ancestry Group
Humans
Hydrazines
*Lactoferrin
*Lactoglobulins
Milk, Human/*analysis
Oligosaccharides
Sialic Acids
Abstract: Human lactoferrin contains 2 asparagine-linked sugar chains in 1 molecule. These sugar chains were released as oligosaccharides by hydrazinolysis from two lactoferrin samples of different races. The two oligosaccharide fractions gave exactly the same fractionation pattern upon paper electrophoresis and Bio-Gel P-4 column chromatography after sialidase digestion. A structural study of the oligosaccharides obtained from the two samples by sequential exoglycosidase digestion in combination with methylation analysis also gave the same results indicating that there is no racial difference both in quality and quantity of the sugar chain moiety of lactoferrin. In addition to the two acidic sugar chains, NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlucNAc and Gal beta 1 leads to 4(Fuc alpha 1 leads to 3)GlcNAc beta 1 leads to 1Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAc, and two novel acidic sugar chains, Gal beta 1 leads to 4(Fuc alpha 1 leads to 3)GlcNAc beta 1 leads to Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 3Gal beta 1 leads to 4GlcNAc beta 1 leads to 2 Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6(GlcNAc and Gal beta 1 leads to 4(Fuc alpha 1 leads to 3)GlcNAc beta 1 leads to 3Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAc, were found to occur in human lactoferrin.
Notes: Matsumoto, A
Yoshima, H
Takasaki, S
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
England
J Biochem. 1982 Jan;91(1):143-55. doi: 10.1093/oxfordjournals.jbchem.a133671.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 564
Author: Matsumoto, K., Maeda, Y., Kato, S. and Yuki, H.
Year: 1994
Title: Alteration of asparagine-linked glycosylation in serum transferrin of patients with hepatocellular carcinoma
Journal: Clin Chim Acta
Volume: 224
Issue: 1
Pages: 1-8
Epub Date: 1994/01/14
Date: Jan 14
Short Title: Alteration of asparagine-linked glycosylation in serum transferrin of patients with hepatocellular carcinoma
Alternate Journal: Clinica chimica acta; international journal of clinical chemistry
ISSN: 0009-8981 (Print)
0009-8981
DOI: 10.1016/0009-8981(94)90115-5
Accession Number: 8174273
Keywords: Asparagine/*metabolism
Biomarkers, Tumor
Carbohydrate Sequence
Carcinoma, Hepatocellular/*blood
Chromatography, High Pressure Liquid
Glucose/*metabolism
Glycoproteins/analysis/metabolism
Glycoside Hydrolases/chemistry
Humans
Hydrolysis
Liver Cirrhosis/blood
Liver Neoplasms/*blood
Molecular Sequence Data
Oligosaccharides
Transferrin/*metabolism
Abstract: The asparagine-linked sugar chains in serum transferrin purified from patients with hepatocellular carcinoma (n = 13), healthy individuals (n = 5) and patients with liver cirrhosis (n = 6) were compared. Sugar chains released with N-glycanase from desialylated and pepsin-digested transferrin were derivatized by reductive pyridylamination. Analysis of the sugar chains by high performance liquid chromatography in combination with exoglycosidase digestion revealed an increase of a biantennary complex-type sugar chain with a fucosylated trimannosyl core; Gal beta 1-4GlcNAc beta 1-2Man alpha 1-6(Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3) Man beta 1-4GlcNAc beta 1-4(Fuc alpha 1-6)GlcNAc in 7 of 13 cancer patients and an increase of a sugar chain with a fucosylated trimannosyl core and bisecting N-acetylglucosamine; Gal beta 1-4GlcNAc beta 1-2Man alpha 1-6(GlcNAc beta 1-4) (Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4(Fuc alpha 1-6)GlcNAc in one of the 13 cancer patients. Further, the fucosylated alteration of the sugar chain was detected also in alpha 1-antitrypsin, hemopexin, alpha 1-acid glycoprotein and alpha 2-HS glycoprotein from one of the patients with increased fucosylated transferrin.
Notes: Matsumoto, K
Maeda, Y
Kato, S
Yuki, H
Journal Article
Netherlands
Clin Chim Acta. 1994 Jan 14;224(1):1-8. doi: 10.1016/0009-8981(94)90115-5.
Author Address: School of Pharmaceutical Sciences, Toho University, Chiba, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 657
Author: Mattu, T. S., Pleass, R. J., Willis, A. C., Kilian, M., Wormald, M. R., Lellouch, A. C., Rudd, P. M., Woof, J. M. and Dwek, R. A.
Year: 1998
Title: The glycosylation and structure of human serum IgA1, Fab, and Fc regions and the role of N-glycosylation on Fcα receptor interactions
Journal: J Biol Chem
Volume: 273
Issue: 4
Pages: 2260-72
Epub Date: 1998/01/27
Date: Jan 23
Short Title: The glycosylation and structure of human serum IgA1, Fab, and Fc regions and the role of N-glycosylation on Fcα receptor interactions
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.4.2260
Accession Number: 9442070
Keywords: Amino Acid Sequence
Animals
Antigens, CD/*chemistry
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Glycosylation
Humans
Immunoglobulin A/*chemistry
Immunoglobulin Fab Fragments/*chemistry
Immunoglobulin Fc Fragments/*chemistry
Mice
Models, Molecular
Molecular Sequence Data
Polysaccharides/analysis
Receptors, Fc/*chemistry
Recombinant Proteins/chemistry
Sequence Alignment
Abstract: The human serum immunoglobulins IgG and IgA1 are produced in bone marrow and both interact with specific cellular receptors that mediate biological events. In contrast to IgA1, the glycosylation of IgG has been well characterized, and its interaction with various Fc receptors (Fc Rs) has been well studied. In this paper, we have analyzed the glycosylation of IgA1 and IgA1 Fab and Fc as well as three recombinant IgA1 molecules, including two N-glycosylation mutants. Amino acid sequencing data of the IgA1 Fc O-glycosylated hinge region indicated that O-glycans are located at Thr228, Ser230, and Ser232, while O-glycan sites at Thr225 and Thr236 are partially occupied. Over 90% of the N-glycans in IgA1 were sialylated, in contrast to IgG, where < 10% contain sialic acid. This paper contains the first report of Fab glycosylation in IgA1, and (in contrast to IgG Fab, which contains only N-linked glycans) both N- and O-linked oligosaccharides were identified. Analysis of the N-glycans attached to recombinant IgA1 indicated that the Cα 2 N-glycosylation site contained mostly biantennary glycans, while the tailpiece site, absent in IgG, contained mostly triantennary structures. Further analysis of these data suggested that processing at one Fc N-glycosylation site affects the other. Neutrophil Fcα R binding studies, using recombinant IgA1, indicated that neither the tailpiece region nor the N-glycans in the C alpha 2 domain contribute to IgA1-neutrophil Fcα R binding. This contrasts with IgG, where removal of the Fc N-glycans reduces binding to the Fcγ R. The primary sequence and disulfide bond pattern of IgA1, together with the crystal structures of IgG1 Fc and mouse IgA Fab and the glycan sequencing data, were used to generate a molecular model of IgA1. As a consequence of both the primary sequence and S-S bond pattern, the N-glycans in IgA1 Fc are not confined within the inter-α-chain space. The accessibility of the Cα 2 N-glycans provides an explanation for the increased sialylation and galactosylation of IgA1 Fc over that of IgG Fc N-glycans, which are confined in the space between the two Cγ 2 domains. This also suggests why in contrast to IgG Fc, the IgA1 N-glycans are not undergalactosylated in rheumatoid arthritis.
Notes: Mattu, T S
Pleass, R J
Willis, A C
Kilian, M
Wormald, M R
Lellouch, A C
Rudd, P M
Woof, J M
Dwek, R A
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1998 Jan 23;273(4):2260-72. doi: 10.1074/jbc.273.4.2260.
Author Address: Department of Biochemistry, Oxford University, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 780
Author: Mattu, T. S., Royle, L., Langridge, J., Wormald, M. R., Van den Steen, P. E., Van Damme, J., Opdenakker, G., Harvey, D. J., Dwek, R. A. and Rudd, P. M.
Year: 2000
Title: O-glycan analysis of natural human neutrophil gelatinase B using a combination of normal phase-HPLC and online tandem mass spectrometry: implications for the domain organization of the enzyme
Journal: Biochemistry
Volume: 39
Issue: 51
Pages: 15695-704
Epub Date: 2000/12/22
Date: Dec 26
Short Title: O-glycan analysis of natural human neutrophil gelatinase B using a combination of normal phase-HPLC and online tandem mass spectrometry: implications for the domain organization of the enzyme
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi001367j
Accession Number: 11123894
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid/methods
Computer Simulation
Fucose/chemistry
Glycoside Hydrolases/chemistry
Humans
Hydrolysis
Mass Spectrometry/methods
Matrix Metalloproteinase 9/blood/*chemistry
Models, Molecular
Neutrophils/*enzymology
Polysaccharides/blood/*chemistry
Protein Structure, Tertiary
Abstract: Gelatinase B is a matrix metalloproteinase (MMP-9) expressed under strict control by many cell types including neutrophils, monocytes, macrophages, and tumor cells. MMP-9 is a key mediator in the physiological maintenance of the extracellular matrix both in tissue remodeling and development, while uncontrolled enzyme activity contributes to pathologies such as cancer and inflammation. Neutrophils release MMP-9 from granules in response to IL-8 stimulation. Human MMP-9 has three potential N-linked glycosylation sites and contains a Ser/Pro/Thr rich domain, known as the type V collagen-like domain, which is expected to be heavily O-glycosylated. Indeed, approximately 85% of the total sugars on human neutrophil MMP-9 are O-linked. This paper presents the detailed analysis of picomole amounts of these O-glycans using a novel HPLC-based strategy for O-glycan analysis that provides linkage and arm specific information in addition to monosaccharide sequence. The initial structural assignments were confirmed using HPLC with online MS/MS fragmentation analysis. Twelve sugars were identified that contained from two to nine monosaccharide residues. Most of these contained type 2 core structures with Galbeta1-4GlcNAc (N-acetyl lactosamine) extensions, with or without sialic acid or fucose. The O-glycans were modeled using the oligosaccharide structural database. On the basis of the structure of gelatinase A (MMP-2), a model of MMP-9 suggests that the type V collagen-like domain in gelatinase B is located on a loop remote from the active site. Fourteen potential O-glycosylation sites are multiply presented on this loop of 52 amino acids. Many of the O-glycans identified contain terminal galactose residues that may provide recognition epitopes. Importantly, heavy glycosylation of this loop region, absent in gelatinase A, has considerable implications for the domain organization of MMP-9.
Notes: Mattu, T S
Royle, L
Langridge, J
Wormald, M R
Van den Steen, P E
Van Damme, J
Opdenakker, G
Harvey, D J
Dwek, R A
Rudd, P M
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2000 Dec 26;39(51):15695-704. doi: 10.1021/bi001367j.
Author Address: Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1706
Author: Maury, J. J., El Farran, C. A., Ng, D., Loh, Y. H., Bi, X., Bardor, M. and Choo, A. B.
Year: 2015
Title: RING1B O-GlcNAcylation regulates gene targeting of polycomb repressive complex 1 in human embryonic stem cells
Journal: Stem Cell Res
Volume: 15
Issue: 1
Pages: 182-9
Epub Date: 2015/06/24
Date: Jul
Short Title: RING1B O-GlcNAcylation regulates gene targeting of polycomb repressive complex 1 in human embryonic stem cells
Alternate Journal: Stem cell research
ISSN: 1873-5061
DOI: 10.1016/j.scr.2015.06.007
Accession Number: 26100231
Keywords: Amino Acid Sequence
Cell Differentiation
Chromosomal Proteins, Non-Histone/metabolism
DNA/metabolism
*Gene Targeting
Glycosylation
Heterogeneous-Nuclear Ribonucleoprotein K/metabolism
Human Embryonic Stem Cells/cytology/*metabolism
Humans
Molecular Sequence Data
Polycomb Repressive Complex 1/chemistry/*metabolism
Protein Binding
Abstract: O-linked-N-acetylglucosamine (O-GlcNAc) post-translationally modifies and regulates thousands of proteins involved in various cellular mechanisms. Recently, O-GlcNAc has been linked to human embryonic stem cells (hESC) differentiation, however the identity and function of O-GlcNAc proteins regulating hESC remain unknown. Here, we firstly identified O-GlcNAc modified human stem cell regulators such as hnRNP K, HP1γ, and especially RING1B/RNF2. Thereafter, we focused our work on RING1B which is the catalytic subunit of the polycomb repressive complex 1 (PRC1) a major epigenetic repressor essential for pluripotency maintenance and differentiation. By point-mutation, we show that T(250)/S(251) and S(278) RING1B residues are bearing O-GlcNAc, and that T(250)/S(251) O-GlcNAcylation decreases during differentiation. O-GlcNAc seems to regulate RING1B-DNA binding as suggested by our ChIP-sequencing results. Non-O-GlcNAcylated RING1B is found to be enriched near cell cycle genes whereas O-GlcNAcylated RING1B seems preferentially enriched near neuronal genes. Our data suggest that during hESC differentiation, the decrease of RING1B O-GlcNAcylation might enable PRC1 to switch its target to induce neuron differentiation. Overall, we demonstrate that O-GlcNAc modifies and regulates an essential epigenetic tool, RING1B, which may contribute to hESC pluripotency maintenance and differentiation.
Notes: 1876-7753
Maury, Julien Jean Pierre
El Farran, Chadi A
Ng, Daniel
Loh, Yuin-Han
Bi, Xuezhi
Bardor, Muriel
Choo, Andre Boon-Hwa
Journal Article
Research Support, Non-U.S. Gov't
England
Stem Cell Res. 2015 Jul;15(1):182-9. doi: 10.1016/j.scr.2015.06.007. Epub 2015 Jun 17.
Author Address: Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), 20 Biopolis Way, #06-01 Centros, 138668 Singapore; Department of Biomedical Engineering, Faculty of Engineering, National University of Singapore, 9 Engineering Drive 1, 117575 Singapore; Institute of Molecular and Cell Biology, Agency for Science, Technology and Research (A*STAR), 61 Biopolis Drive, #08-01 Proteos, 138673 Singapore. Electronic address: jpmaury@imcb.a-star.edu.sg.
Institute of Molecular and Cell Biology, Agency for Science, Technology and Research (A*STAR), 61 Biopolis Drive, #08-01 Proteos, 138673 Singapore; Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, 117543 Singapore.
Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), 20 Biopolis Way, #06-01 Centros, 138668 Singapore.
Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), 20 Biopolis Way, #06-01 Centros, 138668 Singapore; Université de Rouen, Normandie Université, Laboratoire Glycobiologie et Matrice Extracellulaire Végétale (Glyco-MEV) EA 4358, IRIB, VASI, Faculté des Sciences et Techniques, Mont-Saint-Aignan Cédex 76821, France.
Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), 20 Biopolis Way, #06-01 Centros, 138668 Singapore; Department of Biomedical Engineering, Faculty of Engineering, National University of Singapore, 9 Engineering Drive 1, 117575 Singapore. Electronic address: andre_choo@bti.a-star.edu.sg.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1600
Author: Maury, J. J., Ng, D., Bi, X., Bardor, M. and Choo, A. B.
Year: 2014
Title: Multiple reaction monitoring mass spectrometry for the discovery and quantification of O-GlcNAc-modified proteins
Journal: Anal Chem
Volume: 86
Issue: 1
Pages: 395-402
Epub Date: 2013/10/23
Date: Jan 7
Short Title: Multiple reaction monitoring mass spectrometry for the discovery and quantification of O-GlcNAc-modified proteins
Alternate Journal: Analytical chemistry
ISSN: 0003-2700
DOI: 10.1021/ac401821d
Accession Number: 24144119
Keywords: Acetylglucosamine/*analysis/genetics
Amino Acid Sequence
Cyclic AMP Response Element-Binding Protein/*analysis/genetics
Embryonic Stem Cells/chemistry/physiology
Humans
Mass Spectrometry/*methods
Molecular Sequence Data
Protein Processing, Post-Translational/genetics
Abstract: O-linked N-acetylglucosamine (O-GlcNAc) is a post-translational modification regulating proteins involved in a variety of cellular processes and diseases. Unfortunately, O-GlcNAc remains challenging to detect and quantify by shotgun mass spectrometry (MS) where it is time-consuming and tedious. Here, we investigate the potential of Multiple Reaction Monitoring Mass Spectrometry (MRM-MS), a targeted MS method, to detect and quantify native O-GlcNAc modified peptides without extensive labeling and enrichment. We report the ability of MRM-MS to detect a standard O-GlcNAcylated peptide and show that the method is robust to quantify the amount of O-GlcNAcylated peptide with a method detection limit of 3 fmol. In addition, when diluted by 100-fold in a trypsin-digested whole cell lysate, the O-GlcNAcylated peptide remains detectable. Next, we apply this strategy to study glycogen synthase kinase-3 beta (GSK-3β), a kinase able to compete with O-GlcNAc transferase and modify identical site on proteins. We demonstrate that GSK-3β is itself modified by O-GlcNAc in human embryonic stem cells (hESC). Indeed, by only using gel electrophoresis to grossly enrich GSK-3β from whole cell lysate, we discover by MRM-MS a novel O-GlcNAcylated GSK-3β peptide, bearing 3 potential O-GlcNAcylation sites. We confirm our finding by quantifying the increase of O-GlcNAcylation, following hESC treatment with an O-GlcNAc hydrolase inhibitor. This novel O-GlcNAcylation could potentially be involved in an autoinhibition mechanism. To the best of our knowledge, this is the first report utilizing MRM-MS to detect native O-GlcNAc modified peptides. This could potentially facilitate rapid discovery and quantification of new O-GlcNAcylated peptides/proteins.
Notes: 1520-6882
Maury, Julien Jean Pierre
Ng, Daniel
Bi, Xuezhi
Bardor, Muriel
Choo, Andre Boon-Hwa
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Chem. 2014 Jan 7;86(1):395-402. doi: 10.1021/ac401821d. Epub 2013 Dec 16.
Author Address: Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR) , 20 Biopolis Way, #06-01 Centros, Singapore 138668.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 798
Author: Max, M., Shanker, Y. G., Huang, L., Rong, M., Liu, Z., Campagne, F., Weinstein, H., Damak, S. and Margolskee, R. F.
Year: 2001
Title: Tas1r3, encoding a new candidate taste receptor, is allelic to the sweet responsiveness locus Sac
Journal: Nat Genet
Volume: 28
Issue: 1
Pages: 58-63
Epub Date: 2001/04/28
Date: May
Short Title: Tas1r3, encoding a new candidate taste receptor, is allelic to the sweet responsiveness locus Sac
Alternate Journal: Nature genetics
ISSN: 1061-4036 (Print)
1061-4036
DOI: 10.1038/ng0501-58
Accession Number: 11326277
Keywords: Alleles
Amino Acid Sequence
Animals
Chromosome Mapping
Chromosomes/genetics
Humans
Isoenzymes/isolation & purification
Mice
Models, Molecular
Molecular Sequence Data
Phospholipase C beta
Receptors, AMPA
Receptors, Calcium-Sensing
Receptors, Cell Surface/*genetics
*Receptors, G-Protein-Coupled
Sequence Homology, Amino Acid
Species Specificity
*Sweetening Agents
Taste/*genetics
Taste Buds/chemistry/ultrastructure
Transducin/isolation & purification
Type C Phospholipases/isolation & purification
Abstract: The ability to taste the sweetness of carbohydrate-rich foodstuffs has a critical role in the nutritional status of humans. Although several components of bitter transduction pathways have been identified, the receptors and other sweet transduction elements remain unknown. The Sac locus in mouse, mapped to the distal end of chromosome 4 (refs. 7-9), is the major determinant of differences between sweet-sensitive and -insensitive strains of mice in their responsiveness to saccharin, sucrose and other sweeteners. To identify the human Sac locus, we searched for candidate genes within a region of approximately one million base pairs of the sequenced human genome syntenous to the region of Sac in mouse. From this search, we identified a likely candidate: T1R3, a previously unknown G protein-coupled receptor (GPCR) and the only GPCR in this region. Mouse Tas1r3 (encoding T1r3) maps to within 20,000 bp of the marker closest to Sac (ref. 9) and, like human TAS1R3, is expressed selectively in taste receptor cells. By comparing the sequence of Tas1r3 from several independently derived strains of mice, we identified a specific polymorphism that assorts between taster and non-taster strains. According to models of its structure, T1r3 from non-tasters is predicted to have an extra amino-terminal glycosylation site that, if used, would interfere with dimerization.
Notes: Max, M
Shanker, Y G
Huang, L
Rong, M
Liu, Z
Campagne, F
Weinstein, H
Damak, S
Margolskee, R F
DC00310/DC/NIDCD NIH HHS/United States
DC03155/DC/NIDCD NIH HHS/United States
MH57241/MH/NIMH NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Nat Genet. 2001 May;28(1):58-63. doi: 10.1038/ng0501-58.
Author Address: Department of Physiology and Biophysics, Mount Sinai School of Medicine of New York University, New York, New York, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 286
Author: May, L. T., Shaw, J. E., Khanna, A. K., Zabriskie, J. B. and Sehgal, P. B.
Year: 1991
Title: Marked cell-type-specific differences in glycosylation of human interleukin-6
Journal: Cytokine
Volume: 3
Issue: 3
Pages: 204-11
Epub Date: 1991/05/01
Date: May
Short Title: Marked cell-type-specific differences in glycosylation of human interleukin-6
Alternate Journal: Cytokine
ISSN: 1043-4666 (Print)
1043-4666
DOI: 10.1016/1043-4666(91)90018-9
Accession Number: 1883960
Keywords: Amino Acid Sequence
Animals
B-Lymphocytes/immunology
Blotting, Western
Cell Line
Chromatography, Affinity
Fibroblasts/immunology
Glycoside Hydrolases
Glycosylation
Humans
Insecta
Interleukin-6/biosynthesis/*genetics/isolation & purification
Molecular Sequence Data
Molecular Weight
*Protein Processing, Post-Translational
Recombinant Proteins/biosynthesis/isolation & purification
Abstract: Human interleukin-6 (IL-6) secreted by cytokine- or endotoxin-induced fibroblasts, monocytes, keratinocytes, endometrial stromal cells, and endothelial cells, when analyzed under denaturing and reducing conditions, consists of a set of differentially modified phosphoglycoproteins of molecular mass in the range from 23 to 30 kD (a set of at least three O-glycosylated 23- to 25-kD species and a set of at least three N- and O-glycosylated 28- to 30-kD species). The 23- to 25-kD and 28- to 30-kD fibroblast-derived IL-6 species have been separately purified to homogeneity with the use of a combination of lectin and immunoaffinity chromatography. Glycosidase digestion experiments on such purified preparations confirmed that almost all human fibroblast-derived IL-6 species were O-glycosylated; additionally, the 28- to 30-kD species were N-glycosylated. Amino acid sequencing revealed that the major amino terminus in the fibroblast-derived 23- to 25-kD O-glycosylated IL-6 was at Ala28 whereas the major amino terminus in the 28- to 30-kD N- and O-glycosylated IL-6 was at Val30, suggesting that targeting of newly synthesized IL-6 polypeptides into the two different processing pathways in fibroblasts may be keyed to differences in the signal peptide cleavage site. Unexpectedly, IL-6 "constitutively" secreted by the Epstein-Barr virus (EBV)-infected human and primate (tamarin) B-cell lines designated sfBJAB and sfBT, respectively, consisted of a major apparently unglycosylated 21-kD species and a minor 25-kD N-glycosylated species.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: May, L T
Shaw, J E
Khanna, A K
Zabriskie, J B
Sehgal, P B
AI-16262/AI/NIAID NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Cytokine. 1991 May;3(3):204-11. doi: 10.1016/1043-4666(91)90018-9.
Author Address: Rockefeller University, New York, NY 10021.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1610
Author: Mayampurath, A., Yu, C. Y., Song, E., Balan, J., Mechref, Y. and Tang, H.
Year: 2014
Title: Computational framework for identification of intact glycopeptides in complex samples
Journal: Anal Chem
Volume: 86
Issue: 1
Pages: 453-63
Epub Date: 2013/11/28
Date: Jan 7
Short Title: Computational framework for identification of intact glycopeptides in complex samples
Alternate Journal: Analytical chemistry
ISSN: 0003-2700
DOI: 10.1021/ac402338u
Accession Number: 24279413
Keywords: Amino Acid Sequence
Animals
Cattle
Computational Biology/*methods
Glycopeptides/analysis/*blood/*genetics
Humans
Molecular Sequence Data
Swine
Abstract: Glycosylation is an important protein modification that involves enzymatic attachment of sugars to amino acid residues. Understanding the structure of these sugars and the effects of glycosylation are vital for developing indicators of disease development and progression. Although computational methods based on mass spectrometric data have proven to be effective in monitoring changes in the glycome, developing such methods for the glycoproteome are challenging, largely due to the inherent complexity in simultaneously studying glycan structures with their corresponding glycosylation sites. This paper introduces a computational framework for identifying intact N-linked glycopeptides, i.e. glycopeptides with N-linked glycans attached to their glycosylation sites, in complex proteome samples. Scoring algorithms are presented for tandem mass spectra of glycopeptides resulting from collision-induced dissociation (CID), higher-energy C-trap dissociation (HCD), and electron transfer dissociation (ETD) fragmentation modes. An empirical false-discovery rate estimation method, based on a target-decoy search approach, is derived for assigning confidence. The power of our method is further enhanced when multiple data sets are pooled together to increase identification confidence. Using this framework, 103 highly confident N-linked glycopeptides from 53 sites across 33 glycoproteins were identified in complex human serum proteome samples using conventional proteomic platforms with standard depletion of the 7-most abundant proteins. These results indicate that our method is ready to be used for characterizing site-specific protein glycosylation in complex samples.
Notes: 1520-6882
Mayampurath, Anoop
Yu, Chuan-Yih
Song, Ehwang
Balan, Jagadheshwar
Mechref, Yehia
Tang, Haixu
1R01GM093322-03/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
Anal Chem. 2014 Jan 7;86(1):453-63. doi: 10.1021/ac402338u. Epub 2013 Dec 10.
Author Address: School of Informatics & Computing, Indiana University , Bloomington, Indiana 47408, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1219
Author: McElroy, C. A., Dohm, J. A. and Walsh, S. T.
Year: 2009
Title: Structural and biophysical studies of the human IL-7/IL-7Ralpha complex
Journal: Structure
Volume: 17
Issue: 1
Pages: 54-65
Epub Date: 2009/01/15
Date: Jan 14
Short Title: Structural and biophysical studies of the human IL-7/IL-7Ralpha complex
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2008.10.019
PMCID: PMC2654238
NIHMSID: NIHMS92173
Accession Number: 19141282
Keywords: Biophysics
Biosensing Techniques
Enzyme-Linked Immunosorbent Assay
Glycosylation
Humans
Interleukin-7/*chemistry
Interleukin-7 Receptor alpha Subunit/*chemistry
Protein Conformation
Abstract: IL-7 and IL-7Ralpha bind the gamma(c) receptor, forming a complex crucial to several signaling cascades leading to the development and homeostasis of T and B cells. We report that the IL-7Ralpha ectodomain uses glycosylation to modulate its binding constants to IL-7, unlike the other receptors in the gamma(c) family. IL-7 binds glycosylated IL-7Ralpha 300-fold more tightly than unglycosylated IL-7Ralpha, and the enhanced affinity is attributed primarily to an accelerated on rate. Structural comparison of IL-7 in complex to both forms of IL-7Ralpha reveals that glycosylation does not participate directly in the binding interface. The SCID mutations of IL-7Ralpha locate outside the binding interface with IL-7, suggesting that the expressed mutations cause protein folding defects in IL-7Ralpha. The IL-7/IL-7Ralpha structures provide a window into the molecular recognition events of the IL-7 signaling cascade and provide sites to target for designing new therapeutics to treat IL-7-related diseases.
Notes: McElroy, Craig A
Dohm, Julie A
Walsh, Scott T R
R01 AI072142/AI/NIAID NIH HHS/United States
R01 AI072142-01A2/AI/NIAID NIH HHS/United States
R56 AI072142/AI/NIAID NIH HHS/United States
AI72142/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Structure. 2009 Jan 14;17(1):54-65. doi: 10.1016/j.str.2008.10.019.
Author Address: Department of Molecular and Cellular Biochemistry, College of Medicine, Comprehensive Cancer Center, Ohio State University, 467 Hamilton Hall, 1645 Neil Avenue, Columbus, OH 43210, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1425
Author: McEwan, P. A., Yang, W., Carr, K. H., Mo, X., Zheng, X., Li, R. and Emsley, J.
Year: 2011
Title: Quaternary organization of GPIb-IX complex and insights into Bernard-Soulier syndrome revealed by the structures of GPIbβ and a GPIbβ/GPIX chimera
Journal: Blood
Volume: 118
Issue: 19
Pages: 5292-301
Epub Date: 2011/09/13
Date: Nov 10
Short Title: Quaternary organization of GPIb-IX complex and insights into Bernard-Soulier syndrome revealed by the structures of GPIbβ and a GPIbβ/GPIX chimera
Alternate Journal: Blood
ISSN: 0006-4971 (Print)
0006-4971
DOI: 10.1182/blood-2011-05-356253
PMCID: PMC3217411
Accession Number: 21908432
Keywords: Amino Acid Sequence
Animals
Bernard-Soulier Syndrome/*blood/*genetics
CHO Cells
Cricetinae
Cricetulus
Crystallography, X-Ray
Humans
Models, Molecular
Molecular Sequence Data
Mutant Proteins/chemistry/genetics
Mutation, Missense
Platelet Glycoprotein GPIb-IX Complex/*chemistry/*genetics
Protein Structure, Quaternary
Protein Structure, Tertiary
Recombinant Fusion Proteins/chemistry/genetics
Sequence Homology, Amino Acid
Abstract: Platelet GPIb-IX receptor complex has 3 subunits GPIbα, GPIbβ, and GPIX, which assemble with a ratio of 1:2:1. Dysfunction in surface expression of the complex leads to Bernard-Soulier syndrome. We have crystallized the GPIbβ ectodomain (GPIbβ(E)) and determined the structure to show a single leucine-rich repeat with N- and C-terminal disulphide-bonded capping regions. The structure of a chimera of GPIbβ(E) and 3 loops (a,b,c) taken from the GPIX ectodomain sequence was also determined. The chimera (GPIbβ(Eabc)), but not GPIbβ(E), forms a tetramer in the crystal, showing a quaternary interface between GPIbβ and GPIX. Central to this interface is residue Tyr106 from GPIbβ, which inserts into a pocket generated by 2 loops (b,c) from GPIX. Mutagenesis studies confirmed this interface as a valid representation of interactions between GPIbβ and GPIX in the full-length complex. Eight GPIbβ missense mutations identified from patients with Bernard-Soulier syndrome were examined for changes to GPIb-IX complex surface expression. Two mutations, A108P and P74R, were found to maintain normal secretion/folding of GPIbβ(E) but were unable to support GPIX surface expression. The close structural proximity of these mutations to Tyr106 and the GPIbβ(E) interface with GPIX indicates they disrupt the quaternary organization of the GPIb-IX complex.
Notes: 1528-0020
McEwan, Paul A
Yang, Wenjun
Carr, Katherine H
Mo, Xi
Zheng, Xiaofeng
Li, Renhao
Emsley, Jonas
R01 HL082808/HL/NHLBI NIH HHS/United States
C21418/A8573/Cancer Research UK/United Kingdom
RG/07/002/23 132/British Heart Foundation/United Kingdom
RG/07/002/23132/British Heart Foundation/United Kingdom
HL082808/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Blood. 2011 Nov 10;118(19):5292-301. doi: 10.1182/blood-2011-05-356253. Epub 2011 Sep 8.
Author Address: Centre for Biomolecular Sciences, School of Pharmacy, University of Nottingham, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1267
Author: McGrath, A. P., Hilmer, K. M., Collyer, C. A., Shepard, E. M., Elmore, B. O., Brown, D. E., Dooley, D. M. and Guss, J. M.
Year: 2009
Title: Structure and inhibition of human diamine oxidase
Journal: Biochemistry
Volume: 48
Issue: 41
Pages: 9810-22
Epub Date: 2009/09/22
Date: Oct 20
Short Title: Structure and inhibition of human diamine oxidase
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi9014192
PMCID: PMC2791411
NIHMSID: NIHMS158430
Accession Number: 19764817
Keywords: Amine Oxidase (Copper-Containing)/antagonists &
inhibitors/*chemistry/genetics/metabolism
Animals
Binding Sites
Calcium/metabolism
Copper/metabolism
Crystallography, X-Ray
Dimerization
Diminazene/analogs & derivatives/metabolism
Drosophila/enzymology
Humans
Kinetics
Metallothionein/genetics
Models, Molecular
Pentamidine/metabolism
Promoter Regions, Genetic
Protein Conformation
Substrate Specificity
X-Ray Diffraction
Abstract: Humans have three functioning genes that encode copper-containing amine oxidases. The product of the AOC1 gene is a so-called diamine oxidase (hDAO), named for its substrate preference for diamines, particularly histamine. hDAO has been cloned and expressed in insect cells and the structure of the native enzyme determined by X-ray crystallography to a resolution of 1.8 A. The homodimeric structure has the archetypal amine oxidase fold. Two active sites, one in each subunit, are characterized by the presence of a copper ion and a topaquinone residue formed by the post-translational modification of a tyrosine. Although hDAO shares 37.9% sequence identity with another human copper amine oxidase, semicarbazide sensitive amine oxidase or vascular adhesion protein-1, its substrate binding pocket and entry channel are distinctly different in accord with the different substrate specificities. The structures of two inhibitor complexes of hDAO, berenil and pentamidine, have been refined to resolutions of 2.1 and 2.2 A, respectively. They bind noncovalently in the active-site channel. The inhibitor binding suggests that an aspartic acid residue, conserved in all diamine oxidases but absent from other amine oxidases, is responsible for the diamine specificity by interacting with the second amino group of preferred diamine substrates.
Notes: 1520-4995
McGrath, Aaron P
Hilmer, Kimberly M
Collyer, Charles A
Shepard, Eric M
Elmore, Bradley O
Brown, Doreen E
Dooley, David M
Guss, J Mitchell
R01 GM027659-29/GM/NIGMS NIH HHS/United States
GM 27659/GM/NIGMS NIH HHS/United States
R01 GM027659-27/GM/NIGMS NIH HHS/United States
R01 GM027659-28/GM/NIGMS NIH HHS/United States
R01 GM027659/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochemistry. 2009 Oct 20;48(41):9810-22. doi: 10.1021/bi9014192.
Author Address: School of Molecular and Microbial Biosciences, University of Sydney, Sydney, NSW 2006, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 620
Author: McGuire, J. M., Douglas, M. and Smith, K. D.
Year: 1996
Title: The resolution of the neutral N-linked oligosaccharides of IgG by high pH anion-exchange chromatography
Journal: Carbohydr Res
Volume: 292
Pages: 1-9
Epub Date: 1996/10/04
Date: Oct 4
Short Title: The resolution of the neutral N-linked oligosaccharides of IgG by high pH anion-exchange chromatography
Alternate Journal: Carbohydrate research
ISSN: 0008-6215 (Print)
0008-6215
DOI: 10.1016/s0008-6215(96)91015-0
Accession Number: 8870235
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Ion Exchange/methods
Chymotrypsin/metabolism
Hydrogen-Ion Concentration
Immunoglobulin G/*chemistry
Molecular Sequence Data
N-Acetylneuraminic Acid/analysis
Oligosaccharides/chemistry/*isolation & purification
Trypsin/metabolism
Abstract: The introduction of high pH anion-exchange chromatography (HPAEC) has represented a major development in the qualitative analysis of glycoprotein derived oligosaccharides. When coupled with pulsed amperometric detection, the technique permits the detection of picomole quantities of heterogeneous mixtures of oligosaccharide without the need for derivatisation. The applications of HPAEC have generally been limited to the analysis of sialylated oligosaccharides, however, it is now possible to analyse heterogenous mixtures of neutral oligosaccharides with the latest systems. We have used such a system to separate completely a panel of seven commercially available neutral N-linked oligosaccharides and found the influence of monosaccharide substitution on elution position to be identical to that for sialylated structures. A standard monosialylated N-linked oligosaccharide was modified by sequential digestion with specific exoglycosidases to produce a monogalactosylated, diantennary oligosaccharide which is commercially unavailable. This standard's elution position was confirmed by HPAEC. The technique was applied to the identification of neutral N-linked oligosaccharides released from human immunoglobulin G using the enzyme peptide-N-glycosidase F.
Notes: McGuire, J M
Douglas, M
Smith, K D
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Carbohydr Res. 1996 Oct 4;292:1-9. doi: 10.1016/s0008-6215(96)91015-0.
Author Address: Department of Pharmaceutical Sciences, University of Strathclyde, Glasgow, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1841
Author: McMillan, B. J., Zimmerman, B., Egan, E. D., Lofgren, M., Xu, X., Hesser, A. and Blacklow, S. C.
Year: 2017
Title: Structure of human POFUT1, its requirement in ligand-independent oncogenic Notch signaling, and functional effects of Dowling-Degos mutations
Journal: Glycobiology
Volume: 27
Issue: 8
Pages: 777-786
Epub Date: 2017/03/24
Date: Aug 1
Short Title: Structure of human POFUT1, its requirement in ligand-independent oncogenic Notch signaling, and functional effects of Dowling-Degos mutations
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwx020
PMCID: PMC5881682
Accession Number: 28334865
Keywords: Carcinogenesis/genetics/metabolism
Fucosyltransferases/*chemistry/deficiency/*genetics/metabolism
Humans
Hyperpigmentation/*genetics/metabolism
Ligands
*Mutation
Protein Conformation
Receptors, Notch/*genetics/*metabolism
Signal Transduction/*genetics
Skin Diseases, Genetic/*genetics/metabolism
Skin Diseases, Papulosquamous/*genetics/metabolism
Abstract: Protein O-fucosyltransferase-1 (POFUT1), which transfers fucose residues to acceptor sites on serine and threonine residues of epidermal growth factor-like repeats of recipient proteins, is essential for Notch signal transduction in mammals. Here, we examine the consequences of POFUT1 loss on the oncogenic signaling associated with certain leukemia-associated mutations of human Notch1, report the structures of human POFUT1 in free and GDP-fucose bound states, and assess the effects of Dowling-Degos mutations on human POFUT1 function. CRISPR-mediated knockout of POFUT1 in U2OS cells suppresses both normal Notch1 signaling, and the ligand-independent signaling associated with leukemogenic mutations of Notch1. Normal and oncogenic signaling are rescued by wild-type POFUT1 but rescue is impaired by an active-site R240A mutation. The overall structure of the human enzyme closely resembles that of the Caenorhabditis elegans protein, with an overall backbone RMSD of 0.93 Å, despite primary sequence identity of only 39% in the mature protein. GDP-fucose binding to the human enzyme induces limited backbone conformational movement, though the side chains of R43 and D244 reorient to make direct contact with the fucose moiety in the complex. The reported Dowling-Degos mutations of POFUT1, except for M262T, fail to rescue Notch1 signaling efficiently in the CRISPR-engineered POFUT1-/- background. Together, these studies identify POFUT1 as a potential target for cancers driven by Notch1 mutations and provide a structural roadmap for its inhibition.
Notes: 1460-2423
McMillan, Brian J
Zimmerman, Brandon
Egan, Emily D
Lofgren, Michael
Xu, Xiang
Hesser, Anthony
Blacklow, Stephen C
P41 GM103403/GM/NIGMS NIH HHS/United States
R01 CA092433/CA/NCI NIH HHS/United States
S10 RR029205/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Glycobiology. 2017 Aug 1;27(8):777-786. doi: 10.1093/glycob/cwx020.
Author Address: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
Department of Cancer Biology, Dana Farber Cancer Institute, Boston, MA 02215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 425
Author: McMullen, B. A. and Fujikawa, K.
Year: 1985
Title: Amino acid sequence of the heavy chain of human alpha-factor XIIa (activated Hageman factor)
Journal: J Biol Chem
Volume: 260
Issue: 9
Pages: 5328-41
Epub Date: 1985/05/10
Date: May 10
Short Title: Amino acid sequence of the heavy chain of human alpha-factor XIIa (activated Hageman factor)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3886654
Keywords: Amino Acid Sequence
Chromatography, Gel
Chromatography, High Pressure Liquid
Chymotrypsin/metabolism
Cyanogen Bromide/pharmacology
Endopeptidases/metabolism
Factor XII/*analysis
Factor XIIa
Humans
Peptide Fragments/*analysis
Protein Conformation
*Serine Endopeptidases
Trypsin/metabolism
Abstract: The amino acid sequence of the heavy chain of human alpha-factor XIIa (activated Hageman factor) was determined by automated Edman degradation using the peptides produced by chemical and enzymatic cleavages of intact factor XII and alpha-factor XIIa. Combining this sequence with the previously determined sequence of beta-factor XIIa (Fujikawa, K., and McMullen, B. A. (1983) J. Biol. Chem. 258, 10924-10933), the complete amino acid sequence of human factor XII has been established. The heavy chain of alpha-factor XIIa is composed of 353 amino acid residues containing one Asn-linked and six probable O-linked carbohydrate chains. The heavy chain of alpha-factor XIIa appears to contain four different domains including a "kringle," a "growth factor" domain, and the "type I" and "type II" domains of fibronectin. The domain organization of factor XII is analogous to those of several fibrinolytic proteins, including tissue plasminogen activator and urokinase, suggesting that factor XII belongs to the same protease subfamily as these two proteins.
Notes: McMullen, B A
Fujikawa, K
HL 16919/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1985 May 10;260(9):5328-41.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 300
Author: McMullen, B. A., Fujikawa, K. and Davie, E. W.
Year: 1991
Title: Location of the disulfide bonds in human coagulation factor XI: the presence of tandem apple domains
Journal: Biochemistry
Volume: 30
Issue: 8
Pages: 2056-60
Epub Date: 1991/02/26
Date: Feb 26
Short Title: Location of the disulfide bonds in human coagulation factor XI: the presence of tandem apple domains
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00222a008
Accession Number: 1998667
Keywords: Amino Acid Sequence
Chromatography, High Pressure Liquid
Cyanogen Bromide
Disulfides/analysis
Factor XI/*chemistry/isolation & purification
Humans
Molecular Sequence Data
Molecular Weight
Peptide Fragments/isolation & purification
Protein Conformation
Trypsin
Abstract: Factor XI is a plasma glycoprotein that participates in the blood coagulation cascade. Of the 19 disulfide bonds present in each of the subunits of the human protein, 16 were determined by amino acid sequence analysis of peptide fragments produced by chemical and enzymatic digestion. Four apple domains of 90 or 91 amino acids were identified in the tandem repeats present in the amino-terminal portion of each subunit of factor XI. The disulfide bonds in the carboxyl-terminal portion of the molecule were similar to those in the catalytic region of other serine proteases. The two identical subunits of factor XI were connected by a single disulfide bond at Cys321 linking each of the fourth apple domains while each of the Cys residues at position 11 in the first apple domains forms a disulfide bond with another Cys residue.
Notes: McMullen, B A
Fujikawa, K
Davie, E W
HL16919/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1991 Feb 26;30(8):2056-60. doi: 10.1021/bi00222a008.
Author Address: Department of Biochemistry, University of Washington, Seattle 98195.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 700
Author: Mechref, Y., Chen, P. and Novotny, M. V.
Year: 1999
Title: Structural characterization of the N-linked oligosaccharides in bile salt-stimulated lipase originated from human breast milk
Journal: Glycobiology
Volume: 9
Issue: 3
Pages: 227-34
Epub Date: 1999/02/20
Date: Mar
Short Title: Structural characterization of the N-linked oligosaccharides in bile salt-stimulated lipase originated from human breast milk
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/9.3.227
Accession Number: 10024660
Keywords: Carbohydrate Sequence
Female
Glycoproteins/*chemistry
Humans
Milk, Human/*enzymology
Molecular Sequence Data
Oligosaccharides/*chemistry
Sequence Analysis
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Sterol Esterase/*chemistry
alpha-Fetoproteins/chemistry
Abstract: The detailed structures of N- glycans derived from bile salt-stimulated lipase (BSSL) found in human milk were determined by combining exoglycosidase digestion with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The N- glycan structures were conclusively determined in terms of complexity and degree of fucosylation. Ion-exchange chromatography with pulsed amperometric detection, together with mass-spectral analysis of the esterified N- glycans, indicated the presence of monosialylated structures. The molecular mass profile of esterified N- glycans present in BSSL further permitted the more detailed studies through collision-induced dissociation (CID) and sequential exoglycosidase cleavages. The N- glycan structures were elucidated to be complex/dibranched, fucosylated/complex/dibranched, monosialylated/complex/dibranched, and monosialylated/fucosylated/dibranched entities.
Notes: Mechref, Y
Chen, P
Novotny, M V
GM24349/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1999 Mar;9(3):227-34. doi: 10.1093/glycob/9.3.227.
Author Address: Department of Chemistry, Indiana University, Bloomington, IN 47405, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 703
Author: Mechref, Y., Ma, W., Hao, G. and Novotny, M. V.
Year: 1999
Title: N-linked oligosaccharides of vomeromodulin, a putative pheromone transporter in rat
Journal: Biochem Biophys Res Commun
Volume: 255
Issue: 2
Pages: 451-5
Epub Date: 1999/03/02
Date: Feb 16
Short Title: N-linked oligosaccharides of vomeromodulin, a putative pheromone transporter in rat
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1006/bbrc.1999.0176
Accession Number: 10049729
Keywords: Animals
Biological Transport
Carbohydrate Conformation
Carrier Proteins/*chemistry/metabolism
Chromatography, Affinity
Electrophoresis, Polyacrylamide Gel
Female
Glycoproteins/*chemistry/metabolism
Glycoside Hydrolases/metabolism
Glycosylation
Intercellular Signaling Peptides and Proteins
Male
*Membrane Transport Proteins
Oligosaccharides/*chemistry/metabolism
Pheromones/*chemistry/metabolism
Polysaccharides/chemistry/metabolism
Rats
Sialic Acids/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Vomeromodulin, a putative pheromone transporter of the rat vomeronasal organ, was isolated by lectin chromatography, purified, and subjected to a mass spectrometric (MS) system of glycan structural determination. Through a combination of exoglycosidase treatments and measurements by matrix-assisted laser desorption/ionization MS, the N-glycans of vomeromodulin were identified as mainly sialylated and fucosylated biantennary structures. The microheterogeneity of N-glycan structures was also due to the presence of galactose residues with different types of linkages.
Notes: Mechref, Y
Ma, W
Hao, G
Novotny, M V
DC 02418/DC/NIDCD NIH HHS/United States
GM24349/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochem Biophys Res Commun. 1999 Feb 16;255(2):451-5. doi: 10.1006/bbrc.1999.0176.
Author Address: Department of Chemistry, Indiana University, Bloomington 47405, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 480
Author: Mega, T., Lujan, E. and Yoshida, A.
Year: 1980
Title: Studies on the oligosaccharide chains of human alpha 1-protease inhibitor. II. Structure of oligosaccharides
Journal: J Biol Chem
Volume: 255
Issue: 9
Pages: 4057-61
Epub Date: 1980/05/10
Date: May 10
Short Title: Studies on the oligosaccharide chains of human alpha 1-protease inhibitor. II. Structure of oligosaccharides
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6966283
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Glycopeptides/analysis
Humans
*Oligosaccharides
*alpha 1-Antitrypsin
Abstract: Human alpha 1-protease inhibitor has three oligosaccharide side chains attached to 3 separate asparaginyl residues of the protein by N-glycosyl linkages. Two of the three asparaginyl residues link mostly to the A-type oligosaccharide chains which consist of Man3, Gal2, (GlcNAc)4 and (NeuAc)2. One of the three asparaginyl residues also attaches to a B-type oligosaccharide chain, which consists of Man3, Gal3, (GlcNAc)5, and (NeuAc)3. The ratio of A-chain to B-chain in this particular position is about 2:1. The structures of A- and B-chains of the glycoprotein were examined by periodate oxidation, sequential glycosidase digestion, and permethylation. The results unequivocally revealed the following structure for A-chains. (Formula: see text). B-type oligosaccharide chains have an additional trisaccharide, i.e. NeuAc 2 alpha leads to 3 Gal 1 beta leads to 4 GlcNAc attached to mannose at the (a) and/or (b) position of A-type chains by a beta 1 leads to 4 linkage. The sialic acid of human alpha 1-protease inhibitor was determined to be N-acetylneuraminic acid.
Notes: Mega, T
Lujan, E
Yoshida, A
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1980 May 10;255(9):4057-61.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1824
Author: Mehta-D'souza, P., Klopocki, A. G., Oganesyan, V., Terzyan, S., Mather, T., Li, Z., Panicker, S. R., Zhu, C. and McEver, R. P.
Year: 2017
Title: Glycan Bound to the Selectin Low Affinity State Engages Glu-88 to Stabilize the High Affinity State under Force
Journal: J Biol Chem
Volume: 292
Issue: 6
Pages: 2510-2518
Epub Date: 2016/12/25
Date: Feb 10
Short Title: Glycan Bound to the Selectin Low Affinity State Engages Glu-88 to Stabilize the High Affinity State under Force
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.767186
PMCID: PMC5313117
Accession Number: 28011641
Keywords: Animals
CHO Cells
Cricetinae
Cricetulus
Crystallography, X-Ray
Glutamic Acid/*metabolism
Humans
L-Selectin/chemistry/*metabolism
Membrane Glycoproteins/metabolism
Polysaccharides/*metabolism
Protein Conformation
*carbohydrate-binding protein
*cell adhesion
*endothelial cell
*glycobiology
*inflammation
*leukocyte
Abstract: Selectin interactions with fucosylated glycan ligands mediate leukocyte rolling in the vasculature under shear forces. Crystal structures of P- and E-selectin suggest a two-state model in which ligand binding to the lectin domain closes loop 83-89 around the Ca(2+) coordination site, enabling Glu-88 to engage Ca(2+) and fucose. This triggers further allostery that opens the lectin/EGF domain hinge. The model posits that force accelerates transition from the bent (low affinity) to the extended (high affinity) state. However, transition intermediates have not been described, and the role of Glu-88 in force-assisted allostery has not been examined. Here we report the structure of the lectin and EGF domains of L-selectin bound to a fucose mimetic; that is, a terminal mannose on an N-glycan attached to a symmetry-related molecule. The structure is a transition intermediate where loop 83-89 closes to engage Ca(2+) and mannose without triggering allostery that opens the lectin/EGF domain hinge. We used three complementary assays to compare ligand binding to WT selectins and to E88D selectins that replaced Glu-88 with Asp. Soluble P-selectinE88D bound with an ∼9-fold lower affinity to PSGL-1, a physiological ligand, due to faster dissociation. Adhesion frequency experiments with a biomembrane force probe could not detect interactions of P-selectinE88D with PSGL-1. Cells expressing transmembrane P-selectinE88D or L-selectinE88D detached from immobilized ligands immediately after initiating flow. Cells expressing E-selectinE88D rolled but detached faster. Our data support a two-state model for selectins in which Glu-88 must engage ligand to trigger allostery that stabilizes the high affinity state under force.
Notes: 1083-351x
Mehta-D'souza, Padmaja
Klopocki, Arkadiusz G
Oganesyan, Vaheh
Terzyan, Simon
Mather, Timothy
Li, Zhenhai
Panicker, Sumith R
Zhu, Cheng
McEver, Rodger P
Orcid: 0000-0001-5343-2622
R01 HL132019/HL/NHLBI NIH HHS/United States
R01 AI044902/AI/NIAID NIH HHS/United States
R37 HL034363/HL/NHLBI NIH HHS/United States
R01 AI077343/AI/NIAID NIH HHS/United States
P01 HL085607/HL/NHLBI NIH HHS/United States
R01 HL034363/HL/NHLBI NIH HHS/United States
R56 AI077343/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2017 Feb 10;292(6):2510-2518. doi: 10.1074/jbc.M116.767186. Epub 2016 Dec 23.
Author Address: From the Cardiovascular Biology Research Program and.
Crystallography Research Program, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma 73104.
the Coulter Department of Biomedical Engineering.
Woodruff School of Mechanical Engineering, and.
the Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, Georgia 30332.
From the Cardiovascular Biology Research Program and rodger-mcever@omrf.org.
the Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 269
Author: Meikrantz, W., Smith, D. M., Sladicka, M. M. and Schlegel, R. A.
Year: 1991
Title: Nuclear localization of an O-glycosylated protein phosphotyrosine phosphatase from human cells
Journal: J Cell Sci
Volume: 98 ( Pt 3)
Pages: 303-7
Epub Date: 1991/03/01
Date: Mar
Short Title: Nuclear localization of an O-glycosylated protein phosphotyrosine phosphatase from human cells
Alternate Journal: Journal of cell science
ISSN: 0021-9533 (Print)
0021-9533
Accession Number: 1647397
Keywords: Acetylglucosamine/analysis
Cell Line
Cell Nucleus/*enzymology
Chromosomes/enzymology
Cytosol/enzymology
Electrophoresis, Polyacrylamide Gel
Enzyme-Linked Immunosorbent Assay
Glycoproteins/isolation & purification
Humans
Mitosis
Phosphoprotein Phosphatases/chemistry/*isolation & purification/metabolism
Phosphoproteins/antagonists & inhibitors/chemistry/*isolation & purification
Precipitin Tests
Protein Tyrosine Phosphatases
Subcellular Fractions/enzymology
Substrate Specificity
Abstract: Histochemical staining of immunoprecipitates of p65, a component of human M phase-promoting factor, identified the molecule as having phosphatase activity. The enzyme, purified 3400-fold from mitotic cell extracts by (NH4)2SO4 precipitation, DEAE chromatography, and immunoaffinity chromatography on immobilized anti-p65 IgG, was inhibited by Zn2- and Na3VO4 but not NaF or beta-glycerophosphate; 32P-labeled poly(Glu, Tyr) was more efficiently dephosphorylated than phosphorylated histone or phosphorylase a. Indirect immunofluorescence showed most of the phosphatase to be localized in the nucleus of interphase cells, with a fine, granular distribution unaltered by detergent extraction; in mitotic cells, p65 was localized on chromosomes. ELISA of subcellular fractions confirmed this localization. Immunoreactive p65 was recovered from immobilized wheat germ agglutinin (WGA) upon elution with N-acetylglucosamine; similarly, WGA recognized immunoaffinity-purified p65 on blots. Alkaline hydrolysis of blotted protein prevented WGA binding, indicating that phosphatase p65, like a small group of other nuclear proteins, contains O-linked carbohydrate terminating in N-acetylglucosamine.
Notes: Meikrantz, W
Smith, D M
Sladicka, M M
Schlegel, R A
Journal Article
England
J Cell Sci. 1991 Mar;98 ( Pt 3):303-7.
Author Address: Department of Molecular and Cell Biology, Pennsylvania State University, University Park 16802.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 471
Author: Mellis, S. J. and Baenziger, J. U.
Year: 1983
Title: Structures of the O-glycosidically linked oligosaccharides of human IgD
Journal: J Biol Chem
Volume: 258
Issue: 19
Pages: 11557-63
Epub Date: 1983/10/10
Date: Oct 10
Short Title: Structures of the O-glycosidically linked oligosaccharides of human IgD
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6619128
Keywords: Amino Acid Sequence
Amino Acids/analysis
Carbohydrate Conformation
Carbohydrate Sequence
Glycosides/*analysis
Humans
*Immunoglobulin D
Multiple Myeloma/immunology
N-Acetylneuraminic Acid
Oligosaccharides/*isolation & purification
Sialic Acids/analysis
Abstract: In the previous communication (Mellis, S. J., and Baenziger, J. U. (1983) J. Biol. Chem. 258, 11546-11556), the structures of the oligosaccharides present at the 3 asparagine glycosylation sites of a human IgD myeloma protein were defined. In this communication, we present the structures of the O-glycosidically linked oligosaccharides located in the hinge region of IgD:WAH. Three or four threonine residues and one serine residue in the region bear O-glycosidically linked oligosaccharides. Approximately 50% of these molecules have the structure Gal beta 1 leads to 3 GalNAc which is identical with the structure of the predominant oligosaccharide in the hinge region of human IgA1 myeloma proteins (Baenziger, J. U., and Kornfeld, S. (1974) J. Biol. Chem. 249, 7270-7281). The remainder of the oligosaccharides contain 1 or 2 residues of N-acetylneuraminic acid and have the structures NeuAc alpha 2 leads to 3Gal beta 1 leads to 3GalNAc (30%), Gal beta 1 leads to (NeuAc alpha 2 leads to 6)GalNAc (12%), and NeuAc alpha 2 leads to 3Gal beta 1 leads to 3(NeuAc alpha 2 leads to 6)GalNAc (8%). The sialylated molecules have not been encountered previously on other human immunoglobulin heavy chains. These structures, however, have been described on a number of secreted and membrane glycoproteins. Examination of oligosaccharides isolated from different subregions of the IgD hinge indicated that a specific distribution of the sialylated structures among the glycosylated amino acids of the hinge region is not likely.
Notes: Mellis, S J
Baenziger, J U
GM07200/GM/NIGMS NIH HHS/United States
R01 CA21923/CA/NCI NIH HHS/United States
RR00954/RR/NCRR NIH HHS/United States
etc.
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1983 Oct 10;258(19):11557-63.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1601
Author: Meng, L., Forouhar, F., Thieker, D., Gao, Z., Ramiah, A., Moniz, H., Xiang, Y., Seetharaman, J., Milaninia, S., Su, M., Bridger, R., Veillon, L., Azadi, P., Kornhaber, G., Wells, L., Montelione, G. T., Woods, R. J., Tong, L. and Moremen, K. W.
Year: 2013
Title: Enzymatic basis for N-glycan sialylation: structure of rat α2,6-sialyltransferase (ST6GAL1) reveals conserved and unique features for glycan sialylation
Journal: J Biol Chem
Volume: 288
Issue: 48
Pages: 34680-98
Epub Date: 2013/10/25
Date: Nov 29
Short Title: Enzymatic basis for N-glycan sialylation: structure of rat α2,6-sialyltransferase (ST6GAL1) reveals conserved and unique features for glycan sialylation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.519041
PMCID: PMC3843080
Accession Number: 24155237
Keywords: Animals
Bacteria/enzymology/genetics
Binding Sites
*Crystallography, X-Ray
Molecular Docking Simulation
Molecular Dynamics Simulation
Polysaccharides/biosynthesis/*chemistry
Protein Conformation
Rats
Sialic Acids/chemistry/*metabolism
Sialyltransferases/*chemistry/metabolism
Structure-Activity Relationship
Swine/genetics
Amber
Carbohydrate Biosynthesis
Carbohydrate Glycoprotein
Enzyme Structure
Glycam
Glycobiology
Glycoprotein Biosynthesis
Molecular Dynamics
Abstract: Glycan structures on glycoproteins and glycolipids play critical roles in biological recognition, targeting, and modulation of functions in animal systems. Many classes of glycan structures are capped with terminal sialic acid residues, which contribute to biological functions by either forming or masking glycan recognition sites on the cell surface or secreted glycoconjugates. Sialylated glycans are synthesized in mammals by a single conserved family of sialyltransferases that have diverse linkage and acceptor specificities. We examined the enzymatic basis for glycan sialylation in animal systems by determining the crystal structures of rat ST6GAL1, an enzyme that creates terminal α2,6-sialic acid linkages on complex-type N-glycans, at 2.4 Å resolution. Crystals were obtained from enzyme preparations generated in mammalian cells. The resulting structure revealed an overall protein fold broadly resembling the previously determined structure of pig ST3GAL1, including a CMP-sialic acid-binding site assembled from conserved sialylmotif sequence elements. Significant differences in structure and disulfide bonding patterns were found outside the sialylmotif sequences, including differences in residues predicted to interact with the glycan acceptor. Computational substrate docking and molecular dynamics simulations were performed to predict and evaluate the CMP-sialic acid donor and glycan acceptor interactions, and the results were compared with kinetic analysis of active site mutants. Comparisons of the structure with pig ST3GAL1 and a bacterial sialyltransferase revealed a similar positioning of donor, acceptor, and catalytic residues that provide a common structural framework for catalysis by the mammalian and bacterial sialyltransferases.
Notes: 1083-351x
Meng, Lu
Forouhar, Farhad
Thieker, David
Gao, Zhongwei
Ramiah, Annapoorani
Moniz, Heather
Xiang, Yong
Seetharaman, Jayaraman
Milaninia, Sahand
Su, Min
Bridger, Robert
Veillon, Lucas
Azadi, Parastoo
Kornhaber, Gregory
Wells, Lance
Montelione, Gaetano T
Woods, Robert J
Tong, Liang
Moremen, Kelley W
P41GM103390/GM/NIGMS NIH HHS/United States
R01 GM094919/GM/NIGMS NIH HHS/United States
P41 GM103390/GM/NIGMS NIH HHS/United States
P41RR005351/RR/NCRR NIH HHS/United States
P41 RR005351/RR/NCRR NIH HHS/United States
U54 GM094597/GM/NIGMS NIH HHS/United States
R01GM094919/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2013 Nov 29;288(48):34680-98. doi: 10.1074/jbc.M113.519041. Epub 2013 Oct 23.
Author Address: From the Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1328
Author: Meng, W., Brigance, R. P., Chao, H. J., Fura, A., Harrity, T., Marcinkeviciene, J., O'Connor, S. P., Tamura, J. K., Xie, D., Zhang, Y., Klei, H. E., Kish, K., Weigelt, C. A., Turdi, H., Wang, A., Zahler, R., Kirby, M. S. and Hamann, L. G.
Year: 2010
Title: Discovery of 6-(aminomethyl)-5-(2,4-dichlorophenyl)-7-methylimidazo[1,2-a]pyrimidine-2-carboxamides as potent, selective dipeptidyl peptidase-4 (DPP4) inhibitors
Journal: J Med Chem
Volume: 53
Issue: 15
Pages: 5620-8
Epub Date: 2010/08/06
Date: Aug 12
Short Title: Discovery of 6-(aminomethyl)-5-(2,4-dichlorophenyl)-7-methylimidazo[1,2-a]pyrimidine-2-carboxamides as potent, selective dipeptidyl peptidase-4 (DPP4) inhibitors
Alternate Journal: Journal of medicinal chemistry
ISSN: 0022-2623
DOI: 10.1021/jm100634a
Accession Number: 20684603
Keywords: Animals
Catalytic Domain
Crystallography, X-Ray
Diabetes Mellitus, Type 2/drug therapy
Dipeptidyl Peptidase 4/chemistry
*Dipeptidyl-Peptidase IV Inhibitors
Dogs
ERG1 Potassium Channel
Ether-A-Go-Go Potassium Channels/antagonists & inhibitors
Humans
Hypoglycemic Agents/*chemical synthesis/pharmacokinetics/pharmacology
Imidazoles/*chemical synthesis/pharmacokinetics/pharmacology
Male
Mice
Mice, Obese
Models, Molecular
Pyrimidines/*chemical synthesis/pharmacokinetics/pharmacology
Rats
Sodium Channel Blockers/pharmacology
Stereoisomerism
Structure-Activity Relationship
Abstract: Continued structure-activity relationship (SAR) exploration within our previously disclosed azolopyrimidine containing dipeptidyl peptidase-4 (DPP4) inhibitors led us to focus on an imidazolopyrimidine series in particular. Further study revealed that by replacing the aryl substitution on the imidazole ring with a more polar carboxylic ester or amide, these compounds displayed not only increased DPP4 binding activity but also significantly reduced human ether-a-go-go related gene (hERG) and sodium channel inhibitory activities. Additional incremental adjustment of polarity led to permeable molecules which exhibited favorable pharmacokinetic (PK) profiles in preclinical animal species. The active site binding mode of these compounds was determined by X-ray crystallography as exemplified by amide 24c. A subsequent lead molecule from this series, (+)-6-(aminomethyl)-5-(2,4-dichlorophenyl)-N-(1-ethyl-1H-pyrazol-5-yl)-7-methylimidazo[1,2-a]pyrimidine-2-carboxamide (24s), emerged as a potent, selective DPP4 inhibitor that displayed excellent PK profiles and in vivo efficacy in ob/ob mice.
Notes: 1520-4804
Meng, Wei
Brigance, Robert P
Chao, Hannguang J
Fura, Aberra
Harrity, Thomas
Marcinkeviciene, Jovita
O'Connor, Stephen P
Tamura, James K
Xie, Dianlin
Zhang, Yaqun
Klei, Herbert E
Kish, Kevin
Weigelt, Carolyn A
Turdi, Huji
Wang, Aiying
Zahler, Robert
Kirby, Mark S
Hamann, Lawrence G
Journal Article
United States
J Med Chem. 2010 Aug 12;53(15):5620-8. doi: 10.1021/jm100634a.
Author Address: Departments of Discovery Chemistry, Bristol-Myers Squibb, Research and Development, Princeton, New Jersey 08543-5400, USA. wei.meng@bms.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1539
Author: Menting, J. G., Whittaker, J., Margetts, M. B., Whittaker, L. J., Kong, G. K., Smith, B. J., Watson, C. J., Záková, L., Kletvíková, E., Jiráček, J., Chan, S. J., Steiner, D. F., Dodson, G. G., Brzozowski, A. M., Weiss, M. A., Ward, C. W. and Lawrence, M. C.
Year: 2013
Title: How insulin engages its primary binding site on the insulin receptor
Journal: Nature
Volume: 493
Issue: 7431
Pages: 241-5
Epub Date: 2013/01/11
Date: Jan 10
Short Title: How insulin engages its primary binding site on the insulin receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature11781
PMCID: PMC3793637
NIHMSID: NIHMS513687
Accession Number: 23302862
Keywords: Animals
Binding Sites
Calorimetry
Cattle
Cell Line
Crystallography, X-Ray
Humans
Insulin/*chemistry/*metabolism
Leucine/metabolism
Ligands
Models, Molecular
Protein Binding
Protein Structure, Secondary
Receptor, Insulin/*chemistry/*metabolism
Reproducibility of Results
Abstract: Insulin receptor signalling has a central role in mammalian biology, regulating cellular metabolism, growth, division, differentiation and survival. Insulin resistance contributes to the pathogenesis of type 2 diabetes mellitus and the onset of Alzheimer's disease; aberrant signalling occurs in diverse cancers, exacerbated by cross-talk with the homologous type 1 insulin-like growth factor receptor (IGF1R). Despite more than three decades of investigation, the three-dimensional structure of the insulin-insulin receptor complex has proved elusive, confounded by the complexity of producing the receptor protein. Here we present the first view, to our knowledge, of the interaction of insulin with its primary binding site on the insulin receptor, on the basis of four crystal structures of insulin bound to truncated insulin receptor constructs. The direct interaction of insulin with the first leucine-rich-repeat domain (L1) of insulin receptor is seen to be sparse, the hormone instead engaging the insulin receptor carboxy-terminal α-chain (αCT) segment, which is itself remodelled on the face of L1 upon insulin binding. Contact between insulin and L1 is restricted to insulin B-chain residues. The αCT segment displaces the B-chain C-terminal β-strand away from the hormone core, revealing the mechanism of a long-proposed conformational switch in insulin upon receptor engagement. This mode of hormone-receptor recognition is novel within the broader family of receptor tyrosine kinases. We support these findings by photo-crosslinking data that place the suggested interactions into the context of the holoreceptor and by isothermal titration calorimetry data that dissect the hormone-insulin receptor interface. Together, our findings provide an explanation for a wealth of biochemical data from the insulin receptor and IGF1R systems relevant to the design of therapeutic insulin analogues.
Notes: 1476-4687
Menting, John G
Whittaker, Jonathan
Margetts, Mai B
Whittaker, Linda J
Kong, Geoffrey K-W
Smith, Brian J
Watson, Christopher J
Záková, Lenka
Kletvíková, Emília
Jiráček, Jiří
Chan, Shu Jin
Steiner, Donald F
Dodson, Guy G
Brzozowski, Andrzej M
Weiss, Michael A
Ward, Colin W
Lawrence, Michael C
R01 DK040949/DK/NIDDK NIH HHS/United States
DK40949/DK/NIDDK NIH HHS/United States
Biotechnology and Biological Sciences Research Council/United Kingdom
R01 DK013914/DK/NIDDK NIH HHS/United States
UL1 TR000439/TR/NCATS NIH HHS/United States
DK13914/DK/NIDDK NIH HHS/United States
DK20595/DK/NIDDK NIH HHS/United States
P30 DK020595/DK/NIDDK NIH HHS/United States
P60 DK020595/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2013 Jan 10;493(7431):241-5. doi: 10.1038/nature11781.
Author Address: Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Parkville, Victoria 3052, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 612
Author: Meri, S., Lehto, T., Sutton, C. W., Tyynelä, J. and Baumann, M.
Year: 1996
Title: Structural composition and functional characterization of soluble CD59: heterogeneity of the oligosaccharide and glycophosphoinositol (GPI) anchor revealed by laser-desorption mass spectrometric analysis
Journal: Biochem J
Volume: 316 ( Pt 3)
Issue: Pt 3
Pages: 923-35
Epub Date: 1996/06/15
Date: Jun 15
Short Title: Structural composition and functional characterization of soluble CD59: heterogeneity of the oligosaccharide and glycophosphoinositol (GPI) anchor revealed by laser-desorption mass spectrometric analysis
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj3160923
PMCID: PMC1217438
Accession Number: 8670172
Keywords: Amino Acid Sequence
Animals
Antigens, CD/blood/chemistry/urine
CD59 Antigens/blood/*chemistry/urine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Ion Exchange
Complement System Proteins/physiology
Erythrocyte Membrane/immunology
Glycopeptides/chemistry
Glycoside Hydrolases
Glycosylphosphatidylinositols/*analysis/chemistry
Humans
Mammals
Molecular Sequence Data
Molecular Weight
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/chemistry
Peptide Mapping
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
Abstract: CD59 (protectin) is a glycophosphoinositol (GPI)-anchored inhibitor of the membrane attack complex of complement found on blood cells, endothelia and epithelial cells. In addition to the lipid-tailed CD59, soluble lipid-free forms of CD59 are present in human body fluids. We have investigated the detailed structural composition of the naturally occurring soluble urinary CD59 (CD59u) using peptide mapping, anion-exchange chromatography, sequential exoglycosidase digestion and matrix-assisted laser-desorption mass spectrometry (MALDI-MS). CD59u exhibited an average M(r) of 12444 in MALDI-MS. Mass analysis of the isolated C-terminal peptide (T9) indicated that a GPI-anchor (at Asn-77) without an inositol-associated phospholipid was present in soluble CD59u. By using residue-specific exoglycosidases, chemical modification and MALDI-MS structures of seven different GPI-anchor variants were determined. Variant forms of the anchor had deletions and/or extensions of one or more monosaccharide units. Sialic acid linked to an N-acetylhexosamine-galactose arm was found in two GPI-anchor variants. The N-linked carbohydrate side chain of CD59u (at Asn-18) also displayed considerable heterogeneity. The predominant oligosaccharide chains were fucosylated biantennary and triantennary complexes with variable sialylation. Mono Q anion-exchange chromatography resolved urinary CD59 into nine different fractions that bound equally well to the terminal complement SC5b-8 complexes. Despite binding to C5b-8, soluble CD59u inhibited complement lysis at an approx. 200-fold lower efficiency than erythrocyte CD59. These results document the structural heterogeneity of both the GPI anchor and N-linked oligosaccharide of CD59 and demonstrate that the phospholipid tail is needed for the full functional activity of CD59. The site of cleavage between the diradylglycerol phosphate and inositol suggests that a mammalian phospholipase D could be involved in the solubilization of GPI-anchored proteins.
Notes: 1470-8728
Meri, S
Lehto, T
Sutton, C W
Tyynelä, J
Baumann, M
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1996 Jun 15;316 ( Pt 3)(Pt 3):923-35. doi: 10.1042/bj3160923.
Author Address: Department of Bacteriology and Immunology, Haartman Institute, University of Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1031
Author: Mesters, J. R., Barinka, C., Li, W., Tsukamoto, T., Majer, P., Slusher, B. S., Konvalinka, J. and Hilgenfeld, R.
Year: 2006
Title: Structure of glutamate carboxypeptidase II, a drug target in neuronal damage and prostate cancer
Journal: Embo j
Volume: 25
Issue: 6
Pages: 1375-84
Epub Date: 2006/02/10
Date: Mar 22
Short Title: Structure of glutamate carboxypeptidase II, a drug target in neuronal damage and prostate cancer
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7600969
PMCID: PMC1422165
Accession Number: 16467855
Keywords: Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Enzyme Inhibitors/chemistry/metabolism
Glutamate Carboxypeptidase II/antagonists & inhibitors/*chemistry/metabolism
Glutamic Acid/chemistry/metabolism
Glycosylation
Humans
Hydrolysis
Male
Models, Molecular
Molecular Sequence Data
*Neurons/enzymology/pathology
Prostatic Neoplasms/*enzymology/pathology
Protein Conformation
Protein Folding
Recombinant Proteins/chemistry/isolation & purification/metabolism
Abstract: Membrane-bound glutamate carboxypeptidase II (GCPII) is a zinc metalloenzyme that catalyzes the hydrolysis of the neurotransmitter N-acetyl-L-aspartyl-L-glutamate (NAAG) to N-acetyl-L-aspartate and L-glutamate (which is itself a neurotransmitter). Potent and selective GCPII inhibitors have been shown to decrease brain glutamate and provide neuroprotection in preclinical models of stroke, amyotrophic lateral sclerosis, and neuropathic pain. Here, we report crystal structures of the extracellular part of GCPII in complex with both potent and weak inhibitors and with glutamate, the product of the enzyme's hydrolysis reaction, at 2.0, 2.4, and 2.2 A resolution, respectively. GCPII folds into three domains: protease-like, apical, and C-terminal. All three participate in substrate binding, with two of them directly involved in C-terminal glutamate recognition. One of the carbohydrate moieties of the enzyme is essential for homodimer formation of GCPII. The three-dimensional structures presented here reveal an induced-fit substrate-binding mode of this key enzyme and provide essential information for the design of GCPII inhibitors useful in the treatment of neuronal diseases and prostate cancer.
Notes: 1460-2075
Mesters, Jeroen R
Barinka, Cyril
Li, Weixing
Tsukamoto, Takashi
Majer, Pavel
Slusher, Barbara S
Konvalinka, Jan
Hilgenfeld, Rolf
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2006 Mar 22;25(6):1375-84. doi: 10.1038/sj.emboj.7600969. Epub 2006 Feb 9.
Author Address: Institute of Biochemistry, Center for Structural and Cell Biology in Medicine, University of Lübeck, Lübeck, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1099
Author: Mesters, J. R., Henning, K. and Hilgenfeld, R.
Year: 2007
Title: Human glutamate carboxypeptidase II inhibition: structures of GCPII in complex with two potent inhibitors, quisqualate and 2-PMPA
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 63
Issue: Pt 4
Pages: 508-13
Epub Date: 2007/03/21
Date: Apr
Short Title: Human glutamate carboxypeptidase II inhibition: structures of GCPII in complex with two potent inhibitors, quisqualate and 2-PMPA
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s090744490700902x
Accession Number: 17372356
Keywords: Antigens, Surface/chemistry
Binding Sites
Enzyme Inhibitors/*chemistry/pharmacology
Excitatory Amino Acid Agonists/*chemistry/pharmacology
Glutamate Carboxypeptidase II/*antagonists & inhibitors/chemistry
Humans
Hydrogen Bonding
Models, Molecular
Organophosphorus Compounds/*chemistry/pharmacology
Protein Conformation
Quisqualic Acid/*chemistry/pharmacology
Abstract: Human glutamate carboxypeptidase II (GCPII) occurs in the central nervous system as well as in human prostate (where it is called prostate-specific membrane antigen; PSMA). Inhibitors of the enzyme have been shown to provide neuroprotection, but may also be useful for the detection, imaging and treatment of prostate cancer. Crystal structures were determined of the extracellular part of GCPII (amino-acid residues 44-750) in complex with two potent inhibitors, quisqualate and 2-PMPA (the strongest GCPII inhibitor to date), at resolutions of 3.0 and 2.2 A, respectively. In addition, models were constructed for binding of the inhibitors willardiine, homoibotenate, L-2-amino-4-phosphonobutanoic acid and L-serine-O-sulfate to the S1' site of the enzyme. The common denominator for high-affinity binding to the S1' site is the formation of two strong salt bridges.
Notes: Mesters, Jeroen R
Henning, Karen
Hilgenfeld, Rolf
Journal Article
Research Support, Non-U.S. Gov't
United States
Acta Crystallogr D Biol Crystallogr. 2007 Apr;63(Pt 4):508-13. doi: 10.1107/S090744490700902X. Epub 2007 Mar 16.
Author Address: Institute of Biochemistry, Center for Structural and Cell Biology In Medicine, University of Lübeck, Ratzeburger Allee 160, Lübeck 23538, Germany. mesters@biochem.uni-luebeck.de
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 717
Author: Meunier, J. C., Fournillier, A., Choukhi, A., Cahour, A., Cocquerel, L., Dubuisson, J. and Wychowski, C.
Year: 1999
Title: Analysis of the glycosylation sites of hepatitis C virus (HCV) glycoprotein E1 and the influence of E1 glycans on the formation of the HCV glycoprotein complex
Journal: J Gen Virol
Volume: 80 ( Pt 4)
Pages: 887-896
Epub Date: 1999/04/22
Date: Apr
Short Title: Analysis of the glycosylation sites of hepatitis C virus (HCV) glycoprotein E1 and the influence of E1 glycans on the formation of the HCV glycoprotein complex
Alternate Journal: The Journal of general virology
ISSN: 0022-1317 (Print)
0022-1317
DOI: 10.1099/0022-1317-80-4-887
Accession Number: 10211957
Keywords: Animals
Calcium-Binding Proteins/metabolism
Calnexin
Cell Line
Glycoproteins/*metabolism
Glycosylation
Hepacivirus/*chemistry
Mice
Mutagenesis, Site-Directed
Protein Conformation
Rabbits
Structure-Activity Relationship
Viral Envelope Proteins/*metabolism
Abstract: The hepatitis C virus (HCV) genome encodes two membrane-associated envelope glycoproteins (E1 and E2), which are released from the viral polyprotein precursor by host signal peptidase cleavages. These glycoproteins interact to form a noncovalent heterodimeric complex, which is retained in the endoplasmic reticulum. HCV glycoproteins, E1 and E2, are heavily modified by N-linked glycosylation. A recent study has revealed that upon partial deglycosylation with endoglycosidase H only four of the five potential glycosylation sites of HCV glycoprotein E1 are utilized. In this work, the unused glycosylation site on the E1 glycoprotein was identified and the influence of N-linked glycosylation on the formation of the HCV glycoprotein complex was studied by expressing a panel of E1 glycosylation mutants in HepG2 cells. Each of the five potential N-linked glycosylation sites, located at amino acid positions 196, 209, 234, 305 and 325, respectively, on the HCV polyprotein, was mutated separately as well as in combination with the other sites. Expression of the mutated E1 proteins in HepG2 cells indicated that the fifth glycosylation site is not used for the addition of N-linked oligosaccharides and the Pro immediately following the sequon (Asn-Trp-Ser) precludes core glycosylation. The effect of each mutation on the formation of noncovalent E1E2 complexes was also analysed. As determined with the use of a conformation-sensitive monoclonal antibody, mutations at positions N2 and N3 had no, or only minor, effects on the assembly of the E1E2 complex, whereas a mutation at position N1 and predominantly at position N4 dramatically reduced the efficiency of the formation of noncovalent E1E2 complexes.
Notes: Meunier, J C
Fournillier, A
Choukhi, A
Cahour, A
Cocquerel, L
Dubuisson, J
Wychowski, C
Journal Article
Research Support, Non-U.S. Gov't
England
J Gen Virol. 1999 Apr;80 ( Pt 4):887-896. doi: 10.1099/0022-1317-80-4-887.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1687
Author: Mi, L. Z., Brown, C. T., Gao, Y., Tian, Y., Le, V. Q., Walz, T. and Springer, T. A.
Year: 2015
Title: Structure of bone morphogenetic protein 9 procomplex
Journal: Proc Natl Acad Sci U S A
Volume: 112
Issue: 12
Pages: 3710-5
Epub Date: 2015/03/10
Date: Mar 24
Short Title: Structure of bone morphogenetic protein 9 procomplex
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1501303112
PMCID: PMC4378411
Accession Number: 25751889
Keywords: Amino Acid Sequence
Animals
Bone Morphogenetic Protein 7/*chemistry
CHO Cells
Cricetinae
Cricetulus
Growth Differentiation Factor 2
Growth Differentiation Factors/*chemistry
HEK293 Cells
Humans
Intercellular Signaling Peptides and Proteins/metabolism
Molecular Sequence Data
Protein Structure, Secondary
Protein Structure, Tertiary
Sequence Homology, Amino Acid
Signal Transduction
Transforming Growth Factor beta/*chemistry
Abstract: Bone morphogenetic proteins (BMPs) belong to the TGF-β family, whose 33 members regulate multiple aspects of morphogenesis. TGF-β family members are secreted as procomplexes containing a small growth factor dimer associated with two larger prodomains. As isolated procomplexes, some members are latent, whereas most are active; what determines these differences is unknown. Here, studies on pro-BMP structures and binding to receptors lead to insights into mechanisms that regulate latency in the TGF-β family and into the functions of their highly divergent prodomains. The observed open-armed, nonlatent conformation of pro-BMP9 and pro-BMP7 contrasts with the cross-armed, latent conformation of pro-TGF-β1. Despite markedly different arm orientations in pro-BMP and pro-TGF-β, the arm domain of the prodomain can similarly associate with the growth factor, whereas prodomain elements N- and C-terminal to the arm associate differently with the growth factor and may compete with one another to regulate latency and stepwise displacement by type I and II receptors. Sequence conservation suggests that pro-BMP9 can adopt both cross-armed and open-armed conformations. We propose that interactors in the matrix stabilize a cross-armed pro-BMP conformation and regulate transition between cross-armed, latent and open-armed, nonlatent pro-BMP conformations.
Notes: 1091-6490
Mi, Li-Zhi
Brown, Christopher T
Gao, Yijie
Tian, Yuan
Le, Viet Q
Walz, Thomas
Springer, Timothy A
Journal Article
Proc Natl Acad Sci U S A. 2015 Mar 24;112(12):3710-5. doi: 10.1073/pnas.1501303112. Epub 2015 Mar 6.
Author Address: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115; Program in Cellular and Molecular Medicine, Children's Hospital Boston, Boston, MA 02115; Department of Molecular and Structural Biology, School of Life Sciences, Tianjin University, Tianjin 300072, China;
Global BioTherapeutics Technologies, Pfizer Worldwide Research and Development, Cambridge, MA 02139; and.
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115; Program in Cellular and Molecular Medicine, Children's Hospital Boston, Boston, MA 02115;
Howard Hughes Medical Institute and Department of Cell Biology, Harvard Medical School, Boston, MA 02115.
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115; Program in Cellular and Molecular Medicine, Children's Hospital Boston, Boston, MA 02115; timothy.springer@childrens.harvard.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 375
Author: Miedel, M. C., Hulmes, J. D., Weber, D. V., Bailon, P. and Pan, Y. C.
Year: 1988
Title: Structural analysis of recombinant soluble human interleukin-2 receptor. Primary structure, assignment of disulfide bonds and core IL-2 binding structure
Journal: Biochem Biophys Res Commun
Volume: 154
Issue: 1
Pages: 372-9
Epub Date: 1988/07/15
Date: Jul 15
Short Title: Structural analysis of recombinant soluble human interleukin-2 receptor. Primary structure, assignment of disulfide bonds and core IL-2 binding structure
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(88)90695-x
Accession Number: 3134887
Keywords: Amino Acid Sequence
Animals
Cell Line
Cloning, Molecular
Disulfides/analysis
Humans
Interleukin-2/*metabolism
Models, Molecular
Molecular Sequence Data
Peptide Fragments/analysis
Protein Conformation
*Receptors, Immunologic/genetics
Receptors, Interleukin-2
*Recombinant Proteins
Abstract: A purified soluble and functional form of recombinant human interleukin-2 receptor, engineered and expressed in Chinese hamster ovary cells, was structurally characterized. The primary sequence of this 224 amino acid recombinant protein which lacks most of the carboxy-terminal transmembrane and cytoplasmic portions of the intact protein was established by sequence analyses. The disulfide bonds were assigned by comparative peptide mapping of the reduced and non-reduced peptide digests. As in the case of natural interleukin-2 receptor they occur between cysteines 3-147, 46-104, 131-163, and 28/30-59/61. Based on assignment of the disulfide bonds, a structural model of the interleukin-2 receptor for interleukin-2 binding is proposed.
Notes: Miedel, M C
Hulmes, J D
Weber, D V
Bailon, P
Pan, Y C
Journal Article
United States
Biochem Biophys Res Commun. 1988 Jul 15;154(1):372-9. doi: 10.1016/0006-291x(88)90695-x.
Author Address: Department of Protein Biochemistry, Roche Research Center, Hoffmann-La Roche Inc., Nutley, NJ.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1764
Author: Mihara, E., Hirai, H., Yamamoto, H., Tamura-Kawakami, K., Matano, M., Kikuchi, A., Sato, T. and Takagi, J.
Year: 2016
Title: Active and water-soluble form of lipidated Wnt protein is maintained by a serum glycoprotein afamin/α-albumin
Journal: Elife
Volume: 5
Epub Date: 2016/02/24
Date: Feb 23
Short Title: Active and water-soluble form of lipidated Wnt protein is maintained by a serum glycoprotein afamin/α-albumin
Alternate Journal: eLife
ISSN: 2050-084x
DOI: 10.7554/eLife.11621
PMCID: PMC4775226
Accession Number: 26902720
Keywords: Animals
Carrier Proteins/chemistry/*metabolism
Cattle
Glycoproteins/chemistry/*metabolism
Hydrophobic and Hydrophilic Interactions
Protein Binding
Serum Albumin/chemistry/*metabolism
Solubility
Water
Wnt Proteins/chemistry/*metabolism
afamin
biochemistry
developmental biology
epitope tag
human
lipid modification
mouse
purification
serum
stem cells
wnt
Abstract: Wnt plays important role during development and in various diseases. Because Wnts are lipidated and highly hydrophobic, they can only be purified in the presence of detergents, limiting their use in various in vitro and in vivo assays. We purified N-terminally tagged recombinant Wnt3a secreted from cells and accidentally discovered that Wnt3a co-purified with a glycoprotein afamin derived from the bovine serum included in the media. Wnt3a forms a 1:1 complex with afamin, which remains soluble in aqueous buffer after isolation, and can induce signaling in various cellular systems including the intestical stem cell growth assay. By co-expressing with afamin, biologically active afamin-Wnt complex can be easily obtained in large quantity. As afamin can also solubilize Wnt5a, Wnt3, and many more Wnt subtypes, afamin complexation will open a way to put various Wnt ligands and their signaling mechanisms under a thorough biochemical scrutiny that had been difficult for years.
Notes: 2050-084x
Mihara, Emiko
Hirai, Hidenori
Yamamoto, Hideki
Tamura-Kawakami, Keiko
Matano, Mami
Kikuchi, Akira
Sato, Toshiro
Takagi, Junichi
Orcid: 0000-0002-1219-475x
Journal Article
Research Support, Non-U.S. Gov't
Elife. 2016 Feb 23;5:e11621. doi: 10.7554/eLife.11621.
Author Address: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Japan.
Department of Molecular Biology and Biochemistry, Graduate School of Medicine, Osaka University, Suita, Japan.
Department of Gastroenterology, Keio University School of Medicine, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1349
Author: Miknis, Z. J., Magracheva, E., Li, W., Zdanov, A., Kotenko, S. V. and Wlodawer, A.
Year: 2010
Title: Crystal structure of human interferon-λ1 in complex with its high-affinity receptor interferon-λR1
Journal: J Mol Biol
Volume: 404
Issue: 4
Pages: 650-64
Epub Date: 2010/10/12
Date: Dec 10
Short Title: Crystal structure of human interferon-λ1 in complex with its high-affinity receptor interferon-λR1
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2010.09.068
PMCID: PMC2991516
NIHMSID: NIHMS244792
Accession Number: 20934432
Keywords: Amino Acid Sequence
Crystallography, X-Ray
Humans
Interferons
Interleukins/*chemistry/metabolism
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Quaternary
Receptors, Interferon/*chemistry/metabolism
Sequence Alignment
Abstract: Interferon (IFN)-λ1 [also known as interleukin (IL)-29] belongs to the recently discovered group of type III IFNs. All type III IFNs initiate signaling processes through formation of specific heterodimeric receptor complexes consisting of IFN-λR1 and IL-10R2. We have determined the structure of human IFN-λ1 complexed with human IFN-λR1, a receptor unique to type III IFNs. The overall structure of IFN-λ1 is topologically similar to the structure of IL-10 and other members of the IL-10 family of cytokines. IFN-λR1 consists of two distinct domains having fibronectin type III topology. The ligand-receptor interface includes helix A, loop AB, and helix F on the IFN site, as well as loops primarily from the N-terminal domain and inter-domain hinge region of IFN-λR1. Composition and architecture of the interface that includes only a few direct hydrogen bonds support an idea that long-range ionic interactions between ligand and receptor govern the process of initial recognition of the molecules while hydrophobic interactions finalize it.
Notes: 1089-8638
Miknis, Zachary J
Magracheva, Eugenia
Li, Wei
Zdanov, Alexander
Kotenko, Sergei V
Wlodawer, Alexander
HHSN261200800001C/RC/CCR NIH HHS/United States
HHSN261200800001E/CA/NCI NIH HHS/United States
R01 AI057468/AI/NIAID NIH HHS/United States
ZIA BC010348-10/Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, U.S. Gov't, Non-P.H.S.
J Mol Biol. 2010 Dec 10;404(4):650-64. doi: 10.1016/j.jmb.2010.09.068. Epub 2010 Oct 8.
Author Address: Macromolecular Crystallography Laboratory, NCI-Frederick, Frederick, MD 21702, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1808
Author: Mikulecký, P., Zahradník, J., Kolenko, P., Černý, J., Charnavets, T., Kolářová, L., Nečasová, I., Pham, P. N. and Schneider, B.
Year: 2016
Title: Crystal structure of human interferon-γ receptor 2 reveals the structural basis for receptor specificity
Journal: Acta Crystallogr D Struct Biol
Volume: 72
Issue: Pt 9
Pages: 1017-25
Epub Date: 2016/09/08
Date: Sep
Short Title: Crystal structure of human interferon-γ receptor 2 reveals the structural basis for receptor specificity
Alternate Journal: Acta crystallographica. Section D, Structural biology
ISSN: 2059-7983
DOI: 10.1107/s2059798316012237
PMCID: PMC5013595
Accession Number: 27599734
Keywords: Amino Acid Motifs
Crystallography, X-Ray
Disulfides/chemistry
Glycosylation
Humans
Models, Molecular
Protein Conformation
Protein Domains
Protein Folding
Protein Stability
Receptors, Interferon/*chemistry
*class 2 cytokine receptors
*fibronectin type III domain
*interferon-γ receptor 2
Abstract: Interferon-γ receptor 2 is a cell-surface receptor that is required for interferon-γ signalling and therefore plays a critical immunoregulatory role in innate and adaptive immunity against viral and also bacterial and protozoal infections. A crystal structure of the extracellular part of human interferon-γ receptor 2 (IFNγR2) was solved by molecular replacement at 1.8 Å resolution. Similar to other class 2 receptors, IFNγR2 has two fibronectin type III domains. The characteristic structural features of IFNγR2 are concentrated in its N-terminal domain: an extensive π-cation motif of stacked residues KWRWRH, a NAG-W-NAG sandwich (where NAG stands for N-acetyl-D-glucosamine) and finally a helix formed by residues 78-85, which is unique among class 2 receptors. Mass spectrometry and mutational analyses showed the importance of N-linked glycosylation to the stability of the protein and confirmed the presence of two disulfide bonds. Structure-based bioinformatic analysis revealed independent evolutionary behaviour of both receptor domains and, together with multiple sequence alignment, identified putative binding sites for interferon-γ and receptor 1, the ligands of IFNγR2.
Notes: 2059-7983
Mikulecký, Pavel
Zahradník, Jirí
Kolenko, Petr
Černý, Jiří
Charnavets, Tatsiana
Kolářová, Lucie
Nečasová, Iva
Pham, Phuong Ngoc
Schneider, Bohdan
Journal Article
Research Support, Non-U.S. Gov't
Acta Crystallogr D Struct Biol. 2016 Sep;72(Pt 9):1017-25. doi: 10.1107/S2059798316012237. Epub 2016 Aug 18.
Author Address: Institute of Biotechnology CAS, BIOCEV, Prumyslova 595, 252 50 Vestec, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1094
Author: Milder, F. J., Gomes, L., Schouten, A., Janssen, B. J., Huizinga, E. G., Romijn, R. A., Hemrika, W., Roos, A., Daha, M. R. and Gros, P.
Year: 2007
Title: Factor B structure provides insights into activation of the central protease of the complement system
Journal: Nat Struct Mol Biol
Volume: 14
Issue: 3
Pages: 224-8
Epub Date: 2007/02/21
Date: Mar
Short Title: Factor B structure provides insights into activation of the central protease of the complement system
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb1210
Accession Number: 17310251
Keywords: Catalytic Domain
Complement C3-C5 Convertases/chemistry
Complement Factor B/*chemistry/*metabolism
Complement System Proteins/*immunology
Crystallography, X-Ray
Enzyme Activation
Humans
Models, Molecular
Protein Structure, Secondary
Protein Structure, Tertiary
Regulatory Sequences, Nucleic Acid/genetics
Structure-Activity Relationship
von Willebrand Factor/chemistry
Abstract: Factor B is the central protease of the complement system of immune defense. Here, we present the crystal structure of human factor B at 2.3-A resolution, which reveals how the five-domain proenzyme is kept securely inactive. The canonical activation helix of the Von Willebrand factor A (VWA) domain is displaced by a helix from the preceding domain linker. The two helices conformationally link the scissile-activation peptide and the metal ion-dependent adhesion site required for binding of the ligand C3b. The data suggest that C3b binding displaces the three N-terminal control domains and reshuffles the two central helices. Reshuffling of the helices releases the scissile bond for final proteolytic activation and generates a new interface between the VWA domain and the serine protease domain. This allosteric mechanism is crucial for tight regulation of the complement-amplification step in the immune response.
Notes: Milder, Fin J
Gomes, Lucio
Schouten, Arie
Janssen, Bert J C
Huizinga, Eric G
Romijn, Roland A
Hemrika, Wieger
Roos, Anja
Daha, Mohamed R
Gros, Piet
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2007 Mar;14(3):224-8. doi: 10.1038/nsmb1210. Epub 2007 Feb 25.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Faculty of Sciences, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1076
Author: Milder, F. J., Raaijmakers, H. C., Vandeputte, M. D., Schouten, A., Huizinga, E. G., Romijn, R. A., Hemrika, W., Roos, A., Daha, M. R. and Gros, P.
Year: 2006
Title: Structure of complement component C2A: implications for convertase formation and substrate binding
Journal: Structure
Volume: 14
Issue: 10
Pages: 1587-97
Epub Date: 2006/10/10
Date: Oct
Short Title: Structure of complement component C2A: implications for convertase formation and substrate binding
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2006.08.008
Accession Number: 17027507
Keywords: Amino Acids/chemistry/genetics
Catalytic Domain
Complement Activation
Complement C2a/*chemistry/genetics
Humans
Ligands
*Models, Molecular
Mutation
Protein Structure, Secondary
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics
Substrate Specificity
Abstract: C2a provides the catalytic center to the convertase complexes of the classical and lectin-binding pathways of complement activation. We determined two crystal structures of full-length C2a, with and without a pseudo ligand bound. Both structures reveal a near-active conformation of the catalytic center of the serine protease domains, while the von Willebrand factor A-type domains display an intermediate activation state of helix alpha7 with an open, activated metal-ion-dependent adhesion site. The open adhesion site likely serves to enhance the affinity for the ligand C4b, similar to "inside-out" signaling in integrins. Surprisingly, the N-terminal residues of C2a are buried in a crevice near helix alpha7, indicative of a structural switch between C2 and C2a. Extended loops on the protease domain possibly envelop the protruding anaphylatoxin domain of the substrate C3. Together with a putative substrate-induced completion of the oxyanion hole, this may contribute to the high substrate specificity of the convertases.
Notes: Milder, Fin J
Raaijmakers, Hans C A
Vandeputte, Mitja D A A
Schouten, Arie
Huizinga, Eric G
Romijn, Roland A
Hemrika, Wieger
Roos, Anja
Daha, Mohamed R
Gros, Piet
Journal Article
Research Support, Non-U.S. Gov't
United States
Structure. 2006 Oct;14(10):1587-97. doi: 10.1016/j.str.2006.08.008.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 272
Author: Miletich, J. P. and Broze, G. J., Jr.
Year: 1990
Title: Beta protein C is not glycosylated at asparagine 329. The rate of translation may influence the frequency of usage at asparagine-X-cysteine sites
Journal: J Biol Chem
Volume: 265
Issue: 19
Pages: 11397-404
Epub Date: 1990/07/15
Date: Jul 5
Short Title: Beta protein C is not glycosylated at asparagine 329. The rate of translation may influence the frequency of usage at asparagine-X-cysteine sites
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1694179
Keywords: Amino Acid Sequence
Antibodies/immunology
Asparagine/*metabolism
Binding Sites
Blotting, Western
Carcinoma, Hepatocellular/metabolism
*Cysteine
Epitopes/immunology
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Liver Neoplasms/metabolism
Molecular Sequence Data
Oligosaccharides/analysis
*Protein Biosynthesis
Protein C/analysis/genetics/immunology/*metabolism
Thrombin/pharmacology
Tumor Cells, Cultured
Warfarin/pharmacology
Abstract: About 30% of human plasma protein C is smaller than the predominant form as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It has been suggested that this species, referred to as beta protein C, is a degraded molecule. However, beta protein C is secreted in culture by the HepG2 cell line and is present in plasma collected directly into numerous proteinase inhibitors; the percentage of beta protein C does not change with time during culture or after blood collection. Neither thrombin, activated protein C, nor activated factor X converts the alpha form to beta in the presence or absence of calcium and phospholipids. The NH2-terminal sequences of the heavy chains of both forms are identical, and both release the same dodecapeptide and develop a functional active site when cleaved by thrombin. Both also react with antibodies to a synthetic COOH-terminal peptide. Timed digests with N-glycosidase are consistent with the interpretation that beta protein C has three N-linked oligosaccharide chains whereas alpha protein C has four. It is asparagine 329 that is not glycosylated in beta protein C since antibodies to a synthetic peptide based on the sequence around this amino acid react only with beta protein C. This site is unique in having cysteine instead of serine or threonine 2 residues distal. It is likely that the sulfhydryl group can substitute for the usual hydroxyl group as a hydrogen bond acceptor for the glycosylation reaction only until it forms a disulfide bond. The percentage of protein C that is glycosylated at this site may therefore depend at least in part on the rate of disulfide bond formation which may in turn be related to the rate of protein synthesis.
Notes: Miletich, J P
Broze, G J Jr
HL 14147/HL/NHLBI NIH HHS/United States
HL 41963/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1990 Jul 5;265(19):11397-404.
Author Address: Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 887
Author: Miley, M. J., Truscott, S. M., Yu, Y. Y., Gilfillan, S., Fremont, D. H., Hansen, T. H. and Lybarger, L.
Year: 2003
Title: Biochemical features of the MHC-related protein 1 consistent with an immunological function
Journal: J Immunol
Volume: 170
Issue: 12
Pages: 6090-8
Epub Date: 2003/06/10
Date: Jun 15
Short Title: Biochemical features of the MHC-related protein 1 consistent with an immunological function
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
DOI: 10.4049/jimmunol.170.12.6090
Accession Number: 12794138
Keywords: Amino Acid Sequence
Animals
Antibodies, Monoclonal/biosynthesis
Cell Membrane/chemistry/genetics
Epitopes/immunology
H-2 Antigens/genetics
HeLa Cells
Histocompatibility Antigens Class I/biosynthesis/*chemistry/genetics/*immunology
Humans
Intracellular Fluid/chemistry
L Cells
Mice
Mice, Inbred BALB C
Minor Histocompatibility Antigens
Molecular Sequence Data
Peptides/genetics/metabolism
Protein Folding
Protein Structure, Tertiary/genetics
Recombinant Fusion Proteins/chemical synthesis/immunology
Solubility
Spodoptera
Transfection
Tumor Cells, Cultured
beta 2-Microglobulin/physiology
Abstract: MHC-related protein (MR)1 is an MHC class I-related molecule encoded on chromosome 1 that is highly conserved among mammals and is more closely related to classical class I molecules than are other nonclassical class I family members. In this report, we show for the first time that both mouse and human MR1 molecules can associate with the peptide-loading complex and can be detected at low levels at the surface of transfected cells. We also report the production of recombinant human MR1 molecules in insect cells using highly supplemented media and provide evidence that the MR1 H chain can assume a folded conformation and is stoichiometrically associated with beta(2)-microglobulin, similar to class I molecules. Cumulatively, these findings demonstrate that surface expression of MR1 is possible but may be limited by a specific ligand or associated molecule.
Notes: Miley, Michael J
Truscott, Steven M
Yu, Yik Yeung Lawrence
Gilfillan, Susan
Fremont, Daved H
Hansen, Ted H
Lybarger, Lonnie
AI46533/AI/NIAID NIH HHS/United States
T32AI07163/AI/NIAID NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Immunol. 2003 Jun 15;170(12):6090-8. doi: 10.4049/jimmunol.170.12.6090.
Author Address: Department of Pathology, Washington University School of Medicine, St. Louis, MO 63110, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 396
Author: Miller, J., Hatch, J. A., Simonis, S. and Cullen, S. E.
Year: 1988
Title: Identification of the glycosaminoglycan-attachment site of mouse invariant-chain proteoglycan core protein by site-directed mutagenesis
Journal: Proc Natl Acad Sci U S A
Volume: 85
Issue: 5
Pages: 1359-63
Epub Date: 1988/03/01
Date: Mar
Short Title: Identification of the glycosaminoglycan-attachment site of mouse invariant-chain proteoglycan core protein by site-directed mutagenesis
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.85.5.1359
PMCID: PMC279770
Accession Number: 3422739
Keywords: *Antigens, Differentiation, B-Lymphocyte
Chondroitin Sulfate Proteoglycans/*metabolism
DNA Mutational Analysis
Histocompatibility Antigens Class II/*genetics/metabolism
Proteoglycans/*metabolism
Structure-Activity Relationship
Abstract: The invariant chain (Ii), a nonpolymorphic glycoprotein that associates with the immunoregulatory Ia proteins encoded by the major histocompatibility complex, has a proteoglycan form (Ii-CS) that bears a chondroitin sulfate glycosaminoglycan. In this proteoglycan form, Ii may remain associated with Ia at the cell surface. Inhibitors that prevent the addition of glycosaminoglycan to Ii have been found to depress antigen-presenting function. Ii does not have multiple candidate glycosaminoglycan-attachment sites, and we used site-directed mutagenesis to replace a candidate serine glycosaminoglycan-acceptor site with alanine at position 201 in the murine Ii protein. Transfection of the normal or altered gene into Ii-negative COS-7 cells showed that equivalent amounts of core Ii protein and its acidic, terminally glycosylated forms were synthesized, but the Ala-201 mutant Ii did not give rise to Ii-CS. The mutant protein had apparently normal transport through the Golgi compartment and associated stably with Ia molecules. Thus, this mutation directly identifies the site of glycosaminoglycan addition and shows that it can be eliminated without adversely affecting the overall biosynthesis of Ii.
Notes: 1091-6490
Miller, J
Hatch, J A
Simonis, S
Cullen, S E
CA20500/CA/NCI NIH HHS/United States
CA33529/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1988 Mar;85(5):1359-63. doi: 10.1073/pnas.85.5.1359.
Author Address: Laboratory of Immunology, National Institutes of Health, Bethesda, MD 20892.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1648
Author: Miller, P. S. and Aricescu, A. R.
Year: 2014
Title: Crystal structure of a human GABAA receptor
Journal: Nature
Volume: 512
Issue: 7514
Pages: 270-5
Epub Date: 2014/06/10
Date: Aug 21
Short Title: Crystal structure of a human GABAA receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature13293
PMCID: PMC4167603
NIHMSID: EMS57867
Accession Number: 24909990
Keywords: Benzamidines/chemistry/metabolism/pharmacology
Binding Sites
Cell Membrane/chemistry/metabolism
Conserved Sequence
Crystallography, X-Ray
Drug Design
GABA-A Receptor Agonists/chemistry/metabolism/pharmacology
Genetic Predisposition to Disease
Glycosylation
Humans
Models, Molecular
Mutation/genetics
Polysaccharides/chemistry/metabolism
Protein Structure, Quaternary
Protein Structure, Tertiary
Protein Subunits
Receptors, GABA-A/*chemistry/genetics
Synaptic Transmission
Abstract: Type-A γ-aminobutyric acid receptors (GABAARs) are the principal mediators of rapid inhibitory synaptic transmission in the human brain. A decline in GABAAR signalling triggers hyperactive neurological disorders such as insomnia, anxiety and epilepsy. Here we present the first three-dimensional structure of a GABAAR, the human β3 homopentamer, at 3 Å resolution. This structure reveals architectural elements unique to eukaryotic Cys-loop receptors, explains the mechanistic consequences of multiple human disease mutations and shows an unexpected structural role for a conserved N-linked glycan. The receptor was crystallized bound to a previously unknown agonist, benzamidine, opening a new avenue for the rational design of GABAAR modulators. The channel region forms a closed gate at the base of the pore, representative of a desensitized state. These results offer new insights into the signalling mechanisms of pentameric ligand-gated ion channels and enhance current understanding of GABAergic neurotransmission.
Notes: 1476-4687
Miller, Paul S
Aricescu, A Radu
090532/Z/09/Z/Wellcome Trust/United Kingdom
MR/L009609/1/Medical Research Council/United Kingdom
G0700232/Medical Research Council/United Kingdom
090532/Wellcome Trust/United Kingdom
084655/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Nature. 2014 Aug 21;512(7514):270-5. doi: 10.1038/nature13293. Epub 2014 Jun 8.
Author Address: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1562
Author: Mimoto, F., Katada, H., Kadono, S., Igawa, T., Kuramochi, T., Muraoka, M., Wada, Y., Haraya, K., Miyazaki, T. and Hattori, K.
Year: 2013
Title: Engineered antibody Fc variant with selectively enhanced FcγRIIb binding over both FcγRIIa(R131) and FcγRIIa(H131)
Journal: Protein Eng Des Sel
Volume: 26
Issue: 10
Pages: 589-98
Epub Date: 2013/06/08
Date: Oct
Short Title: Engineered antibody Fc variant with selectively enhanced FcγRIIb binding over both FcγRIIa(R131) and FcγRIIa(H131)
Alternate Journal: Protein engineering, design & selection : PEDS
ISSN: 1741-0126 (Print)
1741-0126
DOI: 10.1093/protein/gzt022
PMCID: PMC3785249
Accession Number: 23744091
Keywords: Animals
Crystallography, X-Ray
Humans
Immunoglobulin Fc Fragments/chemistry/*genetics/*metabolism/pharmacology
Immunoglobulin G/chemistry
Mice
Models, Molecular
Mutagenesis
Platelet Activation/drug effects
Platelet Aggregation/drug effects
Protein Binding
Protein Conformation
*Protein Engineering
Receptors, IgG/*metabolism
Recombinant Proteins/chemistry/*genetics/*metabolism/pharmacology
Tumor Necrosis Factor Receptor Superfamily, Member 9/immunology
Fc engineering
FcγRIIb
antibody engineering
inhibitory FcγR
platelet activation
Abstract: Engaging inhibitory FcγRIIb by Fc region has been recently reported to be an attractive approach for improving the efficacy of antibody therapeutics. However, the previously reported S267E/L328F variant with enhanced binding affinity to FcγRIIb, also enhances binding affinity to FcγRIIa(R131) allotype to a similar degree because FcγRIIb and FcγRIIa(R131) are structurally similar. In this study, we applied comprehensive mutagenesis and structure-guided design based on the crystal structure of the Fc/FcγRIIb complex to identify a novel Fc variant with selectively enhanced FcγRIIb binding over both FcγRIIa(R131) and FcγRIIa(H131). This novel variant has more than 200-fold stronger binding affinity to FcγRIIb than wild-type IgG1, while binding affinity to FcγRIIa(R131) and FcγRIIa(H131) is comparable with or lower than wild-type IgG1. This selectivity was achieved by conformational change of the C(H)2 domain by mutating Pro to Asp at position 238. Fc variant with increased binding to both FcγRIIb and FcγRIIa induced platelet aggregation and activation in an immune complex form in vitro while our novel variant did not. When applied to agonistic anti-CD137 IgG1 antibody, our variant greatly enhanced the agonistic activity. Thus, the selective enhancement of FcγRIIb binding achieved by our Fc variant provides a novel tool for improving the efficacy of antibody therapeutics.
Notes: 1741-0134
Mimoto, F
Katada, H
Kadono, S
Igawa, T
Kuramochi, T
Muraoka, M
Wada, Y
Haraya, K
Miyazaki, T
Hattori, K
Journal Article
Research Support, Non-U.S. Gov't
Protein Eng Des Sel. 2013 Oct;26(10):589-98. doi: 10.1093/protein/gzt022. Epub 2013 Jun 5.
Author Address: Research Division, Chugai Pharmaceutical Co., Ltd., Gotemba, Shizuoka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 332
Author: Minamitake, Y., Kodama, S., Katayama, T., Adachi, H., Tanaka, S. and Tsujimoto, M.
Year: 1990
Title: Structure of recombinant human interleukin 5 produced by Chinese hamster ovary cells
Journal: J Biochem
Volume: 107
Issue: 2
Pages: 292-7
Epub Date: 1990/02/01
Date: Feb
Short Title: Structure of recombinant human interleukin 5 produced by Chinese hamster ovary cells
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a123041
Accession Number: 2361960
Keywords: Amino Acid Sequence
Animals
Cricetinae
Disulfides
Female
Glycosylation
Interleukin-5/*isolation & purification
Molecular Sequence Data
Ovary/cytology
Peptide Mapping
Recombinant Proteins/*isolation & purification
Abstract: The complete peptide map of purified recombinant human interleukin 5 (rhIL-5) was determined to verify its primary structure, glycosylation sites, and disulfide bonding structure. Each peptide fragment generated by Achromobacter protease I (API) digestion was purified and characterized by amino acid analysis and amino acid sequence analysis. After digestion with API, we could identify all the peptides which were expected from human IL-5 cDNA sequence. The analyses of sulfhydryl content in rhIL-5 molecule and disulfide-containing peptide obtained from API digestion indicated that active form of rhIL-5 existed as an antiparallel dimer linked by two pairs of Cys-44 and Cys-86. In addition, we concluded that Thr-3 and Asn-28 were glycosylated. The results indicate that primary structure of rhIL-5 is highly homogeneous and observed heterogeneity is due to the difference in the content of carbohydrate.
Notes: Minamitake, Y
Kodama, S
Katayama, T
Adachi, H
Tanaka, S
Tsujimoto, M
Journal Article
England
J Biochem. 1990 Feb;107(2):292-7. doi: 10.1093/oxfordjournals.jbchem.a123041.
Author Address: Suntory Institute for Biomedical Research, Osaka.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 784
Author: Minchiotti, L., Campagnoli, M., Rossi, A., Cosulich, M. E., Monti, M., Pucci, P., Kragh-Hansen, U., Granel, B., Disdier, P., Weiller, P. J. and Galliano, M.
Year: 2001
Title: A nucleotide insertion and frameshift cause albumin Kénitra, an extended and O-glycosylated mutant of human serum albumin with two additional disulfide bridges
Journal: Eur J Biochem
Volume: 268
Issue: 2
Pages: 344-52
Epub Date: 2001/02/13
Date: Jan
Short Title: A nucleotide insertion and frameshift cause albumin Kénitra, an extended and O-glycosylated mutant of human serum albumin with two additional disulfide bridges
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1033.2001.01899.x
Accession Number: 11168369
Keywords: Adenine
Amino Acid Sequence
Codon, Terminator
Cyanogen Bromide
Disulfides/chemistry
Exons
*Frameshift Mutation
Glycoproteins/*chemistry/*genetics
Humans
Jews
Mass Spectrometry
Molecular Sequence Data
Morocco
Oligosaccharides/chemistry
Peptide Fragments/chemistry
Peptide Mapping
Serum Albumin/*chemistry/*genetics
Serum Albumin, Human
Abstract: Albumin Kenitra is a new type of genetic variant of human serum albumin that has been found in two members of a family of Sephardic Jews from Kenitra (Morocco). The slow-migrating variant and the normal protein were isolated by anion-exchange chromatography and, after treatment with CNBr, the digests were analyzed by two-dimensional electrophoresis in a polyacrylamide gel. The CNBr peptides of the variant were purified by reverse-phase high performance liquid chromatography and submitted to sequence analysis. Albumin Kenitra is peculiar because it has an elongated polypeptide chain, 601 residues instead of 585, and its sequence is modified beginning from residue 575. DNA structural studies showed that the variant is caused by a single-base insertion, an adenine at nucleotide position 15 970 in the genomic sequence, which leads to a frameshift with the subsequent translation to the first termination codon of exon 15. Mass spectrometric analyses revealed that the four additional cysteine residues of the variant form two new S-S bridges and showed that albumin Kenitra is partially O-glycosylated by a monosialylated HexHexNAc structure. This oligosaccharide chain has been located to Thr596 by amino-acid sequence analysis of the tryptic fragment 592-597.
Notes: Minchiotti, L
Campagnoli, M
Rossi, A
Cosulich, M E
Monti, M
Pucci, P
Kragh-Hansen, U
Granel, B
Disdier, P
Weiller, P J
Galliano, M
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 2001 Jan;268(2):344-52. doi: 10.1046/j.1432-1033.2001.01899.x.
Author Address: Dipartimento di Biochimica A. Castellani, Università di Pavia, via Taramelli 3B, I-27100 Pavia, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1943
Author: Mire, E., Hocine, M., Bazellières, E., Jungas, T., Davy, A., Chauvet, S. and Mann, F.
Year: 2018
Title: Developmental Upregulation of Ephrin-B1 Silences Sema3C/Neuropilin-1 Signaling during Post-crossing Navigation of Corpus Callosum Axons
Journal: Curr Biol
Volume: 28
Issue: 11
Pages: 1768-1782.e4
Epub Date: 2018/05/22
Date: Jun 4
Short Title: Developmental Upregulation of Ephrin-B1 Silences Sema3C/Neuropilin-1 Signaling during Post-crossing Navigation of Corpus Callosum Axons
Alternate Journal: Current biology : CB
ISSN: 0960-9822
DOI: 10.1016/j.cub.2018.04.026
Accession Number: 29779877
Keywords: Animals
Axons/*physiology
Corpus Callosum/*physiology
Ephrin-B1/*genetics/metabolism
*Gene Expression Regulation, Developmental
Gene Silencing
Mice
Neuropilin-1/*genetics/metabolism
Semaphorins/*genetics/metabolism
*axon guidance
*corpus callosum
*cortex
*ephrins
*neuropilins
*semaphorins
Abstract: The corpus callosum is the largest commissure in the brain, whose main function is to ensure communication between homotopic regions of the cerebral cortex. During fetal development, corpus callosum axons (CCAs) grow toward and across the brain midline and then away on the contralateral hemisphere to their targets. A particular feature of this circuit, which raises a key developmental question, is that the outgoing trajectory of post-crossing CCAs is mirror-symmetric with the incoming trajectory of pre-crossing axons. Here, we show that post-crossing CCAs switch off their response to axon guidance cues, among which the secreted Semaphorin-3C (Sema3C), that act as attractants for pre-crossing axons on their way to the midline. This change is concomitant with an upregulation of the surface protein Ephrin-B1, which acts in CCAs to inhibit Sema3C signaling via interaction with the Neuropilin-1 (Nrp1) receptor. This silencing activity is independent of Eph receptors and involves a N-glycosylation site (N-139) in the extracellular domain of Ephrin-B1. Together, our results reveal a molecular mechanism, involving interaction between the two unrelated guidance receptors Ephrin-B1 and Nrp1, that is used to control the navigation of post-crossing axons in the corpus callosum.
Notes: 1879-0445
Mire, Erik
Hocine, Mélanie
Bazellières, Elsa
Jungas, Thomas
Davy, Alice
Chauvet, Sophie
Mann, Fanny
Journal Article
Research Support, Non-U.S. Gov't
England
Curr Biol. 2018 Jun 4;28(11):1768-1782.e4. doi: 10.1016/j.cub.2018.04.026. Epub 2018 May 17.
Author Address: Aix Marseille Univ, CNRS, IBDM, 13288 Marseille, France. Electronic address: mire.erik@gmail.com.
Aix Marseille Univ, CNRS, IBDM, 13288 Marseille, France.
Centre de Biologie du Développement (CBD), Centre de Biologie Intégrative (CBI), Université de Toulouse, CNRS, UPS, 118 Route de Narbonne, 31062 Toulouse, France.
Aix Marseille Univ, CNRS, IBDM, 13288 Marseille, France. Electronic address: fanny.mann@univ-amu.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 912
Author: Misaki, R., Nagaya, H., Fujiyama, K., Yanagihara, I., Honda, T. and Seki, T.
Year: 2003
Title: N-linked glycan structures of mouse interferon-beta produced by Bombyx mori larvae
Journal: Biochem Biophys Res Commun
Volume: 311
Issue: 4
Pages: 979-86
Epub Date: 2003/11/19
Date: Nov 28
Short Title: N-linked glycan structures of mouse interferon-beta produced by Bombyx mori larvae
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/j.bbrc.2003.10.094
Accession Number: 14623278
Keywords: Animals
Bombyx
Interferon-beta/*chemistry/genetics/*metabolism
Larva
Mice
Molecular Weight
Polysaccharides/*chemistry/*metabolism
Protein Conformation
Protein Engineering/methods
Protein Isoforms/chemistry/metabolism
Protein Structure, Tertiary
Recombinant Proteins
Species Specificity
Structure-Activity Relationship
Abstract: The full-length mouse interferon-beta (mIFN-beta) cDNA, including the secretion signal peptide coding region under control of the polyhedrin promoter, was introduced into Bombyx mori nucleopolyhedrovirus (BmNPV). Recombinant mIFN-beta (rmIFN-beta) was accumulated in the haemolymph of infected silkworm larvae. Western blot analysis showed isoforms of rmIFN-beta, suggesting that rmIFN-beta is glycosylated. The glycan structures of purified rmIFN-beta were determined. The N-glycans were liberated by hydrazinolysis and the resulting oligosaccharides were labeled with 2-aminopyridine. The pyridylaminated (PA) glycans were purified by gel filtration, reversed-phase HPLC, and size-fractionation HPLC. The structures of the PA-sugar chains were identified by a combination of two-dimensional PA-sugar chain mapping, MS analysis, and exoglycosidase digestions.
Notes: Misaki, Ryo
Nagaya, Hidekazu
Fujiyama, Kazuhito
Yanagihara, Itaru
Honda, Takeshi
Seki, Tatsuji
Comparative Study
Journal Article
United States
Biochem Biophys Res Commun. 2003 Nov 28;311(4):979-86. doi: 10.1016/j.bbrc.2003.10.094.
Author Address: The International Center for Biotechnology, Osaka University, 2-1 Yamada-oka, Suita, 565-0871, Osaka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1498
Author: Miura, Y., Sakurai, Y. and Endo, T.
Year: 2012
Title: O-GlcNAc modification affects the ATM-mediated DNA damage response
Journal: Biochim Biophys Acta
Volume: 1820
Issue: 10
Pages: 1678-85
Epub Date: 2012/07/05
Date: Oct
Short Title: O-GlcNAc modification affects the ATM-mediated DNA damage response
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbagen.2012.06.013
Accession Number: 22759405
Keywords: Acetylglucosamine/*metabolism/*physiology
Animals
Ataxia Telangiectasia Mutated Proteins
Cell Cycle Proteins/chemistry/*metabolism/*physiology
DNA Damage/*physiology
DNA Repair/physiology
DNA-Binding Proteins/chemistry/*metabolism/*physiology
Embryo, Mammalian
Enzyme Activation
Glycosylation
HeLa Cells
Humans
Mice
Mice, Inbred ICR
N-Acetylglucosaminyltransferases/*metabolism/physiology
Phosphorylation
Phosphotransferases/metabolism
Protein Processing, Post-Translational/physiology
Protein-Serine-Threonine Kinases/chemistry/*metabolism/*physiology
Tumor Suppressor Proteins/chemistry/*metabolism/*physiology
Abstract: BACKGROUND: O-Linked β-N-acetylglucosamine (O-GlcNAc) is a reversible, post-translational, and regulatory modification of nuclear, mitochondrial, and cytoplasmic proteins that is responsive to cellular stress. The role of O-GlcNAcylation in the ataxia-telangiectasia mutated (ATM)-mediated DNA damage response is unknown. It is unclear whether ATM, which is an early acting and central component of the signal transduction system activated by DNA double strand breaks, is an O-GlcNAc-modified protein. METHODS: The effect of O-GlcNAc modification on ATM activation was examined using two inhibitors, PUGNAc and DON that increase and decrease, respectively, levels of protein O-GlcNAcylation. To assess O-GlcNAcylation of ATM, immunoprecipitation and immunoblot analyses using anti-ATM or anti-O-GlcNAc antibody were performed in HeLa cells and primary cultured neurons. Interaction of ATM with O-GlcNAc transferase (OGT), the enzyme that adds O-GlcNAc to target proteins, was examined by immunoprecipitation and immunoblot analyses using anti-ATM. RESULTS: Enhancement of protein O-GlcNAcylation increased levels of X-irradiation-induced ATM activation. However, decreases in protein O-GlcNAcylation did not affect levels of ATM activation, but these decreases did delay ATM activation and ATM recovery processes based on assessment of de-phosphorylation of phospho-ATM. Thus, activation and recovery of ATM were affected by O-GlcNAcylation. ATM was subjected to O-GlcNAcylation, and ATM interacted with OGT. The steady-state O-GlcNAc level of ATM was not significantly responsive to X-irradiation or oxidative stress. GENERAL SIGNIFICANCE: ATM is an O-GlcNAc modified protein, and dynamic O-GlcNAc modification affects the ATM-mediated DNA damage response.
Notes: Miura, Yuri
Sakurai, Yoko
Endo, Tamao
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2012 Oct;1820(10):1678-85. doi: 10.1016/j.bbagen.2012.06.013. Epub 2012 Jul 1.
Author Address: Research Team for Mechanism of Aging, Tokyo Metropolitan Institute of Gerontology, Tokyo 173-0015, Japan. miura@tmig.or.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1655
Author: Miura, Y., Sato, T., Sakurai, Y., Sakai, R., Hiraoka, W. and Endo, T.
Year: 2014
Title: Hyper-O-GlcNAcylation inhibits the induction of heat shock protein 70 (Hsp 70) by sodium arsenite in HeLa cells
Journal: Biol Pharm Bull
Volume: 37
Issue: 8
Pages: 1308-14
Epub Date: 2014/08/05
Short Title: Hyper-O-GlcNAcylation inhibits the induction of heat shock protein 70 (Hsp 70) by sodium arsenite in HeLa cells
Alternate Journal: Biological & pharmaceutical bulletin
ISSN: 0918-6158
DOI: 10.1248/bpb.b14-00170
Accession Number: 25087952
Keywords: Acetylglucosamine/analogs & derivatives/pharmacology
Acylation
Arsenites/*toxicity
DNA-Binding Proteins/metabolism
Glucosamine/pharmacology
HSP70 Heat-Shock Proteins/genetics/*metabolism
HeLa Cells
Heat Shock Transcription Factors
Humans
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/*metabolism
Oxidative Stress/drug effects
Oximes/pharmacology
Phenylcarbamates/pharmacology
Protein Processing, Post-Translational
RNA, Messenger/metabolism
RNA, Small Interfering/genetics
Sodium Compounds/*toxicity
Transcription Factors/metabolism
Abstract: O-Linked β-N-acetylglucosamine-modification (O-GlcNAcylation) is a reversible, post-translational, and regulatory modification of nuclear, mitochondrial, and cytoplasmic proteins that is responsive to cellular stress. However, the role of O-GlcNAcylation in the induction of heat shock proteins (Hsps) by arsenite remains unclear. We used O-(2-acetamido-2-deoxy-D-glucopyranosylidene) amino N-phenyl carbamate (PUGNAc), an inhibitor of O-GlcNAcase, and glucosamine (GlcN), an enhancer of the hexosamine biosynthesis pathway, or O-GlcNAc transferase (OGT) short interfering RNA (siRNA) to enhance or suppress cellular O-GlcNAcylation levels, respectively, in HeLa cells. The exposure to arsenite increased O-GlcNAcylation and Hsp 70 levels in HeLa cells. However, the pre-treatment with PUGNAc or GlcN, which enhanced O-GlcNAcylation levels, decreased the arsenite-induced expression of Hsp 70. The pre-treatment with OGT siRNA, which suppressed O-GlcNAcylation levels, did not affect the induction of Hsp 70. We then examined the effects of O-GlcNAcylation on the nuclear translocation and phosphorylation of heat shock factor 1 (HSF1), and found that neither the nuclear translocation nor phosphorylation of HSF1 was regulated by O-GlcNAcylation. Finally, Hsp 70 mRNA expression was induced by arsenite, whereas the addition of PUGNAc slightly suppressed its induction. These results indicate that O-GlcNAcylation is related to arsenite-induced Hsp 70 expression, and demonstrated that hyper-O-GlcNAcylation inhibited the induction of Hsp 70 via transcriptional factors instead of HSF1.
Notes: 1347-5215
Miura, Yuri
Sato, Takatoshi
Sakurai, Yoko
Sakai, Ryo
Hiraoka, Wakako
Endo, Tamao
Journal Article
Research Support, Non-U.S. Gov't
Japan
Biol Pharm Bull. 2014;37(8):1308-14. doi: 10.1248/bpb.b14-00170.
Author Address: Research Team for Mechanism of Aging, Tokyo Metropolitan Institute of Gerontology.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1618
Author: Miyano, K. and Sumimoto, H.
Year: 2014
Title: N-Linked glycosylation of the superoxide-producing NADPH oxidase Nox1
Journal: Biochem Biophys Res Commun
Volume: 443
Issue: 3
Pages: 1060-5
Epub Date: 2013/12/25
Date: Jan 17
Short Title: N-Linked glycosylation of the superoxide-producing NADPH oxidase Nox1
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2013.12.086
Accession Number: 24365146
Keywords: Amino Acid Sequence
Animals
Asparagine/metabolism
CHO Cells
Cell Membrane/metabolism
Cricetinae
Cricetulus
Glycosylation
Humans
Membrane Glycoproteins/metabolism
Membrane Proteins/metabolism
Molecular Sequence Data
NADPH Oxidase 1
NADPH Oxidase 2
NADPH Oxidases/chemistry/*metabolism
Polysaccharides/metabolism
Protein Binding
Superoxides/*metabolism
Duox
Er
Endo H
NADPH oxidase
Nox
Nox1
PNGase F
Ros
Superoxide
dual oxidase
endoglycosidase H
endoplasmic reticulum
p22(phox)
peptide: N-glycosidase F
reactive oxygen species
Abstract: Nox1 is a membrane-integrated protein that belongs to the Nox family of superoxide-producing NADPH oxidases. Here we show that human Nox1 undergoes glycosylation at Asn-162 and Asn-236 in the second and third extracellular loops, respectively. Simultaneous threonine substitution for these residues completely abrogates the glycosylation, but does not prevent Nox1 from forming a heterodimer with p22(phox), trafficking to the cell surface, or producing superoxide. In the absence of p22(phox), Nox1 is transported to the plasma membrane mainly as a form with high mannose N-glycans, although their conversion into complex N-glycans is induced by expression of p22(phox). These findings indicate that glycosylation and subsequent N-glycan maturation of Nox1 are both dispensable for its cell surface recruitment. Superoxide production by unglycosylated Nox1 is largely dependent on p22(phox), which is abrogated by glutamine substitution for Pro-156 in p22(phox), a mutation leading to a defective interaction with the Nox1-activating protein Noxo1. Thus p22(phox) directly contributes to Nox1 activation in a glycosylation-independent manner, besides its significant role in Nox1 glycan maturation.
Notes: 1090-2104
Miyano, Kei
Sumimoto, Hideki
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2014 Jan 17;443(3):1060-5. doi: 10.1016/j.bbrc.2013.12.086. Epub 2013 Dec 21.
Author Address: Department of Biochemistry, Kyushu University Graduate School of Medical Sciences, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
Department of Biochemistry, Kyushu University Graduate School of Medical Sciences, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. Electronic address: hsumi@med.kyushu-u.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1567
Author: Mizuguchi-Hata, C., Ogawa, Y., Oka, M. and Yoneda, Y.
Year: 2013
Title: Quantitative regulation of nuclear pore complex proteins by O-GlcNAcylation
Journal: Biochim Biophys Acta
Volume: 1833
Issue: 12
Pages: 2682-2689
Epub Date: 2013/06/20
Date: Dec
Short Title: Quantitative regulation of nuclear pore complex proteins by O-GlcNAcylation
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbamcr.2013.06.008
Accession Number: 23777819
Keywords: Acetylglucosamine/*metabolism
Down-Regulation
Gene Knockdown Techniques
Glycosylation
HeLa Cells
Humans
Membrane Glycoproteins/*metabolism
N-Acetylglucosaminyltransferases
Nuclear Pore Complex Proteins/*metabolism
Protein Binding
RNA, Small Interfering/metabolism
Npc
Nuclear pore complex
Nup62
Nup88
O-GlcNAc transferase
O-GlcNAcase
O-GlcNAcylation
Oga
Ogt
siRNA
Abstract: The nuclear pore complex (NPC) is a macromolecular assembly consisting of approximately 30 different proteins called nucleoporins. Several nucleoporins are O-GlcNAcylated, which is a post-translational modification in which the monosaccharide β-N-acetylglucosamine (GlcNAc) is attached to serine or threonine residues within proteins. However, the biological significance of this modification on nucleoporins remains obscure. Here we found that Nup62 and Nup88 protein levels were significantly decreased upon knockdown of O-GlcNAc transferase (OGT), which catalyzes the O-GlcNAcylation of intracellular proteins. Although Nup88, unlike Nup62, was not recognized by an anti-O-GlcNAc antibody or WGA-HRP, knockdown of Nup62 caused a reduction in Nup88 protein levels, suggesting that the observed decrease in Nup88 in OGT knocked-down cells is due to a decrease in Nup62. Furthermore, we found that Nup88 was preferentially associated with O-GlcNAcylated Nup62 compared with non-O-GlcNAcylated Nup62. These results indicate that Nup62 protein levels are primarily maintained by O-GlcNAcylation and that Nup88 is quantitatively regulated through its interaction with O-GlcNAcylated Nup62.
Notes: Mizuguchi-Hata, Chiaki
Ogawa, Yutaka
Oka, Masahiro
Yoneda, Yoshihiro
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2013 Dec;1833(12):2682-2689. doi: 10.1016/j.bbamcr.2013.06.008. Epub 2013 Jun 15.
Author Address: Department of Frontier Biosciences, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamada-oka, Suita, Osaka 565-0871, Japan.
Department of Frontier Biosciences, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamada-oka, Suita, Osaka 565-0871, Japan; Department of Biochemistry, Graduate School of Medicine, Osaka University, 2-2 Yamada-oka, Suita, Osaka 565-0871, Japan; CREST, JST, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamada-oka, Suita, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1718
Author: Mizuhashi, K., Chaya, T., Kanamoto, T., Omori, Y. and Furukawa, T.
Year: 2015
Title: Obif, a Transmembrane Protein, Is Required for Bone Mineralization and Spermatogenesis in Mice
Journal: PLoS One
Volume: 10
Issue: 7
Pages: e0133704
Epub Date: 2015/07/25
Short Title: Obif, a Transmembrane Protein, Is Required for Bone Mineralization and Spermatogenesis in Mice
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0133704
PMCID: PMC4514473
Accession Number: 26207632
Keywords: Amino Acid Sequence
Animals
Calcification, Physiologic/*genetics
Cells, Cultured
Female
HEK293 Cells
Humans
Male
Membrane Proteins/genetics/*physiology
Mice
Mice, Knockout
Molecular Sequence Data
Sequence Homology, Amino Acid
Spermatocytes/physiology
Spermatogenesis/*genetics
Testis/growth & development/metabolism
Abstract: BACKGROUND: Various kinds of transmembrane and secreted proteins play pivotal roles in development through cell-cell communication. We previously reported that Obif (Osteoblast induction factor, Tmem119), encoding a single transmembrane protein, is expressed in differentiating osteoblasts, and that Obif-/- mice exhibit significantly reduced bone volume in the femur. In the current study, we characterized the Obif protein and further investigated the biological phenotypes of a variety of tissues in Obif-/- mice. RESULTS: First, we found that O-glycosylation of the Obif protein occurs at serine residue 36 in the Obif extracellular domain. Next, we observed that Obif-/- mice exhibit bone dysplasia in association with significantly increased osteoid volume per osteoid surface (OV/OS) and osteoid maturation time (Omt), and significantly decreased mineral apposition rate (MAR) and bone formation rate per bone surface (BFR/BS). In addition, we observed that Obif-/- mice show a significant decrease in testis weight as well as in sperm number. By histological analysis, we found that Obif is expressed in spermatocytes and spermatids in the developing testis and that spermatogenesis is halted at the round spermatid stage in the Obif-/- testis that lacks sperm. However, the number of litters fathered by male mice was slightly reduced in Obif-/- mice compared with wild-type mice, although this was not statistically significant. CONCLUSIONS: Our results, taken together with previous observations, indicate that Obif is a type Ia transmembrane protein whose N-terminal region is O-glycosylated. In addition, we found that Obif is required for normal bone mineralization and late testicular differentiation in vivo. These findings suggest that Obif plays essential roles in the development of multiple tissues.
Notes: 1932-6203
Mizuhashi, Koji
Chaya, Taro
Kanamoto, Takashi
Omori, Yoshihiro
Furukawa, Takahisa
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2015 Jul 24;10(7):e0133704. doi: 10.1371/journal.pone.0133704. eCollection 2015.
Author Address: Laboratory for Molecular and Developmental Biology, Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka, Japan; Department of Developmental Biology, Osaka Bioscience Institute, 6-2-4 Furuedai, Suita, Osaka, Japan.
Laboratory for Molecular and Developmental Biology, Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka, Japan; Department of Developmental Biology, Osaka Bioscience Institute, 6-2-4 Furuedai, Suita, Osaka, Japan; Japan Science and Technology Agency, Core Research for Evolutional Science and Technology, 3-2 Yamadaoka, Suita, Osaka, Japan.
Department of Developmental Biology, Osaka Bioscience Institute, 6-2-4 Furuedai, Suita, Osaka, Japan; Department of Orthopedic Surgery, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 496
Author: Mizuochi, T., Fujii, J., Kisiel, W. and Kobata, A.
Year: 1981
Title: Studies on the structures of the carbohydrate moiety of human prothrombin
Journal: J Biochem
Volume: 90
Issue: 4
Pages: 1023-31
Epub Date: 1981/10/01
Date: Oct
Short Title: Studies on the structures of the carbohydrate moiety of human prothrombin
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a133554
Accession Number: 7309709
Keywords: Carbohydrate Sequence
Chromatography, Gel
Electrophoresis, Paper
Glycoside Hydrolases/metabolism
Humans
Methylation
Prothrombin/*analysis
Abstract: Human prothrombin contains three asparagine-linked sugar chains in one molecule. The sugar chains were quantitatively liberated as radioactive oligosaccharides from the polypeptide moiety by hydrazinolysis followed by N-acetylation and Nab3H4 reduction. All of the oligosaccharides contain N-acetylneuraminic acid. The neutral oligosaccharides obtained from all acidic oligosaccharides by sialidase digestion are identical. By the combination of sequential exoglycosidase digestion and methylation analysis, the structures of the asparagine-linked sugar chains of human prothrombin were confirmed to be as follows: (sequence in text).
Notes: Mizuochi, T
Fujii, J
Kisiel, W
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
England
J Biochem. 1981 Oct;90(4):1023-31. doi: 10.1093/oxfordjournals.jbchem.a133554.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 472
Author: Mizuochi, T., Nishimura, R., Derappe, C., Taniguchi, T., Hamamoto, T., Mochizuki, M. and Kobata, A.
Year: 1983
Title: Structures of the asparagine-linked sugar chains of human chorionic gonadotropin produced in choriocarcinoma. Appearance of triantennary sugar chains and unique biantennary sugar chains
Journal: J Biol Chem
Volume: 258
Issue: 23
Pages: 14126-9
Epub Date: 1983/12/10
Date: Dec 10
Short Title: Structures of the asparagine-linked sugar chains of human chorionic gonadotropin produced in choriocarcinoma. Appearance of triantennary sugar chains and unique biantennary sugar chains
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6643471
Keywords: Adult
*Asparagine
Carbohydrate Sequence
Carbohydrates/analysis
Choriocarcinoma/*analysis
Chorionic Gonadotropin/*analysis
Female
Humans
Pregnancy
Uterine Neoplasms/*analysis
Abstract: Human chorionic gonadotropin (hCG) highly purified from urine of the patient with choriocarcinoma contains four asparagine-linked sugar chains in one molecule. The sugar chains were quantitatively liberated as radioactive oligosaccharides from polypeptide portion by hydrazinolysis followed by N-acetylation and NaB3H4 reduction. The structures of these sugar chains were determined by the combination of sequential glycosidase digestion, periodate oxidation, and methylation analysis. As compared with the sugar chains of normal urinary and placental hCG reported previously, they include several prominent structural differences. More than 97% of the sugar chains of choriocarcinoma hCG was free from sialic acid, while the sugar chains of normal hCG were mostly sialylated. Choriocarcinoma hCG contains unusual biantennary complex-type sugar chains in addition to regular tri-, bi-, and monoantennary sugar chains. These sugar chains have two outer chains linked at the C-2 and C-4 positions of the same alpha-mannosyl residue of the trimannosyl core. Since normal hCG does not contain any triantennary sugar chains, occurrence of Gal beta 1 leads to 4GlcNAc beta 1 leads to 4Man alpha 1 leads to group is another characteristic feature of the sugar chains of choriocarcinoma hCG. The evidence that the monoantennary sugar chain of Man alpha 1 leads to 6(Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAc leads to Asn is not found in normal hCG and the sum total of fucosylated sugar chains is 50%, which is twice as much as normal hCG, indicated that fucosylation is also modified in choriocarcinoma tissue.
Notes: Mizuochi, T
Nishimura, R
Derappe, C
Taniguchi, T
Hamamoto, T
Mochizuki, M
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1983 Dec 10;258(23):14126-9.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1437
Author: Mizushima, T., Yagi, H., Takemoto, E., Shibata-Koyama, M., Isoda, Y., Iida, S., Masuda, K., Satoh, M. and Kato, K.
Year: 2011
Title: Structural basis for improved efficacy of therapeutic antibodies on defucosylation of their Fc glycans
Journal: Genes Cells
Volume: 16
Issue: 11
Pages: 1071-80
Epub Date: 2011/10/26
Date: Nov
Short Title: Structural basis for improved efficacy of therapeutic antibodies on defucosylation of their Fc glycans
Alternate Journal: Genes to cells : devoted to molecular & cellular mechanisms
ISSN: 1356-9597 (Print)
1356-9597
DOI: 10.1111/j.1365-2443.2011.01552.x
PMCID: PMC3258418
Accession Number: 22023369
Keywords: Antibodies/chemistry/immunology/therapeutic use
*Antibody-Dependent Cell Cytotoxicity
Humans
Immunoglobulin Fc Fragments/chemistry/*immunology
Immunoglobulin G/chemistry/*immunology
Polysaccharides/*chemistry/immunology
Protein Conformation
Receptors, IgG/*chemistry/*immunology
Abstract: Removal of the fucose residue from the N-glycans of the Fc portion of immunoglobulin G (IgG) results in a dramatic enhancement of antibody-dependent cellular cytotoxicity (ADCC) through improved affinity for Fcγ receptor IIIa (FcγRIIIa). Here, we present the 2.2-Å structure of the complex formed between nonfucosylated IgG1-Fc and a soluble form of FcγRIIIa (sFcγRIIIa) with two N-glycosylation sites. The crystal structure shows that one of the two N-glycans of sFcγRIIIa mediates the interaction with nonfucosylated Fc, thereby stabilizing the complex. However, fucosylation of the Fc N-glycans inhibits this interaction, because of steric hindrance, and furthermore, negatively affects the dynamics of the receptor binding site. Our results offer a structural basis for improvement in ADCC of therapeutic antibodies by defucosylation.
Notes: 1365-2443
Mizushima, Tsunehiro
Yagi, Hirokazu
Takemoto, Emi
Shibata-Koyama, Mami
Isoda, Yuya
Iida, Shigeru
Masuda, Kazuhiro
Satoh, Mitsuo
Kato, Koichi
Journal Article
Research Support, Non-U.S. Gov't
Genes Cells. 2011 Nov;16(11):1071-80. doi: 10.1111/j.1365-2443.2011.01552.x.
Author Address: Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabe-dori, Mizuho-ku, Nagoya 467-8603, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 983
Author: Mizutani, Y., Kihara, A. and Igarashi, Y.
Year: 2005
Title: Mammalian Lass6 and its related family members regulate synthesis of specific ceramides
Journal: Biochem J
Volume: 390
Issue: Pt 1
Pages: 263-71
Epub Date: 2005/04/13
Date: Aug 15
Short Title: Mammalian Lass6 and its related family members regulate synthesis of specific ceramides
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20050291
PMCID: PMC1184580
Accession Number: 15823095
Keywords: Amino Acid Sequence
Animals
Cell Line
Ceramides/*biosynthesis
Cloning, Molecular
Endoplasmic Reticulum/chemistry
Gene Expression Regulation/*physiology
Humans
Intracellular Membranes/chemistry
Membrane Proteins/genetics/*metabolism
Mice
Molecular Sequence Data
Multigene Family/genetics/*physiology
Oxidoreductases/metabolism
Phylogeny
RNA, Messenger/metabolism
Substrate Specificity
Tissue Distribution
Abstract: The Lass (longevity-assurance homologue) family members, which are highly conserved among eukaryotes, function in ceramide synthesis. In the mouse, there are at least five Lass family members, Lass1, Lass2, Lass4, Lass5 and the hitherto uncharacterized Lass6. To investigate specific roles for each Lass member in ceramide synthesis, we cloned these five mouse proteins. Overproduction of any Lass protein in cultured cells resulted in an increase in cellular ceramide, but the ceramide species produced varied. Overproduction of Lass1 increased C18:0-ceramide levels preferentially, and overproduction of Lass2 and Lass4 increased levels of longer ceramides such as C22:0- and C24:0-ceramides. Lass5 and Lass6 produced shorter ceramide species (C14:0- and C16:0-ceramides); however, their substrate preferences towards saturated/unsaturated fatty acyl-CoA differed. In addition to differences in substrate preferences, we also demonstrated by Northern blotting that Lass family members are differentially expressed among tissues. Additionally, we found that Lass proteins differ with regard to glycosylation. Of the five members, only Lass2, Lass5 and Lass6 were N-glycosylated, each at their N-terminal Asn residue. The occurrence of N-glycosylation of some Lass proteins provides topological insight, indicating that the N-termini of Lass family members probably face the luminal side of the endoplasmic reticulum membrane. Furthermore, based on a proteinase K digestion assay, we demonstrated that the C-terminus of Lass6 faces the cytosolic side of the membrane. From these data we propose topology for the conserved Lag1 motif in Lass family members, namely that the N-terminal region faces the luminal side and the C-terminal region the cytosolic side of the endoplasmic reticulum membrane.
Notes: 1470-8728
Mizutani, Yukiko
Kihara, Akio
Igarashi, Yasuyuki
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2005 Aug 15;390(Pt 1):263-71. doi: 10.1042/BJ20050291.
Author Address: Department of Biomembrane and Biofunctional Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12-jo, Nishi 6-choume, Kita-ku, Sapporo 060-0812, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1080
Author: Mochizuki, K., Kagawa, T., Numari, A., Harris, M. J., Itoh, J., Watanabe, N., Mine, T. and Arias, I. M.
Year: 2007
Title: Two N-linked glycans are required to maintain the transport activity of the bile salt export pump (ABCB11) in MDCK II cells
Journal: Am J Physiol Gastrointest Liver Physiol
Volume: 292
Issue: 3
Pages: G818-28
Epub Date: 2006/11/04
Date: Mar
Short Title: Two N-linked glycans are required to maintain the transport activity of the bile salt export pump (ABCB11) in MDCK II cells
Alternate Journal: American journal of physiology. Gastrointestinal and liver physiology
ISSN: 0193-1857 (Print)
0193-1857
DOI: 10.1152/ajpgi.00415.2006
Accession Number: 17082223
Keywords: ATP Binding Cassette Transporter, Subfamily B, Member 11
ATP-Binding Cassette Transporters/chemistry/*physiology
Amino Acid Substitution
Animals
Bile Acids and Salts/*metabolism
Biological Transport, Active/drug effects
Cell Line
Cysteine Proteinase Inhibitors/pharmacology
Dogs
Epithelial Cells/drug effects/*metabolism
Glycosylation/drug effects
Kinetics
Leupeptins/pharmacology
Luminescent Proteins/genetics/metabolism
Mutation
Organic Anion Transporters, Sodium-Dependent/genetics/metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
Polysaccharides/*physiology
Proteasome Inhibitors
Protein Processing, Post-Translational/drug effects
Rats
Recombinant Fusion Proteins/genetics/metabolism
Symporters/genetics/metabolism
Taurocholic Acid/metabolism
Transfection
Tunicamycin/pharmacology
Abstract: The aim of this study was to determine the role of N-linked glycosylation in protein stability, intracellular trafficking, and bile acid transport activity of the bile salt export pump [Bsep (ATP-binding cassette B11)]. Rat Bsep was fused with yellow fluorescent protein, and the following mutants, in which Asn residues of putative glycosylation sites (Asn(109), Asn(116), Asn(122), and Asn(125)) were sequentially replaced with Gln, were constructed by site-directed mutagenesis: single N109Q, double N109Q + N116Q, triple N109Q + N116Q + N122Q, and quadruple N109Q + N116Q + N122Q + N125Q. Immunoblot and glycosidase cleavage analysis demonstrated that each site was glycosylated. Removal of glycans decreased taurocholate transport activity as determined in polarized MDCK II cells. This decrease resulted from rapid decay of the mutant Bsep protein; biochemical half-lives were 3.76, 3.65, 3.24, 1.35, and 0.52 h in wild-type, single-mutant, double-mutant, triple-mutant, and quadruple-mutant cells, respectively. Wild-type and single- and double-mutant proteins were distributed exclusively along the apical membranes, whereas triple- and quadruple-mutant proteins remained intracellular. MG-132 but not bafilomycin A(1) extended the half-life, suggesting a role for the proteasome in Bsep degradation. To determine whether a specific glycosylation site or the number of glycans was critical for protein stability, we studied the protein expression of combinations of N-glycan-deficient mutants and observed that Bsep with one glycan was considerably unstable compared with Bsep harboring two or more glycans. In conclusion, at least two N-linked glycans are required for Bsep protein stability, intracellular trafficking, and function in the apical membrane.
Notes: Mochizuki, Kaori
Kagawa, Tatehiro
Numari, Asano
Harris, Matthew J
Itoh, Johbu
Watanabe, Norihito
Mine, Tetsuya
Arias, Irwin M
DK-35652/DK/NIDDK NIH HHS/United States
DK-54785/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Am J Physiol Gastrointest Liver Physiol. 2007 Mar;292(3):G818-28. doi: 10.1152/ajpgi.00415.2006. Epub 2006 Nov 2.
Author Address: Department of Internal Medicine, Tokai University School of Medicine, Bohseidai, Isehara, Kanagawa 259-1193, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1586
Author: Moharir, A., Peck, S. H., Budden, T. and Lee, S. Y.
Year: 2013
Title: The role of N-glycosylation in folding, trafficking, and functionality of lysosomal protein CLN5
Journal: PLoS One
Volume: 8
Issue: 9
Pages: e74299
Epub Date: 2013/09/24
Short Title: The role of N-glycosylation in folding, trafficking, and functionality of lysosomal protein CLN5
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0074299
PMCID: PMC3769244
Accession Number: 24058541
Keywords: Endosomes
Glycosylation
HeLa Cells
Humans
Lysosome-Associated Membrane Glycoproteins
Lysosomes/*metabolism
Membrane Proteins/chemistry/*metabolism
Mutant Proteins/metabolism
Mutation/genetics
*Protein Folding
Protein Stability
Protein Transport
Subcellular Fractions/metabolism
rab5 GTP-Binding Proteins/metabolism
Abstract: CLN5 is a soluble lysosomal protein with unknown function. Mutations in CLN5 lead to neuronal ceroid lipofuscinosis, a group of inherited neurodegenerative disorders that mainly affect children. CLN5 has eight potential N-glycosylation sites based on the Asn-X-Thr/Ser consensus sequence. Through site-directed mutagenesis of individual asparagine residues to glutamine on each of the N-glycosylation consensus sites, we showed that all eight putative N-glycosylation sites are utilized in vivo. Additionally, localization studies showed that the lack of N-glycosylation on certain sites (N179, N252, N304, or N320) caused CLN5 retention in the endoplasmic reticulum, indicating that glycosylation is important for protein folding. Interestingly, one particular mutant, N401Q, is mislocalized to the Golgi, suggesting that N401 is not important for protein folding but essential for CLN5 trafficking to the lysosome. Finally, we analyzed several patient mutations in which N-glycosylation is affected. The N192S patient mutant is localized to the lysosome, indicating that this mutant has a functional defect in the lysosome. Our results suggest that there are functional differences in various N-glycosylation sites of CLN5 which affect folding, trafficking, and lysosomal function of CLN5.
Notes: 1932-6203
Moharir, Akshay
Peck, Sun H
Budden, Theodore
Lee, Stella Y
P20 GM103418/GM/NIGMS NIH HHS/United States
P20 RR016475/RR/NCRR NIH HHS/United States
5P20RR016475/RR/NCRR NIH HHS/United States
8P20GM103418/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
PLoS One. 2013 Sep 10;8(9):e74299. doi: 10.1371/journal.pone.0074299. eCollection 2013.
Author Address: Molecular, Cellular and Developmental Biology Program, Division of Biology, Kansas State University, Manhattan, Kansas, United States of America.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 467
Author: Mole, J. E., Anderson, J. K., Davison, E. A. and Woods, D. E.
Year: 1984
Title: Complete primary structure for the zymogen of human complement factor B
Journal: J Biol Chem
Volume: 259
Issue: 6
Pages: 3407-12
Epub Date: 1984/03/25
Date: Mar 25
Short Title: Complete primary structure for the zymogen of human complement factor B
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6546754
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cattle
Chymotrypsinogen
Cloning, Molecular
*Complement C3b/genetics
DNA/analysis
Humans
Peptide Fragments/analysis
Plasmids
Prothrombin
Trypsinogen
Abstract: The entire amino acid sequence of complement factor B has been established combining both protein and DNA sequencing strategies. The zymogen consists of 739 amino acids, has four asparagine-linked carbohydrate sites, and has independently disulfide-bonded NH2- and COOH-terminal regions. The catalytic subunit, Bb, is a unique serine protease containing 259 amino acids that are not integral to any of the classical serine proteases. It is proposed that this region of the Bb fragment functions as a cofactor-binding domain for C3b. The Ba fragment was found to contain three regions of internal sequence homology which were unrelated to the "kringle" regions of prothrombin and plasminogen and which suggest an independent evolution for the B genome. Sequence alignment of the active site of B to the serine proteases was made using the three-dimensional structures of chymotrypsin and trypsin as molecular models. Three stretches within the hypothetical model for B contrast markedly with all known serine proteases in both amino acid sequences and predicted configuration. It is suggested that these "altered" regions contribute at least in part to the formation of the catalytic region of the C3 convertase.
Notes: Mole, J E
Anderson, J K
Davison, E A
Woods, D E
AI 15417/AI/NIAID NIH HHS/United States
AI 19337/AI/NIAID NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1984 Mar 25;259(6):3407-12.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 723
Author: Monné, M., Gafvelin, G., Nilsson, R. and von Heijne, G.
Year: 1999
Title: N-tail translocation in a eukaryotic polytopic membrane protein: synergy between neighboring transmembrane segments
Journal: Eur J Biochem
Volume: 263
Issue: 1
Pages: 264-9
Epub Date: 1999/08/03
Date: Jul
Short Title: N-tail translocation in a eukaryotic polytopic membrane protein: synergy between neighboring transmembrane segments
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.1999.00498.x
Accession Number: 10429212
Keywords: Acetyltransferases
Animals
Binding Sites
Dogs
Fatty Acid Elongases
Glycosylation
In Vitro Techniques
Intracellular Membranes/metabolism
Membrane Proteins/*chemistry/genetics/*metabolism
Microsomes/metabolism
Mutation
Recombinant Proteins/chemistry/genetics/metabolism
Abstract: We have used the natural N-glycosylation site in the N-tail of cig30, a eukaryotic polytopic membrane protein, as a marker for N-tail translocation across the microsomal membrane. Analysis of C-terminally truncated cig30 constructs reveals that the first transmembrane segment is sufficient for translocation of the wild-type N-tail; in contrast, in a mutant with four arginines introduced into the N-tail the second transmembrane segment is also required for efficient N-tail translocation. Our observations imply a non-sequential assembly mechanism in which the ultimate location of the N-tail relative to the membrane may depend on more than one transmembrane segment.
Notes: Monné, M
Gafvelin, G
Nilsson, R
von Heijne, G
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1999 Jul;263(1):264-9. doi: 10.1046/j.1432-1327.1999.00498.x.
Author Address: Department of Biochemistry, Stockholm University, Stockholm, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1880
Author: Moon, S., Lee, Y. K., Lee, S. W. and Um, S. J.
Year: 2017
Title: Suppressive role of OGT-mediated O-GlcNAcylation of BAP1 in retinoic acid signaling
Journal: Biochem Biophys Res Commun
Volume: 492
Issue: 1
Pages: 89-95
Epub Date: 2017/08/15
Date: Oct 7
Short Title: Suppressive role of OGT-mediated O-GlcNAcylation of BAP1 in retinoic acid signaling
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2017.08.029
Accession Number: 28802580
Keywords: Alloxan/pharmacology
Cell Differentiation/drug effects
Cells, Cultured
Glycosylation/drug effects
Humans
N-Acetylglucosaminyltransferases/antagonists & inhibitors/*metabolism
*Signal Transduction
Structure-Activity Relationship
Tretinoin/*metabolism/pharmacology
Tumor Suppressor Proteins/*metabolism
Ubiquitin Thiolesterase/*metabolism
*Bap1
*Differentiation
*Embryonic stem cells
*O-GlcNAcylation
*Ogt
*Retinoic acid
Abstract: BRCA1-associated protein 1 (BAP1) has been implicated in diverse biological functions, including tumor suppression. However, its regulation via glycosylation and its role in embryonic stem (ES) cells are poorly defined. BAP1 was recently reported to interact with O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT). Here, we confirmed the physical interaction and investigated its functional significance. The O-GlcNAcylation of BAP1, which requires OGT, was examined in vivo and in vitro, and was proven using alloxan, an OGT inhibitor. OGT promoted the BAP1-induced repression of retinoic acid (RA)-induced RA receptor (RAR) activation. The repressive activity of BAP1 was relieved by alloxan but exacerbated by PUGNAc, an O-GlcNAcase (OGA) inhibitor. Finally, we addressed the role of O-GlcNAcylation in the RA-induced differentiation of murine ES cells. Alkaline phosphatase staining revealed the cooperation of RA and alloxan for impairing the pluripotency of ES cells. This cooperation was also observed by measuring the size of embryonic bodies and the expression of Sox2, a pluripotency marker. Overall, our data suggest that OGT-mediated O-GlcNAcylation of BAP1 prefers the maintenance of pluripotency, whereas its inhibition facilitates RA-induced differentiation in ES cells.
Notes: 1090-2104
Moon, Seungtae
Lee, Yong-Kyu
Lee, Sang-Wang
Um, Soo-Jong
Journal Article
United States
Biochem Biophys Res Commun. 2017 Oct 7;492(1):89-95. doi: 10.1016/j.bbrc.2017.08.029. Epub 2017 Aug 10.
Author Address: Department of Integrative Bioscience & Biotechnology/Institute of Bioscience, BK21 Graduate Program, Sejong University, Seoul 05006, South Korea.
Department of Integrative Bioscience & Biotechnology/Institute of Bioscience, BK21 Graduate Program, Sejong University, Seoul 05006, South Korea. Electronic address: umsj@sejong.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 740
Author: Morelle, W., Haslam, S. M., Ziak, M., Roth, J., Morris, H. R. and Dell, A.
Year: 2000
Title: Characterization of the N-linked oligosaccharides of megalin (gp330) from rat kidney
Journal: Glycobiology
Volume: 10
Issue: 3
Pages: 295-304
Epub Date: 2000/03/08
Date: Mar
Short Title: Characterization of the N-linked oligosaccharides of megalin (gp330) from rat kidney
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/10.3.295
Accession Number: 10704528
Keywords: Amidohydrolases/metabolism
Animals
Carbohydrate Conformation
Carbohydrate Sequence
Glycoside Hydrolases/metabolism
Glycosylation
Heymann Nephritis Antigenic Complex
Kidney/*chemistry/metabolism
Mass Spectrometry
Membrane Glycoproteins/*chemistry
Molecular Sequence Data
Monosaccharides/chemistry
Oligosaccharides/*chemistry
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Polysaccharides/chemistry
Rats
beta-Galactosidase/metabolism
Abstract: Megalin (gp 330) is a large cell surface receptor expressed on the apical surfaces of epithelial tissues, that mediates the binding and internalization of a number of structurally and functionally distinct ligands. In this paper we report the first detailed structural characterization of megalin-derived oligosaccharides. Using strategies based on mass spectrometric analysis, we have defined the structures of the N-glycans of megalin. The results reveal that megalin glycoprotein is heterogeneously glycosylated. The major N-glycans identified belong to the following two classes: high mannose structures and complex type structures, with complex structures being more abundant than high mannose structures. The major nonreducing epitopes in the complex-type glycans are: GlcNAc, Galbeta1-4GlcNAc (LacNAc), NeuAcalpha2-6Galbeta1-4GlcNAc (sialylated LacNAc), GalNAcbeta1-4[NeuAcalpha2-3]Galbeta1-4GlcNAc (Sd(a)) and Galalpha1-3Galbeta1-4GlcNAc. Most complex structures are characterized by the presence of (alpha1,6)-core fucosylation and the presence of a bisecting GlcNAc residue.
Notes: Morelle, W
Haslam, S M
Ziak, M
Roth, J
Morris, H R
Dell, A
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2000 Mar;10(3):295-304. doi: 10.1093/glycob/10.3.295.
Author Address: Department of Biochemistry, Imperial College, London, SW7 2AY, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1904
Author: Moremen, K. W., Ramiah, A., Stuart, M., Steel, J., Meng, L., Forouhar, F., Moniz, H. A., Gahlay, G., Gao, Z., Chapla, D., Wang, S., Yang, J. Y., Prabhakar, P. K., Johnson, R., Rosa, M. D., Geisler, C., Nairn, A. V., Seetharaman, J., Wu, S. C., Tong, L., Gilbert, H. J., LaBaer, J. and Jarvis, D. L.
Year: 2018
Title: Expression system for structural and functional studies of human glycosylation enzymes
Journal: Nat Chem Biol
Volume: 14
Issue: 2
Pages: 156-162
Epub Date: 2017/12/19
Date: Feb
Short Title: Expression system for structural and functional studies of human glycosylation enzymes
Alternate Journal: Nature chemical biology
ISSN: 1552-4450 (Print)
1552-4450
DOI: 10.1038/nchembio.2539
PMCID: PMC5774587
NIHMSID: EMS75575 Geisler is now an employee of GlycoBac, LLC, a biotechnology spinout that focuses on insect host cell improvements, but that could conceivably profit from the results described herein. The other authors have no competing financial interest.
Accession Number: 29251719
Keywords: Animals
Baculoviridae/metabolism
Crystallography, X-Ray
Cytidine Monophosphate/chemistry
*Gene Expression Profiling
Genetic Vectors
Glycoside Hydrolases/chemistry
Glycosylation
HEK293 Cells
Humans
Insecta
Kinetics
Recombinant Proteins/chemistry
Sialyltransferases/*chemistry
Sulfotransferases/chemistry
Abstract: Vertebrate glycoproteins and glycolipids are synthesized in complex biosynthetic pathways localized predominantly within membrane compartments of the secretory pathway. The enzymes that catalyze these reactions are exquisitely specific, yet few have been extensively characterized because of challenges associated with their recombinant expression as functional products. We used a modular approach to create an expression vector library encoding all known human glycosyltransferases, glycoside hydrolases, and sulfotransferases, as well as other glycan-modifying enzymes. We then expressed the enzymes as secreted catalytic domain fusion proteins in mammalian and insect cell hosts, purified and characterized a subset of the enzymes, and determined the structure of one enzyme, the sialyltransferase ST6GalNAcII. Many enzymes were produced at high yields and at similar levels in both hosts, but individual protein expression levels varied widely. This expression vector library will be a transformative resource for recombinant enzyme production, broadly enabling structure-function studies and expanding applications of these enzymes in glycochemistry and glycobiology.
Notes: 1552-4469
Moremen, Kelley W
Ramiah, Annapoorani
Stuart, Melissa
Steel, Jason
Meng, Lu
Forouhar, Farhad
Moniz, Heather A
Gahlay, Gagandeep
Gao, Zhongwei
Chapla, Digantkumar
Wang, Shuo
Yang, Jeong-Yeh
Prabhakar, Pradeep Kumar
Johnson, Roy
Rosa, Mitche Dela
Geisler, Christoph
Nairn, Alison V
Seetharaman, Jayaraman
Wu, Sheng-Cheng
Tong, Liang
Orcid: 0000-0002-0563-6468
Gilbert, Harry J
LaBaer, Joshua
Jarvis, Donald L
P41 GM103390/GM/NIGMS NIH HHS/United States
P41 GM103490/GM/NIGMS NIH HHS/United States
P01 GM107012/GM/NIGMS NIH HHS/United States
U54 GM094597/GM/NIGMS NIH HHS/United States
322820/European Research Council/International
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Chem Biol. 2018 Feb;14(2):156-162. doi: 10.1038/nchembio.2539. Epub 2017 Dec 18.
Author Address: Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia, USA.
Department of Molecular Biology, University of Wyoming, Laramie, Wyoming, USA.
Biodesign Institute, Arizona State University, Tempe, Arizona, USA.
Department of Biological Sciences, Northeast Structural Genomics Consortium, Columbia University, New York, New York, USA.
Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1223
Author: Morenilla-Palao, C., Pertusa, M., Meseguer, V., Cabedo, H. and Viana, F.
Year: 2009
Title: Lipid raft segregation modulates TRPM8 channel activity
Journal: J Biol Chem
Volume: 284
Issue: 14
Pages: 9215-24
Epub Date: 2009/01/30
Date: Apr 3
Short Title: Lipid raft segregation modulates TRPM8 channel activity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M807228200
PMCID: PMC2666574
Accession Number: 19176480
Keywords: Animals
Cell Line
Electrophysiological Phenomena
Glycosylation
Humans
Membrane Microdomains/*metabolism
Mice
Mutation/genetics
N-Acetylneuraminic Acid
Patch-Clamp Techniques
Protein Transport
TRPM Cation Channels/genetics/*metabolism
Abstract: Transient receptor potential channels are a family of cation channels involved in diverse cellular functions. Most of these channels are expressed in the nervous system and play a key role in sensory physiology. TRPM8 (transient receptor potential melastatine 8), a member of this family, is activated by cold, cooling substances such menthol and icilin and voltage. Although TRPM8 is a thermosensitive channel highly expressed in cold sensory neurons, the mechanisms underlying its temperature sensitivity are still poorly understood. Here we show that, in sensory neurons, TRPM8 channel is localized in cholesterol-rich specialized membrane domains known as lipid rafts. We also show that, in heterologous expression systems, lipid raft segregation of TRPM8 is favored by glycosylation at the Asn(934) residue of the polypeptide. In electrophysiological and imaging experiments, using cold and menthol as agonists, we also demonstrate that lipid raft association modulates TRPM8 channel activity. We found that menthol- and cold-mediated responses of TRPM8 are potentiated when the lipid raft association of the channel is prevented. In addition, lipid raft disruption shifts the threshold for TRPM8 activation to a warmer temperature. In view of these data, we suggest a role for lipid rafts in the activity and temperature sensitivity of TRPM8. We propose a model wherein different lipid membrane environments affect the cold sensing properties of TRPM8, modulating the response of cold thermoreceptors.
Notes: 1083-351x
Morenilla-Palao, Cruz
Pertusa, María
Meseguer, Víctor
Cabedo, Hugo
Viana, Félix
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2009 Apr 3;284(14):9215-24. doi: 10.1074/jbc.M807228200. Epub 2009 Jan 27.
Author Address: Instituto de Neurociencias de Alicante-UMH-CSIC and Unidad de Investigación del Hospital de San Juan de Alicante, San Juan de Alicante 03550, Spain. cruz@umh.es
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 227
Author: Morgan, F. J., Birken, S. and Canfield, R. E.
Year: 1975
Title: The amino acid sequence of human chorionic gonadotropin. The alpha subunit and beta subunit
Journal: J Biol Chem
Volume: 250
Issue: 13
Pages: 5247-58
Epub Date: 1975/07/10
Date: Jul 10
Short Title: The amino acid sequence of human chorionic gonadotropin. The alpha subunit and beta subunit
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1150658
Keywords: Amino Acid Sequence
Carboxypeptidases
Chorionic Gonadotropin/*analysis
Chymotrypsin
Cyanogen Bromide
Glucosamine/analysis
Humans
Thrombin
Trypsin
Abstract: The amino acid sequences of both the alpha and beta subunits of human chorionic gonadotropin have been determined. The amino acid sequence of the alpha subunit is: Ala - Asp - Val - Gln - Asp - Cys - Pro - Glu - Cys-10 - Thr - Leu - Gln - Asp - Pro - Phe - Ser - Gln-20 - Pro - Gly - Ala - Pro - Ile - Leu - Gln - Cys - Met - Gly-30 - Cys - Cys - Phe - Ser - Arg - Ala - Tyr - Pro - Thr - Pro-40 - Leu - Arg - Ser - Lys - Lys - Thr - Met - Leu - Val - Gln-50 - Lys - Asn - Val - Thr - Ser - Glu - Ser - Thr - Cys - Cys-60 - Val - Ala - Lys - Ser - Thr - Asn - Arg - Val - Thr - Val-70 - Met - Gly - Gly - Phe - Lys - Val - Glu - Asn - His - Thr-80 - Ala - Cys - His - Cys - Ser - Thr - Cys - Tyr - Tyr - His-90 - Lys - Ser. Oligosaccharide side chains are attached at residues 52 and 78. In the preparations studied approximately 10 and 30% of the chains lack the initial 2 and 3 NH2-terminal residues, respectively. This sequence is almost identical with that of human luteinizing hormone (Sairam, M. R., Papkoff, H., and Li, C. H. (1972) Biochem. Biophys. Res. Commun. 48, 530-537). The amino acid sequence of the beta subunit is: Ser - Lys - Glu - Pro - Leu - Arg - Pro - Arg - Cys - Arg-10 - Pro - Ile - Asn - Ala - Thr - Leu - Ala - Val - Glu - Lys-20 - Glu - Gly - Cys - Pro - Val - Cys - Ile - Thr - Val - Asn-30 - Thr - Thr - Ile - Cys - Ala - Gly - Tyr - Cys - Pro - Thr-40 - Met - Thr - Arg - Val - Leu - Gln - Gly - Val - Leu - Pro-50 - Ala - Leu - Pro - Gin - Val - Val - Cys - Asn - Tyr - Arg-60 - Asp - Val - Arg - Phe - Glu - Ser - Ile - Arg - Leu - Pro-70 - Gly - Cys - Pro - Arg - Gly - Val - Asn - Pro - Val - Val-80 - Ser - Tyr - Ala - Val - Ala - Leu - Ser - Cys - Gln - Cys-90 - Ala - Leu - Cys - Arg - Arg - Ser - Thr - Thr - Asp - Cys-100 - Gly - Gly - Pro - Lys - Asp - His - Pro - Leu - Thr - Cys-110 - Asp - Asp - Pro - Arg - Phe - Gln - Asp - Ser - Ser - Ser - Ser - Lys - Ala - Pro - Pro - Pro - Ser - Leu - Pro - Ser-130 - Pro - Ser - Arg - Leu - Pro - Gly - Pro - Ser - Asp - Thr-140 - Pro - Ile - Leu - Pro - Gln. Oligosaccharide side chains are found at residues 13, 30, 121, 127, 132, and 138. The proteolytic enzyme, thrombin, which appears to cleave a limited number of arginyl bonds, proved helpful in the determination of the beta sequence.
Notes: Morgan, F J
Birken, S
Canfield, R E
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1975 Jul 10;250(13):5247-58.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 439
Author: Morgan, P. H., Jacobs, H. G., Segrest, J. P. and Cunningham, L. W.
Year: 1970
Title: A comparative study of glycopeptides derived from selected vertebrate collagens. A possible role of the carbohydrate in fibril formation
Journal: J Biol Chem
Volume: 245
Issue: 19
Pages: 5042-8
Epub Date: 1970/10/10
Date: Oct 10
Short Title: A comparative study of glycopeptides derived from selected vertebrate collagens. A possible role of the carbohydrate in fibril formation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4319110
Keywords: Adult
Air Sacs
Amino Acid Sequence
Animals
Chromatography, Gel
Chromatography, Ion Exchange
*Collagen
Cyprinidae
Galactose/analysis
Glucose/analysis
Glycoproteins
Glycosides/isolation & purification
Guinea Pigs
Humans
Microbial Collagenase
Models, Biological
Models, Chemical
Models, Structural
Molecular Weight
Peptides/*analysis
Skin
Trypsin
Notes: Morgan, P H
Jacobs, H G
Segrest, J P
Cunningham, L W
Journal Article
United States
J Biol Chem. 1970 Oct 10;245(19):5042-8.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1532
Author: Mori, M., Kimura, H., Iwamura, Y., Arai, S. and Miyazaki, T.
Year: 2012
Title: Modification of N-glycosylation modulates the secretion and lipolytic function of apoptosis inhibitor of macrophage (AIM)
Journal: FEBS Lett
Volume: 586
Issue: 20
Pages: 3569-74
Epub Date: 2012/12/14
Date: Oct 19
Short Title: Modification of N-glycosylation modulates the secretion and lipolytic function of apoptosis inhibitor of macrophage (AIM)
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1016/j.febslet.2012.08.017
Accession Number: 23236605
Keywords: Animals
Apoptosis Regulatory Proteins
Glycosylation
HEK293 Cells
Humans
*Lipolysis
Mice
Mutagenesis, Site-Directed
Polysaccharides/metabolism
Receptors, Scavenger
Scavenger Receptors, Class B/genetics/*metabolism
Abstract: The mouse macrophage-derived apoptosis inhibitor of macrophage (AIM), which is incorporated into adipocytes and induces lipolysis by suppressing fatty acid synthase (FAS) activity, possesses three potential N-glycosylation sites. Inactivation of N-glycosylation sites revealed that mouse AIM contains two N-glycans in the first and second scavenger receptor cysteine-rich domains, and that depletion of N-glycans decreased AIM secretion from producing cells. Interestingly, the lack of N-glycans increased AIM lipolytic activity through enhancing AIM incorporation into adipocytes. Although human AIM contains no N-glycan, attachment of N-glycans increased AIM secretion. Thus, the N-glycosylation plays important roles in the secretion and lipolytic function of AIM.
Notes: 1873-3468
Mori, Mayumi
Kimura, Hiroki
Iwamura, Yoshihiro
Arai, Satoko
Miyazaki, Toru
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2012 Oct 19;586(20):3569-74. doi: 10.1016/j.febslet.2012.08.017.
Author Address: Laboratory of Molecular Biomedicine for Pathogenesis, Center for Disease Biology and Integrative Medicine, Faculty of Medicine, The University of Tokyo, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1209
Author: Mori, Y., Hamuro, T., Nakashima, T., Hamamoto, T., Natsuka, S., Hase, S. and Iwanaga, S.
Year: 2009
Title: Biochemical characterization of plasma-derived tissue factor pathway inhibitor: post-translational modification of free, full-length form with particular reference to the sugar chain
Journal: J Thromb Haemost
Volume: 7
Issue: 1
Pages: 111-20
Epub Date: 2008/11/20
Date: Jan
Short Title: Biochemical characterization of plasma-derived tissue factor pathway inhibitor: post-translational modification of free, full-length form with particular reference to the sugar chain
Alternate Journal: Journal of thrombosis and haemostasis : JTH
ISSN: 1538-7836
DOI: 10.1111/j.1538-7836.2008.03222.x
Accession Number: 19017259
Keywords: Carbohydrates/*analysis
Chromatography
Glycoproteins/analysis
Humans
Lipoproteins/blood/*chemistry/isolation & purification
Phosphorylation
Protease Inhibitors
*Protein Processing, Post-Translational
Abstract: BACKGROUND: Tissue factor pathway inhibitor (TFPI) is a physiological protease inhibitor that inhibits the initial reactions of the extrinsic blood coagulation pathway. Most TFPI in human plasma is associated with lipoproteins; however, the most functionally active form is thought to be the free, full-length form (f-pTFPI). Cell culture derived TFPI and recombinant TFPI (rTFPI) exhibit variations in their respective anticoagulant activity, which may be caused by post-translational modifications, such as the frequent differences in sugar chain structures among recombinant proteins. Sugar chain structures in rTFPI expressed in Chinese hamster ovary (CHO) cells have been reported previously, but those of plasma TFPI have not been. OBJECTIVES: To purify f-pTFPI and analyze the sugar chain structures. RESULTS AND CONCLUSION: f-pTFPI was purified to homogeneity from blood plasma using a combination of anion-exchange, heparin affinity, immunoaffinity, and reversed-phase chromatographies, resulting in a yield of 76%. f-pTFPI showed a partially phosphorylated glycoprotein comprising a total of 276 amino acids by peptide mapping. The sugar chain structures were analyzed by two-dimensional sugar mapping combined with exoglycosidase digestion of the pyridylamino sugar chains and the following results were obtained. (Sialyl) Galbeta1-3GalNAc was linked to Thr(175), partially to Thr(14) and Ser(174); sialyl complex-type sugar chains to Asn(117) and Asn(167), whereas Asn(228) was not glycosylated. Neuraminidase-resistant acidic sugar chains including sulfated sugar chains were not observed significantly. The protease inhibitory activities of f-pTFPI towards activated factor (F) X and tissue factor-activated FVII complex were identical to those of full-length rTFPI expressed in CHO cells.
Notes: 1538-7836
Mori, Y
Hamuro, T
Nakashima, T
Hamamoto, T
Natsuka, S
Hase, S
Iwanaga, S
Journal Article
England
J Thromb Haemost. 2009 Jan;7(1):111-20. doi: 10.1111/j.1538-7836.2008.03222.x. Epub 2008 Nov 11.
Author Address: Therapeutic Protein Products Research Department, The Chemo-Sero-Therapeutic Research Institute, KAKETSUKEN, Kumamoto, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1731
Author: Moriconi, C., Ordoñez, A., Lupo, G., Gooptu, B., Irving, J. A., Noto, R., Martorana, V., Manno, M., Timpano, V., Guadagno, N. A., Dalton, L., Marciniak, S. J., Lomas, D. A. and Miranda, E.
Year: 2015
Title: Interactions between N-linked glycosylation and polymerisation of neuroserpin within the endoplasmic reticulum
Journal: Febs j
Volume: 282
Issue: 23
Pages: 4565-79
Epub Date: 2015/09/15
Date: Dec
Short Title: Interactions between N-linked glycosylation and polymerisation of neuroserpin within the endoplasmic reticulum
Alternate Journal: The FEBS journal
ISSN: 1742-464X (Print)
1742-464x
DOI: 10.1111/febs.13517
PMCID: PMC4949553
Accession Number: 26367528
Keywords: Amino Acid Sequence
Animals
Biopolymers/genetics/metabolism
COS Cells
Cells, Cultured
Chlorocebus aethiops
Endoplasmic Reticulum/*metabolism
Glycosylation
Humans
Models, Molecular
Mutation
Neuropeptides/genetics/*metabolism
PC12 Cells
*Polymerization
Rats
Serpins/genetics/*metabolism
conformational disease
neurodegeneration
protein aggregation
serpin
serpinopathies
Abstract: The neuronal serpin neuroserpin undergoes polymerisation as a consequence of point mutations that alter its conformational stability, leading to a neurodegenerative dementia called familial encephalopathy with neuroserpin inclusion bodies (FENIB). Neuroserpin is a glycoprotein with predicted glycosylation sites at asparagines 157, 321 and 401. We used site-directed mutagenesis, transient transfection, western blot, metabolic labelling and ELISA to probe the relationship between glycosylation, folding, polymerisation and degradation of neuroserpin in validated cell models of health and disease. Our data show that glycosylation at N157 and N321 plays an important role in maintaining the monomeric state of neuroserpin, and we propose this is the result of steric hindrance or effects on local conformational dynamics that can contribute to polymerisation. Asparagine residue 401 is not glycosylated in wild type neuroserpin and in several polymerogenic variants that cause FENIB, but partial glycosylation was observed in the G392E mutant of neuroserpin that causes severe, early-onset dementia. Our findings indicate that N401 glycosylation reports lability of the C-terminal end of neuroserpin in its native state. This C-terminal lability is not required for neuroserpin polymerisation in the endoplasmic reticulum, but the additional glycan facilitates degradation of the mutant protein during proteasomal impairment. In summary, our results indicate how normal and variant-specific N-linked glycosylation events relate to intracellular folding, misfolding, degradation and polymerisation of neuroserpin.
Notes: 1742-4658
Moriconi, Claudia
Ordoñez, Adriana
Lupo, Giuseppe
Gooptu, Bibek
Irving, James A
Noto, Rosina
Martorana, Vincenzo
Manno, Mauro
Timpano, Valentina
Guadagno, Noemi A
Dalton, Lucy
Marciniak, Stefan J
Lomas, David A
Miranda, Elena
Orcid: 0000-0002-0586-8795
GGP11057/Telethon/Italy
G0601840/Medical Research Council/United Kingdom
G0901786-E01/2/Medical Research Council/United Kingdom
G1002610/Medical Research Council/United Kingdom
100140/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
FEBS J. 2015 Dec;282(23):4565-79. doi: 10.1111/febs.13517. Epub 2015 Oct 3.
Author Address: Department of Biology and Biotechnologies 'Charles Darwin', Sapienza University of Rome, Italy.
Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, UK.
Department of Chemistry, Sapienza University of Rome, Italy.
Division of Asthma, Allergy and Lung Biology, King's College London, UK.
Wolfson Institute for Biomedical Research, University College London, UK.
National Research Council of Italy, Institute of Biophysics, Palermo, Italy.
Pasteur Institute - Cenci Bolognetti Foundation, Sapienza University of Rome, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 633
Author: Morimoto, K., Tsuda, E., Said, A. A., Uchida, E., Hatakeyama, S., Ueda, M. and Hayakawa, T.
Year: 1996
Title: Biological and physicochemical characterization of recombinant human erythropoietins fractionated by Mono Q column chromatography and their modification with sialyltransferase
Journal: Glycoconj J
Volume: 13
Issue: 6
Pages: 1013-20
Epub Date: 1996/12/01
Date: Dec
Short Title: Biological and physicochemical characterization of recombinant human erythropoietins fractionated by Mono Q column chromatography and their modification with sialyltransferase
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/bf01053197
Accession Number: 8981093
Keywords: Animals
CHO Cells/chemistry/metabolism
Carbohydrate Sequence
Chemical Fractionation
Chromatography/methods
Cricetinae
Electrophoresis/methods
Erythropoietin/*chemistry/metabolism/*pharmacology
Female
Humans
Isoelectric Focusing
Mice
Mice, Inbred ICR
Molecular Sequence Data
Polysaccharides/analysis/chemistry
Recombinant Proteins/*chemistry/genetics/metabolism
Sialic Acids/analysis/chemistry
Sialyltransferases/*metabolism
Abstract: Ten erythropoietin (EPO) fractions differing in sialic acid content, ranging from 9.5 to 13.8 mol mol-1 of EPO, were obtained from baby hamster kidney cell-derived recombinant human EPO by Mono Q column chromatography. The mean pI values of the EPO fractions determined by IEF-gel electrophoresis systematically shifted from 4.11 to 3.31, coinciding with the sialic acid content, without a change in the constitution of asialo N-linked oligosaccharides of each fraction. Although a linear relationship between the in vivo bioactivity and the sialic acid content of the fractionated samples was observed until 12.1 mol mol-1 of EPO, there was no further increase in their activity over 12.4 mol mol-1 of EPO. On the other hand, an inverse relationship between the in vitro bioactivity and sialic acid content of EPO was observed. Also, we showed that the in vivo bioactivity of some fractions with low sialic acid contents was increased after treatment with alpha 2,6-sialyltransferase, but the in vivo bioactivity of the other fractions with high sialic acid contents was either decreased or not affected.
Notes: Morimoto, K
Tsuda, E
Said, A A
Uchida, E
Hatakeyama, S
Ueda, M
Hayakawa, T
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 1996 Dec;13(6):1013-20. doi: 10.1007/BF01053197.
Author Address: Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1763
Author: Morita, J., Kano, K., Kato, K., Takita, H., Sakagami, H., Yamamoto, Y., Mihara, E., Ueda, H., Sato, T., Tokuyama, H., Arai, H., Asou, H., Takagi, J., Ishitani, R., Nishimasu, H., Nureki, O. and Aoki, J.
Year: 2016
Title: Structure and biological function of ENPP6, a choline-specific glycerophosphodiester-phosphodiesterase
Journal: Sci Rep
Volume: 6
Pages: 20995
Epub Date: 2016/02/19
Date: Feb 18
Short Title: Structure and biological function of ENPP6, a choline-specific glycerophosphodiester-phosphodiesterase
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/srep20995
PMCID: PMC4757880
Accession Number: 26888014
Keywords: Animals
Crystallography, X-Ray
Endothelial Cells/enzymology
Fatty Liver/enzymology/genetics
Liver/enzymology
Mice
Mice, Knockout
Oligodendroglia/enzymology
Organ Specificity
Phosphatidylcholines/*chemistry/genetics/*metabolism
Phosphoric Diester Hydrolases/*chemistry/*metabolism
Structure-Activity Relationship
Substrate Specificity
Abstract: Choline is an essential nutrient for all living cells and is produced extracellularly by sequential degradation of phosphatidylcholine (PC). However, little is known about how choline is produced extracellularly. Here, we report that ENPP6, a choline-specific phosphodiesterase, hydrolyzes glycerophosphocholine (GPC), a degradation product of PC, as a physiological substrate and participates in choline metabolism. ENPP6 is highly expressed in liver sinusoidal endothelial cells and developing oligodendrocytes, which actively incorporate choline and synthesize PC. ENPP6-deficient mice exhibited fatty liver and hypomyelination, well known choline-deficient phenotypes. The choline moiety of GPC was incorporated into PC in an ENPP6-dependent manner both in vivo and in vitro. The crystal structure of ENPP6 in complex with phosphocholine revealed that the choline moiety of the phosphocholine is recognized by a choline-binding pocket formed by conserved aromatic and acidic residues. The present study provides the molecular basis for ENPP6-mediated choline metabolism at atomic, cellular and tissue levels.
Notes: 2045-2322
Morita, Junko
Kano, Kuniyuki
Kato, Kazuki
Takita, Hiroyuki
Sakagami, Hideki
Yamamoto, Yasuo
Mihara, Emiko
Ueda, Hirofumi
Sato, Takanao
Tokuyama, Hidetoshi
Arai, Hiroyuki
Asou, Hiroaki
Takagi, Junichi
Ishitani, Ryuichiro
Nishimasu, Hiroshi
Nureki, Osamu
Aoki, Junken
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2016 Feb 18;6:20995. doi: 10.1038/srep20995.
Author Address: Department of Biological Sciences, Graduate School of Sciences, The University of Tokyo, 2-11-16, Yayoi, Bunkyo-ku, Tokyo, 113-0032, Japan.
Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3, Aoba, Aramaki, Aoba-ku, Sendai, Miyagi, 980-8578, Japan.
Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1, Hongo, Bumkyo-ku, Tokyo, 113-0033, Japan.
Institute for Protein Research, Osaka University, 3-2, Yamadaoka, Suita-shi, Osaka, 565-0871, Japan.
Center for Kampo Medicine, Keio University School of Medicine, 35, Shinanomachi, Shinjuku-ku, Tokyo, 160-0016, Japan.
PRESTO (Precursory Research for Embryonic Science and Technology), JST (Japan Science and Technology Agency), 4-1-8, Honcho, Kawaguchi, Saitama, 332-0012, Japan.
AMED (Japan Agency for Medical Research and Development)-CREST (Core Research for Evolutional Science and Technology), 1-7-1, Otemachi, Chiyoda-ku, Tokyo, 100-0004, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 779
Author: Moritz, R. L., Hall, N. E., Connolly, L. M. and Simpson, R. J.
Year: 2001
Title: Determination of the disulfide structure and N-glycosylation sites of the extracellular domain of the human signal transducer gp130
Journal: J Biol Chem
Volume: 276
Issue: 11
Pages: 8244-53
Epub Date: 2000/12/01
Date: Mar 16
Short Title: Determination of the disulfide structure and N-glycosylation sites of the extracellular domain of the human signal transducer gp130
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M009979200
Accession Number: 11098061
Keywords: Amino Acid Sequence
Antigens, CD/*chemistry/isolation & purification
Cytokine Receptor gp130
Disulfides/chemistry
Glycosylation
Humans
Membrane Glycoproteins/*chemistry/isolation & purification
Models, Molecular
Molecular Sequence Data
Peptide Mapping
Abstract: gp130 is the common signal transducing receptor subunit for the interleukin-6-type family of cytokines. Its extracellular region (sgp130) is predicted to consist of five fibronectin type III-like domains and an NH2-terminal Ig-like domain. Domains 2 and 3 constitute the cytokine-binding region defined by a set of four conserved cysteines and a WSXWS motif, respectively. Here we determine the disulfide structure of human sgp130 by peptide mapping, in the absence and presence of reducing agent, in combination with Edman degradation and mass spectrometry. Of the 13 cysteines present, 10 form disulfide bonds, two are present as free cysteines (Cys(279) and Cys(469)), and one (Cys(397)) is modified by S-cysteinylation. Of the 11 potential N-glycosylation sites, Asn(21), Asn(61), Asn(109), Asn(135), Asn(205), Asn(357), Asn(361), Asn(531), and Asn(542) are glycosylated but not Asn(224) and Asn(368). The disulfide bonds, Cys(112)-Cys(122) and Cys(150)-Cys(160), are consistent with known cytokine-binding region motifs. Unlike granulocyte colony-stimulating factor receptor, the connectivities of the four cysteines in the NH2-terminal domain of gp130 (Cys(6)-Cys(32) and Cys(26)-Cys(81)) are consistent with known superfamily of Ig-like domains. An eight-residue loop in domain 5 is tethered by Cys(436)-Cys(444). We have created a model predicting that this loop maintains Cys(469) in a reduced form, available for ligand-induced intramolecular disulfide bond formation. Furthermore, we postulate that domain 5 may play a role in the disulfide-linked homodimerization and activation process of gp130.
Notes: Moritz, R L
Hall, N E
Connolly, L M
Simpson, R J
Journal Article
United States
J Biol Chem. 2001 Mar 16;276(11):8244-53. doi: 10.1074/jbc.M009979200. Epub 2000 Nov 29.
Author Address: Joint Protein Structure Laboratory, Ludwig Institute for Cancer Research (Melbourne Tumor Biology Branch) and the Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3050, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1715
Author: Morris, M., Knudsen, G. M., Maeda, S., Trinidad, J. C., Ioanoviciu, A., Burlingame, A. L. and Mucke, L.
Year: 2015
Title: Tau post-translational modifications in wild-type and human amyloid precursor protein transgenic mice
Journal: Nat Neurosci
Volume: 18
Issue: 8
Pages: 1183-9
Epub Date: 2015/07/21
Date: Aug
Short Title: Tau post-translational modifications in wild-type and human amyloid precursor protein transgenic mice
Alternate Journal: Nature neuroscience
ISSN: 1097-6256 (Print)
1097-6256
DOI: 10.1038/nn.4067
PMCID: PMC8049446
NIHMSID: NIHMS702155
Accession Number: 26192747
Keywords: Acetylation
Amyloid beta-Protein Precursor/*metabolism
Animals
Mass Spectrometry
Methylation
Mice
Mice, Inbred C57BL
Mice, Transgenic
Protein Processing, Post-Translational/*physiology
Ubiquitination
tau Proteins/*metabolism
Abstract: The microtubule-associated protein tau has been implicated in the pathogenesis of Alzheimer's disease (AD) and other neurodegenerative disorders. Reducing tau levels ameliorates AD-related synaptic, network, and behavioral abnormalities in transgenic mice expressing human amyloid precursor protein (hAPP). We used mass spectrometry to characterize the post-translational modification of endogenous tau isolated from wild-type and hAPP mice. We identified seven types of tau modifications at 63 sites in wild-type mice. Wild-type and hAPP mice had similar modifications, supporting the hypothesis that neuronal dysfunction in hAPP mice is enabled by physiological forms of tau. Our findings provide clear evidence for acetylation and ubiquitination of the same lysine residues; some sites were also targeted by lysine methylation. Our findings refute the hypothesis of extensive O-linked N-acetylglucosamine (O-GlcNAc) modification of endogenous tau. The complex post-translational modification of physiological tau suggests that tau is regulated by diverse mechanisms.
Notes: 1546-1726
Morris, Meaghan
Knudsen, Giselle M
Maeda, Sumihiro
Orcid: 0000000207016860
Trinidad, Jonathan C
Ioanoviciu, Alexandra
Burlingame, Alma L
Mucke, Lennart
P41 GM103481/GM/NIGMS NIH HHS/United States
R01 NS041787/NS/NINDS NIH HHS/United States
GM103481/GM/NIGMS NIH HHS/United States
NS041787/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Nat Neurosci. 2015 Aug;18(8):1183-9. doi: 10.1038/nn.4067. Epub 2015 Jul 20.
Author Address: 1] Gladstone Institute of Neurological Disease, San Francisco, California, USA. [2] Biochemistry, Cellular and Molecular Biology Graduate Program, Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Mass Spectrometry Facility, Department of Pharmaceutical Chemistry, University of California, San Francisco, California, USA.
1] Gladstone Institute of Neurological Disease, San Francisco, California, USA. [2] Department of Neurology, University of California, San Francisco, California, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1142
Author: Mosyak, L., Georgiadis, K., Shane, T., Svenson, K., Hebert, T., McDonagh, T., Mackie, S., Olland, S., Lin, L., Zhong, X., Kriz, R., Reifenberg, E. L., Collins-Racie, L. A., Corcoran, C., Freeman, B., Zollner, R., Marvell, T., Vera, M., Sum, P. E., Lavallie, E. R., Stahl, M. and Somers, W.
Year: 2008
Title: Crystal structures of the two major aggrecan degrading enzymes, ADAMTS4 and ADAMTS5
Journal: Protein Sci
Volume: 17
Issue: 1
Pages: 16-21
Epub Date: 2007/11/29
Date: Jan
Short Title: Crystal structures of the two major aggrecan degrading enzymes, ADAMTS4 and ADAMTS5
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.073287008
PMCID: PMC2144589
Accession Number: 18042673
Keywords: ADAM Proteins/*chemistry
ADAMTS4 Protein
ADAMTS5 Protein
Binding Sites
Crystallography, X-Ray
Enzyme Inhibitors/chemistry
Humans
Models, Molecular
Procollagen N-Endopeptidase/*chemistry
Protein Conformation
Abstract: Aggrecanases are now believed to be the principal proteinases responsible for aggrecan degradation in osteoarthritis. Given their potential as a drug target, we solved crystal structures of the two most active human aggrecanase isoforms, ADAMTS4 and ADAMTS5, each in complex with bound inhibitor and one wherein the enzyme is in apo form. These structures show that the unliganded and inhibitor-bound enzymes exhibit two essentially different catalytic-site configurations: an autoinhibited, nonbinding, closed form and an open, binding form. On this basis, we propose that mature aggrecanases exist as an ensemble of at least two isomers, only one of which is proteolytically active.
Notes: 1469-896x
Mosyak, Lidia
Georgiadis, Katy
Shane, Tania
Svenson, Kristine
Hebert, Tracy
McDonagh, Thomas
Mackie, Stewart
Olland, Stephane
Lin, Laura
Zhong, Xiaotian
Kriz, Ronald
Reifenberg, Erica L
Collins-Racie, Lisa A
Corcoran, Christopher
Freeman, Bethany
Zollner, Richard
Marvell, Tod
Vera, Matthew
Sum, Phaik-Eng
Lavallie, Edward R
Stahl, Mark
Somers, William
Journal Article
Protein Sci. 2008 Jan;17(1):16-21. doi: 10.1110/ps.073287008. Epub 2007 Nov 27.
Author Address: Department of Chemical and Screening Sciences, Wyeth Research, Cambridge, MA 02140, USA. lmosyak@wyeth.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1072
Author: Mosyak, L., Wood, A., Dwyer, B., Buddha, M., Johnson, M., Aulabaugh, A., Zhong, X., Presman, E., Benard, S., Kelleher, K., Wilhelm, J., Stahl, M. L., Kriz, R., Gao, Y., Cao, Z., Ling, H. P., Pangalos, M. N., Walsh, F. S. and Somers, W. S.
Year: 2006
Title: The structure of the Lingo-1 ectodomain, a module implicated in central nervous system repair inhibition
Journal: J Biol Chem
Volume: 281
Issue: 47
Pages: 36378-90
Epub Date: 2006/09/29
Date: Nov 24
Short Title: The structure of the Lingo-1 ectodomain, a module implicated in central nervous system repair inhibition
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M607314200
Accession Number: 17005555
Keywords: Animals
Axons/metabolism
Biophysics/methods
CHO Cells
Cell Differentiation
Cell Membrane/metabolism
Central Nervous System/*injuries/*pathology
Cricetinae
Crystallography, X-Ray
Humans
Leucine/chemistry
Membrane Proteins/*chemistry/metabolism
Myelin Sheath/chemistry
Nerve Tissue Proteins/*chemistry/metabolism
Oligodendroglia/metabolism
Protein Structure, Tertiary
Abstract: Nogo receptor (NgR)-mediated control of axon growth relies on the central nervous system-specific type I transmembrane protein Lingo-1. Interactions between Lingo-1 and NgR, along with a complementary co-receptor, result in neurite and axonal collapse. In addition, the inhibitory role of Lingo-1 is particularly important in regulation of oligodendrocyte differentiation and myelination, suggesting that pharmacological modulation of Lingo-1 function could be a novel approach for nerve repair and remyelination therapies. Here we report on the crystal structure of the ligand-binding ectodomain of human Lingo-1 and show it has a bimodular, kinked structure composed of leucine-rich repeat (LRR) and immunoglobulin (Ig)-like modules. The structure, together with biophysical analysis of its solution properties, reveals that in the crystals and in solution Lingo-1 persistently associates with itself to form a stable tetramer and that it is its LRR-Ig-composite fold that drives such assembly. Specifically, in the crystal structure protomers of Lingo-1 associate in a ring-shaped tetramer, with each LRR domain filling an open cleft in an adjacent protomer. The tetramer buries a large surface area (9,200 A2) and may serve as an efficient scaffold to simultaneously bind and assemble the NgR complex components during activation on a membrane. Potential functional binding sites that can be identified on the ectodomain surface, including the site of self-recognition, suggest a model for protein assembly on the membrane.
Notes: Mosyak, Lidia
Wood, Andrew
Dwyer, Brian
Buddha, Madhavan
Johnson, Mark
Aulabaugh, Ann
Zhong, Xiaotian
Presman, Eleonora
Benard, Susan
Kelleher, Kerry
Wilhelm, James
Stahl, Mark L
Kriz, Ron
Gao, Ying
Cao, Zixuan
Ling, Huai-Ping
Pangalos, Menelas N
Walsh, Frank S
Somers, William S
Journal Article
United States
J Biol Chem. 2006 Nov 24;281(47):36378-90. doi: 10.1074/jbc.M607314200. Epub 2006 Sep 27.
Author Address: Department of Chemical and Screening sciences, Wyeth Research, Cambridge, Massachusetts 02140, USA. lmosyak@wyeth.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 782
Author: Mowla, S. J., Farhadi, H. F., Pareek, S., Atwal, J. K., Morris, S. J., Seidah, N. G. and Murphy, R. A.
Year: 2001
Title: Biosynthesis and post-translational processing of the precursor to brain-derived neurotrophic factor
Journal: J Biol Chem
Volume: 276
Issue: 16
Pages: 12660-6
Epub Date: 2001/01/22
Date: Apr 20
Short Title: Biosynthesis and post-translational processing of the precursor to brain-derived neurotrophic factor
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M008104200
Accession Number: 11152678
Keywords: Amino Acid Sequence
Animals
Brain-Derived Neurotrophic Factor/*biosynthesis/genetics/*metabolism/pharmacology
COS Cells
Cell Line
Chlorocebus aethiops
Embryo, Mammalian
Glycoside Hydrolases
Glycosylation
Hippocampus/*metabolism
Humans
Mice
Molecular Sequence Data
Mutagenesis, Site-Directed
Neuroglia/metabolism
Neurons/*metabolism
Phosphorylation
Protein Precursors/biosynthesis/genetics/*metabolism
*Protein Processing, Post-Translational
Receptor, trkB/drug effects/physiology
Recombinant Proteins/biosynthesis/metabolism
Transfection
Vaccinia virus/genetics
Abstract: We examined the biosynthesis and post-translational processing of the brain-derived neurotrophic factor precursor (pro-BDNF) in cells infected with a pro-BDNF-encoding vaccinia virus. Metabolic labeling, immunoprecipitation, and SDS-polyacrylamide gel electrophoresis reveal that pro-BDNF is generated as a 32-kDa precursor that is N-glycosylated and glycosulfated on a site, within the pro-domain. Some pro-BDNF is released extracellularly and is biologically active as demonstrated by its ability to mediate TrkB phosphorylation. The precursor undergoes N-terminal cleavage within the trans-Golgi network and/or immature secretory vesicles to generate mature BDNF (14 kDa). Small amounts of a 28-kDa protein that is immunoprecipitated with BDNF antibodies is also evident. This protein is generated in the endoplasmic reticulum through N-terminal cleavage of pro-BDNF at the Arg-Gly-Leu-Thr(57)- downward arrow-Ser-Leu site. Cleavage is abolished when Arg(54) is changed to Ala (R54A) by in vitro mutagenesis. Blocking generation of 28-kDa BDNF has no effect on the level of mature BDNF and blocking generation of mature BDNF with alpha(1)-PDX, an inhibitor of furin-like enzymes, does not lead to accumulation of the 28-kDa form. These data suggest that 28-kDa pro-BDNF is not an obligatory intermediate in the formation of the 14-kDa form in the constitutive secretory pathway.
Notes: Mowla, S J
Farhadi, H F
Pareek, S
Atwal, J K
Morris, S J
Seidah, N G
Murphy, R A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2001 Apr 20;276(16):12660-6. doi: 10.1074/jbc.M008104200. Epub 2001 Jan 10.
Author Address: Salk Institute, La Jolla, California 92037-1099, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1699
Author: Mozzi, A., Forcella, M., Riva, A., Difrancesco, C., Molinari, F., Martin, V., Papini, N., Bernasconi, B., Nonnis, S., Tedeschi, G., Mazzucchelli, L., Monti, E., Fusi, P. and Frattini, M.
Year: 2015
Title: NEU3 activity enhances EGFR activation without affecting EGFR expression and acts on its sialylation levels
Journal: Glycobiology
Volume: 25
Issue: 8
Pages: 855-68
Epub Date: 2015/04/30
Date: Aug
Short Title: NEU3 activity enhances EGFR activation without affecting EGFR expression and acts on its sialylation levels
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwv026
Accession Number: 25922362
Keywords: Cell Line, Tumor
Cell Membrane
Colon/metabolism/pathology
Epithelial Cells/*metabolism/pathology
ErbB Receptors/*genetics/metabolism
G(M3) Ganglioside/metabolism
*Gene Expression Regulation, Neoplastic
Humans
Models, Molecular
Neoplasm Proteins/*genetics/metabolism
Neuraminidase/*genetics/metabolism
Phosphorylation
*Protein Processing, Post-Translational
Sialic Acids/metabolism
Signal Transduction
Transcription, Genetic
Egfr
Neu3
cell lines
colorectal cancer
sialylation
Abstract: Several studies performed over the last decade have focused on the role of sialylation in the progression of cancer and, in particular, on the association between deregulation of sialidases and tumorigenic transformation. The plasma membrane-associated sialidase NEU3 is often deregulated in colorectal cancer (CRC), and it was shown that this enzyme co-immunoprecipitates in HeLa cells with epidermal growth factor receptor (EGFR), the molecular target of most recent monoclonal antibody-based therapies against CRC. To investigate the role of NEU3 sialidase on EGFR deregulation in CRC, we first collected data on NEU3 gene expression levels from a library of commercial colon cell lines, demonstrating that NEU3 transcription is upregulated in these cell lines. We also found EGFR to be hyperphosphorylated in all cell lines, with the exception of SW620 cells and the CCD841 normal intestinal cell line. By comparing the effects induced by overexpression of either the wild-type or the inactive mutant form of NEU3 on EGFR, we demonstrated that the active form of NEU3 enhanced receptor activation without affecting EGFR mRNA or protein expression. Moreover, through western blots and mass spectrometry analysis, we found that EGFR immunoprecipitated from cells overexpressing active NEU3, unlike the receptor from mock cells and cells overexpressing inactive NEU3, is desialylated. On the whole, our data demonstrate that, besides the already reported indirect EGFR activation through GM3, sialidase NEU3 could also play a role on EGFR activation through its desialylation.
Notes: 1460-2423
Mozzi, Alessandra
Forcella, Matilde
Riva, Alice
Difrancesco, Carlotta
Molinari, Francesca
Martin, Vittoria
Papini, Nadia
Bernasconi, Barbara
Nonnis, Simona
Tedeschi, Gabriella
Mazzucchelli, Luca
Monti, Eugenio
Fusi, Paola
Frattini, Milo
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2015 Aug;25(8):855-68. doi: 10.1093/glycob/cwv026. Epub 2015 Apr 28.
Author Address: Department of Biotechnologies and Biosciences, University of Milano-Bicocca, Milano 20126, Italy.
Laboratory of Molecular Pathology, Institute of Pathology, Locarno CH-6600, Switzerland.
Department of Medical Biotechnology and Translational Medicine, University of Milano, Segrate 20090, Italy.
Department of Surgical and Morphological Sciences, University of Insubria, Varese 21100, Italy.
Department of Veterinary Science and Public Health, University of Milano, Milano 20133, Italy Filarete Foundation, Milano 20100, Italy.
Department of Molecular and Translational Medicine (DMTM), University of Brescia, Brescia 25123, Italy eugenio.monti@unibs.it milo.frattini@ti.ch.
Laboratory of Molecular Pathology, Institute of Pathology, Locarno CH-6600, Switzerland eugenio.monti@unibs.it milo.frattini@ti.ch.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 695
Author: Mu, J. and Roach, P. J.
Year: 1998
Title: Characterization of human glycogenin-2, a self-glucosylating initiator of liver glycogen metabolism
Journal: J Biol Chem
Volume: 273
Issue: 52
Pages: 34850-6
Epub Date: 1998/12/18
Date: Dec 25
Short Title: Characterization of human glycogenin-2, a self-glucosylating initiator of liver glycogen metabolism
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.52.34850
Accession Number: 9857012
Keywords: Animals
Fibroblasts/metabolism
Glucose/metabolism
Glucosyltransferases
Glycogen/metabolism
Glycoproteins/genetics/isolation & purification/*metabolism
Glycosylation
Humans
Liver Glycogen/*metabolism
Protein Binding
Protein Isoforms/genetics/isolation & purification/metabolism
Rats
Recombinant Proteins/metabolism
Tissue Distribution
Abstract: Glycogenin-2 is a recently described self-glucosylating protein potentially involved in the initiation of glycogen biosynthesis (Mu, J., Skurat, A. V., and Roach, P. J. (1997) J. Biol. Chem. 272, 27589-27597). In human liver extracts, most of the glycogenin-2 was only detectable after treatment with alpha-amylase. Similarly, purifed high Mr glycogen was only detected after release by alpha-amylase treatment. Based on analysis by polymerase chain reaction, the predominant isoform in liver was glycogenin-2beta. Glycogenin-2 was found in Ewing's sarcoma RD-ES cells where, however, it was not associated with high Mr carbohydrate. Both human liver and human RD-ES cell extracts also contained glycogenin-1. Glycogenin-1 and glycogenin-2 interact with one another, based on in vitro interactions and co-immunoprecipitation from liver and cell extracts. Mutation of Tyr-196 in glycogenin-2 to a Phe residue abolished the ability of glycogenin-2 to self-glucosylate but not to interact with glycogenin-1. Stable overexpression of glycogenin-2alpha in Rat-1 fibroblast cells resulted in a 5-fold increase in the level of glycogen present in the low speed pellet but little change in the low speed supernatant. This result is important since it indicates that the level of glycogenin-2 can determine glycogen accumulation and hence has the potential to control glycogen synthesis.
Notes: Mu, J
Roach, P J
DK20542/DK/NIDDK NIH HHS/United States
DK27221/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1998 Dec 25;273(52):34850-6. doi: 10.1074/jbc.273.52.34850.
Author Address: Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana 46202-5122, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 420
Author: Mueckler, M., Caruso, C., Baldwin, S. A., Panico, M., Blench, I., Morris, H. R., Allard, W. J., Lienhard, G. E. and Lodish, H. F.
Year: 1985
Title: Sequence and structure of a human glucose transporter
Journal: Science
Volume: 229
Issue: 4717
Pages: 941-5
Epub Date: 1985/09/06
Date: Sep 6
Short Title: Sequence and structure of a human glucose transporter
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.3839598
Accession Number: 3839598
Keywords: Amino Acid Sequence
Animals
*Carrier Proteins/genetics/metabolism
Cell Membrane/ultrastructure
Cloning, Molecular
DNA/genetics
Erythrocytes/metabolism
Glucose/*metabolism
Humans
Liver Neoplasms, Experimental/metabolism
*Membrane Proteins/genetics/metabolism
Molecular Weight
Monosaccharide Transport Proteins
Protein Conformation
RNA, Messenger/genetics
Tissue Distribution
Abstract: The amino acid sequence of the glucose transport protein from human HepG2 hepatoma cells was deduced from analysis of a complementary DNA clone. Structural analysis of the purified human erythrocyte glucose transporter by fast atom bombardment mapping and gas phase Edman degradation confirmed the identity of the clone and demonstrated that the HepG2 and erythrocyte transporters are highly homologous and may be identical. The protein lacks a cleavable amino-terminal signal sequence. Analysis of the primary structure suggests the presence of 12 membrane-spanning domains. Several of these may form amphipathic alpha helices and contain abundant hydroxyl and amide side chains that could participate in glucose binding or line a transmembrane pore through which the sugar moves. The amino terminus, carboxyl terminus, and a highly hydrophilic domain in the center of the protein are all predicted to lie on the cytoplasmic face. Messenger RNA species homologous to HepG2 glucose transporter messenger RNA were detected in K562 leukemic cells, HT29 colon adenocarcinoma cells, and human kidney tissue.
Notes: Mueckler, M
Caruso, C
Baldwin, S A
Panico, M
Blench, I
Morris, H R
Allard, W J
Lienhard, G E
Lodish, H F
GM22996/GM/NIGMS NIH HHS/United States
HL32262/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Science. 1985 Sep 6;229(4717):941-5. doi: 10.1126/science.3839598.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1106
Author: Mukhopadhyay, S. S. and Rosen, J. M.
Year: 2007
Title: The C-terminal domain of the nuclear factor I-B2 isoform is glycosylated and transactivates the WAP gene in the JEG-3 cells
Journal: Biochem Biophys Res Commun
Volume: 358
Issue: 3
Pages: 770-6
Epub Date: 2007/05/22
Date: Jul 6
Short Title: The C-terminal domain of the nuclear factor I-B2 isoform is glycosylated and transactivates the WAP gene in the JEG-3 cells
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/j.bbrc.2007.04.185
PMCID: PMC1942171
NIHMSID: NIHMS25427
Accession Number: 17511965
Keywords: Acetylglucosamine/chemistry
Cell Line, Tumor
Glycosylation
Humans
Milk Proteins/chemistry/*genetics
NFI Transcription Factors/*chemistry
Protein Isoforms
Protein Structure, Tertiary
Receptors, Glucocorticoid/chemistry
Recombinant Fusion Proteins/chemistry
STAT5 Transcription Factor/chemistry
Threonine/chemistry
Transcription, Genetic
Transcriptional Activation
Tumor Suppressor Proteins
Abstract: The transcription factor nuclear factor I (NFI) has been shown previously both in vivo and in vitro to be involved in the cooperative regulation of whey acidic protein (WAP) gene transcription along with the glucocorticoid receptor and STAT5. In addition, one of the specific NFI isoforms, NFI-B2, was demonstrated in transient co-transfection experiments in JEG cells, which lack endogenous NFI, to be preferentially involved in the cooperative regulation of WAP gene expression. A comparison of the DNA-binding specificities of the different NFI isoforms only partially explained their differential ability to activate the WAP gene transcription. Here, we analyzed the transactivation regions of two NFI isoforms by making chimeric proteins between the NFI-A and B isoforms. Though, their DNA-binding specificities were not altered as compared to the corresponding wild-type transcription factors, the C-terminal region of the NFI-B isoform was shown to preferentially activate WAP gene transcription in cooperation with GR and STAT5 in transient co-transfection assays in JEG-3 cells. Furthermore, determination of serine and threonine-specific glycosylation (O-linked N-acetylglucosamine) of the C-terminus of the NFI-B isoform suggested that the secondary modification by O-GlcNAc might play a role in the cooperative regulation of WAP gene transcription by NFI-B2 and STAT5.
Notes: 1090-2104
Mukhopadhyay, Sudit S
Rosen, Jeffrey M
R01 CA016303/CA/NCI NIH HHS/United States
R01 CA016303-24/CA/NCI NIH HHS/United States
R37 CA016303/CA/NCI NIH HHS/United States
CA 16303/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Biochem Biophys Res Commun. 2007 Jul 6;358(3):770-6. doi: 10.1016/j.bbrc.2007.04.185. Epub 2007 May 7.
Author Address: Department of Molecular and Cancer Genetics, M.D. Anderson Cancer Center, Houston, TX 77030, USA. suditmukhopadhy@yahoo.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1170
Author: Munz, M., Fellinger, K., Hofmann, T., Schmitt, B. and Gires, O.
Year: 2008
Title: Glycosylation is crucial for stability of tumour and cancer stem cell antigen EpCAM
Journal: Front Biosci
Volume: 13
Pages: 5195-201
Epub Date: 2008/05/30
Date: May 1
Short Title: Glycosylation is crucial for stability of tumour and cancer stem cell antigen EpCAM
Alternate Journal: Frontiers in bioscience : a journal and virtual library
ISSN: 1093-9946 (Print)
1093-4715
DOI: 10.2741/3075
Accession Number: 18508581
Keywords: 3T3 Cells
Animals
Antigens, Neoplasm/genetics/*metabolism
Asparagine/genetics
Cell Adhesion Molecules/genetics/*metabolism
Cell Line
Epithelial Cell Adhesion Molecule
Glycoside Hydrolases/pharmacology
Glycosylation
Humans
Kidney
Mice
Mutation
Neoplasms/*physiopathology
Abstract: Epithelial cell adhesion molecule EpCAM is strongly over-expressed in a variety of carcinomas where it is involved in signalling events resulting in increased expression of target genes such as c-Myc, cyclins and others, eventually conferring cells an oncogenic phenotype. However, EpCAM is also expressed in a series of healthy epithelia, albeit generally to a far lesser extend. We have uncovered differential glycosylation of EpCAM as a means to discriminate normal from malignant tissues. EpCAM was hyperglycosylated in carcinoma tissue as compared with autologous normal epithelia. All three N-glycosylation consensus sequences within EpCAM's extracellular domain were used in human and murine cells. We show that glycosylation at asparagine198 is crucial for protein stability. Mutants of EpCAM that substitute asparagine198 for alanine showed a decreased overall expression and half-life of the molecule at the plasma membrane. This is of considerable importance with respect to EpCAM variants expressed in normal tissue, where it might reveal to be less stable and thus may have repercussions on functionality.
Notes: Munz, Markus
Fellinger, Karin
Hofmann, Tanja
Schmitt, Barbel
Gires, Olivier
Journal Article
Research Support, Non-U.S. Gov't
United States
Front Biosci. 2008 May 1;13:5195-201. doi: 10.2741/3075.
Author Address: Clinical Cooperation Group Molecular Oncology, GSF-Research Center for Environment and Health, and Head and Neck Research Dept. Munich, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 985
Author: Murakami, M., Masuda, S., Shimbara, S., Ishikawa, Y., Ishii, T. and Kudo, I.
Year: 2005
Title: Cellular distribution, post-translational modification, and tumorigenic potential of human group III secreted phospholipase A(2)
Journal: J Biol Chem
Volume: 280
Issue: 26
Pages: 24987-98
Epub Date: 2005/05/03
Date: Jul 1
Short Title: Cellular distribution, post-translational modification, and tumorigenic potential of human group III secreted phospholipase A(2)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M502088200
Accession Number: 15863501
Keywords: Adenoviridae/genetics
Amino Acid Sequence
Animals
Arachidonic Acid/chemistry
Arthritis, Rheumatoid/pathology
Blotting, Northern
Catalytic Domain
Cell Line
Cell Line, Tumor
Cell Proliferation
Colorectal Neoplasms/pathology
Dose-Response Relationship, Drug
Endothelium, Vascular/cytology
Female
Fibroblasts/metabolism
*Gene Expression Regulation, Neoplastic
Glycoside Hydrolases/metabolism
Glycosylation
Group III Phospholipases A2
Humans
Immunohistochemistry
Inflammation
Ischemia/pathology
Lentivirus/metabolism
Lung/pathology
Mice
Mice, Nude
Microcirculation
Microscopy, Confocal
Molecular Sequence Data
Mutagenesis, Site-Directed
Neovascularization, Pathologic
Phospholipases A/*chemistry/*physiology
Point Mutation
Prostaglandins/chemistry
*Protein Processing, Post-Translational
Protein Structure, Tertiary
Reverse Transcriptase Polymerase Chain Reaction
Time Factors
Uterine Neoplasms/pathology
Abstract: Human group III secreted phospholipase A(2) (sPLA(2)-III) consists of a central group III sPLA(2) domain flanked by unique N- and C-terminal domains. We found that the sPLA(2) domain alone was sufficient for its catalytic activity and for its prostaglandin E(2) (PGE(2))-generating functions in various cell types. In several if not all cell types, the N- and C-terminal domains of sPLA(2)-III were proteolytically removed, leading to the production of the form containing only the sPLA(2) domain, which could be further N-glycosylated at two consensus sites. Immunohistochemistry demonstrated that sPLA(2)-III was preferentially expressed in the microvascular endothelium in human tissues with inflammation, ischemic injury, and cancer. In support of this, sPLA(2)-III was induced in cultured microvascular endothelial cells after stimulation with proinflammatory cytokines. Expression of sPLA(2)-III was also associated with various tumor cells, and colorectal cancer cells transfected with sPLA(2)-III exhibited enhanced PGE(2) production and cell proliferation, which required sPLA(2)-III catalytic activity. When implanted into nude mice, the sPLA(2)-III-transfected cells formed larger solid tumors with increased angiogenesis compared with control cells. Moreover, small interfering RNA for sPLA(2)-III significantly reduced PGE(2) production and proliferation of colorectal cancer cells. Taken together, these results reveal unique cell type-specific processing and N-glycosylation of sPLA(2)-III and the potential role of this enzyme in cancer development by stimulating tumor cell growth and angiogenesis.
Notes: Murakami, Makoto
Masuda, Seiko
Shimbara, Satoko
Ishikawa, Yukio
Ishii, Toshiharu
Kudo, Ichiro
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2005 Jul 1;280(26):24987-98. doi: 10.1074/jbc.M502088200. Epub 2005 Apr 29.
Author Address: Department of Health Chemistry, School of Pharmaceutical Sciences, Showa University, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo 142-8555, Japan. mako@pharm.showa-u.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1596
Author: Muta, K., Fukami, T., Nakajima, M. and Yokoi, T.
Year: 2014
Title: N-Glycosylation during translation is essential for human arylacetamide deacetylase enzyme activity
Journal: Biochem Pharmacol
Volume: 87
Issue: 2
Pages: 352-9
Epub Date: 2013/10/16
Date: Jan 15
Short Title: N-Glycosylation during translation is essential for human arylacetamide deacetylase enzyme activity
Alternate Journal: Biochemical pharmacology
ISSN: 0006-2952
DOI: 10.1016/j.bcp.2013.10.001
Accession Number: 24125761
Keywords: Amino Acid Sequence
Carboxylic Ester Hydrolases/genetics/*metabolism
Cell Line
Enzyme Activation/genetics
Glycosylation
Humans
Microsomes, Liver/enzymology
Molecular Sequence Data
Protein Biosynthesis/genetics
Aadac
Arylacetamide deacetylase
Ces
Dmso
Er
Endo H
Enzyme activity
Gpt
GlcN-Ac-1-phosphate transferase
Hlm
N-Glycosylation
PNGase F
Phenacetin
Protein folding
Snp
carboxylesterase
dimethyl sulfoxide
endoglycosidase H
endoplasmic reticulum
human liver microsomes
peptide: N-glycosidase F
single nucleotide polymorphism
α-Glc
α-glucosidase
Abstract: Human arylacetamide deacetylase (AADAC) can hydrolyze clinical drugs such as flutamide, phenacetin, and rifamycins. AADAC is a glycoprotein, but the role of glycosylation remains unclear. In the present study, we investigated the effect of glycosylation on AADAC enzyme activity. Immunoblot analysis of mutant AADACs that contained an asparagine (N, Asn) to glutamine (Q, Gln) substitution at either residue 78 or 282 (N78Q or N282Q) showed a different migration compared with the wild-type protein. A mutant AADAC that contained N to Q substitutions at both residue 78 and 282 (N78Q/N282Q) showed a similar migration to AADAC in human liver microsomes (HLM) treated with endoglycosidase H (Endo H), which produces deglycosylated proteins. This result indicated that AADAC was glycosylated at both N78 and N282. Mutant types of AADAC with the N282Q and the N78Q/N282Q substitutions showed dramatically lower phenacetin hydrolase activity than did the wild-type protein. The treatment of wild-type AADAC-expressing HuH-7 cells with tunicamycin, which produces unglycosylated protein, decreased AADAC enzyme activity. However, the treatment of the HLM with Endo H caused no decrease of AADAC activity. Thus, the oligosaccharide chain, per se, was not important for AADAC activity in the mature form. The mutant types of AADAC containing the N282Q and the N78Q/N282Q substitutions were not detected by immunoblotting analysis after non-reducing SDS-PAGE, suggesting that the glycosylation of AADAC at N282 was important for proper protein folding. Overall, this study found that the translational, but not post-translational, N-glycosylation of AADAC plays a crucial role in regulating AADAC enzyme activity.
Notes: 1873-2968
Muta, Kyotaka
Fukami, Tatsuki
Nakajima, Miki
Yokoi, Tsuyoshi
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem Pharmacol. 2014 Jan 15;87(2):352-9. doi: 10.1016/j.bcp.2013.10.001. Epub 2013 Oct 11.
Author Address: Drug Metabolism and Toxicology, Faculty of Pharmaceutical Sciences, Kanazawa University, Kakuma-machi, Kanazawa, Japan.
Drug Metabolism and Toxicology, Faculty of Pharmaceutical Sciences, Kanazawa University, Kakuma-machi, Kanazawa, Japan. Electronic address: tatsuki@p.kanazawa-u.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1098
Author: Muto, T., Tsuchiya, D., Morikawa, K. and Jingami, H.
Year: 2007
Title: Structures of the extracellular regions of the group II/III metabotropic glutamate receptors
Journal: Proc Natl Acad Sci U S A
Volume: 104
Issue: 10
Pages: 3759-64
Epub Date: 2007/03/16
Date: Mar 6
Short Title: Structures of the extracellular regions of the group II/III metabotropic glutamate receptors
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0611577104
PMCID: PMC1820657
Accession Number: 17360426
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Cysteine/chemistry
Humans
Ligands
Models, Molecular
Molecular Conformation
Molecular Sequence Data
Protein Conformation
Protein Structure, Tertiary
Rats
Receptors, G-Protein-Coupled/metabolism
Receptors, Metabotropic Glutamate/*chemistry/genetics
Sequence Homology, Amino Acid
Abstract: Metabotropic glutamate receptors play major roles in the activation of excitatory synapses in the central nerve system. We determined the crystal structure of the entire extracellular region of the group II receptor and that of the ligand-binding region of the group III receptor. A comparison among groups I, II, and III provides the structural basis that could account for the discrimination of group-specific agonists. Furthermore, the structure of group II includes the cysteine-rich domain, which is tightly linked to the ligand-binding domain by a disulfide bridge, suggesting a potential role in transmitting a ligand-induced conformational change into the downstream transmembrane region. The structure also reveals the lateral interaction between the two cysteine-rich domains, which could stimulate clustering of the dimeric receptors on the cell surface. We propose a general activation mechanism of the dimeric receptor coupled with both ligand-binding and interprotomer rearrangements.
Notes: 1091-6490
Muto, Takanori
Tsuchiya, Daisuke
Morikawa, Kosuke
Jingami, Hisato
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2007 Mar 6;104(10):3759-64. doi: 10.1073/pnas.0611577104. Epub 2007 Feb 26.
Author Address: Biomolecular Engineering Research Institute, 6-2-3 Furuedai, Suita, Osaka 565-0874, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1541
Author: Myers, S. A., Daou, S., Affar el, B. and Burlingame, A.
Year: 2013
Title: Electron transfer dissociation (ETD): the mass spectrometric breakthrough essential for O-GlcNAc protein site assignments-a study of the O-GlcNAcylated protein host cell factor C1
Journal: Proteomics
Volume: 13
Issue: 6
Pages: 982-91
Epub Date: 2013/01/22
Date: Mar
Short Title: Electron transfer dissociation (ETD): the mass spectrometric breakthrough essential for O-GlcNAc protein site assignments-a study of the O-GlcNAcylated protein host cell factor C1
Alternate Journal: Proteomics
ISSN: 1615-9853 (Print)
1615-9853
DOI: 10.1002/pmic.201200332
PMCID: PMC3988289
NIHMSID: NIHMS546749
Accession Number: 23335398
Keywords: Amino Acid Sequence
Animals
Chromatography, High Pressure Liquid
Glycopeptides/chemistry/isolation & purification
Glycosylation
HEK293 Cells
Host Cell Factor C1/*chemistry/isolation & purification/metabolism
Humans
Mice
Molecular Sequence Data
Peptide Fragments/chemistry/isolation & purification
*Protein Processing, Post-Translational
Spectrometry, Mass, Electrospray Ionization/methods
Tandem Mass Spectrometry/methods
Abstract: The development of electron-based, unimolecular dissociation MS, i.e. electron capture and electron transfer dissociation (ECD and ETD, respectively), has greatly increased the speed and reliability of labile PTM site assignment. The field of intracellular O-GlcNAc (O-linked N-acetylglucosamine) signaling has especially advanced with the advent of ETD MS. Only within the last five years have proteomic-scale experiments utilizing ETD allowed the assignment of hundreds of O-GlcNAc sites within cells and subcellular structures. Our ability to identify and unambiguously assign the site of O-GlcNAc modifications using ETD is rapidly increasing our understanding of this regulatory glycosylation and its potential interaction with other PTMs. Here, we discuss the advantages of using ETD, complimented with collisional-activation MS, in a study of the extensively O-GlcNAcylated protein Host Cell Factor C1 (HCF-1). HCF-1 is a transcriptional coregulator that forms a stable complex with O-GlcNAc transferase and controls cell cycle progression. ETD, along with higher energy collisional dissociation (HCD) MS, was employed to assign the PTMs of the HCF-1 protein isolated from HEK293T cells. These include 19 sites of O-GlcNAcylation, two sites of phosphorylation, and two sites bearing dimethylarginine, and showcase the residue-specific, PTM complexity of this regulator of cell proliferation.
Notes: 1615-9861
Myers, Samuel A
Daou, Salima
Affar, El Bachir
Burlingame, Al
Howard Hughes Medical Institute/United States
P41 GM103481/GM/NIGMS NIH HHS/United States
8P41GM103481/GM/NIGMS NIH HHS/United States
MOP115132/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proteomics. 2013 Mar;13(6):982-91. doi: 10.1002/pmic.201200332.
Author Address: Department of Pharmaceutical Chemistry, UCSF, San Francisco, CA, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1755
Author: Nabokina, S. M., Subramanian, V. S. and Said, H. M.
Year: 2016
Title: The human colonic thiamine pyrophosphate transporter (hTPPT) is a glycoprotein and N-linked glycosylation is important for its function
Journal: Biochim Biophys Acta
Volume: 1858
Issue: 4
Pages: 866-71
Epub Date: 2016/02/02
Date: Apr
Short Title: The human colonic thiamine pyrophosphate transporter (hTPPT) is a glycoprotein and N-linked glycosylation is important for its function
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbamem.2016.01.028
PMCID: PMC4779673
NIHMSID: NIHMS756158
Accession Number: 26828122
Keywords: Cell Membrane/chemistry/*metabolism
Colon/*chemistry/metabolism
Epithelial Cells/metabolism
Glycoproteins/chemistry/metabolism
Glycosylation
Humans
Membrane Transport Proteins/chemistry/genetics/*metabolism
Mutagenesis, Site-Directed
*Protein Structure, Secondary
Mutation
Thiamine pyrophosphate
Transport
Uptake
by the authors.
Abstract: The recently identified human thiamine pyrophosphate transporter (hTPPT; product of the SLC44A4 gene) is responsible for absorption of the microbiota-generated TPP in the large intestine. The hTPPT is highly expressed in the colon, but not in other regions of the intestinal tract and is localized exclusively at the apical membrane domain of epithelia. The hTPPT protein is predicted to have multiple TM domains with a number of putative N-glycosylation sites, but it is not known if the protein is actually glycosylated, and if so at which site, and their role in the functionality of the transporter. Using several approaches including inhibiting de novo N-glycosylation in human colonic epithelial NCM460 cells with tunicamycin as well as enzymatic de-glycosylation, we show that the hTPPT protein is, indeed, a glycoprotein. Glycosylation of hTPPT was shown, by mean of site-directed mutagenesis, to occur at Asn(69), Asn(155), Asn(197), Asn(393), and Asn(416). However, only N-glycosylation at Asn(69), Asn(155), and Asn(393) appeared to be important for transporter functionality possibly through an effect on protein conformation and/or interaction with its ligand (but not through changes in expression at the cell membrane as determined by live cell confocal imaging). Results of this study showed, for the first time, that the hTPPT is glycosylated and that N-linked glycosylation occurs at multiple sites with some of them being important for function. The results also provide an indirect support for a membrane topology for hTPPT with 10 transmembrane domains as predicted by the TMHMM transmembrane helixes prediction program.
Notes: Nabokina, Svetlana M
Subramanian, Veedamali S
Said, Hamid M
R37 DK056061/DK/NIDDK NIH HHS/United States
R01 DK056061/DK/NIDDK NIH HHS/United States
AA018071/AA/NIAAA NIH HHS/United States
I01 BX001142/BX/BLRD VA/United States
R01 AA018071/AA/NIAAA NIH HHS/United States
DK-56061/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Biochim Biophys Acta. 2016 Apr;1858(4):866-71. doi: 10.1016/j.bbamem.2016.01.028. Epub 2016 Jan 30.
Author Address: Department of Medicine, University of California, Irvine, CA 92697, United States; Department of Physiology/Biophysics, University of California, Irvine, CA 92697, United States; Department of Veterans Affairs Medical Center, Long Beach, CA 90822, United States.
Department of Medicine, University of California, Irvine, CA 92697, United States; Department of Physiology/Biophysics, University of California, Irvine, CA 92697, United States; Department of Veterans Affairs Medical Center, Long Beach, CA 90822, United States. Electronic address: hmsaid@uci.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 972
Author: Nachon, F., Asojo, O. A., Borgstahl, G. E., Masson, P. and Lockridge, O.
Year: 2005
Title: Role of water in aging of human butyrylcholinesterase inhibited by echothiophate: the crystal structure suggests two alternative mechanisms of aging
Journal: Biochemistry
Volume: 44
Issue: 4
Pages: 1154-62
Epub Date: 2005/01/26
Date: Feb 1
Short Title: Role of water in aging of human butyrylcholinesterase inhibited by echothiophate: the crystal structure suggests two alternative mechanisms of aging
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi048238d
Accession Number: 15667209
Keywords: Animals
Binding Sites
Butyrylcholinesterase/*chemistry/*metabolism
Catalysis
Cholinesterase Inhibitors/*chemistry
Crystallization
Crystallography, X-Ray
Echothiophate Iodide/*chemistry
Glutamic Acid/chemistry
Glycerol/chemistry
Histidine/chemistry
Humans
Hydrolysis
Organophosphorus Compounds/chemistry
Organothiophosphates/chemistry
Peptide Fragments/chemistry/metabolism
Torpedo
Water/*chemistry
Abstract: Organophosphorus poisons (OP) bind covalently to the active-site serine of cholinesterases. The inhibited enzyme can usually be reactivated with powerful nucleophiles such as oximes. However, the covalently bound OP can undergo a suicide reaction (termed aging) yielding nonreactivatable enzyme. In human butyrylcholinesterase (hBChE), aging involves the residues His438 and Glu197 that are proximal to the active-site serine (Ser198). The mechanism of aging is known in detail for the nerve gases soman, sarin, and tabun as well as the pesticide metabolite isomalathion. Aging of soman- and sarin-inhibited acetylcholinesterase occurs by C-O bond cleavage, whereas that of tabun- and isomalathion-inhibited acetylcholinesterase occurs by P-N and P-S bond cleavage, respectively. In this work, the crystal structures of hBChE inhibited by the ophthalmic reagents echothiophate (nonaged and aged) and diisopropylfluorophosphate (aged) were solved and refined to 2.1, 2.25, and 2.2 A resolution, respectively. No appreciable shift in the position of the catalytic triad histidine was observed between the aged and nonaged conjugates of hBChE. This absence of shift contrasts with the aged and nonaged crystal structures of Torpedo californica acetylcholinesterase inhibited by the nerve agent VX. The nonaged hBChE structure shows one water molecule interacting with Glu197 and the catalytic triad histidine (His438). Interestingly, this water molecule is ideally positioned to promote aging by two mechanisms: breaking either a C-O bond or a P-O bond. Pesticides and certain stereoisomers of nerve agents are expected to undergo aging by breaking the P-O bond.
Notes: Nachon, Florian
Asojo, Oluwatoyin A
Borgstahl, Gloria E O
Masson, Patrick
Lockridge, Oksana
P30CA36727-19/CA/NCI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2005 Feb 1;44(4):1154-62. doi: 10.1021/bi048238d.
Author Address: The Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, Nebraska 68198-6805, USA. florian@nachon.net
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1558
Author: Nachon, F., Carletti, E., Ronco, C., Trovaslet, M., Nicolet, Y., Jean, L. and Renard, P. Y.
Year: 2013
Title: Crystal structures of human cholinesterases in complex with huprine W and tacrine: elements of specificity for anti-Alzheimer's drugs targeting acetyl- and butyryl-cholinesterase
Journal: Biochem J
Volume: 453
Issue: 3
Pages: 393-9
Epub Date: 2013/05/18
Date: Aug 1
Short Title: Crystal structures of human cholinesterases in complex with huprine W and tacrine: elements of specificity for anti-Alzheimer's drugs targeting acetyl- and butyryl-cholinesterase
Alternate Journal: The Biochemical journal
ISSN: 0264-6021
DOI: 10.1042/bj20130013
Accession Number: 23679855
Keywords: Acetylcholinesterase/chemistry/metabolism
Alzheimer Disease/*enzymology
Aminoquinolines/*chemistry/pharmacology
Butyrylcholinesterase/chemistry/metabolism
Cholinesterase Inhibitors/*chemistry/pharmacology
Cholinesterases/*chemistry/*metabolism/pharmacology
Crystallography, X-Ray/*methods
Humans
Tacrine/*chemistry
Abstract: The multifunctional nature of Alzheimer's disease calls for MTDLs (multitarget-directed ligands) to act on different components of the pathology, like the cholinergic dysfunction and amyloid aggregation. Such MTDLs are usually on the basis of cholinesterase inhibitors (e.g. tacrine or huprine) coupled with another active molecule aimed at a different target. To aid in the design of these MTDLs, we report the crystal structures of hAChE (human acetylcholinesterase) in complex with FAS-2 (fasciculin 2) and a hydroxylated derivative of huprine (huprine W), and of hBChE (human butyrylcholinesterase) in complex with tacrine. Huprine W in hAChE and tacrine in hBChE reside in strikingly similar positions highlighting the conservation of key interactions, namely, π-π/cation-π interactions with Trp86 (Trp82), and hydrogen bonding with the main chain carbonyl of the catalytic histidine residue. Huprine W forms additional interactions with hAChE, which explains its superior affinity: the isoquinoline moiety is associated with a group of aromatic residues (Tyr337, Phe338 and Phe295 not present in hBChE) in addition to Trp86; the hydroxyl group is hydrogen bonded to both the catalytic serine residue and residues in the oxyanion hole; and the chlorine substituent is nested in a hydrophobic pocket interacting strongly with Trp439. There is no pocket in hBChE that is able to accommodate the chlorine substituent.
Notes: 1470-8728
Nachon, Florian
Carletti, Eugénie
Ronco, Cyril
Trovaslet, Marie
Nicolet, Yvain
Jean, Ludovic
Renard, Pierre-Yves
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem J. 2013 Aug 1;453(3):393-9. doi: 10.1042/BJ20130013.
Author Address: Département de Toxicologie, Institut de Recherche Biomédicale des Armées, 24 Avenue des Maquis du Grésivaudan, BP87, 38702 La Tronche, France. florian@nachon.net
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1964
Author: Nagae, M., Kizuka, Y., Mihara, E., Kitago, Y., Hanashima, S., Ito, Y., Takagi, J., Taniguchi, N. and Yamaguchi, Y.
Year: 2018
Title: Structure and mechanism of cancer-associated N-acetylglucosaminyltransferase-V
Journal: Nat Commun
Volume: 9
Issue: 1
Pages: 3380
Epub Date: 2018/08/25
Date: Aug 23
Short Title: Structure and mechanism of cancer-associated N-acetylglucosaminyltransferase-V
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-018-05931-w
PMCID: PMC6107550
Accession Number: 30140003
Keywords: Antineoplastic Agents/pharmacology/therapeutic use
Biocatalysis
Catalytic Domain/*physiology
Crystallography, X-Ray
Drug Design
Enzyme Assays
Glycoproteins/chemistry/metabolism
Humans
Molecular Docking Simulation
Mutagenesis, Site-Directed
N-Acetylglucosaminyltransferases/antagonists &
inhibitors/chemistry/genetics/*metabolism
Neoplasms/drug therapy/*pathology
Polysaccharides/chemistry/*metabolism
Substrate Specificity
Abstract: N-acetylglucosaminyltransferase-V (GnT-V) alters the structure of specific N-glycans by modifying α1-6-linked mannose with a β1-6-linked N-acetylglucosamine branch. β1-6 branch formation on cell surface receptors accelerates cancer metastasis, making GnT-V a promising target for drug development. However, the molecular basis of GnT-V's catalytic mechanism and substrate specificity are not fully understood. Here, we report crystal structures of human GnT-V luminal domain with a substrate analog. GnT-V luminal domain is composed of a GT-B fold and two accessary domains. Interestingly, two aromatic rings sandwich the α1-6 branch of the acceptor N-glycan and restrain the global conformation, partly explaining the fine branch specificity of GnT-V. In addition, interaction of the substrate N-glycoprotein with GnT-V likely contributes to protein-selective and site-specific glycan modification. In summary, the acceptor-GnT-V complex structure suggests a catalytic mechanism, explains the previously observed inhibition of GnT-V by branching enzyme GnT-III, and provides a basis for the rational design of drugs targeting N-glycan branching.
Notes: 2041-1723
Nagae, Masamichi
Kizuka, Yasuhiko
Mihara, Emiko
Kitago, Yu
Hanashima, Shinya
Ito, Yukishige
Orcid: 0000-0001-6251-7249
Takagi, Junichi
Taniguchi, Naoyuki
Yamaguchi, Yoshiki
15K18496/Ministry of Education, Culture, Sports, Science, and Technology (MEXT)/International
26110724/Ministry of Education, Culture, Sports, Science, and Technology (MEXT)/International
17K07303/Ministry of Education, Culture, Sports, Science, and Technology (MEXT)/International
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2018 Aug 23;9(1):3380. doi: 10.1038/s41467-018-05931-w.
Author Address: Structural Glycobiology Team, Glycobiology Research Group, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan. mnagae@mol.f.u-tokyo.ac.jp.
Graduate School of Pharmaceutical Sciences, The University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo, 113-0033, Japan. mnagae@mol.f.u-tokyo.ac.jp.
Disease Glycomics Team, Systems Glycobiology Research Group, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
Center for Highly Advanced Integration of Nano and Life Sciences (G-CHAIN), Gifu University, 1-1 Yanagido, Gifu-City, Gifu, 501-1193, Japan.
Institute for Protein Research, Osaka University, Suita, Osaka, 565-0871, Japan.
Department of Chemistry, Osaka University, Machikaneyama, Toyonaka, Osaka, 560-0043, Japan.
Synthetic Cellular Chemistry Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
Structural Glycobiology Team, Glycobiology Research Group, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1674
Author: Nagae, M., Morita-Matsumoto, K., Kato, M., Kaneko, M. K., Kato, Y. and Yamaguchi, Y.
Year: 2014
Title: A platform of C-type lectin-like receptor CLEC-2 for binding O-glycosylated podoplanin and nonglycosylated rhodocytin
Journal: Structure
Volume: 22
Issue: 12
Pages: 1711-1721
Epub Date: 2014/12/03
Date: Dec 2
Short Title: A platform of C-type lectin-like receptor CLEC-2 for binding O-glycosylated podoplanin and nonglycosylated rhodocytin
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126
DOI: 10.1016/j.str.2014.09.009
Accession Number: 25458834
Keywords: Binding Sites
Crystallography, X-Ray
Glycosylation
Lectins, C-Type/*metabolism
Membrane Glycoproteins/*metabolism
*Models, Molecular
Platelet Aggregation
Protein Binding
Viper Venoms/*metabolism
Abstract: Podoplanin is a transmembrane O-glycoprotein that binds to C-type lectin-like receptor 2 (CLEC-2). The O-glycan-dependent interaction seems to play crucial roles in various biological processes, such as platelet aggregation. Rhodocytin, a snake venom, also binds to CLEC-2 and aggregates platelets in a glycan-independent manner. To elucidate the structural basis of the glycan-dependent and independent interactions, we performed comparative crystallographic studies of podoplanin and rhodocytin in complex with CLEC-2. Both podoplanin and rhodocytin bind to the noncanonical "side" face of CLEC-2. There is a common interaction mode between consecutive acidic residues on the ligands and the same arginine residues on CLEC-2. Other interactions are ligand-specific. Carboxyl groups from the sialic acid residue on podoplanin and from the C terminus of the rhodocytin α subunit interact differently at this "second" binding site on CLEC-2. The unique and versatile binding modes open a way to understand the functional consequences of CLEC-2-ligand interactions.
Notes: 1878-4186
Nagae, Masamichi
Morita-Matsumoto, Kana
Kato, Masaki
Kaneko, Mika Kato
Kato, Yukinari
Yamaguchi, Yoshiki
Journal Article
Research Support, Non-U.S. Gov't
United States
Structure. 2014 Dec 2;22(12):1711-1721. doi: 10.1016/j.str.2014.09.009. Epub 2014 Nov 6.
Author Address: Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center, RIKEN Global Research Cluster, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Department of Regional Innovation, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai 980-8575, Japan.
Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center, RIKEN Global Research Cluster, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan. Electronic address: yyoshiki@riken.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1472
Author: Nagae, M., Re, S., Mihara, E., Nogi, T., Sugita, Y. and Takagi, J.
Year: 2012
Title: Crystal structure of α5β1 integrin ectodomain: atomic details of the fibronectin receptor
Journal: J Cell Biol
Volume: 197
Issue: 1
Pages: 131-40
Epub Date: 2012/03/28
Date: Apr 2
Short Title: Crystal structure of α5β1 integrin ectodomain: atomic details of the fibronectin receptor
Alternate Journal: The Journal of cell biology
ISSN: 0021-9525 (Print)
0021-9525
DOI: 10.1083/jcb.201111077
PMCID: PMC3317794
Accession Number: 22451694
Keywords: Crystallography, X-Ray
Humans
Integrin alpha5beta1/*chemistry
Models, Molecular
Molecular Dynamics Simulation
Protein Structure, Tertiary
Recombinant Proteins/chemistry
Abstract: Integrin α5β1 is a major cellular receptor for the extracellular matrix protein fibronectin and plays a fundamental role during mammalian development. A crystal structure of the α5β1 integrin headpiece fragment bound by an allosteric inhibitory antibody was determined at a 2.9-Å resolution both in the absence and presence of a ligand peptide containing the Arg-Gly-Asp (RGD) sequence. The antibody-bound β1 chain accommodated the RGD ligand with very limited structural changes, which may represent the initial step of cell adhesion mediated by nonactivated integrins. Furthermore, a molecular dynamics simulation pointed to an important role for Ca(2+) in the conformational coupling between the ligand-binding site and the rest of the molecule. The RGD-binding pocket is situated at the center of a trenchlike exposed surface on the top face of α5β1 devoid of glycosylation sites. The structure also enabled the precise prediction of the acceptor residue for the auxiliary synergy site of fibronectin on the α5 subunit, which was experimentally confirmed by mutagenesis and kinetic binding assays.
Notes: 1540-8140
Nagae, Masamichi
Re, Suyong
Mihara, Emiko
Nogi, Terukazu
Sugita, Yuji
Takagi, Junichi
Journal Article
Research Support, Non-U.S. Gov't
J Cell Biol. 2012 Apr 2;197(1):131-40. doi: 10.1083/jcb.201111077. Epub 2012 Mar 26.
Author Address: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1661
Author: Nagel, A. K. and Ball, L. E.
Year: 2014
Title: O-GlcNAc modification of the runt-related transcription factor 2 (Runx2) links osteogenesis and nutrient metabolism in bone marrow mesenchymal stem cells
Journal: Mol Cell Proteomics
Volume: 13
Issue: 12
Pages: 3381-95
Epub Date: 2014/09/05
Date: Dec
Short Title: O-GlcNAc modification of the runt-related transcription factor 2 (Runx2) links osteogenesis and nutrient metabolism in bone marrow mesenchymal stem cells
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M114.040691
PMCID: PMC4256491
Accession Number: 25187572
Keywords: Acetylglucosamine/*metabolism
Adaptor Proteins, Signal Transducing/genetics/metabolism
Animals
Arginine/metabolism
Bone Marrow Cells/cytology/drug effects/*metabolism
Bone Morphogenetic Protein 2/pharmacology
Bone Morphogenetic Protein 7/pharmacology
Cell Differentiation/drug effects
Cell Line
Core Binding Factor Alpha 1 Subunit/genetics/metabolism
Enzyme Inhibitors/pharmacology
HEK293 Cells
Humans
Membrane Proteins/genetics/metabolism
Mesenchymal Stem Cells/cytology/drug effects/*metabolism
Methylation
Mice
Mice, Inbred C57BL
N-Acetylglucosaminyltransferases/genetics/metabolism
Osteoblasts/cytology/drug effects/*metabolism
Osteogenesis/drug effects/*genetics
*Protein Processing, Post-Translational
Transcriptional Activation/drug effects
Abstract: Runx2 is the master switch controlling osteoblast differentiation and formation of the mineralized skeleton. The post-translational modification of Runx2 by phosphorylation, ubiquitinylation, and acetylation modulates its activity, stability, and interactions with transcriptional co-regulators and chromatin remodeling proteins downstream of osteogenic signals. Characterization of Runx2 by electron transfer dissociation tandem mass spectrometry revealed sites of O-linked N-acetylglucosamine (O-GlcNAc) modification, a nutrient-responsive post-translational modification that modulates the action of numerous transcriptional effectors. O-GlcNAc modification occurs in close proximity to phosphorylated residues and novel sites of arginine methylation within regions known to regulate Runx2 transactivation. An interaction between Runx2 and the O-GlcNAcylated, O-GlcNAc transferase enzyme was also detected. Pharmacological inhibition of O-GlcNAcase (OGA), the enzyme responsible for the removal of O-GlcNAc from Ser/Thr residues, enhanced basal (39.9%) and BMP2/7-induced (43.3%) Runx2 transcriptional activity in MC3T3-E1 pre-osteoblasts. In bone marrow-derived mesenchymal stem cells differentiated for 6 days in osteogenic media, inhibition of OGA resulted in elevated expression (24.3%) and activity (65.8%) of alkaline phosphatase (ALP) an early marker of bone formation and a transcriptional target of Runx2. Osteogenic differentiation of bone marrow-derived mesenchymal stem cells in the presence of BMP2/7 for 8 days culminated in decreased OGA activity (39.0%) and an increase in the abundance of O-GlcNAcylated Runx2, as compared with unstimulated cells. Furthermore, BMP2/7-induced ALP activity was enhanced by 35.6% in bone marrow-derived mesenchymal stem cells differentiated in the presence of the OGA inhibitor, demonstrating that direct or BMP2/7-induced inhibition of OGA is associated with increased ALP activity. Altogether, these findings link O-GlcNAc cycling to the Runx2-dependent regulation of the early ALP marker under osteoblast differentiation conditions.
Notes: 1535-9484
Nagel, Alexis K
Ball, Lauren E
T32 DE017551/DE/NIDCR NIH HHS/United States
T32 DE017551 05/DE/NIDCR NIH HHS/United States
R01 DE020925/DE/NIDCR NIH HHS/United States
S10D010731-01/PHS HHS/United States
S10 OD010731/OD/NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Mol Cell Proteomics. 2014 Dec;13(12):3381-95. doi: 10.1074/mcp.M114.040691. Epub 2014 Sep 3.
Author Address: From the ‡Department of Oral Health Sciences; Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, South Carolina, 29425.
From the ‡Department of Oral Health Sciences; Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, South Carolina, 29425 ballle@musc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 524
Author: Nakada, H., Inoue, M., Numata, Y., Tanaka, N., Funakoshi, I., Fukui, S., Mellors, A. and Yamashina, I.
Year: 1993
Title: Epitopic structure of Tn glycophorin A for an anti-Tn antibody (MLS 128)
Journal: Proc Natl Acad Sci U S A
Volume: 90
Issue: 6
Pages: 2495-9
Epub Date: 1993/03/15
Date: Mar 15
Short Title: Epitopic structure of Tn glycophorin A for an anti-Tn antibody (MLS 128)
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.90.6.2495
PMCID: PMC46114
Accession Number: 7681597
Keywords: Amino Acid Sequence
Animals
Antibodies, Monoclonal
Antigens, Tumor-Associated, Carbohydrate/chemistry/*immunology
Chromatography, Gel
Chromatography, High Pressure Liquid
Epitopes/*analysis/chemistry
Glycopeptides/chemistry/immunology/isolation & purification
Glycophorins/*chemistry/immunology
Humans
Mannheimia haemolytica/enzymology
Metalloendopeptidases
Molecular Sequence Data
Mucins/physiology
Sheep
Abstract: Glycophorin A was digested with glycoprotease (Pasteurella haemolytica) and the digest was fractionated by a combination of high-pressure column chromatographies to produce the glycopeptides GPA-1 to GPA-6. Sequence analysis of the glycopeptides revealed that two serine residues (Ser-14 and Ser-15) are not glycosylated, Thr-17 and Ser-19 being glycosylated instead, in disagreement with the accepted structure. The glycopeptides thus obtained were treated with sialidase and beta-galactosidase. The Tn antigenicity, as assayed by the binding to a monoclonal anti-Tn antibody (MLS 128), was found exclusively in the glycopeptides including three (cluster I) or four (cluster II) consecutive residues of GalNAc-Ser/Thr, whereas the glycopeptide (GPA-2) containing two nonconsecutive GalNAc-Ser/Thr residues had practically no Tn antigenicity. The immunoreactivities of GPA-1 and GPA-3, containing both clusters I and II, and GPA-4, containing cluster II, were 63% (calcd. 67%), 81% (calcd. 86%), and 50% (calcd. 50%), respectively, of the immunoreactivity of GPA-5 or GPA-6, containing cluster I (the average being taken as the basis), based on the reactivity per GalNAc residue. These results indicate that clusters I and II react with the antibody to the same extent. The structure consisting of three consecutive glycosylated Ser/Thr residues may be essential for Tn antigenicity in the light of previous results for ovine submaxillary mucin.
Notes: 1091-6490
Nakada, H
Inoue, M
Numata, Y
Tanaka, N
Funakoshi, I
Fukui, S
Mellors, A
Yamashina, I
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 1993 Mar 15;90(6):2495-9. doi: 10.1073/pnas.90.6.2495.
Author Address: Department of Biotechnology, Faculty of Engineering, Kyoto Sangyo University, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 519
Author: Nakagawa, H., Takahashi, N., Fujikawa, K., Kawamura, Y., Iino, M., Takeya, H., Ogawa, H. and Suzuki, K.
Year: 1995
Title: Identification of the oligosaccharide structures of human coagulation factor X activation peptide at each glycosylation site
Journal: Glycoconj J
Volume: 12
Issue: 2
Pages: 173-81
Epub Date: 1995/04/01
Date: Apr
Short Title: Identification of the oligosaccharide structures of human coagulation factor X activation peptide at each glycosylation site
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/bf00731362
Accession Number: 7620335
Keywords: Asparagine/chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Endopeptidases/metabolism
Factor X/*chemistry
Glycosylation
Humans
Metalloendopeptidases
Molecular Sequence Data
N-Acetylneuraminic Acid
Oligosaccharides/analysis/*chemistry
Peptides/chemistry
Sialic Acids/chemistry
Abstract: Human blood coagulation factor X has two N-linked oligosaccharides at Asn39 and Asn49 residues and two O-linked oligosaccharides at Thr17 and Thr29 residues in the region of the factor X activation peptide (XAP) which is cleaved off during its activation by factor IXa. We determined the structure of oligosaccharides in the XAP region of human factor X. Four glycopeptides each containing a glycosylation site were isolated by digestion of XAP with endoproteinase Asp-N followed by reversed-phase HPLC. N-linked oligosaccharides released from the glycopeptides by glycoamidase A digestion were derivatized with 2-aminopyridine. Pyridylamino(PA)-oligosaccharides were separated by HPLC into neutral and sialyl oligosaccharides using an anion-exchange column. Structures of oligosaccharides and their contents at each glycosylation site were determined by a two-dimensional sugar mapping method. The contents of the neutral oligosaccharides at Asn39 and Asn49 residues were 32.5% and 30.0%, respectively. Six neutral and twelve monosialyl oligosaccharides isolated from both N-linked glycosylation sites showed similar elution profiles composed of bi-, tri- and tetra-antennary complex type oligosaccharides. The predominant component in neutral oligosaccharides was biantennary without a fucose residue. Two major monosialyl oligosaccharides were also biantennary without fucose and with a Neu5Ac alpha 2-->6 residue. In addition, the structures of O-linked oligosaccharides at Thr17 and Thr29 residues were suggested to be disialylated Gal beta 3GalNAc sequences by their component analyses.
Notes: Nakagawa, H
Takahashi, N
Fujikawa, K
Kawamura, Y
Iino, M
Takeya, H
Ogawa, H
Suzuki, K
Journal Article
United States
Glycoconj J. 1995 Apr;12(2):173-81. doi: 10.1007/BF00731362.
Author Address: GlycoLab, Nakano Vinegar Co. Ltd, Aichi, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 605
Author: Nakagawa, H., Zheng, M., Hakomori, S., Tsukamoto, Y., Kawamura, Y. and Takahashi, N.
Year: 1996
Title: Detailed oligosaccharide structures of human integrin alpha 5 beta 1 analyzed by a three-dimensional mapping technique
Journal: Eur J Biochem
Volume: 237
Issue: 1
Pages: 76-85
Epub Date: 1996/04/01
Date: Apr 1
Short Title: Detailed oligosaccharide structures of human integrin alpha 5 beta 1 analyzed by a three-dimensional mapping technique
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1996.0076n.x
Accession Number: 8620897
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, DEAE-Cellulose
Chromatography, High Pressure Liquid
Humans
Hydrolysis
Molecular Sequence Data
Oligosaccharides/*chemistry
Receptors, Fibronectin/*chemistry
Abstract: Structures of N-linked oligosaccharides obtained from human integrin alpha 5 beta 1 are described. Integrin alpha 5 beta 1 (4.5 mg) was purified from human placenta and digested using trypsin and chymotrypsin. N-linked oligosaccharides were released from the glycopeptides by digestion with glycoamidase A (from almond). The reducing ends of the oligosaccharides were derivatized with 2-aminopyridine. The pyridylamino-oligosaccharides were separated and these structures were identified by a three-dimensional HPLC mapping technique on three kinds of HPLC columns [Takahashi, N., Nakagawa, H., Fujikawa, K., Kawamura, Y. & Tomiya, N. (1995) Anal. Biochem. 226, 139-146]. Finally, 35 different oligosaccharide structures were identified, 10 of which were neutral, 6 mono-sialyl, 10 di-sialyl, 7 tri-sialyl and 2 tetra-sialyl. The molar ratio of neutral, mono-sialyl, di-sialyl, tri-sialyl and tetra-sialyl oligosaccharides was 20.8%, 24.8%, 27.7%, 18.1% and 8.6%, respectively. High-mannose-type oligosaccharides accounted for only 1.5% of the total. The remaining oligosaccharides were all complex type. The most predominant structure was the diantennary di-alpha-(2,3)-sialyl fucosyl. Major linking of sialic acid was alpha-(2,3)-linkage, and over 50% of all oligosaccharides were fucosylated at the N-acetylglucosamine residue of the reducing end.
Notes: Nakagawa, H
Zheng, M
Hakomori, S
Tsukamoto, Y
Kawamura, Y
Takahashi, N
CA 42505/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Eur J Biochem. 1996 Apr 1;237(1):76-85. doi: 10.1111/j.1432-1033.1996.0076n.x.
Author Address: GlycoLab, Nakano Central Research Institute, Nakano Vinegar Co. Ltd, Handa-City, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 992
Author: Nakagawa, T., Guichard, A., Castro, C. P., Xiao, Y., Rizen, M., Zhang, H. Z., Hu, D., Bang, A., Helms, J., Bier, E. and Derynck, R.
Year: 2005
Title: Characterization of a human rhomboid homolog, p100hRho/RHBDF1, which interacts with TGF-alpha family ligands
Journal: Dev Dyn
Volume: 233
Issue: 4
Pages: 1315-31
Epub Date: 2005/06/21
Date: Aug
Short Title: Characterization of a human rhomboid homolog, p100hRho/RHBDF1, which interacts with TGF-alpha family ligands
Alternate Journal: Developmental dynamics : an official publication of the American Association of Anatomists
ISSN: 1058-8388 (Print)
1058-8388
DOI: 10.1002/dvdy.20450
Accession Number: 15965977
Keywords: Amino Acid Sequence
Animals
Cell Membrane/chemistry/metabolism
Dimerization
Drosophila/physiology
ErbB Receptors/chemistry/genetics/*metabolism
Humans
Ligands
Membrane Proteins
Mice
Molecular Sequence Data
Multigene Family
Organ Specificity
Protein Isoforms/genetics/metabolism
Protein Structure, Tertiary
Transforming Growth Factor alpha/*metabolism
Abstract: The activity of the TGF-alpha-like ligand Spitz in Drosophila depends on Rhomboid, a seven-transmembrane spanning protein that resides in the Golgi and acts as a serine protease to cleave Spitz, thereby releasing the soluble ligand. Several rhomboids in Drosophila have been implicated in the processing of TGF-alpha-like ligands, and consequent EGF receptor activation. The larger number of TGF-alpha-like ligands in vertebrates raises the possibility that they too might be subject to regulation by rhomboid-like proteins. We present the cDNA cloning and polypeptide sequence of an atypically long human rhomboid, which, based on the absence of critical residues for serine protease activity, is not predicted to act as a serine protease. We examined its tissue distribution, in comparison with TGF-alpha and the TGF-alpha-related protein HB-EGF, and the EGF/TGF-alpha receptor, in mouse embryo. This rhomboid, named p100(hRho) or RHBDF1, is a seven-transmembrane protein with a long N-terminal cytoplasmic extension that comprises half of the polypeptide sequence, and is found in the endoplasmic reticulum and Golgi, but not on the cell surface. It is expressed as two forms with different lengths, forms dimers and interacts with TGF-alpha ligands through a luminal interaction with the EGF core ectodomain. Finally, we evaluated the function of p100(hRho)/RHBDF1 in Drosophila, demonstrating that the short, but not the full-length form has functional activity. The characterization of this protein extends our understanding of the rhomboid family of regulatory proteins.
Notes: Nakagawa, Takatoshi
Guichard, Annabel
Castro, Carolina Perez
Xiao, Yang
Rizen, Michael
Zhang, Hong-Zhong
Hu, Diane
Bang, Anne
Helms, Jill
Bier, Ethan
Derynck, Rik
KO2 DE00421/DE/NIDCR NIH HHS/United States
P60 DE13058/DE/NIDCR NIH HHS/United States
R01 CA54826/CA/NCI NIH HHS/United States
R29 DE12462/DE/NIDCR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Dev Dyn. 2005 Aug;233(4):1315-31. doi: 10.1002/dvdy.20450.
Author Address: Department of Cell and Tissue Biology, Program in Cell Biology, University of California at San Francisco, San Francisco, California 94143-0512, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 610
Author: Nakahara, Y., Miyata, T., Hamuro, T., Funatsu, A., Miyagi, M., Tsunasawa, S. and Kato, H.
Year: 1996
Title: Amino acid sequence and carbohydrate structure of a recombinant human tissue factor pathway inhibitor expressed in Chinese hamster ovary cells: one N-and two O-linked carbohydrate chains are located between Kunitz domains 2 and 3 and one N-linked carbohydrate chain is in Kunitz domain 2
Journal: Biochemistry
Volume: 35
Issue: 20
Pages: 6450-9
Epub Date: 1996/05/21
Date: May 21
Short Title: Amino acid sequence and carbohydrate structure of a recombinant human tissue factor pathway inhibitor expressed in Chinese hamster ovary cells: one N-and two O-linked carbohydrate chains are located between Kunitz domains 2 and 3 and one N-linked carbohydrate chain is in Kunitz domain 2
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi9524880
Accession Number: 8639592
Keywords: Amino Acid Sequence
Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/analysis
Cricetinae
Glycosylation
Humans
Lipoproteins/*chemistry/genetics
Mass Spectrometry
Molecular Sequence Data
Molecular Structure
Recombinant Proteins/chemistry/genetics
Sequence Homology, Amino Acid
Abstract: Human tissue factor pathway inhibitor is a protease inhibitor with three tandem Kunitz-type inhibitory domains. The recombinant protein (r-hTFPI) was produced using Chinese hamster ovary cells, and its polypeptide and carbohydrate chain structures were analyzed. The complete amino acid sequence, composed of 276 residues, was determined using a protein sequencer after protease digestion and it was identical to that predicted from the cDNA sequence. Among three potential N-glycosylation sites, both Asn117 and Asn167 were fully N-glycosylated but Asn228 was not. Thr175 was also fully O-glycosylated, but Ser174 was partially O-glycosylated. Carbohydrate composition and mass spectrometric analyses of the undecapeptide OG-11 (residues Leu 170approximately Leu180) showed that two O-linked carbohydrate chains consisted of a type-1 core structure (Gal-GalNAc-Ser/Thr) with 0-3 mol of N-acetylneuraminic acid(s). The N-linked carbohydrate chains were analyzed by two-dimensional carbohydrate mapping combined with sequential glycosidase digestion, after the reducing-ends of carbohydrate residues were tagged with 2-aminopyridine and non-reducing-end sialic acids were removed with sialidase. All the N-linked structures in r-hTFPI were complex-type carbohydrate chains with one fucose residue attached to the reducing-end GlcNAc and consisted of bi-, tri-, and tetraantennary carbohydrate chains in the ratio 1.9:1.3:1.0. Fucosylated tri- and tetraantennary carbohydrate chains with one or two N-acetyllactosaminyl repeats were also found (30% of carbohydrate chains determined). Thus, the region between Kunitz domains 2 and 3 encoded by exon 7 was highly glycosylated by two O-linked carbohydrate chains at Ser174 and Thr175 and one N-linked carbohydrate chain at Asn167. These results indicated that the region is occupied by a cluster of three bulky and acidic carbohydrate chains.
Notes: Nakahara, Y
Miyata, T
Hamuro, T
Funatsu, A
Miyagi, M
Tsunasawa, S
Kato, H
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1996 May 21;35(20):6450-9. doi: 10.1021/bi9524880.
Author Address: Chemo-Sero-Therapeutic Research Institute, Kumamoto, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1283
Author: Nakajima, M., Koga, T., Sakai, H., Yamanaka, H., Fujiwara, R. and Yokoi, T.
Year: 2010
Title: N-Glycosylation plays a role in protein folding of human UGT1A9
Journal: Biochem Pharmacol
Volume: 79
Issue: 8
Pages: 1165-72
Epub Date: 2009/12/03
Date: Apr 15
Short Title: N-Glycosylation plays a role in protein folding of human UGT1A9
Alternate Journal: Biochemical pharmacology
ISSN: 0006-2952
DOI: 10.1016/j.bcp.2009.11.020
Accession Number: 19951703
Keywords: Cells, Cultured
Electrophoresis, Polyacrylamide Gel
Enzyme Stability
Glucuronosyltransferase/*chemistry/metabolism
Glycosylation
Humans
Indolizines/pharmacology
*Protein Folding
Tunicamycin/pharmacology
Abstract: UDP-glucuronosyltransferases (UGTs) catalyze the glucuronidation of a variety of xeno/endobiotics. UGTs are type I membrane proteins of the endoplasmic reticulum (ER) with a glycosylated luminal domain. In the present study, we investigated the role of N-glycosylation in the function of human UGT1A9. Mutation analysis at the potential N-glycosylation sites at residues 71, 292, and 344 (from asparagine to glutamine) revealed that all of them were glycosylated, but the extent of glycosylation and/or size of the glycan differed. In comparison with the wild-type, these mutants showed decreased enzyme activities in parallel with the extent of the band shift in Western blot analysis. To evaluate the role of glycosylation in the enzyme activity, we produced unglycosylated UGT1A9 by treating HEK293 cells transiently transfected with expression plasmid with tunicamycin. The unglycosylated UGT1A9 was almost inactive, which was not an indirect effect of ER stress. To the contrary, the deglycosylated UGT1A9, which was produced by the treatment with Endo H under the non-denaturing condition, showed the same enzyme kinetics as the control. These results suggest that the glycosylation that occurs during translation is important for the folding of UGT1A9. The thermal stability analysis of the mutated and deglycosylated UGT1A9 proteins supported the findings. In conclusion, we found that the N-glycosylation has an important role in the folding of UGT1A9.
Notes: 1873-2968
Nakajima, Miki
Koga, Toshihisa
Sakai, Haruko
Yamanaka, Hiroyuki
Fujiwara, Ryoichi
Yokoi, Tsuyoshi
Journal Article
England
Biochem Pharmacol. 2010 Apr 15;79(8):1165-72. doi: 10.1016/j.bcp.2009.11.020. Epub 2009 Nov 29.
Author Address: Kanazawa University, Kakuma-machi, Japan. nmiki@kenroku.kanazawa-u.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1154
Author: Nakano, M., Nakagawa, T., Ito, T., Kitada, T., Hijioka, T., Kasahara, A., Tajiri, M., Wada, Y., Taniguchi, N. and Miyoshi, E.
Year: 2008
Title: Site-specific analysis of N-glycans on haptoglobin in sera of patients with pancreatic cancer: a novel approach for the development of tumor markers
Journal: Int J Cancer
Volume: 122
Issue: 10
Pages: 2301-9
Epub Date: 2008/01/25
Date: May 15
Short Title: Site-specific analysis of N-glycans on haptoglobin in sera of patients with pancreatic cancer: a novel approach for the development of tumor markers
Alternate Journal: International journal of cancer
ISSN: 0020-7136
DOI: 10.1002/ijc.23364
Accession Number: 18214858
Keywords: Adult
Aged
Biomarkers, Tumor/*blood
Chromatography, Liquid
Female
Fucose/chemistry/metabolism
Haptoglobins/*chemistry
Humans
Male
Middle Aged
Pancreatic Neoplasms/*blood/diagnosis
Pancreatitis, Chronic/blood/diagnosis
Polysaccharides/*analysis
Spectrometry, Mass, Electrospray Ionization
Abstract: It was found in our previous studies that the concentration of fucosylated haptoglobin had increased in the sera of patients with pancreatic cancer (PC) compared to those of other types of cancer and normal controls. Haptoglobin, an acute phase protein, has four potential N-glycosylation sites, although it remains unknown which site is responsible for the change in fucosylated N-glycans. In the present study, site-specific N-glycan structures of haptoglobin in sera obtained from patients with PC or chronic pancreatitis (CP) were analyzed using liquid chromatography-electrospray ionization mass spectrometry. Mass spectrometry analyses demonstrated that concentrations of total fucosylated di-, tri- and tetra-branched glycans of haptoglobin increased in the sera of PC patients. Tri-antennary N-glycans containing a Lewis X-type fucose markedly increased at the Asn211 site of haptoglobin N-glycans. While fucosylated N-glycans derived from serum haptoglobin of patients with CP slightly increased, di-fucosylated tetra-antennary N-glycans were observed only at this site in PC patients, and were absent in the haptoglobin of normal controls and individuals with CP. Thus, the present study provides evidence that site-specific analyses of N-glycans may be useful as novel tumor markers for PC.
Notes: 1097-0215
Nakano, Miyako
Nakagawa, Tsutomu
Ito, Toshifumi
Kitada, Takatoshi
Hijioka, Taizo
Kasahara, Akinori
Tajiri, Michiko
Wada, Yoshinao
Taniguchi, Naoyuki
Miyoshi, Eiji
Journal Article
Research Support, Non-U.S. Gov't
United States
Int J Cancer. 2008 May 15;122(10):2301-9. doi: 10.1002/ijc.23364.
Author Address: Department of Biochemistry, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 506
Author: Nakano, Y., Noda, K., Endo, T., Kobata, A. and Tomita, M.
Year: 1994
Title: Structural study on the glycosyl-phosphatidylinositol anchor and the asparagine-linked sugar chain of a soluble form of CD59 in human urine
Journal: Arch Biochem Biophys
Volume: 311
Issue: 1
Pages: 117-26
Epub Date: 1994/05/15
Date: May 15
Short Title: Structural study on the glycosyl-phosphatidylinositol anchor and the asparagine-linked sugar chain of a soluble form of CD59 in human urine
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1994.1216
Accession Number: 7514386
Keywords: Antigens, CD/*urine
Asparagine/*chemistry
CD59 Antigens
Carbohydrate Conformation
Carbohydrate Sequence
Carbohydrates/*chemistry
Chromatography, Affinity
Galactose/metabolism
Glycoside Hydrolases/metabolism
Glycosylphosphatidylinositols/*chemistry
Humans
Mannosidases/metabolism
Membrane Glycoproteins/*urine
Methylation
Molecular Sequence Data
Polysaccharides/chemistry
alpha-L-Fucosidase/metabolism
beta-Galactosidase/metabolism
beta-N-Acetylhexosaminidases/metabolism
Abstract: CD59 is an 18-kDa glycoprotein widely expressed on human cells. An important structural feature of CD59 is its attachment to the cell surface via a glycosyl-phosphatidylinositol (GPI) anchor. CD59, like many GPI-anchored proteins, has been found in urine, serum, and other body fluids. The structures of the GPI anchor and the asparagine-linked sugar chain of a soluble form of CD59 in urine, U-CD59, were determined. Purified U-CD59 released 1 mol of inositol per mole of protein by nitrous acid deamination, which cleaved between glucosamine and inositol present commonly in the GPI anchor. This indicates that a GPI anchor, which ended with inositol, is linked at the carboxy terminus of U-CD59. The peptide containing an asparagine-linked sugar chain and the peptide containing a glycan portion of the GPI anchor were isolated after trypsin digestion of U-CD59. The asparagine-linked sugar chains and the glycan portion of the GPI anchor were isolated from these peptides following hydrazinolysis or deamination and dephosphorylation, respectively. Their structures were analyzed by sequential exoglycosidase digestion and methylation analyses. The structures of the asparagine-linked sugar chains of U-CD59 were biantennary complex type, only 4.2% of which are monosialylated. The backbone structure of the GPI anchor was similar to that of Try-panosoma brucei variant surface glycoprotein, but showed significant variations in its side-chain moieties. This is the first detailed structural analysis of the human GPI anchor and the first detailed analysis of the carboxyl-terminal structure of the soluble-form GPI-anchored protein. The results indicate that the backbone structure of the GPI anchor is conserved from parasites to human and that at least a part of the soluble-form GPI-anchored protein has the structure produced by the action of glycan-phosphatidylinositol-specific phospholipase D.
Notes: Nakano, Y
Noda, K
Endo, T
Kobata, A
Tomita, M
Journal Article
United States
Arch Biochem Biophys. 1994 May 15;311(1):117-26. doi: 10.1006/abbi.1994.1216.
Author Address: Department of Physiological Chemistry, School of Pharmaceutical Sciences, Showa University, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 973
Author: Nanao, M. H., Green, T., Stern-Bach, Y., Heinemann, S. F. and Choe, S.
Year: 2005
Title: Structure of the kainate receptor subunit GluR6 agonist-binding domain complexed with domoic acid
Journal: Proc Natl Acad Sci U S A
Volume: 102
Issue: 5
Pages: 1708-13
Epub Date: 2005/01/29
Date: Feb 1
Short Title: Structure of the kainate receptor subunit GluR6 agonist-binding domain complexed with domoic acid
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0409573102
PMCID: PMC547884
Accession Number: 15677325
Keywords: Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray
Kainic Acid/*analogs & derivatives/*pharmacokinetics
Models, Molecular
Molecular Sequence Data
Neurotoxins/pharmacokinetics
Protein Structure, Secondary
Protein Subunits/chemistry/metabolism
Rats
Receptors, AMPA/chemistry/metabolism
Receptors, Kainic Acid/*chemistry/*metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: We report the crystal structure of the glycosylated ligand-binding (S1S2) domain of the kainate receptor subunit GluR6, in complex with the agonist domoate. The structure shows the expected overall homology with AMPA and NMDA receptor subunit structures but reveals an unexpected binding mode for the side chain of domoate, in which contact is made to the larger lobe only (lobe I). In common with the AMPA receptor subunit GluR2, the GluR6 S1S2 domain associates as a dimer, with many of the interdimer contacts being conserved. Subtle differences in these contacts provide a structural explanation for why GluR2 L483Y and GluR3 L507Y are nondesensitizing, but GluR6, which has a tyrosine at that site, is not. The structure incorporates native glycosylation, which has not previously been described for ionotropic glutamate receptors. The position of the sugars near the subunit interface rules out their direct involvement in subunit association but leaves open the possibility of indirect modulation. Finally, we observed several tetrameric assemblies that satisfy topological constraints with respect to connection to the receptor pore, and which are therefore candidates for the native quaternary structure.
Notes: 1091-6490
Nanao, Max H
Green, Tim
Stern-Bach, Yael
Heinemann, Stephen F
Choe, Senyon
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2005 Feb 1;102(5):1708-13. doi: 10.1073/pnas.0409573102. Epub 2005 Jan 26.
Author Address: Structural Biology Laboratory and Molecular Neurobiology Laboratory, The Salk Institute, 10010 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1024
Author: Nandi, A., Sprung, R., Barma, D. K., Zhao, Y., Kim, S. C., Falck, J. R. and Zhao, Y.
Year: 2006
Title: Global identification of O-GlcNAc-modified proteins
Journal: Anal Chem
Volume: 78
Issue: 2
Pages: 452-8
Epub Date: 2006/01/18
Date: Jan 15
Short Title: Global identification of O-GlcNAc-modified proteins
Alternate Journal: Analytical chemistry
ISSN: 0003-2700 (Print)
0003-2700
DOI: 10.1021/ac051207j
Accession Number: 16408927
Keywords: Acetylglucosamine/chemistry/*metabolism
Chromatography, High Pressure Liquid
HeLa Cells
Humans
N-Acetylglucosaminyltransferases/metabolism
*Protein Processing, Post-Translational
Proteins/chemistry/*metabolism
Proteomics/*methods
Tandem Mass Spectrometry
Abstract: The O-linked N-acetylglucosamine (O-GlcNAc) modification of serine/threonine residues is an abundant posttranslational modification present in cytosolic and nuclear proteins. The functions and subproteome of O-GlcNAc modification remain largely undefined. Here we report the application of the tagging-via-substrate (TAS) approach for global identification of O-GlcNAc-modified proteins. The TAS method utilizes an O-GlcNAc azide analogue for metabolic labeling of O-GlcNAc-modified proteins, which can be chemoselectively conjugated for detection and enrichment of the proteins for proteomics studies. Our study led to the identification of 199 putative O-GlcNAc-modified proteins from HeLa cells, among which 23 were confirmed using reciprocal immunoprecipitation. Functional classification shows that proteins with diverse functions are modified by O-GlcNAc, implying that O-GlcNAc might be involved in the regulation of multiple cellular pathways.
Notes: Nandi, Animesh
Sprung, Robert
Barma, Deb K
Zhao, Yingxin
Kim, Sung Chan
Falck, John R
Zhao, Yingming
CA 107943/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Anal Chem. 2006 Jan 15;78(2):452-8. doi: 10.1021/ac051207j.
Author Address: Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9038, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 418
Author: Neame, P. J., Christner, J. E. and Baker, J. R.
Year: 1987
Title: Cartilage proteoglycan aggregates. The link protein and proteoglycan amino-terminal globular domains have similar structures
Journal: J Biol Chem
Volume: 262
Issue: 36
Pages: 17768-78
Epub Date: 1987/12/25
Date: Dec 25
Short Title: Cartilage proteoglycan aggregates. The link protein and proteoglycan amino-terminal globular domains have similar structures
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3693371
Keywords: Amino Acid Sequence
Amino Acids/analysis
Animals
Binding Sites
Cartilage/*analysis
Chickens
Hyaluronic Acid/metabolism
Molecular Sequence Data
Protein Conformation
Proteoglycans/*analysis
Rats
Tumor Cells, Cultured/analysis
Abstract: Cartilage proteoglycan aggregates contain two components (proteoglycan monomer and link protein) which interact with each other and with hyaluronic acid. Data from amino acid sequence analysis are presented that shows that a domain of the proteoglycan, the hyaluronic acid binding region, which interacts with link protein and hyaluronic acid is very similar to link protein in terms of its primary structure. However, the pattern of glycosylation in the hyaluronic acid binding region is different from that found in link protein. After removal of N-linked oligosaccharides, the tryptically prepared hyaluronic acid binding region from rat chondrosarcoma has a mass by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of 43 +/- 2 kDa. The COOH-terminal two-thirds of rat chondrosarcoma link protein, starting at residue 105, has 41.3% identity with a similar region in the hyaluronic acid binding region. We show that, in addition to the hyaluronic acid binding region, proteoglycan contains another region with similarity to the two repeating loop structures in the COOH-terminal two-thirds of link protein. This presumably corresponds to the second globular domain reported in rotary shadowing studies of cartilage proteoglycans. We have deduced the positions of all of the disulfide bonds in the hyaluronic acid binding region and find them to be in the same positions as would be expected from comparison of these sequences with link protein.
Notes: Neame, P J
Christner, J E
Baker, J R
AM 20614/AM/NIADDK NIH HHS/United States
AR 35322/AR/NIAMS NIH HHS/United States
DE 02670/DE/NIDCR NIH HHS/United States
etc.
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1987 Dec 25;262(36):17768-78.
Author Address: Department Medicine, University of Alabama, Birmingham 35294.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1603
Author: Neculai, D., Schwake, M., Ravichandran, M., Zunke, F., Collins, R. F., Peters, J., Neculai, M., Plumb, J., Loppnau, P., Pizarro, J. C., Seitova, A., Trimble, W. S., Saftig, P., Grinstein, S. and Dhe-Paganon, S.
Year: 2013
Title: Structure of LIMP-2 provides functional insights with implications for SR-BI and CD36
Journal: Nature
Volume: 504
Issue: 7478
Pages: 172-6
Epub Date: 2013/10/29
Date: Dec 5
Short Title: Structure of LIMP-2 provides functional insights with implications for SR-BI and CD36
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature12684
Accession Number: 24162852
Keywords: Animals
CD36 Antigens/*metabolism
CHO Cells
Cricetulus
HeLa Cells
Humans
Lysosome-Associated Membrane Glycoproteins/*chemistry/metabolism
*Models, Molecular
Protein Binding
Protein Structure, Tertiary
Abstract: Members of the CD36 superfamily of scavenger receptor proteins are important regulators of lipid metabolism and innate immunity. They recognize normal and modified lipoproteins, as well as pathogen-associated molecular patterns. The family consists of three members: SR-BI (which delivers cholesterol to the liver and steroidogenic organs and is a co-receptor for hepatitis C virus), LIMP-2/LGP85 (which mediates lysosomal delivery of β-glucocerebrosidase and serves as a receptor for enterovirus 71 and coxsackieviruses) and CD36 (a fatty-acid transporter and receptor for phagocytosis of effete cells and Plasmodium-infected erythrocytes). Notably, CD36 is also a receptor for modified lipoproteins and β-amyloid, and has been implicated in the pathogenesis of atherosclerosis and of Alzheimer's disease. Despite their prominent roles in health and disease, understanding the function and abnormalities of the CD36 family members has been hampered by the paucity of information about their structure. Here we determine the crystal structure of LIMP-2 and infer, by homology modelling, the structure of SR-BI and CD36. LIMP-2 shows a helical bundle where β-glucocerebrosidase binds, and where ligands are most likely to bind to SR-BI and CD36. Remarkably, the crystal structure also shows the existence of a large cavity that traverses the entire length of the molecule. Mutagenesis of SR-BI indicates that the cavity serves as a tunnel through which cholesterol(esters) are delivered from the bound lipoprotein to the outer leaflet of the plasma membrane. We provide evidence supporting a model whereby lipidic constituents of the ligands attached to the receptor surface are handed off to the membrane through the tunnel, accounting for the selective lipid transfer characteristic of SR-BI and CD36.
Notes: 1476-4687
Neculai, Dante
Schwake, Michael
Ravichandran, Mani
Zunke, Friederike
Collins, Richard F
Peters, Judith
Neculai, Mirela
Plumb, Jonathan
Loppnau, Peter
Pizarro, Juan Carlos
Seitova, Alma
Trimble, William S
Saftig, Paul
Grinstein, Sergio
Dhe-Paganon, Sirano
MOP-102474/Canadian Institutes of Health Research/Canada
MOP-126069/Canadian Institutes of Health Research/Canada
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2013 Dec 5;504(7478):172-6. doi: 10.1038/nature12684. Epub 2013 Oct 27.
Author Address: Cell Biology Program, The Hospital for Sick Children, Toronto M5G 1X8, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1522
Author: Nelson, C. A., Warren, J. T., Wang, M. W., Teitelbaum, S. L. and Fremont, D. H.
Year: 2012
Title: RANKL employs distinct binding modes to engage RANK and the osteoprotegerin decoy receptor
Journal: Structure
Volume: 20
Issue: 11
Pages: 1971-82
Epub Date: 2012/10/09
Date: Nov 7
Short Title: RANKL employs distinct binding modes to engage RANK and the osteoprotegerin decoy receptor
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2012.08.030
PMCID: PMC3607351
NIHMSID: NIHMS412064
Accession Number: 23039992
Keywords: Amino Acid Sequence
Binding Sites
Humans
Models, Molecular
Molecular Sequence Data
RANK Ligand/chemistry/*metabolism
Sequence Homology, Amino Acid
Abstract: Osteoprotegerin (OPG) and receptor activator of nuclear factor κB (RANK) are members of the tumor necrosis factor receptor (TNFR) superfamily that regulate osteoclast formation and function by competing for RANK ligand (RANKL). RANKL promotes osteoclast development through RANK activation, while OPG inhibits this process by sequestering RANKL. For comparison, we solved crystal structures of RANKL with RANK and RANKL with OPG. Complementary biochemical and functional studies reveal that the monomeric cytokine-binding region of OPG binds RANKL with ∼500-fold higher affinity than RANK and inhibits RANKL-stimulated osteoclastogenesis ∼150 times more effectively, in part because the binding cleft of RANKL makes unique contacts with OPG. Several side chains as well as the C-D and D-E loops of RANKL occupy different orientations when bound to OPG versus RANK. High affinity OPG binding requires a 90s loop Phe residue that is mutated in juvenile Paget's disease. These results suggest cytokine plasticity may help to fine-tune specific tumor necrosis factor (TNF)-family cytokine/receptor pair selectivity.
Notes: 1878-4186
Nelson, Christopher A
Warren, Julia T
Wang, Michael W-H
Teitelbaum, Steven L
Fremont, Daved H
AR032788/AR/NIAMS NIH HHS/United States
R37 AR046523/AR/NIAMS NIH HHS/United States
P41 RR007707/RR/NCRR NIH HHS/United States
F30 AG039896/AG/NIA NIH HHS/United States
R01 AR032788/AR/NIAMS NIH HHS/United States
P30 AR057235/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Structure. 2012 Nov 7;20(11):1971-82. doi: 10.1016/j.str.2012.08.030. Epub 2012 Oct 2.
Author Address: Department of Pathology and Immunology, Washington University School of Medicine, 660 S. Euclid Avenue, St. Louis, MO 63110-1093, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 625
Author: Nelson, D. A., Bryan, J., Wechsler, S., Clement, J. P. th and Aguilar-Bryan, L.
Year: 1996
Title: The high-affinity sulfonylurea receptor: distribution, glycosylation, purification, and immunoprecipitation of two forms from endocrine and neuroendocrine cell lines
Journal: Biochemistry
Volume: 35
Issue: 47
Pages: 14793-9
Epub Date: 1996/11/26
Date: Nov 26
Short Title: The high-affinity sulfonylurea receptor: distribution, glycosylation, purification, and immunoprecipitation of two forms from endocrine and neuroendocrine cell lines
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi960777y
Accession Number: 8942641
Keywords: *ATP-Binding Cassette Transporters
Animals
COS Cells
Cell Line
Chromatography, Affinity
Cloning, Molecular
DNA, Complementary
Endocrine Glands/cytology/*metabolism
Glycosylation
Lectins
Neurosecretory Systems/cytology/*metabolism
Potassium Channels/genetics/isolation & purification/*metabolism
*Potassium Channels, Inwardly Rectifying
Precipitin Tests
Receptors, Drug/genetics/isolation & purification/*metabolism
Solubility
Sulfonylurea Receptors
Abstract: The high-affinity sulfonylurea receptor, a novel member of the ATP-binding cassette superfamily, is one component of the ATP-sensitive K+ channel. The protein is critical for regulation of insulin secretion from pancreatic beta-cells, and mutations in the receptor have been linked to familial hyperinsulinemia, a disorder characterized by unregulated insulin release despite severe hypoglycemia. The sulfonylurea receptor is present in membranes from a number of endocrine and neuroendocrine cell lines, including HIT-T15, RINm5f, alpha TC-6, AtT-20, and GH3 cells. Two forms of the receptor are present in RINm5f and alpha TC-6 cells, with apparent SDS gel molecular masses of 140 and 150 kDa. The two forms have equally high affinity, KD approximately 3 nM, for an iodinated derivative of glyburide, an anti-diabetic sulfonylurea. The receptor is a glycoprotein; treatment of RINm5f or alpha TC-6 cells with tunicamycin reduces the 140 and 150 kDa species to a single approximately 137 kDa protein. The 140 and 150 kDa receptors bind differentially to concanavalin A and wheat germ agglutinin, and lectin-affinity chromatography is ideal for the initial stages of receptor purification. After lectin-affinity chromatography, the same methods can be applied for purifying the 150 kDa form as for the 140 kDa receptor. A transiently expressed receptor with a histidine-tagged carboxy-terminus was purified by Ni-agarose chromatography, and this variant was used to demonstrate that the 140 kDa polypeptide is full length. Anti-peptide antibodies directed against the amino-terminus of the receptor and antibodies against the nucleotide binding folds immunoprecipitate both receptor forms. The results indicate the 140 and 150 kDa receptors are differentially glycosylated forms of the same polypeptide chain.
Notes: Nelson, D A
Bryan, J
Wechsler, S
Clement, J P 4th
Aguilar-Bryan, L
DK41898/DK/NIDDK NIH HHS/United States
DK44311/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1996 Nov 26;35(47):14793-9. doi: 10.1021/bi960777y.
Author Address: Department of Biology, University of North Carolina at Charlotte 28223, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 789
Author: Nemeth, J. F., Hochgesang, G. P., Jr., Marnett, L. J. and Caprioli, R. M.
Year: 2001
Title: Characterization of the glycosylation sites in cyclooxygenase-2 using mass spectrometry
Journal: Biochemistry
Volume: 40
Issue: 10
Pages: 3109-16
Epub Date: 2001/03/22
Date: Mar 13
Short Title: Characterization of the glycosylation sites in cyclooxygenase-2 using mass spectrometry
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi002313c
Accession Number: 11258925
Keywords: Amino Acid Sequence
Animals
Asparagine/metabolism
Cyclooxygenase 2
Genetic Vectors/biosynthesis/chemical synthesis
Glycopeptides/analysis/metabolism
Glycosylation
Isoenzymes/biosynthesis/*chemistry/genetics/*metabolism
Mice
Molecular Sequence Data
Peptide Fragments/analysis/metabolism
Prostaglandin-Endoperoxide Synthases/biosynthesis/*chemistry/genetics/*metabolism
Spectrometry, Mass, Electrospray Ionization
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Trypsin/metabolism
Abstract: Cyclooxygenase is involved in the biosynthesis and function of prostaglandins. It is a glycoprotein located in the endoplasmic reticulum and in the nuclear envelope, and it has been found to have two isoforms termed COX-1 and COX-2. This paper reports on the glycosylation site analysis of recombinant COX-2 using matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) and nanoelectrospray (nanoESI) quadrupole-TOF (Q-TOF) MS. The nanoESI MS analysis of COX-2 revealed the presence of three glycoforms at average molecular masses of 71.4, 72.7, and 73.9 kDa. Each glycoform contained a number of peaks differing by 162 Da indicating heterogeneity and suggesting the presence of high-mannose sugars. The masses of the glycoforms indicate that oligosaccharides occupy two to four sites and a single N-acetylglucosamine (GlcNAc) residue occupied up to two sites. The MALDI MS analysis of a tryptic digest of the protein showed a number of potential glycopeptides. The peptides differed by 162 Da which further suggested high-mannose sugars. Nanoelectrospray MS/MS experiments confirmed glycosylation at the Asn 53 and Asn 130 sites and confirmed the presence of the peptides Asn 396-Arg 414 + GlcNAc and Thr 576-Arg 587 + GlcNAc containing Asn 580. It was not possible to conclusively determine whether the Asn 396 site was glycosylated via an MS/MS experiment, so the tryptic digest was deglycosylated to confirm the presence of the glycopeptides. Finally, a non-glycosylated tryptic peptide was observed containing the Asn 592.
Notes: Nemeth, J F
Hochgesang, G P Jr
Marnett, L J
Caprioli, R M
5R35 CA47479/CA/NCI NIH HHS/United States
GM 58008-02/GM/NIGMS NIH HHS/United States
T32 ES07028/ES/NIEHS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2001 Mar 13;40(10):3109-16. doi: 10.1021/bi002313c.
Author Address: Mass Spectrometry Research Center, and Department of Biochemistry, Vanderbilt University Medical Center, Nashville, Tennesee 37232-6400, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1961
Author: Netsirisawan, P., Chaiyawat, P., Chokchaichamnankit, D., Lirdprapamongkol, K., Srisomsap, C., Svasti, J. and Champattanachai, V.
Year: 2018
Title: Decreasing O-GlcNAcylation affects the malignant transformation of MCF-7 cells via Hsp27 expression and its O-GlcNAc modification
Journal: Oncol Rep
Volume: 40
Issue: 4
Pages: 2193-2205
Epub Date: 2018/08/15
Date: Oct
Short Title: Decreasing O-GlcNAcylation affects the malignant transformation of MCF-7 cells via Hsp27 expression and its O-GlcNAc modification
Alternate Journal: Oncology reports
ISSN: 1021-335x
DOI: 10.3892/or.2018.6617
Accession Number: 30106436
Keywords: Acetylglucosamine/metabolism
*Anoikis
Apoptosis
Biomarkers, Tumor
Breast Neoplasms/metabolism/*pathology
Cell Proliferation
Cell Transformation, Neoplastic/metabolism/*pathology
Female
Glycosylation
HSP27 Heat-Shock Proteins/*metabolism
Heat-Shock Proteins
Humans
Molecular Chaperones
N-Acetylglucosaminyltransferases/*metabolism
*Protein Processing, Post-Translational
Tumor Cells, Cultured
Abstract: O-GlcNAcylation is a dynamic posttranslational modification of nucleoplasmic proteins. Previously, we reported that the O-GlcNAcylation level was increased in primary breast and colorectal cancer tissues. However, its precise roles in cancer development and progression are still largely unexplored. The aim of the present study was to investigate the roles of O-GlcNAcylation in the malignant transformation of cancer cell lines. O-GlcNAcylation level was examined in six cancer cell lines including breast (MCF-7 and MDA-MB-231), colorectal (SW480 and SW620), and liver (SK-Hep1 and HepG2). We found that the levels of O-GlcNAcylation and O-GlcNAc transferase (OGT), an O-GlcNAc catalyzing enzyme, were obviously increased in all cancerous cells, except SK-Hep1, when compared to normal cells. Reducing O-GlcNAcylation using RNA interference against OGT showed a marked reduction in OGT and O-GlcNAcylation levels. Surprisingly, siOGT had no effect on cell growth under conventional monolayer cultures. However, it inhibited anchorage-independent growth in soft agar cultures of all cancer cells, except SK-Hep1. Under anoikis resistance conditions performed by spheroid cultures, siOGT treatment decreased viability only in MCF-7, SW480, and SW620 cells. Among them, OGT knockdown in MCF-7 cells revealed a high inhibitory effect on colony and spheroid cultures. Using two-dimensional gel electrophoresis and mass spectrometric analysis, heat shock protein 27 (Hsp27) was found to be the highest upregulated protein upon OGT knockdown. Immunoblots revealed that the Hsp27 protein level was increased but its O-GlcNAc modification level was decreased in siOGT-treated cells. These changes were associated with the inhibition of MCF-7 cell transformation. Notably, double knockdown of OGT and Hsp27 showed a reversal in the inhibitory effect on colony and spheroid cultures. Collectively, these results indicate that O-GlcNAcylation is required for anoikis resistance and anchorage-independent growth of MCF-7 cells. Blocking this glycosylation by OGT knockdown may regulate both Hsp27 protein expression and its O-GlcNAc modification levels. This alteration may play vital roles in malignant transformation.
Notes: 1791-2431
Netsirisawan, Pukkavadee
Chaiyawat, Parunya
Chokchaichamnankit, Daranee
Lirdprapamongkol, Kriengsak
Srisomsap, Chantragan
Svasti, Jisnuson
Champattanachai, Voraratt
Journal Article
Greece
Oncol Rep. 2018 Oct;40(4):2193-2205. doi: 10.3892/or.2018.6617. Epub 2018 Aug 1.
Author Address: Laboratory of Biochemistry, Chulabhorn Research Institute, Chulabhorn Graduate Institute, Bangkok 10210, Thailand.
Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok 10210, Thailand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1710
Author: Netsirisawan, P., Chokchaichamnankit, D., Srisomsap, C., Svasti, J. and Champattanachai, V.
Year: 2015
Title: Proteomic Analysis Reveals Aberrant O-GlcNAcylation of Extracellular Proteins from Breast Cancer Cell Secretion
Journal: Cancer Genomics Proteomics
Volume: 12
Issue: 4
Pages: 201-9
Epub Date: 2015/07/03
Date: Jul-Aug
Short Title: Proteomic Analysis Reveals Aberrant O-GlcNAcylation of Extracellular Proteins from Breast Cancer Cell Secretion
Alternate Journal: Cancer genomics & proteomics
ISSN: 1109-6535
Accession Number: 26136220
Keywords: Breast Neoplasms/enzymology/*metabolism/pathology
Cell Compartmentation
Cell Line, Tumor
Culture Media, Serum-Free
Electrophoresis, Gel, Two-Dimensional
Extracellular Space/*metabolism
Female
Glycosylation
Humans
Intracellular Space/metabolism
Mass Spectrometry
N-Acetylglucosaminyltransferases/metabolism
Neoplasm Proteins/*metabolism
Proteomics/*methods
Reproducibility of Results
beta-N-Acetylhexosaminidases/metabolism
Breast cancer
O-GlcNAcylation
extracellular proteins
heat-shock 70 kDa
transitional endoplasmic reticulum ATPase
Abstract: BACKGROUND: O-GlcNAcylation is a unique intracellular protein modification; however, few extracellular O-GlcNAc-modified proteins have been discovered. We have previously demonstrated that many cellular proteins were aberrant in O-GlcNAcylation in breast cancer tissues. In the present study, therefore, we investigated whether O-GlcNAc-modified proteins were abnormally secreted from breast cancer cells. MATERIALS AND METHODS: Intracellular and extracellular proteins were prepared from cell lysates of breast cancer cells (MCF-7 and MDA-MB-231) and normal breast cells (HMEC) and from their serum-free media (SFM), respectively. O-GlcNAcylation level was examined by immunoblotting. O-GlcNAc-Modified proteins were identified using two-dimensional gel electrophoresis and Liquid Chromatography-tandem Mass Spectrometry. RESULTS: O-GlcNAcylation level was significantly increased in the extracellular compartment of both types of cancer cells compared to normal cells. Interestingly, O-GlcNAc patterns differed between intracellular and extracellular proteins. Proteomic analysis revealed that many O-GlcNAc spots in MCF-7 secretions were abnormally increased in comparison to those in HMEC secretions. Among these, transitional endoplasmic reticulum ATPase (TER ATPase) and heat-shock 70 kDa (HSP70) were confirmed to be O-GlcNAc-modified. The levels of O-GlcNAc-HSP70 and O-GlcNAc-TER ATPase were higher in SFM from MCF-7 cells than in that from HMEC. CONCLUSION: O-GlcNAcomic study of the extracellular compartments reveals aberrant O-GlcNAc-secreted proteins, which may be of interest as potential biomarkers in breast cancer.
Notes: 1790-6245
Netsirisawan, Pukkavadee
Chokchaichamnankit, Daranee
Srisomsap, Chantragan
Svasti, Jisnuson
Champattanachai, Voraratt
Journal Article
Research Support, Non-U.S. Gov't
Greece
Cancer Genomics Proteomics. 2015 Jul-Aug;12(4):201-9.
Author Address: Applied Biological Sciences Program, Chulabhorn Graduate Institute, Laksi, Bangkok, Thailand.
Laboratory of Biochemistry, Chulabhorn Research Institute, Laksi, Bangkok, Thailand.
Applied Biological Sciences Program, Chulabhorn Graduate Institute, Laksi, Bangkok, Thailand Laboratory of Biochemistry, Chulabhorn Research Institute, Laksi, Bangkok, Thailand.
Applied Biological Sciences Program, Chulabhorn Graduate Institute, Laksi, Bangkok, Thailand Laboratory of Biochemistry, Chulabhorn Research Institute, Laksi, Bangkok, Thailand voraratt@cri.or.th.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 672
Author: Neumann, G. M., Marinaro, J. A. and Bach, L. A.
Year: 1998
Title: Identification of O-glycosylation sites and partial characterization of carbohydrate structure and disulfide linkages of human insulin-like growth factor binding protein 6
Journal: Biochemistry
Volume: 37
Issue: 18
Pages: 6572-85
Epub Date: 1998/06/13
Date: May 5
Short Title: Identification of O-glycosylation sites and partial characterization of carbohydrate structure and disulfide linkages of human insulin-like growth factor binding protein 6
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi972894e
Accession Number: 9572875
Keywords: Amino Acid Sequence
Animals
CHO Cells
COS Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cricetinae
Disulfides/*chemistry
Glycosylation
Humans
Insulin-Like Growth Factor Binding Protein 6/*chemistry/isolation & purification
Mass Spectrometry
Molecular Sequence Data
Peptide Mapping
Abstract: The actions of insulin-like growth factors (IGFs) are modulated by a family of high-affinity binding proteins (IGFBPs), including IGFBP-6, which preferentially binds IGF-II and is O-glycosylated. Glycosylated and nonglycosylated recombinant human IGFBP-6, expressed in Chinese hamster ovary cells and Escherichia coli, respectively, were purified using IGF-II affinity chromatography and reverse-phase medium-pressure chromatography. Electrospray ionization mass spectrometry (ESMS) of glycosylated IGFBP-6 revealed considerable heterogeneity of carbohydrate composition. Major glycoforms contained 8-16 monosaccharides, including N-acetylhexosamine, hexose, and N-acetylneuraminic acid. Glycosylation sites of IGFBP-6 were identified as Thr126, Ser144, Thr145, Thr146, and Ser152 by using a combination of ESMS and Edman sequencing of tryptic fragments separated by reverse-phase high-pressure liquid chromatography. One oligosaccharide chain contained 5-6 monosaccharides, whereas the others contained 2-4 monosaccharides. Glycosylated IGFBP-6 exhibited greater resistance to proteolysis by chymotrypsin and trypsin than nonglycosylated IGFBP-6. Native disulfide bond positions in IGFBP-6 were localized by means of observed disulfide-linked tryptic fragments, revealing that there are two disulfide-linked subdomains within each of the N- and C-terminal regions and confirming a previous suggestion that the latter regions are not interconnected. A model of IGFBP-6 is developed in which these distinct domains are separated by a central region which is O-glycosylated.
Notes: Neumann, G M
Marinaro, J A
Bach, L A
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1998 May 5;37(18):6572-85. doi: 10.1021/bi972894e.
Author Address: School of Biochemistry, La Trobe University, Bundoora, Victoria 3083, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1366
Author: Neutzner, A., Neutzner, M., Benischke, A. S., Ryu, S. W., Frank, S., Youle, R. J. and Karbowski, M.
Year: 2011
Title: A systematic search for endoplasmic reticulum (ER) membrane-associated RING finger proteins identifies Nixin/ZNRF4 as a regulator of calnexin stability and ER homeostasis
Journal: J Biol Chem
Volume: 286
Issue: 10
Pages: 8633-8643
Epub Date: 2011/01/06
Date: Mar 11
Short Title: A systematic search for endoplasmic reticulum (ER) membrane-associated RING finger proteins identifies Nixin/ZNRF4 as a regulator of calnexin stability and ER homeostasis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.197459
PMCID: PMC3048745
Accession Number: 21205830
Keywords: Calnexin/genetics/*metabolism
DNA-Binding Proteins/genetics/*metabolism
Down-Regulation/*physiology
Endoplasmic Reticulum/genetics/*metabolism
Glycosylation
HeLa Cells
Humans
Protein Stability
Abstract: To identify novel regulators of endoplasmic reticulum (ER)-linked protein degradation and ER function, we determined the entire inventory of membrane-spanning RING finger E3 ubiquitin ligases localized to the ER. We identified 24 ER membrane-anchored ubiquitin ligases and found Nixin/ZNRF4 to be central for the regulation of calnexin turnover. Ectopic expression of wild type Nixin induced a dramatic down-regulation of the ER-localized chaperone calnexin that was prevented by inactivation of the Nixin RING domain. Importantly, Nixin physically interacts with calnexin in a glycosylation-independent manner, induces calnexin ubiquitination, and p97-dependent degradation, indicating an ER-associated degradation-like mechanism of calnexin turnover.
Notes: 1083-351x
Neutzner, Albert
Neutzner, Melanie
Benischke, Anne-Sophie
Ryu, Seung-Wook
Frank, Stephan
Youle, Richard J
Karbowski, Mariusz
R01 GM083131/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2011 Mar 11;286(10):8633-8643. doi: 10.1074/jbc.M110.197459. Epub 2011 Jan 4.
Author Address: From the Biochemistry Section, Surgical Neurological Branch, NINDS, National Institutes of Health, Bethesda, Maryland 20892,; the Department of Biomedicine, and the University Eye Clinic, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland,. Electronic address: albert.neutzner@unibas.ch.
the Department of Biomedicine, and the University Eye Clinic, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland,; the Department of Neuropathology, Institute of Pathology, University of Basel, Schönbeinstrasse 40, 4031 Basel, Switzerland.
the Department of Biomedicine, and the University Eye Clinic, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland.
From the Biochemistry Section, Surgical Neurological Branch, NINDS, National Institutes of Health, Bethesda, Maryland 20892,; the Department of Bio and Brain Engineering, Korea Advanced Institute of Science and Technology, Daejeon 305-701, South Korea, and.
the Department of Neuropathology, Institute of Pathology, University of Basel, Schönbeinstrasse 40, 4031 Basel, Switzerland.
From the Biochemistry Section, Surgical Neurological Branch, NINDS, National Institutes of Health, Bethesda, Maryland 20892.
From the Biochemistry Section, Surgical Neurological Branch, NINDS, National Institutes of Health, Bethesda, Maryland 20892,; the Center for Biomedical Engineering and Technology and Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, Maryland 21201. Electronic address: mkarbowski@umaryland.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 897
Author: Neve, E. P., Svensson, K., Fuxe, J. and Pettersson, R. F.
Year: 2003
Title: VIPL, a VIP36-like membrane protein with a putative function in the export of glycoproteins from the endoplasmic reticulum
Journal: Exp Cell Res
Volume: 288
Issue: 1
Pages: 70-83
Epub Date: 2003/07/25
Date: Aug 1
Short Title: VIPL, a VIP36-like membrane protein with a putative function in the export of glycoproteins from the endoplasmic reticulum
Alternate Journal: Experimental cell research
ISSN: 0014-4827 (Print)
0014-4827
DOI: 10.1016/s0014-4827(03)00161-7
Accession Number: 12878160
Keywords: Amino Acid Sequence
Carrier Proteins/genetics
Conserved Sequence
Databases, Nucleic Acid
Endoplasmic Reticulum/*metabolism
Glycoproteins/*metabolism
Humans
Lectins/genetics/*physiology
Mannose-Binding Lectins/genetics/*physiology
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase/metabolism
Membrane Proteins/genetics
Membrane Transport Proteins/genetics/*physiology
Molecular Sequence Data
Protein Transport
RNA, Messenger/metabolism
RNA, Small Interfering/pharmacology
Sequence Alignment
Tissue Distribution
Tumor Cells, Cultured
Abstract: Subsets of glycoproteins are thought to require lectin-like membrane receptors for efficient export out of the endoplasmic reticulum (ER). To identify new members related to two previously characterized intracellular lectins ERGIC-53/p58 and VIP36, we carried out an extensive database search using the conserved carbohydrate recognition domain (CRD) as a search string. A gene, more closely related to VIP36 than to ERGIC-53/p58, and hence called VIPL (VIP36-Like), was identified. VIPL has been conserved through evolution from zebra fish to man. The 2.4-kb VIPL mRNA was widely expressed to varying levels in different tissues. Using an antiserum prepared against the CRD, the 32-kDa VIPL protein was detected in various cell lines. The single N-linked glycan of VIPL remained endoglycosidase H-sensitive during a 2-h pulse-chase, even when the protein was overexpressed or mutated to allow export to the plasma membrane. VIPL localized primarily to the ER and partly to the Golgi complex. Like VIP36, the cytoplasmic tail of VIPL terminates in the sequence KRFY, a motif characteristic for proteins recycling between the ER and ERGIC/cis-Golgi. Mutating the retrograde transport signal KR to AA resulted in transport of VIPL to the cell surface. Finally, knock-down of VIPL mRNA using siRNA significantly slowed down the secretion of two glycoproteins (M(R) 35 and 250 kDa) to the medium, suggesting that VIPL may also function as an ER export receptor.
Notes: Neve, Etienne P A
Svensson, Kerstin
Fuxe, Jonas
Pettersson, Ralf F
Journal Article
United States
Exp Cell Res. 2003 Aug 1;288(1):70-83. doi: 10.1016/s0014-4827(03)00161-7.
Author Address: Ludwig Institute for Cancer Research, Stockholm Branch, Karolinska Institutet, Box 240, SE-17177 Stockholm, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1129
Author: Ngamelue, M. N., Homma, K., Lockridge, O. and Asojo, O. A.
Year: 2007
Title: Crystallization and X-ray structure of full-length recombinant human butyrylcholinesterase
Journal: Acta Crystallogr Sect F Struct Biol Cryst Commun
Volume: 63
Issue: Pt 9
Pages: 723-7
Epub Date: 2007/09/05
Date: Sep 1
Short Title: Crystallization and X-ray structure of full-length recombinant human butyrylcholinesterase
Alternate Journal: Acta crystallographica. Section F, Structural biology and crystallization communications
ISSN: 1744-3091
DOI: 10.1107/s1744309107037335
PMCID: PMC2376307
Accession Number: 17768338
Keywords: Butyrylcholinesterase/*chemistry/isolation & purification
Crystallization
Crystallography, X-Ray
Humans
Models, Molecular
Protein Conformation
Recombinant Proteins/chemistry/isolation & purification
Abstract: Human butyrylcholinesterase (BChE) has been shown to function as an endogenous scavenger of diverse poisons. BChE is a 340 kDa tetrameric glycoprotein that is present in human serum at a concentration of 5 mg l(-1). The well documented therapeutic effects of BChE on cocaine toxicity and organophosphorus agent poisoning has increased the need for effective methods of producing recombinant therapeutic BChE. In order to be therapeutically useful, BChE must have a long circulatory residence time or associate as tetramers. Full-length recombinant BChE produced in Chinese hamster ovary (CHO) cells or human embryonic kidney cells has been shown to associate as monomers, with a shorter circulatory residence time than the naturally occurring tetrameric serum protein. Based on the preceding observation as well as the need to develop novel methodologies to facilitate the mass production of therapeutic recombinant BChE, studies have been initiated to determine the structural basis of tetramer formation. Towards these ends, full-length monomeric recombinant BChE has been crystallized for the first time. A 2.8 A X-ray structure was solved in space group P42(1)2, with unit-cell parameters a = b = 156, c = 146 A.
Notes: 1744-3091
Ngamelue, Michelle N
Homma, Kohei
Lockridge, Oksana
Asojo, Oluwatoyin A
P41 RR007707/RR/NCRR NIH HHS/United States
Journal Article
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2007 Sep 1;63(Pt 9):723-7. doi: 10.1107/S1744309107037335. Epub 2007 Aug 10.
Author Address: Pathology and Microbiology Department, University of Nebraska Medical Center, 986495 Nebraska Medical Center, Omaha, NE 68198-6495, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1387
Author: Nguyen, T. T., Chang, S. C., Evnouchidou, I., York, I. A., Zikos, C., Rock, K. L., Goldberg, A. L., Stratikos, E. and Stern, L. J.
Year: 2011
Title: Structural basis for antigenic peptide precursor processing by the endoplasmic reticulum aminopeptidase ERAP1
Journal: Nat Struct Mol Biol
Volume: 18
Issue: 5
Pages: 604-13
Epub Date: 2011/04/12
Date: May
Short Title: Structural basis for antigenic peptide precursor processing by the endoplasmic reticulum aminopeptidase ERAP1
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb.2021
PMCID: PMC3087843
NIHMSID: NIHMS268283
Accession Number: 21478864
Keywords: Aminopeptidases/*chemistry/metabolism/physiology
*Antigen Presentation
Antigens/*metabolism
Binding Sites
Crystallography, X-Ray
Endoplasmic Reticulum/*metabolism
Humans
Kinetics
Leucine/*analogs & derivatives/chemistry/metabolism
Minor Histocompatibility Antigens
Protein Structure, Tertiary
Structure-Activity Relationship
Substrate Specificity
Abstract: ERAP1 trims antigen precursors to fit into MHC class I proteins. To fulfill this function, ERAP1 has unique substrate preferences, trimming long peptides but sparing shorter ones. To identify the structural basis for ERAP1's unusual properties, we determined the X-ray crystal structure of human ERAP1 bound to bestatin. The structure reveals an open conformation with a large interior compartment. An extended groove originating from the enzyme's catalytic center can accommodate long peptides and has features that explain ERAP1's broad specificity for antigenic peptide precursors. Structural and biochemical analyses suggest a mechanism for ERAP1's length-dependent trimming activity, whereby binding of long rather than short substrates induces a conformational change with reorientation of a key catalytic residue toward the active site. ERAP1's unique structural elements suggest how a generic aminopeptidase structure has been adapted for the specialized function of trimming antigenic precursors.
Notes: 1545-9985
Nguyen, Tina T
Chang, Shih-Chung
Evnouchidou, Irini
York, Ian A
Zikos, Christos
Rock, Kenneth L
Goldberg, Alfred L
Stratikos, Efstratios
Stern, Lawrence J
R37 AI038996-13/AI/NIAID NIH HHS/United States
R37 AI038996-12/AI/NIAID NIH HHS/United States
R01 AI038996/AI/NIAID NIH HHS/United States
P30 DK032520/DK/NIDDK NIH HHS/United States
DK32520/DK/NIDDK NIH HHS/United States
R56 AI038996/AI/NIAID NIH HHS/United States
AI-38996/AI/NIAID NIH HHS/United States
P30 EB009998/EB/NIBIB NIH HHS/United States
R37 AI038996-11A1/AI/NIAID NIH HHS/United States
R01 AI020248/AI/NIAID NIH HHS/United States
R37 AI038996/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Nat Struct Mol Biol. 2011 May;18(5):604-13. doi: 10.1038/nsmb.2021. Epub 2011 Apr 10.
Author Address: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1561
Author: Nicholson, A. M., Finch, N. A., Wojtas, A., Baker, M. C., Perkerson, R. B., 3rd, Castanedes-Casey, M., Rousseau, L., Benussi, L., Binetti, G., Ghidoni, R., Hsiung, G. Y., Mackenzie, I. R., Finger, E., Boeve, B. F., Ertekin-Taner, N., Graff-Radford, N. R., Dickson, D. W. and Rademakers, R.
Year: 2013
Title: TMEM106B p.T185S regulates TMEM106B protein levels: implications for frontotemporal dementia
Journal: J Neurochem
Volume: 126
Issue: 6
Pages: 781-91
Epub Date: 2013/06/08
Date: Sep
Short Title: TMEM106B p.T185S regulates TMEM106B protein levels: implications for frontotemporal dementia
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042 (Print)
0022-3042
DOI: 10.1111/jnc.12329
PMCID: PMC3766501
NIHMSID: NIHMS494079
Accession Number: 23742080
Keywords: Aged
Blotting, Western
Cells, Cultured
Cohort Studies
DNA, Complementary/biosynthesis/genetics
Enzyme-Linked Immunosorbent Assay
Female
Fluorescent Antibody Technique
Frontotemporal Dementia/*genetics/*metabolism
Genotype
Glycosylation
HeLa Cells
Humans
Immunohistochemistry
Intercellular Signaling Peptides and Proteins/biosynthesis/genetics
Isomerism
Lysosomes/metabolism
Male
Membrane Proteins/*genetics/*physiology
Mutagenesis/genetics
Mutation/genetics
Nerve Tissue Proteins/*genetics/*physiology
Polymorphism, Genetic/genetics
Polymorphism, Single Nucleotide/genetics
Progranulins
Real-Time Polymerase Chain Reaction
Tmem106b
frontotemporal dementia
progranulin
Abstract: Frontotemporal lobar degeneration (FTLD) is the second leading cause of dementia in individuals under age 65. In many patients, the predominant pathology includes neuronal cytoplasmic or intranuclear inclusions of ubiquitinated TAR DNA binding protein 43 (FTLD-TDP). Recently, a genome-wide association study identified the first FTLD-TDP genetic risk factor, in which variants in and around the TMEM106B gene (top SNP rs1990622) were significantly associated with FTLD-TDP risk. Intriguingly, the most significant association was in FTLD-TDP patients carrying progranulin (GRN) mutations. Here, we investigated to what extent the coding variant, rs3173615 (p.T185S) in linkage disequilibrium with rs1990622, affects progranulin protein (PGRN) biology and transmembrane protein 106 B (TMEM106B) regulation. First, we confirmed the association of TMEM106B variants with FTLD-TDP in a new cohort of GRN mutation carriers. We next generated and characterized a TMEM106B-specific antibody for investigation of this protein. Enzyme-linked immunoassay analysis of progranulin protein levels showed similar effects upon T185 and S185 TMEM106B over-expression. However, over-expression of T185 consistently led to higher TMEM106B protein levels than S185. Cycloheximide treatment experiments revealed that S185 degrades faster than T185 TMEM106B, potentially due to differences in N-glycosylation at residue N183. Together, our results provide a potential mechanism by which TMEM106B variants lead to differences in FTLD-TDP risk. We studied the p.T185S TMEM106B genetic variant previously implicated in frontotemporal dementia with TAR DNA binding protein 43 pathology caused by progranulin mutations. Our cell culture studies provide evidence that the protective S185 isoform is degraded more rapidly than T185 TMEM106B, potentially due to differences in glycosylation. These findings suggest that low TMEM106B levels might protect against FTLD-TDP in these patients.
Notes: 1471-4159
Nicholson, Alexandra M
Finch, Nicole A
Wojtas, Aleksandra
Baker, Matt C
Perkerson, Ralph B 3rd
Castanedes-Casey, Monica
Rousseau, Linda
Benussi, Luisa
Binetti, Giuliano
Ghidoni, Roberta
Hsiung, Ging-Yuek R
Mackenzie, Ian R
Finger, Elizabeth
Boeve, Bradley F
Ertekin-Taner, Nilüfer
Graff-Radford, Neill R
Dickson, Dennis W
Rademakers, Rosa
P50 AG016574/AG/NIA NIH HHS/United States
179009/Canadian Institutes of Health Research/Canada
R01 NS076471/NS/NINDS NIH HHS/United States
R01 AG032990/AG/NIA NIH HHS/United States
R01 NS080820/NS/NINDS NIH HHS/United States
R01 NS065782/NS/NINDS NIH HHS/United States
R01 AG026251/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Neurochem. 2013 Sep;126(6):781-91. doi: 10.1111/jnc.12329. Epub 2013 Jul 1.
Author Address: Department of Neuroscience, Mayo Clinic, Jacksonville, Florida, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1549
Author: Nicolardi, S., van der Burgt, Y. E., Dragan, I., Hensbergen, P. J. and Deelder, A. M.
Year: 2013
Title: Identification of new apolipoprotein-CIII glycoforms with ultrahigh resolution MALDI-FTICR mass spectrometry of human sera
Journal: J Proteome Res
Volume: 12
Issue: 5
Pages: 2260-8
Epub Date: 2013/03/27
Date: May 3
Short Title: Identification of new apolipoprotein-CIII glycoforms with ultrahigh resolution MALDI-FTICR mass spectrometry of human sera
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr400136p
Accession Number: 23527852
Keywords: Amino Acid Sequence
Apolipoprotein C-III/*blood/chemistry
Blood Proteins/chemistry/metabolism
Carbohydrate Conformation
Carbohydrate Sequence
Fourier Analysis
Glycosylation
Humans
Molecular Sequence Data
Molecular Weight
Protein Processing, Post-Translational
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Apolipoprotein-CIII (apoCIII) is an abundant blood glycoprotein associated with lipoprotein particles. Three different glycoforms have been described, all containing a mucin-type core-1 O-glycosylation with either zero, one or two sialic acids. Changes in the relative abundance of these glycoforms have been observed in a variety of different pathologies. In this study, ultrahigh resolution 15T MALDI Fourier transform ion cyclotron resonance (FTICR) MS was used to analyze apoCIII isoforms in serum protein profiles. For this purpose, serum proteins were purified using both a fully automated RPC18-based magnetic bead method and an RPC4 cartridge-based solid phase extraction method. Six new apoCIII isoforms were identified with low-ppm mass measurement errors and ultrahigh precision. These were characterized by more complex glycan moieties that are fucosylated instead of sialylated. To confirm the glycan moiety and localize the glycosylation site, top-down ESI-FTICR-MS/MS and bottom-up LC-ion trap MS/MS were used. A large variation in the presence and abundance of the fucosylated isoforms was found in a set of 96 serum samples. These findings of fucosylated apolipoprotein-CIII isoforms warrant further research to elucidate the implications these glycoforms may have for the plethora of studies where alterations in apoCIII have been linked to the development of many different pathologies.
Notes: 1535-3907
Nicolardi, Simone
van der Burgt, Yuri E M
Dragan, Irina
Hensbergen, Paul J
Deelder, André M
Journal Article
United States
J Proteome Res. 2013 May 3;12(5):2260-8. doi: 10.1021/pr400136p. Epub 2013 Apr 5.
Author Address: Center for Proteomics and Metabolomics, Leiden University Medical Center (LUMC), Albinusdreef 2, 2300 RC, Leiden, The Netherlands. s.nicolardi@lumc.nl
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 895
Author: Nicolet, Y., Lockridge, O., Masson, P., Fontecilla-Camps, J. C. and Nachon, F.
Year: 2003
Title: Crystal structure of human butyrylcholinesterase and of its complexes with substrate and products
Journal: J Biol Chem
Volume: 278
Issue: 42
Pages: 41141-7
Epub Date: 2003/07/19
Date: Oct 17
Short Title: Crystal structure of human butyrylcholinesterase and of its complexes with substrate and products
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M210241200
Accession Number: 12869558
Keywords: Acetylcholinesterase/chemistry
Animals
Binding Sites
Butyrylcholinesterase/*chemistry
Catalysis
Choline/chemistry
Crystallography, X-Ray
Electrons
Fourier Analysis
Humans
Models, Molecular
Protein Binding
Protein Conformation
Recombinant Proteins/chemistry/metabolism
Serine/chemistry
Torpedo/metabolism
Abstract: Cholinesterases are among the most efficient enzymes known. They are divided into two groups: acetylcholinesterase, involved in the hydrolysis of the neurotransmitter acetylcholine, and butyrylcholinesterase of unknown function. Several crystal structures of the former have shown that the active site is located at the bottom of a deep and narrow gorge, raising the question of how substrate and products enter and leave. Human butyrylcholinesterase (BChE) has attracted attention because it can hydrolyze toxic esters such as cocaine or scavenge organophosphorus pesticides and nerve agents. Here we report the crystal structures of several recombinant truncated human BChE complexes and conjugates and provide a description for mechanistically relevant non-productive substrate and product binding. As expected, the structure of BChE is similar to a previously published theoretical model of this enzyme and to the structure of Torpedo acetylcholinesterase. The main difference between the experimentally determined BChE structure and its model is found at the acyl binding pocket that is significantly bigger than expected. An electron density peak close to the catalytic Ser(198) has been modeled as bound butyrate.
Notes: Nicolet, Yvain
Lockridge, Oksana
Masson, Patrick
Fontecilla-Camps, Juan C
Nachon, Florian
Journal Article
United States
J Biol Chem. 2003 Oct 17;278(42):41141-7. doi: 10.1074/jbc.M210241200. Epub 2003 Jul 17.
Author Address: Laboratoire de Cristallographie et Cristallogénèse des Protéines, Institut de Biologie Structurale "Jean-Pierre Ebel," CEA, UJF, CNRS, 41 rue Jules Horowitz, 38027 Grenoble Cedex 1, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2054
Author: Nie, H., Ju, H., Fan, J., Shi, X., Cheng, Y., Cang, X., Zheng, Z., Duan, X. and Yi, W.
Year: 2020
Title: O-GlcNAcylation of PGK1 coordinates glycolysis and TCA cycle to promote tumor growth
Journal: Nat Commun
Volume: 11
Issue: 1
Pages: 36
Epub Date: 2020/01/09
Date: Jan 7
Short Title: O-GlcNAcylation of PGK1 coordinates glycolysis and TCA cycle to promote tumor growth
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-019-13601-8
PMCID: PMC6946671
Accession Number: 31911580
Keywords: Acetylglucosamine/*metabolism
Amino Acid Motifs
Animals
Cell Line, Tumor
*Citric Acid Cycle
Colonic Neoplasms/*enzymology/genetics/metabolism
*Glycolysis
Humans
Male
Mice
Mice, Nude
Mitochondria/metabolism
Phosphoglycerate Kinase/chemistry/genetics/*metabolism
Pyruvate Dehydrogenase Complex/metabolism
Abstract: Many cancer cells display enhanced glycolysis and suppressed mitochondrial metabolism. This phenomenon, known as the Warburg effect, is critical for tumor development. However, how cancer cells coordinate glucose metabolism through glycolysis and the mitochondrial tricarboxylic acid (TCA) cycle is largely unknown. We demonstrate here that phosphoglycerate kinase 1 (PGK1), the first ATP-producing enzyme in glycolysis, is reversibly and dynamically modified with O-linked N-acetylglucosamine (O-GlcNAc) at threonine 255 (T255). O-GlcNAcylation activates PGK1 activity to enhance lactate production, and simultaneously induces PGK1 translocation into mitochondria. Inside mitochondria, PGK1 acts as a kinase to inhibit pyruvate dehydrogenase (PDH) complex to reduce oxidative phosphorylation. Blocking T255 O-GlcNAcylation of PGK1 decreases colon cancer cell proliferation, suppresses glycolysis, enhances the TCA cycle, and inhibits tumor growth in xenograft models. Furthermore, PGK1 O-GlcNAcylation levels are elevated in human colon cancers. This study highlights O-GlcNAcylation as an important signal for coordinating glycolysis and the TCA cycle to promote tumorigenesis.
Notes: 2041-1723
Nie, Hao
Ju, Haixing
Fan, Jiayi
Shi, Xiaoliu
Cheng, Yaxian
Cang, Xiaohui
Zheng, Zhiguo
Duan, Xiaotao
Yi, Wen
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2020 Jan 7;11(1):36. doi: 10.1038/s41467-019-13601-8.
Author Address: MOE Key Laboratory of Biosystems Homeostasis & Protection, College of Life Sciences; The First Affiliated Hospital, School of Medicine, Zhejiang University, 310058, Hangzhou, China.
Department of Colorectal Surgery, Zhejiang Cancer Hospital, 310022, Hangzhou, China.
Division of Medical Genetics and Genomics, The Children's Hospital, School of Medicine, Zhejiang University, 310058, Hangzhou, China.
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, 100850, Beijing, China.
MOE Key Laboratory of Biosystems Homeostasis & Protection, College of Life Sciences; The First Affiliated Hospital, School of Medicine, Zhejiang University, 310058, Hangzhou, China. wyi@zju.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1102
Author: Nikolov, D., Li, C., Lackmann, M., Jeffrey, P. and Himanen, J.
Year: 2007
Title: Crystal structure of the human ephrin-A5 ectodomain
Journal: Protein Sci
Volume: 16
Issue: 5
Pages: 996-1000
Epub Date: 2007/04/03
Date: May
Short Title: Crystal structure of the human ephrin-A5 ectodomain
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.062665807
PMCID: PMC2206647
Accession Number: 17400922
Keywords: Crystallization
Crystallography, X-Ray
Ephrin-A5/*chemistry/metabolism
Humans
Models, Molecular
Protein Binding
*Protein Structure, Tertiary
Receptor Protein-Tyrosine Kinases/chemistry/metabolism
Abstract: The Eph receptors, the largest subfamily of receptor tyrosine kinases, and their ephrin ligands are important mediators of cell-cell communication regulating cell attachment, pathfinding, and mobility in the nervous and cardiovascular systems. Recent structural studies have revealed unique molecular features that explain many of the biochemical and signaling properties of Ephs and ephrins. Nevertheless, open questions remain, including understanding the precise molecular mechanism underlining their binding-partner preferences and subclass specificity. In this study, we have determined and present the crystal structure of the extracellular domain of ephrin-A5-the first structure of an unbound A-class ephrin. The structure, determined at 2.1 A resolution, is a variation of the Greek key beta-barrel folding topology, containing eight beta-strands, and stabilized by two disulphide bonds. Overall, ephrin-A5 is structurally very similar to ephrin-B1 and ephrin-B2 but, unlike ephrin-B2, it does not show dimerization either in solution or in the crystals. Comparing free ephrin-A5 to the previously published structure of EphB2-bound ephrin-A5 reveals that significant conformational changes occur only around the G-H ephrin loop that upon binding bends toward the receptor. Interestingly, the G-H loop undergoes a very similar conformational rearrangement in ephrin-B2 upon receptor binding. The results of this study further emphasize the importance of the G-H loop for receptor recognition and selectivity, and could serve as a starting point for the development of structure-based Eph antagonists.
Notes: 1469-896x
Nikolov, Dimitar
Li, Chen
Lackmann, Martin
Jeffrey, Philip
Himanen, Juha
R01 NS038486/NS/NINDS NIH HHS/United States
R21 GM075886/GM/NIGMS NIH HHS/United States
GM75886/GM/NIGMS NIH HHS/United States
NS38486/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Protein Sci. 2007 May;16(5):996-1000. doi: 10.1110/ps.062665807. Epub 2007 Mar 30.
Author Address: Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA. nikolovd@mskcc.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1844
Author: Nile, A. H., Mukund, S., Stanger, K., Wang, W. and Hannoush, R. N.
Year: 2017
Title: Unsaturated fatty acyl recognition by Frizzled receptors mediates dimerization upon Wnt ligand binding
Journal: Proc Natl Acad Sci U S A
Volume: 114
Issue: 16
Pages: 4147-4152
Epub Date: 2017/04/06
Date: Apr 18
Short Title: Unsaturated fatty acyl recognition by Frizzled receptors mediates dimerization upon Wnt ligand binding
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1618293114
PMCID: PMC5402412
Accession Number: 28377511
Keywords: Crystallography, X-Ray
Cysteine/*chemistry/metabolism
Fatty Acids, Unsaturated/*chemistry/metabolism
Frizzled Receptors/*chemistry/metabolism
Humans
Ligands
Models, Molecular
Protein Binding
Protein Multimerization
Protein Structure, Tertiary
Wnt Proteins/*chemistry/metabolism
beta Catenin/*chemistry/metabolism
*Frizzled
*Wnt
*cysteine-rich domain
*dimerization
*fatty acid
Abstract: Frizzled (FZD) receptors mediate Wnt signaling in diverse processes ranging from bone growth to stem cell activity. Moreover, high FZD receptor expression at the cell surface contributes to overactive Wnt signaling in subsets of pancreatic, ovarian, gastric, and colorectal tumors. Despite the progress in biochemical understanding of Wnt-FZD receptor interactions, the molecular basis for recognition of Wnt cis-unsaturated fatty acyl groups by the cysteine-rich domain (CRD) of FZD receptors remains elusive. Here, we determined a crystal structure of human FZD7 CRD unexpectedly bound to a 24-carbon fatty acid. We also report a crystal structure of human FZD5 CRD bound to C16:1 cis-Δ9 unsaturated fatty acid. Both structures reveal a dimeric arrangement of the CRD. The lipid-binding groove exhibits flexibility and spans both monomers, adopting a U-shaped geometry that accommodates the fatty acid. Re-evaluation of the published mouse FZD8 CRD structure reveals that it also shares the same architecture as FZD5 and FZD7 CRDs. Our results define a common molecular mechanism for recognition of the cis-unsaturated fatty acyl group, a necessary posttranslational modification of Wnts, by multiple FZD receptors. The fatty acid bridges two CRD monomers, implying that Wnt binding mediates FZD receptor dimerization. Our data uncover possibilities for the arrangement of Wnt-FZD CRD complexes and shed structural insights that could aide in the identification of pharmacological strategies to modulate FZD receptor function.
Notes: 1091-6490
Nile, Aaron H
Mukund, Susmith
Stanger, Karen
Wang, Weiru
Hannoush, Rami N
S10 OD021832/OD/NIH HHS/United States
Journal Article
Proc Natl Acad Sci U S A. 2017 Apr 18;114(16):4147-4152. doi: 10.1073/pnas.1618293114. Epub 2017 Apr 4.
Author Address: Department of Early Discovery Biochemistry, Genentech, South San Francisco, CA 94080.
Department of Structural Biology, Genentech, South San Francisco, CA 94080.
Department of Early Discovery Biochemistry, Genentech, South San Francisco, CA 94080; hannoush.rami@gene.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1316
Author: Nilsson, J., Nilsson, J., Larson, G. and Grahn, A.
Year: 2010
Title: Characterization of site-specific O-glycan structures within the mucin-like domain of alpha-dystroglycan from human skeletal muscle
Journal: Glycobiology
Volume: 20
Issue: 9
Pages: 1160-9
Epub Date: 2010/05/29
Date: Sep
Short Title: Characterization of site-specific O-glycan structures within the mucin-like domain of alpha-dystroglycan from human skeletal muscle
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwq082
Accession Number: 20507882
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Catalytic Domain
Dystroglycans/analysis/*chemistry/metabolism
Glycosylation
Humans
Molecular Sequence Data
Mucins/*chemistry
Muscle, Skeletal/chemistry/*metabolism
N-Acetylneuraminic Acid/chemistry/metabolism
Peptide Mapping
Polysaccharides/chemistry/*metabolism
Protein Processing, Post-Translational
Protein Structure, Tertiary/physiology
Structure-Activity Relationship
Substrate Specificity
Abstract: The glycosylation of the extracellular protein alpha-dystroglycan is important for its ligand-binding activity, and altered or blocked glycosylation is associated with several forms of congenital muscular dystrophies. By immunoprecipitation and sialic acid capture-and-release enrichment strategies, we isolated tryptic glycopeptides of alpha-dystroglycan from human skeletal muscle. Nano-liquid chromatography tandem mass spectrometry was used to identify both glycopeptides and peptides corresponding to the mucin-like and C-terminal domain of alpha-dystroglycan. The O-glycans found had either Hex-O-Thr or HexNAc-O-Ser/Thr anchored structures, which were often elongated and frequently, but not always, terminated with sialic acid. The HexNAc-O-Ser/Thr, but not Hex-O-Thr glycopeptides, displayed heterogeneity regarding glycan core structures and level of glycosylation site occupancy. We demonstrate for the first time glycan attachment sites of the NeuAcHexHexNAcHex-O structure corresponding to the anticipated Neu5Acalpha3Galbeta4GlcNAcbeta2Man-O-glycan (sLacNAc-Man), within the mucin-like domain of human alpha-dystroglycan from human skeletal muscle. Twenty-five glycopeptides were characterized from human alpha-dystroglycan, which provide insight to the complex in vivo O-glycosylation of alpha-dystroglycan.
Notes: 1460-2423
Nilsson, Johanna
Nilsson, Jonas
Larson, Göran
Grahn, Ammi
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2010 Sep;20(9):1160-9. doi: 10.1093/glycob/cwq082. Epub 2010 May 27.
Author Address: Institute of Biomedicine, Department of Clinical Chemistry and Transfusion Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1271
Author: Nilsson, J., Rüetschi, U., Halim, A., Hesse, C., Carlsohn, E., Brinkmalm, G. and Larson, G.
Year: 2009
Title: Enrichment of glycopeptides for glycan structure and attachment site identification
Journal: Nat Methods
Volume: 6
Issue: 11
Pages: 809-11
Epub Date: 2009/10/20
Date: Nov
Short Title: Enrichment of glycopeptides for glycan structure and attachment site identification
Alternate Journal: Nature methods
ISSN: 1548-7091
DOI: 10.1038/nmeth.1392
Accession Number: 19838169
Keywords: Cerebrospinal Fluid Proteins/*analysis
Glycopeptides/analysis
Glycoproteins/*chemistry/*isolation & purification
Glycosylation
Humans
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
Polysaccharides/analysis/chemistry
Proteomics
Tandem Mass Spectrometry
Abstract: We present a method to enrich for glycoproteins from proteomic samples. Sialylated glycoproteins were selectively periodate-oxidized, captured on hydrazide beads, trypsinized and released by acid hydrolysis of sialic acid glycosidic bonds. Mass spectrometric fragment analysis allowed identification of glycan structures, and additional fragmentation of deglycosylated ions yielded peptide sequence information, which allowed glycan attachment site and protein identification. We identified 36 N-linked and 44 O-linked glycosylation sites on glycoproteins from human cerebrospinal fluid.
Notes: 1548-7105
Nilsson, Jonas
Rüetschi, Ulla
Halim, Adnan
Hesse, Camilla
Carlsohn, Elisabet
Brinkmalm, Gunnar
Larson, Göran
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Methods. 2009 Nov;6(11):809-11. doi: 10.1038/nmeth.1392. Epub 2009 Oct 18.
Author Address: Department of Clinical Chemistry and Transfusion Medicine, Institute of Biomedicine, University of Gothenburg, Sahlgrenska University Hospital, Gothenburg, Sweden. jonas.nilsson@clinchem.gu.se
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 596
Author: Nimtz, M., Martin, W., Wray, V., Klöppel, K. D., Augustin, J. and Conradt, H. S.
Year: 1993
Title: Structures of sialylated oligosaccharides of human erythropoietin expressed in recombinant BHK-21 cells
Journal: Eur J Biochem
Volume: 213
Issue: 1
Pages: 39-56
Epub Date: 1993/04/01
Date: Apr 1
Short Title: Structures of sialylated oligosaccharides of human erythropoietin expressed in recombinant BHK-21 cells
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb17732.x
Accession Number: 8477709
Keywords: Amino Acid Sequence
Animals
Carbohydrate Sequence
Cells, Cultured
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Cloning, Molecular
Cricetinae
Erythropoietin/*chemistry/genetics
Glycosylation
Humans
Kidney/chemistry
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Molecular Structure
N-Acetylneuraminic Acid
Oligosaccharides/*chemistry
Polysaccharides/analysis
Recombinant Proteins/chemistry/genetics
Sialic Acids/*chemistry
Spectrometry, Mass, Fast Atom Bombardment
Abstract: The native structures of the Asn-linked oligosaccharides and the O-glycans at Ser126 of human erythropoietin expressed from recombinant BHK cells have been elucidated. Enzymatically released N-glycans were studied by methylation analyses, fast-atom-bombardment mass spectrometry as well as one- and two-dimensional 1H-NMR spectrometry at 600 MHz. Many (82.7%) were found to be tetraantennary N-acetyllactosamine-type (22.8% with one, 3.6% with two and 0.4% with three N-acetyllactosamine repeats) being tetrasialylated (41%), trisialylated (29.6%) and disialylated (12.2%). A few (9.7%; 4.1% 2,4-branched, 5.6%, 2,6-branched) of the chains were triantennary (5.4% trisialyl, 4.3% disialyl) and 4.6% were of the disialyl diantennary type. Almost all of the innermost GlcNAc residues were alpha 1-6 fucosylated and NeuAc was exclusively alpha 2-3 linked to Gal beta 1-4GlcNAc-R; 60% of the protein was found to be O-glycosylated at Ser126; structures were monosialylated (70%) or disialylated (30%) forms of the Gal beta 1-3GalNAc core type. Glycosylation patterns at individual Asn-Xaa-Thr/Ser sites were determined by analytical high-pH anion-exchange chromatography with pulsed amperometric detection. Only tetraantennary chains with 0-3 N-acetyllactosamine repeats were detected at Asn38 and Asn83, while almost all of the di- and triantennary oligosaccharides were attached to Asn24. Batch analysis of different preparations of recombinant erythropoietin revealed the high reproducibility of the production procedure. Structures containing terminal GalNAc-GlcNAc were detected in small amounts in a few batches.
Notes: Nimtz, M
Martin, W
Wray, V
Klöppel, K D
Augustin, J
Conradt, H S
Journal Article
England
Eur J Biochem. 1993 Apr 1;213(1):39-56. doi: 10.1111/j.1432-1033.1993.tb17732.x.
Author Address: Department of Cell Biology and Genetics, GBF-Gesellschaft für Biotechnologische Forschung mbH, Braunschweig, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 325
Author: Nimtz, M., Noll, G., Pâques, E. P. and Conradt, H. S.
Year: 1990
Title: Carbohydrate structures of a human tissue plasminogen activator variant expressed in recombinant Chinese hamster ovary cells
Journal: FEBS Lett
Volume: 271
Issue: 1-2
Pages: 14-8
Epub Date: 1990/10/01
Date: Oct 1
Short Title: Carbohydrate structures of a human tissue plasminogen activator variant expressed in recombinant Chinese hamster ovary cells
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(90)80361-l
Accession Number: 2226797
Keywords: Animals
Asparagine/genetics
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Clone Cells
Cricetinae
Cricetulus
Genetic Variation
Glutamine/genetics
Glycoproteins/biosynthesis/*chemistry/genetics
Glycosylation
Humans
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry
Plasminogen Activators/biosynthesis/*chemistry/genetics
Recombinant Proteins
Abstract: The carbohydrate structures of a genetically engineered human tissue plasminogen activator variant bearing a single N-glycosylation site at Asn 448 are reported. After isolation of the tryptic glycopeptide and liberation of the N-linked carbohydrates by polypeptide:N-glycosidase F, 6 major oligosaccharide fractions were separated by HPLC on NH2-bonded phase. Their structures were determined by compositional and methylation analyses combined with fast atom bombardment mass spectrometry. Seventy percent of the carbohydrates were of the biantennary complex type with fucose at the proximal GlcNAc and zero, one or two alpha 2-3 linked NeuAc. The remainder were triantennary structures with one, two or three NeuAc.
Notes: Nimtz, M
Noll, G
Pâques, E P
Conradt, H S
Journal Article
England
FEBS Lett. 1990 Oct 1;271(1-2):14-8. doi: 10.1016/0014-5793(90)80361-l.
Author Address: Department of Cell Biology and Genetics, GBF-Gesellschaft für Biotechnologische Forschung mbH, Braunschweig, FRG.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 538
Author: Nimtz, M., Wray, V., Rüdiger, A. and Conradt, H. S.
Year: 1995
Title: Identification and structural characterization of a mannose-6-phosphate containing oligomannosidic N-glycan from human erythropoietin secreted by recombinant BHK-21 cells
Journal: FEBS Lett
Volume: 365
Issue: 2-3
Pages: 203-8
Epub Date: 1995/05/29
Date: May 29
Short Title: Identification and structural characterization of a mannose-6-phosphate containing oligomannosidic N-glycan from human erythropoietin secreted by recombinant BHK-21 cells
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(95)00473-m
Accession Number: 7781780
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Chromatography, High Pressure Liquid
Cricetinae
Erythropoietin/biosynthesis/*chemistry
Glycosylation
Humans
Kidney
Magnetic Resonance Spectroscopy
Mannosephosphates/*analysis
Mass Spectrometry
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Recombinant Proteins/biosynthesis/chemistry
Transfection
Abstract: A sialidase resistant mono-charged N-glycan was isolated from glycosylation site I (Asn-24) of recombinant human erythropoietin expressed from baby hamster kidney (BHK-21) cells and constituted approximately 2-4% of the oligosaccharide material at this glycosylation site. Mass spectrometry and both 1- and 2-dimensional NMR techniques revealed a high mannose type structure (Man6) with a phospho-diesterbridged N-acetylglucosamine as follows: [formula: see text]
Notes: Nimtz, M
Wray, V
Rüdiger, A
Conradt, H S
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1995 May 29;365(2-3):203-8. doi: 10.1016/0014-5793(95)00473-m.
Author Address: Department of Molecular and Instrumental Structure Research, GBF-Gesellschaft für Biotechnologische Forschung mbH, Braunschweig, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1682
Author: Nin, D. S., Huang, W., Ali, M., Yew, C. W., Kutateladze, T. G. and Deng, L. W.
Year: 2015
Title: O-GlcNAcylation of MLL5β is essential for MLL5β-AP-1 transcription complex assembly at the HPV16/18-long control region
Journal: J Mol Cell Biol
Volume: 7
Issue: 2
Pages: 180-3
Epub Date: 2015/02/12
Date: Apr
Short Title: O-GlcNAcylation of MLL5β is essential for MLL5β-AP-1 transcription complex assembly at the HPV16/18-long control region
Alternate Journal: Journal of molecular cell biology
ISSN: 1674-2788 (Print)
1759-4685
DOI: 10.1093/jmcb/mjv009
PMCID: PMC4425831
Accession Number: 25670814
Keywords: Acetylglucosamine/metabolism
DNA-Binding Proteins/*metabolism
Female
Gene Expression Regulation, Neoplastic
Glycosylation
HeLa Cells
Human papillomavirus 16/*genetics
Human papillomavirus 18/*genetics
Humans
Locus Control Region
Protein Multimerization
Protein Processing, Post-Translational
Transcription Factor AP-1/*metabolism
Uterine Cervical Neoplasms/*metabolism/virology
Notes: 1759-4685
Nin, Dawn Sijin
Huang, Weitai
Ali, Muzaffar
Yew, Chow Wenn
Kutateladze, Tatiana G
Deng, Lih-Wen
R01 GM101664/GM/NIGMS NIH HHS/United States
GM101664/GM/NIGMS NIH HHS/United States
Letter
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Mol Cell Biol. 2015 Apr;7(2):180-3. doi: 10.1093/jmcb/mjv009. Epub 2015 Feb 10.
Author Address: Department of Biochemistry, Yong Loo Lin School of Medicine, National University Health System, National University of Singapore, Singapore 117596, Singapore.
Department of Pharmacology, University of Colorado School of Medicine, Aurora, CO 80045, USA.
Department of Biochemistry, Yong Loo Lin School of Medicine, National University Health System, National University of Singapore, Singapore 117596, Singapore bchdlw@nus.edu.sg.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1789
Author: Ning, X., Tao, T., Shen, J., Ji, Y., Xie, L., Wang, H., Liu, N., Xu, X., Sun, C., Zhang, D., Shen, A. and Ke, K.
Year: 2017
Title: The O-GlcNAc Modification of CDK5 Involved in Neuronal Apoptosis Following In Vitro Intracerebral Hemorrhage
Journal: Cell Mol Neurobiol
Volume: 37
Issue: 3
Pages: 527-536
Epub Date: 2016/06/19
Date: Apr
Short Title: The O-GlcNAc Modification of CDK5 Involved in Neuronal Apoptosis Following In Vitro Intracerebral Hemorrhage
Alternate Journal: Cellular and molecular neurobiology
ISSN: 0272-4340
DOI: 10.1007/s10571-016-0391-y
Accession Number: 27316643
Keywords: Acetylglucosamine/*metabolism
Animals
*Apoptosis/drug effects
Cerebral Hemorrhage/*enzymology/*pathology
Cyclin-Dependent Kinase 5/*metabolism
Glycosylation/drug effects
HEK293 Cells
Hemin/pharmacology
Humans
Models, Biological
Neurons/drug effects/*enzymology/*pathology
PC12 Cells
Rats
Cdk5
Intracerebral hemorrhage
Neuronal apoptosis
O-GlcNAc modification
Abstract: Contrary to cell cycle-associated cyclin-dependent kinases, CDK5 is best known for its regulation of signaling processes in regulating mammalian CNS development. Studies of CDK5 have focused on its phosphorylation, although the diversity of CDK5 functions in the brain suggests additional forms of regulation. Here we expanded on the functional roles of CDK5 glycosylation in neurons. We showed that CDK5 was dynamically modified with O-GlcNAc in response to neuronal activity and that glycosylation represses CDK5-dependent apoptosis by impairing its association with p53 pathway. Blocking glycosylation of CDK5 alters cellular function and increases neuronal apoptosis in the cell model of the ICH. Our findings demonstrated a new role for O-glycosylation in neuronal apoptosis and provided a mechanistic understanding of how glycosylation contributes to critical neuronal functions. Moreover, we identified a previously unknown mechanism for the regulation of activity-dependent gene expression, neural development, and apoptosis.
Notes: 1573-6830
Ning, Xiaojin
Tao, Tao
Shen, Jianhong
Ji, Yuteng
Xie, Lili
Wang, Hongmei
Liu, Ning
Xu, Xide
Sun, Chi
Zhang, Dongmei
Shen, Aiguo
Ke, Kaifu
Journal Article
United States
Cell Mol Neurobiol. 2017 Apr;37(3):527-536. doi: 10.1007/s10571-016-0391-y. Epub 2016 Jun 17.
Author Address: Department of Neurology, Affiliated Hospital of Nantong University, Nantong, 226001, China.
Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong, 226001, China.
Department of Neurosurgery, Affiliated Hospital of Nantong University, Nantong, 226001, China.
Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong, 226001, China. shag@ntu.edu.cn.
Co-Innovation Center of Neuroregeneration, Nantong University, Nantong, 226001, China. shag@ntu.edu.cn.
Department of Neurology, Affiliated Hospital of Nantong University, Nantong, 226001, China. kekaifu_nt@126.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1369
Author: Nishimasu, H., Okudaira, S., Hama, K., Mihara, E., Dohmae, N., Inoue, A., Ishitani, R., Takagi, J., Aoki, J. and Nureki, O.
Year: 2011
Title: Crystal structure of autotaxin and insight into GPCR activation by lipid mediators
Journal: Nat Struct Mol Biol
Volume: 18
Issue: 2
Pages: 205-12
Epub Date: 2011/01/18
Date: Feb
Short Title: Crystal structure of autotaxin and insight into GPCR activation by lipid mediators
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.1998
Accession Number: 21240269
Keywords: Amino Acid Sequence
Animals
Binding Sites
Cell Line
Crystallography, X-Ray
Humans
Hydrophobic and Hydrophilic Interactions
Lysophospholipids/chemistry/*metabolism
Mice
Models, Molecular
Molecular Sequence Data
Multienzyme Complexes/*chemistry/*metabolism
Phosphodiesterase I/*chemistry/*metabolism
Phosphoric Diester Hydrolases
Protein Conformation
Pyrophosphatases/*chemistry/*metabolism
Receptors, G-Protein-Coupled/*metabolism
Substrate Specificity
Abstract: Autotaxin (ATX, also known as Enpp2) is a secreted lysophospholipase D that hydrolyzes lysophosphatidylcholine to generate lysophosphatidic acid (LPA), a lipid mediator that activates G protein-coupled receptors to evoke various cellular responses. Here, we report the crystal structures of mouse ATX alone and in complex with LPAs with different acyl-chain lengths and saturations. These structures reveal that the multidomain architecture helps to maintain the structural rigidity of the lipid-binding pocket, which accommodates the respective LPA molecules in distinct conformations. They indicate that a loop region in the catalytic domain is a major determinant for the substrate specificity of the Enpp family enzymes. Furthermore, along with biochemical and biological data, these structures suggest that the produced LPAs are delivered from the active site to cognate G protein-coupled receptors through a hydrophobic channel.
Notes: 1545-9985
Nishimasu, Hiroshi
Okudaira, Shinichi
Hama, Kotaro
Mihara, Emiko
Dohmae, Naoshi
Inoue, Asuka
Ishitani, Ryuichiro
Takagi, Junichi
Aoki, Junken
Nureki, Osamu
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2011 Feb;18(2):205-12. doi: 10.1038/nsmb.1998. Epub 2011 Jan 16.
Author Address: Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 343
Author: Nishimura, H., Kawabata, S., Kisiel, W., Hase, S., Ikenaka, T., Takao, T., Shimonishi, Y. and Iwanaga, S.
Year: 1989
Title: Identification of a disaccharide (Xyl-Glc) and a trisaccharide (Xyl2-Glc) O-glycosidically linked to a serine residue in the first epidermal growth factor-like domain of human factors VII and IX and protein Z and bovine protein Z
Journal: J Biol Chem
Volume: 264
Issue: 34
Pages: 20320-5
Epub Date: 1989/12/05
Date: Dec 5
Short Title: Identification of a disaccharide (Xyl-Glc) and a trisaccharide (Xyl2-Glc) O-glycosidically linked to a serine residue in the first epidermal growth factor-like domain of human factors VII and IX and protein Z and bovine protein Z
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2511201
Keywords: Amino Acid Sequence
Animals
*Blood Proteins
Carbohydrate Sequence
Cattle
Disaccharides/*isolation & purification
*Epidermal Growth Factor
*Factor IX
*Factor VII
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Oligopeptides/*isolation & purification
*Serine
Trisaccharides/*isolation & purification
Abstract: We have recently described a unique trisaccharide linked to a serine residue in the first epidermal growth factor-like domains of bovine blood coagulation factors VII (Ser-52) and IX (Ser-53) (Hase, S., Kawabata, S., Nishimura, H., Takeya, H., Sueyoshi, T., Miyata, T., Iwanaga, S., Takao, T., Shimonishi, Y., and Ikenaka, T. (1988) J. Biochem. (Tokyo) 104, 867-868). The sugar chain identified in these clotting factors consists of 1 mol of hexose (glucose (Glc] and 2 mol of pentose (xylose (Xyl]. We report here that human factors VII and IX and protein Z and bovine protein Z also contain such carbohydrate moieties linked to a serine residue at the same position found in bovine factors VII and IX. A glycopeptide derived from each of these proteins was subjected to amino acid sequence and component sugar analyses and fast atom bombardment mass spectrometric analysis. The results indicate that the glycopeptide derived from human factor IX contains 1 mol each of Glc and Xyl. The reducing end of this disaccharide was identified as Glc by analyzing the disaccharide generated by hydrazinolysis. In contrast, human factor VII and protein Z yielded two different glycopeptides which contained Glc and Xyl at molar ratios of 1:1 and 1:2, respectively, suggesting microheterogeneity of these O-linked sugar chains. Bovine protein Z glycopeptide contained 1 mol of Glc and 2 mol of Xyl. These sugar compositions were confirmed by analyses of the intact proteins. In relation to the trisaccharide sugar chain previously discovered in bovine factors VII and IX, these findings indicate the existence of a Xyl2-Glc-Ser and a Xyl-Glc-Ser structure in the first epidermal growth factor-like domains of human factors VII and IX and protein Z in addition to that of bovine protein Z. Whether these carbohydrate moieties contribute to the biological activities of these proteins is unknown.
Notes: Nishimura, H
Kawabata, S
Kisiel, W
Hase, S
Ikenaka, T
Takao, T
Shimonishi, Y
Iwanaga, S
HL35246/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1989 Dec 5;264(34):20320-5.
Author Address: Department of Biology, Faculty of Science, Kyushu University 33, Fukuoka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 252
Author: Nishimura, H., Takao, T., Hase, S., Shimonishi, Y. and Iwanaga, S.
Year: 1992
Title: Human factor IX has a tetrasaccharide O-glycosidically linked to serine 61 through the fucose residue
Journal: J Biol Chem
Volume: 267
Issue: 25
Pages: 17520-5
Epub Date: 1992/09/05
Date: Sep 5
Short Title: Human factor IX has a tetrasaccharide O-glycosidically linked to serine 61 through the fucose residue
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1517205
Keywords: Amino Acid Sequence
Carbohydrates/analysis
Chromatography, Gel
Chromatography, High Pressure Liquid
Factor IXa/*chemistry
*Fucose
Glycopeptides/chemistry/isolation & purification
Glycosides
Humans
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Sequence Homology, Nucleic Acid
*Serine
Spectrometry, Mass, Fast Atom Bombardment
Abstract: We have recently discovered unusual sugar chains (xylose (Xyl)-glucose (Glc) and (Xyl)2-Glc) linked to a serine residue in the epidermal growth factor (EGF)-like domains of human and bovine clotting factors VII (Ser-52), IX (Ser-53), and protein Z (Ser-53), in addition to bovine platelet glycoprotein thrombospondin. We now have evidence of another modification in the first EGF-like domain of human factor IX, which proved to be a tetrasaccharide O-fucosidically linked to Ser-61. Two large peptides containing Ser-61 (positions 44-63), named hIX-GP1 and hIX-GP2, were first isolated from the lysyl endopeptidase-digest of human factor IX, by reversed-phase high performance liquid chromatography (HPLC). Data on the component sugar analysis after pyridylamination (PA) and sialic acid analysis of the isolated peptides indicated that they contained 1 mol each of galactose (Gal), fucose (Fuc), N-acetylglucosamine (GlcNAc), and N-acetylneuraminic acid (NeuAc), in addition to Glc and Xyl. hIX-GP1 was further digested with asparaginyl endopeptidase, and two glycopeptides containing Ser-61, named N-3 (positions 59-63) and N-9 (positions 55-63), were isolated, respectively. These glycopeptides were both composed of 1 mol each of Gal, Fuc, GlcNAc, and NeuAc but did not contain Xyl and Glc. Moreover, the data on beta-elimination for N-9 and of the fast atom bombardment mass spectrometric analysis on peptide N-3 suggested the presence of a tetrasaccharide linked to Ser-61. An analysis of the PA-oligosaccharide released from hIX-GP1 by hydrazinolysis followed by pyridylamination revealed that the reducing end was PA-Fuc. All the results support the proposal that human factor IX has a novel tetrasaccharide consisting of 1 mol each of Gal, Fuc, GlcNAc, and NeuAc, which is O-glycosidically linked to Ser-61 through the Fuc residue.
Notes: Nishimura, H
Takao, T
Hase, S
Shimonishi, Y
Iwanaga, S
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1992 Sep 5;267(25):17520-5.
Author Address: Department of Molecular Biology, Graduate School of Medical Science, Kyushu University, Fukuoka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1792
Author: Nishimura, K., Han, L., Bianchi, E., Wright, G. J., de Sanctis, D. and Jovine, L.
Year: 2016
Title: The structure of sperm Izumo1 reveals unexpected similarities with Plasmodium invasion proteins
Journal: Curr Biol
Volume: 26
Issue: 14
Pages: R661-2
Epub Date: 2016/07/05
Date: Jul 25
Short Title: The structure of sperm Izumo1 reveals unexpected similarities with Plasmodium invasion proteins
Alternate Journal: Current biology : CB
ISSN: 0960-9822 (Print)
0960-9822
DOI: 10.1016/j.cub.2016.06.028
PMCID: PMC4963209
Accession Number: 27374339
Keywords: Animals
Immunoglobulins/chemistry/*genetics/metabolism
Male
Membrane Proteins/chemistry/*genetics/metabolism
Mice/*genetics/metabolism
Plasmodium/*growth & development
Protozoan Proteins/chemistry/*genetics
Sequence Analysis, Protein
Spermatozoa/metabolism
Abstract: Fertilization, the culminating event in sexual reproduction, occurs when haploid sperm and egg recognize each other and fuse to form a diploid zygote. In mammals this process critically depends on the interaction between Izumo1, a protein exposed on the equatorial segment of acrosome-reacted sperm, and the egg plasma-membrane-anchored receptor Juno [1,2]. The molecular mechanism triggering gamete fusion is unresolved because both Izumo1 and Juno lack sequence similarity to known membrane fusogens. Here we report the crystal structure of Izumo1, which reveals a membrane distal region composed of a four-helix bundle connected to a carboxy-terminal immunoglobulin (Ig)-like domain through a β-hairpin stabilized by disulfide bonds. Remarkably, different regions of Izumo1 display significant structural similarities to two proteins expressed by the invasive sporozoite stage of Plasmodium parasites: SPECT1, which is essential for host cell traversal and hepatocyte invasion [3]; and TRAP, which is necessary for gliding motility and invasion [4]. These observations suggest a link between the molecular mechanisms underlying host cell invasion by the malaria parasite and gamete membrane fusion at fertilization.
Notes: 1879-0445
Nishimura, Kaoru
Han, Ling
Bianchi, Enrica
Wright, Gavin J
de Sanctis, Daniele
Jovine, Luca
260759/European Research Council/International
MR/M012468/1/Medical Research Council/United Kingdom
Letter
Research Support, Non-U.S. Gov't
Curr Biol. 2016 Jul 25;26(14):R661-2. doi: 10.1016/j.cub.2016.06.028. Epub 2016 Jun 30.
Author Address: Department of Biosciences and Nutrition & Center for Innovative Medicine, Karolinska Institutet, Huddinge, SE-141 83, Sweden.
Cell Surface Signalling Laboratory, Wellcome Trust Sanger Institute, Hinxton, Cambridge, CB10 1SA, UK.
ESRF - The European Synchrotron, Grenoble 38000, France.
Department of Biosciences and Nutrition & Center for Innovative Medicine, Karolinska Institutet, Huddinge, SE-141 83, Sweden. Electronic address: luca.jovine@ki.se.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 665
Author: Nissler, K., Kreusch, S., Rommerskirch, W., Strubel, W., Weber, E. and Wiederanders, B.
Year: 1998
Title: Sorting of non-glycosylated human procathepsin S in mammalian cells
Journal: Biol Chem
Volume: 379
Issue: 2
Pages: 219-24
Epub Date: 1998/04/02
Date: Feb
Short Title: Sorting of non-glycosylated human procathepsin S in mammalian cells
Alternate Journal: Biological chemistry
ISSN: 1431-6730 (Print)
1431-6730
DOI: 10.1515/bchm.1998.379.2.219
Accession Number: 9524075
Keywords: Animals
Cathepsins/genetics/*metabolism
Cell Fractionation
Cell Line
Cell Membrane/metabolism
Enzyme Activation
Enzyme Precursors/genetics/*metabolism
Gene Expression
Glycosylation
Humans
Mammals
Mutagenesis, Site-Directed
Abstract: Cathepsin S, a lysosomal cysteine protease, is synthesized as inactive precursor. It is activated in the lysosomes by a proteolytic cleavage of the propeptide. HEK 293-cells which do not express cathepsin S were transfected with cDNA of either wild type human procathepsin S or a mutant procathepsin S in which Asn of the only glycosylation site in the proregion was replaced by Gln. The cells expressed glycosylated and non-glycosylated procathepsin S, respectively. Large amounts of the precursors were secreted into the culture media by both transfectants. Secreted wild type procathepsin S contained Man-6-phosphate in the oligosaccharide chain. Wild type procathepsin S was activated in the cells but no maturation occurred in the culture media. In vitro processing of glycosylated as well as of non-glycosylated procathepsin S gave fully active enzymes thus indicating that the oligosaccharide chain was not necessary for proper folding. A reuptake of the glycosylated and non-glycosylated procathepsin S by HEK 293-cells could be observed. Small amounts of mature cathepsin S were detected in the lysosomes of the mutant transfectants. Subcellular fractionation showed non-glycosylated procathepsin S in the membrane fraction. Non-glycosylated procathepsin S was bound to the plasma membrane at 2 degrees C, suggesting an additional sorting motif in the cathepsin S molecule besides the Man-6-phosphate residue.
Notes: Nissler, K
Kreusch, S
Rommerskirch, W
Strubel, W
Weber, E
Wiederanders, B
Journal Article
Research Support, Non-U.S. Gov't
Germany
Biol Chem. 1998 Feb;379(2):219-24. doi: 10.1515/bchm.1998.379.2.219.
Author Address: Institut für Biochemie, Friedrich-Schiller-Universität, Jena, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 843
Author: Nitanai, Y., Satow, Y., Adachi, H. and Tsujimoto, M.
Year: 2002
Title: Crystal structure of human renal dipeptidase involved in beta-lactam hydrolysis
Journal: J Mol Biol
Volume: 321
Issue: 2
Pages: 177-84
Epub Date: 2002/07/30
Date: Aug 9
Short Title: Crystal structure of human renal dipeptidase involved in beta-lactam hydrolysis
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/s0022-2836(02)00632-0
Accession Number: 12144777
Keywords: Anti-Bacterial Agents/*metabolism
Binding Sites
Crystallography, X-Ray
Dimerization
Dipeptidases/*chemistry/*metabolism
Humans
Hydrolysis
Models, Molecular
Protein Structure, Quaternary
Protein Structure, Secondary
beta-Lactams
Abstract: Human renal dipeptidase is a membrane-bound glycoprotein hydrolyzing dipeptides and is involved in hydrolytic metabolism of penem and carbapenem beta-lactam antibiotics. The crystal structures of the saccharide-trimmed enzyme are determined as unliganded and inhibitor-liganded forms. They are informative for designing new antibiotics that are not hydrolyzed by this enzyme. The active site in each of the (alpha/beta)(8) barrel subunits of the homodimeric molecule is composed of binuclear zinc ions bridged by the Glu125 side-chain located at the bottom of the barrel, and it faces toward the microvillar membrane of a kidney tubule. A dipeptidyl moiety of the therapeutically used cilastatin inhibitor is fully accommodated in the active-site pocket, which is small enough for precise recognition of dipeptide substrates. The barrel and active-site architectures utilizing catalytic metal ions exhibit unexpected similarities to those of the murine adenosine deaminase and the catalytic domain of the bacterial urease.
Notes: Nitanai, Yasushi
Satow, Yoshinori
Adachi, Hideki
Tsujimoto, Masafumi
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2002 Aug 9;321(2):177-84. doi: 10.1016/s0022-2836(02)00632-0.
Author Address: Faculty of Pharmaceutical Sciences, University of Tokyo, Hongo, Bunkyo-ku, Tokyo, Japan. nitanai@spring8.or.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1913
Author: Niwa, Y., Nakano, Y., Suzuki, T., Yamagishi, M., Otani, K., Dohmae, N. and Simizu, S.
Year: 2018
Title: Topological analysis of DPY19L3, a human C-mannosyltransferase
Journal: Febs j
Volume: 285
Issue: 6
Pages: 1162-1174
Epub Date: 2018/02/07
Date: Mar
Short Title: Topological analysis of DPY19L3, a human C-mannosyltransferase
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/febs.14398
Accession Number: 29405629
Keywords: Amino Acid Sequence
Asparagine/chemistry/genetics/*metabolism
Binding Sites/genetics
Computer Simulation
Endoplasmic Reticulum/*metabolism
Glycosylation
HEK293 Cells
Humans
Mannose/metabolism
Mannosyltransferases/chemistry/genetics/*metabolism
Membrane Proteins/chemistry/genetics/*metabolism
Mutation
Protein Domains
Sequence Homology, Amino Acid
*C-mannosylation
*glycosyltransferase
*multipass membrane protein
*re-entrant loop
*redox-sensitive luciferase assay
Abstract: C-mannosylation is a rare type of protein glycosylation, the functions and mechanisms of which remain unclear. Recently, we identified DPY19L3 as a C-mannosyltransferase of R-spondin1 in human cells. DPY19L3 is predicted to be a multipass transmembrane protein that localizes to the endoplasmic reticulum (ER); however, its structure is undetermined. In this study, we propose a topological structure of DPY19L3 by in silico analysis and experimental methods such as redox-sensitive luciferase assay and introduction of N-glycosylation sites, suggesting that DPY19L3 comprises 11 transmembrane regions and two re-entrant loops with the N- and C-terminal ends facing the cytoplasm and ER lumen, respectively. Furthermore, DPY19L3 has four predicted N-glycosylation sites, and we have demonstrated that DPY19L3 is N-glycosylated at Asn(118) and Asn(704) but not Asn(319) and Asn(439) , supporting our topological model. By mass spectrometry, we measured the C-mannosyltransferase activity of N-glycosylation-defective mutants of DPY19L3 and isoform2, a splice variant, which lacks the C-terminal luminal region of DPY19L3. Isoform2 does not possess C-mannosyltransferase activity, indicating the importance of the C-terminal region; however, N-glycosylations of DPY19L3 do not have any roles for its enzymatic activity. These novel findings on DPY19L3 provide important insights into the mechanism of C-mannosylation.
Notes: 1742-4658
Niwa, Yuki
Nakano, Yoshihiko
Suzuki, Takehiro
Yamagishi, Mizuo
Otani, Kei
Dohmae, Naoshi
Simizu, Siro
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2018 Mar;285(6):1162-1174. doi: 10.1111/febs.14398. Epub 2018 Feb 16.
Author Address: Department of Applied Chemistry, Faculty of Science and Technology, Keio University, Yokohama, Japan.
Biomolecular Characterization Unit, RIKEN Center for Sustainable Resource Science, Wako, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1514
Author: Niwa, Y., Suzuki, T., Dohmae, N., Umezawa, K. and Simizu, S.
Year: 2012
Title: Determination of cathepsin V activity and intracellular trafficking by N-glycosylation
Journal: FEBS Lett
Volume: 586
Issue: 20
Pages: 3601-7
Epub Date: 2012/09/13
Date: Oct 19
Short Title: Determination of cathepsin V activity and intracellular trafficking by N-glycosylation
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1016/j.febslet.2012.08.001
Accession Number: 22967898
Keywords: Amino Acid Sequence
Binding Sites
Cathepsins/chemistry/*metabolism
Cell Line, Tumor
Cysteine Endopeptidases/chemistry/*metabolism
Glycosylation
Humans
Intracellular Space/*metabolism
Molecular Sequence Data
Protein Transport
Abstract: Cathepsin V (L2), a lysosomal cysteine protease, is a member of cathepsin family, relating to cancer invasion and metastasis. Cathepsin V contains two predicted N-glycosylation sites, but it has not been reported whether cathepsin V is glycosylated or not. In this study, we clarified the role of N-glycosylation of cathepsin V for its functions. We demonstrated that cathepsin V is N-glycosylated at both Asn(221) and Asn(292) using mass spectrometry and site-directed mutagenesis. N-glycosylation of cathepsin V was important for transportation to lysosome, secretion, and activity in HT1080 cells. These data demonstrated that functions of cathepsin V are controlled by N-glycosylation.
Notes: 1873-3468
Niwa, Yuki
Suzuki, Takehiro
Dohmae, Naoshi
Umezawa, Kazuo
Simizu, Siro
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2012 Oct 19;586(20):3601-7. doi: 10.1016/j.febslet.2012.08.001. Epub 2012 Aug 9.
Author Address: Department of Applied Chemistry, Faculty of Science and Technology, Keio University, Yokohama 223-8522, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1348
Author: Nogi, T., Yasui, N., Mihara, E., Matsunaga, Y., Noda, M., Yamashita, N., Toyofuku, T., Uchiyama, S., Goshima, Y., Kumanogoh, A. and Takagi, J.
Year: 2010
Title: Structural basis for semaphorin signalling through the plexin receptor
Journal: Nature
Volume: 467
Issue: 7319
Pages: 1123-7
Epub Date: 2010/10/01
Date: Oct 28
Short Title: Structural basis for semaphorin signalling through the plexin receptor
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature09473
Accession Number: 20881961
Keywords: Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray
HEK293 Cells
Humans
Ligands
Mice
Models, Molecular
Molecular Sequence Data
Nerve Tissue Proteins/*chemistry/genetics/*metabolism
Protein Binding
Protein Structure, Tertiary
Receptors, Cell Surface/*chemistry/genetics/*metabolism
Semaphorins/*chemistry/genetics/*metabolism
*Signal Transduction
Structure-Activity Relationship
Abstract: Semaphorins and their receptor plexins constitute a pleiotropic cell-signalling system that is used in a wide variety of biological processes, and both protein families have been implicated in numerous human diseases. The binding of soluble or membrane-anchored semaphorins to the membrane-distal region of the plexin ectodomain activates plexin's intrinsic GTPase-activating protein (GAP) at the cytoplasmic region, ultimately modulating cellular adhesion behaviour. However, the structural mechanism underlying the receptor activation remains largely unknown. Here we report the crystal structures of the semaphorin 6A (Sema6A) receptor-binding fragment and the plexin A2 (PlxnA2) ligand-binding fragment in both their pre-signalling (that is, before binding) and signalling (after complex formation) states. Before binding, the Sema6A ectodomain was in the expected 'face-to-face' homodimer arrangement, similar to that adopted by Sema3A and Sema4D, whereas PlxnA2 was in an unexpected 'head-on' homodimer arrangement. In contrast, the structure of the Sema6A-PlxnA2 signalling complex revealed a 2:2 heterotetramer in which the two PlxnA2 monomers dissociated from one another and docked onto the top face of the Sema6A homodimer using the same interface as the head-on homodimer, indicating that plexins undergo 'partner exchange'. Cell-based activity measurements using mutant ligands/receptors confirmed that the Sema6A face-to-face dimer arrangement is physiologically relevant and is maintained throughout signalling events. Thus, homodimer-to-heterodimer transitions of cell-surface plexin that result in a specific orientation of its molecular axis relative to the membrane may constitute the structural mechanism by which the ligand-binding 'signal' is transmitted to the cytoplasmic region, inducing GAP domain rearrangements and activation.
Notes: 1476-4687
Nogi, Terukazu
Yasui, Norihisa
Mihara, Emiko
Matsunaga, Yukiko
Noda, Masanori
Yamashita, Naoya
Toyofuku, Toshihiko
Uchiyama, Susumu
Goshima, Yoshio
Kumanogoh, Atsushi
Takagi, Junichi
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2010 Oct 28;467(7319):1123-7. doi: 10.1038/nature09473. Epub 2010 Sep 29.
Author Address: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1457
Author: Noinaj, N., Easley, N. C., Oke, M., Mizuno, N., Gumbart, J., Boura, E., Steere, A. N., Zak, O., Aisen, P., Tajkhorshid, E., Evans, R. W., Gorringe, A. R., Mason, A. B., Steven, A. C. and Buchanan, S. K.
Year: 2012
Title: Structural basis for iron piracy by pathogenic Neisseria
Journal: Nature
Volume: 483
Issue: 7387
Pages: 53-8
Epub Date: 2012/02/14
Date: Feb 12
Short Title: Structural basis for iron piracy by pathogenic Neisseria
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature10823
PMCID: PMC3292680
NIHMSID: NIHMS349012
Accession Number: 22327295
Keywords: Animals
Apoproteins/chemistry/metabolism
Bacterial Proteins/*chemistry/metabolism/ultrastructure
Binding Sites
Biological Transport
Cattle
Crystallography, X-Ray
Humans
Iron/*metabolism
Mice
Models, Molecular
Molecular Dynamics Simulation
Neisseria/*metabolism/pathogenicity
Protein Conformation
Scattering, Small Angle
Species Specificity
Structure-Activity Relationship
Transferrin/chemistry/metabolism/ultrastructure
Transferrin-Binding Protein A/*chemistry/*metabolism/ultrastructure
Transferrin-Binding Protein B/*chemistry/*metabolism/ultrastructure
X-Ray Diffraction
Abstract: Neisseria are obligate human pathogens causing bacterial meningitis, septicaemia and gonorrhoea. Neisseria require iron for survival and can extract it directly from human transferrin for transport across the outer membrane. The transport system consists of TbpA, an integral outer membrane protein, and TbpB, a co-receptor attached to the cell surface; both proteins are potentially important vaccine and therapeutic targets. Two key questions driving Neisseria research are how human transferrin is specifically targeted, and how the bacteria liberate iron from transferrin at neutral pH. To address these questions, we solved crystal structures of the TbpA-transferrin complex and of the corresponding co-receptor TbpB. We characterized the TbpB-transferrin complex by small-angle X-ray scattering and the TbpA-TbpB-transferrin complex by electron microscopy. Our studies provide a rational basis for the specificity of TbpA for human transferrin, show how TbpA promotes iron release from transferrin, and elucidate how TbpB facilitates this process.
Notes: 1476-4687
Noinaj, Nicholas
Easley, Nicole C
Oke, Muse
Mizuno, Naoko
Gumbart, James
Boura, Evzen
Steere, Ashley N
Zak, Olga
Aisen, Philip
Tajkhorshid, Emad
Evans, Robert W
Gorringe, Andrew R
Mason, Anne B
Steven, Alasdair C
Buchanan, Susan K
ZIA DK036143-04/ImNIH/Intramural NIH HHS/United States
P41 RR005969/RR/NCRR NIH HHS/United States
R01 DK021739/DK/NIDDK NIH HHS/United States
R01 GM086749/GM/NIGMS NIH HHS/United States
U54-GM087519/GM/NIGMS NIH HHS/United States
U54 GM087519/GM/NIGMS NIH HHS/United States
R01-GM086749/GM/NIGMS NIH HHS/United States
R01-DK21739/DK/NIDDK NIH HHS/United States
P41 GM104601/GM/NIGMS NIH HHS/United States
P41-RR05969/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Nature. 2012 Feb 12;483(7387):53-8. doi: 10.1038/nature10823.
Author Address: Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, US National Institutes of Health, Bethesda, Maryland 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1680
Author: Nørskov-Lauritsen, L., Jørgensen, S. and Bräuner-Osborne, H.
Year: 2015
Title: N-glycosylation and disulfide bonding affects GPRC6A receptor expression, function, and dimerization
Journal: FEBS Lett
Volume: 589
Issue: 5
Pages: 588-97
Epub Date: 2015/01/27
Date: Feb 27
Short Title: N-glycosylation and disulfide bonding affects GPRC6A receptor expression, function, and dimerization
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1016/j.febslet.2015.01.019
Accession Number: 25617829
Keywords: Amino Acid Sequence
Disulfides/*chemistry
Glycosylation
HEK293 Cells
Humans
Molecular Sequence Data
Protein Multimerization
Receptors, G-Protein-Coupled/*chemistry/*metabolism
Dimerization
Disulfide
G protein-coupled receptor (GPCR)
Gprc6a
N-linked glycosylation
Post-translational modification (PTM)
Signal transduction
Abstract: Investigation of post-translational modifications of receptor proteins is important for our understanding of receptor pharmacology and disease physiology. However, our knowledge about post-translational modifications of class C G protein-coupled receptors and how these modifications regulate expression and function is very limited. Herein, we show that the nutrient-sensing class C G protein-coupled receptor GPRC6A carries seven N-glycans and that one of these sites modulates surface expression whereas mutation of another site affects receptor function. GPRC6A has been speculated to form covalently linked dimers through cysteine disulfide linkage in the extracellular amino-terminal domain and here we show that GPRC6A indeed is a homodimer and that a disulfide bridge between the C131 residues is formed.
Notes: 1873-3468
Nørskov-Lauritsen, Lenea
Jørgensen, Stine
Bräuner-Osborne, Hans
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2015 Feb 27;589(5):588-97. doi: 10.1016/j.febslet.2015.01.019. Epub 2015 Jan 21.
Author Address: Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark.
Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark. Electronic address: hbo@sund.ku.dk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 673
Author: Noso, N., Bartels, J., Mallet, A. I., Mochizuki, M., Christophers, E. and Schröder, J. M.
Year: 1998
Title: Delayed production of biologically active O-glycosylated forms of human eotaxin by tumor-necrosis-factor-alpha-stimulated dermal fibroblasts
Journal: Eur J Biochem
Volume: 253
Issue: 1
Pages: 114-22
Epub Date: 1998/05/13
Date: Apr 1
Short Title: Delayed production of biologically active O-glycosylated forms of human eotaxin by tumor-necrosis-factor-alpha-stimulated dermal fibroblasts
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.1998.2530114.x
Accession Number: 9578468
Keywords: Amino Acid Sequence
Cells, Cultured
Chemokine CCL11
Chemokine CCL5/pharmacology
Chemokine CCL7
*Chemokines, CC
Chemotactic Factors, Eosinophil/*biosynthesis/chemistry/pharmacology
Chemotaxis, Leukocyte/drug effects
Cytokines/*biosynthesis/chemistry/pharmacology
Eosinophils/drug effects/physiology
Fibroblasts/drug effects/metabolism
Glycosylation
Humans
In Vitro Techniques
Molecular Sequence Data
Monocyte Chemoattractant Proteins/pharmacology
Recombinant Proteins/pharmacology
Sequence Homology, Amino Acid
Skin/*drug effects/*metabolism
Tumor Necrosis Factor-alpha/*pharmacology
Abstract: Since a number of inflammatory skin diseases are characterized by selective eosinophil infiltration preferentially in the dermis, we speculated that dermal fibroblasts might represent a potential cellular source of eosinophil-selective attractants. Cultivated dermal fibroblasts treated with tumor necrosis factor alpha secreted, not before day 3 of stimulation, eosinophil-specific chemotactic activity. Purification of this activity revealed a heparin-binding protein with an apparent molecular mass of 13 kDa in SDS/polyacrylamide gel electrophoresis. Peptide mapping with subsequent amino acid sequence analyses revealed it to be human eotaxin. Natural eotaxin preparations contain 50% N-terminally truncated forms missing two or three amino acids. It is O-glycosylated at Thr71, resulting in at least two sialylated O-glycosylated variants. Electrospray ionization mass spectrometric analyses revealed the natural eotaxin preparation to be heterogeneous with principal masses of 9033 Da and 9317 Da. Natural eotaxin stimulated eosinophil chemotaxis with identical potency and efficacy as recombinant human eotaxin. Neither neutrophils, monocytes or lymphocytes responded towards natural eotaxin preparations indicating that N-terminal truncation and O-glycosylation did not affect the cell-specificity of chemotactic activity. Treatment of eosinophils with natural eotaxin desensitizes chemotactic responses towards eotaxin, regulated an normal T-lymphocyte expressed and secreted (RANTES) and monocyte chemotactic protein 3 (MCP-3), whereas RANTES and MCP-3 were unable to desensitize natural eotaxin-dependent responses.
Notes: Noso, N
Bartels, J
Mallet, A I
Mochizuki, M
Christophers, E
Schröder, J M
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1998 Apr 1;253(1):114-22. doi: 10.1046/j.1432-1327.1998.2530114.x.
Author Address: Department of Dermatology, University of Kiel, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 565
Author: Novick, D., Cohen, B. and Rubinstein, M.
Year: 1994
Title: The human interferon alpha/beta receptor: characterization and molecular cloning
Journal: Cell
Volume: 77
Issue: 3
Pages: 391-400
Epub Date: 1994/05/06
Date: May 6
Short Title: The human interferon alpha/beta receptor: characterization and molecular cloning
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/0092-8674(94)90154-6
Accession Number: 8181059
Keywords: 3T3 Cells
Amino Acid Sequence
Amino Acids/analysis
Animals
Antibodies
Base Sequence
Cell Membrane/chemistry
Cloning, Molecular
DNA, Complementary/analysis
Humans
Interferon Type I/*metabolism
Membrane Proteins
Mice
Molecular Sequence Data
Neutralization Tests
Protein-Tyrosine Kinases/analysis
Rabbits
Radioligand Assay
Receptor, Interferon alpha-beta
Receptors, Interferon/analysis/chemistry/*genetics/immunology/isolation &
purification/*metabolism
Sequence Analysis
Sequence Analysis, DNA
Abstract: We describe a universal ligand-binding receptor for human interferons alpha and interferon beta (type I IFNs). A soluble 40 kDa IFN-alpha/beta receptor (p40) that blocks the activity of type I IFNs was purified from urine and sequenced. Antibodies raised against p40 completely block the activity of several type I IFNs and immuno-precipitate both a cellular 102 kDa IFN-alpha/beta receptor and its cross-linked complexes with IFN-alpha 2. The receptor is a disulfide-linked dimer, consisting of 51 kDa subunits. We isolated and expressed a 1.5 kb cDNA, coding for the IFN-alpha/beta receptor. Its 331 amino acid sequence includes a leader and a transmembrane region, while its ectodomain corresponds to p40. IFN-alpha/beta receptor is physically associated with the cytoplasmic Tyr kinase JAK1, hence, in addition to ligand binding, it is directly involved in signal transduction.
Notes: Novick, D
Cohen, B
Rubinstein, M
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell. 1994 May 6;77(3):391-400. doi: 10.1016/0092-8674(94)90154-6.
Author Address: Department of Molecular Genetics and Virology Weizmann Institute of Science, Rehovot, Israel.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 830
Author: Numao, S., Maurus, R., Sidhu, G., Wang, Y., Overall, C. M., Brayer, G. D. and Withers, S. G.
Year: 2002
Title: Probing the role of the chloride ion in the mechanism of human pancreatic alpha-amylase
Journal: Biochemistry
Volume: 41
Issue: 1
Pages: 215-25
Epub Date: 2002/01/05
Date: Jan 8
Short Title: Probing the role of the chloride ion in the mechanism of human pancreatic alpha-amylase
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi0115636
Accession Number: 11772019
Keywords: Base Sequence
Binding Sites
Chlorides/*physiology
Crystallization
Escherichia coli/enzymology
Humans
Kinetics
Models, Chemical
Molecular Sequence Data
Mutation
Pancreas/*enzymology
alpha-Amylases/*metabolism
Abstract: Human pancreatic alpha-amylase (HPA) is a member of the alpha-amylase family involved in the degradation of starch. Some members of this family, including HPA, require chloride for maximal activity. To determine the mechanism of chloride activation, a series of mutants (R195A, R195Q, N298S, R337A, and R337Q) were made in which residues in the chloride ion binding site were replaced. Mutations in this binding site were found to severely affect the ability of HPA to bind chloride ions with no binding detected for the R195 and R337 mutant enzymes. X-ray crystallographic analysis revealed that these mutations did not result in significant structural changes. However, the introduction of these mutations did alter the kinetic properties of the enzyme. Mutations to residue R195 resulted in a 20-450-fold decrease in the activity of the enzyme toward starch and shifted the pH optimum to a more basic pH. Interestingly, replacement of R337 with a nonbasic amino acid resulted in an alpha-amylase that no longer required chloride for catalysis and has a pH profile similar to that of wild-type HPA. In contrast, a mutation at residue N298 resulted in an enzyme that had much lower binding affinity for chloride but still required chloride for maximal activity. We propose that the chloride is required to increase the pK(a) of the acid/base catalyst, E233, which would otherwise be lower due to the presence of R337, a positively charged residue.
Notes: Numao, Shin
Maurus, Robert
Sidhu, Gary
Wang, Yili
Overall, Christopher M
Brayer, Gary D
Withers, Stephen G
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2002 Jan 8;41(1):215-25. doi: 10.1021/bi0115636.
Author Address: Department of Chemistry, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z1.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 676
Author: Nyman, T. A., Kalkkinen, N., Tölö, H. and Helin, J.
Year: 1998
Title: Structural characterisation of N-linked and O-linked oligosaccharides derived from interferon-alpha2b and interferon-alpha14c produced by Sendai-virus-induced human peripheral blood leukocytes
Journal: Eur J Biochem
Volume: 253
Issue: 2
Pages: 485-93
Epub Date: 1998/07/08
Date: Apr 15
Short Title: Structural characterisation of N-linked and O-linked oligosaccharides derived from interferon-alpha2b and interferon-alpha14c produced by Sendai-virus-induced human peripheral blood leukocytes
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.1998.2530485.x
Accession Number: 9654101
Keywords: Glycosylation
Humans
Interferon-alpha/isolation & purification/*metabolism
Leukocytes, Mononuclear/*metabolism/virology
Mass Spectrometry
Oligosaccharides/*chemistry/isolation & purification
Polysaccharides/chemistry
*Respirovirus
Abstract: We have previously isolated and partially characterised the components of a highly purified interferon-alpha (IFN-alpha) preparation produced by Sendai-virus-induced human peripheral blood leukocytes. Nine IFN-alpha species were identified, and two of these were found to be glycosylated [Nyman, T. A., Tölö, H., Parkkinen, J. & Kalkkinen, N. (1998) Identification of nine interferon-alpha subtypes produced by Sendai-virus-induced human peripheral blood leukocytes, Biochem. J. 329, 295-302]. Here, we isolated the N-linked oligosaccharides of IFN-alpha14c and the O-linked chains of IFN-alpha2b, and the glycans were characterised by electrospray tandem mass spectrometry and by specific glycosidase digestions monitored by matrix-assisted laser desorption ionisation time of flight mass spectrometry. The IFN-alpha14c N-glycans were shown to exhibit core-fucosylated biantennary glycans, with about 10% carrying an additional alpha1,3-linked fucose unit at the antennae. The IFN-alpha2b was shown to carry about 50% core type-1 disialyltetrasaccharides, 30% core type-1 monosialyltrisaccharides and 20% core type-2 monosialylpentasaccharides.
Notes: Nyman, T A
Kalkkinen, N
Tölö, H
Helin, J
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1998 Apr 15;253(2):485-93. doi: 10.1046/j.1432-1327.1998.2530485.x.
Author Address: Institute of Biotechnology, Protein Chemistry Laboratory, University of Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 656
Author: Nyman, T. A., Tölö, H., Parkkinen, J. and Kalkkinen, N.
Year: 1998
Title: Identification of nine interferon-alpha subtypes produced by Sendai virus-induced human peripheral blood leucocytes
Journal: Biochem J
Volume: 329 ( Pt 2)
Issue: Pt 2
Pages: 295-302
Epub Date: 1998/02/28
Date: Jan 15
Short Title: Identification of nine interferon-alpha subtypes produced by Sendai virus-induced human peripheral blood leucocytes
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj3290295
PMCID: PMC1219044
Accession Number: 9425112
Keywords: Amino Acid Sequence
Cells, Cultured
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Humans
Interferon-alpha/biosynthesis/chemistry/*classification/isolation & purification
Leukocytes/*metabolism
Molecular Sequence Data
Respirovirus/*physiology
Sequence Homology, Amino Acid
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: The human interferon-alpha (IFN-alpha) family is encoded by 13 different functional genes, and including all cloned sequence variants there are 28 potential IFN-alpha proteins. To find out which of the described sequences are expressed in normal human leucocytes, we have isolated and partly characterized the components of a highly purified IFN-alpha preparation produced by Sendai virus-induced human peripheral blood leucocytes. The identification protocol consisted of N-terminal sequencing and mass mapping of the proteins separated by reverse-phase HPLC and/or SDS/PAGE. The highly purified leucocyte IFN-alpha preparation was found to contain at least nine different IFN-alpha species: IFN-alpha1a, IFN-alpha2b, IFN-alpha4b, IFN-alpha7a, IFN-alpha8b, IFN-alpha10a, IFN-alpha14c, IFN-alpha17b and IFN-alpha21b. IFN-alpha1a was the major subtype, comprising approx. 30% of total leucocyte IFN-alpha. IFN-alpha14c, the only subtype containing potential N-glycosylation sites, was shown to be glycosylated at Asn-72. Molecular mass determination of the intact proteins by electrospray ionization MS showed that there are no other post-translational modifications in the IFN-alpha subtypes than the glycosylation of IFN-alpha2b and IFN-alpha14c. Only one sequence variant was found for each subtype, suggesting that the other described gene sequences represent allelic variants or mutations that are more rarely found in the general population.
Notes: 1470-8728
Nyman, T A
Tölö, H
Parkkinen, J
Kalkkinen, N
Journal Article
Biochem J. 1998 Jan 15;329 ( Pt 2)(Pt 2):295-302. doi: 10.1042/bj3290295.
Author Address: Institute of Biotechnology, Protein Chemistry Laboratory, P.O. Box 56 (Viikinkaari 9), FIN-00014 University of Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1409
Author: Öberg, F., Sjöhamn, J., Fischer, G., Moberg, A., Pedersen, A., Neutze, R. and Hedfalk, K.
Year: 2011
Title: Glycosylation increases the thermostability of human aquaporin 10 protein
Journal: J Biol Chem
Volume: 286
Issue: 36
Pages: 31915-23
Epub Date: 2011/07/08
Date: Sep 9
Short Title: Glycosylation increases the thermostability of human aquaporin 10 protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.242677
PMCID: PMC3173105
Accession Number: 21733844
Keywords: Aquaporins/*chemistry/metabolism
Binding Sites
Biological Transport
Glycosylation
Hot Temperature
Humans
Pichia/genetics
Protein Stability
Abstract: Human aquaporin10 (hAQP10) is a transmembrane facilitator of both water and glycerol transport in the small intestine. This aquaglyceroporin is located in the apical membrane of enterocytes and is believed to contribute to the passage of water and glycerol through these intestinal absorptive cells. Here we overproduced hAQP10 in the yeast Pichia pastoris and observed that the protein is glycosylated at Asn-133 in the extracellular loop C. This finding confirms one of three predicted glycosylation sites for hAQP10, and its glycosylation is unique for the human aquaporins overproduced in this host. Nonglycosylated protein was isolated using both glycan affinity chromatography and through mutating asparagine 133 to a glutamine. All three forms of hAQP10 where found to facilitate the transport of water, glycerol, erythritol, and xylitol, and glycosylation had little effect on functionality. In contrast, glycosylated hAQP10 showed increased thermostability of 3-6 °C compared with the nonglycosylated protein, suggesting a stabilizing effect of the N-linked glycan. Because only one third of hAQP10 was glycosylated yet the thermostability titration was mono-modal, we suggest that the presence of at least one glycosylated protein within each tetramer is sufficient to convey an enhanced structural stability to the remaining hAQP10 protomers of the tetramer.
Notes: 1083-351x
Öberg, Fredrik
Sjöhamn, Jennie
Fischer, Gerhard
Moberg, Andreas
Pedersen, Anders
Neutze, Richard
Hedfalk, Kristina
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 Sep 9;286(36):31915-23. doi: 10.1074/jbc.M111.242677. Epub 2011 Jul 6.
Author Address: Department of Chemistry/Biochemistry, University of Gothenburg, P. O. Box 462, SE-405 30 Göteborg, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 365
Author: O'Brien, J. S., Kretz, K. A., Dewji, N., Wenger, D. A., Esch, F. and Fluharty, A. L.
Year: 1988
Title: Coding of two sphingolipid activator proteins (SAP-1 and SAP-2) by same genetic locus
Journal: Science
Volume: 241
Issue: 4869
Pages: 1098-101
Epub Date: 1988/08/26
Date: Aug 26
Short Title: Coding of two sphingolipid activator proteins (SAP-1 and SAP-2) by same genetic locus
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.2842863
Accession Number: 2842863
Keywords: Amino Acid Sequence
Animals
Base Sequence
Carcinoma, Hepatocellular/analysis
Chromosome Mapping
Chromosomes, Human, Pair 10
DNA/genetics/isolation & purification
Glycoproteins/analysis/*genetics
Humans
Liver Neoplasms/analysis
Male
Mice
Mice, Nude
Molecular Sequence Data
Nucleic Acid Hybridization
Protein Conformation
Protein Precursors/analysis/genetics
Protein Processing, Post-Translational
Rats
Saposins
Sphingolipid Activator Proteins
Tissue Distribution
Abstract: Several complementary DNAs (cDNAs) coding for sphingolipid activator protein-2 (SAP-2) were isolated from a lambda gt-11 human hepatoma library by means of polyclonal antibodies. The nucleotide sequence of the largest cDNA was colinear with the derived amino acid sequence of SAP-2 and with the nucleotide sequence of the cDNA coding for the 70-kilodalton precursor of SAP-1 (SAP precursor cDNA). The coding sequence for mature SAP-2 was located 3' to that coding for SAP-1 in the SAP precursor cDNA. Both SAP-1 and SAP-2 appeared to be derived by proteolytic processing from a common precursor that is coded by a genetic locus on human chromosome 10. Two other domains similar to SAP-1 and SAP-2 were also identified in SAP precursor protein. Each of the four domains was approximately 80 amino acid residues long, had nearly identical placement of cysteine residues, potential glycosylation sites, and proline residues. Each domain also contained internal amino acid sequences capable of forming amphipathic helices separated by helix breakers to give a cylindrical hydrophobic domain that is probably stabilized by disulfide bridges. Protein immunoblotting experiments indicated that SAP precursor protein (70 kilodaltons) as well as immunoreactive SAP-like proteins of intermediate sizes (65, 50, and 31 kilodaltons) are present in most human tissues.
Notes: O'Brien, J S
Kretz, K A
Dewji, N
Wenger, D A
Esch, F
Fluharty, A L
DK 38795/DK/NIDDK NIH HHS/United States
HD 18983/HD/NICHD NIH HHS/United States
NS 08682/NS/NINDS NIH HHS/United States
etc.
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Science. 1988 Aug 26;241(4869):1098-101. doi: 10.1126/science.2842863.
Author Address: Department of Neurosciences, University of California, San Diego, La Jolla 92093.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 735
Author: Oefner, C., D'Arcy, A., Hennig, M., Winkler, F. K. and Dale, G. E.
Year: 2000
Title: Structure of human neutral endopeptidase (Neprilysin) complexed with phosphoramidon
Journal: J Mol Biol
Volume: 296
Issue: 2
Pages: 341-9
Epub Date: 2000/02/12
Date: Feb 18
Short Title: Structure of human neutral endopeptidase (Neprilysin) complexed with phosphoramidon
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1006/jmbi.1999.3492
Accession Number: 10669592
Keywords: Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Disulfides/metabolism
Enzyme Inhibitors/chemistry/*metabolism/pharmacology
Glycopeptides/chemistry/*metabolism/pharmacology
Humans
Hydrogen Bonding
Models, Molecular
Molecular Sequence Data
Neprilysin/antagonists & inhibitors/*chemistry/classification/*metabolism
Peptide Fragments/antagonists & inhibitors/chemistry/metabolism
Protein Conformation
Recombinant Proteins/antagonists & inhibitors/chemistry/metabolism
Sequence Alignment
Solubility
Substrate Specificity
Abstract: Neutral endopeptidase is a mammalian type II integral membrane zinc-containing endopeptidase, which degrades and inactivates a number of bioactive peptides. The range of substrates cleaved by neutral endopeptidase in vitro includes the enkephalins, substance P, endothelin, bradykinin and atrial natriuretic factor. Due to the physiological importance of neutral endopeptidase in the modulation of nociceptive and pressor responses there is considerable interest in inhibitors of this enzyme as novel analgesics and anti-hypertensive agents. Here we describe the crystal structure of the extracellular domain (residues 52-749) of human NEP complexed with the generic metalloproteinase inhibitor phosphoramidon at 2.1 A resolution. The structure reveals two multiply connected folding domains which embrace a large central cavity containing the active site. The inhibitor is bound to one side of this cavity and its binding mode provides a detailed understanding of the ligand-binding and specificity determinants.
Notes: Oefner, C
D'Arcy, A
Hennig, M
Winkler, F K
Dale, G E
Journal Article
England
J Mol Biol. 2000 Feb 18;296(2):341-9. doi: 10.1006/jmbi.1999.3492.
Author Address: Pharma Preclinical Research, F. Hoffmann-La Roche Ltd., Basel, CH-4070, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1126
Author: Oefner, C., Pierau, S., Schulz, H. and Dale, G. E.
Year: 2007
Title: Structural studies of a bifunctional inhibitor of neprilysin and DPP-IV
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 63
Issue: Pt 9
Pages: 975-81
Epub Date: 2007/08/21
Date: Sep
Short Title: Structural studies of a bifunctional inhibitor of neprilysin and DPP-IV
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s0907444907036281
Accession Number: 17704566
Keywords: Adenosine Deaminase/metabolism
*Adenosine Deaminase Inhibitors
Binding, Competitive
Crystallography, X-Ray
Diabetes Mellitus, Type 2/drug therapy
Dipeptidyl Peptidase 4/metabolism
*Dipeptidyl-Peptidase IV Inhibitors
Glucagon-Like Peptide 1/metabolism
Glycoproteins/*antagonists & inhibitors/metabolism
Humans
Models, Molecular
Molecular Structure
Neprilysin/*antagonists & inhibitors/chemistry/*metabolism
Oligopeptides/*chemistry/*pharmacology
Peptide Fragments/chemistry
Protein Binding
Protein Structure, Tertiary
Zinc/chemistry
Abstract: Neutral endopeptidase (NEP) is the major enzyme involved in the metabolic inactivation of a number of bioactive peptides including the enkephalins, substance P, endothelin, bradykinin and atrial natriuretic factor, as well as the incretin hormone glucagon-like peptide 1 (GLP-1), which is a potent stimulator of insulin secretion. The activity of GLP-1 is also rapidly abolished by the serine protease dipeptidyl peptidase IV (DPP-IV), which led to an elevated interest in inhibitors of this enzyme for the treatment of type II diabetes. A dual NEP/DPP-IV inhibitor concept is proposed, offering an alternative strategy for the treatment of type 2 diabetes. Here, the synthesis and crystal structures of the soluble extracellular domain of human NEP (residues 52-749) complexed with the NEP, competitive and potent dual NEP/DPP-IV inhibitor MCB3937 are described.
Notes: Oefner, Christian
Pierau, Sabine
Schulz, Henk
Dale, Glenn E
Journal Article
United States
Acta Crystallogr D Biol Crystallogr. 2007 Sep;63(Pt 9):975-81. doi: 10.1107/S0907444907036281. Epub 2007 Aug 17.
Author Address: Morphochem AG, CH-4058, Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 920
Author: Oefner, C., Roques, B. P., Fournie-Zaluski, M. C. and Dale, G. E.
Year: 2004
Title: Structural analysis of neprilysin with various specific and potent inhibitors
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 60
Issue: Pt 2
Pages: 392-6
Epub Date: 2004/01/30
Date: Feb
Short Title: Structural analysis of neprilysin with various specific and potent inhibitors
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s0907444903027410
Accession Number: 14747736
Keywords: Binding, Competitive
Crystallography, X-Ray
Electrons
Enkephalins/chemistry
Humans
Models, Chemical
Models, Molecular
Neprilysin/*chemistry
Peptides/chemistry
Protein Binding
Protein Structure, Tertiary
Zinc/chemistry
Abstract: Neutral endopeptidase (NEP) is the major enzyme involved in the metabolic inactivation of a number of bioactive peptides including the enkephalins, substance P, endothelin, bradykinin and atrial natriuretic factor. Owing to the physiological importance of NEP in the modulation of nociceptive and pressor responses, there is considerable interest in inhibitors of this enzyme as novel analgesics and antihypertensive agents. Here, the crystal structures of the soluble extracellular domain of human NEP (residues 52-749) complexed with various potent and competitive inhibitors are described. The structures unambiguously reveal the binding mode of the different zinc-chelating groups and the subsite specificity of the enzyme.
Notes: Oefner, Christian
Roques, Bernard P
Fournie-Zaluski, Marie-Claude
Dale, Glenn E
Journal Article
United States
Acta Crystallogr D Biol Crystallogr. 2004 Feb;60(Pt 2):392-6. doi: 10.1107/S0907444903027410. Epub 2004 Jan 23.
Author Address: Morphochem AG, CH-4058 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1868
Author: Oertwig, K., Ulbricht, D., Hanke, S., Pippel, J., Bellmann-Sickert, K., Sträter, N. and Heiker, J. T.
Year: 2017
Title: Glycosylation of human vaspin (SERPINA12) and its impact on serpin activity, heparin binding and thermal stability
Journal: Biochim Biophys Acta Proteins Proteom
Volume: 1865
Issue: 9
Pages: 1188-1194
Epub Date: 2017/07/03
Date: Sep
Short Title: Glycosylation of human vaspin (SERPINA12) and its impact on serpin activity, heparin binding and thermal stability
Alternate Journal: Biochimica et biophysica acta. Proteins and proteomics
ISSN: 1570-9639 (Print)
1570-9639
DOI: 10.1016/j.bbapap.2017.06.020
Accession Number: 28668641
Keywords: Asparagine/metabolism
Glycosylation
HEK293 Cells
Heparin/metabolism
Humans
Kallikreins/metabolism
Models, Molecular
Protein Binding
Protein Conformation
Protein Processing, Post-Translational
Protein Stability
Recombinant Proteins/metabolism
Serpins/*chemistry/metabolism
Structure-Activity Relationship
*Adipokine
*Glycosylation
*Kallikrein 7
*Serine protease
*Serpin
*Vaspin
Abstract: Vaspin is a glycoprotein with three predicted glycosylation sites at asparagine residues located in proximity to the reactive center loop and close to domains that play important roles in conformational changes underlying serpin function. In this study, we have investigated the glycosylation of human vaspin and its effects on biochemical properties relevant to vaspin function. We show that vaspin is modified at all three sites and biochemical data demonstrate that glycosylation does not hinder inhibition of the target protease kallikrein 7. Although binding affinity to heparin is slightly decreased, the protease inhibition reaction is still significantly accelerated in the presence of heparin. Glycosylation did not affect thermal stability.
Notes: Oertwig, Kathrin
Ulbricht, David
Hanke, Stefanie
Pippel, Jan
Bellmann-Sickert, Kathrin
Sträter, Norbert
Heiker, John T
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta Proteins Proteom. 2017 Sep;1865(9):1188-1194. doi: 10.1016/j.bbapap.2017.06.020. Epub 2017 Jun 29.
Author Address: Institute of Biochemistry, Faculty of Biosciences, Pharmacy and Psychology, University of Leipzig, 04103 Leipzig, Germany.
Institute of Bioanalytical Chemistry, Center for Biotechnology and Biomedicine, University of Leipzig, 04103 Leipzig, Germany.
Institute of Biochemistry, Faculty of Biosciences, Pharmacy and Psychology, University of Leipzig, 04103 Leipzig, Germany. Electronic address: jheiker@uni-leipzig.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 838
Author: Oganesyan, V., Oganesyan, N., Terzyan, S., Qu, D., Dauter, Z., Esmon, N. L. and Esmon, C. T.
Year: 2002
Title: The crystal structure of the endothelial protein C receptor and a bound phospholipid
Journal: J Biol Chem
Volume: 277
Issue: 28
Pages: 24851-4
Epub Date: 2002/05/30
Date: Jul 12
Short Title: The crystal structure of the endothelial protein C receptor and a bound phospholipid
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.C200163200
Accession Number: 12034704
Keywords: Animals
*Blood Coagulation Factors
CHO Cells
Cricetinae
Crystallography, X-Ray
Models, Molecular
Mutagenesis
Phospholipids/*chemistry/metabolism
Protein C/*metabolism
Receptors, Cell Surface/*chemistry/genetics/metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Abstract: The endothelial cell protein C receptor (EPCR) shares approximately 20% sequence identity with the major histocompatibility complex class 1/CD1 family of molecules, accelerates the thrombin-thrombomodulin-dependent generation of activated protein C, a natural anticoagulant, binds to activated neutrophils, and can undergo translocation from the plasma membrane to the nucleus. Blocking protein C/activated protein C binding to the receptor inhibits not only protein C activation but the ability of the host to respond appropriately to bacterial challenge, exacerbating both the coagulant and inflammatory responses. To understand how EPCR accomplishes these multiple tasks, we solved the crystal structure of EPCR alone and in complex with the phospholipid binding domain of protein C. The structures were strikingly similar to CD1d. A tightly bound phospholipid resides in the groove typically involved in antigen presentation. The protein C binding site is outside this conserved groove and is distal from the membrane-spanning domain. Extraction of the lipid resulted in loss of protein C binding, which could be restored by lipid reconstitution. CD1d augments the immune response by presenting glycolipid antigens. The EPCR structure is a model for how CD1d binds lipids and further suggests additional potential functions for EPCR in immune regulation, possibly including the anti-phospholipid syndrome.
Notes: Oganesyan, Vaheh
Oganesyan, Natalia
Terzyan, Simon
Qu, Dongfeng
Dauter, Zbigniew
Esmon, Naomi L
Esmon, Charles T
P50 HL54502/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2002 Jul 12;277(28):24851-4. doi: 10.1074/jbc.C200163200. Epub 2002 May 28.
Author Address: Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City 73104, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 938
Author: Ogawa, H., Qiu, Y., Ogata, C. M. and Misono, K. S.
Year: 2004
Title: Crystal structure of hormone-bound atrial natriuretic peptide receptor extracellular domain: rotation mechanism for transmembrane signal transduction
Journal: J Biol Chem
Volume: 279
Issue: 27
Pages: 28625-31
Epub Date: 2004/05/01
Date: Jul 2
Short Title: Crystal structure of hormone-bound atrial natriuretic peptide receptor extracellular domain: rotation mechanism for transmembrane signal transduction
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M313222200
Accession Number: 15117952
Keywords: Animals
Atrial Natriuretic Factor/*chemistry
Binding Sites
Cricetinae
Crystallography, X-Ray
Dimerization
Glycosylation
Hormones/*chemistry
Kinetics
Ligands
Models, Molecular
Peptides
Protein Binding
Protein Conformation
Protein Structure, Quaternary
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Atrial Natriuretic Factor/*chemistry
*Signal Transduction
Abstract: A cardiac hormone, atrial natriuretic peptide (ANP), plays a major role in blood pressure and volume regulation. ANP activities are mediated by a single span transmembrane receptor carrying intrinsic guanylate cyclase activity. ANP binding to its extracellular domain stimulates guanylate cyclase activity by an as yet unknown mechanism. Here we report the crystal structure of dimerized extracellular hormone-binding domain in complex with ANP. The structural comparison with the unliganded receptor reveals that hormone binding causes the two receptor monomers to undergo an intermolecular twist with little intramolecular conformational change. This motion produces a Ferris wheel-like translocation of two juxtamembrane domains in the dimer with essentially no change in the interdomain distance. This movement alters the relative orientation of the two domains by a shift equivalent to counterclockwise rotation of each by 24 degrees. These results suggest that transmembrane signaling by the ANP receptor is initiated via a hormone-induced rotation mechanism.
Notes: Ogawa, Haruo
Qiu, Yue
Ogata, Craig M
Misono, Kunio S
HL 54329/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2004 Jul 2;279(27):28625-31. doi: 10.1074/jbc.M313222200. Epub 2004 Apr 26.
Author Address: Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH 44195, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 513
Author: Ogawa, H., Yoneda, A., Seno, N., Hayashi, M., Ishizuka, I., Hase, S. and Matsumoto, I.
Year: 1995
Title: Structures of the N-linked oligosaccharides on human plasma vitronectin
Journal: Eur J Biochem
Volume: 230
Issue: 3
Pages: 994-1000
Epub Date: 1995/06/15
Date: Jun 15
Short Title: Structures of the N-linked oligosaccharides on human plasma vitronectin
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1995.tb20647.x
Accession Number: 7541354
Keywords: Carbohydrate Sequence
Glycoproteins/blood/*chemistry/metabolism
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Vitronectin
Abstract: The structures of N-linked oligosaccharides present on human plasma vitronectin were elucidated. Oligosaccharides were released from the vitronectin by N-glycosidase F digestion and tagged with 2-aminopyridine; the pyridylamino-oligosaccharides were then fractionated by anion-exchange and reverse-phase HPLC. Ten major pyridylamino-oligosaccharides were isolated. The linkages and locations of sialic acid residues were determined by desialylation with Salmonella sialidase in combination with acid. The asialo forms were then analyzed by two-dimensional sugar mapping, component sugar analysis and 400-MHz 1H-NMR spectroscopy. The major oligosaccharides of human vitronectin were of the diantennary N-acetyllactosamine type, with a lesser amount of the tri- and a small amount of the mono-antennary type, to which 1-3 mol sialic acid residues were linked, mostly through alpha 2-6 linkages, although alpha 2-3 linkages were also present. The possibility that several binding activities of vitronectin can be ascribed to its glycan moiety was discussed, based on the specific features of the N-linked oligosaccharides on human vitronectin revealed here.
Notes: Ogawa, H
Yoneda, A
Seno, N
Hayashi, M
Ishizuka, I
Hase, S
Matsumoto, I
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1995 Jun 15;230(3):994-1000. doi: 10.1111/j.1432-1033.1995.tb20647.x.
Author Address: Department of Chemistry, Ochanomizu University, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 853
Author: Ogiso, H., Ishitani, R., Nureki, O., Fukai, S., Yamanaka, M., Kim, J. H., Saito, K., Sakamoto, A., Inoue, M., Shirouzu, M. and Yokoyama, S.
Year: 2002
Title: Crystal structure of the complex of human epidermal growth factor and receptor extracellular domains
Journal: Cell
Volume: 110
Issue: 6
Pages: 775-87
Epub Date: 2002/09/26
Date: Sep 20
Short Title: Crystal structure of the complex of human epidermal growth factor and receptor extracellular domains
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/s0092-8674(02)00963-7
Accession Number: 12297050
Keywords: Amino Acid Sequence
Amino Acids, Aromatic/chemistry
Animals
Binding Sites
CHO Cells
Cricetinae
Crystallization
Crystallography, X-Ray
Dimerization
Disulfides/chemistry
Epidermal Growth Factor/*chemistry/*metabolism
ErbB Receptors/*chemistry/*metabolism
Humans
Ligands
*Models, Molecular
Molecular Structure
Mutagenesis, Site-Directed
Phosphorylation
Protein Binding
Protein Structure, Tertiary
Selenium/chemistry
Sequence Analysis, DNA
Abstract: Epidermal growth factor (EGF) regulates cell proliferation and differentiation by binding to the EGF receptor (EGFR) extracellular region, comprising domains I-IV, with the resultant dimerization of the receptor tyrosine kinase. In this study, the crystal structure of a 2:2 complex of human EGF and the EGFR extracellular region has been determined at 3.3 A resolution. EGFR domains I-III are arranged in a C shape, and EGF is docked between domains I and III. The 1:1 EGF*EGFR complex dimerizes through a direct receptor*receptor interaction, in which a protruding beta-hairpin arm of each domain II holds the body of the other. The unique "receptor-mediated dimerization" was verified by EGFR mutagenesis.
Notes: Ogiso, Hideo
Ishitani, Ryuichiro
Nureki, Osamu
Fukai, Shuya
Yamanaka, Mari
Kim, Jae-Hoon
Saito, Kazuki
Sakamoto, Ayako
Inoue, Mio
Shirouzu, Mikako
Yokoyama, Shigeyuki
Journal Article
United States
Cell. 2002 Sep 20;110(6):775-87. doi: 10.1016/s0092-8674(02)00963-7.
Author Address: RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, 230-0045, Yokohama, Kanagawa, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 624
Author: Oh, Y., Nagalla, S. R., Yamanaka, Y., Kim, H. S., Wilson, E. and Rosenfeld, R. G.
Year: 1996
Title: Synthesis and characterization of insulin-like growth factor-binding protein (IGFBP)-7. Recombinant human mac25 protein specifically binds IGF-I and -II
Journal: J Biol Chem
Volume: 271
Issue: 48
Pages: 30322-5
Epub Date: 1996/11/29
Date: Nov 29
Short Title: Synthesis and characterization of insulin-like growth factor-binding protein (IGFBP)-7. Recombinant human mac25 protein specifically binds IGF-I and -II
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.48.30322
Accession Number: 8939990
Keywords: Gene Expression
Humans
Insulin-Like Growth Factor Binding Proteins/*biosynthesis/chemistry
Insulin-Like Growth Factor I/*metabolism
Insulin-Like Growth Factor II/*metabolism
Peptides/metabolism
RNA, Messenger/genetics
Recombinant Proteins
Tissue Distribution
Abstract: The mac25 cDNA was originally cloned from leptomeningial cells and subsequently reisolated through differential display as a sequence preferentially expressed in senescent human mammary epithelial cells. The deduced amino acid sequence of the human mac25 propeptide shares a 20-25% identity to human insulin-like growth factor-binding proteins (IGFBPs), suggesting that mac25 could be another member of the IGFBP family. In the present study, we have generated recombinant human mac25 (rh-mac25) in a baculovirus expression system and assessed its affinity for IGFs and have evaluated the pattern of expression of the mac25 gene in human tissues. Binding of 125I-IGF-I and 125I-IGF-II to rh-mac25 was demonstrated by Western ligand blotting after nondenaturing polyacrylamide gel electrophoresis and by affinity cross-linking with as little as 2 nM rh-mac25. Specificity of rh-mac25 binding to 125I-IGFs was demonstrated by competition for rh-mac25 binding with unlabeled IGFs, but not with [QAYLL]IGF-II analog, which has 100-fold less affinity for IGFBPs. In comparison with IGFBP-3, rh-mac25 has at least a 5-6-fold lower affinity for IGF-I and 20-25-fold lower affinity for IGF-II. mac25 mRNA was detectable in a wide range of normal human tissues, with decreased expression in breast, prostate, colon, and lung cancer cell lines. In conclusion, mac25 specifically binds IGFs and constitutes a new member of the IGFBP family, IGFBP-7. Its wider distribution in normal tissue and lower expression in several cancer cells indicate that IGFBP-7 may function as a growth-suppressing factor, as well as an IGF-binding protein.
Notes: Oh, Y
Nagalla, S R
Yamanaka, Y
Kim, H S
Wilson, E
Rosenfeld, R G
CA58110/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1996 Nov 29;271(48):30322-5. doi: 10.1074/jbc.271.48.30322.
Author Address: Department of Pediatrics, School of Medicine, Oregon Health Sciences University, Portland, Oregon 97201, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 905
Author: Ohe, Y., Ohnishi, H., Okazawa, H., Tomizawa, K., Kobayashi, H., Okawa, K. and Matozaki, T.
Year: 2003
Title: Characterization of nucleotide pyrophosphatase-5 as an oligomannosidic glycoprotein in rat brain
Journal: Biochem Biophys Res Commun
Volume: 308
Issue: 4
Pages: 719-25
Epub Date: 2003/08/21
Date: Sep 5
Short Title: Characterization of nucleotide pyrophosphatase-5 as an oligomannosidic glycoprotein in rat brain
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/s0006-291x(03)01454-2
Accession Number: 12927778
Keywords: Amino Acid Sequence
Animals
Blotting, Northern
Brain/metabolism
Cell Adhesion
Cell Communication
Cell Membrane/*metabolism
DNA, Complementary/metabolism
Detergents/pharmacology
Dimerization
Glutathione Transferase/metabolism
Glycoproteins/*chemistry
Glycosylation
Immunoblotting
Mass Spectrometry
Mice
Molecular Sequence Data
Neurons/metabolism
Octoxynol/pharmacology
Peptides
Phosphoric Diester Hydrolases/chemistry
Precipitin Tests
Protein Structure, Tertiary
Pyrophosphatases/*chemistry/*physiology
Rats
Recombinant Fusion Proteins/metabolism
Silver Staining
Sodium-Potassium-Exchanging ATPase/chemistry
Tissue Distribution
Abstract: Membrane glycoproteins of neural cells play crucial roles in axon guidance, synaptogenesis, and neuronal transmission. We have here characterized membrane glycoproteins containing terminal alpha-mannose residues in rat brain membranes. Affinity purification using Galanthus nivalis agglutinin, that is highly specific for terminal alpha-mannose residues, revealed a 50-kDa protein as well as 80-kDa SHPS-1 and 45-kDa beta2 subunit of Na,K-ATPase in rat brain membranes. Combination of N-terminal peptide sequencing and mass spectrometry indicated that the 50-kDa protein was rat nucleotide pyrophosphatase-5 (NPP-5). In contrast to other NPPs, NPP-5 was a type-I transmembrane protein. Northern blot analysis showed that NPP-5 was highly expressed in brain, but also expressed in other peripheral tissues. However, we could not detect either the NPP activity or the lysophospholipase D activity in the immunoprecipitates with antibodies to NPP-5 from rat brain membranes. These data, therefore, suggest that NPP-5 is a neural oligomannosidic glycoprotein that may participate in neural cell communications.
Notes: Ohe, Yoshihide
Ohnishi, Hiroshi
Okazawa, Hideki
Tomizawa, Kyoko
Kobayashi, Hisae
Okawa, Katsuya
Matozaki, Takashi
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2003 Sep 5;308(4):719-25. doi: 10.1016/s0006-291x(03)01454-2.
Author Address: Biosignal Research Center, Institute for Molecular and Cellular Regulation, Gunma University, 3-39-15 Showa-Machi, Maebashi, Gunma 371-8512, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1448
Author: Ohgomori, T., Nanao, T., Morita, A. and Ikekita, M.
Year: 2012
Title: Asn54-linked glycan is critical for functional folding of intercellular adhesion molecule-5
Journal: Glycoconj J
Volume: 29
Issue: 1
Pages: 47-55
Epub Date: 2011/12/22
Date: Jan
Short Title: Asn54-linked glycan is critical for functional folding of intercellular adhesion molecule-5
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080
DOI: 10.1007/s10719-011-9363-0
Accession Number: 22187327
Keywords: Animals
Asparagine/chemistry/*metabolism
Cell Adhesion Molecules/chemistry/genetics/*metabolism
Cell Line, Tumor
Endoplasmic Reticulum/metabolism
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Immunohistochemistry
Mannose/metabolism
Mice
Microscopy, Fluorescence
Mutagenesis, Site-Directed/methods
Nerve Tissue Proteins/chemistry/genetics/*metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
Polysaccharides/chemistry/*metabolism
*Protein Folding
Protein Processing, Post-Translational
Protein Transport
Pseudopodia/chemistry
Rats
Transfection/methods
Abstract: Intercellular adhesion molecule-5 (ICAM-5, telencephalin) is a dendritically polarized type I membrane glycoprotein, and promotes dendritic filopodia formation. Although we have determined the N-glycan structures of ICAM-5 in a previous report, their function is unknown. Here, we produced fifteen ICAM-5 gene constructs, in which each potential N-glycosylation site was mutated, to elucidate the function of the N-glycans of ICAM-5, and observed the effects of transfection of them on a neuronal cell line, Neuro-2a (N2a). Only the N54Q mutant, which is the mutant for the most N-terminal glycosylation site, failed to induce filopodia-like protrusions in N2a cells. Immunofluorescence staining and cell surface biotinylation revealed that N54Q ICAM-5 was confined to the ER and also could not be expressed on the cell surface. This is further supported by the biochemical evidence that almost all N-glycans of N54Q ICAM-5 were digested by Endo glycosidase H and peptide:N-glycanase, indicating that almost all of them retain high-mannose-type structures in ER. In additon, it also failed to form disulfide bonds or functional protein complexes. The stable transformants of N54Q ICAM-5 showed retarded cell growth, but it was interesting that there was no apparent ER stress, because the mutant was sequentially degraded via ER associated degradation pathway by comparing the susceptibilities of the responses to various inhibitors of this pathway in wild-type and N54Q ICAM-5 transfectants. Taken together, the Asn(54)-linked glycan is necessary for normal trafficking and function of ICAM-5, but is unassociated with ER-associated degradation of it.
Notes: 1573-4986
Ohgomori, Tomohiro
Nanao, Tomohisa
Morita, Akinori
Ikekita, Masahiko
Journal Article
United States
Glycoconj J. 2012 Jan;29(1):47-55. doi: 10.1007/s10719-011-9363-0. Epub 2011 Dec 21.
Author Address: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Yamazaki, Noda, Chiba, Japan. ohgomori@med.nagoya-u.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 769
Author: Ohki, R., Nemoto, J., Murasawa, H., Oda, E., Inazawa, J., Tanaka, N. and Taniguchi, T.
Year: 2000
Title: Reprimo, a new candidate mediator of the p53-mediated cell cycle arrest at the G2 phase
Journal: J Biol Chem
Volume: 275
Issue: 30
Pages: 22627-30
Epub Date: 2000/08/10
Date: Jul 28
Short Title: Reprimo, a new candidate mediator of the p53-mediated cell cycle arrest at the G2 phase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.C000235200
Accession Number: 10930422
Keywords: Amino Acid Sequence
Animals
CDC2 Protein Kinase/metabolism
Cell Cycle/*physiology
Cell Cycle Proteins/chemistry/*genetics
Cell Line
Cyclin B/metabolism
Cyclin B1
DNA, Complementary
*G2 Phase
*Genes, Suppressor
Glycoproteins/chemistry/*genetics
Humans
Molecular Sequence Data
Sequence Homology, Amino Acid
Tumor Suppressor Protein p53/*physiology
Abstract: A novel gene, Reprimo, in which induction in cells exposed to X-irradiation is dependent on p53 expression, has been isolated. Ectopic p53 expression results in the induction of its mRNA. Reprimo is a highly glycosylated protein and, when ectopically expressed, it is localized in the cytoplasm and induces G(2) arrest of the cell cycle. In the arrested cells, both Cdc2 activity and nuclear translocation of cyclin B1 are inhibited, suggesting the involvement of Reprimo in the Cdc2.cyclin B1 regulation pathway. Thus, Reprimo may be a new member involved in the regulation of p53-dependent G(2) arrest of the cell cycle.
Notes: Ohki, R
Nemoto, J
Murasawa, H
Oda, E
Inazawa, J
Tanaka, N
Taniguchi, T
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2000 Jul 28;275(30):22627-30. doi: 10.1074/jbc.C000235200.
Author Address: Department of Immunology, Graduate School of Medicine and Faculty of Medicine, University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 813
Author: Ohnishi, T., Muroi, M. and Tanamoto, K.
Year: 2001
Title: N-linked glycosylations at Asn(26) and Asn(114) of human MD-2 are required for toll-like receptor 4-mediated activation of NF-kappaB by lipopolysaccharide
Journal: J Immunol
Volume: 167
Issue: 6
Pages: 3354-9
Epub Date: 2001/09/07
Date: Sep 15
Short Title: N-linked glycosylations at Asn(26) and Asn(114) of human MD-2 are required for toll-like receptor 4-mediated activation of NF-kappaB by lipopolysaccharide
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
DOI: 10.4049/jimmunol.167.6.3354
Accession Number: 11544325
Keywords: *Acute-Phase Proteins
Antigens, Surface/*chemistry/physiology
Asparagine/*chemistry
Carrier Proteins/genetics/physiology
Cell Line
Cell Membrane/metabolism
*Drosophila Proteins
Electrophoresis, Polyacrylamide Gel
Gene Expression Regulation/drug effects
Genes, Reporter
Glycosylation
Humans
Kidney
Lipopolysaccharides/*pharmacology
Luciferases/analysis/genetics
Lymphocyte Antigen 96
Membrane Glycoproteins/*physiology
Mutagenesis, Site-Directed
NF-kappa B/*metabolism
Protein Binding
Protein Processing, Post-Translational
Protein Transport
Receptors, Cell Surface/*physiology
Recombinant Fusion Proteins/chemistry/physiology
Signal Transduction
Structure-Activity Relationship
Toll-Like Receptor 4
Toll-Like Receptors
Transfection
Abstract: MD-2 is physically associated with Toll-like receptor 4 (TLR4) and is required for TLR4-mediated LPS signaling. Western blotting analysis revealed the presence of three forms of human (h)MD-2 with different electrophoretic mobilities. After N-glycosidase treatment of the cellular extract prepared from cells expressing hMD-2, only a single form with the fastest mobility was detected. Mutation of either one of two potential glycosylation sites (Asn(26) and Asn(114)) of MD-2 resulted in the disappearance of the slowest mobility form, and only the fastest form was detected in hMD-2 carrying mutations at both Asn(26) and Asn(114). Although these mutants were expressed on the cell surface and maintained its ability to associate with human TLR4, these mutations or tunicamycin treatment substantially impaired the ability of MD-2 to complement TLR4-mediated activation of NF-kappaB by LPS. LPS binding to cells expressing CD14, TLR4, and MD-2 was unaffected by these mutations. These observations demonstrate that hMD-2 undergoes N-linked glycosylation at Asn(26) and Asn(114), and that these glycosylations are crucial for TLR4-mediated signal transduction of LPS.
Notes: Ohnishi, T
Muroi, M
Tanamoto, K
Journal Article
United States
J Immunol. 2001 Sep 15;167(6):3354-9. doi: 10.4049/jimmunol.167.6.3354.
Author Address: Division of Microbiology, National Institute of Health Sciences, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 739
Author: Ohta, M., Ohnishi, T., Ioannou, Y. A., Hodgson, M. E., Matsuura, F. and Desnick, R. J.
Year: 2000
Title: Human alpha-N-acetylgalactosaminidase: site occupancy and structure of N-linked oligosaccharides
Journal: Glycobiology
Volume: 10
Issue: 3
Pages: 251-61
Epub Date: 2000/03/08
Date: Mar
Short Title: Human alpha-N-acetylgalactosaminidase: site occupancy and structure of N-linked oligosaccharides
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/10.3.251
Accession Number: 10704524
Keywords: Animals
COS Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Hexosaminidases/*chemistry/genetics/metabolism
Humans
Molecular Sequence Data
Mutagenesis, Site-Directed
Oligosaccharides/*chemistry/metabolism
alpha-N-Acetylgalactosaminidase
Abstract: Human alpha-N-acetylgalactosaminidase (alpha-GalNAc; also known as alpha-galactosidase B) is the lysosomal exoglycohydrolase that cleaves alpha-N-acetylgalactosaminyl moieties in glycoconjugates. Mutagenesis studies indicated that the first five (N124, N177, N201, N359, and N385) of the six potential N-glycosylation sites were occupied. Site 3 occupancy was important for enzyme function and stability. Characterization of the N-linked oligosaccharide structures on the secreted enzyme overexpressed in Chinese hamster ovary cells revealed highly heterogeneous structures consisting of complex (approximately 53%), hybrid (approximately 12%), and high mannose-type (approximately 33%) oligosaccharides. The complex structures were mono-, bi-, 2,4-tri-, 2,6-tri-, and tetraantennary, among which the biantennary structures were most predominant (approximately 53%). Approximately 80% of the complex oligo-saccharides had a core-region fucose and 50% of the complex oligosaccharides were sialylated exclusively with alpha-2,3-linked sialic acid residues. The majority of hybrid type oligo-saccharides were GalGlcNAcMan(6)GlcNAc-Fuc(0-1)GlcNAc. Approximately 54% of the hybrid oligosaccharide were phosphorylated and one-third of these structures were further sialylated, the latter representing unique phosphorylated and sialylated structures. Of the high mannose oligosaccharides, Man(5-7)GlcNAc(2) were the predominant species (approximately 90%) and about 50% of the high mannose oligosaccharides were phosphorylated, exclusively as monoesters whose positions were determined. Comparison of the oligosaccharide structures of alpha-GalNAc and alpha-galactosidase A, an evolutionary-related and highly homologous exoglycosidase, indicated that alpha-GalNAc had more completed complex chains, presumably due to differences in enzyme structure/domains, rate of biosynthesis, and/or aggregation of the overexpressed recombinant enzymes.
Notes: Ohta, M
Ohnishi, T
Ioannou, Y A
Hodgson, M E
Matsuura, F
Desnick, R J
5 R01 DK34045/DK/NIDDK NIH HHS/United States
M01 RR00071/RR/NCRR NIH HHS/United States
P30 HD28822/HD/NICHD NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 2000 Mar;10(3):251-61. doi: 10.1093/glycob/10.3.251.
Author Address: Department of Biotechnology, Fukuyama University, Fukuyama, Hiroshima 729-0292, Japan
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1230
Author: Ohta, T., Ikemoto, Y., Usami, A., Koide, T. and Wakabayashi, S.
Year: 2009
Title: High affinity interaction between histidine-rich glycoprotein and the cell surface type ATP synthase on T-cells
Journal: Biochim Biophys Acta
Volume: 1788
Issue: 5
Pages: 1099-107
Epub Date: 2009/03/17
Date: May
Short Title: High affinity interaction between histidine-rich glycoprotein and the cell surface type ATP synthase on T-cells
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbamem.2009.03.005
Accession Number: 19285951
Keywords: Amino Acid Sequence
Biophysical Phenomena
Cell Line
Cell Membrane/metabolism
Chromatography, Affinity
Humans
Mitochondria/enzymology
Molecular Sequence Data
Peptide Fragments/chemistry/genetics/metabolism
Peptide Mapping
Protein Binding
Protein Subunits
Proteins/chemistry/genetics/*metabolism
Proton-Translocating ATPases/chemistry/genetics/*metabolism
T-Lymphocytes/*metabolism
Abstract: Histidine-rich glycoprotein (HRG) is a plasma protein implicated in the innate immune system. In recent studies, we showed that either HRG, or the Arg23-Lys66 glycopeptide derived from HRG, in concert with concanavalin A (Con A), promotes a morphological change and adhesion of the human leukemic T-cell line MOLT-4 to culture dishes, and that cell surface glycosaminoglycan or Fcgamma receptors do not participate in this cellular event. In the present study, we identified the alpha-subunit of ATP synthase as one of the HRG-binding proteins on the surface of T-cells by HRG-derived glycopeptide affinity chromatography and by a peptide mass finger printing method. HRG specifically interacted with mitochondrial ATP synthase with a dissociation constant of 66 nM. The presence of alpha- and beta-subunits of ATP synthase on the plasma membrane of MOLT-4 cell was demonstrated by immunofluorescent staining and FACS analysis. The HRG/Con A-induced morphological changes of MOLT-4 cells were specifically inhibited by a monoclonal antibody against the beta-subunit of ATP synthase. These results strongly suggest that the cell surface ATP synthase functions as a binding protein for HRG on MOLT-4 cells, which is required for the morphological changes observed in MOLT-4 cells following treatment with HRG/Con A.
Notes: Ohta, Takeshi
Ikemoto, Yoshitaka
Usami, Ayako
Koide, Takehiko
Wakabayashi, Sadao
Journal Article
Netherlands
Biochim Biophys Acta. 2009 May;1788(5):1099-107. doi: 10.1016/j.bbamem.2009.03.005. Epub 2009 Mar 12.
Author Address: Department of Life Science, Graduate School of Life Science, University of Hyogo, Harima Science Garden City, Hyogo 678-1297, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1114
Author: Ohto, U., Fukase, K., Miyake, K. and Satow, Y.
Year: 2007
Title: Crystal structures of human MD-2 and its complex with antiendotoxic lipid IVa
Journal: Science
Volume: 316
Issue: 5831
Pages: 1632-4
Epub Date: 2007/06/16
Date: Jun 15
Short Title: Crystal structures of human MD-2 and its complex with antiendotoxic lipid IVa
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1139111
Accession Number: 17569869
Keywords: Binding Sites
Crystallography, X-Ray
Fatty Acids/chemistry
Glycolipids/*chemistry/metabolism
Glycosylation
Humans
Hydrophobic and Hydrophilic Interactions
Ligands
Lipid A/*analogs & derivatives/chemistry/metabolism
Lymphocyte Antigen 96/*chemistry/metabolism
Models, Molecular
Phosphorylation
Protein Conformation
Protein Folding
Protein Structure, Secondary
Abstract: Endotoxic lipopolysaccharide (LPS) with potent immunostimulatory activity is recognized by the receptor complex of MD-2 and Toll-like receptor 4. Crystal structures of human MD-2 and its complex with the antiendotoxic tetra-acylated lipid A core of LPS have been determined at 2.0 and 2.2 angstrom resolutions, respectively. MD-2 shows a deep hydrophobic cavity sandwiched by two beta sheets, in which four acyl chains of the ligand are fully confined. The phosphorylated glucosamine moieties are located at the entrance to the cavity. These structures suggest that MD-2 plays a principal role in endotoxin recognition and provide a basis for antiseptic drug development.
Notes: 1095-9203
Ohto, Umeharu
Fukase, Koichi
Miyake, Kensuke
Satow, Yoshinori
Journal Article
Research Support, Non-U.S. Gov't
United States
Science. 2007 Jun 15;316(5831):1632-4. doi: 10.1126/science.1139111.
Author Address: Graduate School of Pharmaceutical Sciences, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1430
Author: Ohto, U., Miyake, K. and Shimizu, T.
Year: 2011
Title: Crystal structures of mouse and human RP105/MD-1 complexes reveal unique dimer organization of the toll-like receptor family
Journal: J Mol Biol
Volume: 413
Issue: 4
Pages: 815-25
Epub Date: 2011/10/01
Date: Nov 4
Short Title: Crystal structures of mouse and human RP105/MD-1 complexes reveal unique dimer organization of the toll-like receptor family
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2011.09.020
Accession Number: 21959264
Keywords: Animals
Antigens, CD/*chemistry/metabolism
Antigens, Surface/*chemistry/metabolism
Crystallography, X-Ray
Humans
Membrane Glycoproteins/*chemistry/metabolism
Mice
Models, Molecular
Protein Conformation
*Protein Multimerization
Toll-Like Receptors/*chemistry/metabolism
Abstract: The Toll-like receptor (TLR) 4/MD-2 heterodimer senses lipopolysaccharide (LPS). RP105 (radioprotective 105 kDa), a TLR-related molecule, is similar to TLR4 in that the extracellular leucine-rich repeats associate with MD-1, the MD-2-like molecule. MD-2 has a unique hydrophobic cavity that directly binds to lipid A, the active center of LPS. LPS-bound MD-2 opens the secondary interface with TLR4, leading to dimerization of TLR4/MD-2. MD-1 also has a hydrophobic cavity that accommodates lipid IVa, a precursor of lipid A, suggesting a role for the RP105/MD-1 heterodimer in sensing LPS or related microbial products. Little is known, however, about the structure of the RP105/MD-1 heterodimer or its oligomer. Here, we have determined the crystal structures of mouse and human RP105/MD-1 complexes at 1.9 and 2.8 Å resolutions, respectively. Both mouse and human RP105/MD-1 exhibit dimerization of the 1:1 RP105/MD-1 complex, demonstrating a novel organization. The "m"-shaped 2:2 RP105/MD-1 complex exhibits an inverse arrangement, with N-termini interacting in the middle. Thus, the dimerization interface of RP105/MD-1 is located on the opposite side of the complex, compared to the 2:2 TLR4/MD-2 complex. These results demonstrate that the 2:2 RP105/MD-1 complex is distinct from previously reported TLR dimers, including TLR4/MD-2, TLR1/TLR2, TLR2/TLR6, and TLR3, all of which facilitate homotypic or heterotypic interaction of the C-terminal cytoplasmic signaling domain.
Notes: 1089-8638
Ohto, Umeharu
Miyake, Kensuke
Shimizu, Toshiyuki
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2011 Nov 4;413(4):815-25. doi: 10.1016/j.jmb.2011.09.020. Epub 2011 Sep 19.
Author Address: Graduate School of Pharmaceutical Sciences, University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1683
Author: Ohto, U., Shibata, T., Tanji, H., Ishida, H., Krayukhina, E., Uchiyama, S., Miyake, K. and Shimizu, T.
Year: 2015
Title: Structural basis of CpG and inhibitory DNA recognition by Toll-like receptor 9
Journal: Nature
Volume: 520
Issue: 7549
Pages: 702-5
Epub Date: 2015/02/18
Date: Apr 30
Short Title: Structural basis of CpG and inhibitory DNA recognition by Toll-like receptor 9
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature14138
Accession Number: 25686612
Keywords: Animals
Base Sequence
CpG Islands/*immunology
Crystallography, X-Ray
DNA/*chemistry/genetics/*immunology/metabolism
Humans
Ligands
Models, Molecular
Nucleic Acid Conformation
Protein Structure, Tertiary
Structure-Activity Relationship
Toll-Like Receptor 9/agonists/antagonists & inhibitors/*chemistry/*immunology
Abstract: Innate immunity serves as the first line of defence against invading pathogens such as bacteria and viruses. Toll-like receptors (TLRs) are examples of innate immune receptors, which sense specific molecular patterns from pathogens and activate immune responses. TLR9 recognizes bacterial and viral DNA containing the cytosine-phosphate-guanine (CpG) dideoxynucleotide motif. The molecular basis by which CpG-containing DNA (CpG-DNA) elicits immunostimulatory activity via TLR9 remains to be elucidated. Here we show the crystal structures of three forms of TLR9: unliganded, bound to agonistic CpG-DNA, and bound to inhibitory DNA (iDNA). Agonistic-CpG-DNA-bound TLR9 formed a symmetric TLR9-CpG-DNA complex with 2:2 stoichiometry, whereas iDNA-bound TLR9 was a monomer. CpG-DNA was recognized by both protomers in the dimer, in particular by the amino-terminal fragment (LRRNT-LRR10) from one protomer and the carboxy-terminal fragment (LRR20-LRR22) from the other. The iDNA, which formed a stem-loop structure suitable for binding by intramolecular base pairing, bound to the concave surface from LRR2-LRR10. This structure serves as an important basis for improving our understanding of the functional mechanisms of TLR9.
Notes: 1476-4687
Ohto, Umeharu
Shibata, Takuma
Tanji, Hiromi
Ishida, Hanako
Krayukhina, Elena
Uchiyama, Susumu
Miyake, Kensuke
Shimizu, Toshiyuki
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2015 Apr 30;520(7549):702-5. doi: 10.1038/nature14138. Epub 2015 Feb 9.
Author Address: Graduate School of Pharmaceutical Sciences, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
1] Division of Innate Immunity, Department of Microbiology and Immunology, Laboratory of Innate Immunity, Center for Experimental Medicine and Systems Biology, The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan [2] Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Saitama 332-0012, Japan.
1] Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan [2] U-Medico Corporation, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Division of Innate Immunity, Department of Microbiology and Immunology, Laboratory of Innate Immunity, Center for Experimental Medicine and Systems Biology, The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
1] Graduate School of Pharmaceutical Sciences, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan [2] Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Saitama 332-0012, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1443
Author: Ohto, U., Usui, K., Ochi, T., Yuki, K., Satow, Y. and Shimizu, T.
Year: 2012
Title: Crystal structure of human β-galactosidase: structural basis of Gm1 gangliosidosis and morquio B diseases
Journal: J Biol Chem
Volume: 287
Issue: 3
Pages: 1801-12
Epub Date: 2011/12/01
Date: Jan 13
Short Title: Crystal structure of human β-galactosidase: structural basis of Gm1 gangliosidosis and morquio B diseases
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.293795
PMCID: PMC3265862
Accession Number: 22128166
Keywords: 1-Deoxynojirimycin/*analogs & derivatives/chemistry
Crystallography, X-Ray
G(M1) Ganglioside/genetics/metabolism
Galactose/*chemistry/genetics/metabolism
Gangliosidosis, GM1/*enzymology/genetics
Humans
Hydrolysis
Keratan Sulfate/genetics/metabolism
Lysosomes/*enzymology/genetics
Models, Molecular
Mucopolysaccharidosis IV/*enzymology/genetics
Mutation
Protein Structure, Tertiary
beta-Galactosidase/*chemistry/genetics/metabolism
Abstract: G(M1) gangliosidosis and Morquio B are autosomal recessive lysosomal storage diseases associated with a neurodegenerative disorder or dwarfism and skeletal abnormalities, respectively. These diseases are caused by deficiencies in the lysosomal enzyme β-d-galactosidase (β-Gal), which lead to accumulations of the β-Gal substrates, G(M1) ganglioside, and keratan sulfate. β-Gal is an exoglycosidase that catalyzes the hydrolysis of terminal β-linked galactose residues. This study shows the crystal structures of human β-Gal in complex with its catalytic product galactose or with its inhibitor 1-deoxygalactonojirimycin. Human β-Gal is composed of a catalytic TIM barrel domain followed by β-domain 1 and β-domain 2. To gain structural insight into the molecular defects of β-Gal in the above diseases, the disease-causing mutations were mapped onto the three-dimensional structure. Finally, the possible causes of the diseases are discussed.
Notes: 1083-351x
Ohto, Umeharu
Usui, Kimihito
Ochi, Toshinari
Yuki, Kenjiro
Satow, Yoshinori
Shimizu, Toshiyuki
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Jan 13;287(3):1801-12. doi: 10.1074/jbc.M111.293795. Epub 2011 Nov 28.
Author Address: From the Graduate School of Pharmaceutical Sciences, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1023
Author: Ohtsubo, K., Takamatsu, S., Minowa, M. T., Yoshida, A., Takeuchi, M. and Marth, J. D.
Year: 2005
Title: Dietary and genetic control of glucose transporter 2 glycosylation promotes insulin secretion in suppressing diabetes
Journal: Cell
Volume: 123
Issue: 7
Pages: 1307-21
Epub Date: 2005/12/27
Date: Dec 29
Short Title: Dietary and genetic control of glucose transporter 2 glycosylation promotes insulin secretion in suppressing diabetes
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2005.09.041
Accession Number: 16377570
Keywords: Animals
Cells, Cultured
Diabetes Mellitus, Type 2/*enzymology/genetics/*prevention & control
*Diet
Dietary Fats/administration & dosage/pharmacology
Glucose Transporter Type 2/genetics/*metabolism
Glycosylation
Insulin/*biosynthesis/metabolism
Insulin Secretion
Insulin-Secreting Cells/chemistry
Mice
Mice, Inbred C57BL
Mice, Transgenic
N-Acetylglucosaminyltransferases/deficiency/genetics/*metabolism
Up-Regulation
Abstract: Pancreatic beta cell-surface expression of glucose transporter 2 (Glut-2) is essential for glucose-stimulated insulin secretion, thereby controlling blood glucose homeostasis in response to dietary intake. We show that the murine GlcNAcT-IVa glycosyltransferase is required for Glut-2 residency on the beta cell surface by constructing a cell-type- and glycoprotein-specific N-glycan ligand for pancreatic lectin receptors. Loss of GlcNAcT-IVa, or the addition of glycan-ligand mimetics, attenuates Glut-2 cell-surface half-life, provoking endocytosis with redistribution into endosomes and lysosomes. The ensuing impairment of glucose-stimulated insulin secretion leads to metabolic dysfunction diagnostic of type 2 diabetes. Remarkably, the induction of diabetes by chronic ingestion of a high-fat diet is associated with reduced GlcNAcT-IV expression and attenuated Glut-2 glycosylation coincident with Glut-2 endocytosis. We infer that beta cell glucose-transporter glycosylation mediates a link between diet and insulin production that typically suppresses the pathogenesis of type 2 diabetes.
Notes: Ohtsubo, Kazuaki
Takamatsu, Shinji
Minowa, Mari T
Yoshida, Aruto
Takeuchi, Makoto
Marth, Jamey D
DK48247/DK/NIDDK NIH HHS/United States
GM62116/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Cell. 2005 Dec 29;123(7):1307-21. doi: 10.1016/j.cell.2005.09.041.
Author Address: Howard Hughes Medical Institute and Department of Cellular and Molecular Medicine, 9500 Gilman Drive, University of California, San Diego, La Jolla, CA 92093, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2056
Author: Ohyama, Y., Yamaguchi, H., Nakajima, K., Mizuno, T., Fukamachi, Y., Yokoi, Y., Tsuboi, N., Inaguma, D., Hasegawa, M., Renfrow, M. B., Novak, J., Yuzawa, Y. and Takahashi, K.
Year: 2020
Title: Analysis of O-glycoforms of the IgA1 hinge region by sequential deglycosylation
Journal: Sci Rep
Volume: 10
Issue: 1
Pages: 671
Epub Date: 2020/01/22
Date: Jan 20
Short Title: Analysis of O-glycoforms of the IgA1 hinge region by sequential deglycosylation
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/s41598-020-57510-z
PMCID: PMC6971281
Accession Number: 31959827
Keywords: Glomerulonephritis, IGA/*etiology
Glycomics/*methods
Glycopeptides/*chemistry
Glycosylation
High-Throughput Screening Assays/*methods
Humans
Immunoglobulin A/*chemistry/*metabolism
Polysaccharides/*chemistry
Protein Isoforms
Tandem Mass Spectrometry
Abstract: A common renal disease, immunoglobulin A (IgA) nephropathy (IgAN), is associated with glomerular deposition of IgA1-containing immune complexes. IgA1 hinge region (HR) has up to six clustered O-glycans consisting of Ser/Thr-linked N-acetylgalactosamine with β1,3-linked galactose and variable sialylation. IgA1 glycoforms with some galactose-deficient (Gd) HR O-glycans play a key role in IgAN pathogenesis. The clustered and variable O-glycans make the IgA1 glycomic analysis challenging and better approaches are needed. Here, we report a comprehensive analytical workflow for IgA1 HR O-glycoform analysis. We combined an automated quantitative analysis of the HR O-glycopeptide profiles with sequential deglycosylation to remove all but Gd O-glycans from the HR. The workflow was tested using serum IgA1 from healthy subjects. Twelve variants of glycopeptides corresponding to the HR with three to six O-glycans were detected; nine glycopeptides carried up to three Gd O-glycans. Sites with Gd O-glycans were unambiguously identified by electron-transfer/higher-energy collision dissociation tandem mass spectrometry. Extracted ion chromatograms of isomeric glycoforms enabled quantitative assignment of Gd sites. The most frequent Gd site was T(236), followed by S(230), T(233), T(228), and S(232). The new workflow for quantitative profiling of IgA1 HR O-glycoforms with site-specific resolution will enable identification of pathogenic IgA1 HR O-glycoforms in IgAN.
Notes: 2045-2322
Ohyama, Yukako
Yamaguchi, Hisateru
Nakajima, Kazuki
Mizuno, Tomohiro
Fukamachi, Yukihiro
Yokoi, Yasuto
Tsuboi, Naotake
Inaguma, Daijo
Orcid: 0000-0002-3977-5933
Hasegawa, Midori
Renfrow, Matthew B
Orcid: 0000-0002-4193-7305
Novak, Jan
Orcid: 0000-0002-9211-6670
Yuzawa, Yukio
Takahashi, Kazuo
R01 GM098539/GM/NIGMS NIH HHS/United States
R01 DK078244/DK/NIDDK NIH HHS/United States
R01 DK082753/DK/NIDDK NIH HHS/United States
R56 DK078244/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Sci Rep. 2020 Jan 20;10(1):671. doi: 10.1038/s41598-020-57510-z.
Author Address: Department of Nephrology, Fujita Health University School of Medicine, Toyoake, Japan.
Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Japan.
Center for Research Promotion and Support, Fujita Health University, Toyoake, Japan.
Analytical Pharmacology, Faculty of Pharmacy, Meijo University, Nagoya, Japan.
Mitsui Knowledge Industry, Tokyo, Japan.
Departments of Biochemistry and Molecular Genetics and Microbiology, University of Alabama at Birmingham, Birmingham, AL, USA.
Department of Nephrology, Fujita Health University School of Medicine, Toyoake, Japan. kazuot@fujita-hu.ac.jp.
Departments of Biochemistry and Molecular Genetics and Microbiology, University of Alabama at Birmingham, Birmingham, AL, USA. kazuot@fujita-hu.ac.jp.
Department of Biomedical Molecular Sciences, Fujita Health University School of Medicine, Toyoake, Japan. kazuot@fujita-hu.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 619
Author: Oinonen, C., Tikkanen, R., Rouvinen, J. and Peltonen, L.
Year: 1995
Title: Three-dimensional structure of human lysosomal aspartylglucosaminidase
Journal: Nat Struct Biol
Volume: 2
Issue: 12
Pages: 1102-8
Epub Date: 1995/12/01
Date: Dec
Short Title: Three-dimensional structure of human lysosomal aspartylglucosaminidase
Alternate Journal: Nature structural biology
ISSN: 1072-8368 (Print)
1072-8368
DOI: 10.1038/nsb1295-1102
Accession Number: 8846222
Keywords: Aspartylglucosylaminase/*chemistry/ultrastructure
Humans
Ligands
Lysosomes/chemistry/*enzymology
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Abstract: The high resolution crystal structure of human lysosomal aspartylglucosaminidase (AGA) has been determined. This lysosomal enzyme is synthesized as a single polypeptide precursor, which is immediately post-translationally cleaved into alpha- and beta-subunits. Two alpha- and beta-chains are found to pack together forming the final heterotetrameric structure. The catalytically essential residue, the N-terminal threonine of the beta-chain is situated in the deep pocket of the funnel-shaped active site. On the basis of the structure of the enzyme-product complex we present a catalytic mechanism for this lysosomal enzyme with an exceptionally high pH optimum. The three-dimensional structure also allows the prediction of the structural consequences of human mutations resulting in aspartylglucosaminuria (AGU), a lysosomal storage disease.
Notes: Oinonen, C
Tikkanen, R
Rouvinen, J
Peltonen, L
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Biol. 1995 Dec;2(12):1102-8. doi: 10.1038/nsb1295-1102.
Author Address: Department of Chemistry, University of Joensuu, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1827
Author: Okuda, T.
Year: 2017
Title: Western blot data using two distinct anti-O-GlcNAc monoclonal antibodies showing unique glycosylation status on cellular proteins under 2-deoxy-d-glucose treatment
Journal: Data Brief
Volume: 10
Pages: 449-453
Epub Date: 2017/01/06
Date: Feb
Short Title: Western blot data using two distinct anti-O-GlcNAc monoclonal antibodies showing unique glycosylation status on cellular proteins under 2-deoxy-d-glucose treatment
Alternate Journal: Data in brief
ISSN: 2352-3409 (Print)
2352-3409
DOI: 10.1016/j.dib.2016.12.001
PMCID: PMC5198851
Accession Number: 28054006
Keywords: 2-deoxy-d-glucose
O-GlcNAcylation
Sp1
Teratocarcinoma
Abstract: Protein modification by O-linked N-acetylglucosamine (O-GlcNAcylation) is one of the post transcriptional modifications occurring on cellular proteins. This paper provides a data set relating to the O-GlcNAcylation of cellular proteins detected by RL2 and CTD110.6 antibodies, which are commonly used for detection of protein O-GlcNAcylation, in 2-deoxy-d-glucose (2DG)-treated human teratocarcinoma NCCIT cells in support of the research article entitled "A novel, promoter-based, target-specific assay identifies 2-deoxy-d-glucose as an inhibitor of globotriaosylceramide biosynthesis" (Okuda et al., 2009) [1]. The main article described a suppressive effect of 2DG on an Sp1 target gene in NCCIT cells and discussed the relationship between the effect of 2DG and O-GlcNAcylation status of Sp1. The data in this paper complements this relationship by Western blotting and clearly showed that the 2DG treatment increased O-GlcNAcylation of cellular proteins in NCCIT cells, whereas the RL2 and CTD110.6 epitopes were detected in a different manner. The RL2 epitope was detected on Sp1 during 2DG treatment, and the level was transiently increased at 24 h. In contrast, the CTD110.6 epitope became detectable on Sp1 over 72 h after 2DG treatment, and then the other proteins containing CTD110.6 epitopes also appeared in the cell lysates and the anti-Sp1 antibody precipitates.
Notes: 2352-3409
Okuda, Tetsuya
Journal Article
Data Brief. 2016 Dec 9;10:449-453. doi: 10.1016/j.dib.2016.12.001. eCollection 2017 Feb.
Author Address: Bio-design research group, Bioproduction research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Central 6, 1-1-1 Higashi, Tsukuba 305-8566, Ibaraki, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1638
Author: Olivier-Van Stichelen, S., Dehennaut, V., Buzy, A., Zachayus, J. L., Guinez, C., Mir, A. M., El Yazidi-Belkoura, I., Copin, M. C., Boureme, D., Loyaux, D., Ferrara, P. and Lefebvre, T.
Year: 2014
Title: O-GlcNAcylation stabilizes β-catenin through direct competition with phosphorylation at threonine 41
Journal: Faseb j
Volume: 28
Issue: 8
Pages: 3325-38
Epub Date: 2014/04/20
Date: Aug
Short Title: O-GlcNAcylation stabilizes β-catenin through direct competition with phosphorylation at threonine 41
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638 (Print)
0892-6638
DOI: 10.1096/fj.13-243535
PMCID: PMC4101651
Accession Number: 24744147
Keywords: Acetylglucosamine/*metabolism
Adenocarcinoma/etiology/metabolism
Adherens Junctions/metabolism/pathology
Amino Acid Sequence
Animals
Colon/metabolism
Colorectal Neoplasms/etiology/metabolism
Dietary Carbohydrates/metabolism/toxicity
Enzyme Inhibitors/pharmacology
Glucose/metabolism
Glycosylation
HEK293 Cells
Humans
Hyperglycemia/complications/metabolism
Intestinal Mucosa/metabolism
MCF-7 Cells
Male
Mice
Mice, Inbred C57BL
Molecular Sequence Data
N-Acetylglucosaminyltransferases/antagonists & inhibitors/physiology
Neoplasm Proteins/chemistry/metabolism
Phosphorylation
Protein Interaction Mapping
*Protein Processing, Post-Translational
Protein Stability
Proteolysis
RNA, Small Interfering/pharmacology
Threonine/*chemistry
Wnt Signaling Pathway
alpha Catenin/metabolism
beta Catenin/*chemistry/metabolism
beta-N-Acetylhexosaminidases/antagonists & inhibitors/physiology
Etd-ms/ms
Wnt signaling
cancer
Abstract: Dysfunctions in Wnt signaling increase β-catenin stability and are associated with cancers, including colorectal cancer. In addition, β-catenin degradation is decreased by nutrient-dependent O-GlcNAcylation. Human colon tumors and colons from mice fed high-carbohydrate diets exhibited higher amounts of β-catenin and O-GlcNAc relative to healthy tissues and mice fed a standard diet, respectively. Administration of the O-GlcNAcase inhibitor thiamet G to mice also increased colonic expression of β-catenin. By ETD-MS/MS, we identified 4 O-GlcNAcylation sites at the N terminus of β-catenin (S23/T40/T41/T112). Furthermore, mutation of serine and threonine residues within the D box of β-catenin reduced O-GlcNAcylation by 75%. Interestingly, elevating O-GlcNAcylation in human colon cell lines drastically reduced phosphorylation at T41, a key residue of the D box responsible for β-catenin stability. Analyses of β-catenin O-GlcNAcylation mutants reinforced T41 as the most crucial residue that controls the β-catenin degradation rate. Finally, inhibiting O-GlcNAcylation decreased the β-catenin/α-catenin interaction necessary for mucosa integrity, whereas O-GlcNAcase silencing improved this interaction. These results suggest that O-GlcNAcylation regulates not only the stability of β-catenin, but also affects its localization at the level of adherens junctions. Accordingly, we propose that O-GlcNAcylation of β-catenin is a missing link between the glucose metabolism deregulation observed in metabolic disorders and the development of cancer.
Notes: 1530-6860
Olivier-Van Stichelen, Stéphanie
Dehennaut, Vanessa
Buzy, Armelle
Zachayus, Jean-Luc
Guinez, Céline
Mir, Anne-Marie
El Yazidi-Belkoura, Ikram
Copin, Marie-Christine
Boureme, Didier
Loyaux, Denis
Ferrara, Pascual
Lefebvre, Tony
Journal Article
Research Support, Non-U.S. Gov't
FASEB J. 2014 Aug;28(8):3325-38. doi: 10.1096/fj.13-243535. Epub 2014 Apr 17.
Author Address: Unit of Structural and Functional Glycobiology, Institut Fédératif de Recherche (IFR) 147, Centre National de la Recherche Scientifique-Unité Mixte de Recherche (CNRS-UMR) 8576, University of Lille 1, Villeneuve d'Ascq, France;
Exploratory Unit/Systems Biology, Sanofi-Aventis Research and Development, Toulouse, France; and.
Tumor Bank, Regional Reference Center in Cancer, Centre Hospitalier Régional Universitaire de Lille (CHRU), University of Lille 2, Lille, France.
Unit of Structural and Functional Glycobiology, Institut Fédératif de Recherche (IFR) 147, Centre National de la Recherche Scientifique-Unité Mixte de Recherche (CNRS-UMR) 8576, University of Lille 1, Villeneuve d'Ascq, France; tony.lefebvre@univ-lille1.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1550
Author: Olivier-Van Stichelen, S., Drougat, L., Dehennaut, V., El Yazidi-Belkoura, I., Guinez, C., Mir, A. M., Michalski, J. C., Vercoutter-Edouart, A. S. and Lefebvre, T.
Year: 2012
Title: Serum-stimulated cell cycle entry promotes ncOGT synthesis required for cyclin D expression
Journal: Oncogenesis
Volume: 1
Issue: 12
Pages: e36
Epub Date: 2012/01/01
Date: Dec 10
Short Title: Serum-stimulated cell cycle entry promotes ncOGT synthesis required for cyclin D expression
Alternate Journal: Oncogenesis
ISSN: 2157-9024 (Print)
2157-9024
DOI: 10.1038/oncsis.2012.36
PMCID: PMC3545199
Accession Number: 23552487
Abstract: Nuclear and cytoplasmic O-GlcNAc transferase (OGT) is a unique and universally expressed enzyme catalyzing O-GlcNAcylation of thousands of proteins. Although OGT interferes with many crucial intracellular processes, including cell cycle, only few studies have focused on elucidating the precise role of the glycosyltransferase during cell cycle entry. We first demonstrated that starved MCF7 cells reincubated with serum quickly induced a significant OGT increase concomitantly to activation of PI3K and MAPK pathways. Co-immunoprecipitation experiments performed upon serum stimulation showed a progressive interaction between OGT and β-catenin, a major factor in the regulation of cell cycle. OGT expression was also observed in starved HeLa cells reincubated with serum. In these cells, the O-GlcNAcylation status of the β-catenin-2XFLAG was increased following stimulation. Moreover, β-catenin-2XFLAG was heavily O-GlcNAcylated in exponentially proliferating HeLa cells when compared to confluent cells. Furthermore, blocking OGT activity using the potent inhibitor Ac-5SGlcNAc prevented serum-stimulated cyclin D1 synthesis and slightly delayed cell proliferation. At last, interfering with OGT expression (siOGT) blocked cyclin D1 expression and decreased PI3K and MAPK activation. Together, our data indicate that expression and catalytic activity of OGT are necessary and essential for G0/G1 transition.
Notes: 2157-9024
Olivier-Van Stichelen, S
Drougat, L
Dehennaut, V
El Yazidi-Belkoura, I
Guinez, C
Mir, A-M
Michalski, J-C
Vercoutter-Edouart, A-S
Lefebvre, T
Journal Article
Oncogenesis. 2012 Dec 10;1(12):e36. doi: 10.1038/oncsis.2012.36.
Author Address: CNRS-UMR 8576, Unit of Structural and Functional Glycobiology (UGSF), IFR 147, University of Lille 1, CitÃ© Scientifique, Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 655
Author: Olsen, E. H., Rahbek-Nielsen, H., Thogersen, I. B., Roepstorff, P. and Enghild, J. J.
Year: 1998
Title: Posttranslational modifications of human inter-alpha-inhibitor: identification of glycans and disulfide bridges in heavy chains 1 and 2
Journal: Biochemistry
Volume: 37
Issue: 1
Pages: 408-16
Epub Date: 1998/02/07
Date: Jan 6
Short Title: Posttranslational modifications of human inter-alpha-inhibitor: identification of glycans and disulfide bridges in heavy chains 1 and 2
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi971137d
Accession Number: 9425062
Keywords: Alpha-Globulins/chemistry/*metabolism
Asparagine/chemistry
Carbohydrate Conformation
Cysteine/chemistry
Disulfides/chemistry/*metabolism
Glycosylation
Humans
Monosaccharides/chemistry/metabolism
Oligosaccharides/chemistry/metabolism
Peptides/chemistry/isolation & purification/metabolism
Polysaccharides/chemistry/*metabolism
*Protein Processing, Post-Translational
Abstract: Inter-alpha-inhibitor (IalphaI) is a serine proteinase inhibitor found in high concentrations in human plasma. The protein is composed of a light inhibitory chain called bikunin and two heavy chains of unknown function. The three polypeptide chains are covalently assembled via a carbohydrate cross-link [Enghild, J. J., Salvesen, G., Hefta, S. A., Thogersen, I. B., Rutherfurd, S., & Pizzo, S. V. (1991) J. Biol. Chem. 266, 747-751]. The aim of this study was to complete the primary structure by characterizing additional covalent posttranslational modifications of the heavy chains. Analysis revealed three N-linked oligosaccharides located on Asn251 and Asn554 of heavy chain 1 and on Asn64 of heavy chain 2: all these were complex biantennary structures composed of (Asn)-GlcNAc2-Man-(Man-GlcNAc-Gal-SA)2. In addition, the IalphaI heavy chains carried several O-linked glycans located on Thr619 of heavy chain 1 and a cluster of four O-linked oligosaccharides on Thr612, Ser619, Thr621, and Thr637 of heavy chain 2. The oligosaccharides were short (Ser/Thr)-GalNAc-Gal-SA trisaccharides. The IalphaI heavy chains contain nine Cys residues, of which eight are involved in disulfide bridges. The unpaired Cys residue residing on heavy chain 1, Cys26, appears to be modified by dihexosylation. The other Cys residues exclusively form intrachain disulfide bridges. In heavy chain 1 the two disulfide bonds are formed between Cys210 and Cys213 and between Cys234 and Cys506, and in heavy chain 2, between Cys207 and Cys210 and between Cys596 and Cys597. Interestingly, three of these four disulfides are formed between Cys residues that are either adjacent or only two amino acid residues apart.
Notes: Olsen, E H
Rahbek-Nielsen, H
Thogersen, I B
Roepstorff, P
Enghild, J J
HL49542/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1998 Jan 6;37(1):408-16. doi: 10.1021/bi971137d.
Author Address: Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 819
Author: Olsen, J. G., Kadziola, A., Lauritzen, C., Pedersen, J., Larsen, S. and Dahl, S. W.
Year: 2001
Title: Tetrameric dipeptidyl peptidase I directs substrate specificity by use of the residual pro-part domain
Journal: FEBS Lett
Volume: 506
Issue: 3
Pages: 201-6
Epub Date: 2001/10/17
Date: Oct 12
Short Title: Tetrameric dipeptidyl peptidase I directs substrate specificity by use of the residual pro-part domain
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/s0014-5793(01)02911-8
Accession Number: 11602245
Keywords: Animals
Cathepsin C/chemistry/*metabolism
Models, Molecular
Protein Conformation
Rats
Recombinant Proteins/chemistry/metabolism
Substrate Specificity
Abstract: The crystal structure of mature dipeptidyl peptidase I reveals insight into the unique tetrameric structure, substrate binding and activation of this atypical papain family peptidase. Each subunit is composed of three peptides. The heavy and light chains form the catalytic domain, which adopts the papain fold. The residual pro-part forms a beta-barrel with the carboxylate group of Asp1 pointing towards the substrate amino-terminus. The tetrameric structure appears to stabilize the association of the two domains and encloses a 12700 A3 spherical cavity. The tetramer contains six chloride ions, one buried in each S2 pocket and two at subunit interfaces.
Notes: Olsen, J G
Kadziola, A
Lauritzen, C
Pedersen, J
Larsen, S
Dahl, S W
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2001 Oct 12;506(3):201-6. doi: 10.1016/s0014-5793(01)02911-8.
Author Address: Center for Crystallographic Studies, University of Copenhagen, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1976
Author: Oosterheert, W., van Bezouwen, L. S., Rodenburg, R. N. P., Granneman, J., Förster, F., Mattevi, A. and Gros, P.
Year: 2018
Title: Cryo-EM structures of human STEAP4 reveal mechanism of iron(III) reduction
Journal: Nat Commun
Volume: 9
Issue: 1
Pages: 4337
Epub Date: 2018/10/20
Date: Oct 18
Short Title: Cryo-EM structures of human STEAP4 reveal mechanism of iron(III) reduction
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-018-06817-7
PMCID: PMC6194020
Accession Number: 30337524
Keywords: Binding Sites
Biocatalysis
*Cryoelectron Microscopy
Electrons
Flavin-Adenine Dinucleotide/metabolism
Heme/metabolism
Humans
Iron/*metabolism
Membrane Proteins/chemistry/metabolism/*ultrastructure
NADP/metabolism
NADPH Oxidases/metabolism
Oxidation-Reduction
Oxidoreductases/chemistry/metabolism/*ultrastructure
Protein Domains
Substrate Specificity
Abstract: Enzymes of the six-transmembrane epithelial antigen of the prostate (STEAP) family reduce Fe(3+) and Cu(2+) ions to facilitate metal-ion uptake by mammalian cells. STEAPs are highly upregulated in several types of cancer, making them potential therapeutic targets. However, the structural basis for STEAP-catalyzed electron transfer through an array of cofactors to metals at the membrane luminal side remains elusive. Here, we report cryo-electron microscopy structures of human STEAP4 in absence and presence of Fe(3+)-NTA. Domain-swapped, trimeric STEAP4 orients NADPH bound to a cytosolic domain onto axially aligned flavin-adenine dinucleotide (FAD) and a single b-type heme that cross the transmembrane-domain to enable electron transfer. Substrate binding within a positively charged ring indicates that iron gets reduced while in complex with its chelator. These molecular principles of iron reduction provide a basis for exploring STEAPs as therapeutic targets.
Notes: 2041-1723
Oosterheert, Wout
van Bezouwen, Laura S
Rodenburg, Remco N P
Granneman, Joke
Förster, Friedrich
Mattevi, Andrea
Orcid: 0000-0002-9523-7128
Gros, Piet
Orcid: 0000-0002-7782-2585
IG19808/Associazione Italiana per la Ricerca sul Cancro (Italian Association for Cancer Research)/International
731.015.201/Nederlandse Organisatie voor Wetenschappelijk Onderzoek (Netherlands Organisation for Scientific Research)/International
653706/EC | Horizon 2020 (European Union Framework Programme for Research and Innovation)/International
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2018 Oct 18;9(1):4337. doi: 10.1038/s41467-018-06817-7.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
Cryo-Electron Microscopy, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
Department of Biology and Biotechnology 'L. Spallanzani', University of Pavia, 27100, Pavia, Italy.
Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands. p.gros@uu.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1032
Author: Oostra, M., de Haan, C. A., de Groot, R. J. and Rottier, P. J.
Year: 2006
Title: Glycosylation of the severe acute respiratory syndrome coronavirus triple-spanning membrane proteins 3a and M
Journal: J Virol
Volume: 80
Issue: 5
Pages: 2326-36
Epub Date: 2006/02/14
Date: Mar
Short Title: Glycosylation of the severe acute respiratory syndrome coronavirus triple-spanning membrane proteins 3a and M
Alternate Journal: Journal of virology
ISSN: 0022-538X (Print)
0022-538x
DOI: 10.1128/jvi.80.5.2326-2336.2006
PMCID: PMC1395384
Accession Number: 16474139
Keywords: Amino Acid Sequence
Coronavirus M Proteins
Gene Expression
Glycosylation
Molecular Sequence Data
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
*Protein Modification, Translational
*Protein Processing, Post-Translational
Radioisotopes
SARS Virus/*chemistry
Sialic Acids/analysis
Staining and Labeling
Viral Envelope Proteins
Viral Matrix Proteins/chemistry/*metabolism
Viral Proteins/chemistry/*metabolism
Viroporin Proteins
Abstract: The severe acute respiratory syndrome coronavirus (SARS-CoV) open reading frame 3a protein has recently been shown to be a structural protein. The protein is encoded by one of the so-called group-specific genes and has no sequence homology with any of the known structural or group-specific proteins of coronaviruses. It does, however, have several similarities to the coronavirus M proteins; (i) they are triple membrane spanning with the same topology, (ii) they have similar intracellular localizations (predominantly Golgi), (iii) both are viral structural proteins, and (iv) they appear to interact with the E and S proteins, as well as with each other. The M protein plays a crucial role in coronavirus assembly and is glycosylated in all coronaviruses, either by N-linked or by O-linked oligosaccharides. The conserved glycosylation of the coronavirus M proteins and the resemblance of the 3a protein to them led us to investigate the glycosylation of these two SARS-CoV membrane proteins. The proteins were expressed separately using the vaccinia virus T7 expression system, followed by metabolic labeling. Pulse-chase analysis showed that both proteins were modified, although in different ways. While the M protein acquired cotranslationally oligosaccharides that could be removed by PNGaseF, the 3a protein acquired its modifications posttranslationally, and they were not sensitive to the N-glycosidase enzyme. The SARS-CoV 3a protein, however, was demonstrated to contain sialic acids, indicating the presence of oligosaccharides. O-glycosylation of the 3a protein was indeed confirmed using an in situ O-glycosylation assay of endoplasmic reticulum-retained mutants. In addition, we showed that substitution of serine and threonine residues in the ectodomain of the 3a protein abolished the addition of the O-linked sugars. Thus, the SARS-CoV 3a protein is an O-glycosylated glycoprotein, like the group 2 coronavirus M proteins but unlike the SARS-CoV M protein, which is N glycosylated.
Notes: 1098-5514
Oostra, M
de Haan, C A M
de Groot, R J
Rottier, P J M
Journal Article
J Virol. 2006 Mar;80(5):2326-36. doi: 10.1128/JVI.80.5.2326-2336.2006.
Author Address: Virology Division, Department of Infectious Diseases and Immunology, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1192
Author: Oppizzi, M. L., Akhavan, A., Singh, M., Fata, J. E. and Muschler, J. L.
Year: 2008
Title: Nuclear translocation of beta-dystroglycan reveals a distinctive trafficking pattern of autoproteolyzed mucins
Journal: Traffic
Volume: 9
Issue: 12
Pages: 2063-72
Epub Date: 2008/09/04
Date: Dec
Short Title: Nuclear translocation of beta-dystroglycan reveals a distinctive trafficking pattern of autoproteolyzed mucins
Alternate Journal: Traffic (Copenhagen, Denmark)
ISSN: 1398-9219 (Print)
1398-9219
DOI: 10.1111/j.1600-0854.2008.00822.x
PMCID: PMC2950207
NIHMSID: NIHMS240157
Accession Number: 18764929
Keywords: Active Transport, Cell Nucleus
Amino Acid Sequence
Animals
Carcinoma/metabolism
Cell Line, Tumor
Cell Nucleus/*metabolism
Cytoplasm/metabolism
Dystroglycans/chemistry/genetics/*metabolism
Humans
Molecular Sequence Data
Mucins/chemistry/*metabolism
Sequence Alignment
Abstract: Dystroglycan (DG) is an extracellular matrix receptor implicated in muscular dystrophies and cancers. DG belongs to the membrane-tethered mucin family and is composed of extracellular (alpha-DG) and transmembrane (beta-DG) subunits stably coupled at the cell surface. These two subunits are generated by autoproteolysis of a monomeric precursor within a distinctive protein motif called sea urchin-enterokinase-agrin (SEA) domain, yet the purpose of this cleavage and heterodimer creation is uncertain. In this study, we identify a functional nuclear localization signal within beta-DG and show that, in addition to associating with alpha-DG at the cell surface, the full-length and glycosylated beta-DG autonomously traffics to the cytoplasm and nucleoplasm in a process that occurs independent of alpha-DG ligand binding. The trafficking pattern of beta-DG mirrors that of MUC1-C, the transmembrane subunit of the related MUC1 oncoprotein, also a heterodimeric membrane-tethered mucin created by SEA autoproteolysis. We show that the transmembrane subunits of both MUC1 and DG transit the secretory pathway prior to nuclear targeting and that their monomeric precursors maintain the capacity for nuclear trafficking. A screen of breast carcinoma cell lines of distinct pathophysiological origins revealed considerable variability in the nuclear partitioning of beta-DG, indicating that nuclear localization of beta-DG is regulated, albeit independent of extracellular ligand binding. These findings point to novel intracellular functions for beta-DG, with possible disease implications. They also reveal an evolutionarily conserved role for SEA autoproteolysis, serving to enable independent functions of mucin transmembrane subunits, enacted by a shared and poorly understood pathway of segregated subunit trafficking.
Notes: 1600-0854
Oppizzi, Maria Luisa
Akhavan, Armin
Singh, Manisha
Fata, Jimmie E
Muschler, John L
R01 CA109579/CA/NCI NIH HHS/United States
R01 CA109579-05S2/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Traffic. 2008 Dec;9(12):2063-72. doi: 10.1111/j.1600-0854.2008.00822.x. Epub 2008 Sep 18.
Author Address: California Pacific Medical Center Research Institute, 475 Brannan Street, Suite 220, San Francisco, CA 94107, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 258
Author: Orberger, G., Geyer, R., Stirm, S. and Tauber, R.
Year: 1992
Title: Structure of the N-linked oligosaccharides of the human transferrin receptor
Journal: Eur J Biochem
Volume: 205
Issue: 1
Pages: 257-67
Epub Date: 1992/04/01
Date: Apr 1
Short Title: Structure of the N-linked oligosaccharides of the human transferrin receptor
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1992.tb16776.x
Accession Number: 1555586
Keywords: Agglutinins/metabolism
Blotting, Western
Carbohydrate Sequence
Chromatography, Affinity
Chromatography, Ion Exchange
Electrophoresis, Polyacrylamide Gel
Female
Glycosylation
Humans
Liver Neoplasms
Mass Spectrometry/methods
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry
Oxidation-Reduction
Placenta/chemistry
Polysaccharides/isolation & purification/metabolism
Pregnancy
Receptors, Transferrin/*chemistry/isolation & purification
Tumor Cells, Cultured
Abstract: Human transferrin receptor was isolated from placenta and from the hepatocarcinoma cell line Hep G2. Asparagine-linked oligosaccharides were released by treatment of tryptic glycopeptides with endo-beta-N-acetylglucosaminidase H or peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. Oligosaccharide alditols were fractionated by anion-exchange high-performance liquid chromatography and by high-pH anion-exchange chromatography. Glycans from placental transferrin receptor were further characterized, after desialylation, by methylation analysis and, in part, by liquid secondary-ion mass spectrometry. Sialylation of placental transferrin receptor was examined by lectin affinity blotting with Sambucus nigra agglutinin and Maackia amurensis agglutinin. In order to trace possible inter-individual differences in N-glycosylation of the receptor, two preparations of placental transferrin receptor purified from two donors were compared. The results demonstrate that human transferrin receptor from placenta predominantly carries diantennary and triantennary N-acetyllactosaminic glycans as well as hybrid-type species, the galactose residues of which being almost completely substituted with (alpha 2-3)-linked sialic acid residues. Distinct differences were noted in the glycosylation pattern of the receptor from different individuals. Transferrin receptor from donor A carried predominantly diantennary and triantennary complex-type glycans, in part fucosylated at the innermost N-acetylglucosamine residue, in addition to small amounts of bisected and of incomplete diantennary species. Placental transferrin receptor from donor B predominantly carried triantennary N-acetyllactosaminic glycans without fucose and hybrid-type oligosaccharides with four or five mannose residues. Distinct from placental transferrin receptor, the receptor from Hep G2 cells contained larger amounts of oligomannosidic glycans with six to nine mannose residues and tetrasialylated complex-type oligosaccharides apart from mono-, di- and trisialylated species.
Notes: Orberger, G
Geyer, R
Stirm, S
Tauber, R
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1992 Apr 1;205(1):257-67. doi: 10.1111/j.1432-1033.1992.tb16776.x.
Author Address: Institut für Klinische Chemie und Biochemie, Universitätsklinikum Rudolf-Virchow, Freie Universität Berlin, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1782
Author: Orlando, B. J. and Malkowski, M. G.
Year: 2016
Title: Substrate-selective Inhibition of Cyclooxygeanse-2 by Fenamic Acid Derivatives Is Dependent on Peroxide Tone
Journal: J Biol Chem
Volume: 291
Issue: 29
Pages: 15069-81
Epub Date: 2016/05/27
Date: Jul 15
Short Title: Substrate-selective Inhibition of Cyclooxygeanse-2 by Fenamic Acid Derivatives Is Dependent on Peroxide Tone
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.725713
PMCID: PMC4946924
Accession Number: 27226593
Keywords: Amino Acid Substitution
Binding Sites
Crystallography, X-Ray
Cyclooxygenase 2/chemistry/genetics/*metabolism
Cyclooxygenase 2 Inhibitors/*chemistry/*pharmacology
Endocannabinoids/metabolism
Fenamates/chemistry/*pharmacology
Heme/chemistry
Humans
In Vitro Techniques
Models, Molecular
Mutagenesis, Site-Directed
Peroxides/metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Substrate Specificity
Tryptophan/chemistry
*crystal structure
*cyclooxygenase (COX)
*electron paramagnetic resonance (EPR)
*endocannabinoid
*nonsteroidal anti-inflammatory drugs
*structural biology
Abstract: Cyclooxygenase-2 (COX-2) catalyzes the oxygenation of arachidonic acid (AA) and endocannabinoid substrates, placing the enzyme at a unique junction between the eicosanoid and endocannabinoid signaling pathways. COX-2 is a sequence homodimer, but the enzyme displays half-of-site reactivity, such that only one monomer of the dimer is active at a given time. Certain rapid reversible, competitive nonsteroidal anti-inflammatory drugs (NSAIDs) have been shown to inhibit COX-2 in a substrate-selective manner, with the binding of inhibitor to a single monomer sufficient to inhibit the oxygenation of endocannabinoids but not arachidonic acid. The underlying mechanism responsible for substrate-selective inhibition has remained elusive. We utilized structural and biophysical methods to evaluate flufenamic acid, meclofenamic acid, mefenamic acid, and tolfenamic acid for their ability to act as substrate-selective inhibitors. Crystal structures of each drug in complex with human COX-2 revealed that the inhibitor binds within the cyclooxygenase channel in an inverted orientation, with the carboxylate group interacting with Tyr-385 and Ser-530 at the top of the channel. Tryptophan fluorescence quenching, continuous-wave electron spin resonance, and UV-visible spectroscopy demonstrate that flufenamic acid, mefenamic acid, and tolfenamic acid are substrate-selective inhibitors that bind rapidly to COX-2, quench tyrosyl radicals, and reduce higher oxidation states of the heme moiety. Substrate-selective inhibition was attenuated by the addition of the lipid peroxide 15-hydroperoxyeicosatertaenoic acid. Collectively, these studies implicate peroxide tone as an important mechanistic component of substrate-selective inhibition by flufenamic acid, mefenamic acid, and tolfenamic acid.
Notes: 1083-351x
Orlando, Benjamin J
Malkowski, Michael G
P41 GM103521/GM/NIGMS NIH HHS/United States
R01 GM115386/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2016 Jul 15;291(29):15069-81. doi: 10.1074/jbc.M116.725713. Epub 2016 May 20.
Author Address: From the Department of Structural Biology, The State University of New York at Buffalo and.
From the Department of Structural Biology, The State University of New York at Buffalo and the Hauptman-Woodward Medical Research Institute, Buffalo, New York 14203 mgm22@buffalo.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1812
Author: Orlando, B. J. and Malkowski, M. G.
Year: 2016
Title: Crystal structure of rofecoxib bound to human cyclooxygenase-2
Journal: Acta Crystallogr F Struct Biol Commun
Volume: 72
Issue: Pt 10
Pages: 772-776
Epub Date: 2016/10/07
Date: Oct 1
Short Title: Crystal structure of rofecoxib bound to human cyclooxygenase-2
Alternate Journal: Acta crystallographica. Section F, Structural biology communications
ISSN: 2053-230x
DOI: 10.1107/s2053230x16014230
PMCID: PMC5053162
Accession Number: 27710942
Keywords: Amino Acid Motifs
Binding Sites
Celecoxib/*chemistry
Crystallography, X-Ray
Cyclooxygenase 2/*chemistry/genetics/metabolism
Cyclooxygenase 2 Inhibitors/*chemistry
Gene Expression
Humans
Kinetics
Lactones/*chemistry
Models, Molecular
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Recombinant Proteins/chemistry/genetics/metabolism
Safety-Based Drug Withdrawals
Substrate Specificity
Sulfones/*chemistry
*Vioxx
*crystal structure
*cyclooxygenase
*nonsteroidal anti-inflammatory drugs
*rofecoxib
Abstract: Rofecoxib (Vioxx) was one of the first selective cyclooxygenase-2 (COX-2) inhibitors (coxibs) to be approved for use in humans. Within five years after its release to the public, Vioxx was withdrawn from the market owing to the adverse cardiovascular effects of the drug. Despite the widespread knowledge of the development and withdrawal of Vioxx, relatively little is known at the molecular level about how the inhibitor binds to COX-2. Vioxx is unique in that the inhibitor contains a methyl sulfone moiety in place of the sulfonamide moiety found in other coxibs such as celecoxib and valdecoxib. Here, new crystallization conditions were identified that allowed the structural determination of human COX-2 in complex with Vioxx and the structure was subsequently determined to 2.7 Å resolution. The crystal structure provides the first atomic level details of the binding of Vioxx to COX-2. As anticipated, Vioxx binds with its methyl sulfone moiety located in the side pocket of the cyclooxygenase channel, providing support for the isoform selectivity of this drug.
Notes: 2053-230x
Orlando, Benjamin J
Malkowski, Michael G
R01 GM077176/GM/NIGMS NIH HHS/United States
R01 GM115386/GM/NIGMS NIH HHS/United States
Journal Article
Acta Crystallogr F Struct Biol Commun. 2016 Oct 1;72(Pt 10):772-776. doi: 10.1107/S2053230X16014230. Epub 2016 Sep 22.
Author Address: Department of Structural Biology, The State University of New York at Buffalo and the Hauptman-Woodward Medical Research Institute, 700 Ellicott Street, Buffalo, NY 14203, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1631
Author: Ortiz-Meoz, R. F., Merbl, Y., Kirschner, M. W. and Walker, S.
Year: 2014
Title: Microarray discovery of new OGT substrates: the medulloblastoma oncogene OTX2 is O-GlcNAcylated
Journal: J Am Chem Soc
Volume: 136
Issue: 13
Pages: 4845-8
Epub Date: 2014/03/04
Date: Apr 2
Short Title: Microarray discovery of new OGT substrates: the medulloblastoma oncogene OTX2 is O-GlcNAcylated
Alternate Journal: Journal of the American Chemical Society
ISSN: 0002-7863 (Print)
0002-7863
DOI: 10.1021/ja500451w
PMCID: PMC3988687
Accession Number: 24580054
Keywords: Acylation
Cell Line, Tumor
Cerebellar Neoplasms/*metabolism
Humans
Medulloblastoma/*metabolism
N-Acetylglucosaminyltransferases/*metabolism
Otx Transcription Factors/*metabolism
Protein Array Analysis
Abstract: O-GlcNAc transferase (OGT) is a serine/threonine glycosyltransferase that is essential for development and continues to be critically important throughout life. Understanding OGT's complex biology requires identifying its substrates. Here we demonstrate the utility of a microarray approach for discovering novel OGT substrates. We also report a rapid method to validate OGT substrates that combines in vitro transcription-translation with O-GlcNAc mass tagging. Among the validated new OGT targets is Orthodenticle homeobox 2 (OTX2), a transcription factor critical for brain development, which is primarily expressed only during early embryogenesis and in medulloblastomas, where it functions as an oncogene. We show that endogenous OTX2 from a medulloblastoma cell line is O-GlcNAcylated at several sites. Our results demonstrate that protein microarray technology, combined with the target validation strategy we report, is useful for identifying biologically important OGT substrates, including substrates not present in most tissue types or cell lines.
Notes: 1520-5126
Ortiz-Meoz, Rodrigo F
Merbl, Yifat
Kirschner, Marc W
Walker, Suzanne
R01 GM094263/GM/NIGMS NIH HHS/United States
R01 GM026875/GM/NIGMS NIH HHS/United States
F32 GM100539/GM/NIGMS NIH HHS/United States
GM094263/GM/NIGMS NIH HHS/United States
GM26875/GM/NIGMS NIH HHS/United States
GM100539/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Am Chem Soc. 2014 Apr 2;136(13):4845-8. doi: 10.1021/ja500451w. Epub 2014 Mar 17.
Author Address: Department of Microbiology and Immunobiology, and ‡Department of Systems Biology, Harvard Medical School , Boston, Massachusetts 02115, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 860
Author: Ortlund, E., LaCount, M. W. and Lebioda, L.
Year: 2003
Title: Crystal structures of human prostatic acid phosphatase in complex with a phosphate ion and alpha-benzylaminobenzylphosphonic acid update the mechanistic picture and offer new insights into inhibitor design
Journal: Biochemistry
Volume: 42
Issue: 2
Pages: 383-9
Epub Date: 2003/01/15
Date: Jan 21
Short Title: Crystal structures of human prostatic acid phosphatase in complex with a phosphate ion and alpha-benzylaminobenzylphosphonic acid update the mechanistic picture and offer new insights into inhibitor design
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi0265067
Accession Number: 12525165
Keywords: Acid Phosphatase
Animals
Binding Sites
Carbohydrates/chemistry
Catalysis
Crystallography, X-Ray
Dimerization
Enzyme Inhibitors/*chemical synthesis
Humans
Macromolecular Substances
Male
Models, Chemical
Models, Molecular
Organophosphonates/*chemical synthesis/*chemistry
Phosphates/*chemistry
Polyethylene Glycols/chemistry
Protein Tyrosine Phosphatases/*antagonists & inhibitors/*chemistry
Rats
Semen/enzymology
Abstract: The X-ray crystal structure of human prostatic acid phosphatase (PAP) in complex with a phosphate ion has been determined at 2.4 A resolution. This structure offers a snapshot of the final intermediate in the catalytic mechanism and does not support the role of Asp 258 as a proton donor in catalysis. A total of eight hydrogen bonds serve to strongly bind the phosphate ion within the active site. Bound PEG molecules from the crystallization matrix have allowed the identification of a channel within the molecule that likely plays a role in molecular recognition and in macromolecular substrate selectivity. Additionally, the structure of PAP in complex with a phosphate derivative, alpha-benzylaminobenzylphosphonic acid, a potent inhibitor (IC(50) = 4 nM), has been determined to 2.9 A resolution. This structure gives new insight into the determinants of binding hydrophobic ligands within the active site and allows us to explain PAP's preference for aromatic substrates.
Notes: Ortlund, Eric
LaCount, Michael W
Lebioda, Lukasz
GM42898/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2003 Jan 21;42(2):383-9. doi: 10.1021/bi0265067.
Author Address: Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 581
Author: Otto, J. C., DeWitt, D. L. and Smith, W. L.
Year: 1993
Title: N-glycosylation of prostaglandin endoperoxide synthases-1 and -2 and their orientations in the endoplasmic reticulum
Journal: J Biol Chem
Volume: 268
Issue: 24
Pages: 18234-42
Epub Date: 1993/08/25
Date: Aug 25
Short Title: N-glycosylation of prostaglandin endoperoxide synthases-1 and -2 and their orientations in the endoplasmic reticulum
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8349699
Keywords: Amino Acid Sequence
Animals
Base Sequence
Blotting, Western
Cell Line
Conserved Sequence
Electrophoresis, Polyacrylamide Gel
Endoplasmic Reticulum/*enzymology/ultrastructure
Glycosylation
Heme/pharmacology
Indomethacin/pharmacology
Intracellular Membranes/*enzymology/ultrastructure
Isoenzymes/biosynthesis/isolation & purification/*metabolism
Mice
Microsomes/*enzymology/ultrastructure
Molecular Sequence Data
Mutagenesis, Site-Directed
Oligodeoxyribonucleotides
Peptide Mapping
Peroxidases/metabolism
Prostaglandin-Endoperoxide Synthases/biosynthesis/isolation &
purification/*metabolism
Protein Conformation
Restriction Mapping
Sheep
Transfection
Trypsin/metabolism
Abstract: Using site-directed mutagenesis, we have determined that Asn68, Asn144, and Asn410 of ovine prostaglandin endoperoxide (PGH) synthase-1 are N-glycosylated. A fourth consensus N-glycosylation sequence at Asn104 is not glycosylated. Glycosylation of PGH synthase-1 at Asn410 and at either Asn68 or Asn144 was required for expression of both the cyclooxygenase and the peroxidase activities of the enzyme. Inactive PGH synthase-1 glycosylation site mutant proteins do not appear to achieve their native conformations. However, the native enzyme, once in an active conformation, does not appear to require attached carbohydrate for cyclooxygenase or peroxidase activities. N-Glycosylation consensus sequences corresponding to the three glycosylation sites of ovine PGH synthase-1 are conserved in the deduced amino acid sequences of PGH synthases-2. Using site-directed mutagenesis, we determined that there is an additional site of N-glycosylation in murine PGH synthase-2 located at Asn580. This site is N-glycosylated in about 50% of PGH synthase-2 molecules, resulting in two peptide bands on SDS-polyacrylamide gel electrophoresis (72 and 74 kDa). Glycosylation of PGH synthase-2 is necessary for expression of enzyme activity, but glycosylation of PGH synthase-2 at Asn580 per se does not affect activity. Assuming that the N-glycosylation sites of PGH synthase-1 are on the luminal side of the endoplasmic reticulum (ER), and that the site of tryptic cleavage of ovine PGH synthase-1 (Arg277) is on the cytoplasmic side of the ER, we propose that both the NH2 and COOH termini of PGH synthase-1 are located in the lumen of the ER and that there are two transmembrane domains located between Asn144 and Arg277 and between Arg277 and Asn410, respectively. A similar orientation is predicted for PGH synthase-2.
Notes: Otto, J C
DeWitt, D L
Smith, W L
DK22042/DK/NIDDK NIH HHS/United States
DK42509/DK/NIDDK NIH HHS/United States
HL-07404/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1993 Aug 25;268(24):18234-42.
Author Address: Department of Biochemistry, Michigan State University, East Lansing 48824.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1064
Author: Otto, V. I., Damoc, E., Cueni, L. N., Schürpf, T., Frei, R., Ali, S., Callewaert, N., Moise, A., Leary, J. A., Folkers, G. and Przybylski, M.
Year: 2006
Title: N-glycan structures and N-glycosylation sites of mouse soluble intercellular adhesion molecule-1 revealed by MALDI-TOF and FTICR mass spectrometry
Journal: Glycobiology
Volume: 16
Issue: 11
Pages: 1033-44
Epub Date: 2006/08/01
Date: Nov
Short Title: N-glycan structures and N-glycosylation sites of mouse soluble intercellular adhesion molecule-1 revealed by MALDI-TOF and FTICR mass spectrometry
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwl032
Accession Number: 16877748
Keywords: Animals
CHO Cells
Consensus Sequence
Cricetinae
Cricetulus
Glycosylation
Intercellular Adhesion Molecule-1/genetics/*metabolism
Mass Spectrometry
Mice
Mutation
N-Acetylneuraminic Acid/metabolism
Polysaccharides/*metabolism
Protein Isoforms/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Intercellular adhesion molecule-1 (ICAM-1) is a heavily N-glycosylated transmembrane protein comprising five extracellular Ig-like domains. The soluble isoform of ICAM-1 (sICAM-1), consisting of its extracellular part, is elevated in the cerebrospinal fluid of patients with severe brain trauma. In mouse astrocytes, recombinant mouse sICAM-1 induces the production of the CXC chemokine macrophage inflammatory protein-2 (MIP-2). MIP-2 induction is glycosylation dependent, as it is strongly enhanced when sICAM-1 carries sialylated, complex-type N-glycans as synthesized by wild-type Chinese hamster ovary (CHO) cells. The present study was aimed at elucidating the N-glycosylation of mouse sICAM-1 expressed in wild-type CHO cells with regard to sialylation, N-glycan profile, and N-glycosylation sites. Ion-exchange chromatography and matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) of the released N-glycans showed that sICAM-1 mostly carried di- and trisialylated complex-type N-glycans with or without one fucose. In some sialylated N-glycans, one N-acetylneuraminic acid was replaced by N-glycolylneuraminic acid, and approximately 4% carried a higher number of sialic acid residues than of antennae. The N-glycosylation sites of mouse sICAM-1 were analyzed by MALDI-Fourier transform ion cyclotron resonance (FTICR)-MS and nanoLC-ESI-FTICR-MS of tryptic digests of mouse sICAM-1 expressed in the Lec1 mutant of CHO cells. All nine consensus sequences for N-glycosylation were found to be glycosylated. These results show that the N-glycans that enhance the MIP-2-inducing activity of mouse sICAM-1 are mostly di- and trisialylated complex-type N-glycans including a small fraction carrying more sialic acid residues than antennae and that the nine N-glycosylation sites of mouse sICAM-1 are all glycosylated.
Notes: Otto, Vivianne I
Damoc, Eugen
Cueni, Leah N
Schürpf, Thomas
Frei, Renate
Ali, Sarah
Callewaert, Nico
Moise, Adrian
Leary, Julie A
Folkers, Gerd
Przybylski, Michael
Journal Article
England
Glycobiology. 2006 Nov;16(11):1033-44. doi: 10.1093/glycob/cwl032. Epub 2006 Jul 28.
Author Address: Institute of Pharmaceutical Sciences, Department of Chemistry and Applied Biosciences, ETH Zurich, Switzerland. vivianne.otto@pharma.ethz.ch
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1477
Author: Overath, T., Kuckelkorn, U., Henklein, P., Strehl, B., Bonar, D., Kloss, A., Siele, D., Kloetzel, P. M. and Janek, K.
Year: 2012
Title: Mapping of O-GlcNAc sites of 20 S proteasome subunits and Hsp90 by a novel biotin-cystamine tag
Journal: Mol Cell Proteomics
Volume: 11
Issue: 8
Pages: 467-77
Epub Date: 2012/05/05
Date: Aug
Short Title: Mapping of O-GlcNAc sites of 20 S proteasome subunits and Hsp90 by a novel biotin-cystamine tag
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M111.015966
PMCID: PMC3412975
Accession Number: 22556278
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Binding Sites
Biotin/metabolism
Blotting, Western
Carbon Radioisotopes/metabolism
Cattle
Chromatography, Liquid
Cystamine/metabolism
Glucose/pharmacology
Glycosylation/drug effects
HSP90 Heat-Shock Proteins/*metabolism
Mice
Mice, Inbred C57BL
Molecular Sequence Data
Peptides/*metabolism
Proteasome Endopeptidase Complex/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
alpha-Crystallins/metabolism
Abstract: The post-translational modification of proteins with O-GlcNAc is involved in various cellular processes including signal transduction, transcription, translation, and nuclear transport. This transient protein modification enables cells or tissues to adapt to nutrient conditions or stress. O-Glycosylation of the 26 S proteasome ATPase subunit Rpt2 is known to influence the stability of proteins by reducing their proteasome-dependent degradation. In contrast, knowledge of the sites of O-GlcNAcylation on the subunits of the catalytic core of the 26 S proteasome, the 20 S proteasome, and the impact on proteasome activity is very limited. This is predominantly because O-GlcNAc modifications are often substoichiometric and because 20 S proteasomes represent a complex protein mixture of different subtypes. Therefore, identification of O-GlcNAcylation sites on proteasome subunits essentially requires effective enrichment strategies. Here we describe an adapted β-elimination-based derivatization method of O-GlcNAc peptides using a novel biotin-cystamine tag. The specificity of the reaction was increased by differential isotopic labeling with either "light" biotin-cystamine or deuterated "heavy" biotin-cystamine. The enriched peptides were analyzed by LC-MALDI-TOF/TOF-MS and relatively quantified. The method was optimized using bovine α-crystallin and then applied to murine 20 S proteasomes isolated from spleen and brain and murine Hsp90 isolated from liver. Using this approach, we identified five novel and one known O-GlcNAc sites within the murine 20 S proteasome core complex that are located on five different subunits and in addition two novel O-GlcNAc sites on murine Hsp90β, of which one corresponds to a previously described phosphorylation site.
Notes: 1535-9484
Overath, Thorsten
Kuckelkorn, Ulrike
Henklein, Petra
Strehl, Britta
Bonar, David
Kloss, Alexander
Siele, Dagmar
Kloetzel, Peter-Michael
Janek, Katharina
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2012 Aug;11(8):467-77. doi: 10.1074/mcp.M111.015966. Epub 2012 May 3.
Author Address: Institut für Biochemie, Charité-Universitätsmedizin Berlin, 13347 Berlin, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 857
Author: Overgaard, M. T., Sorensen, E. S., Stachowiak, D., Boldt, H. B., Kristensen, L., Sottrup-Jensen, L. and Oxvig, C.
Year: 2003
Title: Complex of pregnancy-associated plasma protein-A and the proform of eosinophil major basic protein. Disulfide structure and carbohydrate attachment
Journal: J Biol Chem
Volume: 278
Issue: 4
Pages: 2106-17
Epub Date: 2002/11/08
Date: Jan 24
Short Title: Complex of pregnancy-associated plasma protein-A and the proform of eosinophil major basic protein. Disulfide structure and carbohydrate attachment
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M208777200
Accession Number: 12421832
Keywords: Amino Acid Sequence
Amino Acids/chemistry
Animals
Blood Proteins/*chemistry/metabolism
Blotting, Western
Carbohydrates/chemistry
Cations
Chromatography, Gel
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Cyanogen Bromide/pharmacology
Cysteine/chemistry
DNA, Complementary/metabolism
Disulfides/chemistry
Electrophoresis, Polyacrylamide Gel
Eosinophil Granule Proteins
Humans
Mass Spectrometry
Molecular Sequence Data
Mutagenesis, Site-Directed
Peptides/chemistry
Pregnancy-Associated Plasma Protein-A/*chemistry/metabolism
Protein Binding
Protein Folding
Protein Structure, Tertiary
Recombinant Fusion Proteins/chemistry
Recombinant Proteins/chemistry
*Ribonucleases
Sequence Homology, Amino Acid
Transfection
Abstract: Pregnancy-associated plasma protein-A (PAPP-A) is a metzincin superfamily metalloproteinase responsible for cleavage of insulin-like growth factor-binding protein-4, thus causing release of bound insulin-like growth factor. PAPP-A is secreted as a dimer of 400 kDa but circulates in pregnancy as a disulfide-bound 500-kDa 2:2 complex with the proform of eosinophil major basic protein (pro-MBP), recently shown to function as a proteinase inhibitor of PAPP-A. Except for PAPP-A2, PAPP-A does not share global similarity with other proteins. Three lin-notch (LNR or LIN-12) modules and five complement control protein modules (also known as SCR modules) have been identified in PAPP-A by sequence similarity with other proteins, but no data are available that allow unambiguous prediction of disulfide bonds of these modules. To establish the connectivities of cysteine residues of the PAPP-A.pro-MBP complex, biochemical analyses of peptides derived from purified protein were performed. The PAPP-A subunit contains a total of 82 cysteine residues, of which 81 have been accounted for. The pro-MBP subunit contains 12 cysteine residues, of which 10 have been accounted for. Within the 2:2 complex, PAPP-A is dimerized by a single disulfide bond; pro-MBP is dimerized by two disulfides, and each PAPP-A subunit is connected to a pro-MBP subunit by two disulfide bonds. All other disulfides are intrachain bridges. We also show that of 13 potential sites for N-linked carbohydrate substitution of the PAPP-A subunit, 11 are occupied. The large number of disulfide bonds of the PAPP-A.pro-MBP complex imposes many restraints on polypeptide folding, and knowledge of the disulfide pattern of PAPP-A will facilitate structural studies based on recombinant expression of individual, putative PAPP-A domains. Furthermore, it will allow rational experimental design of functional studies aimed at understanding the formation of the PAPP-A.pro-MBP complex, as well as the inhibitory mechanism of pro-MBP.
Notes: Overgaard, Michael T
Sorensen, Esben S
Stachowiak, Damian
Boldt, Henning B
Kristensen, Lene
Sottrup-Jensen, Lars
Oxvig, Claus
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2003 Jan 24;278(4):2106-17. doi: 10.1074/jbc.M208777200. Epub 2002 Nov 5.
Author Address: Department of Molecular Biology, Science Park, University of Aarhus, Gustav Wieds Vej 10C, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1284
Author: Owczarek, S., Kiryushko, D., Larsen, M. H., Kastrup, J. S., Gajhede, M., Sandi, C., Berezin, V., Bock, E. and Soroka, V.
Year: 2010
Title: Neuroplastin-55 binds to and signals through the fibroblast growth factor receptor
Journal: Faseb j
Volume: 24
Issue: 4
Pages: 1139-50
Epub Date: 2009/12/03
Date: Apr
Short Title: Neuroplastin-55 binds to and signals through the fibroblast growth factor receptor
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638
DOI: 10.1096/fj.09-140509
Accession Number: 19952283
Keywords: Amino Acid Motifs
Animals
Antidepressive Agents/chemistry/metabolism/pharmacology
Calcium/*metabolism
Cell Line
Crystallography, X-Ray
Enzyme Activation/drug effects/physiology
Humans
Membrane Glycoproteins/chemistry/*metabolism/pharmacology
Neurites/metabolism
Neuronal Plasticity/drug effects/physiology
Peptides/chemistry/*metabolism/pharmacology
Physical Conditioning, Animal
Protein Binding/drug effects/physiology
Protein Structure, Tertiary
Rats
Rats, Wistar
Receptor, Fibroblast Growth Factor, Type 1/*metabolism
Signal Transduction/*physiology
Swimming
Synapses/*metabolism
Abstract: Neuroplastin (Np) is a glycoprotein belonging to the immunoglobulin superfamily of cell adhesion molecules (CAMs) and existing in two isoforms, Np55 and Np65, named according to their molecular weights. The extracellular part of Np65 contains three immunoglobulin (Ig)-like modules (Ig1, Ig2, and Ig3), whereas Np55 lacks the Ig1 module. Of these two isoforms, only Np65 is involved in homophilic interactions resulting in cell adhesion, whereas the role of Np55 is poorly understood. The present study reports for the first time the crystal structure of the ectodomain of Np55 at 1.95-A resolution and demonstrates that Np55 binds to and activates the fibroblast growth factor receptor 1 (FGFR1). Furthermore, we identify a sequence motif in the Ig2 module of Np55 interacting with FGFR1 and show that a synthetic peptide encompassing this motif, termed narpin, binds to and activates FGFR1. We show that both Np55 and the narpin peptide induce neurite outgrowth through FGFR1 activation and that Np55 increases synaptic calcium concentration in an FGFR1-dependent manner. Moreover, we demonstrate that narpin has an antidepressive-like effect in rats subjected to the forced swim test, suggesting that Np55-induced signaling may be involved in synaptic plasticity in vivo. Owczarek, S., Kiryushko, D., Larsen, M. H., Kastrup, J. S., Gajhede, M., Sandi, C., Berezin, V., Bock, E., Soroka, V. Neuroplastin-55 binds to and signals through the fibroblast growth factor receptor.
Notes: 1530-6860
Owczarek, Sylwia
Kiryushko, Darya
Larsen, Marianne Hald
Kastrup, Jette Sandholm
Gajhede, Michael
Sandi, Carmen
Berezin, Vladimir
Bock, Elisabeth
Soroka, Vladislav
Journal Article
Research Support, Non-U.S. Gov't
United States
FASEB J. 2010 Apr;24(4):1139-50. doi: 10.1096/fj.09-140509. Epub 2009 Dec 1.
Author Address: Protein Laboratory, Department of Neuroscience and Pharmacology, Panum Institute, University of Copenhagen, Blegdamsvej 3C, Bldg. 24.2, 2200 Copenhagen, Denmark. sylwia@sund.ku.dk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 508
Author: Oxvig, C., Haaning, J., Højrup, P. and Sottrup-Jensen, L.
Year: 1994
Title: Location and nature of carbohydrate groups in proform of human major basic protein isolated from pregnancy serum
Journal: Biochem Mol Biol Int
Volume: 33
Issue: 2
Pages: 329-36
Epub Date: 1994/05/01
Date: May
Short Title: Location and nature of carbohydrate groups in proform of human major basic protein isolated from pregnancy serum
Alternate Journal: Biochemistry and molecular biology international
ISSN: 1039-9712 (Print)
1039-9712
Accession Number: 7524900
Keywords: Amino Acid Sequence
Blood Proteins/*chemistry/metabolism
Carbohydrate Sequence
Carbohydrates/*analysis
Chromatography, Gel
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Eosinophil Granule Proteins
Female
Glycosaminoglycans/metabolism
Glycosylation
Humans
Molecular Sequence Data
Molecular Weight
Pregnancy/*blood
Pregnancy-Associated Plasma Protein-A/*metabolism
Protein Precursors/blood/*chemistry/metabolism
*Ribonucleases
Abstract: From human pregnancy serum we have isolated the proform of eosinophil major basic protein (proMBP), which forms a complex with pregnancy-associated plasma protein-A (PAPP-A), PAPP-A/proMBP. It is shown that proMBP contains O-linked glycan bound to Ser-24, Thr-25 (fully substituted), and to Thr-23 and Thr-34 (partially substituted). N-linked glycan is bound to Asn-86 and O-linked glycosaminoglycan is bound to Ser-62 (both fully substituted). From the RP-HPLC elution profile and mass spectra of tryptic peptides it is found that proMBP is extremely heterogeneous with respect to glycosylation.
Notes: Oxvig, C
Haaning, J
Højrup, P
Sottrup-Jensen, L
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem Mol Biol Int. 1994 May;33(2):329-36.
Author Address: Department of Molecular Biology, University of Aarhus, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 906
Author: Ozaslan, D., Wang, S., Ahmed, B. A., Kocabas, A. M., McCastlain, J. C., Bene, A. and Kilic, F.
Year: 2003
Title: Glycosyl modification facilitates homo- and hetero-oligomerization of the serotonin transporter. A specific role for sialic acid residues
Journal: J Biol Chem
Volume: 278
Issue: 45
Pages: 43991-4000
Epub Date: 2003/08/29
Date: Nov 7
Short Title: Glycosyl modification facilitates homo- and hetero-oligomerization of the serotonin transporter. A specific role for sialic acid residues
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M306360200
PMCID: PMC3042025
NIHMSID: NIHMS271643
Accession Number: 12944413
Keywords: Animals
CHO Cells
Carrier Proteins/*chemistry/genetics/metabolism
Choriocarcinoma
Cricetinae
Cyclic GMP/pharmacology
Cyclic GMP-Dependent Protein Kinases/metabolism
Female
Gene Expression
Glycosylation
Macromolecular Substances
Membrane Glycoproteins/*chemistry/genetics/metabolism
*Membrane Transport Proteins
Mutagenesis, Site-Directed
N-Acetylneuraminic Acid/chemistry/genetics
*Nerve Tissue Proteins
Nonmuscle Myosin Type IIA/genetics/metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
Phosphorylation
Pregnancy
Protein Conformation
Rats
Serotonin/metabolism
Serotonin Plasma Membrane Transport Proteins
Structure-Activity Relationship
Transfection
Tumor Cells, Cultured
Uterine Neoplasms
Abstract: The serotonin transporter (SERT) is an oligomeric glycoprotein with two sialic acid residues on each of two complex oligosaccharide molecules. In this study, we investigated the contribution of N-glycosyl modification to the structure and function of SERT in two model systems: wild-type SERT expressed in sialic acid-defective Lec4 Chinese hamster ovary (CHO) cells and a mutant form (after site-directed mutagenesis of Asn-208 and Asn-217 to Gln) of SERT, QQ, expressed in parental CHO cells. In both systems, SERT monomers required modification with both complex oligosaccharide residues to associate with each other and to function in homo-oligomeric forms. However, defects in sialylated N-glycans did not alter surface expression of the SERT protein. Furthermore, in heterologous (CHO and Lec4 cells) and endogenous (placental choriocarcinoma JAR cells) expression systems, we tested whether glycosyl modification also manipulates the hetero-oligomeric interactions of SERT, specifically with myosin IIA. SERT is phosphorylated by cGMP-dependent protein kinase G through interactions with anchoring proteins, and myosin is a protein kinase G-anchoring protein. A physical interaction between myosin and SERT was apparent; however, defects in sialylated N-glycans impaired association of SERT with myosin as well as the stimulation of the serotonin uptake function in the cGMP-dependent pathway. We propose that sialylated N-glycans provide a favorable conformation to SERT that allows the transporter to function most efficiently via its protein-protein interactions.
Notes: 1083-351x
Ozaslan, Deniz
Wang, Sophie
Ahmed, Billow A
Kocabas, Arif M
McCastlain, John C
Bene, Anca
Kilic, Fusun
R03 HD053477/HD/NICHD NIH HHS/United States
R03 HD053477-01/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Retracted Publication
J Biol Chem. 2003 Nov 7;278(45):43991-4000. doi: 10.1074/jbc.M306360200. Epub 2003 Aug 27.
Author Address: Department of Biochemistry and Molecular Biology, College of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 759
Author: Pabon, A., Chan, K. W., Sui, J. L., Wu, X., Logothetis, D. E. and Thornhill, W. B.
Year: 2000
Title: Glycosylation of GIRK1 at Asn119 and ROMK1 at Asn117 has different consequences in potassium channel function
Journal: J Biol Chem
Volume: 275
Issue: 39
Pages: 30677-82
Epub Date: 2000/07/13
Date: Sep 29
Short Title: Glycosylation of GIRK1 at Asn119 and ROMK1 at Asn117 has different consequences in potassium channel function
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M005338200
Accession Number: 10889209
Keywords: Amidohydrolases/metabolism
Asparagine/*metabolism
Cloning, Molecular
G Protein-Coupled Inwardly-Rectifying Potassium Channels
Glycoproteins/genetics/*metabolism
Glycosylation
Hexosaminidases/metabolism
Humans
Mutation
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Potassium Channels/genetics/*metabolism
*Potassium Channels, Inwardly Rectifying
Protein Conformation
RNA, Complementary
Receptor, Muscarinic M2
Receptors, Muscarinic
Recombinant Proteins/metabolism
Abstract: GIRK (G protein-gated inward rectifier K(+) channel) proteins play critical functional roles in heart and brain physiology. Using antibodies directed to either GIRK1 or GIRK4, site-directed mutagenesis, and specific glycosidases, we have investigated the effects of glycosylation in the biosynthesis and heteromerization of these proteins expressed in oocytes. Both GIRK1 and GIRK4 have one extracellular consensus N-glycosylation site. Using chimeras between GIRK1 and GIRK4 as well as a GIRK1 N-glycosylation mutant, we report that GIRK1 was glycosylated at Asn(119), whereas GIRK4 was not glycosylated at Asn(132). GIRK1 membrane-spanning domain 1 was required for optimal glycosylation at Asn(119) because a chimera that contained GIRK4 membrane-spanning domain 1 significantly reduced the addition of a carbohydrate structure at this site. This finding may partly account for the reason that GIRK4 is not glycosylated at Asn(132), either as a homomer or when coexpressed with GIRK1. When the GIRK1(N119Q) mutant was coexpressed with GIRK4, the biophysical properties of the heteromeric channel and the magnitude of the agonist-induced currents were similar to those of controls. Thus, N-glycosylation of GIRK1 at Asn(119) does not appear to affect its physical association with GIRK4, the routing of the heteromer to the cell surface, or heteromeric channel function, unlike the dramatic functional effects of N-glycosylation of ROMK1 at Asn(117) (Schwalbe, R. A., Wang, Z., Wible, B. A., and Brown, A. M. (1995) J. Biol. Chem. 270, 15336-15340).
Notes: Pabon, A
Chan, K W
Sui, J L
Wu, X
Logothetis, D E
Thornhill, W B
HL54185/HL/NHLBI NIH HHS/United States
NS29633/NS/NINDS NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2000 Sep 29;275(39):30677-82. doi: 10.1074/jbc.M005338200.
Author Address: Department of Physiology and Biophysics, Mount Sinai School of Medicine, New York, New York 10029, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1693
Author: Pabst, M., Küster, S. K., Wahl, F., Krismer, J., Dittrich, P. S. and Zenobi, R.
Year: 2015
Title: A Microarray-Matrix-assisted Laser Desorption/Ionization-Mass Spectrometry Approach for Site-specific Protein N-glycosylation Analysis, as Demonstrated for Human Serum Immunoglobulin M (IgM)
Journal: Mol Cell Proteomics
Volume: 14
Issue: 6
Pages: 1645-56
Epub Date: 2015/03/25
Date: Jun
Short Title: A Microarray-Matrix-assisted Laser Desorption/Ionization-Mass Spectrometry Approach for Site-specific Protein N-glycosylation Analysis, as Demonstrated for Human Serum Immunoglobulin M (IgM)
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.O114.046748
PMCID: PMC4458726 the microarray for mass spectrometry technology.
Accession Number: 25802287
Keywords: Chromatography, Liquid
Glycosylation
Humans
Immunoglobulin M/blood/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: We demonstrate a new approach for the site-specific identification and characterization of protein N-glycosylation. It is based on a nano-liquid chromatography microarray-matrix assisted laser desorption/ionization-MS platform, which employs droplet microfluidics for on-plate nanoliter reactions. A chromatographic separation of a proteolytic digest is deposited at a high frequency on the microarray. In this way, a short separation run is archived into thousands of nanoliter reaction cavities, and chromatographic peaks are spread over multiple array spots. After fractionation, each other spot is treated with PNGaseF to generate two correlated traces within one run, one with treated spots where glycans are enzymatically released from the peptides, and one containing the intact glycopeptides. Mining for distinct glycosites is performed by searching for the predicted deglycosylated peptides in the treated trace. An identified peptide then leads directly to the position of the "intact" glycopeptide clusters, which are located in the adjacent spots. Furthermore, the deglycosylated peptide can be sequenced efficiently in a simple collision-induced dissociation-MS experiment. We applied the microarray approach to a detailed site-specific glycosylation analysis of human serum IgM. By scanning the treated spots with low-resolution matrix assisted laser desorption/ionization-time-of-flight-MS, we observed all five deglycosylated peptides, including the one originating from the secretory chain. A detailed glycopeptide characterization was then accomplished on the adjacent, untreated spots with high mass resolution and high mass accuracy using a matrix assisted laser desorption ionization-Fourier transform-MS. We present the first detailed and comprehensive mass spectrometric analysis on the glycopeptide level for human polyclonal IgM with high mass accuracy. Besides complex type glycans on Asn 395, 332, 171, and on the J chain, we observed oligomannosidic glycans on Asn 563, Asn 402 and minor amounts of oligomannosidic glycans on the glycosite Asn 171. Furthermore, hybrid type glycans were found on Asn 402, Asn 171 and in traces Asn 332.
Notes: 1535-9484
Pabst, Martin
Küster, Simon Karl
Wahl, Fabian
Krismer, Jasmin
Dittrich, Petra S
Zenobi, Renato
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2015 Jun;14(6):1645-56. doi: 10.1074/mcp.O114.046748. Epub 2015 Mar 23.
Author Address: From the ‡Department of Chemistry and Applied Biosciences, ETH Zürich, Vladimir-Prelog-Weg 3, 8093 Zürich, Switzerland;
§Sigma-Aldrich Chemie GmbH, Industriestrasse 25, 9471 Buchs (SG), Switzerland.
From the ‡Department of Chemistry and Applied Biosciences, ETH Zürich, Vladimir-Prelog-Weg 3, 8093 Zürich, Switzerland; zenobi@org.chem.ethz.ch.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1748
Author: Paddock, C., Zhou, D., Lertkiatmongkol, P., Newman, P. J. and Zhu, J.
Year: 2016
Title: Structural basis for PECAM-1 homophilic binding
Journal: Blood
Volume: 127
Issue: 8
Pages: 1052-61
Epub Date: 2015/12/25
Date: Feb 25
Short Title: Structural basis for PECAM-1 homophilic binding
Alternate Journal: Blood
ISSN: 0006-4971 (Print)
0006-4971
DOI: 10.1182/blood-2015-07-660092
PMCID: PMC4768429
Accession Number: 26702061
Keywords: Amino Acid Sequence
Animals
Cell Adhesion/physiology
Crystallography, X-Ray
Electrophoresis, Polyacrylamide Gel
Endothelial Cells/*metabolism
Humans
Intercellular Junctions/*metabolism
Mice
*Models, Molecular
Molecular Sequence Data
Platelet Endothelial Cell Adhesion Molecule-1/*chemistry/*metabolism
Protein Binding/physiology
Protein Structure, Tertiary
Species Specificity
Transfection
Abstract: Platelet endothelial cell adhesion molecule-1 (PECAM-1) is a 130-kDa member of the immunoglobulin gene superfamily (IgSF) that is present on the surface of circulating platelets and leukocytes, and highly expressed at the junctions of confluent endothelial cell monolayers. PECAM-1-mediated homophilic interactions, known to be mediated by its 2 amino-terminal immunoglobulin homology domains, are essential for concentrating PECAM-1 at endothelial cell intercellular junctions, where it functions to facilitate diapedesis, maintain vascular integrity, and transmit survival signals into the cell. Given the importance of PECAM-1-mediated homophilic interactions in mediating each of these cell physiological events, and to reveal the nature and orientation of the PECAM-1-PECAM-1 homophilic-binding interface, we undertook studies aimed at determining the crystal structure of the PECAM-1 homophilic-binding domain, which is composed of amino-terminal immunoglobulin homology domains 1 and 2 (IgD1 and IgD2). The crystal structure revealed that both IgD1 and IgD2 exhibit a classical IgSF fold, having a β-sandwich topology formed by 2 sheets of antiparallel β strands stabilized by the hallmark disulfide bond between the B and F strands. Interestingly, despite previous assignment to the C2 class of immunoglobulin-like domains, the structure of IgD1 reveals that it actually belongs to the I2 set of IgSF folds. Both IgD1 and IgD2 participate importantly in the formation of the trans homophilic-binding interface, with a total buried interface area of >2300 Å(2). These and other unique structural features of PECAM-1 allow for the development of an atomic-level model of the interactions that PECAM-1 forms during assembly of endothelial cell intercellular junctions.
Notes: 1528-0020
Paddock, Cathy
Zhou, Dongwen
Lertkiatmongkol, Panida
Newman, Peter J
Zhu, Jieqing
R01 HL040926/HL/NHLBI NIH HHS/United States
R01 HL131836/HL/NHLBI NIH HHS/United States
R56 HL122985/HL/NHLBI NIH HHS/United States
HL40926/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Blood. 2016 Feb 25;127(8):1052-61. doi: 10.1182/blood-2015-07-660092. Epub 2015 Dec 23.
Author Address: Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, WI; and.
Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, WI; and Department of Pharmacology and.
Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, WI; and Department of Biochemistry, Medical College of Wisconsin, Milwaukee, WI.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1058
Author: Pak, J. E., Arnoux, P., Zhou, S., Sivarajah, P., Satkunarajah, M., Xing, X. and Rini, J. M.
Year: 2006
Title: X-ray crystal structure of leukocyte type core 2 beta1,6-N-acetylglucosaminyltransferase. Evidence for a convergence of metal ion-independent glycosyltransferase mechanism
Journal: J Biol Chem
Volume: 281
Issue: 36
Pages: 26693-701
Epub Date: 2006/07/11
Date: Sep 8
Short Title: X-ray crystal structure of leukocyte type core 2 beta1,6-N-acetylglucosaminyltransferase. Evidence for a convergence of metal ion-independent glycosyltransferase mechanism
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M603534200
Accession Number: 16829524
Keywords: Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray
Ions/*chemistry
Metals/*chemistry
Mice
Models, Molecular
Molecular Sequence Data
N-Acetylglucosaminyltransferases/*chemistry/genetics/metabolism
Oligosaccharides/chemistry/metabolism
Polysaccharides/metabolism
*Protein Structure, Tertiary
Recombinant Fusion Proteins/chemistry/genetics/metabolism
Sequence Alignment
Substrate Specificity
Uridine Diphosphate Glucuronic Acid/metabolism
Abstract: Leukocyte type core 2 beta1,6-N-acetylglucosaminyltransferase (C2GnT-L) is a key enzyme in the biosynthesis of branched O-glycans. It is an inverting, metal ion-independent family 14 glycosyltransferase that catalyzes the formation of the core 2 O-glycan (Galbeta1-3[GlcNAcbeta1-6]GalNAc-O-Ser/Thr) from its donor and acceptor substrates, UDP-GlcNAc and the core 1 O-glycan (Galbeta1-3GalNAc-O-Ser/Thr), respectively. Reported here are the x-ray crystal structures of murine C2GnT-L in the absence and presence of the acceptor substrate Galbeta1-3GalNAc at 2.0 and 2.7A resolution, respectively. C2GnT-L was found to possess the GT-A fold; however, it lacks the characteristic metal ion binding DXD motif. The Galbeta1-3GalNAc complex defines the determinants of acceptor substrate binding and shows that Glu-320 corresponds to the structurally conserved catalytic base found in other inverting GT-A fold glycosyltransferases. Comparison of the C2GnT-L structure with that of other GT-A fold glycosyltransferases further suggests that Arg-378 and Lys-401 serve to electrostatically stabilize the nucleoside diphosphate leaving group, a role normally played by metal ion in GT-A structures. The use of basic amino acid side chains in this way is strikingly similar to that seen in a number of metal ion-independent GT-B fold glycosyltransferases and suggests a convergence of catalytic mechanism shared by both GT-A and GT-B fold glycosyltransferases.
Notes: Pak, John E
Arnoux, Pascal
Zhou, Sihong
Sivarajah, Prashanth
Satkunarajah, Malathy
Xing, Xuekun
Rini, James M
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Sep 8;281(36):26693-701. doi: 10.1074/jbc.M603534200. Epub 2006 Jul 7.
Author Address: Department of Molecular, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1211
Author: Pal, A., Kraetzner, R., Gruene, T., Grapp, M., Schreiber, K., Grønborg, M., Urlaub, H., Becker, S., Asif, A. R., Gärtner, J., Sheldrick, G. M. and Steinfeld, R.
Year: 2009
Title: Structure of tripeptidyl-peptidase I provides insight into the molecular basis of late infantile neuronal ceroid lipofuscinosis
Journal: J Biol Chem
Volume: 284
Issue: 6
Pages: 3976-84
Epub Date: 2008/11/29
Date: Feb 6
Short Title: Structure of tripeptidyl-peptidase I provides insight into the molecular basis of late infantile neuronal ceroid lipofuscinosis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M806947200
Accession Number: 19038966
Keywords: Aminopeptidases
Cell Line
Crystallography, X-Ray
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases
Endopeptidases/*chemistry/genetics
Glycosylation
Humans
Mutation
Neuronal Ceroid-Lipofuscinoses/*enzymology/genetics
*Protein Folding
Protein Structure, Tertiary/genetics
Serine Proteases
Structural Homology, Protein
Structure-Activity Relationship
Abstract: Late infantile neuronal ceroid lipofuscinosis, a fatal neurodegenerative disease of childhood, is caused by mutations in the TPP1 gene that encodes tripeptidyl-peptidase I. We show that purified TPP1 requires at least partial glycosylation for in vitro autoprocessing and proteolytic activity. We crystallized the fully glycosylated TPP1 precursor under conditions that implied partial autocatalytic cleavage between the prosegment and the catalytic domain. X-ray crystallographic analysis at 2.35 angstroms resolution reveals a globular structure with a subtilisin-like fold, a Ser475-Glu272-Asp360 catalytic triad, and an octahedrally coordinated Ca2+-binding site that are characteristic features of the S53 sedolisin family of peptidases. In contrast to other S53 peptidases, the TPP1 structure revealed steric constraints on the P4 substrate pocket explaining its preferential cleavage of tripeptides from the unsubstituted N terminus of proteins. Two alternative conformations of the catalytic Asp276 are associated with the activation status of TPP1. 28 disease-causing missense mutations are analyzed in the light of the TPP1 structure providing insight into the molecular basis of late infantile neuronal ceroid lipofuscinosis.
Notes: Pal, Aritra
Kraetzner, Ralph
Gruene, Tim
Grapp, Marcel
Schreiber, Kathrin
Grønborg, Mads
Urlaub, Henning
Becker, Stefan
Asif, Abdul R
Gärtner, Jutta
Sheldrick, George M
Steinfeld, Robert
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2009 Feb 6;284(6):3976-84. doi: 10.1074/jbc.M806947200. Epub 2008 Nov 26.
Author Address: Department of Structural Chemistry, University of Göttingen, Tammannstrasse 4, 34077 Göttingen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1502
Author: Palakodeti, A., Sandstrom, A., Sundaresan, L., Harly, C., Nedellec, S., Olive, D., Scotet, E., Bonneville, M. and Adams, E. J.
Year: 2012
Title: The molecular basis for modulation of human Vγ9Vδ2 T cell responses by CD277/butyrophilin-3 (BTN3A)-specific antibodies
Journal: J Biol Chem
Volume: 287
Issue: 39
Pages: 32780-90
Epub Date: 2012/08/01
Date: Sep 21
Short Title: The molecular basis for modulation of human Vγ9Vδ2 T cell responses by CD277/butyrophilin-3 (BTN3A)-specific antibodies
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.384354
PMCID: PMC3463320
Accession Number: 22846996
Keywords: Antibodies/chemistry/*immunology
Antigens, CD/chemistry/genetics/*immunology
Butyrophilins
Humans
*Lymphocyte Activation
Protein Isoforms/chemistry/genetics/immunology
Protein Structure, Tertiary
Receptors, Antigen, T-Cell, gamma-delta/chemistry/genetics/*immunology
Structural Homology, Protein
Structure-Activity Relationship
T-Lymphocytes/chemistry/*immunology
Abstract: Human Vγ9Vδ2 T cells are well known for their rapid and potent response to infection and tumorigenesis when in the presence of endogenous or exogenous phosphoisoprenoids. However, the molecular mechanisms behind the activation of this γδ T cell population remains unclear. Evidence pointing to a role for the CD277/butyrophilin-3 (BTN3A) molecules in this response led us to investigate the structures of these molecules and their modifications upon binding to an agonist antibody (20.1) that mimics phosphoisoprenoid-mediated Vγ9Vδ2 activation and an antagonist antibody (103.2) that inhibits this reactivity. We find that the three BTN3A isoforms: BTN3A1, BTN3A2, and BTN3A3, have high structural homology to the B7 superfamily of proteins and exist as V-shaped homodimers in solution, associating through the membrane proximal C-type Ig domain. The 20.1 and 103.2 antibodies bind to separate epitopes on the BTN3A Ig-V domain with high affinity but likely with different valencies based on their binding orientation. These structures directly complement functional studies of this system that demonstrate that BTN3A1 is necessary for Vγ9Vδ2 activation and begin to unravel the extracellular events that occur during stimulation through the Vγ9Vδ2 T cell receptor.
Notes: 1083-351x
Palakodeti, Aparna
Sandstrom, Andrew
Sundaresan, Lakshmi
Harly, Christelle
Nedellec, Steven
Olive, Daniel
Scotet, Emmanuel
Bonneville, Marc
Adams, Erin J
R01 AI073922/AI/NIAID NIH HHS/United States
R01AI073922/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Sep 21;287(39):32780-90. doi: 10.1074/jbc.M112.384354. Epub 2012 Jul 30.
Author Address: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois 60637, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1594
Author: Palaniappan, K. K., Hangauer, M. J., Smith, T. J., Smart, B. P., Pitcher, A. A., Cheng, E. H., Bertozzi, C. R. and Boyce, M.
Year: 2013
Title: A chemical glycoproteomics platform reveals O-GlcNAcylation of mitochondrial voltage-dependent anion channel 2
Journal: Cell Rep
Volume: 5
Issue: 2
Pages: 546-52
Epub Date: 2013/10/15
Date: Oct 31
Short Title: A chemical glycoproteomics platform reveals O-GlcNAcylation of mitochondrial voltage-dependent anion channel 2
Alternate Journal: Cell reports
DOI: 10.1016/j.celrep.2013.08.048
PMCID: PMC3869705
NIHMSID: NIHMS524960
Accession Number: 24120863
Keywords: Acetylglucosamine/metabolism
Animals
Cell Line
Electrophoresis, Gel, Pulsed-Field
Fluorescent Dyes/chemistry
Glycoproteins/metabolism
Glycosylation
HEK293 Cells
Humans
Jurkat Cells
Mice
Mitochondria/*metabolism
Proteomics
Substrate Specificity
Voltage-Dependent Anion Channel 2/deficiency/genetics/*metabolism
Abstract: Protein modification by O-linked β-N-acetylglucosamine (O-GlcNAc) is a critical cell signaling modality, but identifying signal-specific O-GlcNAcylation events remains a significant experimental challenge. Here, we describe a method for visualizing and analyzing organelle- and stimulus-specific O-GlcNAcylated proteins and use it to identify the mitochondrial voltage-dependent anion channel 2 (VDAC2) as an O-GlcNAc substrate. VDAC2(-/-) cells resist the mitochondrial dysfunction and apoptosis caused by global O-GlcNAc perturbation, demonstrating a functional connection between O-GlcNAc signaling and mitochondrial physiology through VDAC2. More broadly, our method will enable the discovery of signal-specific O-GlcNAcylation events in a wide array of experimental contexts.
Notes: 2211-1247
Palaniappan, Krishnan K
Hangauer, Matthew J
Smith, Timothy J
Smart, Brian P
Pitcher, Austin A
Cheng, Emily H
Bertozzi, Carolyn R
Boyce, Michael
Howard Hughes Medical Institute/United States
R01 GM066047/GM/NIGMS NIH HHS/United States
GM066047/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Cell Rep. 2013 Oct 31;5(2):546-52. doi: 10.1016/j.celrep.2013.08.048. Epub 2013 Oct 10.
Author Address: Department of Chemistry, University of California, Berkeley, Berkeley, CA 94720, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 979
Author: Pallarès, I., Bonet, R., García-Castellanos, R., Ventura, S., Avilés, F. X., Vendrell, J. and Gomis-Rüth, F. X.
Year: 2005
Title: Structure of human carboxypeptidase A4 with its endogenous protein inhibitor, latexin
Journal: Proc Natl Acad Sci U S A
Volume: 102
Issue: 11
Pages: 3978-83
Epub Date: 2005/03/02
Date: Mar 15
Short Title: Structure of human carboxypeptidase A4 with its endogenous protein inhibitor, latexin
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0500678102
PMCID: PMC554826
Accession Number: 15738388
Keywords: Amino Acid Sequence
Antigens/*chemistry/metabolism
Carboxypeptidases A/*chemistry/metabolism
Crystallography
Humans
Molecular Sequence Data
Protein Structure, Tertiary
Sequence Alignment
Abstract: The only endogenous protein inhibitor known for metallocarboxypeptidases (MCPs) is latexin, a 25-kDa protein discovered in the rat brain. Latexin, alias endogenous carboxypeptidase inhibitor, inhibits human CPA4 (hCPA4), whose expression is induced in prostate cancer cells after treatment with histone deacetylase inhibitors. hCPA4 is a member of the A/B subfamily of MCPs and displays the characteristic alpha/beta-hydrolase fold. Human latexin consists of two topologically equivalent subdomains, reminiscent of cystatins, consisting of an alpha-helix enveloped by a curved beta-sheet. These subdomains are packed against each other through the helices and linked by a connecting segment encompassing a third alpha-helix. The enzyme is bound at the interface of these subdomains. The complex occludes a large contact surface but makes rather few contacts, despite a nanomolar inhibition constant. This low specificity explains the flexibility of latexin in inhibiting all vertebrate A/B MCPs tested, even across species barriers. In contrast, modeling studies reveal why the N/E subfamily of MCPs and invertebrate A/B MCPs are not inhibited. Major differences in the loop segments shaping the border of the funnel-like access to the protease active site impede complex formation with latexin. Several sequences ascribable to diverse tissues and organs have been identified in vertebrate genomes as being highly similar to latexin. They are proposed to constitute the latexin family of potential inhibitors. Because they are ubiquitous, latexins could represent for vertebrate A/B MCPs the counterparts of tissue inhibitors of metalloproteases for matrix metalloproteinases.
Notes: 1091-6490
Pallarès, Irantzu
Bonet, Roman
García-Castellanos, Raquel
Ventura, Salvador
Avilés, Francesc X
Vendrell, Josep
Gomis-Rüth, F Xavier
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2005 Mar 15;102(11):3978-83. doi: 10.1073/pnas.0500678102. Epub 2005 Feb 28.
Author Address: Institut de Biologia Molecular de Barcelona, Centro de Investigación y Desarrollo-Consejo Superior de Investigaciones Científicas, C/Jordi Girona, 18-26, E-08034 Barcelona, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 682
Author: Pan, C. J., Lei, K. J. and Chou, J. Y.
Year: 1998
Title: Asparagine-linked oligosaccharides are localized to a luminal hydrophilic loop in human glucose-6-phosphatase
Journal: J Biol Chem
Volume: 273
Issue: 34
Pages: 21658-62
Epub Date: 1998/08/15
Date: Aug 21
Short Title: Asparagine-linked oligosaccharides are localized to a luminal hydrophilic loop in human glucose-6-phosphatase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.34.21658
Accession Number: 9705299
Keywords: Amino Acid Sequence
Asparagine/*chemistry
Blotting, Western
Cell Line
Glucose-6-Phosphatase/*chemistry/genetics
Glycosylation
Humans
Molecular Sequence Data
Mutagenesis, Site-Directed
Oligosaccharides/*chemistry
Protein Structure, Secondary
Structure-Activity Relationship
Abstract: Deficiency of glucose-6-phosphatase (G6Pase), an endoplasmic reticulum transmembrane glycoprotein, causes glycogen storage disease type 1a. We have recently shown that human G6Pase contains an odd number of transmembrane segments, supporting a nine-transmembrane helical model for this enzyme. Sequence analysis predicts the presence of three potential asparagine (N)-linked glycosylation sites, N96TS, N203AS, and N276SS, conserved among mammalian G6Pases. According to this model, Asn96, located in a 37-residue luminal loop, is a potential acceptor for oligosaccharides, whereas Asn203 and Asn276, located in a 12-residue cytoplasmic loop and helix 7, respectively, would not be utilized for this purpose. We therefore characterized mutant G6Pases lacking one, two, or all three potential N-linked glycosylation sites. Western blot and in vitro translation studies showed that G6Pase is glycosylated only at Asn96, further validating the nine-transmembrane topology model. Substituting Asn96 with an Ala (N96A) moderately reduced enzymatic activity and had no effect on G6Pase synthesis or degradation, suggesting that oligosaccharide chains do not play a major role in protecting the enzyme from proteolytic degradation. In contrast, mutation of Asn276 to an Ala (N276A) destabilized the enzyme and markedly reduced enzymatic activity. We present additional evidence suggesting that the integrity of transmembrane helices is essential for G6Pase stability and catalytic activity.
Notes: Pan, C J
Lei, K J
Chou, J Y
Journal Article
United States
J Biol Chem. 1998 Aug 21;273(34):21658-62. doi: 10.1074/jbc.273.34.21658.
Author Address: Heritable Disorders Branch, NICHD, National Institutes of Health, Bethesda, Maryland 20892-1830, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2007
Author: Pan, X., Li, Z., Huang, X., Huang, G., Gao, S., Shen, H., Liu, L., Lei, J. and Yan, N.
Year: 2019
Title: Molecular basis for pore blockade of human Na(+) channel Na(v)1.2 by the μ-conotoxin KIIIA
Journal: Science
Volume: 363
Issue: 6433
Pages: 1309-1313
Epub Date: 2019/02/16
Date: Mar 22
Short Title: Molecular basis for pore blockade of human Na(+) channel Na(v)1.2 by the μ-conotoxin KIIIA
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.aaw2999
Accession Number: 30765605
Keywords: Amino Acid Sequence
Conotoxins/*chemistry
Cryoelectron Microscopy
HEK293 Cells
Humans
NAV1.2 Voltage-Gated Sodium Channel/*chemistry
Protein Conformation
Voltage-Gated Sodium Channel Blockers/*chemistry
Voltage-Gated Sodium Channel beta-2 Subunit/chemistry
Abstract: The voltage-gated sodium channel Na(v)1.2 is responsible for the initiation and propagation of action potentials in the central nervous system. We report the cryo-electron microscopy structure of human Na(v)1.2 bound to a peptidic pore blocker, the μ-conotoxin KIIIA, in the presence of an auxiliary subunit, β2, to an overall resolution of 3.0 angstroms. The immunoglobulin domain of β2 interacts with the shoulder of the pore domain through a disulfide bond. The 16-residue KIIIA interacts with the extracellular segments in repeats I to III, placing Lys(7) at the entrance to the selectivity filter. Many interacting residues are specific to Na(v)1.2, revealing a molecular basis for KIIIA specificity. The structure establishes a framework for the rational design of subtype-specific blockers for Na(v) channels.
Notes: 1095-9203
Pan, Xiaojing
Orcid: 0000-0003-1882-5361
Li, Zhangqiang
Orcid: 0000-0002-3296-2293
Huang, Xiaoshuang
Orcid: 0000-0002-3329-5496
Huang, Gaoxingyu
Gao, Shuai
Orcid: 0000-0002-8734-2489
Shen, Huaizong
Orcid: 0000-0002-1216-5393
Liu, Lei
Orcid: 0000-0001-6290-8602
Lei, Jianlin
Orcid: 0000-0002-9384-8742
Yan, Nieng
Orcid: 0000-0003-4829-7416
Journal Article
Research Support, Non-U.S. Gov't
United States
Science. 2019 Mar 22;363(6433):1309-1313. doi: 10.1126/science.aaw2999. Epub 2019 Feb 14.
Author Address: State Key Laboratory of Membrane Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China.
Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China.
Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China.
Department of Chemistry, Tsinghua University, Beijing 100084, China.
Technology Center for Protein Sciences, Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China.
State Key Laboratory of Membrane Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China. nyan@princeton.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1966
Author: Pan, X., Li, Z., Zhou, Q., Shen, H., Wu, K., Huang, X., Chen, J., Zhang, J., Zhu, X., Lei, J., Xiong, W., Gong, H., Xiao, B. and Yan, N.
Year: 2018
Title: Structure of the human voltage-gated sodium channel Na(v)1.4 in complex with β1
Journal: Science
Volume: 362
Issue: 6412
Epub Date: 2018/09/08
Date: Oct 19
Short Title: Structure of the human voltage-gated sodium channel Na(v)1.4 in complex with β1
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.aau2486
Accession Number: 30190309
Keywords: Allosteric Regulation
Amino Acid Sequence
Channelopathies/genetics/metabolism
Cryoelectron Microscopy
Drug Discovery
HEK293 Cells
Humans
Mutation
NAV1.4 Voltage-Gated Sodium Channel/*chemistry/genetics/ultrastructure
Protein Domains
Voltage-Gated Sodium Channel beta-4 Subunit/*chemistry/genetics/ultrastructure
Abstract: Voltage-gated sodium (Na(v)) channels, which are responsible for action potential generation, are implicated in many human diseases. Despite decades of rigorous characterization, the lack of a structure of any human Na(v) channel has hampered mechanistic understanding. Here, we report the cryo-electron microscopy structure of the human Na(v)1.4-β1 complex at 3.2-Å resolution. Accurate model building was made for the pore domain, the voltage-sensing domains, and the β1 subunit, providing insight into the molecular basis for Na(+) permeation and kinetic asymmetry of the four repeats. Structural analysis of reported functional residues and disease mutations corroborates an allosteric blocking mechanism for fast inactivation of Na(v) channels. The structure provides a path toward mechanistic investigation of Na(v) channels and drug discovery for Na(v) channelopathies.
Notes: 1095-9203
Pan, Xiaojing
Orcid: 0000-0003-1882-5361
Li, Zhangqiang
Orcid: 0000-0002-3296-2293
Zhou, Qiang
Orcid: 0000-0002-6237-8813
Shen, Huaizong
Orcid: 0000-0002-1216-5393
Wu, Kun
Orcid: 0000-0002-3075-8272
Huang, Xiaoshuang
Chen, Jiaofeng
Zhang, Juanrong
Zhu, Xuechen
Orcid: 0000-0001-5003-8531
Lei, Jianlin
Orcid: 0000-0002-9384-8742
Xiong, Wei
Gong, Haipeng
Orcid: 0000-0002-5532-1640
Xiao, Bailong
Yan, Nieng
Orcid: 0000-0003-4829-7416
Journal Article
Research Support, Non-U.S. Gov't
United States
Science. 2018 Oct 19;362(6412):eaau2486. doi: 10.1126/science.aau2486. Epub 2018 Sep 6.
Author Address: State Key Laboratory of Membrane Biology, Tsinghua University, Beijing 100084, China.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua University, Beijing 100084, China.
Tsinghua-Peking Joint Center for Life Sciences, Tsinghua University, Beijing 100084, China.
School of Medicine, Tsinghua University, Beijing 100084, China.
School of Life Sciences, Tsinghua University, Beijing 100084, China.
School of Pharmaceutical Sciences, Tsinghua University, Beijing 100084, China.
Technology Center for Protein Sciences, Ministry of Education Key Laboratory of Protein Sciences, Tsinghua University, Beijing 100084, China.
State Key Laboratory of Membrane Biology, Tsinghua University, Beijing 100084, China. nyan@princeton.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1350
Author: Pang, S. S., Berry, R., Chen, Z., Kjer-Nielsen, L., Perugini, M. A., King, G. F., Wang, C., Chew, S. H., La Gruta, N. L., Williams, N. K., Beddoe, T., Tiganis, T., Cowieson, N. P., Godfrey, D. I., Purcell, A. W., Wilce, M. C., McCluskey, J. and Rossjohn, J.
Year: 2010
Title: The structural basis for autonomous dimerization of the pre-T-cell antigen receptor
Journal: Nature
Volume: 467
Issue: 7317
Pages: 844-8
Epub Date: 2010/10/15
Date: Oct 14
Short Title: The structural basis for autonomous dimerization of the pre-T-cell antigen receptor
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature09448
Accession Number: 20944746
Keywords: Crystallography, X-Ray
Gene Rearrangement, T-Lymphocyte/genetics
Humans
Models, Molecular
Mutation
Protein Folding
*Protein Multimerization
Protein Structure, Tertiary
Receptors, Antigen, T-Cell/*chemistry/genetics/*metabolism
Receptors, Antigen, T-Cell, alpha-beta/chemistry/metabolism
Signal Transduction
Solutions
T-Lymphocytes/cytology/immunology/metabolism
Abstract: The pre-T-cell antigen receptor (pre-TCR), expressed by immature thymocytes, has a pivotal role in early T-cell development, including TCR β-selection, survival and proliferation of CD4(-)CD8(-) double-negative thymocytes, and subsequent αβ T-cell lineage differentiation. Whereas αβTCR ligation by the peptide-loaded major histocompatibility complex initiates T-cell signalling, pre-TCR-induced signalling occurs by means of a ligand-independent dimerization event. The pre-TCR comprises an invariant α-chain (pre-Tα) that pairs with any TCR β-chain (TCRβ) following successful TCR β-gene rearrangement. Here we provide the basis of pre-Tα-TCRβ assembly and pre-TCR dimerization. The pre-Tα chain comprised a single immunoglobulin-like domain that is structurally distinct from the constant (C) domain of the TCR α-chain; nevertheless, the mode of association between pre-Tα and TCRβ mirrored that mediated by the Cα-Cβ domains of the αβTCR. The pre-TCR had a propensity to dimerize in solution, and the molecular envelope of the pre-TCR dimer correlated well with the observed head-to-tail pre-TCR dimer. This mode of pre-TCR dimerization enabled the pre-Tα domain to interact with the variable (V) β domain through residues that are highly conserved across the Vβ and joining (J) β gene families, thus mimicking the interactions at the core of the αβTCR's Vα-Vβ interface. Disruption of this pre-Tα-Vβ dimer interface abrogated pre-TCR dimerization in solution and impaired pre-TCR expression on the cell surface. Accordingly, we provide a mechanism of pre-TCR self-association that allows the pre-Tα chain to simultaneously 'sample' the correct folding of both the V and C domains of any TCR β-chain, regardless of its ultimate specificity, which represents a critical checkpoint in T-cell development. This unusual dual-chaperone-like sensing function of pre-Tα represents a unique mechanism in nature whereby developmental quality control regulates the expression and signalling of an integral membrane receptor complex.
Notes: 1476-4687
Pang, Siew Siew
Berry, Richard
Chen, Zhenjun
Kjer-Nielsen, Lars
Perugini, Matthew A
King, Glenn F
Wang, Christina
Chew, Sock Hui
La Gruta, Nicole L
Williams, Neal K
Beddoe, Travis
Tiganis, Tony
Cowieson, Nathan P
Godfrey, Dale I
Purcell, Anthony W
Wilce, Matthew C J
McCluskey, James
Rossjohn, Jamie
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2010 Oct 14;467(7317):844-8. doi: 10.1038/nature09448.
Author Address: The Protein Crystallography Unit, Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2057
Author: Pap, A., Tasnadi, E., Medzihradszky, K. F. and Darula, Z.
Year: 2020
Title: Novel O-linked sialoglycan structures in human urinary glycoproteins
Journal: Mol Omics
Volume: 16
Issue: 2
Pages: 156-164
Epub Date: 2020/02/06
Date: Apr 1
Short Title: Novel O-linked sialoglycan structures in human urinary glycoproteins
Alternate Journal: Molecular omics
ISSN: 2515-4184
DOI: 10.1039/c9mo00160c
Accession Number: 32022078
Keywords: Chromatography, Liquid
Computational Biology/*methods
Glycoproteins/*chemistry/*urine
Glycosylation
Humans
Search Engine
Tandem Mass Spectrometry
Abstract: Glycopeptides represent cross-linked structures between chemically and physically different biomolecules. Mass spectrometric analysis of O-glycopeptides may reveal the identity of the peptide, the composition of the glycan and even the connection between certain sugar units, but usually only the combination of different MS/MS techniques provides sufficient information for reliable assignment. Currently, HCD analysis followed by diagnostic sugar fragment-triggered ETD or EThcD experiments is the most promising data acquisition protocol. However, the information content of the different MS/MS data is handled separately by search engines. We are convinced that these data should be used in concert, as we demonstrate in the present study. First, glycopeptides bearing the most common glycans can be identified from EThcD and/or HCD data. Then, searching for Y(0) (the gas-phase deglycosylated peptide) in HCD spectra, the potential glycoforms of these glycopeptides could be lined up. Finally, these spectra and the corresponding EThcD data can be used to verify or discard the tentative assignments and to obtain further structural information about the glycans. We present 18 novel human urinary sialoglycan structures deciphered using this approach. To accomplish this in an automated fashion further software development is necessary.
Notes: 2515-4184
Pap, Adam
Tasnadi, Ervin
Medzihradszky, Katalin F
Darula, Zsuzsanna
Journal Article
Research Support, Non-U.S. Gov't
England
Mol Omics. 2020 Apr 1;16(2):156-164. doi: 10.1039/c9mo00160c. Epub 2020 Feb 5.
Author Address: Laboratory of Proteomics Research, Biological Research Centre, Temesvari krt. 62, H-6726 Szeged, Hungary. darula.zsuzsanna@brc.hu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2043
Author: Paredes, F., Williams, H. C., Quintana, R. A. and San Martin, A.
Year: 2020
Title: Mitochondrial Protein Poldip2 (Polymerase Delta Interacting Protein 2) Controls Vascular Smooth Muscle Differentiated Phenotype by O-Linked GlcNAc (N-Acetylglucosamine) Transferase-Dependent Inhibition of a Ubiquitin Proteasome System
Journal: Circ Res
Volume: 126
Issue: 1
Pages: 41-56
Epub Date: 2019/10/28
Date: Jan 3
Short Title: Mitochondrial Protein Poldip2 (Polymerase Delta Interacting Protein 2) Controls Vascular Smooth Muscle Differentiated Phenotype by O-Linked GlcNAc (N-Acetylglucosamine) Transferase-Dependent Inhibition of a Ubiquitin Proteasome System
Alternate Journal: Circulation research
ISSN: 0009-7330 (Print)
0009-7330
DOI: 10.1161/circresaha.119.315932
PMCID: PMC7251964
NIHMSID: NIHMS1549111
Accession Number: 31656131
Keywords: Animals
Cell Differentiation
Cells, Cultured
Gene Expression Regulation
Humans
Hyperplasia
Kruppel-Like Transcription Factors/biosynthesis/genetics
Male
Mice
Mice, Inbred C57BL
Mitochondria/metabolism
Mitochondrial Proteins/deficiency/genetics/*physiology
Muscle, Smooth, Vascular/*cytology
Myocytes, Smooth Muscle/cytology/*metabolism
Neointima
Nuclear Proteins/biosynthesis/deficiency/genetics/*physiology
Phenotype
Proteasome Endopeptidase Complex/metabolism
Serum Response Factor/biosynthesis/genetics
Trans-Activators/biosynthesis/genetics
Ubiquitin/metabolism
*aneurysm
*hyperplasia
*metabolism
*mitochondria
*muscle
*smooth
*vascular
article.
Abstract: RATIONALE: The mitochondrial Poldip2 (protein polymerase interacting protein 2) is required for the activity of the tricarboxylic acid cycle. As a consequence, Poldip2 deficiency induces metabolic reprograming with repressed mitochondrial respiration and increased glycolytic activity. Though homozygous deletion of Poldip2 is lethal, heterozygous mice are viable and show protection against aneurysm and injury-induced neointimal hyperplasia, diseases linked to loss of vascular smooth muscle differentiation. Thus, we hypothesize that the metabolic reprograming induced by Poldip2 deficiency controls VSMC differentiation. OBJECTIVE: To determine the role of Poldip2-mediated metabolic reprograming in phenotypic modulation of VSMC. METHODS AND RESULTS: We show that Poldip2 deficiency in vascular smooth muscle in vitro and in vivo induces the expression of the SRF (serum response factor), myocardin, and MRTFA (myocardin-related transcription factor A) and dramatically represses KLF4 (Krüppel-like factor 4). Consequently, Poldip2-deficient VSMC and mouse aorta express high levels of contractile proteins and, more significantly, these cells do not dedifferentiate nor acquire macrophage-like characteristics when exposed to cholesterol or PDGF (platelet-derived growth factor). Regarding the mechanism, we found that Poldip2 deficiency upregulates the hexosamine biosynthetic pathway and OGT (O-linked N-acetylglucosamine transferase)-mediated protein O-GlcNAcylation. Increased protein glycosylation causes the inhibition of a nuclear ubiquitin proteasome system responsible for SRF stabilization and KLF4 repression and is required for the establishment of the differentiated phenotype in Poldip2-deficient cells. CONCLUSIONS: Our data show that Poldip2 deficiency induces a highly differentiated phenotype in VSMCs through a mechanism that involves regulation of metabolism and proteostasis. Additionally, our study positions mitochondria-initiated signaling as key element of the VSMC differentiation programs that can be targeted to modulate VSMC phenotype during vascular diseases.
Notes: 1524-4571
Paredes, Felipe
Williams, Holly C
Quintana, Raymundo A
San Martin, Alejandra
P01 HL095070/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Circ Res. 2020 Jan 3;126(1):41-56. doi: 10.1161/CIRCRESAHA.119.315932. Epub 2019 Oct 28.
Author Address: From the Department of Medicine, Division of Cardiology, Emory University, Atlanta, GA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 345
Author: Parekh, R. B., Dwek, R. A., Thomas, J. R., Opdenakker, G., Rademacher, T. W., Wittwer, A. J., Howard, S. C., Nelson, R., Siegel, N. R., Jennings, M. G. and et al.
Year: 1989
Title: Cell-type-specific and site-specific N-glycosylation of type I and type II human tissue plasminogen activator
Journal: Biochemistry
Volume: 28
Issue: 19
Pages: 7644-62
Epub Date: 1989/09/19
Date: Sep 19
Short Title: Cell-type-specific and site-specific N-glycosylation of type I and type II human tissue plasminogen activator
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00445a021
Accession Number: 2514791
Keywords: Amino Acids/analysis
Animals
Carbohydrate Conformation
Carbohydrate Sequence
Cells, Cultured
Chromatography, Gel
Gene Expression
Glycopeptides/isolation & purification
Glycosylation
Humans
Hydrolysis
Methylation
Mice
Molecular Sequence Data
Oligosaccharides/genetics/isolation & purification/pharmacokinetics
Protein Processing, Post-Translational
Tissue Plasminogen Activator/*analysis/isolation & purification/*metabolism
Tumor Cells, Cultured
Abstract: Tissue plasminogen activator (t-PA) is an important initiator of fibrinolysis. The t-PA polypeptide has four potential N-glycosylation sites of which three are occupied in type I (Asn-117, -184, and -448) and two in type II (Asn-117 and -448). In an effort to elucidate the factors controlling the expression of N-linked oligosaccharides on this polypeptide, we have used a combination of sequential exoglycosidase digestion, methylation analysis, and controlled acetolysis to determine the oligosaccharide structures at each of the N-glycosylation sites of type I and type II t-PA when isolated from a human colon fibroblast cell strain and from a Bowes melanoma cell line. Our results suggest the following: (i) type I and type II t-PA are N-glycosylated in an identical way at Asn-117 and Asn-448, when isolated from the same cell line; (ii) Asn-117 is predominantly associated with oligomannose-type structures in all cases; (iii) Asn-184 and Asn-448 are predominantly associated with complex-type structures when t-PA is isolated from fibroblast cells, but with both complex- and oligomannose-type structures when isolated from melanoma cells; (iv) fibroblast cell derived t-PA is associated with both neutral and sialylated oligosaccharides, while melanoma cell derived t-PA is also associated with sulfated oligosaccharides, which are located exclusively at Asn-448 of type II t-PA; (v) no complex-type structures occur in common between t-PA from the two cell lines. These results indicate that the t-PA glycoprotein is secreted by each cell line as a set of glycoforms, each glycoform being unique with respect to the nature and disposition of oligosaccharides on a common polypeptide. Further, the two cell lines express no glycoform in common, despite expressing the same t-PA polypeptide. The implications of these results for both the control of oligosaccharide processing in different cell lines and the genetic engineering of mammalian glycoproteins are discussed.
Notes: Parekh, R B
Dwek, R A
Thomas, J R
Opdenakker, G
Rademacher, T W
Wittwer, A J
Howard, S C
Nelson, R
Siegel, N R
Jennings, M G
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1989 Sep 19;28(19):7644-62. doi: 10.1021/bi00445a021.
Author Address: Department of Biochemistry, University of Oxford, U.K.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1228
Author: Park, B. S., Song, D. H., Kim, H. M., Choi, B. S., Lee, H. and Lee, J. O.
Year: 2009
Title: The structural basis of lipopolysaccharide recognition by the TLR4-MD-2 complex
Journal: Nature
Volume: 458
Issue: 7242
Pages: 1191-5
Epub Date: 2009/03/03
Date: Apr 30
Short Title: The structural basis of lipopolysaccharide recognition by the TLR4-MD-2 complex
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature07830
Accession Number: 19252480
Keywords: Binding Sites
Crystallography, X-Ray
Escherichia coli/chemistry
Humans
Hydrophobic and Hydrophilic Interactions
Lipopolysaccharides/*chemistry/*immunology
Lymphocyte Antigen 96/*chemistry/*immunology
Models, Molecular
Protein Binding
Protein Multimerization
Structure-Activity Relationship
Toll-Like Receptor 4/*chemistry/*immunology
Abstract: The lipopolysaccharide (LPS) of Gram negative bacteria is a well-known inducer of the innate immune response. Toll-like receptor (TLR) 4 and myeloid differentiation factor 2 (MD-2) form a heterodimer that recognizes a common 'pattern' in structurally diverse LPS molecules. To understand the ligand specificity and receptor activation mechanism of the TLR4-MD-2-LPS complex we determined its crystal structure. LPS binding induced the formation of an m-shaped receptor multimer composed of two copies of the TLR4-MD-2-LPS complex arranged symmetrically. LPS interacts with a large hydrophobic pocket in MD-2 and directly bridges the two components of the multimer. Five of the six lipid chains of LPS are buried deep inside the pocket and the remaining chain is exposed to the surface of MD-2, forming a hydrophobic interaction with the conserved phenylalanines of TLR4. The F126 loop of MD-2 undergoes localized structural change and supports this core hydrophobic interface by making hydrophilic interactions with TLR4. Comparison with the structures of tetra-acylated antagonists bound to MD-2 indicates that two other lipid chains in LPS displace the phosphorylated glucosamine backbone by approximately 5 A towards the solvent area. This structural shift allows phosphate groups of LPS to contribute to receptor multimerization by forming ionic interactions with a cluster of positively charged residues in TLR4 and MD-2. The TLR4-MD-2-LPS structure illustrates the remarkable versatility of the ligand recognition mechanisms employed by the TLR family, which is essential for defence against diverse microbial infection.
Notes: 1476-4687
Park, Beom Seok
Song, Dong Hyun
Kim, Ho Min
Choi, Byong-Seok
Lee, Hayyoung
Lee, Jie-Oh
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2009 Apr 30;458(7242):1191-5. doi: 10.1038/nature07830. Epub 2009 Mar 1.
Author Address: Department of Chemistry, KAIST, Daejeon, 305-701, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1352
Author: Park, S. Y., Kim, H. S., Kim, N. H., Ji, S., Cha, S. Y., Kang, J. G., Ota, I., Shimada, K., Konishi, N., Nam, H. W., Hong, S. W., Yang, W. H., Roth, J., Yook, J. I. and Cho, J. W.
Year: 2010
Title: Snail1 is stabilized by O-GlcNAc modification in hyperglycaemic condition
Journal: Embo j
Volume: 29
Issue: 22
Pages: 3787-96
Epub Date: 2010/10/21
Date: Nov 17
Short Title: Snail1 is stabilized by O-GlcNAc modification in hyperglycaemic condition
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2010.254
PMCID: PMC2989108
Accession Number: 20959806
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Cadherins/genetics/metabolism
Cell Line, Tumor
Cell Movement
Epithelial-Mesenchymal Transition
Gene Expression Regulation
Glucose/metabolism
HEK293 Cells
HeLa Cells
Humans
Hyperglycemia/*metabolism
Molecular Sequence Data
Phosphorylation
Protein Stability
RNA, Messenger/genetics
Serine/metabolism
Snail Family Transcription Factors
Transcription Factors/genetics/*metabolism
Abstract: Protein O-phosphorylation often occurs reciprocally with O-GlcNAc modification and represents a regulatory principle for proteins. O-phosphorylation of serine by glycogen synthase kinase-3β on Snail1, a transcriptional repressor of E-cadherin and a key regulator of the epithelial-mesenchymal transition (EMT) programme, results in its proteasomal degradation. We show that by suppressing O-phosphorylation-mediated degradation, O-GlcNAc at serine112 stabilizes Snail1 and thus increases its repressor function, which in turn attenuates E-cadherin mRNA expression. Hyperglycaemic condition enhances O-GlcNAc modification and initiates EMT by transcriptional suppression of E-cadherin through Snail1. Thus, dynamic reciprocal O-phosphorylation and O-GlcNAc modification of Snail1 constitute a molecular link between cellular glucose metabolism and the control of EMT.
Notes: 1460-2075
Park, Sang Yoon
Kim, Hyun Sil
Kim, Nam Hee
Ji, Suena
Cha, So Young
Kang, Jeong Gu
Ota, Ichiro
Shimada, Keiji
Konishi, Noboru
Nam, Hyung Wook
Hong, Soon Won
Yang, Won Ho
Roth, Jürgen
Yook, Jong In
Cho, Jin Won
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2010 Nov 17;29(22):3787-96. doi: 10.1038/emboj.2010.254. Epub 2010 Oct 19.
Author Address: Department of Biology, Yonsei University, Seodaemun-gu, Seoul, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1245
Author: Parkash, V. and Goldman, A.
Year: 2009
Title: Comparison of GFL-GFRalpha complexes: further evidence relating GFL bend angle to RET signalling
Journal: Acta Crystallogr Sect F Struct Biol Cryst Commun
Volume: 65
Issue: Pt 6
Pages: 551-8
Epub Date: 2009/05/30
Date: Jun 1
Short Title: Comparison of GFL-GFRalpha complexes: further evidence relating GFL bend angle to RET signalling
Alternate Journal: Acta crystallographica. Section F, Structural biology and crystallization communications
ISSN: 1744-3091
DOI: 10.1107/s1744309109017722
PMCID: PMC2688408
Accession Number: 19478429
Keywords: 3T3 Cells
Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray
Data Collection
Dimerization
Glial Cell Line-Derived Neurotrophic Factor/chemistry/isolation &
purification/*metabolism
Glial Cell Line-Derived Neurotrophic Factor Receptors/chemistry/isolation &
purification/*metabolism
Ligands
Mice
Models, Chemical
Models, Molecular
Molecular Sequence Data
Molecular Weight
Nerve Tissue Proteins/chemistry/genetics/metabolism
Protein Binding
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Proto-Oncogene Proteins c-ret/chemistry/*metabolism
Reproducibility of Results
Rotation
Sequence Homology, Amino Acid
Signal Transduction/*physiology
Statistics as Topic
X-Ray Diffraction
Abstract: Glial cell line-derived neurotrophic factor (GDNF) activates the receptor tyrosine kinase RET by binding to the GDNF-family receptor alpha1 (GFRalpha1) and forming the GDNF(2)-GFRalpha1(2)-RET(2) heterohexamer complex. A previous crystal structure of the GDNF(2)-GFRalpha1(2) complex (PDB code 2v5e) suggested that differences in signalling in GDNF-family ligand (GFL) complexes might arise from differences in the bend angle between the two monomers in the GFL homodimer. Here, a 2.35 A resolution structure of the GDNF(2)-GFRalpha1(2) complex crystallized with new cell dimensions is reported. The structure was refined to a final R factor of 22.5% (R(free) = 28%). The structures of both biological tetrameric complexes in the asymmetric unit are very similar to 2v5e and different from the artemin-GFRalpha3 structure, even though there is a small change in the structure of the GDNF. By comparison of all known GDNF and artemin structures, it is concluded that GDNF is more bent and more flexible than artemin and that this may be related to RET signalling. Comparisons also suggest that the differences between artemin and GDNF arise from the increased curvature of the artemin ;fingers', which both increases the buried surface area in the monomer-monomer interface and changes the intermonomer bend angle. From sequence comparison, it is suggested that neuturin (the second GFL) adopts an artemin-like conformation, while persephin has a different conformation to the other three.
Notes: 1744-3091
Parkash, Vimal
Goldman, Adrian
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2009 Jun 1;65(Pt 6):551-8. doi: 10.1107/S1744309109017722. Epub 2009 May 23.
Author Address: Institute of Biotechnology, University of Helsinki, Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1362
Author: Parker, B. L., Gupta, P., Cordwell, S. J., Larsen, M. R. and Palmisano, G.
Year: 2011
Title: Purification and identification of O-GlcNAc-modified peptides using phosphate-based alkyne CLICK chemistry in combination with titanium dioxide chromatography and mass spectrometry
Journal: J Proteome Res
Volume: 10
Issue: 4
Pages: 1449-58
Epub Date: 2010/12/17
Date: Apr 1
Short Title: Purification and identification of O-GlcNAc-modified peptides using phosphate-based alkyne CLICK chemistry in combination with titanium dioxide chromatography and mass spectrometry
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr100565j
Accession Number: 21158410
Keywords: Acetylglucosamine/*chemistry
Alkynes/*chemistry
Amino Acid Sequence
Animals
Cattle
Chromatography/*methods
Glycosylation
HeLa Cells
Humans
Mass Spectrometry/*methods
Mice
Molecular Sequence Data
Molecular Structure
Nuclear Proteins/chemistry/genetics/isolation & purification
Peptides/*chemistry/genetics/*isolation & purification
Proteomics/methods
Titanium/*chemistry
alpha-Crystallins/chemistry/genetics
Abstract: A selective method for the enrichment of O-GlcNAcylated peptides using a novel CLICK chemistry reagent is described. Peptides modified by O-GlcNAc were enzymatically labeled with N-azidoacetylgalactosamine. The azide was then reacted with a phospho-alkyne using CLICK chemistry and O-GlcNAcGalNAzPO(4)-containing peptides were enriched using titanium dioxide chromatography. Modified peptides were analyzed using a combination of higher energy collision dissociation for identification and electron transfer dissociation to localize the site of O-GlcNAc attachment. The enrichment method was developed and optimized using an alpha-crystallin standard protein and then applied to a soluble protein preparation of mouse brain tissue and a nuclear preparation generated from HeLa cells. A total of 42 unique O-GlcNAcylated peptides were identified, including 7 novel O-GlcNAc sites.
Notes: 1535-3907
Parker, Benjamin L
Gupta, Pankaj
Cordwell, Stuart J
Larsen, Martin R
Palmisano, Giuseppe
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2011 Apr 1;10(4):1449-58. doi: 10.1021/pr100565j. Epub 2011 Feb 22.
Author Address: Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, 5230 Odense M, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1590
Author: Parker, B. L., Thaysen-Andersen, M., Solis, N., Scott, N. E., Larsen, M. R., Graham, M. E., Packer, N. H. and Cordwell, S. J.
Year: 2013
Title: Site-specific glycan-peptide analysis for determination of N-glycoproteome heterogeneity
Journal: J Proteome Res
Volume: 12
Issue: 12
Pages: 5791-800
Epub Date: 2013/10/05
Date: Dec 6
Short Title: Site-specific glycan-peptide analysis for determination of N-glycoproteome heterogeneity
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr400783j
Accession Number: 24090084
Keywords: Amino Acid Sequence
Animals
Brain/*metabolism
Brain Chemistry
Carbohydrate Sequence
Chromatography, Liquid/instrumentation/methods
Databases, Factual
Genetic Heterogeneity
Glycomics/instrumentation/*methods
Glycosylation
Humans
Hydrogen-Ion Concentration
Male
Molecular Sequence Annotation
Molecular Sequence Data
Peptide Mapping/*methods
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/chemistry
*Protein Processing, Post-Translational
Proteome/*analysis/chemistry
Rats
Rats, Inbred Lew
Tandem Mass Spectrometry
Abstract: A combined glycomics and glycoproteomics strategy was developed for the site-specific analysis of N-linked glycosylation heterogeneity from a complex mammalian protein mixture. Initially, global characterization of the N-glycome was performed using porous graphitized carbon liquid chromatography-tandem mass spectrometry (PGC-LC-MS/MS) and the data used to create an N-glycan modification database. In the next step, tryptic glycopeptides were enriched using zwitterionic hydrophilic interaction liquid chromatography (Zic-HILIC) and fractionated by reversed-phase liquid chromatography (RPLC; pH 7.9). The resulting fractions were each separated into two equal aliquots. The first set of aliquots were treated with peptide-N-glycosidase F (PNGase F) to remove N-glycans and the former N-glycopeptides analyzed by nano-RPLC-MS/MS (pH 2.7) and identified by Mascot database search. This enabled the creation of a glycopeptide-centric concatenated database for each fraction. The second set of aliquots was analyzed directly by nanoRPLC-MS/MS (pH 2.7), employing fragmentation by CID and HCD. The assignment of glycan compositions to peptide sequences was achieved by searching the N-glycopeptide HCD MS/MS spectra against the glycopeptide-centric concatenated databases employing the N-glycan modification database. CID spectra were used to assign glycan structures identified in the glycomic analysis to peptide sequences. This multidimensional approach allowed confident identification of 863 unique intact N-linked glycopeptides from 161 rat brain glycoproteins.
Notes: 1535-3907
Parker, Benjamin L
Thaysen-Andersen, Morten
Solis, Nestor
Scott, Nichollas E
Larsen, Martin R
Graham, Mark E
Packer, Nicolle H
Cordwell, Stuart J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2013 Dec 6;12(12):5791-800. doi: 10.1021/pr400783j. Epub 2013 Nov 1.
Author Address: Discipline of Pathology, School of Medical Sciences, The University of Sydney , Sydney 2006, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 869
Author: Pasternack, A., Coleman, S. K., Féthière, J., Madden, D. R., LeCaer, J. P., Rossier, J., Pasternack, M. and Keinänen, K.
Year: 2003
Title: Characterization of the functional role of the N-glycans in the AMPA receptor ligand-binding domain
Journal: J Neurochem
Volume: 84
Issue: 5
Pages: 1184-92
Epub Date: 2003/02/27
Date: Mar
Short Title: Characterization of the functional role of the N-glycans in the AMPA receptor ligand-binding domain
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042 (Print)
0022-3042
DOI: 10.1046/j.1471-4159.2003.01611.x
Accession Number: 12603841
Keywords: Amino Acid Substitution
Animals
Binding Sites/physiology
Binding, Competitive/physiology
Cell Line
Circular Dichroism
Glycosylation/drug effects
Humans
Kidney/cytology/metabolism
Ligands
Mutagenesis, Site-Directed
Oligopeptides
Peptides/genetics
Polysaccharides/chemistry/*metabolism
Protein Denaturation/physiology
Protein Structure, Tertiary/physiology
Protein Subunits/chemistry/genetics/metabolism
Receptors, Glutamate/*chemistry/genetics/*metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Structure-Activity Relationship
Temperature
Tunicamycin/pharmacology
Abstract: The ligand-binding domains of AMPA receptor subunits carry two conserved N-glycosylation sites. In order to gain insight into the functional role of the corresponding N-glycans, we examined how the elimination of glycosylation at these sites (N407 and N414) affects the ligand-binding characteristics, structural stability, cell-surface expression, and channel properties of homomeric GluR-D (GluR4) receptor and its soluble ligand-binding domain (S1S2). GluR-D S1S2 protein expressed as a secreted protein in insect cells was found to be glycosylated at N407 and N414. No major differences in the ligand-binding properties were observed between the 'wild-type' S1S2 and non-glycosylated N407D/N414Q double mutant, or between S1S2 proteins expressed in the presence or absence of tunicamycin, an inhibitor of N-glycosylation. Purified glycosylated and non-glycosylated S1S2 proteins also showed similar thermostabilities as determined by CD spectroscopy. Full-length homomeric GluR-D receptor with N407D/N414Q mutation was expressed on the surface of HEK293 cells like the wild-type GluR-D. In outside-out patches, GluR-D and the N407D/N414Q mutant produced similar rapidly desensitizing current responses to glutamate and AMPA. We therefore report that the two conserved ligand-binding domain glycans do not play any major role in receptor-ligand interactions, do not impart a stabilizing effect on the ligand-binding domain, and are not critical for the formation and surface localization of homomeric GluR-D AMPA receptors in HEK293 cells.
Notes: Pasternack, Arja
Coleman, Sarah K
Féthière, James
Madden, Dean R
LeCaer, Jean-Pierre
Rossier, Jean
Pasternack, Michael
Keinänen, Kari
Journal Article
Research Support, Non-U.S. Gov't
England
J Neurochem. 2003 Mar;84(5):1184-92. doi: 10.1046/j.1471-4159.2003.01611.x.
Author Address: Viikki Biocenter, Department of Biosciences (Division of Biochemistry), Viikinkaari 5D, FIN-00014 University of Helsinki, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1454
Author: Pathak, S., Borodkin, V. S., Albarbarawi, O., Campbell, D. G., Ibrahim, A. and van Aalten, D. M.
Year: 2012
Title: O-GlcNAcylation of TAB1 modulates TAK1-mediated cytokine release
Journal: Embo j
Volume: 31
Issue: 6
Pages: 1394-404
Epub Date: 2012/02/07
Date: Mar 21
Short Title: O-GlcNAcylation of TAB1 modulates TAK1-mediated cytokine release
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2012.8
PMCID: PMC3321193
Accession Number: 22307082
Keywords: Acylation
Adaptor Proteins, Signal Transducing/*metabolism
Animals
Cells, Cultured
Fibroblasts/metabolism
HEK293 Cells
Humans
Interleukin-1/*metabolism
Interleukin-6/*metabolism
MAP Kinase Kinase Kinases/*metabolism
Mice
NF-kappa B/metabolism
Osmotic Pressure/physiology
Phosphorylation
Phosphotransferases (Alcohol Group Acceptor)/*metabolism
Protein Transport
Protein-Serine-Threonine Kinases/metabolism
Signal Transduction/physiology
Tumor Necrosis Factor-alpha/*metabolism
Abstract: Transforming growth factor (TGF)-β-activated kinase 1 (TAK1) is a key serine/threonine protein kinase that mediates signals transduced by pro-inflammatory cytokines such as transforming growth factor-β, tumour necrosis factor (TNF), interleukin-1 (IL-1) and wnt family ligands. TAK1 is found in complex with binding partners TAB1-3, phosphorylation and ubiquitination of which has been found to regulate TAK1 activity. In this study, we show that TAB1 is modified with N-acetylglucosamine (O-GlcNAc) on a single site, Ser395. With the help of a novel O-GlcNAc site-specific antibody, we demonstrate that O-GlcNAcylation of TAB1 is induced by IL-1 and osmotic stress, known inducers of the TAK1 signalling cascade. By reintroducing wild-type or an O-GlcNAc-deficient mutant TAB1 (S395A) into Tab1(-/-) mouse embryonic fibroblasts, we determined that O-GlcNAcylation of TAB1 is required for full TAK1 activation upon stimulation with IL-1/osmotic stress, for downstream activation of nuclear factor κB and finally production of IL-6 and TNFα. This is one of the first examples of a single O-GlcNAc site on a signalling protein modulating a key innate immunity signalling pathway.
Notes: 1460-2075
Pathak, Shalini
Borodkin, Vladimir S
Albarbarawi, Osama
Campbell, David G
Ibrahim, Adel
van Aalten, Daan Mf
087590/Wellcome Trust/United Kingdom
G0900138/Medical Research Council/United Kingdom
G1100713/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2012 Mar 21;31(6):1394-404. doi: 10.1038/emboj.2012.8. Epub 2012 Feb 3.
Author Address: Division of Cell Signalling and Immunology, College of Life Sciences, University of Dundee, Dundee, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1104
Author: Pearce, M. M., Wang, Y., Kelley, G. G. and Wojcikiewicz, R. J.
Year: 2007
Title: SPFH2 mediates the endoplasmic reticulum-associated degradation of inositol 1,4,5-trisphosphate receptors and other substrates in mammalian cells
Journal: J Biol Chem
Volume: 282
Issue: 28
Pages: 20104-15
Epub Date: 2007/05/16
Date: Jul 13
Short Title: SPFH2 mediates the endoplasmic reticulum-associated degradation of inositol 1,4,5-trisphosphate receptors and other substrates in mammalian cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M701862200
Accession Number: 17502376
Keywords: Animals
Endoplasmic Reticulum/genetics/*metabolism
Gene Expression Regulation/*physiology
HeLa Cells
Humans
Inositol 1,4,5-Trisphosphate Receptors/genetics/*metabolism
Membrane Proteins/antagonists & inhibitors/genetics/*metabolism
Proteasome Endopeptidase Complex/genetics/metabolism
Protein Binding/physiology
RNA, Small Interfering/genetics
Rats
TNF Receptor-Associated Factor 2
Tumor Necrosis Factor Receptor-Associated Peptides and Proteins/genetics/metabolism
Ubiquitin/genetics/metabolism
Abstract: Inositol 1,4,5-trisphosphate (IP(3)) receptors are endoplasmic reticulum (ER) membrane calcium channels that, upon activation, become substrates for the ER-associated degradation (ERAD) pathway. Although it is clear that IP(3) receptors are polyubiquitinated upon activation and are transferred to the proteasome by a p97-based complex, currently nothing is known about the proteins that initially select activated IP(3) receptors for ERAD. Here, we sought to identify novel proteins that associate with and mediate the ERAD of endogenous activated IP(3) receptors. SPFH2, an uncharacterized SPFH domain-containing protein, rapidly associated with IP(3) receptors in a manner that preceded significant polyubiquitination and the association of p97 and related proteins. SPFH2 was found to be an ER membrane protein largely residing within the ER lumen and in resting and stimulated cells was linked to ERAD pathway components, apparently via endogenous substrates undergoing degradation. Suppression of SPFH2 expression by RNA interference markedly inhibited IP(3) receptor polyubiquitination and degradation and the processing of other ERAD substrates. Overall, these studies identify SPFH2 as a key ERAD pathway component and suggest that it may act as a substrate recognition factor.
Notes: Pearce, Margaret M P
Wang, Yuan
Kelley, Grant G
Wojcikiewicz, Richard J H
DK 49194/DK/NIDDK NIH HHS/United States
DK 56294/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
J Biol Chem. 2007 Jul 13;282(28):20104-15. doi: 10.1074/jbc.M701862200. Epub 2007 May 14.
Author Address: Departments of Pharmacology and Medicine, State University of New York Upstate Medical University, Syracuse, NY 13210, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2038
Author: Pedersen, D. V., Gadeberg, T. A. F., Thomas, C., Wang, Y., Joram, N., Jensen, R. K., Mazarakis, S. M. M., Revel, M., El Sissy, C., Petersen, S. V., Lindorff-Larsen, K., Thiel, S., Laursen, N. S., Fremeaux-Bacchi, V. and Andersen, G. R.
Year: 2019
Title: Structural Basis for Properdin Oligomerization and Convertase Stimulation in the Human Complement System
Journal: Front Immunol
Volume: 10
Pages: 2007
Epub Date: 2019/09/12
Short Title: Structural Basis for Properdin Oligomerization and Convertase Stimulation in the Human Complement System
Alternate Journal: Frontiers in immunology
ISSN: 1664-3224
DOI: 10.3389/fimmu.2019.02007
PMCID: PMC6713926
Accession Number: 31507604
Keywords: Binding Sites
Complement C3-C5 Convertases/*chemistry/metabolism
Complement System Proteins/*chemistry
HEK293 Cells
Humans
Models, Molecular
Mutation
Properdin/*chemistry/metabolism
Protein Binding
Protein Conformation
*Protein Multimerization
Protein Processing, Post-Translational
Proteolysis
Structure-Activity Relationship
*complement
*complement component C3
*convertase
*crystal structure
*factor B
*properdin
*regulation
Abstract: Properdin (FP) is a positive regulator of the immune system stimulating the activity of the proteolytically active C3 convertase C3bBb in the alternative pathway of the complement system. Here we present two crystal structures of FP and two structures of convertase bound FP. A structural core formed by three thrombospondin repeats (TSRs) and a TB domain harbors the convertase binding site in FP that mainly interacts with C3b. Stabilization of the interaction between the C3b C-terminus and the MIDAS bound Mg(2+) in the Bb protease by FP TSR5 is proposed to underlie FP convertase stabilization. Intermolecular contacts between FP and the convertase subunits suggested by the structure were confirmed by binding experiments. FP is shown to inhibit C3b degradation by FI due to a direct competition for a common binding site on C3b. FP oligomers are held together by two sets of intermolecular contacts, where the first is formed by the TB domain from one FP molecule and TSR4 from another. The second and largest interface is formed by TSR1 and TSR6 from the same two FP molecules. Flexibility at four hinges between thrombospondin repeats is suggested to enable the oligomeric, polydisperse, and extended architecture of FP. Our structures rationalize the effects of mutations associated with FP deficiencies and provide a structural basis for the analysis of FP function in convertases and its possible role in pattern recognition.
Notes: 1664-3224
Pedersen, Dennis V
Gadeberg, Trine A F
Thomas, Caroline
Wang, Yong
Joram, Nicolas
Jensen, Rasmus K
Mazarakis, Sofia M M
Revel, Margot
El Sissy, Carine
Petersen, Steen V
Lindorff-Larsen, Kresten
Thiel, Steffen
Laursen, Nick S
Fremeaux-Bacchi, Véronique
Andersen, Gregers R
Journal Article
Research Support, Non-U.S. Gov't
Front Immunol. 2019 Aug 22;10:2007. doi: 10.3389/fimmu.2019.02007. eCollection 2019.
Author Address: Department of Molecular Biology and Genetics, Center for Structural Biology, Aarhus University, Aarhus, Denmark.
Service d'Oncologie Pédiatrique, CHU Nantes, Hôpital Mère Enfant, Nantes, France.
Department of Biology, Linderstrøm-Lang Centre for Protein Science, University of Copenhagen, Copenhagen, Denmark.
Service de Réanimation Pédiatrique, CHU Nantes, Nantes, France.
Centre de Recherche des Cordeliers, INSERM, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, Paris, France.
Service d'Immunologie Biologique, Assistance Publique - Hôpitaux de Paris, Hôpital Européen Georges Pompidou, Paris, France.
Department of Biomedicine, Aarhus University, Aarhus, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 482
Author: Peeters, B., Rombauts, W., Mous, J. and Heyns, W.
Year: 1981
Title: Structural studies on rat prostatic binding protein. The primary structure of its glycosylated component C3
Journal: Eur J Biochem
Volume: 115
Issue: 1
Pages: 115-21
Epub Date: 1981/03/16
Date: Mar 16
Short Title: Structural studies on rat prostatic binding protein. The primary structure of its glycosylated component C3
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
Accession Number: 7014218
Keywords: Amino Acid Sequence
Amino Acids/analysis
Animals
Carrier Proteins/*analysis
Chemical Phenomena
Chemistry
Glycopeptides/*analysis
Male
Peptide Fragments/analysis
Peptide Hydrolases
Prostate/*analysis
Rats
Staphylococcus aureus/enzymology
Abstract: The amino acid sequence of the glycosylated component C3 of rat prostatic binding protein has been determined. The peptides obtained by digestion of the S-carboxamidomethylated or S-aminoethylated glycoprotein with trypsin and Staphylococcus aureus protease were sequenced by manual Edman degradation. The alignment of the fragments was further established with overlapping peptides obtained by enzymic hydrolysis of the modified protein with chymotrypsin and thermolysin, and by chemical cleavage with cyanogen bromide. The glycopeptide C3 contains 77 amino acids corresponding to a molecular weight of 8653. the oligosaccharide chain is attached to the peptide by an N-glycosidic bond to asparagine-17. C3 is an acidic polypeptide due to the presence of ten acidic residues; its three cysteine residues are located at both extremities and in the middle of the molecule.
Notes: Peeters, B
Rombauts, W
Mous, J
Heyns, W
Journal Article
England
Eur J Biochem. 1981 Mar 16;115(1):115-21.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1652
Author: Pekkurnaz, G., Trinidad, J. C., Wang, X., Kong, D. and Schwarz, T. L.
Year: 2014
Title: Glucose regulates mitochondrial motility via Milton modification by O-GlcNAc transferase
Journal: Cell
Volume: 158
Issue: 1
Pages: 54-68
Epub Date: 2014/07/06
Date: Jul 3
Short Title: Glucose regulates mitochondrial motility via Milton modification by O-GlcNAc transferase
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2014.06.007
PMCID: PMC4224014
NIHMSID: NIHMS604258
Accession Number: 24995978
Keywords: Adaptor Proteins, Vesicular Transport/*metabolism
Animals
Axons/metabolism
Carrier Proteins
Drosophila melanogaster
Gene Knockdown Techniques
Glucose/*metabolism
Hippocampus/cytology/metabolism
Humans
Male
Mice
Mice, Inbred C57BL
Mitochondria/*metabolism
N-Acetylglucosaminyltransferases/genetics/*metabolism
Rats
Sequence Alignment
Abstract: Cells allocate substantial resources toward monitoring levels of nutrients that can be used for ATP generation by mitochondria. Among the many specialized cell types, neurons are particularly dependent on mitochondria due to their complex morphology and regional energy needs. Here, we report a molecular mechanism by which nutrient availability in the form of extracellular glucose and the enzyme O-GlcNAc Transferase (OGT), whose activity depends on glucose availability, regulates mitochondrial motility in neurons. Activation of OGT diminishes mitochondrial motility. We establish the mitochondrial motor-adaptor protein Milton as a required substrate for OGT to arrest mitochondrial motility by mapping and mutating the key O-GlcNAcylated serine residues. We find that the GlcNAcylation state of Milton is altered by extracellular glucose and that OGT alters mitochondrial motility in vivo. Our findings suggest that, by dynamically regulating Milton GlcNAcylation, OGT tailors mitochondrial dynamics in neurons based on nutrient availability.
Notes: 1097-4172
Pekkurnaz, Gulcin
Trinidad, Jonathan C
Wang, Xinnan
Kong, Dong
Schwarz, Thomas L
P30DK46200/DK/NIDDK NIH HHS/United States
K01 DK094943/DK/NIDDK NIH HHS/United States
P30 DK046200/DK/NIDDK NIH HHS/United States
P30 HD018655/HD/NICHD NIH HHS/United States
P30HD18655/HD/NICHD NIH HHS/United States
R01 GM069808/GM/NIGMS NIH HHS/United States
5R01GM069808-08/GM/NIGMS NIH HHS/United States
K01DK094943/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell. 2014 Jul 3;158(1):54-68. doi: 10.1016/j.cell.2014.06.007.
Author Address: The F.M. Kirby Neurobiology Center, Boston Children's Hospital and Department of Neurobiology, Harvard Medical School, Boston, MA 02115, USA.
Department of Chemistry, Biological Mass Spectrometry Facility, Indiana University, Bloomington, IN 47405, USA.
Department of Neurosurgery, Stanford University, Stanford, CA 94304, USA.
Division of Endocrinology, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA.
The F.M. Kirby Neurobiology Center, Boston Children's Hospital and Department of Neurobiology, Harvard Medical School, Boston, MA 02115, USA. Electronic address: thomas.schwarz@childrens.harvard.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1890
Author: Peng, C., Zhu, Y., Zhang, W., Liao, Q., Chen, Y., Zhao, X., Guo, Q., Shen, P., Zhen, B., Qian, X., Yang, D., Zhang, J. S., Xiao, D., Qin, W. and Pei, H.
Year: 2017
Title: Regulation of the Hippo-YAP Pathway by Glucose Sensor O-GlcNAcylation
Journal: Mol Cell
Volume: 68
Issue: 3
Pages: 591-604.e5
Epub Date: 2017/11/04
Date: Nov 2
Short Title: Regulation of the Hippo-YAP Pathway by Glucose Sensor O-GlcNAcylation
Alternate Journal: Molecular cell
ISSN: 1097-2765
DOI: 10.1016/j.molcel.2017.10.010
Accession Number: 29100056
Keywords: Adaptor Proteins, Signal Transducing/genetics/*metabolism
Animals
Cell Proliferation
Cell Transformation, Neoplastic/genetics/*metabolism/pathology
Gene Expression Regulation, Neoplastic
Glucose/*metabolism
Glycosylation
HEK293 Cells
HeLa Cells
Humans
Mice, Nude
N-Acetylglucosaminyltransferases/*metabolism
Neoplasms/*enzymology/genetics/pathology
Phosphoproteins/genetics/*metabolism
Phosphorylation
*Protein Processing, Post-Translational
Protein-Serine-Threonine Kinases/genetics/metabolism
Serine
Signal Transduction
Time Factors
Transcription Factors
Transcription, Genetic
Transcriptional Activation
Hippo
O-GlcNAcylation
Ogt
Yap
pancreatic cancer
Abstract: The Hippo pathway is crucial in organ size control and tissue homeostasis, with deregulation leading to cancer. An extracellular nutrition signal, such as glucose, regulates the Hippo pathway activation. However, the mechanisms are still not clear. Here, we found that the Hippo pathway is directly regulated by the hexosamine biosynthesis pathway (HBP) in response to metabolic nutrients. Mechanistically, the core component of Hippo pathway (YAP) is O-GlcNAcylated by O-GlcNAc transferase (OGT) at serine 109. YAP O-GlcNAcylation disrupts its interaction with upstream kinase LATS1, prevents its phosphorylation, and activates its transcriptional activity. And this activation is not dependent on AMPK. We also identified OGT as a YAP-regulated gene that forms a feedback loop. Finally, we confirmed that glucose-induced YAP O-GlcNAcylation and activation promoted tumorigenesis. Together, our data establish a molecular mechanism and functional significance of the HBP in directly linking extracellular glucose signal to the Hippo-YAP pathway and tumorigenesis.
Notes: 1097-4164
Peng, Changmin
Zhu, Yue
Zhang, Wanjun
Liao, Qinchao
Chen, Yali
Zhao, Xinyuan
Guo, Qiang
Shen, Pan
Zhen, Bei
Qian, Xiaohong
Yang, Dong
Zhang, Jin-San
Xiao, Dongguang
Qin, Weijie
Pei, Huadong
Journal Article
United States
Mol Cell. 2017 Nov 2;68(3):591-604.e5. doi: 10.1016/j.molcel.2017.10.010.
Author Address: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Lab, College of Biotechnology, Tianjin University of Science and Technology, No 29, 13ST. TEDA, Tianjin 300457, China; State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing 102206, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing 102206, China; Anhui Medical University, Hefei 230032, China; Beijing Institute of Radiation Medicine, Beijing 102206, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing 102206, China.
Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Lab, College of Biotechnology, Tianjin University of Science and Technology, No 29, 13ST. TEDA, Tianjin 300457, China.
Cell Signaling and Epigenetics Laboratory, School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou 325035, Zhejiang, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing 102206, China. Electronic address: aunp_dna@126.com.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing 102206, China; Department of Biochemistry and Molecular Medicine, George Washington University School of Medicine and Health Science, 2300 Eye Street, N.W., Washington, DC 20037, USA. Electronic address: peihuadong@hotmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1135
Author: Penuela, S., Bhalla, R., Gong, X. Q., Cowan, K. N., Celetti, S. J., Cowan, B. J., Bai, D., Shao, Q. and Laird, D. W.
Year: 2007
Title: Pannexin 1 and pannexin 3 are glycoproteins that exhibit many distinct characteristics from the connexin family of gap junction proteins
Journal: J Cell Sci
Volume: 120
Issue: Pt 21
Pages: 3772-83
Epub Date: 2007/10/11
Date: Nov 1
Short Title: Pannexin 1 and pannexin 3 are glycoproteins that exhibit many distinct characteristics from the connexin family of gap junction proteins
Alternate Journal: Journal of cell science
ISSN: 0021-9533 (Print)
0021-9533
DOI: 10.1242/jcs.009514
Accession Number: 17925379
Keywords: Amino Acid Sequence
Animals
Cell Line
Connexin 43/genetics/*metabolism
Connexins/genetics/*metabolism
Gap Junctions/chemistry/*metabolism
Glycoproteins/chemistry/genetics/*metabolism
Glycosylation
Humans
Mice
Molecular Sequence Data
Nerve Tissue Proteins/chemistry/genetics/*metabolism
Protein Conformation
Rats
Recombinant Fusion Proteins/genetics/metabolism
Sequence Alignment
Skin/cytology/metabolism
Spleen/cytology/metabolism
Abstract: Pannexins are mammalian orthologs of the invertebrate gap junction proteins innexins and thus have been proposed to play a role in gap junctional intercellular communication. Localization of exogenously expressed pannexin 1 (Panx1) and pannexin 3 (Panx3), together with pharmacological studies, revealed a cell surface distribution profile and life cycle dynamics that were distinct from connexin 43 (Cx43, encoded by Gja1). Furthermore, N-glycosidase treatment showed that both Panx1 (approximately 41-48 kD species) and Panx3 (approximately 43 kD) were glycosylated, whereas N-linked glycosylation-defective mutants exhibited a decreased ability to be transported to the cell surface. Tissue surveys revealed the expression of Panx1 in several murine tissues--including in cartilage, skin, spleen and brain--whereas Panx3 expression was prevalent in skin and cartilage with a second higher-molecular-weight species present in a broad range of tissues. Tissue-specific localization patterns of Panx1 and Panx3 ranging from distinct cell surface clusters to intracellular profiles were revealed by immunostaining of skin and spleen sections. Finally, functional assays in cultured cells transiently expressing Panx1 and Panx3 were incapable of forming intercellular channels, but assembled into functional cell surface channels. Collectively, these studies show that Panx1 and Panx3 have many characteristics that are distinct from Cx43 and that these proteins probably play an important biological role as single membrane channels.
Notes: Penuela, Silvia
Bhalla, Ruchi
Gong, Xiang-Qun
Cowan, Kyle N
Celetti, Steven J
Cowan, Bryce J
Bai, Donglin
Shao, Qing
Laird, Dale W
Journal Article
Research Support, Non-U.S. Gov't
England
J Cell Sci. 2007 Nov 1;120(Pt 21):3772-83. doi: 10.1242/jcs.009514. Epub 2007 Oct 9.
Author Address: Department of Anatomy and Cell Biology, University of Western Ontario, London, ON, N6A 5C1, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 849
Author: Peoc'h, K., Serres, C., Frobert, Y., Martin, C., Lehmann, S., Chasseigneaux, S., Sazdovitch, V., Grassi, J., Jouannet, P., Launay, J. M. and Laplanche, J. L.
Year: 2002
Title: The human "prion-like" protein Doppel is expressed in both Sertoli cells and spermatozoa
Journal: J Biol Chem
Volume: 277
Issue: 45
Pages: 43071-8
Epub Date: 2002/08/30
Date: Nov 8
Short Title: The human "prion-like" protein Doppel is expressed in both Sertoli cells and spermatozoa
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M206357200
Accession Number: 12200435
Keywords: Animals
Base Sequence
Brain/metabolism
CHO Cells
Cricetinae
GPI-Linked Proteins
Glycosylation
Humans
Male
Organ Specificity
Plasmids
Prions/*genetics/*metabolism
Recombinant Proteins/metabolism
Restriction Mapping
Sertoli Cells/*metabolism
Spermatozoa/*metabolism
Testis/cytology/metabolism
Transfection
Abstract: The prion-like Doppel protein (Dpl) has many biochemical and structural properties in common with the cellular prion protein (PrP(c)), and the physiological role of neither protein is known. Experimental data suggest either direct or indirect interaction between the two proteins. In this study, we investigated the expression pattern and biochemical characteristics of Dpl in human tissues and in Chinese hamster ovary cells transfected with wild-type or variant human Dpl gene constructs. Human Dpl appears to be a glycosylphosphatidylinositol-anchored glycoprotein with N- and O-linked sugars. It was found on Sertoli cells in the testis, on the flagella of epididymal and mature spermatozoa, and in seminal plasma. Dpl coexists only with N-terminally truncated isoforms of PrP(c) on mature spermatozoa. The localization of human Dpl on both Sertoli cells (somatic cells) and spermatozoa (germinal cells) strongly suggests that this protein may play a major role in human male fertility. Finally, our data indicate that spermatozoa are thus an interesting model for studies of the potential interaction between Dpl and PrP(c).
Notes: Peoc'h, Katell
Serres, Catherine
Frobert, Yveline
Martin, Caroline
Lehmann, Sylvain
Chasseigneaux, Stephanie
Sazdovitch, Veronique
Grassi, Jacques
Jouannet, Pierre
Launay, Jean-Marie
Laplanche, Jean-Louis
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2002 Nov 8;277(45):43071-8. doi: 10.1074/jbc.M206357200. Epub 2002 Aug 27.
Author Address: Service de Biochimie et Biologie Moléculaire, Hôpital Lariboisière, 2, rue Ambroise Paré, 75475 Paris Cedex 10, France. katell.peoch@lrb.ap-hop-paris.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 566
Author: Pepys, M. B., Rademacher, T. W., Amatayakul-Chantler, S., Williams, P., Noble, G. E., Hutchinson, W. L., Hawkins, P. N., Nelson, S. R., Gallimore, J. R., Herbert, J. and et al.
Year: 1994
Title: Human serum amyloid P component is an invariant constituent of amyloid deposits and has a uniquely homogeneous glycostructure
Journal: Proc Natl Acad Sci U S A
Volume: 91
Issue: 12
Pages: 5602-6
Epub Date: 1994/06/07
Date: Jun 7
Short Title: Human serum amyloid P component is an invariant constituent of amyloid deposits and has a uniquely homogeneous glycostructure
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.91.12.5602
PMCID: PMC44044
Accession Number: 8202534
Keywords: Amyloid/*chemistry
Animals
Asialoglycoproteins/metabolism
Carbohydrate Sequence
Female
Humans
Metabolic Clearance Rate
Mice
Mice, Inbred CBA
Molecular Sequence Data
Serum Amyloid P-Component/*chemistry/metabolism
Abstract: Human serum amyloid P component (SAP) is a normal plasma protein and the precursor of amyloid P component (AP), a universal constituent of the abnormal tissue deposits in amyloidosis, including Alzheimer disease. We show here that its single N-linked biantennary oligosaccharide does not display the microheterogeneity usually characteristic of glycoproteins. The protein and the glycan structures of AP were also invariant, their resistance to degradation suggesting a role in persistence of amyloid deposits. Asialo-SAP was rapidly cleared from the circulation in mice by a mechanism dependent on terminal galactose residues and was catabolized in hepatocytes. However blockade of this pathway did not affect the clearance of native SAP. Rapid hepatic uptake and catabolism of human asialo-SAP in man were also directly demonstrated. The protein and glycan homogeneity of SAP and the integrity of AP suggest that the complete glycoprotein structure is important for the normal and the pathophysiological functions of this molecule.
Notes: 1091-6490
Pepys, M B
Rademacher, T W
Amatayakul-Chantler, S
Williams, P
Noble, G E
Hutchinson, W L
Hawkins, P N
Nelson, S R
Gallimore, J R
Herbert, J
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 1994 Jun 7;91(12):5602-6. doi: 10.1073/pnas.91.12.5602.
Author Address: Immunological Medicine Unit, Royal Postgraduate Medical School, Hammersmith Hospital, London, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1399
Author: Perdivara, I., Peddada, S. D., Miller, F. W., Tomer, K. B. and Deterding, L. J.
Year: 2011
Title: Mass spectrometric determination of IgG subclass-specific glycosylation profiles in siblings discordant for myositis syndromes
Journal: J Proteome Res
Volume: 10
Issue: 7
Pages: 2969-78
Epub Date: 2011/05/26
Date: Jul 1
Short Title: Mass spectrometric determination of IgG subclass-specific glycosylation profiles in siblings discordant for myositis syndromes
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr200397h
PMCID: PMC3137520
NIHMSID: NIHMS302619
Accession Number: 21609021
Keywords: Adult
Age of Onset
Asymptomatic Diseases
Case-Control Studies
Child
Chromatography, Liquid
Electrophoresis, Polyacrylamide Gel
Female
Galactose/*metabolism
Genetic Predisposition to Disease
Glycopeptides/*analysis/chemistry
Glycosylation
Humans
Immunoglobulin G/*blood/immunology
Immunoglobulin Isotypes/analysis/*blood
Male
Mass Spectrometry
*Myositis/blood/diagnosis/genetics/immunology/physiopathology
Peptide Fragments/*analysis/chemistry
Siblings
Twins/*blood/genetics/immunology
Abstract: Many autoimmune conditions are believed to result from chronic inflammation as a consequence of the interaction of genetic and environmental factors in susceptible individuals. One common feature in some autoimmune diseases is the decrease in terminal galactosylation of the constant region N-glycan of the total plasma immunoglobulin. To determine whether a similar pattern is characteristic for the autoimmune disorder myositis, we analyzed the antibody subclass specific glycosylation in patients with myositis, their asymptomatic siblings, and healthy unrelated age- and sex-matched controls. The antibody subclass specific glycosylation was determined from the LC-MS analyses of the IgG glycopeptides generated by trypsin digestion of the antibody heavy chain. The glycosylation profiles of the IgG subclasses were determined relative to the total abundance of all glycoforms. We found elevated amounts of glycoforms lacking terminal galactose in myositis patients. Pairwise statistical analyses reveals that galactosylation is statistically different between the myositis patients and control groups. Furthermore, the trend analysis for glycosylation indicates a pattern of decreasing galactosylation in the order controls ≥ siblings ≥ myositis patients, suggesting the existence of a genetic, immune-related predisposition in the group of asymptomatic siblings that can be detected before the onset of clinical symptoms at the level of plasma proteins.
Notes: 1535-3907
Perdivara, Irina
Peddada, Shyamal D
Miller, Frederick W
Tomer, Kenneth B
Deterding, Leesa J
Z01 ES045005-11/ImNIH/Intramural NIH HHS/United States
Z01 ES050171-09/ImNIH/Intramural NIH HHS/United States
ES101074-07/ES/NIEHS NIH HHS/United States
Z01 ES045005/ImNIH/Intramural NIH HHS/United States
Z01 ES101074/ImNIH/Intramural NIH HHS/United States
Z99 ES999999/ImNIH/Intramural NIH HHS/United States
Z01 ES101074-07/ImNIH/Intramural NIH HHS/United States
Z01 ES050171/ImNIH/Intramural NIH HHS/United States
ES045005-14/ES/NIEHS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
J Proteome Res. 2011 Jul 1;10(7):2969-78. doi: 10.1021/pr200397h. Epub 2011 Jun 8.
Author Address: Laboratory of Structural Biology, NIH/DHHS, Research Triangle Park, North Carolina 27709, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1275
Author: Perez-Caballero, D., Zang, T., Ebrahimi, A., McNatt, M. W., Gregory, D. A., Johnson, M. C. and Bieniasz, P. D.
Year: 2009
Title: Tetherin inhibits HIV-1 release by directly tethering virions to cells
Journal: Cell
Volume: 139
Issue: 3
Pages: 499-511
Epub Date: 2009/11/03
Date: Oct 30
Short Title: Tetherin inhibits HIV-1 release by directly tethering virions to cells
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2009.08.039
PMCID: PMC2844890
NIHMSID: NIHMS186264
Accession Number: 19879838
Keywords: Animals
Antigens, CD/chemistry/genetics/*metabolism
Cell Line
Cell Membrane/metabolism
Ebolavirus/metabolism
GPI-Linked Proteins
HIV-1/*metabolism
Human Immunodeficiency Virus Proteins/metabolism
Humans
Membrane Glycoproteins/chemistry/genetics/*metabolism
Mutagenesis
Protein Structure, Tertiary
Rats
Viral Matrix Proteins/metabolism
Viral Regulatory and Accessory Proteins/metabolism
Virion/*metabolism
Virus Replication
Abstract: Tetherin is an interferon-induced protein whose expression blocks the release of HIV-1 and other enveloped viral particles. The underlying mechanism by which tetherin functions and whether it directly or indirectly causes virion retention are unknown. Here, we elucidate the mechanism by which tetherin exerts its antiviral activity. We demonstrate, through mutational analyses and domain replacement experiments, that tetherin configuration rather than primary sequence is critical for antiviral activity. These findings allowed the design of a completely artificial protein, lacking sequence homology with native tetherin, that nevertheless mimicked its antiviral activity. We further show that tetherin is incorporated into HIV-1 particles as a parallel homodimer using either of its two membrane anchors. These results indicate that tetherin functions autonomously and directly and that infiltration of virion envelopes by one or both of tetherin's membrane anchors is necessary, and likely sufficient, to tether enveloped virus particles that bud through the plasma membrane.
Notes: 1097-4172
Perez-Caballero, David
Zang, Trinity
Ebrahimi, Alaleh
McNatt, Matthew W
Gregory, Devon A
Johnson, Marc C
Bieniasz, Paul D
R01 AI073098-02/AI/NIAID NIH HHS/United States
R01AI50111/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 AI050111-09/AI/NIAID NIH HHS/United States
R01 AI050111/AI/NIAID NIH HHS/United States
R01 AI073098/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell. 2009 Oct 30;139(3):499-511. doi: 10.1016/j.cell.2009.08.039.
Author Address: Aaron Diamond AIDS Research Center, The Rockefeller University, New York, NY 10016, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 919
Author: Perez-Vilar, J., Randell, S. H. and Boucher, R. C.
Year: 2004
Title: C-Mannosylation of MUC5AC and MUC5B Cys subdomains
Journal: Glycobiology
Volume: 14
Issue: 4
Pages: 325-37
Epub Date: 2004/01/14
Date: Apr
Short Title: C-Mannosylation of MUC5AC and MUC5B Cys subdomains
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwh041
Accession Number: 14718370
Keywords: Amino Acid Sequence
Animals
COS Cells
Gene Expression
Mannose/*metabolism
Molecular Sequence Data
Mucin 5AC
Mucin-5B
Mucins/biosynthesis/*chemistry/genetics/*metabolism
Peptide Fragments/chemistry/genetics/metabolism
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Homology, Amino Acid
Abstract: We expressed recombinant Cys subdomains in COS-7 cells to examine the role of this highly conserved protein domain in mucin biosynthesis. The entire Cys1 and Cys5 and Cys1 and Cys3 subdomains in MUC5AC and MUC5B, respectively, each with six carboxyl terminal histidine residues, were pulse-labeled with [(35)S]cysteine/methionine, and the labeled proteins were examined in the culture medium. Under nonreducing conditions, secreted Cys subdomains were monomers, indicating the absence of interchain disulfide bonds. Cross-linking studies suggested the domains are able to interact through very weak noncovalent interactions. Though the domains had apparent M(r) consistent with the absence of N- and O-glycans, they could be purified with mannose-specific lectins. Lectin binding was prevented by mutation of the first tryptophan residue in the putative C-mannosylation acceptor motif WXXW, indicating that C-mannosylation is responsible for lectin binding. As judged by pulse-chase experiments, C-mannosylation occurred very early during the domain biosynthesis, likely in the endoplasmic reticulum (ER). Mutation of the WXXW motif or expression of the unmutated domain in CHO-Lec35.1 cells, a C-mannosylation-defective cell line, resulted in reduced secretion of the corresponding Cys subdomains. Live cell imaging of green fluorescent protein fused to the Cys subdomains clearly revealed increased presence of Cys subdomains in the ER of CHO-Lec35.1 cells when compared to the same domains expressed in CHO-K1 cells. Considered together, these studies suggest that the Cys subdomains of MUC5AC and MUC5B are C-mannosylated in their respective WXXW motifs. C-mannosylation is likely required for proper folding of the Cys subdomains and/or for some aspect of ER export during mucin biosynthesis.
Notes: Perez-Vilar, Juan
Randell, Scott H
Boucher, Richard C
HL58345/HL/NHLBI NIH HHS/United States
HL60280/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 2004 Apr;14(4):325-37. doi: 10.1093/glycob/cwh041. Epub 2004 Jan 12.
Author Address: Cystic Fibrosis/Pulmonary Research and Treatment Center, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA. juan_vilar@med.unc.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2051
Author: Petrus, P., Lecoutre, S., Dollet, L., Wiel, C., Sulen, A., Gao, H., Tavira, B., Laurencikiene, J., Rooyackers, O., Checa, A., Douagi, I., Wheelock, C. E., Arner, P., McCarthy, M., Bergo, M. O., Edgar, L., Choudhury, R. P., Aouadi, M., Krook, A. and Rydén, M.
Year: 2020
Title: Glutamine Links Obesity to Inflammation in Human White Adipose Tissue
Journal: Cell Metab
Volume: 31
Issue: 2
Pages: 375-390.e11
Epub Date: 2019/12/24
Date: Feb 4
Short Title: Glutamine Links Obesity to Inflammation in Human White Adipose Tissue
Alternate Journal: Cell metabolism
ISSN: 1550-4131
DOI: 10.1016/j.cmet.2019.11.019
Accession Number: 31866443
Keywords: Acetylglucosamine
*Adipocytes/metabolism/pathology
*Adipose Tissue, White/metabolism/pathology
Adult
Animals
Cells, Cultured
Cohort Studies
Female
Glucose/metabolism
*Glutamine/metabolism/pharmacology
Glycosylation/drug effects
Humans
Inflammation/*metabolism
Male
Mice, Inbred C57BL
Middle Aged
N-Acetylglucosaminyltransferases/metabolism
Obesity/*metabolism
*adipocyte
*adipokine
*epigenetics
*inflammation
*leukocyte
*macrophage
*metabolomics
*obesity
since June 2019 taken up a position as Senior Director and Staff Scientist at
Genentech, Inc. and is a holder of Roche stock. However, neither he nor any other
author of this work has any conflict of interest to report.
Abstract: While obesity and associated metabolic complications are linked to inflammation of white adipose tissue (WAT), the causal factors remain unclear. We hypothesized that the local metabolic environment could be an important determinant. To this end, we compared metabolites released from WAT of 81 obese and non-obese women. This identified glutamine to be downregulated in obesity and inversely associated with a pernicious WAT phenotype. Glutamine administration in vitro and in vivo attenuated both pro-inflammatory gene and protein levels in adipocytes and WAT and macrophage infiltration in WAT. Metabolomic and bioenergetic analyses in human adipocytes suggested that glutamine attenuated glycolysis and reduced uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) levels. UDP-GlcNAc is the substrate for the post-translational modification O-linked β-N-acetylglucosamine (O-GlcNAc) mediated by the enzyme O-GlcNAc transferase. Functional studies in human adipocytes established a mechanistic link between reduced glutamine, O-GlcNAcylation of nuclear proteins, and a pro-inflammatory transcriptional response. Altogether, glutamine metabolism is linked to WAT inflammation in obesity.
Notes: 1932-7420
Petrus, Paul
Lecoutre, Simon
Dollet, Lucile
Wiel, Clotilde
Sulen, André
Gao, Hui
Tavira, Beatriz
Laurencikiene, Jurga
Rooyackers, Olav
Checa, Antonio
Douagi, Iyadh
Wheelock, Craig E
Arner, Peter
McCarthy, Mark
Bergo, Martin O
Edgar, Laurienne
Choudhury, Robin P
Aouadi, Myriam
Krook, Anna
Rydén, Mikael
PG/18/53/33895/BHF_/British Heart Foundation/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell Metab. 2020 Feb 4;31(2):375-390.e11. doi: 10.1016/j.cmet.2019.11.019. Epub 2019 Dec 19.
Author Address: Department of Medicine (H7), Karolinska Institutet, Stockholm 141 86, Sweden.
Department of Physiology and Pharmacology, Integrative Physiology, Karolinska Institutet, Stockholm, Sweden.
Department of Biosciences and Nutrition (BioNut), H2, Karolinska Institutet, Stockholm 141 86, Sweden.
Integrated Cardio Metabolic Centre, Department of Medicine, Karolinska Institutet, Huddinge, Sweden.
Perioperative Medicine and Intensive Care, B31, Karolinska University Hospital, Huddinge, 141 86 Stockholm, Sweden.
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, 171 77 Stockholm, Sweden.
Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford, Churchill Hospital, Old Road, Headington, Oxford OX3 7LJ, UK; Wellcome Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK; Oxford NIHR Biomedical Research Centre, Oxford University Hospitals NHS Foundation Trust, John Radcliffe Hospital, Oxford OX3 9DU, UK.
Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, UK.
Department of Medicine (H7), Karolinska Institutet, Stockholm 141 86, Sweden. Electronic address: mikael.ryden@ki.se.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 344
Author: Pfeiffer, G., Schmidt, M., Strube, K. H. and Geyer, R.
Year: 1989
Title: Carbohydrate structure of recombinant human uterine tissue plasminogen activator expressed in mouse epithelial cells
Journal: Eur J Biochem
Volume: 186
Issue: 1-2
Pages: 273-86
Epub Date: 1989/12/08
Date: Dec 8
Short Title: Carbohydrate structure of recombinant human uterine tissue plasminogen activator expressed in mouse epithelial cells
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1989.tb15206.x
Accession Number: 2513186
Keywords: Amino Acid Sequence
Animals
Asialoglycoproteins/isolation & purification
Carbohydrate Conformation
Female
Glycopeptides/isolation & purification
Humans
Methylation
Mice
Oligosaccharides/isolation & purification
Recombinant Proteins
*Tissue Plasminogen Activator
Uterus
Abstract: Recombinant human uterine tissue plasminogen activator (tPA), in part metabolically labeled with [6-3H]glucosamine or [35S]sulfate, was isolated from mouse epithelial cells (C127). Oligosaccharides present were liberated by treatment of tryptic glycopeptides with endo-beta-N-acetylglucosaminidase H or peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F and fractionated by high-performance liquid chromatography. The glycans were characterized by digestion with exoglycosidases, methylation analysis and, in part, by acetolysis and 1H-NMR spectroscopy. Glycopeptides comprising individual glycosylation sites were identified by N-terminal amino acid sequencing. The results demonstrate that recombinant tPA from C127 cells carries at Asn117 oligomannosidic glycans with 5-8 mannose residues as well as small amounts of hybrid-type species. Asn184 is only partially glycosylated and substituted by fucosylated triantennary and small amounts of diantennary N-acetyllactosaminic glycans. Likewise, Asn448 carries predominantly fucosylated triantennary species, in addition to, small amounts of diantennary and tetraantennary oligosaccharides. As a characteristic feature, part of the triantennary glycans at Asn184 and Asn448 contain additional Gal(alpha 1-3) substituents and/or sulfate groups linked to position six of beta-galactosyl residues forming NeuAc(alpha 2-3)[HO3S-6]Gal(beta 1-4) units. Oligosaccharides attached to Asn448 are almost completely substituted by (alpha 2-3)- or (alpha 2-6)-linked sialic acid residues and carry the majority of sulfate groups present. Glycans at Asn184 were found to be less sialylated and sulfated.
Notes: Pfeiffer, G
Schmidt, M
Strube, K H
Geyer, R
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1989 Dec 8;186(1-2):273-86. doi: 10.1111/j.1432-1033.1989.tb15206.x.
Author Address: Biochemisches Institut am Klinikum, Universität Giessen, FRG.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1983
Author: Phoomak, C., Park, D., Silsirivanit, A., Sawanyawisuth, K., Vaeteewoottacharn, K., Detarya, M., Wongkham, C., Lebrilla, C. B. and Wongkham, S.
Year: 2019
Title: O-GlcNAc-induced nuclear translocation of hnRNP-K is associated with progression and metastasis of cholangiocarcinoma
Journal: Mol Oncol
Volume: 13
Issue: 2
Pages: 338-357
Epub Date: 2018/11/18
Date: Feb
Short Title: O-GlcNAc-induced nuclear translocation of hnRNP-K is associated with progression and metastasis of cholangiocarcinoma
Alternate Journal: Molecular oncology
ISSN: 1574-7891 (Print)
1574-7891
DOI: 10.1002/1878-0261.12406
PMCID: PMC6360360
Accession Number: 30444036
Keywords: Bile Duct Neoplasms/*metabolism/*pathology
Biomarkers, Tumor/metabolism
Cell Line, Tumor
Cell Movement
Cell Nucleus/*metabolism
Cell Proliferation
Cholangiocarcinoma/*metabolism/*pathology
*Disease Progression
Female
Glucosamine/*metabolism
Glycosylation
Heterogeneous-Nuclear Ribonucleoprotein K/*metabolism
Humans
Male
Middle Aged
Multivariate Analysis
Neoplasm Invasiveness
Neoplasm Metastasis
Protein Transport
Treatment Outcome
*O-GlcNAcylated proteins
*bile duct cancer
*heterogeneous nuclear ribonucleoprotein-K
*metastasis
Abstract: O-GlcNAcylation is a key post-translational modification that modifies the functions of proteins. Associations between O-GlcNAcylation, shorter survival of cholangiocarcinoma (CCA) patients, and increased migration/invasion of CCA cell lines have been reported. However, the specific O-GlcNAcylated proteins (OGPs) that participate in promotion of CCA progression are poorly understood. OGPs were isolated from human CCA cell lines, KKU-213 and KKU-214, using a click chemistry-based enzymatic labeling system, identified using LC-MS/MS, and searched against an OGP database. From the proteomic analysis, a total of 21 OGPs related to cancer progression were identified, of which 12 have not been previously reported. Among these, hnRNP-K, a multifaceted RNA- and DNA-binding protein known as a pre-mRNA-binding protein, was one of the most abundantly expressed, suggesting its involvement in CCA progression. O-GlcNAcylation of hnRNP-K was further verified by anti-OGP/anti-hnRNP-K immunoprecipitations and sWGA pull-down assays. The perpetuation of CCA by hnRNP-K was evaluated using siRNA, which revealed modulation of cyclin D1, XIAP, EMT markers, and MMP2 and MMP7 expression. In native CCA cells, hnRNP-K was primarily localized in the nucleus; however, when O-GlcNAcylation was suppressed, hnRNP-K was retained in the cytoplasm. These data signify an association between nuclear accumulation of hnRNP-K and the migratory capabilities of CCA cells. In human CCA tissues, expression of nuclear hnRNP-K was positively correlated with high O-GlcNAcylation levels, metastatic stage, and shorter survival of CCA patients. This study demonstrates the significance of O-GlcNAcylation on the nuclear translocation of hnRNP-K and its impact on the progression of CCA.
Notes: 1878-0261
Phoomak, Chatchai
Park, Dayoung
Silsirivanit, Atit
Sawanyawisuth, Kanlayanee
Vaeteewoottacharn, Kulthida
Detarya, Marutpong
Wongkham, Chaisiri
Lebrilla, Carlito B
Wongkham, Sopit
DBG5980004/Thailand Research Fund and Medical Research Council-UK/International
59151/Post-Doctoral Training Program from Research Affairs and Graduate School, Khon Kaen University, Thailand/International
(NRU592009)/National Research University Grant, Khon Kaen University/International
R01GM049077/US National Institutes of Health/International
R01 GM049077/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Oncol. 2019 Feb;13(2):338-357. doi: 10.1002/1878-0261.12406. Epub 2019 Jan 10.
Author Address: Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Thailand.
Cholangiocarcinoma Research Institute, Khon Kaen University, Thailand.
Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA.
Department of Chemistry, University of California, Davis, CA, USA.
Center for Translational Medicine, Faculty of Medicine, Khon Kaen University, Thailand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1837
Author: Phoomak, C., Vaeteewoottacharn, K., Silsirivanit, A., Saengboonmee, C., Seubwai, W., Sawanyawisuth, K., Wongkham, C. and Wongkham, S.
Year: 2017
Title: High glucose levels boost the aggressiveness of highly metastatic cholangiocarcinoma cells via O-GlcNAcylation
Journal: Sci Rep
Volume: 7
Pages: 43842
Epub Date: 2017/03/07
Date: Mar 6
Short Title: High glucose levels boost the aggressiveness of highly metastatic cholangiocarcinoma cells via O-GlcNAcylation
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/srep43842
PMCID: PMC5338328
Accession Number: 28262738
Keywords: Acetylglucosamine/*metabolism
Acylation
Bile Duct Neoplasms/*metabolism/pathology
Cell Line, Tumor
Cell Movement/*drug effects
Cholangiocarcinoma/*metabolism/pathology
Dose-Response Relationship, Drug
Glucose/metabolism/*pharmacology
Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing)/metabolism
Humans
N-Acetylglucosaminyltransferases/metabolism
Neoplasm Metastasis
Nitrogenous Group Transferases
Vimentin/metabolism
Abstract: Increased glucose utilization is a feature of cancer cells to support cell survival, proliferation, and metastasis. An association between diabetes mellitus and cancer progression was previously demonstrated in cancers including cholangiocarcinoma (CCA). This study was aimed to determine the effects of high glucose on protein O-GlcNAcylation and metastatic potentials of CCA cells. Two pairs each of the parental low metastatic and highly metastatic CCA sublines were cultured in normal (5.6 mM) or high (25 mM) glucose media. The migration and invasion abilities were determined and underlying mechanisms were explored. Results revealed that high glucose promoted migration and invasion of CCA cells that were more pronounced in the highly metastatic sublines. Concomitantly, high glucose increased global O-GlcNAcylated proteins, the expressions of vimentin, hexokinase, glucosamine-fructose-6-phosphate amidotransferase (GFAT) and O-GlcNAc transferase of CCA cells. The glucose level that promoted migration/invasion was shown to be potentiated by the induction of GFAT, O-GlcNAcylation and an increase of O-GlcNAcylated vimentin and vimentin expression. Treatment with a GFAT inhibitor reduced global O-GlcNAcylated proteins, vimentin expression, and alleviated cell migration. Altogether, these results suggested the role of high glucose enhanced CCA metastasis via modulation of O-GlcNAcylation, through the expressions of GFAT and vimentin.
Notes: 2045-2322
Phoomak, Chatchai
Vaeteewoottacharn, Kulthida
Silsirivanit, Atit
Saengboonmee, Charupong
Seubwai, Wunchana
Sawanyawisuth, Kanlayanee
Wongkham, Chaisiri
Wongkham, Sopit
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2017 Mar 6;7:43842. doi: 10.1038/srep43842.
Author Address: Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002, Thailand.
Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002, Thailand.
Department of Forensic Medicine, Faculty of Medicine, Khon Kaen University, Khon Kaen, 40002, Thailand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1597
Author: Phueaouan, T., Chaiyawat, P., Netsirisawan, P., Chokchaichamnankit, D., Punyarit, P., Srisomsap, C., Svasti, J. and Champattanachai, V.
Year: 2013
Title: Aberrant O-GlcNAc-modified proteins expressed in primary colorectal cancer
Journal: Oncol Rep
Volume: 30
Issue: 6
Pages: 2929-36
Epub Date: 2013/10/16
Date: Dec
Short Title: Aberrant O-GlcNAc-modified proteins expressed in primary colorectal cancer
Alternate Journal: Oncology reports
ISSN: 1021-335x
DOI: 10.3892/or.2013.2794
Accession Number: 24126823
Keywords: Acetylglucosamine/*metabolism
Colorectal Neoplasms/genetics/*metabolism/pathology
*Gene Expression Regulation, Neoplastic
Glycosylation
Hexosamines/metabolism
Humans
N-Acetylglucosaminyltransferases/metabolism
Neoplasm Proteins/*isolation & purification/metabolism
Protein Processing, Post-Translational
Tandem Mass Spectrometry
beta-N-Acetylhexosaminidases/metabolism
Abstract: O-GlcNAcylation is a post-translational modification of serine and threonine residues which is dynamically regulated by 2 enzymes; O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) that catalyze the addition and removal of a single N-acetylglucosamine (GlcNAc) molecule, respectively. This modification is thought to be a nutrient sensor in highly proliferating cells via the hexosamine biosynthesis pathway, a minor branch of glycolysis. Although emerging evidence suggests that O-GlcNAc modification is associated with many types of cancer, identification of O-GlcNAc-modified proteins and their role in cancer remain unexplored. In the present study, we demonstrated that O-GlcNAcylation is increased in primary colorectal cancer tissues, and that this augmentation is associated with an increased expression of OGT levels. Using 2-dimensional O-GlcNAc immunoblotting and LC-MS/MS analysis, 16 proteins were successfully identified and 8 proteins showed an increase in O-GlcNAcylation, including cytokeratin 18, heterogeneous nuclear ribonucleoproteins A2/B1 (hnRNP A2/B1), hnRNP H, annexin A2, annexin A7, laminin-binding protein, α-tubulin and protein DJ-1. Among these identified proteins, annexin A2 was further confirmed to show overexpression of O-GlcNAc in all cancer samples. The results, therefore, indicate that aberrant O-GlcNAcylation of proteins is associated with colorectal cancer and that identification of O-GlcNAc-modified proteins may provide novel biomarkers of cancer.
Notes: 1791-2431
Phueaouan, Thanong
Chaiyawat, Parunya
Netsirisawan, Pukkavadee
Chokchaichamnankit, Daranee
Punyarit, Phaibul
Srisomsap, Chantragan
Svasti, Jisnuson
Champattanachai, Voraratt
Journal Article
Research Support, Non-U.S. Gov't
Greece
Oncol Rep. 2013 Dec;30(6):2929-36. doi: 10.3892/or.2013.2794. Epub 2013 Oct 11.
Author Address: Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok, Thailand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1197
Author: Picariello, G., Ferranti, P., Mamone, G., Roepstorff, P. and Addeo, F.
Year: 2008
Title: Identification of N-linked glycoproteins in human milk by hydrophilic interaction liquid chromatography and mass spectrometry
Journal: Proteomics
Volume: 8
Issue: 18
Pages: 3833-47
Epub Date: 2008/09/10
Date: Sep
Short Title: Identification of N-linked glycoproteins in human milk by hydrophilic interaction liquid chromatography and mass spectrometry
Alternate Journal: Proteomics
ISSN: 1615-9853
DOI: 10.1002/pmic.200701057
Accession Number: 18780401
Keywords: Chromatography, Liquid/methods
Female
Glycopeptides/analysis
Glycoproteins/*analysis
Glycosylation
Humans
Milk Proteins/*analysis
Milk, Human/*chemistry
Proteomics
Spectrometry, Mass, Electrospray Ionization/methods
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Breastfeeding is now generally recognized as a critical factor in protecting newborns against infections. An important mechanism responsible for the antibacterial and antiviral effects of breast milk is the prevention of pathogen adhesion to host cell membranes mediated by a number of glycoconjugates, also including glycoproteins. A number of approaches to describe the complexity of human milk proteome have provided only a partial characterization of restricted classes of N-linked glycoproteins. To achieve this objective, profiling N-linked glycoproteins of human milk was performed by Hydrophilic Interaction LC (HILIC) and MS analysis. Glycopeptides were selectively enriched from the protein tryptic digest of human milk samples. Oligosaccharide-free peptides obtained by peptide N-glycosidase F (PNGase F) treatment were characterized by a shotgun MS-based approach, allowing the identification of N-glycosylated sites localized on proteins. Using this strategy, 32 different glycoproteins were identified and 63 N-glycosylated sites encrypted in them were located. The glycoproteins include immunocompetent factors, membrane fat globule-associated proteins, enzymes involved in lipid degradation and cell differentiation, specific receptors, and other gene products with still unknown functions.
Notes: 1615-9861
Picariello, Gianluca
Ferranti, Pasquale
Mamone, Gianfranco
Roepstorff, Peter
Addeo, Francesco
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2008 Sep;8(18):3833-47. doi: 10.1002/pmic.200701057.
Author Address: Istituto di Scienze dell'Alimentazione-CNR, Avellino, Italy. picariello@isa.cnr.it
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 793
Author: Pickford, A. R., Smith, S. P., Staunton, D., Boyd, J. and Campbell, I. D.
Year: 2001
Title: The hairpin structure of the (6)F1(1)F2(2)F2 fragment from human fibronectin enhances gelatin binding
Journal: Embo j
Volume: 20
Issue: 7
Pages: 1519-29
Epub Date: 2001/04/04
Date: Apr 2
Short Title: The hairpin structure of the (6)F1(1)F2(2)F2 fragment from human fibronectin enhances gelatin binding
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/20.7.1519
PMCID: PMC145459
Accession Number: 11285216
Keywords: Binding Sites
Crystallography, X-Ray
Fibronectins/*chemistry/metabolism
Gelatin/*chemistry/metabolism
Humans
Models, Molecular
Peptide Fragments/*chemistry/metabolism
Protein Structure, Secondary
Protein Structure, Tertiary
Solutions
Abstract: The solution structure of the (6)F1(1)F2(2)F2 fragment from the gelatin-binding region of fibronectin has been determined (Protein Data Bank entry codes 1e88 and 1e8b). The structure reveals an extensive hydrophobic interface between the non-contiguous (6)F1 and (2)F2 modules. The buried surface area between (6)F1 and (2)F2 ( approximately 870 A(2)) is the largest intermodule interface seen in fibronectin to date. The dissection of (6)F1(1)F2(2)F2 into the (6)F1(1)F2 pair and (2)F2 results in near-complete loss of gelatin-binding activity. The hairpin topology of (6)F1(1)F2(2)F2 may facilitate intramolecular contact between the matrix assembly regions flanking the gelatin-binding domain. This is the first high-resolution study to reveal a compact, globular arrangement of modules in fibronectin. This arrangement is not consistent with the view that fibronectin is simply a linear 'string of beads'.
Notes: 1460-2075
Pickford, A R
Smith, S P
Staunton, D
Boyd, J
Campbell, I D
Journal Article
EMBO J. 2001 Apr 2;20(7):1519-29. doi: 10.1093/emboj/20.7.1519.
Author Address: Department of Biochemistry and Oxford Centre for Molecular Sciences, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 358
Author: Pierce-Crétel, A., Decottignies, J. P., Wieruszeski, J. M., Strecker, G., Montreuil, J. and Spik, G.
Year: 1989
Title: Primary structure of twenty three neutral and monosialylated oligosaccharides O-glycosidically linked to the human secretory immunoglobulin A hinge region determined by a combination of permethylation analysis and 400-MHz 1H-NMR spectroscopy
Journal: Eur J Biochem
Volume: 182
Issue: 2
Pages: 457-76
Epub Date: 1989/06/15
Date: Jun 15
Short Title: Primary structure of twenty three neutral and monosialylated oligosaccharides O-glycosidically linked to the human secretory immunoglobulin A hinge region determined by a combination of permethylation analysis and 400-MHz 1H-NMR spectroscopy
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1989.tb14853.x
Accession Number: 2737211
Keywords: Chromatography, High Pressure Liquid
Glycosides/*analysis
Humans
Immunoglobulin A/*analysis
Magnetic Resonance Spectroscopy/methods
Mass Spectrometry/methods
Methylation
Milk, Human/*analysis/immunology
Molecular Structure
Oligosaccharides/*analysis
Sugar Alcohols/analysis
Abstract: Sialooligosaccharide and asialooligosaccharide alditols, derived from the human milk secretory immunoglobulin A hinge region, have been purified by HPLC using, successively, an amino-bonded silica column and an octadecyl-bonded silica column. Their primary structures were completely resolved by applying a combination of sugar analysis, methylation mass spectrometry and 400-MHz 1H-NMR spectroscopy. In the present report, twenty three oligosaccharide alditols are described and all possess a type two core consisting of the branched trisaccharide: Gal(beta 1-3)[GlcNAc(beta 1-6)]GalNAc-ol. The elongation of this core arises by chain lengthening only on the GlcNAc(beta 1-6) branch for the sialylated compounds, leading to a dodecasaccharide, and on both branches for the neutral compounds leading to a nonasaccharide. Sixteen of the described oligosaccharide structures are original. Moreover, some of the fucosylated structures were found to support Le(a), Le(b) and X blood group determinants.
Notes: Pierce-Crétel, A
Decottignies, J P
Wieruszeski, J M
Strecker, G
Montreuil, J
Spik, G
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1989 Jun 15;182(2):457-76. doi: 10.1111/j.1432-1033.1989.tb14853.x.
Author Address: Laboratoire de Chimie Biologique, Université des Sciences et Techniques de Lille Flandres-Artois, Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 491
Author: Pierce-Cretel, A., Pamblanco, M., Strecker, G., Montreuil, J. and Spik, G.
Year: 1981
Title: Heterogeneity of the glycans O-glycosidically linked to the hinge region of secretory immunoglobulins from human milk
Journal: Eur J Biochem
Volume: 114
Issue: 1
Pages: 169-78
Epub Date: 1981/01/01
Short Title: Heterogeneity of the glycans O-glycosidically linked to the hinge region of secretory immunoglobulins from human milk
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1981.tb06188.x
Accession Number: 7215351
Keywords: Amino Acids/analysis
Carbohydrates/analysis
Chemical Phenomena
Chemistry
Humans
Immunoglobulin A/*analysis
Immunoglobulin A, Secretory/*analysis
Magnetic Resonance Spectroscopy
Mass Spectrometry
Methylation
Milk, Human/*analysis
Oligosaccharides/*analysis
Abstract: Pure secretory immunoglobulin A was isolated from human milk by fractionation in gradients of pH and (NH4)2SO4 concentration followed by gel filtration. The hinge region containing all the O-glycosidically linked oligosaccharides was isolated en bloc after trypsin and pepsin hydrolysis and separated by gel filtration. The mixture of O-glycosidically linked oligosaccharides contained N-acetylneuraminic acid (NeuAc), fucose (Fuc), galactose (Gal), N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc) in the molar ratio of 0.2:0.5:2.5:2:1 respectively. After beta-elimination several oligosaccharides were separated by a combination of ion-exchange chromatography and gel-filtration chromatography. The complete structure of four of these oligosaccharides was determined by methanolysis, methylation and mass spectrometry. The structure of the four oligosaccharides which are linked to serine or threonine residues of the hinge region are as follows: beta-Gal-(1 leads to 3)-GalNAc-ol; alpha-HeuAc-(2 leads to 3)-beta-Gal-(1 leads to 3)-GalNAc-ol; beta-Gal-(1 leads to 3)-[beta-GlcNAc-(1 leads to 6)]-GalNAc-ol; beta-Gal-(1 leads to 3)-[beta-Gal-(1 leads to 4)-beta-GlcNac-(1 leads to 6)]-Gal-NAc-ol. These oligosaccharides are more complex and heterogenous than the oligosaccharides linked to serine residues of the hinge region from myeloma serum immunoglobulin A1.
Notes: Pierce-Cretel, A
Pamblanco, M
Strecker, G
Montreuil, J
Spik, G
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1981;114(1):169-78. doi: 10.1111/j.1432-1033.1981.tb06188.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 490
Author: Pierce-Cretel, A., Pamblanco, M., Strecker, G., Montreuil, J., Spik, G., Dorland, L., Van Halbeek, H. and Vliegenthart, J. F.
Year: 1982
Title: Primary structure of the N-glycosidically linked sialoglycans of secretory immunoglobulins A from human milk
Journal: Eur J Biochem
Volume: 125
Issue: 2
Pages: 383-8
Epub Date: 1982/07/01
Date: Jul
Short Title: Primary structure of the N-glycosidically linked sialoglycans of secretory immunoglobulins A from human milk
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1982.tb06694.x
Accession Number: 7117239
Keywords: Alkalies
Chemical Phenomena
Chemistry
Chromatography, Gel
Chromatography, Ion Exchange
Glycosides/*isolation & purification
Humans
Immunoglobulin A/*isolation & purification
Immunoglobulin A, Secretory/*isolation & purification
Milk, Human/*immunology
Sialoglycoproteins/*isolation & purification
Abstract: The alkali-stable sialoglycopeptides of secretory immunoglobulins A from human milk have been separated from the alkali-labile glycopeptides by gel filtration and from the asialoglycopeptides by ion-exchange chromatography. The structures of five of them have been determined on the basis of the results obtained by methylation analysis, mass spectrometry and 360 MHz 1H-NMR spectroscopy. For glycopeptide B, the following structure has been found: (formula; see text) The other glycopeptides can be considered as extensions of this structure. The following extensions to Gal-6' are proposed: NeuAc(alpha 2-6) (glycopeptide A), Gal(beta 1-3) (glycopeptide D) and Fuc(alpha 1-6) (glycopeptide E). Furthermore, in glycopeptide C a fucose residue in (alpha 1-3) linkage to GlcNAc-5' could be traced.
Notes: Pierce-Cretel, A
Pamblanco, M
Strecker, G
Montreuil, J
Spik, G
Dorland, L
Van Halbeek, H
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1982 Jul;125(2):383-8. doi: 10.1111/j.1432-1033.1982.tb06694.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 305
Author: Piller, F., Le Deist, F., Weinberg, K. I., Parkman, R. and Fukuda, M.
Year: 1991
Title: Altered O-glycan synthesis in lymphocytes from patients with Wiskott-Aldrich syndrome
Journal: J Exp Med
Volume: 173
Issue: 6
Pages: 1501-10
Epub Date: 1991/06/01
Date: Jun 1
Short Title: Altered O-glycan synthesis in lymphocytes from patients with Wiskott-Aldrich syndrome
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.173.6.1501
PMCID: PMC2190829
Accession Number: 2033371
Keywords: Antigens, CD/chemistry/*metabolism
Carbohydrate Sequence
Flow Cytometry
Glycosylation
Hexosyltransferases/metabolism
Humans
Leukosialin
Lymphocyte Activation
Lymphocyte Subsets/chemistry
Lymphocytes/*metabolism
Molecular Sequence Data
Molecular Weight
Oligosaccharides/chemistry
Protein Processing, Post-Translational
Sialoglycoproteins/chemistry/*metabolism
Wiskott-Aldrich Syndrome/*metabolism
Abstract: The only molecular defect reported for the X-linked immunodeficiency Wiskott-Aldrich syndrome (WAS) is the abnormal electrophoretic behavior of the major T lymphocyte sialoglycoprotein CD43. Since the 70 to 80 O-linked carbohydrate chains of CD43 are known to influence markedly its electrophoretic mobility, we analyzed the structure and the biosynthesis of O-glycans of CD43 in lymphocytes from patients with WAS. Immunofluorescence analysis with the carbohydrate dependent anti-CD43 antibody T305 revealed that in 10 out of the 12 WAS patients tested increased numbers of T lymphocytes carry on CD43 an epitope which on normal lymphocytes is expressed only after activation. Other activation antigens were absent from WAS lymphocytes. Western blots of WAS cell lysates displayed a high molecular mass form of CD43 which reacted with the T305 antibody and which could be found on in vivo activated lymphocytes but was absent from normal unstimulated lymphocytes. To examine the O-glycan structures, carbohydrate labeled CD43 was immunoprecipitated and the released oligosaccharides identified. WAS lymphocyte CD43 was found to carry predominantly the branched structure NeuNAc alpha 2----3Gal beta 1----3 (NeuNAc alpha 2----3Gal beta 1----4G1cNAc beta 1----6) GalNAcOH whereas normal lymphocytes carry the structure NeuNAc alpha 2----3Gal beta 1----3 (NeuNAc alpha 2----6) GalNAcOH. Only after activation NeuNAc alpha 2----3Gal beta 1----3 (NeuNAc alpha 2----3Gal beta 1----4GlcNAc beta 1----6) GalNAcOH becomes the principal oligosaccharide on CD43 from normal lymphocytes. Analyzing the six glycosyltransferases involved in the biosynthesis of these O-glycan structures it was found that in WAS lymphocytes high levels of beta 1----6 N-acetyl-glucosaminyl transferase are responsible for the expression of NeuNAc alpha 2----3Gal beta 1----3 (NeuNAc alpha 2----3Gal beta 1----4GlcNAc beta 1----6) GalNAcOH on CD43. The gene responsible for WAS has not yet been identified but the results presented in this study suggest that the primary defect in WAS may affect a gene which is involved in the regulation of O-glycosylation.
Notes: 1540-9538
Piller, F
Le Deist, F
Weinberg, K I
Parkman, R
Fukuda, M
CA-33895/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
J Exp Med. 1991 Jun 1;173(6):1501-10. doi: 10.1084/jem.173.6.1501.
Author Address: La Jolla Cancer Research Foundation, California 92037.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 368
Author: Piller, F., Piller, V., Fox, R. I. and Fukuda, M.
Year: 1988
Title: Human T-lymphocyte activation is associated with changes in O-glycan biosynthesis
Journal: J Biol Chem
Volume: 263
Issue: 29
Pages: 15146-50
Epub Date: 1988/10/15
Date: Oct 15
Short Title: Human T-lymphocyte activation is associated with changes in O-glycan biosynthesis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2971663
Keywords: *Antigens, CD
Carbohydrate Conformation
Carbohydrate Sequence
Humans
Immune Sera
In Vitro Techniques
Leukosialin
*Lymphocyte Activation
Molecular Sequence Data
Oligosaccharides/isolation & purification
Polysaccharides/*biosynthesis
Sialoglycoproteins/*biosynthesis/isolation & purification
T-Lymphocytes/*immunology/metabolism
Abstract: The activation of human T-lymphocytes by anti-CD3 antibodies and interleukin-2 results in a marked increase in apparent molecular weight of the major cell-surface sialoglycoprotein. Both forms of the sialoglycoprotein were identified as leukosialin by a monospecific antiserum, and the differences in molecular weight were found to be due to changes in the carbohydrate structures. Our results suggest that resting T-lymphocytes express on leukosialin the disialotetrasaccharides NeuNAc alpha 2----3Gal beta 1----3(NeuNAc alpha 2----6)Gal-NAc-Ser/Thr, whereas activated human T-cells carry on leukosialin exclusively the more complex structures NeuNAc alpha 2----3Gal beta 1----3(NeuNAc alpha 2----3Gal beta 1----4GlcNAc beta 1----6)GalNAc-Ser/Thr. The radical shift in the biosynthetic pathway of O-glycans in activated T-lymphocytes compared to resting cells is apparently caused by a decrease of alpha 2----6 sialyltransferase activity and by the parallel dramatic stimulation of the beta 1----6GlcNAc-transferase. Since both enzymes compete for the same precursor substrate, the coordinate changes in their activities are most likely responsible for the complete change of the carbohydrate structures on leukosialin during the activation of human T-lymphocytes.
Notes: Piller, F
Piller, V
Fox, R I
Fukuda, M
CA33895/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1988 Oct 15;263(29):15146-50.
Author Address: Cancer Research Center, La Jolla Cancer Research Foundation, California.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1716
Author: Piper, D. E., Romanow, W. G., Gunawardane, R. N., Fordstrom, P., Masterman, S., Pan, O., Thibault, S. T., Zhang, R., Meininger, D., Schwarz, M., Wang, Z., King, C., Zhou, M. and Walker, N. P.
Year: 2015
Title: The high-resolution crystal structure of human LCAT
Journal: J Lipid Res
Volume: 56
Issue: 9
Pages: 1711-9
Epub Date: 2015/07/22
Date: Sep
Short Title: The high-resolution crystal structure of human LCAT
Alternate Journal: Journal of lipid research
ISSN: 0022-2275 (Print)
0022-2275
DOI: 10.1194/jlr.M059873
PMCID: PMC4548775
Accession Number: 26195816
Keywords: Cholesterol/genetics/*metabolism
Crystallography, X-Ray
Humans
Mutation
Phosphatidylcholine-Sterol O-Acyltransferase/*chemistry/*genetics/metabolism
Protein Binding
Protein Conformation
Signal Transduction
X-ray crystallography
antibodies
cholesterol
high density lipoprotein
lecithin:cholesterol acyltransferase
Abstract: LCAT is intimately involved in HDL maturation and is a key component of the reverse cholesterol transport (RCT) pathway which removes excess cholesterol molecules from the peripheral tissues to the liver for excretion. Patients with loss-of-function LCAT mutations exhibit low levels of HDL cholesterol and corneal opacity. Here we report the 2.65 Å crystal structure of the human LCAT protein. Crystallization required enzymatic removal of N-linked glycans and complex formation with a Fab fragment from a tool antibody. The crystal structure reveals that LCAT has an α/β hydrolase core with two additional subdomains that play important roles in LCAT function. Subdomain 1 contains the region of LCAT shown to be required for interfacial activation, while subdomain 2 contains the lid and amino acids that shape the substrate binding pocket. Mapping the naturally occurring mutations onto the structure provides insight into how they may affect LCAT enzymatic activity.
Notes: 1539-7262
Piper, Derek E
Romanow, William G
Gunawardane, Ruwanthi N
Fordstrom, Preston
Masterman, Stephanie
Pan, Oscar
Thibault, Stephen T
Zhang, Richard
Meininger, David
Schwarz, Margrit
Wang, Zhulun
King, Chadwick
Zhou, Mingyue
Walker, Nigel P C
Journal Article
J Lipid Res. 2015 Sep;56(9):1711-9. doi: 10.1194/jlr.M059873. Epub 2015 Jul 20.
Author Address: Therapeutic Discovery Amgen Inc., South San Francisco, CA 94080.
Therapeutic Discovery, Amgen Inc., Seattle, WA 98119.
Metabolic Disorders, Amgen Inc., South San Francisco, CA 94080.
Therapeutic Discovery, Amgen Inc., Burnaby, BC V5A1V7, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 638
Author: Pirie-Shepherd, S. R., Stevens, R. D., Andon, N. L., Enghild, J. J. and Pizzo, S. V.
Year: 1997
Title: Evidence for a novel O-linked sialylated trisaccharide on Ser-248 of human plasminogen 2
Journal: J Biol Chem
Volume: 272
Issue: 11
Pages: 7408-11
Epub Date: 1997/03/14
Date: Mar 14
Short Title: Evidence for a novel O-linked sialylated trisaccharide on Ser-248 of human plasminogen 2
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.11.7408
Accession Number: 9054441
Keywords: Amino Acid Sequence
Chromatography, High Pressure Liquid
Glycosylation
Humans
Molecular Sequence Data
Plasminogen/*chemistry
Trisaccharides/*chemistry
Abstract: Human plasminogen, the inactive precursor of plasmin, exists in two major glycoforms. Plasminogen 1 contains an N-linked oligosaccharide at Asn-289 and an O-linked oligosaccharide at Thr-345. Plasminogen 2 is known to contain only an O-linked oligosaccharide at Thr-345. However, plasminogen 2 displays a further well documented microheterogeneity dependent on the N-acetylneuraminic acid content, which has functional consequences with regard to activation of plasminogen. The proposed structure and number of known oligosaccharide linkages in plasminogen 2 is insufficient to account for this microheterogeneity. In the present study, a combination of trypsin digestion, lectin affinity chromatography, Edman degradation amino acid sequence analysis, carbohydrate composition analysis, and mass spectrometry revealed the existence of a novel site for O-linked glycosylation on plasminogen 2 at Ser-248. Direct evidence for the structure of the carbohydrate was obtained from a combination of lectin affinity chromatography, desialylation experiments, and mass spectrometry analysis. These findings provide a structural basis for some of the observed microheterogeneity, and have implications with regard to the known functional consequences of the extent of sialylation of plasminogen.
Notes: Pirie-Shepherd, S R
Stevens, R D
Andon, N L
Enghild, J J
Pizzo, S V
HL-24066/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1997 Mar 14;272(11):7408-11. doi: 10.1074/jbc.272.11.7408.
Author Address: Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1992
Author: Pirro, M., Schoof, E., van Vliet, S. J., Rombouts, Y., Stella, A., de Ru, A., Mohammed, Y., Wuhrer, M., van Veelen, P. A. and Hensbergen, P. J.
Year: 2019
Title: Glycoproteomic Analysis of MGL-Binding Proteins on Acute T-Cell Leukemia Cells
Journal: J Proteome Res
Volume: 18
Issue: 3
Pages: 1125-1132
Epub Date: 2018/12/26
Date: Mar 1
Short Title: Glycoproteomic Analysis of MGL-Binding Proteins on Acute T-Cell Leukemia Cells
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/acs.jproteome.8b00796
PMCID: PMC6399673
Accession Number: 30582698
Keywords: Acetylgalactosamine/genetics/metabolism
Antigens, Tumor-Associated, Carbohydrate/genetics/metabolism
Carrier Proteins/genetics/immunology
Glycoproteins/*genetics/immunology
Glycosylation
Humans
Immunity, Innate/genetics
Jurkat Cells
Lectins, C-Type/*genetics/immunology
Leukocyte Common Antigens/genetics
Leukosialin/genetics
Ligands
Precursor T-Cell Lymphoblastic
Leukemia-Lymphoma/*genetics/immunology/metabolism/pathology
*O-glycosylation
*Tn antigen
*glycoproteomics
*lectin
*leukemia
Abstract: C-type lectins are a diverse group of proteins involved in many human physiological and pathological processes. Most C-type lectins are glycan-binding proteins, some of which are pivotal for innate immune responses against pathogens. Other C-type lectins, such as the macrophage galactose-type lectin (MGL), have been shown to induce immunosuppressive responses upon the recognition of aberrant glycosylation on cancer cells. MGL is known to recognize terminal N-acetylgalactosamine (GalNAc), such as the Tn antigen, which is commonly found on malignant cells. Even though this glycan specificity of MGL is well described, there is a lack of understanding of the actual glycoproteins that bind MGL. We present a glycoproteomic workflow for the identification of MGL-binding proteins, which we applied to study MGL ligands on the human Jurkat leukemia cell line. In addition to the known MGL ligands and Tn antigen-carrying proteins CD43 and CD45 on these cells, we have identified a set of novel cell-surface ligands for MGL. Importantly, for several of these, O-glycosylation has hitherto not been described. Altogether, our data provide new insight into the identification and structure of novel MGL ligands that presumably act as modulatory molecules in cancer immune responses.
Notes: 1535-3907
Pirro, Martina
Orcid: 0000-0001-9562-2395
Schoof, Esmee
van Vliet, Sandra J
Orcid: 0000-0003-1811-2687
Rombouts, Yoann
Orcid: 0000-0003-4482-2199
Stella, Alexandre
de Ru, Arnoud
Mohammed, Yassene
Orcid: 0000-0003-3265-3332
Wuhrer, Manfred
Orcid: 0000-0002-0814-4995
van Veelen, Peter A
Hensbergen, Paul J
Orcid: 0000-0002-3193-5445
Journal Article
Research Support, Non-U.S. Gov't
J Proteome Res. 2019 Mar 1;18(3):1125-1132. doi: 10.1021/acs.jproteome.8b00796. Epub 2019 Jan 9.
Author Address: Center for Proteomics and Metabolomics , Leiden University Medical Center , 2300 RC Leiden , The Netherlands.
Amsterdam UMC, Vrije Universiteit Amsterdam, Dept. of Molecular Cell Biology and Immunology, Cancer Center Amsterdam, Amsterdam Infection & Immunity Institute, 1007 MB Amsterdam , The Netherlands.
Institut de Pharmacologie et de Biologie Structurale , Université de Toulouse, CNRS, UPS , Toulouse 31062 , France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 571
Author: Pisano, A., Redmond, J. W., Williams, K. L. and Gooley, A. A.
Year: 1993
Title: Glycosylation sites identified by solid-phase Edman degradation: O-linked glycosylation motifs on human glycophorin A
Journal: Glycobiology
Volume: 3
Issue: 5
Pages: 429-35
Epub Date: 1993/10/01
Date: Oct
Short Title: Glycosylation sites identified by solid-phase Edman degradation: O-linked glycosylation motifs on human glycophorin A
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/3.5.429
Accession Number: 8286855
Keywords: Alleles
Amino Acid Sequence
Binding Sites/genetics
Chromatography, High Pressure Liquid
Glycophorins/*chemistry/genetics
Glycosylation
Humans
Molecular Sequence Data
Molecular Structure
Peptide Fragments/chemistry/isolation & purification
Trypsin
Abstract: The human red blood cell sialoglycoprotein, glycophorin A (GpA), contains a 'mucin-like' extensively O-glycosylated extracellular domain which carries the MN blood group antigens. We have revised the sites of O-glycosylation in the extracellular domain of GpA by automated solid-phase Edman degradation, which allowed positive identification and quantitation of O-glycosylated Ser and Thr residues, as well as the single N-glycosylation site. One N-linked and 16 O-linked sites were identified. Carbohydrate was absent on Ser1, Ser14, Ser15, Ser23, Thr28 and Thr58 in GpA. We propose that the glycosyltransferases present in erythrocytes recognize specific flanking sequences around potential O-glycosylation sites. All 16 O-glycosylation sites are explained on the basis of four motifs. Three motifs are associated with Thr-glycosylation: Xaa-Pro-Xaa-Xaa where at least one Xaa = Thr; Thr-Xaa-Xaa-Xaa where at least one Xaa = Thr; Xaa-Xaa-Thr-Xaa where at least one X = Arg or Lys. The fourth motif is associated with Ser-glycosylation: Ser-Xaa-Xaa-Xaa where at least one Xaa = Ser. These simple rules explain the glycosylation (or lack of it) on 21 of 22 Ser/Thr in the extracellular domain of GpA.
Notes: Pisano, A
Redmond, J W
Williams, K L
Gooley, A A
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1993 Oct;3(5):429-35. doi: 10.1093/glycob/3.5.429.
Author Address: Macquarie University Centre for Analytical Biotechnology, Macquarie, NSW, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1689
Author: Plomp, R., Dekkers, G., Rombouts, Y., Visser, R., Koeleman, C. A., Kammeijer, G. S., Jansen, B. C., Rispens, T., Hensbergen, P. J., Vidarsson, G. and Wuhrer, M.
Year: 2015
Title: Hinge-Region O-Glycosylation of Human Immunoglobulin G3 (IgG3)
Journal: Mol Cell Proteomics
Volume: 14
Issue: 5
Pages: 1373-84
Epub Date: 2015/03/12
Date: May
Short Title: Hinge-Region O-Glycosylation of Human Immunoglobulin G3 (IgG3)
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M114.047381
PMCID: PMC4424406
Accession Number: 25759508
Keywords: Adult
Amino Acid Sequence
Carbohydrate Sequence
Female
Gene Expression
Glycosylation
Humans
Immunoglobulin G/*analysis/chemistry/genetics
Male
Middle Aged
Molecular Sequence Data
Mutation
Peptides/*analysis
Proteolysis
Recombinant Proteins/analysis/chemistry/genetics
Spectrometry, Mass, Electrospray Ionization
Tandem Mass Spectrometry
Threonine/*chemistry/metabolism
Trypsin/chemistry
Abstract: Immunoglobulin G (IgG) is one of the most abundant proteins present in human serum and a fundamental component of the immune system. IgG3 represents ∼8% of the total amount of IgG in human serum and stands out from the other IgG subclasses because of its elongated hinge region and enhanced effector functions. This study reports partial O-glycosylation of the IgG3 hinge region, observed with nanoLC-ESI-IT-MS(/MS) analysis after proteolytic digestion. The repeat regions within the IgG3 hinge were found to be in part O-glycosylated at the threonine in the triple repeat motif. Non-, mono- and disialylated core 1-type O-glycans were detected in various IgG3 samples, both poly- and monoclonal. NanoLC-ESI-IT-MS/MS with electron transfer dissociation fragmentation and CE-MS/MS with CID fragmentation were used to determine the site of IgG3 O-glycosylation. The O-glycosylation site was further confirmed by the recombinant production of mutant IgG3 in which potential O-glycosylation sites had been knocked out. For IgG3 samples from six donors we found similar O-glycan structures and site occupancies, whereas for the same samples the conserved N-glycosylation of the Fc CH2 domain showed considerable interindividual variation. The occupancy of each of the three O-glycosylation sites was found to be ∼10% in six serum-derived IgG3 samples and ∼13% in two monoclonal IgG3 allotypes.
Notes: 1535-9484
Plomp, Rosina
Dekkers, Gillian
Rombouts, Yoann
Visser, Remco
Koeleman, Carolien A M
Kammeijer, Guinevere S M
Jansen, Bas C
Rispens, Theo
Hensbergen, Paul J
Vidarsson, Gestur
Wuhrer, Manfred
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2015 May;14(5):1373-84. doi: 10.1074/mcp.M114.047381. Epub 2015 Mar 10.
Author Address: From the ‡Center for Proteomics and Metabolomics.
¶Department of Experimental Immunohematology.
From the ‡Center for Proteomics and Metabolomics, §Department of Rheumatology, Leiden University Medical Center, Leiden, The Netherlands;
¶¶Department of Immunopathology, Sanquin Research, and Landsteiner Laboratory, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands;
From the ‡Center for Proteomics and Metabolomics, **Division of BioAnalytical Chemistry, VU University Amsterdam, Amsterdam, The Netherlands m.wuhrer@lumc.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1614
Author: Plomp, R., Hensbergen, P. J., Rombouts, Y., Zauner, G., Dragan, I., Koeleman, C. A., Deelder, A. M. and Wuhrer, M.
Year: 2014
Title: Site-specific N-glycosylation analysis of human immunoglobulin e
Journal: J Proteome Res
Volume: 13
Issue: 2
Pages: 536-46
Epub Date: 2013/12/07
Date: Feb 7
Short Title: Site-specific N-glycosylation analysis of human immunoglobulin e
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr400714w
Accession Number: 24308486
Keywords: Chromatography, Reverse-Phase
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Immunoglobulin E/chemistry/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tandem Mass Spectrometry
Abstract: Immunoglobulin E (IgE) is a heterodimeric glycoprotein involved in antiparasitic and allergic immune reactions. IgE glycosylation is known to exhibit significant interindividual variation, and several reports have indicated its relevance in determining IgE activity. Here, we present site-specific glycosylation analysis of IgE from three different sources: IgE from the serum of a hyperimmune donor, from the pooled serum of multiple nondiseased donors, and from the pooled serum of 2 patients with IgE myeloma. The heavy chains were isolated and digested with either trypsin, proteinase K, or chymotrypsin, which permitted coverage of all seven potential N-glycosylation sites. The resulting (glyco-)peptides were analyzed by nano-reversed-phase-LC-MS/MS and MALDI-TOF/TOF-MS/MS. Site Asn264 was shown to be unoccupied. In all three samples, site Asn275 contained exclusively oligomannosidic structures with between 2 and 9 mannoses, whereas sites Asn21, Asn49, Asn99, Asn146, and Asn252 contained exclusively complex-type glycans. For the nonmyeloma IgE, the majority of these glycans were biantennary and core-fucosylated and contained one or two terminal N-acetylneuraminic acids. In contrast, myeloma IgE showed a higher abundance of triantennary and tetraantennary glycan structures and a low abundance of species with a bisecting N-acetylglucosamine. Our approach allows comparison of the glycosylation of IgE samples in a site-specific manner.
Notes: 1535-3907
Plomp, Rosina
Hensbergen, Paul J
Rombouts, Yoann
Zauner, Gerhild
Dragan, Irina
Koeleman, Carolien A M
Deelder, André M
Wuhrer, Manfred
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2014 Feb 7;13(2):536-46. doi: 10.1021/pr400714w. Epub 2013 Dec 13.
Author Address: Center for Proteomics and Metabolomics, ‡Department of Rheumatology, Leiden University Medical Center , 2300 RC Leiden, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 431
Author: Podolsky, D. K.
Year: 1985
Title: Oligosaccharide structures of human colonic mucin
Journal: J Biol Chem
Volume: 260
Issue: 14
Pages: 8262-71
Epub Date: 1985/07/15
Date: Jul 15
Short Title: Oligosaccharide structures of human colonic mucin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4008490
Keywords: Carbohydrate Conformation
Chromatography, DEAE-Cellulose
Chromatography, Gel
Chromatography, High Pressure Liquid
Colon/*analysis
Humans
Intestinal Mucosa/analysis
Mucins/*analysis
Oligosaccharides/*analysis
Abstract: Purified human colonic mucin was separated into six distinct components by DEAE-cellulose chromatography, and the structures of oligosaccharide side chains from the three most abundant species were determined. Oligosaccharide side chains were isolated from colonic mucin species III, IV, and V after alkaline borohydride reductive cleavage in the presence of sodium borotritide. After initial separation of acidic and neutral oligosaccharides by ion exchange chromatography, individual oligosaccharides were isolated by sequential chromatography on Bio-Gel P-4 and Bio-Gel P-2 resins followed by preparative normal phase high performance liquid chromatography. Composition and structure of individual oligosaccharides were determined by combination of gas chromatography, methylation analysis, and sequential glycosidase digestion. Collectively, 21 discrete oligosaccharide structures were identified in the major human colonic mucin species including 10 acidic oligosaccharides and 11 neutral structures which ranged in size from 2 to 12 sugar residues. Although detailed structures were defined for each oligosaccharide, the majority of the structures identified were variations of a relatively small number of "basic" structures, and several generalizations pertained. First, many oligosaccharides represented variations of a biantennary structure in which branch chains arise in N-acetylglucosaminyl residues linked to C3 and C6 of a galactosyl residue linked in turn to a GlcNAc beta (1-3)GalNAc core; second, non-branched oligosaccharides appeared to be linear chain derivatives of the same core structure; third, all acidic oligosaccharides could be derived from neutral structures present in the mucin species; fourth, sialic acid substitution was limited to few sites and always included substitution in alpha 2-6 linkage to the reducing terminal N-acetylgalactosamine, and finally several structures contained both sialic acid and fucose residues. Individually, mucin species III, IV, and V were found to contain unique mixtures of 13, 14, and 10 oligosaccharide structures, respectively. These data demonstrate that human colonic mucin contain a wide range of oligosaccharides reflecting variations of common core oligosaccharide structures. The major chromatographically defined constituents of normal colonic mucin appear to possess characteristic and distinguishable combinations of oligosaccharide structures. These findings support the concept that colonic mucin contains structurally and functionally distinct subpopulations.
Notes: Podolsky, D K
AM01257/AM/NIADDK NIH HHS/United States
AM34422/AM/NIADDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1985 Jul 15;260(14):8262-71.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 435
Author: Podolsky, D. K.
Year: 1985
Title: Oligosaccharide structures of isolated human colonic mucin species
Journal: J Biol Chem
Volume: 260
Issue: 29
Pages: 15510-5
Epub Date: 1985/12/15
Date: Dec 15
Short Title: Oligosaccharide structures of isolated human colonic mucin species
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 4066681
Keywords: Carbohydrate Sequence
Chromatography, Gas
Chromatography, High Pressure Liquid
Colon/*analysis
Glycoside Hydrolases/metabolism
Humans
Methylation
Mucins/*analysis
Oligosaccharides/*analysis
Abstract: Purified human colonic mucin contains six distinct components which may be separated by DEAE-cellulose chromatography. Past studies defined the structure of oligosaccharide side chains from the most abundant species III, IV, and V which elute at intermediate salt concentrations. In these studies the structures of oligosaccharide side chains liberated from the remaining early and late eluting species I, II, and VI were determined after isolation by sequential conventional and high performance liquid chromatography through combination of gas chromatography, methylation analysis, and sequential glycosidase digestion. Mucin species I, II, and VI contained a less varied array of discrete oligosaccharide structures than that observed in the major mucin components. Mucin species I and II contained five and 10 structures, respectively, which account for 68 and 71% of total oligosaccharide content in these fractions. The predominant oligosaccharides of mucin species I included three neutral structures: a disaccharide GlcNAc beta (1-3)GalNAc-ol, a trisaccharide Gal beta (1-4)GlcNAc beta (1-3)GalNAc-ol, and a tetrasaccharide GlcNAc beta (1-4)Gal beta (1-4)GlcNAc beta (1-3)GalNAc-ol as well as two acidic components representing the sialylated forms of two of these oligosaccharides. Mucin species II contained these same oligosaccharides as well as four additional acidic structures, notably a disaccharide Neu alpha (2-6)GalNAc-ol and a hexasaccharide Gal beta (1-4)GlcNAc beta (1-3)Gal beta (1-4)GlcNAc beta (1-3) (NeuAc alpha (2-6))-GalNAc-ol, not identified in any other mucin species. The late eluting mucin species VI contained at least five discrete neutral oligosaccharides and six major acidic structures. While the majority of these structures had been previously isolated from the earlier eluting mucin species IV and V, species VI also contained di- and trisialylated oligosaccharides not identified in other mucin species. In conjunction with earlier studies of the major mucin species III, IV, and V, these data define the range of oligosaccharide structures present in human colonic mucin. These studies demonstrate that human colonic mucin possesses species with characteristic and distinguishable combinations of oligosaccharides which reflect variations of common core structures.
Notes: Podolsky, D K
AM01257/AM/NIADDK NIH HHS/United States
AM34422/AM/NIADDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1985 Dec 15;260(29):15510-5.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 326
Author: Pohl, J., Pereira, H. A., Martin, N. M. and Spitznagel, J. K.
Year: 1990
Title: Amino acid sequence of CAP37, a human neutrophil granule-derived antibacterial and monocyte-specific chemotactic glycoprotein structurally similar to neutrophil elastase
Journal: FEBS Lett
Volume: 272
Issue: 1-2
Pages: 200-4
Epub Date: 1990/10/15
Date: Oct 15
Short Title: Amino acid sequence of CAP37, a human neutrophil granule-derived antibacterial and monocyte-specific chemotactic glycoprotein structurally similar to neutrophil elastase
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(90)80484-z
Accession Number: 2226832
Keywords: Amino Acid Sequence
Antimicrobial Cationic Peptides
Blood Bactericidal Activity
Blood Proteins/*chemistry
*Carrier Proteins
Chemotactic Factors/*chemistry
Chromatography, High Pressure Liquid
Disulfides
Endopeptidases/metabolism
Glycoproteins/*chemistry
Glycosylation
Leukocyte Elastase
Metalloendopeptidases
Molecular Sequence Data
Pancreatic Elastase/*chemistry
Peptide Fragments/chemistry
Sequence Homology, Nucleic Acid
Trypsin/metabolism
Abstract: We report the amino acid sequence of CAP37, a human neutrophil granule protein with antibacterial and monocyte-specific chemotactic activity. CAP37 is a single-chain protein consisting of 222 amino acid residues. It has three N-glycosylation sites, at Asn residues 100, 114 and 145. Some species of CAP37 are glycosylated at all three sites; some at Asn-114 alone, others at Asn-114 and Asn-110 or Asn-145. CAP37 has 45% sequence identity to human neutrophil elastase, and 30-37% identity to several other granule serine proteinases. Despite these similarities, CAP37 is not a serine proteinase because the active site residues serine and histidine are replaced.
Notes: Pohl, J
Pereira, H A
Martin, N M
Spitznagel, J K
AI17662/AI/NIAID NIH HHS/United States
AI26589/AI/NIAID NIH HHS/United States
AI28018/AI/NIAID NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
FEBS Lett. 1990 Oct 15;272(1-2):200-4. doi: 10.1016/0014-5793(90)80484-z.
Author Address: Microchemical Facility, Winship Cancer Center, Atlanta, GA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1665
Author: Pompach, P., Ashline, D. J., Brnakova, Z., Benicky, J., Sanda, M. and Goldman, R.
Year: 2014
Title: Protein and site specificity of fucosylation in liver-secreted glycoproteins
Journal: J Proteome Res
Volume: 13
Issue: 12
Pages: 5561-9
Epub Date: 2014/09/30
Date: Dec 5
Short Title: Protein and site specificity of fucosylation in liver-secreted glycoproteins
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr5005482
PMCID: PMC4261953
Accession Number: 25265424
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Glycoproteins/*blood/metabolism
Glycosylation
Hepatitis C/blood
Humans
Liver/*metabolism
Liver Cirrhosis/blood/virology
Male
Middle Aged
Molecular Sequence Data
*Protein Processing, Post-Translational
MSn structural analysis
N-glycans
exoglycosidase treatment
fucosylation
glycopeptides
hepatocellular carcinoma
mass spectrometry
microheterogeneity
permethylation
site specificity
Abstract: Chronic liver diseases are a serious health problem worldwide. One of the frequently reported glycan alterations in liver disease is aberrant fucosylation, which was suggested as a marker for noninvasive serologic monitoring. We present a case study that compares site specific glycoforms of four proteins including haptoglobin, complement factor H, kininogen-1, and hemopexin isolated from the same patient. Our exoglycosidase-assisted LC-MS/MS analysis confirms the high degree of fucosylation of some of the proteins but shows that microheterogeneity is protein- and site-specific. MSn analysis of permethylated detached glycans confirms the presence of LeY glycoforms on haptoglobin, which cannot be detected in hemopexin or complement factor H; all three proteins carry Lewis and H epitopes. Core fucosylation is detectable in only trace amounts in haptoglobin but with confidence on hemopexin and complement factor H, where core fucosylation of the bi-antennary glycans on select glycopeptides reaches 15-20% intensity. These protein-specific differences in fucosylation, observed in proteins isolated from the same patient source, suggest that factors other than up-regulation of enzymatic activity regulate the microheterogeneity of glycoforms. This has implications for selection of candidate proteins for disease monitoring and suggests that site-specific glycoforms have structural determinants, which could lead to functional consequences for specific subsets of proteins or their domains.
Notes: 1535-3907
Pompach, Petr
Ashline, David J
Brnakova, Zuzana
Benicky, Julius
Sanda, Miloslav
Goldman, Radoslav
P01 HL107146/HL/NHLBI NIH HHS/United States
P30 CA051008/CA/NCI NIH HHS/United States
U01 CA168926/CA/NCI NIH HHS/United States
R01 CA135069/CA/NCI NIH HHS/United States
P30 CA51008/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Proteome Res. 2014 Dec 5;13(12):5561-9. doi: 10.1021/pr5005482. Epub 2014 Oct 10.
Author Address: Institute of Microbiology v.v.i., Czech Academy of Sciences , Videnska 1083, Prague 142 20, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 677
Author: Ponce-Castañeda, M. V., Esparza-López, J., Vilchis-Landeros, M. M., Mendoza, V. and López-Casillas, F.
Year: 1998
Title: Murine betaglycan primary structure, expression and glycosaminoglycan attachment sites
Journal: Biochim Biophys Acta
Volume: 1384
Issue: 2
Pages: 189-96
Epub Date: 1998/07/11
Date: May 19
Short Title: Murine betaglycan primary structure, expression and glycosaminoglycan attachment sites
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/s0167-4838(98)00033-8
Accession Number: 9659379
Keywords: Amino Acid Sequence
Animals
Base Sequence
Binding Sites
Glycosaminoglycans/*metabolism
Mice
Molecular Sequence Data
Myocardium/metabolism
Proteoglycans/*genetics/metabolism
Receptors, Transforming Growth Factor beta/*genetics/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: The primary structure of murine betaglycan, also known as transforming growth factor beta (TGF-beta) type III receptor, was deduced from the nucleotide sequence of a cDNA clone isolated from a heart library. Murine betaglycan is a single spanning membrane polypeptide of 850 amino acids which is highly similar to betaglycan of other species. Transfection of this cDNA into COS1 cells resulted in the expression of a membrane proteoglycan that binds TGF-beta and is recognized by antibodies raised against rat betaglycan. COS1 cells transfected with the double mutant Ser533Ala; Ser544Ala of the murine betaglycan cDNA produced a TGF-beta type III receptor devoid of glycosaminoglycan chains.
Notes: Ponce-Castañeda, M V
Esparza-López, J
Vilchis-Landeros, M M
Mendoza, V
López-Casillas, F
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 1998 May 19;1384(2):189-96. doi: 10.1016/s0167-4838(98)00033-8.
Author Address: Departmento de Biología Celular, Universidad Nacional Autónoma de México, México City.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 720
Author: Powell, M. S., Barton, P. A., Emmanouilidis, D., Wines, B. D., Neumann, G. M., Peitersz, G. A., Maxwell, K. F., Garrett, T. P. and Hogarth, P. M.
Year: 1999
Title: Biochemical analysis and crystallisation of Fc gamma RIIa, the low affinity receptor for IgG
Journal: Immunol Lett
Volume: 68
Issue: 1
Pages: 17-23
Epub Date: 1999/07/09
Date: May 3
Short Title: Biochemical analysis and crystallisation of Fc gamma RIIa, the low affinity receptor for IgG
Alternate Journal: Immunology letters
ISSN: 0165-2478 (Print)
0165-2478
DOI: 10.1016/s0165-2478(99)00025-5
Accession Number: 10397151
Keywords: Animals
Antibody Affinity
Antigens, CD/*chemistry/*isolation & purification/metabolism
Binding Sites, Antibody
CHO Cells
Cricetinae
Crystallization
Crystallography, X-Ray
Humans
Immunoglobulin G/*metabolism
Mass Spectrometry
Receptors, IgG/*chemistry/*isolation & purification/metabolism
Recombinant Proteins/isolation & purification
Solubility
Abstract: Fc gamma RIIa is one of a family of specific cell surface receptors for immunoglobulin. Fc gamma RIIa, which binds immune complexes of certain IgG isotypes, plays important roles in immune homeostasis. However, the precise characteristics of IgG binding and three-dimensional structure of Fc gamma RIIa have not been reported. This study describes the affinity of the Fc gamma RIIa:IgG interaction as well as biochemical characterisation of recombinant Fc gamma RIIa that has been used to generate high quality crystals. Equilibrium binding analysis of the Fc gamma RII:IgG interaction found, IgG3 binds with an affinity of K(D) = 0.6 microM, as expected. Unlike other Fc gamma R, IgG4 also bound to Fc gamma RIIa, K(D) = 3 microM, clearly establishing Fc gamma RIIa as an IgG4 receptor. Biochemical analysis of mammalian and insect cell derived Fc gamma RIIa established the genuine N-terminus with Q being the first amino acid in the sequence Q, A, A, A, P... extending the N-terminus further than previously thought. Furthermore, both potential N-linked glycosylation sites are occupied. Electrospray ionisation mass spectrometry (ESMS) indicate that the N-glycans of baculovirus derived Fc gamma RIIa are core mannose oligosaccharide side chains. Finally, we describe the first crystallisation of diffraction quality crystals of soluble Fc gamma RIIa. Orthorhombic crystals diffract X-rays beyond 2.1 A resolution in the space group P2(1)2(1)2 with cell dimensions a = 78.8 A, b = 100.5 A, c = 27.8 A. This marks a significant advance towards understanding the three-dimensional structure of Fc gamma RIIa and related FcR proteins that share high amino acid identity with Fc gamma RIIa.
Notes: Powell, M S
Barton, P A
Emmanouilidis, D
Wines, B D
Neumann, G M
Peitersz, G A
Maxwell, K F
Garrett, T P
Hogarth, P M
Journal Article
Netherlands
Immunol Lett. 1999 May 3;68(1):17-23. doi: 10.1016/s0165-2478(99)00025-5.
Author Address: Helen M. Schutt Trust Laboratory for Immunology, Austin Research Institute, Austin and Repatriation Medical Centre, Heidelberg, Victoria, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 236
Author: Power, S. G., Bocchinfuso, W. P., Pallesen, M., Warmels-Rodenhiser, S., Van Baelen, H. and Hammond, G. L.
Year: 1992
Title: Molecular analyses of a human sex hormone-binding globulin variant: evidence for an additional carbohydrate chain
Journal: J Clin Endocrinol Metab
Volume: 75
Issue: 4
Pages: 1066-70
Epub Date: 1992/10/01
Date: Oct
Short Title: Molecular analyses of a human sex hormone-binding globulin variant: evidence for an additional carbohydrate chain
Alternate Journal: The Journal of clinical endocrinology and metabolism
ISSN: 0021-972X (Print)
0021-972x
DOI: 10.1210/jcem.75.4.1400872
Accession Number: 1400872
Keywords: Amino Acid Sequence
Animals
Base Sequence
Blotting, Western
CHO Cells
Concanavalin A/metabolism
Cricetinae
DNA/genetics
Exons/genetics
Gene Expression
Glycosylation
Humans
Molecular Sequence Data
Point Mutation
Polymerase Chain Reaction
Protein Binding
Sex Hormone-Binding Globulin/*genetics/metabolism
Abstract: Genomic DNA was isolated from an individual who is homozygous for a sex hormone-binding globulin (SHBG) variant that resolves into three molecular weight forms of 56K, 52K, and 48K during electrophoresis under denaturing conditions (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). This material was amplified using intron-specific oligonucleotide primers in a polymerase chain reaction to obtain the eight exons encoding SHBG. Sequence analysis of these exons revealed a point mutation encoding an amino acid substitution (Asp --> Asn) at residue 327 in the SHBG polypeptide, and the same mutation was identified in three siblings who also appear to be homozygous for this trait. This mutation introduces an additional consensus site for N-glycosylation at this position, and to confirm its utilization we introduced it into a human SHBG complementary DNA. The mutated complementary DNA was inserted into the pRc/CMV expression vector, and transfected into Chinese hamster ovary (CHO) cells. The product was secreted normally but proportionally less of it (54%) bound to concanavalin A when compared to normal SHBG produced by CHO cells (85%), or SHBG in the serum of either a normal individual or those who produce an electrophoretic variant (98%). Furthermore, when subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting, the SHBG variant produced by CHO cells consisted of a 60K subunit as well as the heavy (52K) and light (48K) subunits associated with normal SHBG produced by CHO cells or in serum. This additional subunit is larger than the variant in serum and probably reflects a greater degree of complexity in the carbohydrate structures added to recombinant SHBG during synthesis in CHO cells. Nevertheless, its steroid-binding affinity was equal to normal SHBG produced by CHO cells or SHBG in serum.
Notes: Power, S G
Bocchinfuso, W P
Pallesen, M
Warmels-Rodenhiser, S
Van Baelen, H
Hammond, G L
Journal Article
Research Support, Non-U.S. Gov't
United States
J Clin Endocrinol Metab. 1992 Oct;75(4):1066-70. doi: 10.1210/jcem.75.4.1400872.
Author Address: Department of Obstetrics and Gynecology, University of Western Ontario, London, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 668
Author: Prakobphol, A., Thomsson, K. A., Hansson, G. C., Rosen, S. D., Singer, M. S., Phillips, N. J., Medzihradszky, K. F., Burlingame, A. L., Leffler, H. and Fisher, S. J.
Year: 1998
Title: Human low-molecular-weight salivary mucin expresses the sialyl lewisx determinant and has L-selectin ligand activity
Journal: Biochemistry
Volume: 37
Issue: 14
Pages: 4916-27
Epub Date: 1998/05/16
Date: Apr 7
Short Title: Human low-molecular-weight salivary mucin expresses the sialyl lewisx determinant and has L-selectin ligand activity
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi972612a
Accession Number: 9538010
Keywords: Carbohydrate Sequence
Humans
L-Selectin/*metabolism
Ligands
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Molecular Weight
Mucins/chemistry/*immunology/metabolism
Oligosaccharides/*metabolism
Protein Binding
Saliva/*chemistry
Sialyl Lewis X Antigen
Spectrometry, Mass, Fast Atom Bombardment
Abstract: Previously we showed that the low-molecular-weight mucin (MG2, encoded by MUC7), a major component of human submandibular/sublingual saliva, is a bacterial receptor that coats the tooth surface. Here we tested the hypothesis that the structure of its carbohydrate residues contains important information about its function. Purified MG2 (Mr 120 000) was digested with trypsin, and the resulting Mr 90 000 fragment, which carried primarily O-linked oligosaccharides, was subjected to reductive beta-elimination. The released oligosaccharides were characterized by using nuclear magnetic resonance spectroscopy and mass spectrometry. Of the 41 different structures we detected, the most prominent included NeuAcalpha2-->3Galbeta1-->3GalNAc-ol (sialyl-T antigen), Galbeta1-->4(Fucalpha1-->3)GlcNAcbeta1-->6(Galbeta1 -->3)GalNAc-ol [type 2 core with Lewisx (Lex) determinant], and NeuAcalpha2-->3Galbeta1-->4(Fucalpha1-->3)GlcNAcbet a1-->6(Galbeta1--> 3) GalNAc-ol [type 2 core with sialyl Lex (sLex) determinant]. We also detected di-, tri-, and pentasaccharides with one sulfate group. Lex, sLex, and related sulfated structures are ligands for selectins, adhesion molecules that mediate leukocyte trafficking. Therefore, we investigated whether MG2 was a selectin ligand. In an enzyme-linked immunosorbent assay, L-selectin chimeras interacted with immobilized MG2 in a Ca2+-dependent manner. L-Selectin chimeras also bound to MG2 immobilized on nitrocellulose. Together, these results suggest that the saccharides that MG2 carries could specify some of its important functions, which may include mediating leukocyte interactions in the oral cavity.
Notes: Prakobphol, A
Thomsson, K A
Hansson, G C
Rosen, S D
Singer, M S
Phillips, N J
Medzihradszky, K F
Burlingame, A L
Leffler, H
Fisher, S J
DE 07244/DE/NIDCR NIH HHS/United States
GM23547/GM/NIGMS NIH HHS/United States
RR 01614/RR/NCRR NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1998 Apr 7;37(14):4916-27. doi: 10.1021/bi972612a.
Author Address: Department of Stomatology, University of California, San Francisco, San Francisco, California 94143, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 483
Author: Prasad, R. V., Butkowski, R. J., Hamilton, J. W. and Ebner, K. E.
Year: 1982
Title: Amino acid sequence of rat alpha-lactalbumin: a unique alpha-lactalbumin
Journal: Biochemistry
Volume: 21
Issue: 7
Pages: 1479-82
Epub Date: 1982/03/30
Date: Mar 30
Short Title: Amino acid sequence of rat alpha-lactalbumin: a unique alpha-lactalbumin
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00536a002
Accession Number: 7044414
Keywords: Amino Acid Sequence
Animals
Carboxypeptidase B
Carboxypeptidases
Cyanogen Bromide
Endopeptidases
*Lactalbumin/isolation & purification
*Metalloendopeptidases
Peptide Fragments/isolation & purification
Rats
Abstract: The amino acid sequence of rat alpha-lactalbumin has been determined. Unlike other alpha-lactalbumins which contain 122 or 123 amino acids, rat alpha-lactalbumin is unique in that it contains 140 amino acids. The extra amino acids are a 17 amino acid extension at the carboxyl terminus. The amino acid sequence of this extension is Gly124-Ala-Pro-Ala-Leu-Val-Val130-Pro-Ala-Leu-Asp-Gly135-Glu-Thr-Pro-Val-Pro140 . The extension is proline rich, which may contribute to the anomalous structural properties of rat alpha-lactalbumin. The amino acid sequence from residues 1 to 123 is similar to that of other alpha-lactalbumins. One possible explanation for the 17 amino acid extension is a mutation at the termination codon.
Notes: Prasad, R V
Butkowski, R J
Hamilton, J W
Ebner, K E
AM 18257/AM/NIADDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1982 Mar 30;21(7):1479-82. doi: 10.1021/bi00536a002.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1708
Author: Preston, R. C., Jakob, R. P., Binder, F. P., Sager, C. P., Ernst, B. and Maier, T.
Year: 2016
Title: E-selectin ligand complexes adopt an extended high-affinity conformation
Journal: J Mol Cell Biol
Volume: 8
Issue: 1
Pages: 62-72
Epub Date: 2015/06/29
Date: Feb
Short Title: E-selectin ligand complexes adopt an extended high-affinity conformation
Alternate Journal: Journal of molecular cell biology
ISSN: 1674-2788 (Print)
1759-4685
DOI: 10.1093/jmcb/mjv046
PMCID: PMC4710209
Accession Number: 26117840
Keywords: E-Selectin/*chemistry/*metabolism
Glycolipids/chemistry/pharmacology
Humans
Protein Binding
Protein Conformation
Protein Structure, Secondary
E-selectin
conformational change
glycomimetic antagonist
inflammation
leukocyte adhesion
sialyl Lewisx
Abstract: E-selectin is a cell-adhesion molecule of the vascular endothelium that promotes essential leukocyte rolling in the early inflammatory response by binding to glycoproteins containing the tetrasaccharide sialyl Lewis(x) (sLe(x)). Efficient leukocyte recruitment under vascular flow conditions depends on an increased lifetime of E-selectin/ligand complexes under tensile force in a so-called catch-bond binding mode. Co-crystal structures of a representative fragment of the extracellular E-selectin region with sLe(x) and a glycomimetic antagonist thereof reveal an extended E-selectin conformation, which is identified as a high-affinity binding state of E-selectin by molecular dynamics simulations. Small-angle X-ray scattering experiments demonstrate a direct link between ligand binding and E-selectin conformational transition under static conditions in solution. This permits tracing a series of concerted structural changes connecting ligand binding to conformational stretching as the structural basis of E-selectin catch-bond-mediated leukocyte recruitment. The detailed molecular view of the binding site paves the way for the design of a new generation of selectin antagonists. This is of special interest, since their therapeutic potential was recently demonstrated with the pan-selectin antagonists GMI-1070 (Rivipansel).
Notes: 1759-4685
Preston, Roland C
Jakob, Roman P
Binder, Florian P C
Sager, Christoph P
Ernst, Beat
Maier, Timm
Journal Article
Research Support, Non-U.S. Gov't
J Mol Cell Biol. 2016 Feb;8(1):62-72. doi: 10.1093/jmcb/mjv046. Epub 2015 Jun 27.
Author Address: Institute of Molecular Pharmacy, Universität Basel, 4056 Basel, Switzerland.
Biozentrum, Universität Basel, 4056 Basel, Switzerland.
Institute of Molecular Pharmacy, Universität Basel, 4056 Basel, Switzerland timm.maier@unibas.ch beat.ernst@unibas.ch.
Biozentrum, Universität Basel, 4056 Basel, Switzerland timm.maier@unibas.ch beat.ernst@unibas.ch.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 970
Author: Preusser-Kunze, A., Mariappan, M., Schmidt, B., Gande, S. L., Mutenda, K., Wenzel, D., von Figura, K. and Dierks, T.
Year: 2005
Title: Molecular characterization of the human Calpha-formylglycine-generating enzyme
Journal: J Biol Chem
Volume: 280
Issue: 15
Pages: 14900-10
Epub Date: 2005/01/20
Date: Apr 15
Short Title: Molecular characterization of the human Calpha-formylglycine-generating enzyme
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M413383200
Accession Number: 15657036
Keywords: Amino Acid Sequence
Binding Sites
Blotting, Western
Catalytic Domain
Cell Line, Tumor
Cross-Linking Reagents/pharmacology
Cysteine/chemistry
DNA, Complementary/metabolism
Dimerization
Disulfides/chemistry
Endoplasmic Reticulum/metabolism
Ethylmaleimide/pharmacology
Fibroblasts/metabolism
Fluorescent Antibody Technique, Indirect
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Microscopy, Immunoelectron
Molecular Sequence Data
Oxidation-Reduction
Oxidoreductases Acting on Sulfur Group Donors
Peptides/chemistry
Plasmids/metabolism
Polysaccharides/chemistry
Proline/chemistry
Protein Binding
Protein Processing, Post-Translational
Protein Structure, Tertiary
Recombinant Proteins/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Sulfatases/*chemistry
Thermolysin/chemistry
Time Factors
Trypsin/pharmacology
Two-Hybrid System Techniques
Abstract: Calpha-formylglycine (FGly) is the catalytic residue in the active site of sulfatases. In eukaryotes, it is generated in the endoplasmic reticulum by post-translational modification of a conserved cysteine residue. The FGly-generating enzyme (FGE), performing this modification, is an endoplasmic reticulum-resident enzyme that upon overexpression is secreted. Recombinant FGE was purified from cells and secretions to homogeneity. Intracellular FGE contains a high mannose type N-glycan, which is processed to the complex type in secreted FGE. Secreted FGE shows partial N-terminal trimming up to residue 73 without loosing catalytic activity. FGE is a calcium-binding protein containing an N-terminal (residues 86-168) and a C-terminal (residues 178-374) protease-resistant domain. The latter is stabilized by three disulfide bridges arranged in a clamp-like manner, which links the third to the eighth, the fourth to the seventh, and the fifth to the sixth cysteine residue. The innermost cysteine pair is partially reduced. The first two cysteine residues are located in the sequence preceding the N-terminal protease-resistant domain. They can form intramolecular or intermolecular disulfide bonds, the latter stabilizing homodimers. The C-terminal domain comprises the substrate binding site, as evidenced by yeast two-hybrid interaction assays and photocross-linking of a substrate peptide to proline 182. Peptides derived from all known human sulfatases served as substrates for purified FGE indicating that FGE is sufficient to modify all sulfatases of the same species.
Notes: Preusser-Kunze, Andrea
Mariappan, Malaiyalam
Schmidt, Bernhard
Gande, Santosh Lakshmi
Mutenda, Kudzai
Wenzel, Dirk
von Figura, Kurt
Dierks, Thomas
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2005 Apr 15;280(15):14900-10. doi: 10.1074/jbc.M413383200. Epub 2005 Jan 18.
Author Address: Institut für Biochemie und Molekulare Zellbiologie, Abteilung Biochemie II, Universität Göttingen, Heinrich-Düker-Weg 12, 37073 Göttingen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1696
Author: Pronker, M. F., Bos, T. G., Sharp, T. H., Thies-Weesie, D. M. and Janssen, B. J.
Year: 2015
Title: Olfactomedin-1 Has a V-shaped Disulfide-linked Tetrameric Structure
Journal: J Biol Chem
Volume: 290
Issue: 24
Pages: 15092-101
Epub Date: 2015/04/24
Date: Jun 12
Short Title: Olfactomedin-1 Has a V-shaped Disulfide-linked Tetrameric Structure
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M115.653485
PMCID: PMC4463452
Accession Number: 25903135
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Extracellular Matrix Proteins/*chemistry/metabolism
Glycoproteins/*chemistry/metabolism
HEK293 Cells
Humans
Mice
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Quaternary
Sequence Homology, Amino Acid
analytical ultracentrifugation
cell signaling
coiled coil
development
disulfide
electron tomography
neurobiology
olfactomedin-1 (Olfm1)
small angle x-ray scattering (SAXS)
x-ray crystallography
Abstract: Olfactomedin-1 (Olfm1; also known as noelin and pancortin) is a member of the olfactomedin domain-containing superfamily and a highly expressed neuronal glycoprotein important for nervous system development. It binds a number of secreted proteins and cell surface-bound receptors to induce cell signaling processes. Using a combined approach of x-ray crystallography, solution scattering, analytical ultracentrifugation, and electron microscopy we determined that full-length Olfm1 forms disulfide-linked tetramers with a distinctive V-shaped architecture. The base of the "V" is formed by two disulfide-linked dimeric N-terminal domains. Each of the two V legs consists of a parallel dimeric disulfide-linked coiled coil with a C-terminal β-propeller dimer at the tips. This agrees with our crystal structure of a C-terminal coiled-coil segment and β-propeller combination (Olfm1(coil-Olf)) that reveals a disulfide-linked dimeric arrangement with the β-propeller top faces in an outward exposed orientation. Similar to its family member myocilin, Olfm1 is stabilized by calcium. The dimer-of-dimers architecture suggests a role for Olfm1 in clustering receptors to regulate signaling and sheds light on the conformation of several other olfactomedin domain family members.
Notes: 1083-351x
Pronker, Matti F
Bos, Trusanne G A A
Sharp, Thomas H
Thies-Weesie, Dominique M E
Janssen, Bert J C
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2015 Jun 12;290(24):15092-101. doi: 10.1074/jbc.M115.653485. Epub 2015 Apr 21.
Author Address: From the Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research and.
Section Electron Microscopy, Department of Molecular Cell Biology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.
Van't Hoff Laboratory for Physical and Colloid Chemistry, Debye Institute of Nanomaterials Science, Department of Chemistry, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands and.
From the Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research and b.j.c.janssen@uu.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1819
Author: Pronker, M. F., Lemstra, S., Snijder, J., Heck, A. J., Thies-Weesie, D. M., Pasterkamp, R. J. and Janssen, B. J.
Year: 2016
Title: Structural basis of myelin-associated glycoprotein adhesion and signalling
Journal: Nat Commun
Volume: 7
Pages: 13584
Epub Date: 2016/12/07
Date: Dec 6
Short Title: Structural basis of myelin-associated glycoprotein adhesion and signalling
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms13584
PMCID: PMC5150538
Accession Number: 27922006
Keywords: Animals
Cell Adhesion
Crystallography, X-Ray
HEK293 Cells
Hippocampus/cytology
Humans
Mice
Models, Biological
Models, Molecular
Myelin-Associated Glycoprotein/*chemistry/*metabolism
N-Acetylneuraminic Acid/metabolism
Neurites/metabolism
Protein Domains
Protein Multimerization
Protein Processing, Post-Translational
Scattering, Small Angle
*Signal Transduction
Solutions
Structure-Activity Relationship
X-Ray Diffraction
Abstract: Myelin-associated glycoprotein (MAG) is a myelin-expressed cell-adhesion and bi-directional signalling molecule. MAG maintains the myelin-axon spacing by interacting with specific neuronal glycolipids (gangliosides), inhibits axon regeneration and controls myelin formation. The mechanisms underlying MAG adhesion and signalling are unresolved. We present crystal structures of the MAG full ectodomain, which reveal an extended conformation of five Ig domains and a homodimeric arrangement involving membrane-proximal domains Ig4 and Ig5. MAG-oligosaccharide complex structures and biophysical assays show how MAG engages axonal gangliosides at domain Ig1. Two post-translational modifications were identified-N-linked glycosylation at the dimerization interface and tryptophan C-mannosylation proximal to the ganglioside binding site-that appear to have regulatory functions. Structure-guided mutations and neurite outgrowth assays demonstrate MAG dimerization and carbohydrate recognition are essential for its regeneration-inhibiting properties. The combination of trans ganglioside binding and cis homodimerization explains how MAG maintains the myelin-axon spacing and provides a mechanism for MAG-mediated bi-directional signalling.
Notes: 2041-1723
Pronker, Matti F
Lemstra, Suzanne
Snijder, Joost
Heck, Albert J R
Orcid: 0000-0002-2405-4404
Thies-Weesie, Dominique M E
Pasterkamp, R Jeroen
Janssen, Bert J C
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2016 Dec 6;7:13584. doi: 10.1038/ncomms13584.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Department for Translational Neuroscience, Brain Center Rudolf Magnus, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands.
Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Department of Chemistry and Department of Pharmaceutical Sciences, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Van't Hoff Laboratory for Physical and Colloid Chemistry, Debye Institute of Nanomaterials Science, Department of Chemistry, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1889
Author: Pronker, M. F., Tas, R. P., Vlieg, H. C. and Janssen, B. J. C.
Year: 2017
Title: Nogo Receptor crystal structures with a native disulfide pattern suggest a novel mode of self-interaction
Journal: Acta Crystallogr D Struct Biol
Volume: 73
Issue: Pt 11
Pages: 860-876
Epub Date: 2017/11/03
Date: Nov 1
Short Title: Nogo Receptor crystal structures with a native disulfide pattern suggest a novel mode of self-interaction
Alternate Journal: Acta crystallographica. Section D, Structural biology
ISSN: 2059-7983
DOI: 10.1107/s2059798317013791
Accession Number: 29095159
Keywords: Amino Acid Sequence
Animals
Catalytic Domain
Crystallization
*Crystallography, X-Ray
Disulfides/*chemistry/metabolism
Mice
Models, Molecular
Nogo Receptors/*chemistry/metabolism
Protein Conformation
Protein Multimerization
Sequence Homology
Nogo Receptor
X-ray crystallography
analytical ultracentrifugation
myelin-associated inhibitors
neuronal plasticity
small-angle X-ray scattering
Abstract: The Nogo Receptor (NgR) is a glycophosphatidylinositol-anchored cell-surface protein and is a receptor for three myelin-associated inhibitors of regeneration: myelin-associated glycoprotein, Nogo66 and oligodendrocyte myelin glycoprotein. In combination with different co-receptors, NgR mediates signalling that reduces neuronal plasticity. The available structures of the NgR ligand-binding leucine-rich repeat (LRR) domain have an artificial disulfide pattern owing to truncated C-terminal construct boundaries. NgR has previously been shown to self-associate via its LRR domain, but the structural basis of this interaction remains elusive. Here, crystal structures of the NgR LRR with a longer C-terminal segment and a native disulfide pattern are presented. An additional C-terminal loop proximal to the C-terminal LRR cap is stabilized by two newly formed disulfide bonds, but is otherwise mostly unstructured in the absence of any stabilizing interactions. NgR crystallized in six unique crystal forms, three of which share a crystal-packing interface. NgR crystal-packing interfaces from all eight unique crystal forms are compared in order to explore how NgR could self-interact on the neuronal plasma membrane.
Notes: 2059-7983
Pronker, Matti F
Tas, Roderick P
Vlieg, Hedwich C
Janssen, Bert J C
723.012.002/Nederlandse Organisatie voor Wetenschappelijk Onderzoek/
Journal Article
United States
Acta Crystallogr D Struct Biol. 2017 Nov 1;73(Pt 11):860-876. doi: 10.1107/S2059798317013791. Epub 2017 Oct 19.
Author Address: Crystal and Structural Chemistry, Bijvoet Center for Biomolecular Research, Department of Chemistry, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 989
Author: Prorok-Hamon, M., Notel, F., Mathieu, S., Langlet, C., Fukuda, M. and El-Battari, A.
Year: 2005
Title: N-glycans of core2 beta(1,6)-N-acetylglucosaminyltransferase-I (C2GnT-I) but not those of alpha(1,3)-fucosyltransferase-VII (FucT-VII) are required for the synthesis of functional P-selectin glycoprotein ligand-1 (PSGL-1): effects on P-, L- and E-selectin binding
Journal: Biochem J
Volume: 391
Issue: Pt 3
Pages: 491-502
Epub Date: 2005/06/02
Date: Nov 1
Short Title: N-glycans of core2 beta(1,6)-N-acetylglucosaminyltransferase-I (C2GnT-I) but not those of alpha(1,3)-fucosyltransferase-VII (FucT-VII) are required for the synthesis of functional P-selectin glycoprotein ligand-1 (PSGL-1): effects on P-, L- and E-selectin binding
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20050344
PMCID: PMC1276950
Accession Number: 15926890
Keywords: Animals
Cell Line
Cricetinae
E-Selectin/*metabolism
Fucosyltransferases/chemistry/genetics/metabolism
Gene Expression Regulation, Enzymologic
Glycosylation
Humans
L-Selectin/*metabolism
Membrane Glycoproteins/*biosynthesis
Mutation
N-Acetylglucosaminyltransferases/*chemistry/genetics/*metabolism
P-Selectin/*metabolism
Polysaccharides/*metabolism
Protein Binding
Abstract: C2GnT-I [core2 beta(1,6)-N-acetyglucosaminyltransferase-I] and FucT-VII [alpha(1,3)-fucosyltransferase-VII] are the key enzymes for the biosynthesis of sialyl-Lewis x determinants on selectin ligands and therefore they represent good drug targets for the treatment of inflammatory disorders and other pathologies involving selectins. In the present study, we examined the importance of N-glycosylation for the ability of C2GnT-I and FucT-VII to generate functional selectin ligands, particularly the PSGL-1 (P-selectin glycoprotein ligand-1). We found that (i) both enzymes have their two N-glycosylation sites occupied, (ii) for C2GnT-I, the N-glycan chain linked to Asn-95 significantly contributes to the synthesis of functional PSGL-1 and is required to localize the enzyme to the cis/medial-Golgi compartment, (iii) all N-glycosylation-deficient proteins of FucT-VII displayr a dramatic impairment of their in vitro enzymatic activities, but retain their ability to fucosylate the core2-modified PSGL-I and to generate P- and L-selectin binding, and (iv) the glycomutants of FucT-VII fail to synthesize sialyl-Lewis x or to generate E-selectin binding unless core2-modified PSGL-1 is present. All combined, our results show a differential functional impact of N-glycosylation on C2GnT-1 and FucT-VII and disclose that a strongly reduced FucT-VII activity retains the ability to fucosylate PSGL-1 on the core2-based binding site(s) for the three selectins.
Notes: 1470-8728
Prorok-Hamon, Maëlle
Notel, Frédéric
Mathieu, Sylvie
Langlet, Claire
Fukuda, Minoru
El-Battari, Assou
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2005 Nov 1;391(Pt 3):491-502. doi: 10.1042/BJ20050344.
Author Address: INSERM U-559/IFR-125 Université de la Méditerranée, 27 Bd. Jean Moulin, 13385 Marseille Cedex 05, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 842
Author: Prota, A. E., Sage, D. R., Stehle, T. and Fingeroth, J. D.
Year: 2002
Title: The crystal structure of human CD21: Implications for Epstein-Barr virus and C3d binding
Journal: Proc Natl Acad Sci U S A
Volume: 99
Issue: 16
Pages: 10641-6
Epub Date: 2002/07/18
Date: Aug 6
Short Title: The crystal structure of human CD21: Implications for Epstein-Barr virus and C3d binding
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.162360499
PMCID: PMC124999
Accession Number: 12122212
Keywords: Carbohydrate Sequence
Complement C3d/*chemistry/immunology
Crystallography, X-Ray
Herpesvirus 4, Human/*chemistry/immunology
Humans
Models, Molecular
Molecular Sequence Data
Receptors, Complement 3d/*chemistry/genetics/immunology
Abstract: Human complement receptor type 2 (CD21) is the cellular receptor for Epstein-Barr virus (EBV), a human tumor virus. The N-terminal two short consensus repeats (SCR1-SCR2) of the receptor interact with the EBV glycoprotein gp350/220 and also with the natural CD21 ligand C3d. Here we present the crystal structure of the CD21 SCR1-SCR2 fragment in the absence of ligand and demonstrate that it is able to bind EBV. Based on a functional analysis of wild-type and mutant CD21 and molecular modeling, we identify a likely region for EBV attachment and demonstrate that this region is not involved in the interaction with C3d. A comparison with the previously determined structure of CD21 SCR1-SCR2 in complex with C3d shows that, in both cases, CD21 assumes compact V-shaped conformations. However, our analysis reveals a surprising degree of flexibility at the SCR1-SCR2 interface, suggesting interactions between the two domains are not specific. We present evidence that the V-shaped conformation is induced by deglycosylation of the protein, and that physiologic glycosylation of CD21 would result in a more extended conformation, perhaps with additional epitopes for C3d binding.
Notes: 1091-6490
Prota, Andrea E
Sage, David R
Stehle, Thilo
Fingeroth, Joyce D
R01 AI045716/AI/NIAID NIH HHS/United States
AI45716/AI/NIAID NIH HHS/United States
DE12186/DE/NIDCR NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2002 Aug 6;99(16):10641-6. doi: 10.1073/pnas.162360499. Epub 2002 Jul 16.
Author Address: Harvard Medical School, Division of Experimental Medicine and Infectious Diseases, Beth Israel Deaconess Medical Center, Harvard Institutes of Medicine, 4 Blackfan Circle, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1200
Author: Protty, M. B., Watkins, N. A., Colombo, D., Thomas, S. G., Heath, V. L., Herbert, J. M., Bicknell, R., Senis, Y. A., Ashman, L. K., Berditchevski, F., Ouwehand, W. H., Watson, S. P. and Tomlinson, M. G.
Year: 2009
Title: Identification of Tspan9 as a novel platelet tetraspanin and the collagen receptor GPVI as a component of tetraspanin microdomains
Journal: Biochem J
Volume: 417
Issue: 1
Pages: 391-400
Epub Date: 2008/09/18
Date: Jan 1
Short Title: Identification of Tspan9 as a novel platelet tetraspanin and the collagen receptor GPVI as a component of tetraspanin microdomains
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20081126
PMCID: PMC2652832
Accession Number: 18795891
Keywords: Animals
Blood Platelets/*metabolism
Blotting, Western
Cell Line
Cell Line, Tumor
Cells, Cultured
Humans
Megakaryocytes/cytology/*metabolism
Membrane Microdomains/metabolism
Membrane Proteins/genetics/*metabolism
Mice
Microscopy, Confocal
Oligonucleotide Array Sequence Analysis
Platelet Membrane Glycoproteins/genetics/*metabolism
RNA, Messenger/genetics/metabolism
Tetraspanins
Abstract: Platelets are essential for wound healing and inflammatory processes, but can also play a deleterious role by causing heart attack and stroke. Normal platelet activation is dependent on tetraspanins, a superfamily of glycoproteins that function as 'organisers' of cell membranes by recruiting other receptors and signalling proteins into tetraspanin-enriched microdomains. However, our understanding of how tetraspanin microdomains regulate platelets is hindered by the fact that only four of the 33 mammalian tetraspanins have been identified in platelets. This is because of a lack of antibodies to most tetraspanins and difficulties in measuring mRNA, due to low levels in this anucleate cell. To identify potentially platelet-expressed tetraspanins, mRNA was measured in their nucleated progenitor cell, the megakaryocyte, using serial analysis of gene expression and DNA microarrays. Amongst 19 tetraspanins identified in megakaryocytes, Tspan9, a previously uncharacterized tetraspanin, was relatively specific to these cells. Through generating the first Tspan9 antibodies, Tspan9 expression was found to be tightly regulated in platelets. The relative levels of CD9, CD151, Tspan9 and CD63 were 100, 14, 6 and 2 respectively. Since CD9 was expressed at 49000 cell surface copies per platelet, this suggested a copy number of 2800 Tspan9 molecules. Finally, Tspan9 was shown to be a component of tetraspanin microdomains that included the collagen receptor GPVI (glycoprotein VI) and integrin alpha6beta1, but not the von Willebrand receptor GPIbalpha or the integrins alphaIIbbeta3 or alpha2beta1. These findings suggest a role for Tspan9 in regulating platelet function in concert with other platelet tetraspanins and their associated proteins.
Notes: 1470-8728
Protty, Majd B
Watkins, Nicholas A
Colombo, Dario
Thomas, Steven G
Heath, Victoria L
Herbert, John M J
Bicknell, Roy
Senis, Yotis A
Ashman, Leonie K
Berditchevski, Fedor
Ouwehand, Willem H
Watson, Steve P
Tomlinson, Michael G
FS/08/034/25085/British Heart Foundation/United Kingdom
G0400247/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2009 Jan 1;417(1):391-400. doi: 10.1042/BJ20081126.
Author Address: Centre for Cardiovascular Sciences, Institute of Biomedical Research, The Medical School, University of Birmingham, Birmingham B152TT, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 220
Author: Purkayastha, S., Rao, C. V. and Lamm, M. E.
Year: 1979
Title: Structure of the carbohydrate chain of free secretory component from human milk
Journal: J Biol Chem
Volume: 254
Issue: 14
Pages: 6583-7
Epub Date: 1979/07/25
Date: Jul 25
Short Title: Structure of the carbohydrate chain of free secretory component from human milk
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 447737
Keywords: Amino Acids/analysis
Asparagine
Aspartic Acid/analysis
Carbohydrates/analysis
Female
Glycopeptides/isolation & purification
Humans
*Immunoglobulin Fragments/isolation & purification
Methylation
Milk, Human/*immunology
Pregnancy
Pronase
*Secretory Component/isolation & purification
Abstract: Secretory component from human milk was found to contain 23.4% carbohydrate, which includes galactose, mannose, fucose, glucosamine, and sialic acid. Secretory component could be degraded by pronase or base-borohydride to yield the same, single type of carbohydrate chain. In the glycopeptide produced by pronase digestion, aspartic acid was the only amino acid present in molar quantities after amino acid analysis, which suggests that the carbohydrate moiety is linked to the polypeptide chain at asparagine residues. The positions of links between the various sugar units were studied by methylation analyses of: secretory component, periodate-oxidized and reduced secretory component, the fragment produced by base-borohydride treatment, and the pronase glycopeptide after treatment with specific glycosidases. Sugars released from the glycopeptide by various glycosidases were also quantitated. From the results of these studies a branched chain structure was assigned to the carbohydrate chain of secretory component.
Notes: Purkayastha, S
Rao, C V
Lamm, M E
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1979 Jul 25;254(14):6583-7.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 446
Author: Putnam, F. W., Florent, G., Paul, C., Shinoda, T. and Shimizu, A.
Year: 1973
Title: Complete amino acid sequence of the Mu heavy chain of a human IgM immunoglobulin
Journal: Science
Volume: 182
Issue: 4109
Pages: 287-91
Epub Date: 1973/10/19
Date: Oct 19
Short Title: Complete amino acid sequence of the Mu heavy chain of a human IgM immunoglobulin
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.182.4109.287
Accession Number: 4742735
Keywords: Amino Acid Sequence
Autoanalysis
Humans
Hydrolysis
Immunoglobulin Fragments/*analysis
Immunoglobulin M/*analysis
Trypsin
Abstract: The amino acid sequence of the micro, chain of a human IgM immunoglobulin, including the location of all disulfide bridges and oligosaccharides, has been determined. The homology of the constant regions of immunoglobulin micro, gamma, alpha, and epsilon heavy chains reveals evolutionary relationships and suggests that two genes code for each heavy chain.
Notes: Putnam, F W
Florent, G
Paul, C
Shinoda, T
Shimizu, A
Journal Article
United States
Science. 1973 Oct 19;182(4109):287-91. doi: 10.1126/science.182.4109.287.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1988
Author: Pyo, K. E., Kim, C. R., Lee, M., Kim, J. S., Kim, K. I. and Baek, S. H.
Year: 2018
Title: ULK1 O-GlcNAcylation Is Crucial for Activating VPS34 via ATG14L during Autophagy Initiation
Journal: Cell Rep
Volume: 25
Issue: 10
Pages: 2878-2890.e4
Epub Date: 2018/12/06
Date: Dec 4
Short Title: ULK1 O-GlcNAcylation Is Crucial for Activating VPS34 via ATG14L during Autophagy Initiation
Alternate Journal: Cell reports
DOI: 10.1016/j.celrep.2018.11.042
Accession Number: 30517873
Keywords: AMP-Activated Protein Kinases/metabolism
Adaptor Proteins, Vesicular Transport/*metabolism
Amino Acid Sequence
Autophagosomes/metabolism
*Autophagy
Autophagy-Related Protein-1 Homolog/*metabolism
Autophagy-Related Proteins/*metabolism
Cell Line
Class III Phosphatidylinositol 3-Kinases/*metabolism
Glucosamine/*metabolism
Glucose/deficiency
Glycosylation
Humans
Intracellular Signaling Peptides and Proteins/*metabolism
N-Acetylglucosaminyltransferases/metabolism
Phosphorylation
Threonine/metabolism
*Ampk
*O-GlcNACylation
*Pp1
*Ulk1
*autophagy initiation
*mTORC1
*phagophore formation
Abstract: Unc-51-like-kinase 1 (ULK1) is a target of both the mechanistic target of rapamycin (mTOR) and AMP-activated protein kinase (AMPK), whose role is to facilitate the initiation of autophagy in response to starvation. Upon glucose starvation, dissociation of mTOR from ULK1 and phosphorylation by AMPK leads to the activation of ULK1 activity. Here, we provide evidence that ULK1 is the attachment of O-linked N-acetylglucosamine (O-GlcNAcylated) on the threonine 754 site by O-linked N-acetylglucosamine transferase (OGT) upon glucose starvation. ULK1 O-GlcNAcylation occurs after dephosphorylation of adjacent mTOR-dependent phosphorylation on the serine 757 site by protein phosphatase 1 (PP1) and phosphorylation by AMPK. ULK1 O-GlcNAcylation is crucial for binding and phosphorylation of ATG14L, allowing the activation of lipid kinase VPS34 and leading to the production of phosphatidylinositol-(3)-phosphate (PI(3)P), which is required for phagophore formation and initiation of autophagy. Our findings provide insights into the crosstalk between dephosphorylation and O-GlcNAcylation during autophagy and specify a molecular framework for potential therapeutic intervention in autophagy-related diseases.
Notes: 2211-1247
Pyo, Ki Eun
Kim, Chang Rok
Lee, Minkyoung
Kim, Jong-Seo
Kim, Keun Il
Baek, Sung Hee
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell Rep. 2018 Dec 4;25(10):2878-2890.e4. doi: 10.1016/j.celrep.2018.11.042.
Author Address: Creative Research Initiatives Center for Epigenetic Code and Diseases, Department of Biological Sciences, Seoul National University, Seoul 08826, South Korea.
Center for RNA Research, Institute for Basic Science, Department of Biological Sciences, Seoul National University, Seoul 08826, South Korea.
Department of Biological Sciences, Sookmyung Women's University, Seoul 04310, South Korea. Electronic address: kikim@sookmyung.ac.kr.
Creative Research Initiatives Center for Epigenetic Code and Diseases, Department of Biological Sciences, Seoul National University, Seoul 08826, South Korea. Electronic address: sbaek@snu.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1493
Author: Qian, Y., Zhang, X., Zhou, L., Yun, X., Xie, J., Xu, J., Ruan, Y. and Ren, S.
Year: 2012
Title: Site-specific N-glycosylation identification of recombinant human lectin-like oxidized low density lipoprotein receptor-1 (LOX-1)
Journal: Glycoconj J
Volume: 29
Issue: 5-6
Pages: 399-409
Epub Date: 2012/06/13
Date: Aug
Short Title: Site-specific N-glycosylation identification of recombinant human lectin-like oxidized low density lipoprotein receptor-1 (LOX-1)
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080
DOI: 10.1007/s10719-012-9408-z
Accession Number: 22688517
Keywords: Carbohydrate Sequence
Cell Line, Tumor
Gene Expression
Glycosylation
Humans
Molecular Sequence Data
Pronase/chemistry
Recombinant Proteins/chemistry/genetics/metabolism
Scavenger Receptors, Class E/*chemistry/genetics/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tandem Mass Spectrometry
Abstract: Human LOX-1/OLR 1 plays a key role in atherogenesis and endothelial dysfunction. The N-glycosylation of LOX-1 has been shown to affect its biological functions in vivo and modulate the pathogenesis of atherosclerosis. However, the N-glycosylation pattern of LOX-1 has not been described yet. The present study was aimed at elucidating the N-glycosylation of recombinant human LOX-1 with regard to N-glycan profile and N-glycosylation sites. Here, an approach using nonspecific protease (Pronase E) digestion followed by MALDI-QIT-TOF MS and multistage MS (MS(3)) analysis is explored to obtain site-specific N-glycosylation information of recombinant human LOX-1, in combination with glycan structure confirmation through characterizing released glycans using tandem MS. The results reveal that N-glycans structures as well as their corresponding attached site of LOX-1 can be identified simultaneously by direct MS analysis of glycopeptides from non-specific protease digestion. With this approach, one potential glycosylation site of recombinant human LOX-1 on Asn(139) is readily identified and found to carry heterogeneous complex type N-glycans. In addition, manual annotation of multistage MS data utilizing diagnostic ions, which were found to be particularly useful in defining the structure of glycopeptides and glycans was addressed for proper spectra interpretation. The findings described herein will shed new light on further research of the structure-function relationships of LOX-1 N-glycan.
Notes: 1573-4986
Qian, Yifan
Zhang, Xingwang
Zhou, Lei
Yun, Xiaojing
Xie, Jianhui
Xu, Jiejie
Ruan, Yuanyuan
Ren, Shifang
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 2012 Aug;29(5-6):399-409. doi: 10.1007/s10719-012-9408-z. Epub 2012 Jun 12.
Author Address: Key Laboratory of Glycoconjugate Research Ministry of Public Health, Shanghai Medical College, Fudan University, Shanghai, People's Republic of China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1754
Author: Qiao, Y., Zhang, X., Zhang, Y., Wang, Y., Xu, Y., Liu, X., Sun, F. and Wang, J.
Year: 2016
Title: High Glucose Stimulates Tumorigenesis in Hepatocellular Carcinoma Cells Through AGER-Dependent O-GlcNAcylation of c-Jun
Journal: Diabetes
Volume: 65
Issue: 3
Pages: 619-32
Epub Date: 2016/01/31
Date: Mar
Short Title: High Glucose Stimulates Tumorigenesis in Hepatocellular Carcinoma Cells Through AGER-Dependent O-GlcNAcylation of c-Jun
Alternate Journal: Diabetes
ISSN: 0012-1797
DOI: 10.2337/db15-1057
Accession Number: 26825459
Keywords: Acylation
Animals
Blood Glucose/*metabolism
Blotting, Western
Carcinogenesis/*metabolism
Carcinoma, Hepatocellular/*metabolism
Cell Line, Tumor
Cell Proliferation
Diabetes Mellitus, Experimental/*metabolism
Fluorescent Antibody Technique
Gas Chromatography-Mass Spectrometry
Hep G2 Cells
Humans
Immunoblotting
Immunohistochemistry
Immunoprecipitation
Liver Neoplasms/*metabolism
Mice
Mice, Inbred BALB C
N-Acetylglucosaminyltransferases/*metabolism
Proto-Oncogene Proteins c-jun/*metabolism
Receptor for Advanced Glycation End Products/*metabolism
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
Abstract: Epidemiologic studies suggest that hepatocellular carcinoma (HCC) has a strong relationship with diabetes. However, the underlying molecular mechanisms still remain unclear. Here, we demonstrated that high glucose (HG), one of the main characteristics of diabetes, was capable of accelerating tumorigenesis in HCC cells. Advanced glycosylation end product-specific receptor (AGER) was identified as a stimulator during this process. Mechanistically, AGER activated a hexosamine biosynthetic pathway, leading to enhanced O-GlcNAcylation of target proteins. Notably, AGER was capable of increasing activity and stability of proto-oncoprotein c-Jun via O-GlcNAcylation of this protein at Ser73. Interestingly, c-Jun can conversely enhance AGER transcription. Thereby, a positive autoregulatory feedback loop that stimulates diabetic HCC was established. Finally, we found that AG490, an inhibitor of Janus kinase, has the ability to impair AGER expression and its functions in HCC cells. In conclusion, AGER and its functions to stimulate O-GlcNAcylation are important during liver tumorigenesis, when high blood glucose levels are inadequately controlled.
Notes: 1939-327x
Qiao, Yongxia
Zhang, Xiao
Zhang, Yue
Wang, Yulan
Xu, Yanfeng
Liu, Xiangfan
Sun, Fenyong
Wang, Jiayi
Journal Article
Research Support, Non-U.S. Gov't
United States
Diabetes. 2016 Mar;65(3):619-32. doi: 10.2337/db15-1057. Epub 2016 Jan 29.
Author Address: School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai, China.
Department of Central Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai, China.
Department of Pharmacy, Shanghai Municipal Hospital of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, China.
Faculty of Medical Laboratory Science, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai, China sunfenyong@126.com karajan2@163.com.
Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai, China Tongji University Advanced Institute of Translational Medicine, Shanghai, China sunfenyong@126.com karajan2@163.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1953
Author: Qin, K., Zhu, Y., Qin, W., Gao, J., Shao, X., Wang, Y. L., Zhou, W., Wang, C. and Chen, X.
Year: 2018
Title: Quantitative Profiling of Protein O-GlcNAcylation Sites by an Isotope-Tagged Cleavable Linker
Journal: ACS Chem Biol
Volume: 13
Issue: 8
Pages: 1983-1989
Epub Date: 2018/07/31
Date: Aug 17
Short Title: Quantitative Profiling of Protein O-GlcNAcylation Sites by an Isotope-Tagged Cleavable Linker
Alternate Journal: ACS chemical biology
ISSN: 1554-8929
DOI: 10.1021/acschembio.8b00414
Accession Number: 30059200
Keywords: Acetylglucosamine/*chemistry
Animals
Biotin/analogs & derivatives
Carbon Isotopes
Female
Glycoproteins/*analysis/chemistry
Glycosylation
HeLa Cells
Humans
Isotope Labeling/*methods
Male
Mice
Molecular Probes/*chemistry
Nitrogen Isotopes
Proteome/*analysis/chemistry
Proteomics/*methods
Triazoles/chemistry
Abstract: Large-scale quantification of protein O-linked β- N-acetylglucosamine (O-GlcNAc) modification in a site-specific manner remains a key challenge in studying O-GlcNAc biology. Herein, we developed an isotope-tagged cleavable linker (isoTCL) strategy, which enabled isotopic labeling of O-GlcNAc through bioorthogonal conjugation of affinity tags. We demonstrated the application of the isoTCL in mapping and quantification of O-GlcNAcylation sites in HeLa cells. Furthermore, we investigated the O-GlcNAcylation sensitivity to the sugar donor by quantifying the levels of modification under different concentrations of the O-GlcNAc labeling probe in a site-specific manner. In addition, we applied isoTCL to compare the O-GlcNAcylation stoichiometry levels of more than 100 modification sites between placenta samples from male and female mice and confirmed site-specifically that female placenta has a higher O-GlcNAcylation than its male counterpart. The isoTCL platform provides a powerful tool for quantitative profiling of O-GlcNAc modification.
Notes: 1554-8937
Qin, Ke
Zhu, Yuntao
Qin, Wei
Gao, Jinjun
Orcid: 0000-0002-7390-5578
Shao, Xuan
Wang, Yan-Ling
Zhou, Wen
Wang, Chu
Orcid: 0000-0002-6925-1268
Chen, Xing
Orcid: 0000-0002-3058-7370
Journal Article
Research Support, Non-U.S. Gov't
United States
ACS Chem Biol. 2018 Aug 17;13(8):1983-1989. doi: 10.1021/acschembio.8b00414. Epub 2018 Jul 30.
Author Address: State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology , Chinese Academy of Sciences , Beijing , 100101 , China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1876
Author: Qin, W., Lv, P., Fan, X., Quan, B., Zhu, Y., Qin, K., Chen, Y., Wang, C. and Chen, X.
Year: 2017
Title: Quantitative time-resolved chemoproteomics reveals that stable O-GlcNAc regulates box C/D snoRNP biogenesis
Journal: Proc Natl Acad Sci U S A
Volume: 114
Issue: 33
Pages: E6749-e6758
Epub Date: 2017/08/02
Date: Aug 15
Short Title: Quantitative time-resolved chemoproteomics reveals that stable O-GlcNAc regulates box C/D snoRNP biogenesis
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1702688114
PMCID: PMC5565422
Accession Number: 28760965
Keywords: Acetylglucosamine/chemistry/*metabolism
Animals
Antibiotics, Antineoplastic/pharmacology
Cell Survival/drug effects
Chromosomal Proteins, Non-Histone/genetics/metabolism
Doxorubicin/pharmacology
HeLa Cells
Humans
Isotope Labeling/methods
MCF-7 Cells
Male
Methylation
Mice
Mice, Inbred BALB C
Mice, Nude
NIH 3T3 Cells
Neoplasms/drug therapy/genetics/metabolism
Protein Processing, Post-Translational
Proteome/genetics/*metabolism
Proteomics/*methods
RNA, Ribosomal/genetics/metabolism
Ribonucleoproteins, Small Nucleolar/genetics/*metabolism
Xenograft Model Antitumor Assays
*O-GlcNAcylation
*metabolic labeling
*protein stability
*proteomics
*snoRNP
Abstract: O-linked GlcNAcylation (O-GlcNAcylation), a ubiquitous posttranslational modification on intracellular proteins, is dynamically regulated in cells. To analyze the turnover dynamics of O-GlcNAcylated proteins, we developed a quantitative time-resolved O-linked GlcNAc proteomics (qTOP) strategy based on metabolic pulse-chase labeling with an O-GlcNAc chemical reporter and stable isotope labeling with amino acids in cell culture (SILAC). Applying qTOP, we quantified the turnover rates of 533 O-GlcNAcylated proteins in NIH 3T3 cells and discovered that about 14% exhibited minimal removal of O-GlcNAc or degradation of protein backbones. The stability of those hyperstable O-GlcNAcylated proteins was more sensitive to O-GlcNAcylation inhibition compared with the more dynamic populations. Among the hyperstable population were three core proteins of box C/D small nucleolar ribonucleoprotein complexes (snoRNPs): fibrillarin (FBL), nucleolar protein 5A (NOP56), and nucleolar protein 5 (NOP58). We showed that O-GlcNAcylation stabilized these proteins and was essential for snoRNP assembly. Blocking O-GlcNAcylation on FBL altered the 2'-O-methylation of rRNAs and impaired cancer cell proliferation and tumor formation in vivo.
Notes: 1091-6490
Qin, Wei
Lv, Pinou
Fan, Xinqi
Quan, Baiyi
Zhu, Yuntao
Qin, Ke
Chen, Ying
Wang, Chu
Chen, Xing
Orcid: 0000-0002-3058-7370
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2017 Aug 15;114(33):E6749-E6758. doi: 10.1073/pnas.1702688114. Epub 2017 Jul 31.
Author Address: College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Peking-Tsinghua Center for Life Sciences, Peking University, Beijing 100871, China.
College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China; chuwang@pku.edu.cn xingchen@pku.edu.cn.
Beijing National Laboratory for Molecular Sciences, Peking University, Beijing 100871, China.
Synthetic and Functional Biomolecules Center, Peking University, Beijing 100871, China.
Key Laboratory of Bioorganic Chemistry and Molecular Engineering of the Ministry of Education, Peking University, Beijing 100871, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1902
Author: Qin, W., Qin, K., Fan, X., Peng, L., Hong, W., Zhu, Y., Lv, P., Du, Y., Huang, R., Han, M., Cheng, B., Liu, Y., Zhou, W., Wang, C. and Chen, X.
Year: 2018
Title: Artificial Cysteine S-Glycosylation Induced by Per-O-Acetylated Unnatural Monosaccharides during Metabolic Glycan Labeling
Journal: Angew Chem Int Ed Engl
Volume: 57
Issue: 7
Pages: 1817-1820
Epub Date: 2017/12/14
Date: Feb 12
Short Title: Artificial Cysteine S-Glycosylation Induced by Per-O-Acetylated Unnatural Monosaccharides during Metabolic Glycan Labeling
Alternate Journal: Angewandte Chemie (International ed. in English)
ISSN: 1433-7851
DOI: 10.1002/anie.201711710
Accession Number: 29237092
Keywords: Acetylation
Alkynes/chemistry
Azides/chemistry
Biotin/chemistry/metabolism
Cysteine/*chemistry/metabolism
Glutathione/chemistry
Glycosylation
HeLa Cells
Humans
Metabolic Engineering
Monosaccharides/*chemistry/metabolism
Peptides/analysis
Tandem Mass Spectrometry
*O-GlcNAc
*S-glycosylation
*chemoproteomics
*labeling
*unnatural monosaccharides
Abstract: The unexpected, non-enzymatic S-glycosylation of cysteine residues in various proteins by per-O-acetylated monosaccharides is described. This artificial S-glycosylation greatly compromises the specificity and validity of metabolic glycan labeling in living cells by per-O-acetylated azido and alkynyl sugars, which has been overlooked in the field for decades. It is demonstrated that the use of unacetylated unnatural sugars can avoid the artifact formation and a corrected list of O-GlcNAcylated proteins and O-GlcNAc sites in HeLa cells has been assembled by using N-azidoacetylgalactosamine (GalNAz).
Notes: 1521-3773
Qin, Wei
Qin, Ke
Fan, Xinqi
Peng, Linghang
Hong, Weiyao
Zhu, Yuntao
Lv, Pinou
Du, Yifei
Huang, Rongbing
Han, Mengting
Cheng, Bo
Liu, Yuan
Zhou, Wen
Wang, Chu
Chen, Xing
Orcid: 0000-0002-3058-7370
Journal Article
Research Support, Non-U.S. Gov't
Germany
Angew Chem Int Ed Engl. 2018 Feb 12;57(7):1817-1820. doi: 10.1002/anie.201711710. Epub 2018 Jan 5.
Author Address: College of Chemistry and Molecular Engineering, Peking-Tsinghua Center for Life Science, Beijing National Laboratory for Molecular Sciences, Synthetic and Functional Biomolecules Center, and Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University, Beijing, 100871, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1216
Author: Quistgaard, E. M., Madsen, P., Grøftehauge, M. K., Nissen, P., Petersen, C. M. and Thirup, S. S.
Year: 2009
Title: Ligands bind to Sortilin in the tunnel of a ten-bladed beta-propeller domain
Journal: Nat Struct Mol Biol
Volume: 16
Issue: 1
Pages: 96-8
Epub Date: 2009/01/06
Date: Jan
Short Title: Ligands bind to Sortilin in the tunnel of a ten-bladed beta-propeller domain
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.1543
Accession Number: 19122660
Keywords: Adaptor Proteins, Vesicular Transport/*chemistry/*metabolism
Animals
Apoptosis
Binding Sites
Ligands
Mammals
Models, Molecular
Neurons/physiology
Neurotensin/chemistry/*metabolism
Protein Binding
Protein Conformation
Abstract: The structure of the Sortilin ectodomain in complex with neurotensin has been determined at 2-A resolution, revealing that the C-terminal part of neurotensin binds in the tunnel of a ten-bladed beta-propeller domain. Binding competition studies suggest that additional binding sites, for example, for the prodomain of nerve growth factor-beta, are present in the tunnel and that competition for binding relates to the restricted space inside the propeller.
Notes: 1545-9985
Quistgaard, Esben M
Madsen, Peder
Grøftehauge, Morten K
Nissen, Poul
Petersen, Claus M
Thirup, Søren S
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2009 Jan;16(1):96-8. doi: 10.1038/nsmb.1543. Epub 2009 Jan 4.
Author Address: MIND Centre, Department of Molecular Biology, University of Aarhus, Gustav Wieds Vej 10C, DK 8000 Arhus C, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1091
Author: Ragolia, L., Hall, C. E. and Palaia, T.
Year: 2007
Title: Post-translational modification regulates prostaglandin D2 synthase apoptotic activity: characterization by site-directed mutagenesis
Journal: Prostaglandins Other Lipid Mediat
Volume: 83
Issue: 1-2
Pages: 25-32
Epub Date: 2007/01/30
Date: Feb
Short Title: Post-translational modification regulates prostaglandin D2 synthase apoptotic activity: characterization by site-directed mutagenesis
Alternate Journal: Prostaglandins & other lipid mediators
ISSN: 1098-8823 (Print)
1098-8823
DOI: 10.1016/j.prostaglandins.2006.09.006
PMCID: PMC1805777
NIHMSID: NIHMS18356
Accession Number: 17259069
Keywords: Amino Acid Sequence
Animals
*Apoptosis/drug effects
Cell Proliferation/drug effects
Electrophoresis, Gel, Two-Dimensional
Glucose/pharmacology
Humans
Hyperglycemia/enzymology
Intramolecular Oxidoreductases/chemistry/*metabolism/pharmacology
Isoenzymes/chemistry/metabolism/pharmacology
Lipocalins
Male
Molecular Sequence Data
Muscle, Smooth, Vascular/cytology/drug effects/enzymology
*Mutagenesis, Site-Directed
Mutation/genetics
*Protein Processing, Post-Translational/drug effects
Rats
Rats, Wistar
Abstract: Lipocalin-type prostaglandin D(2) synthase (L-PGDS) is a highly glycosylated protein found in several body fluids. Elevated L-PGDS levels have been observed in the serum of patients with renal impairment, diabetes mellitus, and hypertension. Recently, we demonstrated the ability of L-PGDS to induce apoptosis in a variety of cell types including epithelial cells, neuronal cells, and vascular smooth muscle cells (VSMCs). The aim of this study was to investigate the effect several site-directed mutations had on L-PGDS-induced apoptosis in order to identify potential sites of regulation. Point mutations created in a glycosylation site (Asn51), a protein kinase C phosphorylation site (Ser106), and the enzymatic active site (Cys65) all inhibited L-PGDS-induced apoptosis as determined by both terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL) and caspase3 activity. We also compared the L-PGDS isoforms present in GK rat serum to WKY control serum using two-dimensional gel electrophoresis and observed distinct differences which vanished after PNGase F glycolytic digestion. We conclude that post-translational modification of L-PGDS, by either glycosylation or phosphorylation, enhances its apoptotic activity and inhibits VSMC hyperproliferation and postulate that this process is altered in type 2 diabetes.
Notes: Ragolia, Louis
Hall, Christopher E
Palaia, Thomas
R01 HL067953/HL/NHLBI NIH HHS/United States
R01HL067953-01A2/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Prostaglandins Other Lipid Mediat. 2007 Feb;83(1-2):25-32. doi: 10.1016/j.prostaglandins.2006.09.006. Epub 2006 Nov 7.
Author Address: Vascular Biology Laboratory, Winthrop-University Hospital, Mineola, NY 11501, USA. LRagolia@winthrop.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1048
Author: Ramachandran, P., Boontheung, P., Xie, Y., Sondej, M., Wong, D. T. and Loo, J. A.
Year: 2006
Title: Identification of N-linked glycoproteins in human saliva by glycoprotein capture and mass spectrometry
Journal: J Proteome Res
Volume: 5
Issue: 6
Pages: 1493-503
Epub Date: 2006/06/03
Date: Jun
Short Title: Identification of N-linked glycoproteins in human saliva by glycoprotein capture and mass spectrometry
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr050492k
Accession Number: 16740002
Keywords: Amino Acid Sequence
Electrophoresis, Gel, Two-Dimensional
Glycoproteins/*analysis
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
Proteomics
Saliva/*chemistry
Abstract: Glycoproteins make up a major and important part of the salivary proteome and play a vital role in maintaining the health of the oral cavity. Because changes in the physiological state of a person are reflected as changes in the glycoproteome composition, mapping the salivary glycoproteome will provide insights into various processes in the body. Salivary glycoproteins were identified by the hydrazide coupling and release method. In this approach, glycoproteins were coupled onto a hydrazide resin, the proteins were then digested and formerly N-glycosylated peptides were selectively released with the enzyme PNGase F and analyzed by LC-MS/MS. Employing this method, coupled with in-solution isoelectric focusing separation as an additional means for pre-fractionation, we identified 84 formerly N-glycosylated peptides from 45 unique N-glycoproteins. Of these, 16 glycoproteins have not been reported previously in saliva. In addition, we identified 44 new sites of N-linked glycosylation on the proteins.
Notes: Ramachandran, Prasanna
Boontheung, Pinmannee
Xie, Yongming
Sondej, Melissa
Wong, David T
Loo, Joseph A
GM07185/GM/NIGMS NIH HHS/United States
U01 DE016275/DE/NIDCR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
J Proteome Res. 2006 Jun;5(6):1493-503. doi: 10.1021/pr050492k.
Author Address: Department of Chemistry and Biochemistry, University of California-Los Angeles, Los Angeles, California, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1579
Author: Ramakrishnan, P., Clark, P. M., Mason, D. E., Peters, E. C., Hsieh-Wilson, L. C. and Baltimore, D.
Year: 2013
Title: Activation of the transcriptional function of the NF-κB protein c-Rel by O-GlcNAc glycosylation
Journal: Sci Signal
Volume: 6
Issue: 290
Pages: ra75
Epub Date: 2013/08/29
Date: Aug 27
Short Title: Activation of the transcriptional function of the NF-κB protein c-Rel by O-GlcNAc glycosylation
Alternate Journal: Science signaling
ISSN: 1945-0877
DOI: 10.1126/scisignal.2004097
PMCID: PMC4066889
NIHMSID: NIHMS578080
Accession Number: 23982206
Keywords: Acetylglucosamine/*metabolism
Acylation
Animals
Binding Sites/genetics
Blotting, Western
Cell Line, Tumor
Cells, Cultured
Glycosylation
Granulocyte-Macrophage Colony-Stimulating Factor/genetics/metabolism
HEK293 Cells
Humans
Interferon-gamma/genetics/metabolism
Interleukin-2/genetics/metabolism
Jurkat Cells
Mice
Mice, Inbred C57BL
Mice, Knockout
Mutation
NF-kappa B/genetics/*metabolism
Protein Binding
Proto-Oncogene Proteins c-rel/genetics/*metabolism
RNA Interference
Reverse Transcriptase Polymerase Chain Reaction
Serine/genetics/metabolism
Signal Transduction
Abstract: The transcription factor nuclear factor κB (NF-κB) rapidly reprograms gene expression in response to various stimuli, and its activity is regulated by several posttranslational modifications, including phosphorylation, methylation, and acetylation. The addition of O-linked β-N-acetylglucosamine (a process known as O-GlcNAcylation) is an abundant posttranslational modification that is enhanced in conditions such as hyperglycemia and cellular stress. We report that the NF-κB subunit c-Rel is modified and activated by O-GlcNAcylation. We identified serine 350 as the site of O-GlcNAcylation, which was required for the DNA binding and transactivation functions of c-Rel. Blocking the O-GlcNAcylation of this residue abrogated c-Rel-mediated expression of the cytokine-encoding genes IL2, IFNG, and CSF2 in response to T cell receptor (TCR) activation, whereas increasing the extent of O-GlcNAcylation of cellular proteins enhanced the expression of these genes. TCR- or tumor necrosis factor (TNF)-induced expression of other NF-κB target genes, such as NFKBIA (which encodes IκBα) and TNFAIP3 (which encodes A20), occurred independently of the O-GlcNAcylation of c-Rel. Our findings suggest a stimulus-specific role for hyperglycemia-induced O-GlcNAcylation of c-Rel in promoting T cell-mediated autoimmunity in conditions such as type 1 diabetes by enhancing the production of T helper cell cytokines.
Notes: 1937-9145
Ramakrishnan, Parameswaran
Clark, Peter M
Mason, Daniel E
Peters, Eric C
Hsieh-Wilson, Linda C
Baltimore, David
R01 GM039458/GM/NIGMS NIH HHS/United States
R01 GM084724/GM/NIGMS NIH HHS/United States
2R01 GM039458/GM/NIGMS NIH HHS/United States
2R01 GM084724/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Sci Signal. 2013 Aug 27;6(290):ra75. doi: 10.1126/scisignal.2004097.
Author Address: Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1456
Author: Rambaruth, N. D., Greenwell, P. and Dwek, M. V.
Year: 2012
Title: The lectin Helix pomatia agglutinin recognizes O-GlcNAc containing glycoproteins in human breast cancer
Journal: Glycobiology
Volume: 22
Issue: 6
Pages: 839-48
Epub Date: 2012/02/11
Date: Jun
Short Title: The lectin Helix pomatia agglutinin recognizes O-GlcNAc containing glycoproteins in human breast cancer
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cws051
Accession Number: 22322011
Keywords: Acetylglucosamine/analysis/antagonists & inhibitors/*metabolism
Antibodies, Monoclonal/pharmacology
Breast Neoplasms/*metabolism/pathology
Female
Glycoproteins/analysis/*metabolism
Humans
Lectins/analysis/*metabolism
Tumor Cells, Cultured
Abstract: There has been considerable interest in understanding the epitopes that bind the lectin Helix pomatia agglutinin (HPA) in breast cancer as the lectin has been shown to identify glycosylation changes associated with the development of metastatic disease. HPA has previously been shown to recognize aberrant O-linked α-N-acetylgalactosamine (GalNAcα)/mucin glycosylation in cancer, including exposed Tn epitopes. However, recent glycan-array analysis reported that diverse epitopes are also recognized by the lectin, e.g. consortium for functional glycomics (CFG) data: GalNAcα1,3Gal; β-GalNAc; GlcNAcβ1,4Gal. The intriguing observations from the CFG array led to this study, in which HPA-binding epitopes were localized and characterized in an in vitro model of breast cancer metastasis. HMT3522 (benign disease), BT474 (primary cancer) and T47D/MCF7 (metastatic cancer) cells were assessed in confocal microscopy-based co-localization studies and a glycoproteomic analysis based on 2-dimensional electrophoresis (2DE), western blotting and mass spectrometry was adopted. HPA binding correlated with levels of integrin α6, transcription factors heterogeneous nuclear ribonuclear protein (HnRNP) H1, HnRNP D-like, HnRNP A2/B1 as well as heat shock protein 27 (Hsp27), glial fibrillary acidic protein and enolase 1 (ENO1). These glycoproteins were non-detectable in the non-metastatic breast cancer cell lines. The recognition of HnRNPs, Hsp27 and ENO1 by HPA correlated with O-GlcNAcylation of these proteins. Integrin α6 was the most abundant HPA glycoprotein in the breast cancer cells with a metastatic phenotype; this concurred with previous findings in colorectal cancer. This is the first report in which HPA has been shown to bind O-GlcNAcylated transcription factors. This class of proteins represents a new means by which HPA differentiates cancer cells with an aggressive metastatic phenotype.
Notes: 1460-2423
Rambaruth, Neela D S
Greenwell, Pamela
Dwek, Miriam V
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2012 Jun;22(6):839-48. doi: 10.1093/glycob/cws051. Epub 2012 Feb 9.
Author Address: Department of Molecular and Applied Biosciences, University of Westminster, London W1W 6UW, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2063
Author: Ramirez, D. H., Aonbangkhen, C., Wu, H. Y., Naftaly, J. A., Tang, S., O'Meara, T. R. and Woo, C. M.
Year: 2020
Title: Engineering a Proximity-Directed O-GlcNAc Transferase for Selective Protein O-GlcNAcylation in Cells
Journal: ACS Chem Biol
Volume: 15
Issue: 4
Pages: 1059-1066
Epub Date: 2020/03/03
Date: Apr 17
Short Title: Engineering a Proximity-Directed O-GlcNAc Transferase for Selective Protein O-GlcNAcylation in Cells
Alternate Journal: ACS chemical biology
ISSN: 1554-8929 (Print)
1554-8929
DOI: 10.1021/acschembio.0c00074
PMCID: PMC7296736
NIHMSID: NIHMS1588277
Accession Number: 32119511
Keywords: Glycosylation
HEK293 Cells
Humans
N-Acetylglucosaminyltransferases/genetics/*metabolism
Protein Engineering
Protein Processing, Post-Translational
Proteome/chemistry/metabolism
Recombinant Fusion Proteins/genetics/*metabolism
Single-Domain Antibodies/genetics/*metabolism
alpha-Synuclein/chemistry/*metabolism
Abstract: O-Linked β-N-acetylglucosamine (O-GlcNAc) is a monosaccharide that plays an essential role in cellular signaling throughout the nucleocytoplasmic proteome of eukaryotic cells. Strategies for selectively increasing O-GlcNAc levels on a target protein in cells would accelerate studies of this essential modification. Here, we report a generalizable strategy for introducing O-GlcNAc into selected target proteins in cells using a nanobody as a proximity-directing agent fused to O-GlcNAc transferase (OGT). Fusion of a nanobody that recognizes GFP (nGFP) or a nanobody that recognizes the four-amino acid sequence EPEA (nEPEA) to OGT yielded nanobody-OGT constructs that selectively delivered O-GlcNAc to a series of tagged target proteins (e.g., JunB, cJun, and Nup62). Truncation of the tetratricopeptide repeat domain as in OGT(4) increased selectivity for the target protein through the nanobody by reducing global elevation of O-GlcNAc levels in the cell. Quantitative chemical proteomics confirmed the increase in O-GlcNAc to the target protein by nanobody-OGT(4). Glycoproteomics revealed that nanobody-OGT(4) or full-length OGT produced a similar glycosite profile on the target protein JunB and Nup62. Finally, we demonstrate the ability to selectively target endogenous α-synuclein for O-GlcNAcylation in HEK293T cells. These first proximity-directed OGT constructs provide a flexible strategy for targeting additional proteins and a template for further engineering of OGT and the O-GlcNAc proteome in the future. The use of a nanobody to redirect OGT substrate selection for glycosylation of desired proteins in cells may further constitute a generalizable strategy for controlling a broader array of post-translational modifications in cells.
Notes: 1554-8937
Ramirez, Daniel H
Orcid: 0000-0003-3288-9951
Aonbangkhen, Chanat
Orcid: 0000-0002-7378-1341
Wu, Hung-Yi
Naftaly, Jeffrey A
Tang, Stephanie
Orcid: 0000-0002-2705-2201
O'Meara, Timothy R
Woo, Christina M
Orcid: 0000-0001-8687-9105
U01 CA242098/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
ACS Chem Biol. 2020 Apr 17;15(4):1059-1066. doi: 10.1021/acschembio.0c00074. Epub 2020 Mar 2.
Author Address: Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts 02138, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1742
Author: Ramkumar, N., Harvey, B. M., Lee, J. D., Alcorn, H. L., Silva-Gagliardi, N. F., McGlade, C. J., Bestor, T. H., Wijnholds, J., Haltiwanger, R. S. and Anderson, K. V.
Year: 2015
Title: Protein O-Glucosyltransferase 1 (POGLUT1) Promotes Mouse Gastrulation through Modification of the Apical Polarity Protein CRUMBS2
Journal: PLoS Genet
Volume: 11
Issue: 10
Pages: e1005551
Epub Date: 2015/10/27
Date: Oct
Short Title: Protein O-Glucosyltransferase 1 (POGLUT1) Promotes Mouse Gastrulation through Modification of the Apical Polarity Protein CRUMBS2
Alternate Journal: PLoS genetics
ISSN: 1553-7390 (Print)
1553-7390
DOI: 10.1371/journal.pgen.1005551
PMCID: PMC4619674
Accession Number: 26496195
Keywords: Animals
Embryo, Mammalian
Embryonic Development
Eye Proteins/*genetics/metabolism
Gastrulation/genetics
Glucosyltransferases/*genetics/metabolism
Glycosylation
Humans
Membrane Proteins/*genetics/metabolism
Mice
Nerve Tissue Proteins/*genetics/metabolism
Phenotype
Protein Processing, Post-Translational/genetics
Receptor, Notch1/genetics/*metabolism
Signal Transduction
Abstract: Crumbs family proteins are apical transmembrane proteins with ancient roles in cell polarity. Mouse Crumbs2 mutants arrest at midgestation with abnormal neural plate morphology and a deficit of mesoderm caused by defects in gastrulation. We identified an ENU-induced mutation, wsnp, that phenocopies the Crumbs2 null phenotype. We show that wsnp is a null allele of Protein O-glucosyltransferase 1 (Poglut1), which encodes an enzyme previously shown to add O-glucose to EGF repeats in the extracellular domain of Drosophila and mammalian Notch, but the role of POGLUT1 in mammalian gastrulation has not been investigated. As predicted, we find that POGLUT1 is essential for Notch signaling in the early mouse embryo. However, the loss of mouse POGLUT1 causes an earlier and more dramatic phenotype than does the loss of activity of the Notch pathway, indicating that POGLUT1 has additional biologically relevant substrates. Using mass spectrometry, we show that POGLUT1 modifies EGF repeats in the extracellular domain of full-length mouse CRUMBS2. CRUMBS2 that lacks the O-glucose modification fails to be enriched on the apical plasma membrane and instead accumulates in the endoplasmic reticulum. The data demonstrate that CRUMBS2 is the target of POGLUT1 for the gastrulation epithelial-to-mesenchymal transitions (EMT) and that all activity of CRUMBS2 depends on modification by POGLUT1. Mutations in human POGLUT1 cause Dowling-Degos Disease, POGLUT1 is overexpressed in a variety of tumor cells, and mutations in the EGF repeats of human CRUMBS proteins are associated with human congenital nephrosis, retinitis pigmentosa and retinal degeneration, suggesting that O-glucosylation of CRUMBS proteins has broad roles in human health.
Notes: 1553-7404
Ramkumar, Nitya
Harvey, Beth M
Lee, Jeffrey D
Alcorn, Heather L
Silva-Gagliardi, Nancy F
McGlade, C Jane
Bestor, Timothy H
Wijnholds, Jan
Haltiwanger, Robert S
Anderson, Kathryn V
T32 GM008468/GM/NIGMS NIH HHS/United States
P30 CA008748/CA/NCI NIH HHS/United States
R37 HD035455/HD/NICHD NIH HHS/United States
R37 HD03455/HD/NICHD NIH HHS/United States
R01 GM061126/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
PLoS Genet. 2015 Oct 23;11(10):e1005551. doi: 10.1371/journal.pgen.1005551. eCollection 2015 Oct.
Author Address: Developmental Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, New York, United States of America; Program in Biochemistry and Structural Biology, Cell and Developmental Biology, and Molecular Biology, Weill Cornell Graduate School of Medical Sciences, Cornell University, New York, New York, United States of America.
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, New York, United States of America.
Developmental Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, New York, United States of America.
The Hospital for Sick Children, Arthur and Sonia Labatt Brain Tumor Research Center and Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada.
Department of Genetics and Development, College of Physicians and Surgeons of Columbia University, New York, New York, United States of America.
Department of Neuromedical Genetics, Netherlands Institute for Neuroscience, Amsterdam, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1025
Author: Ramsland, P. A., Terzyan, S. S., Cloud, G., Bourne, C. R., Farrugia, W., Tribbick, G., Geysen, H. M., Moomaw, C. R., Slaughter, C. A. and Edmundson, A. B.
Year: 2006
Title: Crystal structure of a glycosylated Fab from an IgM cryoglobulin with properties of a natural proteolytic antibody
Journal: Biochem J
Volume: 395
Issue: 3
Pages: 473-81
Epub Date: 2006/01/21
Date: May 1
Short Title: Crystal structure of a glycosylated Fab from an IgM cryoglobulin with properties of a natural proteolytic antibody
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20051739
PMCID: PMC1462693
Accession Number: 16422668
Keywords: Amino Acid Sequence
Binding Sites
Catalysis
Cold Temperature
Cryoglobulins/*chemistry/*metabolism
Crystallography, X-Ray
Gels/chemistry
Glycosylation
Humans
Immunoglobulin Fab Fragments/*chemistry/*metabolism
Immunoglobulin M/*chemistry/*metabolism
Lysine/chemistry/metabolism
Models, Molecular
Molecular Sequence Data
Protein Structure, Tertiary
Sequence Alignment
Structural Homology, Protein
Water/chemistry
Abstract: The 2.6 A (1 A=0.1 nm) resolution structure has been determined for the glycosylated Fab (fragment antigen binding) of an IgM (Yvo) obtained from a subject with Waldenström's macroglobulinaemia. Dynamic light scattering was used to estimate the gel point and monitor the formation of an ordered hydroscopic gel of Yvo IgM upon cooling. If a cryoglobulin forms gels in peripheral tissues and organs, the associated swelling and damage to microvasculature can result in considerable morbidity and mortality. The three-dimensional structure of the branched N-linked oligosaccharide associated with the CH1 domain (first constant domain of heavy chain) is reported. The carbohydrate may act to shield part of the lateral surface of the CH1 domain and crowd the junction between the CH1 and CH2 domains, thereby limiting the segmental flexibility of the Fab arms in intact Yvo IgM, especially at low temperatures. Recently, Yvo IgM was shown to have the properties of a naturally occurring proteolytic antibody [Paul, Karle, Planque, Taguchi, Salas, Nishiyama, Handy, Hunter, Edmundson and Hanson (2004) J. Biol. Chem. 279, 39611-39619; Planque, Bangale, Song, Karle, Taguchi, Poindexter, Bick, Edmundson, Nishiyama and Paul (2004) J. Biol Chem. 279, 14024-14032]. The Yvo protein displayed the ability to cleave, by a nucleophilic mechanism, the amide bonds of a variety of serine protease substrates and the gp120 coat protein of HIV. An atypical serine, arginine and glutamate motif is located in the middle of the Yvo antigen-binding site and displays an overall geometry that mimics the classical serine, histidine and aspartate catalytic triad of serine proteases. Our present findings indicate that pre-existing or natural antibodies can utilize at least one novel strategy for the cleavage of peptide bonds.
Notes: 1470-8728
Ramsland, Paul A
Terzyan, Simon S
Cloud, Gwendolyn
Bourne, Christina R
Farrugia, William
Tribbick, Gordon
Geysen, H Mario
Moomaw, Carolyn R
Slaughter, Clive A
Edmundson, Allen B
CA72803/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochem J. 2006 May 1;395(3):473-81. doi: 10.1042/BJ20051739.
Author Address: Structural Immunology Laboratory, Austin Research Institute, Kronheimer Building, Austin Health, Studley Road, Heidelberg, VIC 3084, Australia. p.ramsland@ari.unimelb.edu.au
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1412
Author: Rana, N. A., Nita-Lazar, A., Takeuchi, H., Kakuda, S., Luther, K. B. and Haltiwanger, R. S.
Year: 2011
Title: O-glucose trisaccharide is present at high but variable stoichiometry at multiple sites on mouse Notch1
Journal: J Biol Chem
Volume: 286
Issue: 36
Pages: 31623-37
Epub Date: 2011/07/16
Date: Sep 9
Short Title: O-glucose trisaccharide is present at high but variable stoichiometry at multiple sites on mouse Notch1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.268243
PMCID: PMC3173066
Accession Number: 21757702
Keywords: Amino Acid Motifs
Amino Acid Sequence
Animals
*Consensus Sequence
Glucose/*analysis
Glycosylation
Mice
Receptor, Notch1/*chemistry/physiology
Trisaccharides/*analysis
Abstract: Notch activity is regulated by both O-fucosylation and O-glucosylation, and Notch receptors contain multiple predicted sites for both. Here we examine the occupancy of the predicted O-glucose sites on mouse Notch1 (mN1) using the consensus sequence C(1)XSXPC(2). We show that all of the predicted sites are modified, although the efficiency of modifying O-glucose sites is site- and cell type-dependent. For instance, although most sites are modified at high stoichiometries, the site at EGF 27 is only partially glucosylated, and the occupancy of the site at EGF 4 varies with cell type. O-Glucose is also found at a novel, non-traditional consensus site at EGF 9. Based on this finding, we propose a revision of the consensus sequence for O-glucosylation to allow alanine N-terminal to cysteine 2: C(1)XSX(A/P)C(2). We also show through biochemical and mass spectral analyses that serine is the only hydroxyamino acid that is modified with O-glucose on EGF repeats. The O-glucose at all sites is efficiently elongated to the trisaccharide Xyl-Xyl-Glc. To establish the functional importance of individual O-glucose sites in mN1, we used a cell-based signaling assay. Elimination of most individual sites shows little or no effect on mN1 activation, suggesting that the major effects of O-glucose are mediated by modification of multiple sites. Interestingly, elimination of the site in EGF 28, found in the Abruptex region of Notch, does significantly reduce activity. These results demonstrate that, like O-fucose, the O-glucose modifications of EGF repeats occur extensively on mN1, and they play important roles in Notch function.
Notes: 1083-351x
Rana, Nadia A
Nita-Lazar, Aleksandra
Takeuchi, Hideyuki
Kakuda, Shinako
Luther, Kelvin B
Haltiwanger, Robert S
R01 GM061126/GM/NIGMS NIH HHS/United States
R01 GM061126-12/GM/NIGMS NIH HHS/United States
T32 CA009176/CA/NCI NIH HHS/United States
GM61126/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2011 Sep 9;286(36):31623-37. doi: 10.1074/jbc.M111.268243. Epub 2011 Jul 8.
Author Address: Department of Biochemistry and Cell Biology, Institute of Cell and Developmental Biology, Stony Brook University, Stony Brook, New York 11794-5215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1483
Author: Ranuncolo, S. M., Ghosh, S., Hanover, J. A., Hart, G. W. and Lewis, B. A.
Year: 2012
Title: Evidence of the involvement of O-GlcNAc-modified human RNA polymerase II CTD in transcription in vitro and in vivo
Journal: J Biol Chem
Volume: 287
Issue: 28
Pages: 23549-61
Epub Date: 2012/05/19
Date: Jul 6
Short Title: Evidence of the involvement of O-GlcNAc-modified human RNA polymerase II CTD in transcription in vitro and in vivo
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.330910
PMCID: PMC3390630
Accession Number: 22605332
Keywords: Acetylglucosamine/analogs & derivatives/*metabolism/pharmacology
Acetylglucosaminidase/antagonists & inhibitors/metabolism
Binding Sites
Blotting, Western
Cell Line, Tumor
Chromatin Immunoprecipitation
Enzyme Inhibitors/pharmacology
Gene Expression Regulation/drug effects
HeLa Cells
Humans
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/metabolism
Oximes/pharmacology
Phenylcarbamates/pharmacology
Promoter Regions, Genetic/genetics
Protein Binding
*Protein Processing, Post-Translational
Protein Subunits/metabolism
RNA Interference
RNA Polymerase II/*metabolism
Reverse Transcriptase Polymerase Chain Reaction
Serine/metabolism
Transcription, Genetic/drug effects/*genetics
Abstract: The RNA polymerase II C-terminal domain (CTD), which serves as a scaffold to recruit machinery involved in transcription, is modified post-translationally. Although the O-GlcNAc modification of RNA polymerase II CTD was documented in 1993, its functional significance remained obscure. We show that O-GlcNAc transferase (OGT) modified CTD serine residues 5 and 7. Drug inhibition of OGT and OGA (N-acetylglucosaminidase) blocked transcription during preinitiation complex assembly. Polymerase II and OGT co-immunoprecipitated, and OGT is a component of the preinitiation complex. OGT shRNA experiments showed that reduction of OGT causes a reduction in transcription and RNA polymerase II occupancy at several B-cell promoters. These data suggest that the cycling of O-GlcNAc on and off of polymerase II occurs during assembly of the preinitiation complex. Our results define unexpected roles for both the CTD and O-GlcNAc in the regulation of transcription initiation in higher eukaryotes.
Notes: 1083-351x
Ranuncolo, Stella M
Ghosh, Salil
Hanover, John A
Hart, Gerald W
Lewis, Brian A
P30 DK079637/DK/NIDDK NIH HHS/United States
R01-DK61671/DK/NIDDK NIH HHS/United States
Intramural NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
NIH R01-CA42486/CA/NCI NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Jul 6;287(28):23549-61. doi: 10.1074/jbc.M111.330910. Epub 2012 May 17.
Author Address: Metabolism Branch, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1735
Author: Rao, X., Duan, X., Mao, W., Li, X., Li, Z., Li, Q., Zheng, Z., Xu, H., Chen, M., Wang, P. G., Wang, Y., Shen, B. and Yi, W.
Year: 2015
Title: O-GlcNAcylation of G6PD promotes the pentose phosphate pathway and tumor growth
Journal: Nat Commun
Volume: 6
Pages: 8468
Epub Date: 2015/09/25
Date: Sep 24
Short Title: O-GlcNAcylation of G6PD promotes the pentose phosphate pathway and tumor growth
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms9468
PMCID: PMC4598839
Accession Number: 26399441
Keywords: Acetylglucosamine/*metabolism
Animals
Cell Line, Tumor
Cell Proliferation
Glucose/*metabolism
Glucosephosphate Dehydrogenase/*metabolism
Glycosylation
HEK293 Cells
Hep G2 Cells
Humans
Hypoxia/*metabolism
In Vitro Techniques
Lung Neoplasms/*metabolism
MCF-7 Cells
Mice
N-Acetylglucosaminyltransferases/*metabolism
Neoplasm Transplantation
*Pentose Phosphate Pathway
Reactive Oxygen Species/*metabolism
Up-Regulation
Abstract: The pentose phosphate pathway (PPP) plays a critical role in macromolecule biosynthesis and maintaining cellular redox homoeostasis in rapidly proliferating cells. Upregulation of the PPP has been shown in several types of cancer. However, how the PPP is regulated to confer a selective growth advantage on cancer cells is not well understood. Here we show that glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the PPP, is dynamically modified with an O-linked β-N-acetylglucosamine sugar in response to hypoxia. Glycosylation activates G6PD activity and increases glucose flux through the PPP, thereby providing precursors for nucleotide and lipid biosynthesis, and reducing equivalents for antioxidant defense. Blocking glycosylation of G6PD reduces cancer cell proliferation in vitro and impairs tumor growth in vivo. Importantly, G6PD glycosylation is increased in human lung cancers. Our findings reveal a mechanistic understanding of how O-glycosylation directly regulates the PPP to confer a selective growth advantage to tumours.
Notes: 2041-1723
Rao, Xiongjian
Duan, Xiaotao
Mao, Weimin
Li, Xuexia
Li, Zhonghua
Li, Qian
Zheng, Zhiguo
Xu, Haimiao
Chen, Min
Wang, Peng G
Wang, Yingjie
Shen, Binghui
Yi, Wen
R01 CA085344/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2015 Sep 24;6:8468. doi: 10.1038/ncomms9468.
Author Address: Institute of Biochemistry, College of Life Sciences, Zhejiang University, Hangzhou 310058, China.
Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou 310003, China.
State Key Laboratory of Toxicology and Medical Countermeasures, Beijing Institute of Pharmacology and Toxicology, Beijing 100850, China.
Zhejiang Cancer Hospital, Zhejiang Cancer Research Institute, Hangzhou 310022, China.
School of Life Science and the State Key Laboratory of Microbial Technology, National Glycoengineering Research Center, Shandong University, Shandong 250100, China.
Department of Chemistry, Georgia State University, Atlanta, Georgia 30303, USA.
State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital of Zhejiang University, Hangzhou 310003, China.
Department of Radiation Biology, City of Hope National Medical Center, Duarte, California 91010, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1973
Author: Rasila, T., Saavalainen, O., Attalla, H., Lankila, P., Haglund, C., Hölttä, E. and Andersson, L. C.
Year: 2019
Title: Astroprincin (FAM171A1, C10orf38): A Regulator of Human Cell Shape and Invasive Growth
Journal: Am J Pathol
Volume: 189
Issue: 1
Pages: 177-189
Epub Date: 2018/10/13
Date: Jan
Short Title: Astroprincin (FAM171A1, C10orf38): A Regulator of Human Cell Shape and Invasive Growth
Alternate Journal: The American journal of pathology
ISSN: 0002-9440
DOI: 10.1016/j.ajpath.2018.09.006
Accession Number: 30312582
Keywords: 3T3 Cells
ADAM10 Protein/genetics/metabolism
Animals
COS Cells
Carrier Proteins/genetics/metabolism
*Cell Shape
Chlorocebus aethiops
Gene Knockdown Techniques
HEK293 Cells
Humans
MCF-7 Cells
Membrane Proteins/genetics/*metabolism
Mice
Neoplasm Invasiveness/genetics/pathology
Neoplasm Proteins/genetics/*metabolism
Nerve Tissue Proteins/genetics/metabolism
Proteins
Rabbits
Stress Fibers/genetics/metabolism/pathology
Two-Hybrid System Techniques
Abstract: Our group originally found and cloned cDNA for a 98-kDa type 1 transmembrane glycoprotein of unknown function. Because of its abundant expression in astrocytes, it was called the protein astroprincin (APCN). Two thirds of the evolutionarily conserved protein is intracytoplasmic, whereas the extracellular domain carries two N-glycosidic side chains. APCN is physiologically expressed in placental trophoblasts, skeletal and hearth muscle, and kidney and pancreas. Overexpression of APCN (cDNA) in various cell lines induced sprouting of slender projections, whereas knockdown of APCN expression by siRNA caused disappearance of actin stress fibers. Immunohistochemical staining of human cancers for endogenous APCN showed elevated expression in invasive tumor cells compared with intratumoral cells. Human melanoma cells (SK-MEL-28) transfected with APCN cDNA acquired the ability of invasive growth in semisolid medium (Matrigel) not seen with control cells. A conserved carboxyterminal stretch of 21 amino acids was found to be essential for APCN to induce cell sprouting and invasive growth. Yeast two-hybrid screening revealed several interactive partners, of which ornithine decarboxylase antizyme-1, NEEP21 (NSG1), and ADAM10 were validated by coimmunoprecipitation. This is the first functional description of APCN. These data show that APCN regulates the dynamics of the actin cytoskeletal and, thereby, the cell shape and invasive growth potential of tumor cells.
Notes: 1525-2191
Rasila, Tiina
Saavalainen, Olga
Attalla, Hesham
Lankila, Petri
Haglund, Caj
Hölttä, Erkki
Andersson, Leif C
Journal Article
Research Support, Non-U.S. Gov't
United States
Am J Pathol. 2019 Jan;189(1):177-189. doi: 10.1016/j.ajpath.2018.09.006. Epub 2018 Oct 10.
Author Address: Department of Pathology, University of Helsinki, Helsinki, Finland.
Research Programs Unit, Translational Cancer Biology, University of Helsinki, Helsinki, Finland; HUSLAB, Helsinki University Hospital, Helsinki, Finland.
Department of Pathology, University of Helsinki, Helsinki, Finland. Electronic address: leif.andersson@helsinki.fi.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 859
Author: Rasmussen, H. B., Branner, S., Wiberg, F. C. and Wagtmann, N.
Year: 2003
Title: Crystal structure of human dipeptidyl peptidase IV/CD26 in complex with a substrate analog
Journal: Nat Struct Biol
Volume: 10
Issue: 1
Pages: 19-25
Epub Date: 2002/12/17
Date: Jan
Short Title: Crystal structure of human dipeptidyl peptidase IV/CD26 in complex with a substrate analog
Alternate Journal: Nature structural biology
ISSN: 1072-8368 (Print)
1072-8368
DOI: 10.1038/nsb882
Accession Number: 12483204
Keywords: Binding Sites
Catalysis
Catalytic Domain
Crystallography, X-Ray/methods
Dipeptidyl Peptidase 4/*chemistry/*metabolism
Humans
Models, Molecular
Protein Structure, Tertiary
Substrate Specificity
Abstract: Dipeptidyl peptidase IV (DPP-IV/CD26) is a multifunctional type II transmembrane serine peptidase. This enzyme contributes to the regulation of various physiological processes, including blood sugar homeostasis, by cleaving peptide hormones, chemokines and neuropeptides. We have determined the 2.5 A structure of the extracellular region of DPP-IV in complex with the inhibitor valine-pyrrolidide. The catalytic site is located in a large cavity formed between the alpha/beta-hydrolase domain and an eight-bladed beta-propeller domain. Both domains participate in inhibitor binding. The structure indicates how substrate specificity is achieved and reveals a new and unexpected opening to the active site.
Notes: Rasmussen, Hanne B
Branner, Sven
Wiberg, Finn C
Wagtmann, Nicolai
Journal Article
United States
Nat Struct Biol. 2003 Jan;10(1):19-25. doi: 10.1038/nsb882.
Author Address: Protein Chemistry, Research and Development, Novo Nordisk A/S, Novo Allé, DK-2880 Bagsvaerd, Denmark. Hbrm@novonordisk.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 280
Author: Rassouli, M., Sambasivam, H., Azadi, P., Dell, A., Morris, H. R., Nagpurkar, A., Mookerjea, S. and Murray, R. K.
Year: 1992
Title: Derivation of the amino acid sequence of rat C-reactive protein from cDNA cloning with additional studies on the nature of its dimeric component
Journal: J Biol Chem
Volume: 267
Issue: 5
Pages: 2947-54
Epub Date: 1992/02/15
Date: Feb 15
Short Title: Derivation of the amino acid sequence of rat C-reactive protein from cDNA cloning with additional studies on the nature of its dimeric component
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1737750
Keywords: Amino Acid Sequence
Animals
Base Sequence
C-Reactive Protein/*genetics/isolation & purification/metabolism
Chromatography, High Pressure Liquid
Cloning, Molecular
DNA/*genetics
Electrophoresis, Polyacrylamide Gel
Gene Library
Humans
Kinetics
Macromolecular Substances
Male
Mass Spectrometry
Molecular Sequence Data
Molecular Weight
RNA, Messenger/genetics
Rats
Rats, Inbred Strains
Recombinant Proteins/isolation & purification/metabolism
Sequence Homology, Nucleic Acid
Abstract: Rat C-reactive protein (CRP) is unique among mammalian CRPs in being a glycoprotein and in containing a covalently linked dimer in its pentameric structure. To investigate these features, cDNA clones encoding rat CRP were isolated from an expression library, and the primary structure of the protein was derived. Taken along with the results of Northern blotting, we conclude that a single mRNA of approximately 2,500 nucleotides codes for a precursor of rat CRP with a signal sequence of 19 amino acids and a polypeptide of 211 amino acids, the latter sharing extensive homology with human, rabbit, and mouse CRPs. The deduced sequence agreed with results obtained from partial microsequencing and mapping by fast atom bombardment-mass spectrometry. Two potential sites for N-glycosylation (Asn-128 and Asn-147) and a C-terminal heptapeptide (Leu-205 to Ser-211, containing two cysteines at positions 208 and 209) were unique to rat CRP. The protein was also shown to be composed of five apparently identical monomers, two of which form a dimer linked by two interchain disulfide bonds involving Cys-208 and Cys-209. These same cysteines form an intrachain disulfide bond in the other three monomers. The primary structure of rat CRP and the basis of dimer formation have, therefore, been elucidated.
Notes: Rassouli, M
Sambasivam, H
Azadi, P
Dell, A
Morris, H R
Nagpurkar, A
Mookerjea, S
Murray, R K
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1992 Feb 15;267(5):2947-54.
Author Address: Department of Biochemistry, University of Toronto, Ontario, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 230
Author: Rauch, U., Karthikeyan, L., Maurel, P., Margolis, R. U. and Margolis, R. K.
Year: 1992
Title: Cloning and primary structure of neurocan, a developmentally regulated, aggregating chondroitin sulfate proteoglycan of brain
Journal: J Biol Chem
Volume: 267
Issue: 27
Pages: 19536-47
Epub Date: 1992/09/25
Date: Sep 25
Short Title: Cloning and primary structure of neurocan, a developmentally regulated, aggregating chondroitin sulfate proteoglycan of brain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1326557
Keywords: Aggrecans
Amino Acid Sequence
Animals
Base Sequence
Calcium-Binding Proteins/chemistry
Chondroitin Sulfate Proteoglycans/chemistry/*genetics
Chondroitin Sulfates/metabolism
Cloning, Molecular
DNA/genetics
Disulfides/chemistry
*Extracellular Matrix Proteins
Gelsolin
Gene Expression
Lectins, C-Type
Microfilament Proteins/chemistry
Molecular Sequence Data
Nerve Tissue Proteins/chemistry/*genetics
Oligodeoxyribonucleotides/chemistry
Polymerase Chain Reaction
Proteoglycans/chemistry
RNA, Messenger/genetics
Rats
Repetitive Sequences, Nucleic Acid
Restriction Mapping
Sequence Alignment
Tissue Distribution
Versicans
Abstract: We have obtained the complete coding sequence of neurocan, a chondroitin sulfate proteoglycan of rat brain which is developmentally regulated with respect to its molecular size, concentration, carbohydrate composition, sulfation, and immunocytochemical localization. Two degenerate oligonucleotides, based on amino acid sequence data from the proteoglycan isolated from adult brain by immunoaffinity chromatography with the 1D1 monoclonal antibody, were used as sense and antisense primers in the polymerase chain reaction with a brain cDNA library as template to generate an unambiguous cDNA probe. A second probe for the N-terminal portion of the early postnatal form of the proteoglycan was obtained by reverse transcription/polymerase chain reaction. The composite sequence of overlapping cDNA clones is 5.2-kilobases (kb) long, including 1.3 kb of 3'-untranslated sequence and 76 base pairs of 5'-untranslated sequence. An open reading frame of 1257 amino acids encodes a protein with a molecular mass of 136 kDa containing 10 peptide sequences present in the adult and/or early postnatal brain proteoglycans. The deduced amino acid sequence revealed a 22-amino acid signal peptide followed by an immunoglobulin domain, tandem repeats characteristic of the hyaluronic acid-binding region of aggregating proteoglycans, and an RGDS sequence. The C-terminal portion (amino acids 951-1215) has approximately 60% identity to regions in the C termini of the fibroblast and cartilage proteoglycans, versican and aggrecan, including two epidermal growth factor-like domains, a lectin-like domain, and a complement regulatory protein-like sequence. The central 595-amino acid portion of neurocan has no homology with other reported protein sequences. The proteoglycan contains six potential N-glycosylation sites and 25 potential threonine O-glycosylation sites. In the adult form of the proteoglycan (which represents the C-terminal half of neurocan) a single 32-kDa chondroitin 4-sulfate chain is linked at serin-944, whereas three additional potential chondroitin sulfate attachment sites (only two of which are utilized) are present in the larger proteoglycan species. A probe corresponding to a region of neurocan having no homology with versican or aggrecan hybridized with a single band at approximately 7.5 kb on Northern blots of mRNA from both 4-day and adult rat brain (but not with muscle, kidney, liver, or lung mRNA), indicating that the 1D1 proteoglycan of adult brain, containing a 68-kDa core protein, is generated by a developmentally regulated in vivo proteolytic processing of the 136-kDa species which is predominant in early postnatal brain.(ABSTRACT TRUNCATED AT 400 WORDS)
Notes: Rauch, U
Karthikeyan, L
Maurel, P
Margolis, R U
Margolis, R K
NS-09348/NS/NINDS NIH HHS/United States
NS-13876/NS/NINDS NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Sep 25;267(27):19536-47.
Author Address: Department of Pharmacology, New York University Medical Center, New York 10016.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 552
Author: Ravanat, C., Gachet, C., Herbert, J. M., Schuhler, S., Guillemot, J. C., Uzabiaga, F., Picard, C., Ferrara, P., Freund, M. and Cazenave, J. P.
Year: 1994
Title: Rat platelets contain glycosylated and non-glycosylated forms of platelet factor 4. Identification and characterization by mass spectrometry
Journal: Eur J Biochem
Volume: 223
Issue: 1
Pages: 203-10
Epub Date: 1994/07/01
Date: Jul 1
Short Title: Rat platelets contain glycosylated and non-glycosylated forms of platelet factor 4. Identification and characterization by mass spectrometry
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1994.tb18984.x
Accession Number: 8033893
Keywords: Amino Acid Sequence
Animals
Blood
Blood Platelets/*metabolism
Carbohydrate Metabolism
Electrophoresis, Polyacrylamide Gel
Glycosylation
Heparin/metabolism
Immunochemistry
Mass Spectrometry
Molecular Sequence Data
Platelet Factor 4/isolation & purification/*metabolism
Rats
Abstract: Platelet factor 4 is a heparin-binding protein released from the alpha granules of activated platelets. This study describes the purification and identification of two forms of rat platelet factor 4, the previously characterized non-glycosylated form of 7 kDa and an additional glycosylated form of molecular mass 9 kDa. The two proteins both neutralized the antithrombin-III-dependent inhibitory activity of heparin. Although their amino acid composition was found to be the same, in the N-terminal sequence of the 9-kDa protein, the second threonine residue could not be detected and a difference of 976Da was determined by mass spectrometry. After digestion with O-glycanase and sialidase, the two proteins showed the same molecular mass. Overall consideration of these data led to identification of the higher-molecular-mass protein as a glycosylated form of rat platelet factor 4 with O-glycosylation at the second N-terminal amino acid, while the structure of the oligosaccharide core was established by mass spectrometry and sugar differentiation with lectins. The two forms of platelet factor 4 are both present in platelets and secreted after platelet activation.
Notes: Ravanat, C
Gachet, C
Herbert, J M
Schuhler, S
Guillemot, J C
Uzabiaga, F
Picard, C
Ferrara, P
Freund, M
Cazenave, J P
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1994 Jul 1;223(1):203-10. doi: 10.1111/j.1432-1033.1994.tb18984.x.
Author Address: INSERM U311, Centre Régional de Transfusion Sanguine, Strasbourg, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 251
Author: Reason, A. J., Morris, H. R., Panico, M., Marais, R., Treisman, R. H., Haltiwanger, R. S., Hart, G. W., Kelly, W. G. and Dell, A.
Year: 1992
Title: Localization of O-GlcNAc modification on the serum response transcription factor
Journal: J Biol Chem
Volume: 267
Issue: 24
Pages: 16911-21
Epub Date: 1992/08/25
Date: Aug 25
Short Title: Localization of O-GlcNAc modification on the serum response transcription factor
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1512232
Keywords: Acetylglucosamine/*analysis/metabolism
Amino Acid Sequence
Animals
Cell Line
Chromatography, High Pressure Liquid
DNA-Binding Proteins/*chemistry/genetics/isolation & purification
Genes, fos
Glycosylation
Insecta
Molecular Sequence Data
Nuclear Proteins/*chemistry/genetics/isolation & purification
Peptide Fragments/isolation & purification
Sequence Homology, Nucleic Acid
Serum Response Factor
Spectrometry, Mass, Fast Atom Bombardment
Transcription Factors
Transfection
Abstract: A unique form of nucleoplasmic and cytoplasmic protein glycosylation, O-linked GlcNAc, has previously been detected, using Gal transferase labeling techniques, on a myriad of proteins (for review see Hart, G. W., Haltiwanger, R. S., Holt, G. D., and Kelly, W. G. (1989a) Annu. Rev. Biochem. 58, 841-874), including many RNA polymerase II transcription factors (Jackson, S. P., and Tjian, R. (1988) Cell 55, 125-133). However, virtually nothing is known about the degree of glycosylation at individual sites, or, indeed, the actual sites of attachment of O-GlcNAc on transcription factors. In this paper we provide rigorous evidence for the occurrence and locations of O-GlcNAc on the c-fos transcription factor, serum response factor (SRF), expressed in an insect cell line. Fast atom bombardment mass spectrometry (FAB-MS) of proteolytic digests of SRF provides evidence for the presence of a single substoichiometric O-GlcNAc residue on each of four peptides isolated after sequential cyanogen bromide, tryptic, and proline specific enzyme digestion: these peptides are 306VSASVSP312, 274GTTSTIQTAP283, 313SAVSSADGTVLK324, and 374DSSTDLTQTSSSGTVTLP391. Using an array of techniques, including manual Edman degradation, aminopeptidase, and elastase digestion, together with FAB-MS, the major sites of O-GlcNAc attachment were shown to be serine residues within short tandem repeat regions. The highest level of glycosylation was found on the SSS tandem repeat of peptide (374-391) which is situated within the transcriptional activation domain of SRF. The other glycosylation sites observed in SRF are located in the region of the protein between the DNA binding domain and the transcriptional activation domain. Glycosylation of peptides (274-283) and (313-324) was found to occur on the serine in the TTST tandem repeat and on serine 316 in the SS repeat, respectively. The lowest level of glycosylation was recovered in peptide (306-312) which lacks tandem repeats. All the glycosylation sites identified in SRF are situated in a relatively short region of the primary sequence close to or within the transcriptional activation domain which is distant from the major sites of phosphorylation catalyzed by casein kinase II.
Notes: Reason, A J
Morris, H R
Panico, M
Marais, R
Treisman, R H
Haltiwanger, R S
Hart, G W
Kelly, W G
Dell, A
CA 42486/CA/NCI NIH HHS/United States
HD 13563/HD/NICHD NIH HHS/United States
etc.
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Aug 25;267(24):16911-21.
Author Address: Department of Biochemistry, Imperial College of Science, Technology, and Medicine, London, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 422
Author: Reddy, M. S., Levine, M. J. and Prakobphol, A.
Year: 1985
Title: Oligosaccharide structures of the low-molecular-weight salivary mucin from a normal individual and one with cystic fibrosis
Journal: J Dent Res
Volume: 64
Issue: 1
Pages: 33-6
Epub Date: 1985/01/01
Date: Jan
Short Title: Oligosaccharide structures of the low-molecular-weight salivary mucin from a normal individual and one with cystic fibrosis
Alternate Journal: Journal of dental research
ISSN: 0022-0345 (Print)
0022-0345
DOI: 10.1177/00220345850640010601
Accession Number: 3855417
Keywords: Adolescent
Adult
Chromatography, Gel
Chromatography, Paper
Cystic Fibrosis/*metabolism
Electrophoresis, Paper
Female
Fucose/analysis
Galactose/analysis
Humans
Hydrolysis
Male
Methylation
Molecular Weight
Mucins/*analysis
Oligosaccharides/*analysis
Oxidation-Reduction
Salivary Proteins and Peptides/*analysis
Abstract: Studies were begun to compare the oligosaccharide structures of the low-molecular-weight mucin purified from the submandibular-sublingual saliva of a normal individual with that from one with cystic fibrosis. Following alkaline/borotritide cleavage, neutral and sialic acid-containing chains were purified by a combination of gel filtration, paper chromatography, and high-voltage paper electrophoresis. Oligosaccharides ranged in size from a disaccharide to a heptasaccharide. Approximately 80% of the oligosaccharides were GalB1, 3GalNAc; Fuc alpha 1,2GalB1,3GalNAc; and NeuAc alpha 2,3GalB1, 3GalNAc. The other structures were Fuc alpha 1,2GalB1,4(Fuc alpha 1,3)GlcNAcB1,6(GalB1,3)GalNAc or GalB1,4(Fuc alpha 1,3)GlcNAcB1,6(Fuc alpha 1,2GalB1,3)GAlNAc; GalB1,4(Fuc alpha 1,3)GlcNAcB1,6(NeuAc alpha 2,3GalB1,3)GalNAc; and Fuc alpha 1,2GalB1,4(Fuc alpha 1,3)GlcNAcB1,6 (NeuAc alpha 2,3GalB1,3)GalNAc. There were no apparent qualitative differences in the neutral and sialic acid-containing units recovered from the normal and cystic fibrosis samples.
Notes: Reddy, M S
Levine, M J
Prakobphol, A
DEO4518/DE/NIDCR NIH HHS/United States
DEO4971/DE/NIDCR NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Dent Res. 1985 Jan;64(1):33-6. doi: 10.1177/00220345850640010601.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 223
Author: Reid, K. B.
Year: 1979
Title: Complete amino acid sequences of the three collagen-like regions present in subcomponent C1q of the first component of human complement
Journal: Biochem J
Volume: 179
Issue: 2
Pages: 367-71
Epub Date: 1979/05/01
Date: May 1
Short Title: Complete amino acid sequences of the three collagen-like regions present in subcomponent C1q of the first component of human complement
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj1790367
PMCID: PMC1186634
Accession Number: 486087
Keywords: Amino Acid Sequence
*Complement C1
Humans
Abstract: The sequences of amino acid residues 38--51 of the A-chain, and residues 42--90 of the C-chain, of human subcomponent C1q are given. These results, along with previously published sequence data [Reid (1974) Biochem.J. 141, 189--203; Reid (1977) Biochem.J. 161, 247--251; Reid & Thompson (1978) Biochem.J. 173, 863--868] allow the presentation, and comparison with each other, of the complete amino acid sequences of the collagen-like regions found in the A-, B- and C-chains of human subcomponent C1q. Each chain has the continuity of its collagen-like Gly-X-Y repeating triplet amino acid sequence broken. The B- and C-chains have alanine residues at positions B-9 and C-36 where glycine might be expected. The A-chain has a threonine residue at position A-39, which is located between two Gly-X-Y triplets.
Notes: 1470-8728
Reid, K B
Journal Article
Biochem J. 1979 May 1;179(2):367-71. doi: 10.1042/bj1790367.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1405
Author: Reiling, J. H., Clish, C. B., Carette, J. E., Varadarajan, M., Brummelkamp, T. R. and Sabatini, D. M.
Year: 2011
Title: A haploid genetic screen identifies the major facilitator domain containing 2A (MFSD2A) transporter as a key mediator in the response to tunicamycin
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 29
Pages: 11756-65
Epub Date: 2011/06/17
Date: Jul 19
Short Title: A haploid genetic screen identifies the major facilitator domain containing 2A (MFSD2A) transporter as a key mediator in the response to tunicamycin
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1018098108
PMCID: PMC3141996
Accession Number: 21677192
Keywords: Blotting, Western
Cell Line
Cell Survival
Endoplasmic Reticulum/*metabolism
Glycosylation/drug effects
Humans
Immunoprecipitation
Membrane Transport Proteins/*metabolism
Microscopy, Confocal
Protein Folding/*drug effects
Signal Transduction/*physiology
Symporters
Tumor Suppressor Proteins/genetics/*metabolism
Tunicamycin/*pharmacology
Abstract: Tunicamycin (TM) inhibits eukaryotic asparagine-linked glycosylation, protein palmitoylation, ganglioside production, proteoglycan synthesis, 3-hydroxy-3-methylglutaryl coenzyme-A reductase activity, and cell wall biosynthesis in bacteria. Treatment of cells with TM elicits endoplasmic reticulum stress and activates the unfolded protein response. Although widely used in laboratory settings for many years, it is unknown how TM enters cells. Here, we identify in an unbiased genetic screen a transporter of the major facilitator superfamily, major facilitator domain containing 2A (MFSD2A), as a critical mediator of TM toxicity. Cells without MFSD2A are TM-resistant, whereas MFSD2A-overexpressing cells are hypersensitive. Hypersensitivity is associated with increased cellular TM uptake concomitant with an enhanced endoplasmic reticulum stress response. Furthermore, MFSD2A mutant analysis reveals an important function of the C terminus for correct intracellular localization and protein stability, and it identifies transmembrane helical amino acid residues essential for mediating TM sensitivity. Overall, our data uncover a critical role for MFSD2A by acting as a putative TM transporter at the plasma membrane.
Notes: 1091-6490
Reiling, Jan H
Clish, Clary B
Carette, Jan E
Varadarajan, Malini
Brummelkamp, Thijn R
Sabatini, David M
R01 CA103866/CA/NCI NIH HHS/United States
R21 HG004938/HG/NHGRI NIH HHS/United States
CA103866/CA/NCI NIH HHS/United States
Howard Hughes Medical Institute/United States
CA129105/CA/NCI NIH HHS/United States
R01 CA129105/CA/NCI NIH HHS/United States
R21 HG004938-01/HG/NHGRI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Jul 19;108(29):11756-65. doi: 10.1073/pnas.1018098108. Epub 2011 Jun 15.
Author Address: Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1147
Author: Rengifo, J., Gibson, C. J., Winkler, E., Collin, T. and Ehrlich, B. E.
Year: 2007
Title: Regulation of the inositol 1,4,5-trisphosphate receptor type I by O-GlcNAc glycosylation
Journal: J Neurosci
Volume: 27
Issue: 50
Pages: 13813-21
Epub Date: 2007/12/14
Date: Dec 12
Short Title: Regulation of the inositol 1,4,5-trisphosphate receptor type I by O-GlcNAc glycosylation
Alternate Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience
ISSN: 0270-6474 (Print)
0270-6474
DOI: 10.1523/jneurosci.2069-07.2007
PMCID: PMC6673603
Accession Number: 18077693
Keywords: Animals
Calcium Signaling/drug effects/physiology
Cells, Cultured
Cerebellum/chemistry/metabolism
Glucosamine/*analogs & derivatives/chemistry/metabolism
Glycosylation
Inositol 1,4,5-Trisphosphate/analogs & derivatives/metabolism/pharmacology
Inositol 1,4,5-Trisphosphate Receptors/*chemistry/*metabolism
Interneurons/drug effects/*metabolism
Lipid Bilayers/chemistry
Mice
Microsomes/chemistry
N-Acetylglucosaminyltransferases/chemistry/metabolism
Organ Culture Techniques
Patch-Clamp Techniques
Rats
Ultraviolet Rays
Uridine Diphosphate N-Acetylglucosamine/metabolism/pharmacology
beta-N-Acetylhexosaminidases/chemistry
Abstract: The inositol 1,4,5-trisphosphate (InsP3) receptor type I (InsP3R-I) is the principle channel for intracellular calcium (Ca2+) release in many cell types, including central neurons. It is regulated by endogenous compounds like Ca2+ and ATP, by protein partners, and by posttranslational modification. We report that the InsP3R-I is modified by O-linked glycosylation of serine or threonine residues with beta-N-acetylglucosamine (O-GlcNAc). The level of O-GlcNAcylation can be altered in vitro by the addition of the enzymes which add [OGT (O-GlcNActransferase)] or remove (O-GlcNAcase) this sugar or by loading cells with UDP-GlcNAc. We monitored the effects of this modification on InsP3R function at the single-channel level and on intracellular Ca2+ transients. Single-channel activity was monitored with InsP3R incorporated into bilayers; Ca2+ signaling was monitored using cells loaded with a Ca2+-sensitive fluorophore. We found that channel activity was decreased by the addition of O-GlcNAc and that this decrease was reversed by removal of the sugar. Similarly, cells loaded with UDP-GlcNAc had an attenuated response to uncaging of InsP3. These results show that O-GlcNAcylation is an important regulator of the InsP3R-I and suggest a mechanism for neuronal dysfunction under conditions in which O-GlcNAc is high, such as diabetes or physiological stress.
Notes: 1529-2401
Rengifo, Juliana
Gibson, Craig J
Winkler, Eva
Collin, Thibault
Ehrlich, Barbara E
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Neurosci. 2007 Dec 12;27(50):13813-21. doi: 10.1523/JNEUROSCI.2069-07.2007.
Author Address: Department of Pharmacology and Cellular and Molecular Physiology, Yale University, New Haven, Connecticut 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 932
Author: Reverter, D., Maskos, K., Tan, F., Skidgel, R. A. and Bode, W.
Year: 2004
Title: Crystal structure of human carboxypeptidase M, a membrane-bound enzyme that regulates peptide hormone activity
Journal: J Mol Biol
Volume: 338
Issue: 2
Pages: 257-69
Epub Date: 2004/04/07
Date: Apr 23
Short Title: Crystal structure of human carboxypeptidase M, a membrane-bound enzyme that regulates peptide hormone activity
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2004.02.058
Accession Number: 15066430
Keywords: Amino Acid Sequence
Animals
Catalytic Domain
Cell Membrane/metabolism
Crystallography, X-Ray
GPI-Linked Proteins
Glycosylphosphatidylinositols
Humans
Metalloendopeptidases/*chemistry/genetics/metabolism
Models, Molecular
Molecular Sequence Data
Peptide Hormones/*metabolism
Protein Structure, Secondary
*Protein Structure, Tertiary
Sequence Alignment
Abstract: Carboxypeptidase M (CPM), an extracellular glycosylphosphatidyl-inositol(GPI)-anchored membrane glycoprotein belonging to the CPN/E subfamily of "regulatory" metallo-carboxypeptidases, specifically removes C-terminal basic residues from peptides and proteins. Due to its wide distribution in human tissues, CPM is believed to play important roles in the control of peptide hormone and growth factor activity at the cell surface, and in the membrane-localized degradation of extracellular proteins. We have crystallized human GPI-free CPM, and have determined and refined its 3.0A crystal structure. The structure analysis reveals that CPM consists of a 295 residue N-terminal catalytic domain similar to that of duck CPD-2 (but only distantly related to CPA/B), an adjacent 86 residue beta-sandwich C-terminal domain characteristic of the CPN/E family but more conically shaped than the equivalent domain in CPD-2, and a unique, partially disordered 25 residue C-terminal extension to which the GPI membrane-anchor is post-translationally attached. Through this GPI anchor, and presumably via some positively charged side-chains of the C-terminal domain, the CPM molecule may interact with the membrane in such a way that its active centre will face alongside, i.e. well suited to interact with other membrane-bound protein substrates or small peptides. Modelling of the C-terminal part of the natural substrate Arg(6)-Met-enkephalin into the active site shows that the S1' pocket of CPM is particularly well designed to accommodate P1'-Arg residues, in agreement with the preference of CPM for cleaving C-terminal Arg.
Notes: Reverter, David
Maskos, Klaus
Tan, Fulong
Skidgel, Randal A
Bode, Wolfram
DK41431/DK/NIDDK NIH HHS/United States
HL60678/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
J Mol Biol. 2004 Apr 23;338(2):257-69. doi: 10.1016/j.jmb.2004.02.058.
Author Address: Abteilung für Strukturforschung, Max-Planck-Institut für Biochemie, Am Klopferspitz 18a, D-82152 Planegg-Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1325
Author: Rexach, J. E., Rogers, C. J., Yu, S. H., Tao, J., Sun, Y. E. and Hsieh-Wilson, L. C.
Year: 2010
Title: Quantification of O-glycosylation stoichiometry and dynamics using resolvable mass tags
Journal: Nat Chem Biol
Volume: 6
Issue: 9
Pages: 645-51
Epub Date: 2010/07/27
Date: Sep
Short Title: Quantification of O-glycosylation stoichiometry and dynamics using resolvable mass tags
Alternate Journal: Nature chemical biology
ISSN: 1552-4450 (Print)
1552-4450
DOI: 10.1038/nchembio.412
PMCID: PMC2924450
NIHMSID: NIHMS210959 financial interests. Reprints and permissions information is available online at http://npg.nature.com/reprintsandpermissions/
Accession Number: 20657584
Keywords: Acetylglucosamine/*analysis/chemistry/*metabolism
Glycosylation
Kinetics
Mass Spectrometry
Methyl-CpG-Binding Protein 2/genetics/metabolism
Phosphorylation
Polyethylene Glycols/analysis/*chemistry
*Protein Processing, Post-Translational/genetics
Transcription, Genetic
Abstract: Mechanistic studies of O-GlcNAc glycosylation have been limited by an inability to monitor the glycosylation stoichiometries of proteins obtained from cells. Here we describe a powerful method to visualize the O-GlcNAc-modified protein subpopulation using resolvable polyethylene glycol mass tags. This approach enables rapid quantification of in vivo glycosylation levels on endogenous proteins without the need for protein purification, advanced instrumentation or expensive radiolabels. In addition, it establishes the glycosylation state (for example, mono-, di-, tri-) of proteins, providing information regarding overall O-GlcNAc site occupancy that cannot be obtained using mass spectrometry. Finally, we apply this strategy to rapidly assess the complex interplay between glycosylation and phosphorylation and discover an unexpected reverse 'yin-yang' relationship on the transcriptional repressor MeCP2 that was undetectable by traditional methods. We anticipate that this mass-tagging strategy will advance our understanding of O-GlcNAc glycosylation, as well as other post-translational modifications and poorly understood glycosylation motifs.
Notes: 1552-4469
Rexach, Jessica E
Rogers, Claude J
Yu, Seok-Ho
Tao, Jifang
Sun, Yi E
Hsieh-Wilson, Linda C
R01 GM084724-07/GM/NIGMS NIH HHS/United States
R01 GM084724/GM/NIGMS NIH HHS/United States
R01 GM084724-07S1/GM/NIGMS NIH HHS/United States
F31 NS056525/NS/NINDS NIH HHS/United States
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Chem Biol. 2010 Sep;6(9):645-51. doi: 10.1038/nchembio.412. Epub 2010 Jul 25.
Author Address: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 241
Author: Rice, K. G., Takahashi, N., Namiki, Y., Tran, A. D., Lisi, P. J. and Lee, Y. C.
Year: 1992
Title: Quantitative mapping of the N-linked sialyloligosaccharides of recombinant erythropoietin: combination of direct high-performance anion-exchange chromatography and 2-aminopyridine derivatization
Journal: Anal Biochem
Volume: 206
Issue: 2
Pages: 278-87
Epub Date: 1992/11/01
Date: Nov 1
Short Title: Quantitative mapping of the N-linked sialyloligosaccharides of recombinant erythropoietin: combination of direct high-performance anion-exchange chromatography and 2-aminopyridine derivatization
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/0003-2697(92)90367-g
Accession Number: 1443598
Keywords: Amidohydrolases
Aminopyridines
Animals
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Ion Exchange/methods
Cricetinae
Electrophoresis, Polyacrylamide Gel
Erythropoietin/*chemistry
Glycopeptides/isolation & purification
Humans
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Recombinant Proteins/*chemistry
Abstract: A rapid quantitative analysis of the sialylated N-linked oligosaccharides of recombinant erythropoietin (EPO) expressed in Chinese hamster ovary (CHO) cells has been developed. The procedure utilizes a glycoamidase (glycopeptidase F) to release all of the N-linked oligosaccharides from the native glycoprotein, followed by direct chromatographic analysis using high-performance anion-exchange chromatography (HPAEC) with pulsed amperometric detection. The eight sialyloligosaccharides isolated from HPAEC were characterized by derivatizing with 2-aminopyridine followed by two-dimensional HPLC mapping of the pyridylaminated asialooligosaccharides (Tomiya et al., 1988, Anal. Biochem. 171, 73-90). Seven kinds of complex-type asialooligosaccharides were identified ranging from a biantennary structure to N-acetyllactosamine-extended tetraantennary structure. Approximately 3% of the terminal galactose residues of the oligosaccharides released from EPO were not sialylated whereas 97% contained an alpha(2-->3)-linked sialic acid. Quantitative oligosaccharide mapping of four different lots of EPO from CHO cells was performed to quantify the molar balance and distribution of the N-linked oligosaccharides. The sialyloligosaccharides were distributed with approximately 5% disialylated (single type), 20% trisialylated (six types), and 75% tetrasialylated (four types) oligosaccharides with an average molar recovery of 85% starting from 750 pmol of EPO.
Notes: Rice, K G
Takahashi, N
Namiki, Y
Tran, A D
Lisi, P J
Lee, Y C
DK09970/DK/NIDDK NIH HHS/United States
GM13013-01/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Anal Biochem. 1992 Nov 1;206(2):278-87. doi: 10.1016/0003-2697(92)90367-g.
Author Address: Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1273
Author: Richards, A. A., Colgrave, M. L., Zhang, J., Webster, J., Simpson, F., Preston, E., Wilks, D., Hoehn, K. L., Stephenson, M., Macdonald, G. A., Prins, J. B., Cooney, G. J., Xu, A. and Whitehead, J. P.
Year: 2010
Title: Sialic acid modification of adiponectin is not required for multimerization or secretion but determines half-life in circulation
Journal: Mol Endocrinol
Volume: 24
Issue: 1
Pages: 229-39
Epub Date: 2009/10/27
Date: Jan
Short Title: Sialic acid modification of adiponectin is not required for multimerization or secretion but determines half-life in circulation
Alternate Journal: Molecular endocrinology (Baltimore, Md.)
ISSN: 0888-8809 (Print)
0888-8809
DOI: 10.1210/me.2009-0133
PMCID: PMC5428139
Accession Number: 19855092
Keywords: Adiponectin/blood/chemistry/genetics/*metabolism
Animals
Asialoglycoprotein Receptor/metabolism
Carbohydrate Conformation
Cell Line
Cells, Cultured
Cricetinae
Cricetulus
Glycosylation
Half-Life
Hepatocytes/metabolism
Humans
Insulin Resistance
Male
Mice
Mice, Inbred C57BL
Mice, Knockout
*Protein Multimerization
*Protein Processing, Post-Translational
Rats
Rats, Sprague-Dawley
Rats, Wistar
Recombinant Proteins/metabolism
Sialic Acids/*metabolism
Sialyltransferases/*metabolism
Abstract: Adiponectin is an adipocyte-secreted, insulin-sensitizing hormone the circulating levels of which are reduced in conditions of insulin resistance and diabetes. Previous work has demonstrated the importance of posttranslational modifications, such as proline hydroxylation and lysine hydroxylation/glycosylation, in adiponectin oligomerization, secretion, and function. Here we describe the first functional characterization of adiponectin sialylation. Using a variety of biochemical approaches we demonstrated that sialylation occurs on previously unidentified O-linked glycans on Thr residues of the variable domain in human adiponectin. Enzymatic removal of sialic acid or its underlying O-linked sugars did not affect adiponectin multimer composition. Expression of mutant forms of adiponectin (lacking the modified Thr residues) or of wild-type adiponectin in cells defective in sialylation did not compromise multimer formation or secretion, arguing against a structural role for this modification. Activity of desialylated adiponectin was comparable to control adiponectin in L6 myotubes and acute assays in adiponectin(-/-) mice. In contrast, plasma clearance of desialylated adiponectin was accelerated compared with that of control adiponectin, implicating a role for this modification in determining the half-life of circulating adiponectin. Uptake of desialylated adiponectin by isolated primary rat hepatocytes was also accelerated, suggesting a role for the hepatic asialoglycoprotein receptor. Finally, after chronic administration in adiponectin(-/-) mice steady-state levels of desialylated adiponectin were lower than control adiponectin and failed to recapitulate the improvements in glucose and insulin tolerance tests observed with control adiponectin. These data suggest an important role for sialic acid content in the regulation of circulating adiponectin levels and highlight the importance of understanding mechanisms regulating adiponectin sialylation/desialylation.
Notes: 1944-9917
Richards, Ayanthi A
Colgrave, Michelle L
Zhang, Jialiang
Webster, Julie
Simpson, Fiona
Preston, Elaine
Wilks, Donna
Hoehn, Kyle L
Stephenson, Matthew
Macdonald, Graeme A
Prins, John B
Cooney, Gregory J
Xu, Aimin
Whitehead, Jonathan P
Journal Article
Research Support, Non-U.S. Gov't
Mol Endocrinol. 2010 Jan;24(1):229-39. doi: 10.1210/me.2009-0133. Epub 2009 Oct 23.
Author Address: Diamantina Institute for Cancer, Immunology and Metabolic Medicine, University of Queensland, Brisbane, Queensland, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1036
Author: Richards, A. A., Stephens, T., Charlton, H. K., Jones, A., Macdonald, G. A., Prins, J. B. and Whitehead, J. P.
Year: 2006
Title: Adiponectin multimerization is dependent on conserved lysines in the collagenous domain: evidence for regulation of multimerization by alterations in posttranslational modifications
Journal: Mol Endocrinol
Volume: 20
Issue: 7
Pages: 1673-87
Epub Date: 2006/02/25
Date: Jul
Short Title: Adiponectin multimerization is dependent on conserved lysines in the collagenous domain: evidence for regulation of multimerization by alterations in posttranslational modifications
Alternate Journal: Molecular endocrinology (Baltimore, Md.)
ISSN: 0888-8809 (Print)
0888-8809
DOI: 10.1210/me.2005-0390
Accession Number: 16497731
Keywords: 2,2'-Dipyridyl/pharmacology
Adiponectin/chemistry/metabolism
Amino Acid Sequence
Animals
Bacteria/genetics/metabolism
CHO Cells
Collagen/metabolism
Conserved Sequence
Cricetinae
*Dimerization
Gene Expression
Glucose/pharmacology
Glycosylation
Humans
Hydroxylation/drug effects
Lysine/metabolism
Mass Spectrometry
Models, Biological
Molecular Sequence Data
Multiprotein Complexes/*metabolism
Proline/metabolism
Protein Isoforms/metabolism
*Protein Processing, Post-Translational
Protein Structure, Tertiary
Recombinant Proteins/chemistry
Abstract: Adiponectin is a secreted, multimeric protein with insulin-sensitizing, antiatherogenic, and antiinflammatory properties. Serum adiponectin consists of trimer, hexamer, and larger high-molecular-weight (HMW) multimers, and these HMW multimers appear to be the more bioactive forms. Multimer composition of adiponectin appears to be regulated; however, the molecular mechanisms involved are unknown. We hypothesize that regulation of adiponectin multimerization and secretion occurs via changes in posttranslational modifications (PTMs). Although a structural role for intertrimer disulfide bonds in the formation of hexamers and HMW multimers is established, the role of other PTMs is unknown. PTMs identified in murine and bovine adiponectin include hydroxylation of multiple conserved proline and lysine residues and glycosylation of hydroxylysines. By mass spectrometry, we confirmed the presence of these PTMs in human adiponectin and identified three additional hydroxylations on Pro71, Pro76, and Pro95. We also investigated the role of the five modified lysines in multimer formation and secretion of recombinant human adiponectin expressed in mammalian cell lines. Mutation of modified lysines in the collagenous domain prevented formation of HMW multimers, whereas a pharmacological inhibitor of prolyl- and lysyl-hydroxylases, 2,2'-dipyridyl, inhibited formation of hexamers and HMW multimers. Bacterially expressed human adiponectin displayed a complete lack of differentially modified isoforms and failed to form bona fide trimers and larger multimers. Finally, glucose-induced increases in HMW multimer production from human adipose explants correlated with changes in the two-dimensional electrophoresis profile of adiponectin isoforms. Collectively, these data suggest that adiponectin multimer composition is affected by changes in PTM in response to physiological factors.
Notes: Richards, Ayanthi A
Stephens, Tim
Charlton, Hayley K
Jones, Alun
Macdonald, Graeme A
Prins, Johannes B
Whitehead, Jonathan P
Journal Article
Research Support, Non-U.S. Gov't
United States
Mol Endocrinol. 2006 Jul;20(7):1673-87. doi: 10.1210/me.2005-0390. Epub 2006 Feb 23.
Author Address: Centre for Diabetes and Endocrine Research, University of Queensland, Princess Alexandra Hospital, Brisbane, Queensland 4102, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 770
Author: Richter, R., Schulz-Knappe, P., John, H. and Forssmann, W. G.
Year: 2000
Title: Posttranslationally processed forms of the human chemokine HCC-1
Journal: Biochemistry
Volume: 39
Issue: 35
Pages: 10799-805
Epub Date: 2000/09/09
Date: Sep 5
Short Title: Posttranslationally processed forms of the human chemokine HCC-1
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi992488q
Accession Number: 10978165
Keywords: Amino Acid Sequence
Blood Proteins/genetics/isolation & purification/metabolism
Blotting, Western
Chemokines, CC/blood/*genetics/isolation & purification/*metabolism
Hemofiltration
Humans
Molecular Sequence Data
Peptide Fragments/blood/genetics/isolation & purification/metabolism
Peptide Library
*RNA Processing, Post-Transcriptional
Radioimmunoassay
Abstract: HCC-1 is the only CC-chemokine known so far which circulates in nanomolar concentrations in human plasma. Its physiological function is not well defined. Posttranslational processing of HCC-1 was shown to modulate its biological properties. In this study several different processed forms of HCC-1 were isolated. Western blot analysis of human plasma extracts revealed a HCC-1 immunoreactive double band at 8-10 kDa indicating the presence of two distinct HCC-1 peptides. These peptides were isolated from a peptide library of human blood filtrate and represent predominantly HCC-1 (1-74) and glycosylated HCC-1 (1-74). Glycosylated HCC-1 exhibits a molecular mass of 9621 Da due to O-glycosylation at position 7 (Ser-7) with two N-acetylneuraminic acids and the disaccharide N-acetylgalactosamine galactose. Furthermore N-terminally truncated HCC-1 (3-74) and HCC-1 (4-74) were identified in the peptide library. In hemofiltrate approximately 3% of total HCC-1 represents HCC-1 (3-74) and approximately 1% represents HCC-1 (4-74) whereas the major products are nonglycosylated HCC-1 (1-74) and glycosylated HCC-1 (1-74). Our data imply that HCC-1 (1-74), HCC-1 (3-74), HCC-1 (4-74) and glycosylated HCC-1 (1-74) circulate in human blood. The N-terminal processing and modification of HCC-1 might be of importance in displaying its full biological activity.
Notes: Richter, R
Schulz-Knappe, P
John, H
Forssmann, W G
Journal Article
United States
Biochemistry. 2000 Sep 5;39(35):10799-805. doi: 10.1021/bi992488q.
Author Address: Lower Saxony Institute for Peptide Research, Feodor-Lynen Strasse 31, D-30625 Hannover, Germany. rudorichter@gmx.de
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1101
Author: Ricketts, L. M., Dlugosz, M., Luther, K. B., Haltiwanger, R. S. and Majerus, E. M.
Year: 2007
Title: O-fucosylation is required for ADAMTS13 secretion
Journal: J Biol Chem
Volume: 282
Issue: 23
Pages: 17014-23
Epub Date: 2007/03/31
Date: Jun 8
Short Title: O-fucosylation is required for ADAMTS13 secretion
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M700317200
Accession Number: 17395589
Keywords: ADAM Proteins/genetics/*metabolism
ADAMTS13 Protein
Amino Acid Sequence
Base Sequence
Cell Line
DNA Primers
Fucose/*metabolism
Humans
Molecular Sequence Data
Mutagenesis, Site-Directed
RNA, Small Interfering
Abstract: ADAMTS13 is a plasma metalloproteinase that cleaves von Willebrand factor to smaller, less thrombogenic forms. Deficiency of ADAMTS13 activity in plasma leads to thrombotic thrombocytopenic purpura. ADAMTS13 contains eight thrombospondin type 1 repeats (TSR), seven of which contain a consensus sequence for the direct addition of fucose to the hydroxyl group of serine or threonine. Mass spectral analysis of tryptic peptides derived from human ADAMTS13 indicate that at least six of the TSRs are modified with an O-fucose disaccharide. Analysis of [(3)H]fucose metabolically incorporated into ADAMTS13 demonstrated that the disaccharide has the structure glucose-beta1,3-fucose. Mutation of the modified serine to alanine in TSR2, TSR5, TSR7, and TSR8 reduced the secretion of ADAMTS13. Mutation of more than one site dramatically reduced secretion regardless of the sites mutated. When the expression of protein O-fucosyltransferase 2 (POFUT2), the enzyme that transfers fucose to serines in TSRs, was reduced using siRNA, the secretion of ADAMTS13 decreased. A similar outcome was observed when ADAMTS13 was expressed in a cell line unable to synthesize the donor for fucose addition, GDP-fucose. Although overexpression of POFUT2 did not affect the secretion of wild-type ADAMTS13, it did increase the secretion of the ADAMTS13 TSR1,2 double mutant but not that of ADAMTS13 TSR1-8 mutant. Together these findings indicate that O-fucosylation is functionally significant for secretion of ADAMTS13.
Notes: Ricketts, Lindsay M
Dlugosz, Malgosia
Luther, Kelvin B
Haltiwanger, Robert S
Majerus, Elaine M
R01 CA123071/CA/NCI NIH HHS/United States
R01 CA123071-01A1/CA/NCI NIH HHS/United States
GM61126/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2007 Jun 8;282(23):17014-23. doi: 10.1074/jbc.M700317200. Epub 2007 Mar 29.
Author Address: Department of Internal Medicine, Division of Hematology, Washington University School of Medicine, St. Louis, Missouri, 63110, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1828
Author: Riethmueller, S., Somasundaram, P., Ehlers, J. C., Hung, C. W., Flynn, C. M., Lokau, J., Agthe, M., Düsterhöft, S., Zhu, Y., Grötzinger, J., Lorenzen, I., Koudelka, T., Yamamoto, K., Pickhinke, U., Wichert, R., Becker-Pauly, C., Rädisch, M., Albrecht, A., Hessefort, M., Stahnke, D., Unverzagt, C., Rose-John, S., Tholey, A. and Garbers, C.
Year: 2017
Title: Proteolytic Origin of the Soluble Human IL-6R In Vivo and a Decisive Role of N-Glycosylation
Journal: PLoS Biol
Volume: 15
Issue: 1
Pages: e2000080
Epub Date: 2017/01/07
Date: Jan
Short Title: Proteolytic Origin of the Soluble Human IL-6R In Vivo and a Decisive Role of N-Glycosylation
Alternate Journal: PLoS biology
ISSN: 1544-9173 (Print)
1544-9173
DOI: 10.1371/journal.pbio.2000080
PMCID: PMC5218472
Accession Number: 28060820
Keywords: ADAM10 Protein/metabolism
ADAM17 Protein/metabolism
Alternative Splicing/genetics
Amino Acid Sequence
Amyloid Precursor Protein Secretases/metabolism
Cell Line
Cell Membrane/metabolism
Glycosylation
Humans
Intracellular Space/metabolism
Mass Spectrometry
Membrane Proteins/metabolism
Mutation/genetics
Polysaccharides/metabolism
Proline/metabolism
Protein Domains
Protein Transport
*Proteolysis
RNA, Messenger/genetics/metabolism
Receptors, Interleukin-6/blood/chemistry/genetics/*metabolism
Signal Transduction
Solubility
Valine/metabolism
Abstract: Signaling of the cytokine interleukin-6 (IL-6) via its soluble IL-6 receptor (sIL-6R) is responsible for the proinflammatory properties of IL-6 and constitutes an attractive therapeutic target, but how the sIL-6R is generated in vivo remains largely unclear. Here, we use liquid chromatography-mass spectrometry to identify an sIL-6R form in human serum that originates from proteolytic cleavage, map its cleavage site between Pro-355 and Val-356, and determine the occupancy of all O- and N-glycosylation sites of the human sIL-6R. The metalloprotease a disintegrin and metalloproteinase 17 (ADAM17) uses this cleavage site in vitro, and mutation of Val-356 is sufficient to completely abrogate IL-6R proteolysis. N- and O-glycosylation were dispensable for signaling of the IL-6R, but proteolysis was orchestrated by an N- and O-glycosylated sequon near the cleavage site and an N-glycan exosite in domain D1. Proteolysis of an IL-6R completely devoid of glycans is significantly impaired. Thus, glycosylation is an important regulator for sIL-6R generation.
Notes: 1545-7885
Riethmueller, Steffen
Somasundaram, Prasath
Ehlers, Johanna C
Hung, Chien-Wen
Flynn, Charlotte M
Lokau, Juliane
Agthe, Maria
Düsterhöft, Stefan
Zhu, Yijue
Grötzinger, Joachim
Lorenzen, Inken
Koudelka, Tomas
Yamamoto, Kosuke
Pickhinke, Ute
Wichert, Rielana
Becker-Pauly, Christoph
Rädisch, Marisa
Albrecht, Alexander
Hessefort, Markus
Stahnke, Dominik
Unverzagt, Carlo
Rose-John, Stefan
Tholey, Andreas
Garbers, Christoph
Journal Article
Research Support, Non-U.S. Gov't
PLoS Biol. 2017 Jan 6;15(1):e2000080. doi: 10.1371/journal.pbio.2000080. eCollection 2017 Jan.
Author Address: Institute of Biochemistry, Kiel University, Kiel, Germany.
Systematic Proteomics & Bioanalytics, Institute for Experimental Medicine, Kiel University, Kiel, Germany.
Institute of Biochemistry, Unit for Degradomics of the Protease Web, Kiel University, Kiel, Germany.
Bioorganic Chemistry, Gebaeude NWI, University of Bayreuth, Bayreuth, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2016
Author: Riley, N. M., Hebert, A. S., Westphall, M. S. and Coon, J. J.
Year: 2019
Title: Capturing site-specific heterogeneity with large-scale N-glycoproteome analysis
Journal: Nat Commun
Volume: 10
Issue: 1
Pages: 1311
Epub Date: 2019/03/23
Date: Mar 21
Short Title: Capturing site-specific heterogeneity with large-scale N-glycoproteome analysis
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-019-09222-w
PMCID: PMC6428843 Thermo Fisher Scientific. The other authors declare no competing interests.
Accession Number: 30899004
Keywords: Animals
Brain/*metabolism
Brain Chemistry
Datasets as Topic
Female
Gene Expression
Glycopeptides/*chemistry/classification/genetics/isolation & purification
Glycosylation
Mice
Mice, Inbred C57BL
Nerve Tissue Proteins/*chemistry/classification/genetics/isolation & purification
Polysaccharides/*chemistry/isolation & purification
*Protein Processing, Post-Translational
Proteome/*chemistry/classification/genetics/isolation & purification
Tandem Mass Spectrometry/instrumentation/methods
Abstract: Protein glycosylation is a highly important, yet poorly understood protein post-translational modification. Thousands of possible glycan structures and compositions create potential for tremendous site heterogeneity. A lack of suitable analytical methods for large-scale analyses of intact glycopeptides has limited our abilities both to address the degree of heterogeneity across the glycoproteome and to understand how this contributes biologically to complex systems. Here we show that N-glycoproteome site-specific microheterogeneity can be captured via large-scale glycopeptide profiling methods enabled by activated ion electron transfer dissociation (AI-ETD), ultimately characterizing 1,545 N-glycosites (>5,600 unique N-glycopeptides) from mouse brain tissue. Our data reveal that N-glycosylation profiles can differ between subcellular regions and structural domains and that N-glycosite heterogeneity manifests in several different forms, including dramatic differences in glycosites on the same protein. Moreover, we use this large-scale glycoproteomic dataset to develop several visualizations that will prove useful for analyzing intact glycopeptides in future studies.
Notes: 2041-1723
Riley, Nicholas M
Orcid: 0000-0002-1536-2966
Hebert, Alexander S
Westphall, Michael S
Coon, Joshua J
K00 CA212454/CA/NCI NIH HHS/United States
R35 GM118110/GM/NIGMS NIH HHS/United States
R35 GM118110/NH/NIH HHS/United States
P41 GM108538/NH/NIH HHS/United States
P41 GM108538/GM/NIGMS NIH HHS/United States
F99 CA212454/CA/NCI NIH HHS/United States
F99 CA212454/NH/NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Nat Commun. 2019 Mar 21;10(1):1311. doi: 10.1038/s41467-019-09222-w.
Author Address: Genome Center of Wisconsin, University of Wisconsin-Madison, Madison, WI, 53706, USA.
Department of Chemistry, University of Wisconsin-Madison, Madison, WI, 53706, USA.
Department of Chemistry, Stanford University, Stanford, CA, 94305, USA.
Genome Center of Wisconsin, University of Wisconsin-Madison, Madison, WI, 53706, USA. jcoon@chem.wisc.edu.
Department of Chemistry, University of Wisconsin-Madison, Madison, WI, 53706, USA. jcoon@chem.wisc.edu.
Department of Biomolecular Chemistry, University of Wisconsin-Madison, Madison, WI, 53706, USA. jcoon@chem.wisc.edu.
Morgridge Institute for Research, Madison, WI, 53715, USA. jcoon@chem.wisc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 464
Author: Rinderknecht, E., O'Connor, B. H. and Rodriguez, H.
Year: 1984
Title: Natural human interferon-gamma. Complete amino acid sequence and determination of sites of glycosylation
Journal: J Biol Chem
Volume: 259
Issue: 11
Pages: 6790-7
Epub Date: 1984/06/10
Date: Jun 10
Short Title: Natural human interferon-gamma. Complete amino acid sequence and determination of sites of glycosylation
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6427223
Keywords: Amino Acid Sequence
*Carbohydrate Metabolism
Carboxypeptidases/metabolism
Chromatography, Gel
Chromatography, High Pressure Liquid
Dioxanes/pharmacology
Endopeptidases/metabolism
Humans
Interferon-gamma/*analysis
Molecular Weight
*Serine Endopeptidases
Trypsin/metabolism
Abstract: Fresh human peripheral blood lymphocytes were induced with desacetylthymosin -alpha 1 and staphylococcal enterotoxin B. The induced gamma interferon (or IFN-gamma, immune interferon, type II interferon) was purified to homogeneity utilizing controlled-pore glass, concanavalin A-Sepharose, Bio-Gel P100, or Sephacryl S-200, and reversed phase high performance liquid chromatography. This procedure resulted in two active species with apparent Mr = 20,000 and 25,000 as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Both species were found to have identical amino acid sequences with a pyroglutamate residue as NH2-terminus. In both cases six different COOH termini were found. They are, at least qualitatively, identical in both species. There are two possible Asn-X-Ser/Thr glycosylation sites. Both carry carbohydrates in the Mr = 25,000 species whereas in the Mr = 20,000 species only one site is glycosylated. This likely explains the difference in apparent molecular weight between the two species and the expected molecular weight based upon the amino acid sequence.
Notes: Rinderknecht, E
O'Connor, B H
Rodriguez, H
Journal Article
United States
J Biol Chem. 1984 Jun 10;259(11):6790-7.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 229
Author: Rip, J. W., Coulter-Mackie, M. B., Rupar, C. A. and Gordon, B. A.
Year: 1992
Title: Purification and structure of human liver aspartylglucosaminidase
Journal: Biochem J
Volume: 288 ( Pt 3)
Issue: Pt 3
Pages: 1005-10
Epub Date: 1992/12/15
Date: Dec 15
Short Title: Purification and structure of human liver aspartylglucosaminidase
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2881005
PMCID: PMC1131987
Accession Number: 1281977
Keywords: Amino Acid Sequence
Aspartylglucosylaminase/chemistry/*isolation & purification/metabolism
Dextrans
Electrophoresis, Polyacrylamide Gel
Ethanolamines
Glycoside Hydrolases/metabolism
Humans
Hydrolases/isolation & purification
Liver/*enzymology
Macromolecular Substances
Molecular Sequence Data
Sepharose
Sodium Dodecyl Sulfate
Abstract: We have recently diagnosed aspartylglucosaminuria (AGU) in four members of a Canadian family. AGU is a lysosomal storage disease in which asparagine-linked glycopeptides accumulate to particularly high concentrations in liver, spleen and thyroid of affected individuals. A lesser accumulation of these glycopeptides is seen in the kidney and brain, and they are also excreted in the urine. The altered metabolism in AGU results from a deficiency of the enzyme aspartylglucosaminidase (1-aspartamido-beta-N-acetylglucosamine amidohydrolase), which hydrolyses the asparagine to N-acetylglucosamine linkages of glycoproteins and glycopeptides. We have used human liver as a source of material for the purification of aspartylglucosaminidase. The enzyme has been purified to homogeneity by using heat treatment, (NH4)2SO4 fractionation, and chromatography on concanavalin A-Sepharose, DEAE-Sepharose, sulphopropyl-Sephadex, hydroxyapatite, DEAE-cellulose and Sephadex G-100. Enzyme activity was followed by measuring colorimetrically the N-acetylglucosamine released from aspartylglucosamine at 56 degrees C. The purified enzyme protein ran at a 'native' molecular mass of 56 kDa in SDS/12.5%-PAGE gels, and the enzyme activity could be quantitatively recovered at this molecular mass by using gel slices as enzyme source in the assay. After denaturation by boiling in SDS the 56 kDa protein was lost with the corresponding appearance of polypeptides alpha,beta and beta 1, lacking enzyme activity, at 24.6, 18.4 and 17.4 kDa respectively. Treatment of heat-denatured enzyme with N-glycosidase F resulted in the following decreases in molecular mass; 24.6 to 23 kDa and 18.4 and 17.4 to 15.8 kDa. These studies indicate that human liver aspartylglucosaminidase is composed of two non-identical polypeptides, each of which is glycosylated. The N-termini of alpha,beta and beta 1 were directly accessible for sequencing, and the first 21, 26 and 22 amino acids respectively were identified.
Notes: 1470-8728
Rip, J W
Coulter-Mackie, M B
Rupar, C A
Gordon, B A
Journal Article
Biochem J. 1992 Dec 15;288 ( Pt 3)(Pt 3):1005-10. doi: 10.1042/bj2881005.
Author Address: Children's Psychiatric Research Institute, London, ON, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 367
Author: Ripoche, J., Day, A. J., Harris, T. J. and Sim, R. B.
Year: 1988
Title: The complete amino acid sequence of human complement factor H
Journal: Biochem J
Volume: 249
Issue: 2
Pages: 593-602
Epub Date: 1988/01/15
Date: Jan 15
Short Title: The complete amino acid sequence of human complement factor H
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2490593
PMCID: PMC1148743
Accession Number: 2963625
Keywords: Amino Acid Sequence
Base Sequence
*Complement C3b Inactivator Proteins/genetics
Complement Factor H
DNA, Circular/genetics
Humans
Molecular Sequence Data
Abstract: The complete amino acid sequence of the human complement system regulatory protein, factor H, has been derived from sequencing three overlapping cDNA clones. The sequence consists of 1213 amino acids arranged in 20 homologous units, each about 60 amino acids long, and an 18-residue leader sequence. The 60-amino-acid-long repetitive units are homologous with those found in a large number of other complement and non-complement proteins. Two basic C-terminal residues, deduced from the cDNA sequence, are absent from factor H isolated from outdated plasma. A tyrosine/histidine polymorphism was observed within the seventh homologous repeat unit of factor H. This is likely to represent a difference between the two major allelic variants of factor H. The nature of the cDNA clones indicates that there is likely to be an alternative splicing mechanism, resulting in the formation of at least two species of factor H mRNA.
Notes: 1470-8728
Ripoche, J
Day, A J
Harris, T J
Sim, R B
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1988 Jan 15;249(2):593-602. doi: 10.1042/bj2490593.
Author Address: Department of Biochemistry, University of Oxford, U.K.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1671
Author: Risteli, M., Ruotsalainen, H., Bergmann, U., Venkatraman Girija, U., Wallis, R. and Myllylä, R.
Year: 2014
Title: Lysyl hydroxylase 3 modifies lysine residues to facilitate oligomerization of mannan-binding lectin
Journal: PLoS One
Volume: 9
Issue: 11
Pages: e113498
Epub Date: 2014/11/25
Short Title: Lysyl hydroxylase 3 modifies lysine residues to facilitate oligomerization of mannan-binding lectin
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0113498
PMCID: PMC4242627 this manuscript have the following competing interests: US7951562 Method for glycosylating hydroxylysine residues in polypeptides or peptides; and US7341853 Method for glycosylating hydroxylysine residues. This does not alter the authors' adherence to PLOS ONE policies on sharing data and materials.
Accession Number: 25419660
Keywords: Animals
Cells, Cultured
Embryo, Mammalian/cytology
Fibroblasts/cytology/metabolism
Glycosylation
Immunoblotting
Lysine/genetics/*metabolism
Mannose-Binding Lectin/*chemistry/genetics/*metabolism
Mice, Inbred C57BL
Mice, Knockout
Mutation
Peptides/chemistry/metabolism
Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase/genetics/*metabolism
*Protein Multimerization
Rats
Recombinant Proteins/chemistry/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Trypsin/metabolism
Abstract: Lysyl hydroxylase 3 (LH3) is a multifunctional protein with lysyl hydroxylase, galactosyltransferase and glucosyltransferase activities. The LH3 has been shown to modify the lysine residues both in collagens and also in some collagenous proteins. In this study we show for the first time that LH3 is essential for catalyzing formation of the glucosylgalactosylhydroxylysines of mannan-binding lectin (MBL), the first component of the lectin pathway of complement activation. Furthermore, loss of the terminal glucose units on the derivatized lysine residues in mouse embryonic fibroblasts lacking the LH3 protein leads to defective disulphide bonding and oligomerization of rat MBL-A, with a decrease in the proportion of the larger functional MBL oligomers. The oligomerization could be completely restored with the full length LH3 or the amino-terminal fragment of LH3 that possesses the glycosyltransferase activities. Our results confirm that LH3 is the only enzyme capable of glucosylating the galactosylhydroxylysine residues in proteins with a collagenous domain. In mice lacking the lysyl hydroxylase activity of LH3, but with untouched galactosyltransferase and glucosyltransferase activities, reduced circulating MBL-A levels were observed. Oligomerization was normal, however and residual lysyl hydroxylation was compensated in part by other lysyl hydroxylase isoenzymes. Our data suggest that LH3 is commonly involved in biosynthesis of collagenous proteins and the glucosylation of galactosylhydroxylysines residues by LH3 is crucial for the formation of the functional high-molecular weight MBL oligomers.
Notes: 1932-6203
Risteli, Maija
Ruotsalainen, Heli
Bergmann, Ulrich
Venkatraman Girija, Umakhanth
Wallis, Russell
Myllylä, Raili
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2014 Nov 24;9(11):e113498. doi: 10.1371/journal.pone.0113498. eCollection 2014.
Author Address: Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland; Department of Diagnostics and Oral Medicine, Institute of Dentistry, University of Oulu, Oulu, Finland; Medical Research Center Oulu, Oulu University Hospital and University of Oulu, Oulu, Finland.
Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland; Biocenter Oulu, University of Oulu, Oulu, Finland.
Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland; Biocenter Oulu, Mass Spectrometry Core Facility, University of Oulu, Oulu, Finland.
Department of Infection, Immunity, and Inflammation, University of Leicester, Leicester, United Kingdom.
Department of Infection, Immunity, and Inflammation, University of Leicester, Leicester, United Kingdom; Department of Biochemistry, University of Leicester, Leicester, United Kingdom.
Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 390
Author: Ritonja, A., Popović, T., Kotnik, M., Machleidt, W. and Turk, V.
Year: 1988
Title: Amino acid sequences of the human kidney cathepsins H and L
Journal: FEBS Lett
Volume: 228
Issue: 2
Pages: 341-5
Epub Date: 1988/02/15
Date: Feb 15
Short Title: Amino acid sequences of the human kidney cathepsins H and L
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(88)80028-0
Accession Number: 3342889
Keywords: Amino Acid Sequence
Cathepsin B
Cathepsin H
Cathepsin L
*Cathepsins
Cyanogen Bromide
*Cysteine Endopeptidases
*Endopeptidases
Humans
Molecular Sequence Data
Papain
Peptide Fragments
Sequence Homology, Nucleic Acid
Abstract: The complete amino acid sequences of human kidney cathepsin H (EC 3.4.22.16) and human kidney cathepsin L (EC 3.4.22.15) were determined. Cathepsin H contains 230 residues and has an Mr of 25116. The sequence was obtained by sequencing the light, heavy and mini chain and the peptides produced by cyanogen bromide cleavage of the single-chain form of the enzyme. The glycosylated mini chain is a proteolytic fragment of the propeptide of cathepsin H. Human cathepsin L has 217 amino acid residues and an Mr of 23720. Its amino acid sequence was deduced from N-terminal sequences of the heavy and light chains and from the sequences of cyanogen bromide fragments of the heavy chain. The fragments were aligned by comparison with known sequences of cathepsins H and L from other species. Cathepsins H and L exhibit a high degree of sequence homology to cathepsin B (EC 3.4.22.1) and other cysteine proteinases of the papain superfamily.
Notes: Ritonja, A
Popović, T
Kotnik, M
Machleidt, W
Turk, V
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1988 Feb 15;228(2):341-5. doi: 10.1016/0014-5793(88)80028-0.
Author Address: Department of Biochemistry, J. Stefan Institute, Ljubljana, Yugoslavia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 646
Author: Ritter, G., Cohen, L. S., Nice, E. C., Catimel, B., Burgess, A. W., Moritz, R. L., Ji, H., Heath, J. K., White, S. J., Welt, S., Old, L. J. and Simpson, R. J.
Year: 1997
Title: Characterization of posttranslational modifications of human A33 antigen, a novel palmitoylated surface glycoprotein of human gastrointestinal epithelium
Journal: Biochem Biophys Res Commun
Volume: 236
Issue: 3
Pages: 682-6
Epub Date: 1997/07/30
Date: Jul 30
Short Title: Characterization of posttranslational modifications of human A33 antigen, a novel palmitoylated surface glycoprotein of human gastrointestinal epithelium
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1006/bbrc.1997.6966
Accession Number: 9245713
Keywords: Acylation
Amino Acid Sequence
Animals
Antibodies, Monoclonal
Antigens, Neoplasm/*metabolism
Blotting, Western
Colonic Neoplasms/immunology
Digestive System/*immunology
Electrophoresis, Polyacrylamide Gel
Epithelium/immunology
Glycosylation
Humans
Immunosorbent Techniques
Membrane Glycoproteins/*metabolism
Mice
Palmitic Acid/*metabolism
*Protein Processing, Post-Translational
Abstract: Monoclonal antibody (mAb) A33 recognizes a differentiation antigen (A33) expressed in normal human gastrointestinal epithelium and in 95% of human colon cancers. Murine mAb A33 shows specific targeting of colon cancer in humans and a humanized A33 antibody is currently being evaluated in the clinic. The cDNA for the human A33 antigen has recently been cloned, and sequence comparison indicated that the A33 antigen is a novel human cell surface molecule of the immunoglobulin superfamily. Because mAb A33 recognizes a conformational epitope, only a partial characterization of the A33 antigen has been carried out to date. In this report we show that the A33 antigen is (I) N-glycosylated, containing approximately 8 K of N-linked carbohydrate and there is no evidence for O-glycosylation, sialylation or glycophosphatidylinositol, and (ii) S-acylated in vitro, incorporating [3H] palmitic acid linked through a hydroxylamine-sensitive thioester bond. The S-palmitoylation may be involved in regulating the internalization process initiated by binding of mAb A33 to cell surface A33 antigen.
Notes: Ritter, G
Cohen, L S
Nice, E C
Catimel, B
Burgess, A W
Moritz, R L
Ji, H
Heath, J K
White, S J
Welt, S
Old, L J
Simpson, R J
Journal Article
United States
Biochem Biophys Res Commun. 1997 Jul 30;236(3):682-6. doi: 10.1006/bbrc.1997.6966.
Author Address: Ludwig Institute for Cancer Research, New York Branch at Memorial Sloan-Kettering Cancer Center, New York 10021, USA. ritterg@mskcc.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1511
Author: Rivera-Colón, Y., Schutsky, E. K., Kita, A. Z. and Garman, S. C.
Year: 2012
Title: The structure of human GALNS reveals the molecular basis for mucopolysaccharidosis IV A
Journal: J Mol Biol
Volume: 423
Issue: 5
Pages: 736-51
Epub Date: 2012/09/04
Date: Nov 9
Short Title: The structure of human GALNS reveals the molecular basis for mucopolysaccharidosis IV A
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2012.08.020
PMCID: PMC3472114
NIHMSID: NIHMS404526
Accession Number: 22940367
Keywords: Animals
Catalytic Domain
Cell Line
Chondroitinsulfatases/*chemistry/genetics/metabolism
Crystallography, X-Ray
Humans
Insecta
Kinetics
Ligands
Models, Molecular
Mucopolysaccharidosis IV/*genetics
Mutation, Missense
Protein Binding
Protein Conformation
Abstract: Lysosomal enzymes catalyze the breakdown of macromolecules in the cell. In humans, loss of activity of a lysosomal enzyme leads to an inherited metabolic defect known as a lysosomal storage disorder. The human lysosomal enzyme galactosamine-6-sulfatase (GALNS, also known as N-acetylgalactosamine-6-sulfatase and GalN6S; E.C. 3.1.6.4) is deficient in patients with the lysosomal storage disease mucopolysaccharidosis IV A (also known as MPS IV A and Morquio A). Here, we report the three-dimensional structure of human GALNS, determined by X-ray crystallography at 2.2Å resolution. The structure reveals a catalytic gem diol nucleophile derived from modification of a cysteine side chain. The active site of GALNS is a large, positively charged trench suitable for binding polyanionic substrates such as keratan sulfate and chondroitin-6-sulfate. Enzymatic assays on the insect-cell-expressed human GALNS indicate activity against synthetic substrates and inhibition by both substrate and product. Mapping 120 MPS IV A missense mutations onto the structure reveals that a majority of mutations affect the hydrophobic core of the structure, indicating that most MPS IV A cases result from misfolding of GALNS. Comparison of the structure of GALNS to paralogous sulfatases shows a wide variety of active-site geometries in the family but strict conservation of the catalytic machinery. Overall, the structure and the known mutations establish the molecular basis for MPS IV A and for the larger MPS family of diseases.
Notes: 1089-8638
Rivera-Colón, Yadilette
Schutsky, Emily K
Kita, Adriana Z
Garman, Scott C
R01 DK76877/DK/NIDDK NIH HHS/United States
T32 GM008515/GM/NIGMS NIH HHS/United States
P41 GM103403/GM/NIGMS NIH HHS/United States
P41 RR015301/RR/NCRR NIH HHS/United States
R01 DK076877/DK/NIDDK NIH HHS/United States
5P41RR015301-10/RR/NCRR NIH HHS/United States
Howard Hughes Medical Institute/United States
(8 P41 GM103403-10/GM/NIGMS NIH HHS/United States
Y01 GM000080/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Mol Biol. 2012 Nov 9;423(5):736-51. doi: 10.1016/j.jmb.2012.08.020. Epub 2012 Aug 29.
Author Address: Department of Biochemistry and Molecular Biology, University of Massachusetts Amherst, Amherst, MA 01003, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 461
Author: Robb, R. J., Kutny, R. M., Panico, M., Morris, H. R. and Chowdhry, V.
Year: 1984
Title: Amino acid sequence and post-translational modification of human interleukin 2
Journal: Proc Natl Acad Sci U S A
Volume: 81
Issue: 20
Pages: 6486-90
Epub Date: 1984/10/01
Date: Oct
Short Title: Amino acid sequence and post-translational modification of human interleukin 2
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.81.20.6486
PMCID: PMC391949
Accession Number: 6333684
Keywords: Amino Acid Sequence
Disulfides
Glycoproteins/analysis
Humans
*Interleukin-2/metabolism
Protein Processing, Post-Translational
Abstract: Human interleukin 2 was separated into multiple molecular forms by selective immunoaffinity chromatography and chromatofocusing. For the most part, this heterogeneity was attributed to variations in glycosylation of the threonine residue in position 3 of the polypeptide chain. The various molecular forms of interleukin 2 had nearly identical specific activities in the in vitro proliferation assay, indicating that the glycosylation had no significant effect on this response. The entire primary sequence of interleukin 2, including the location of the intramolecular disulfide bridge, was determined by a combination of peptide mapping and protein sequencing. This information should aid in the determination of the active site(s) of the molecule.
Notes: 1091-6490
Robb, R J
Kutny, R M
Panico, M
Morris, H R
Chowdhry, V
Journal Article
Proc Natl Acad Sci U S A. 1984 Oct;81(20):6486-90. doi: 10.1073/pnas.81.20.6486.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 247
Author: Robertson, D. A., Freeman, C., Morris, C. P. and Hopwood, J. J.
Year: 1992
Title: A cDNA clone for human glucosamine-6-sulphatase reveals differences between arylsulphatases and non-arylsulphatases
Journal: Biochem J
Volume: 288 ( Pt 2)
Issue: Pt 2
Pages: 539-44
Epub Date: 1992/12/01
Date: Dec 1
Short Title: A cDNA clone for human glucosamine-6-sulphatase reveals differences between arylsulphatases and non-arylsulphatases
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2880539
PMCID: PMC1132044
Accession Number: 1463457
Keywords: Amino Acid Sequence
Arylsulfatases/chemistry
Base Sequence
Cloning, Molecular
DNA/genetics
Humans
Liver/enzymology
Molecular Sequence Data
Molecular Weight
Oligodeoxyribonucleotides/chemistry
Sequence Alignment
Structure-Activity Relationship
Substrate Specificity
Sulfatases/chemistry/*genetics/metabolism
Abstract: Glucosamine-6-sulphatase is an exo-hydrolase required for the lysosomal degradation of heparan sulphate and keratan sulphate. Deficiency of glucosamine-6-sulphatase activity leads to the lysosomal storage of the glycosaminoglycan, heparan sulphate and the monosaccharide sulphate N-acetylglucosamine 6-sulphate and the autosomal recessive genetic disorder mucopolysaccharidosis type IIID. Glucosamine-6-sulphatase can be classified as a non-arylsulphatase since, relative to arylsulphatase B, it shows negligible activity toward 4-methylumbelliferyl sulphate. We have isolated human cDNA clones and derived amino acid sequence coding for the entire glucosamine-6-sulphatase protein. The predicted sequence has 552 amino acids with a leader peptide of 36 amino acids and contains 13 potential N-glycosylation sites, of which it is likely that 10 are used. Glucosamine-6-sulphatase shows strong sequence similarity to other sulphatases such as the family of arylsulphatases, although the degree of similarity is not as high as that between members of the arylsulphatase family. This pattern of inter- and intra-family similarity delineates regions and amino acid residues that may be critical for sulphatase function and substrate specificity.
Notes: 1470-8728
Robertson, D A
Freeman, C
Morris, C P
Hopwood, J J
Comparative Study
Journal Article
Biochem J. 1992 Dec 1;288 ( Pt 2)(Pt 2):539-44. doi: 10.1042/bj2880539.
Author Address: Department of Chemical Pathology, Adelaide Children's Hospital, South Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 476
Author: Robinson, E. A. and Appella, E.
Year: 1980
Title: Complete amino acid sequence of a mouse immunoglobulin alpha chain (MOPC 511)
Journal: Proc Natl Acad Sci U S A
Volume: 77
Issue: 8
Pages: 4909-13
Epub Date: 1980/08/01
Date: Aug
Short Title: Complete amino acid sequence of a mouse immunoglobulin alpha chain (MOPC 511)
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.77.8.4909
PMCID: PMC349958
Accession Number: 6776528
Keywords: Amino Acid Sequence
Animals
Glycoproteins
*Immunoglobulin Heavy Chains/analysis
*Immunoglobulin alpha-Chains/analysis/genetics
Mice
Mutation
*Myeloma Proteins
Oligosaccharides/analysis
Abstract: The complete amino acid sequence of the 432-residue heavy (alpha) chain of mouse myeloma MOPC 511 has been determined. The variable region of the alpha chain of IgA 511, a phosphocholine-binding protein, is highly homologous to that of the other phosphocholine-binding immunoglobulins. Comparison of the 511 alpha chain constant region with that of other mouse and human heavy chains shows that sequence divain. The CH3 domain disulfide bridge of the 511 alpha chain, for example, consists of only 28 amino acid residues compared to 60 residues for other chains and domains. Sequence divergences are alsos apparent at the CH2/CH3 domain boundary, an area where a number of frameshift mutations have occurred. One mutant, mouse IgA 47A, lacks the entire CH3 domain. Comparison of the 511 alppha chain with the 47A alpha chain reveals two noncconservative amino acid changes at the COOH terminus of the 47A chain, Ser-Gln for VAl-Thr in the 511 chain. These changes and the deletion of the CH3 domain can be explained by a single genetic event--namely, a frameshift mutation followed by premature chain termination. The remainder of the 47A constant region, including the hinge region, is identical to the 511 alpha chain, except for two conservative changes in the CH1 domain: serine-126 and theonine-197 in the 511 alpha chain are both replaced by alanine in the 47A chain.
Notes: 1091-6490
Robinson, E A
Appella, E
Journal Article
Proc Natl Acad Sci U S A. 1980 Aug;77(8):4909-13. doi: 10.1073/pnas.77.8.4909.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1887
Author: Roig-Zamboni, V., Cobucci-Ponzano, B., Iacono, R., Ferrara, M. C., Germany, S., Bourne, Y., Parenti, G., Moracci, M. and Sulzenbacher, G.
Year: 2017
Title: Structure of human lysosomal acid α-glucosidase-a guide for the treatment of Pompe disease
Journal: Nat Commun
Volume: 8
Issue: 1
Pages: 1111
Epub Date: 2017/10/25
Date: Oct 24
Short Title: Structure of human lysosomal acid α-glucosidase-a guide for the treatment of Pompe disease
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-017-01263-3
PMCID: PMC5653652
Accession Number: 29061980
Keywords: Acetylcysteine/chemistry/metabolism
Catalytic Domain
Glycogen Storage Disease Type II/*enzymology/genetics
Humans
Lysosomes/chemistry/enzymology/genetics
Models, Molecular
Protein Conformation
alpha-Glucosidases/*chemistry/genetics/metabolism
Abstract: Pompe disease, a rare lysosomal storage disease caused by deficiency of the lysosomal acid α-glucosidase (GAA), is characterized by glycogen accumulation, triggering severe secondary cellular damage and resulting in progressive motor handicap and premature death. Numerous disease-causing mutations in the gaa gene have been reported, but the structural effects of the pathological variants were unknown. Here we present the high-resolution crystal structures of recombinant human GAA (rhGAA), the standard care of Pompe disease. These structures portray the unbound form of rhGAA and complexes thereof with active site-directed inhibitors, providing insight into substrate recognition and the molecular framework for the rationalization of the deleterious effects of disease-causing mutations. Furthermore, we report the structure of rhGAA in complex with the allosteric pharmacological chaperone N-acetylcysteine, which reveals the stabilizing function of this chaperone at the structural level.
Notes: 2041-1723
Roig-Zamboni, Véronique
Cobucci-Ponzano, Beatrice
Orcid: 0000-0002-8211-2297
Iacono, Roberta
Orcid: 0000-0002-3586-4322
Ferrara, Maria Carmina
Orcid: 0000-0001-8905-2488
Germany, Stanley
Bourne, Yves
Parenti, Giancarlo
Moracci, Marco
Orcid: 0000-0002-9846-2531
Sulzenbacher, Gerlind
Orcid: 0000-0003-4844-2820
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2017 Oct 24;8(1):1111. doi: 10.1038/s41467-017-01263-3.
Author Address: Centre National de la Recherche Scientifique (CNRS), Aix-Marseille Univ, AFMB, 163 Avenue de Luminy, 13288, Marseille, France.
Institute of Biosciences and Bioresources, National Research Council of Italy, Via P. Castellino 111, 80131, Naples, Italy.
Telethon Institute of Genetics and Medicine (TIGEM), Via Campi Flegrei 34, Pozzuoli, 80078, Naples, Italy.
Department of Translational Medical Sciences, Federico II University, Via Pansini 5, 80131, Naples, Italy.
Department of Biology, Federico II University, Complesso Universitario di Monte S. Angelo, Via Cintia 21, 80126, Naples, Italy.
Centre National de la Recherche Scientifique (CNRS), Aix-Marseille Univ, AFMB, 163 Avenue de Luminy, 13288, Marseille, France. Gerlind.Sulzenbacher@afmb.univ-mrs.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1944
Author: Rong, Y., Bansal, P. K., Wei, P., Guo, H., Correia, K., Parris, J. and Morgan, J. I.
Year: 2018
Title: Glycosylation of Cblns attenuates their receptor binding
Journal: Brain Res
Volume: 1694
Pages: 129-139
Epub Date: 2018/05/22
Date: Sep 1
Short Title: Glycosylation of Cblns attenuates their receptor binding
Alternate Journal: Brain research
ISSN: 0006-8993 (Print)
0006-8993
DOI: 10.1016/j.brainres.2018.05.022
PMCID: PMC6026032
NIHMSID: NIHMS971627
Accession Number: 29782851
Keywords: Animals
Cells, Cultured
Cerebellum/*metabolism
*Glycosylation
Mice, Inbred C57BL
Mutation/genetics
Nerve Tissue Proteins/genetics/*metabolism
Neurons/metabolism
Protein Precursors/genetics/*metabolism
Synapses/*physiology
*Locomotor activity
*S4 domain
*Transgenic
Abstract: Cbln1 is the prototype of a family (Cbln1-Cbln4) of secreted glycoproteins and is essential for normal synapse structure and function in cerebellum by bridging presynaptic Nrxn to postsynaptic Grid2. Here we report the effects of glycosylation on the in vitro receptor binding properties of Cblns. Cbln1, 2 and 4 harbor two N-linked glycosylation sites, one at the N-terminus is in a region implicated in Nrxn binding and the second is in the C1q domain, a region involved in Grid2 binding. Mutation (asparagine to glutamine) of the N-terminal site, increased neurexin binding whereas mutation of the C1q site markedly increased Grid2 binding. These mutations did not influence subunit composition of Cbln trimeric complexes (mediated through the C1q domain) nor their assembly into hexamers (mediated by the N-terminal region). Therefore, glycosylation likely masks the receptor binding interfaces of Cblns. As Cbln4 has undetectable Grid2 binding in vitro we assessed whether transgenic expression of wild type Cbln4 or its glycosylation mutants rescued the Cbln1-null phenotype in vivo. Cbln4 partially rescued and both glycosylation mutants completely rescued ataxia in cbln1-null mice. Thus Cbln4 has intrinsic Grid2 binding that is attenuated by glycosylation, and glycosylation mutants exhibit gain of function in vivo.
Notes: 1872-6240
Rong, Yongqi
Bansal, Parmil K
Wei, Peng
Guo, Hong
Correia, Kristen
Parris, Jennifer
Morgan, James I
P30 CA021765/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Brain Res. 2018 Sep 1;1694:129-139. doi: 10.1016/j.brainres.2018.05.022. Epub 2018 May 18.
Author Address: Department of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
Department of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA. Electronic address: jim.morgan@stjude.org.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1730
Author: Rønningen, T., Shah, A., Oldenburg, A. R., Vekterud, K., Delbarre, E., Moskaug, JØ and Collas, P.
Year: 2015
Title: Prepatterning of differentiation-driven nuclear lamin A/C-associated chromatin domains by GlcNAcylated histone H2B
Journal: Genome Res
Volume: 25
Issue: 12
Pages: 1825-35
Epub Date: 2015/09/12
Date: Dec
Short Title: Prepatterning of differentiation-driven nuclear lamin A/C-associated chromatin domains by GlcNAcylated histone H2B
Alternate Journal: Genome research
ISSN: 1088-9051 (Print)
1088-9051
DOI: 10.1101/gr.193748.115
PMCID: PMC4665004
Accession Number: 26359231
Keywords: Acetylation
Adipogenesis
*Cell Differentiation
Chromatin/genetics/*metabolism
Chromatin Assembly and Disassembly
Chromatin Immunoprecipitation
Gene Expression Regulation
Glycolysis/genetics
Glycosylation
High-Throughput Nucleotide Sequencing
Histones/chemistry/*metabolism
Humans
Lamin Type A/*metabolism
Protein Binding
Protein Interaction Domains and Motifs
Abstract: Dynamic interactions of nuclear lamins with chromatin through lamin-associated domains (LADs) contribute to spatial arrangement of the genome. Here, we provide evidence for prepatterning of differentiation-driven formation of lamin A/C LADs by domains of histone H2B modified on serine 112 by the nutrient sensor O-linked N-acetylglucosamine (H2BS112GlcNAc), which we term GADs. We demonstrate a two-step process of lamin A/C LAD formation during in vitro adipogenesis, involving spreading of lamin A/C-chromatin interactions in the transition from progenitor cell proliferation to cell-cycle arrest, and genome-scale redistribution of these interactions through a process of LAD exchange within hours of adipogenic induction. Lamin A/C LADs are found both in active and repressive chromatin contexts that can be influenced by cell differentiation status. De novo formation of adipogenic lamin A/C LADs occurs nonrandomly on GADs, which consist of megabase-size intergenic and repressive chromatin domains. Accordingly, whereas predifferentiation lamin A/C LADs are gene-rich, post-differentiation LADs harbor repressive features reminiscent of lamin B1 LADs. Release of lamin A/C from genes directly involved in glycolysis concurs with their transcriptional up-regulation after adipogenic induction, and with downstream elevations in H2BS112GlcNAc levels and O-GlcNAc cycling. Our results unveil an epigenetic prepatterning of adipogenic LADs by GADs, suggesting a coupling of developmentally regulated lamin A/C-genome interactions to a metabolically sensitive chromatin modification.
Notes: 1549-5469
Rønningen, Torunn
Shah, Akshay
Oldenburg, Anja R
Vekterud, Kristin
Delbarre, Erwan
Moskaug, Jan Øivind
Collas, Philippe
Journal Article
Research Support, Non-U.S. Gov't
Genome Res. 2015 Dec;25(12):1825-35. doi: 10.1101/gr.193748.115. Epub 2015 Sep 10.
Author Address: Department of Molecular Medicine, Institute of Basic Medical Sciences, Faculty of Medicine, University of Oslo, 0317 Oslo, Norway;
Department of Molecular Medicine, Institute of Basic Medical Sciences, Faculty of Medicine, University of Oslo, 0317 Oslo, Norway; Norwegian Center for Stem Cell Research, Oslo University Hospital, 0317 Oslo, Norway.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 652
Author: Roos, M. D., Su, K., Baker, J. R. and Kudlow, J. E.
Year: 1997
Title: O glycosylation of an Sp1-derived peptide blocks known Sp1 protein interactions
Journal: Mol Cell Biol
Volume: 17
Issue: 11
Pages: 6472-80
Epub Date: 1997/10/29
Date: Nov
Short Title: O glycosylation of an Sp1-derived peptide blocks known Sp1 protein interactions
Alternate Journal: Molecular and cellular biology
ISSN: 0270-7306 (Print)
0270-7306
DOI: 10.1128/mcb.17.11.6472
PMCID: PMC232500
Accession Number: 9343410
Keywords: Amino Acid Sequence
Chromatography, High Pressure Liquid
DNA-Binding Proteins/*metabolism
Genes, Reporter
Glycopeptides/genetics/isolation & purification/*metabolism
Glycosylation
HeLa Cells
Humans
Models, Genetic
Molecular Sequence Data
Mutation
Peptide Fragments/genetics/isolation & purification/*metabolism
Protein Binding/drug effects
Recombinant Proteins/metabolism
Sequence Analysis
Sp1 Transcription Factor/*metabolism
*TATA-Binding Protein Associated Factors
Trans-Activators/*metabolism
*Transcription Factor TFIID
Transcription, Genetic
Abstract: The O-linked N-acetylglucosamine (O-GlcNAc) modification of proteins is dynamic and abundant in the nucleus and cytosol. Several transcription factors, including Sp1, have been shown to contain this modification; however, the functional role of O-GlcNAc in these proteins has not been determined. In this paper we describe the use of the previously characterized glutamine-rich transactivation domain of Sp1 (B-c) as a model to investigate the role of O-GlcNAc in Sp1's transcriptionally relevant protein-to-protein interactions with the TATA-binding-protein-associated factor (TAF110) and holo-Sp1. When the model Sp1 peptide was overexpressed in primate cells, this 97-amino-acid domain of Sp1 was found to contain a dominant O-GlcNAc residue at high stoichiometry, which allowed the mapping and mutagenesis of this glycosylation site. In vitro interaction studies between this segment of Sp1 and Drosophila TAF110 or holo-Sp1 indicate that the O-GlcNAc modification functions to inhibit the largely hydrophobic interactions between these proteins. In HeLa cells, the mutation at the mapped glycosylation site was permissive for transcriptional activation. We propose the hypothesis that the removal of O-GlcNAc from an interaction domain can be a signal for protein association. O-GlcNAc may thereby prevent untimely and ectopic interactions.
Notes: 1098-5549
Roos, M D
Su, K
Baker, J R
Kudlow, J E
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Biol. 1997 Nov;17(11):6472-80. doi: 10.1128/MCB.17.11.6472.
Author Address: Department of Cell Biology, University of Alabama at Birmingham, 35294, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 609
Author: Roquemore, E. P., Chevrier, M. R., Cotter, R. J. and Hart, G. W.
Year: 1996
Title: Dynamic O-GlcNAcylation of the small heat shock protein alpha B-crystallin
Journal: Biochemistry
Volume: 35
Issue: 11
Pages: 3578-86
Epub Date: 1996/03/19
Date: Mar 19
Short Title: Dynamic O-GlcNAcylation of the small heat shock protein alpha B-crystallin
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi951918j
Accession Number: 8639509
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Astrocytes/metabolism
Crystallins/*metabolism
Electrophoresis, Gel, Two-Dimensional
Heat-Shock Proteins/*metabolism
Humans
Lens, Crystalline/metabolism
Macaca mulatta
Molecular Sequence Data
Myocardium/metabolism
Peptide Fragments/chemistry
Protein Processing, Post-Translational
Rats
Rats, Sprague-Dawley
Threonine/chemistry
Tumor Cells, Cultured
Abstract: alphaB-Crystallin, originally described as a structural lens protein, is now known to be a member of the small heat shock protein family and is expressed in a number of nonlens tissues. This highly conserved 20 kDa protein aggregates with homologous proteins, including alphaA-crystallin and the small heat shock protein HSP28, to form large heteromeric complexes. Recently, Roquemore et al. (1992) have established that both phosphorylated and unphosphorylated forms of lens alphaB-crystallin are modified with O-linked N-acetylglucosamine, a dynamic posttranslational modification abundant on nuclear and cytoplasmic proteins. In this paper, we have identified the major site of O-GlcNAcylation on lens alphaB as Thr 170. We have further shown that this modification is not restricted to lens alphaB-crystallin but occurs on alphaB isolated from rat heart tissue and human astroglioma cells. Two-dimensional electrophoresis of rat heart alphaB-crystallin revealed two O-GlcNAcylated forms with mobilities corresponding to the unphosphorylated form (alphaB2) and an unidentified, slightly more acidic form. Phosphorylated alphaB-crystallin (alphaB1) was not detected in the rat heart preparation. The major O-GlcNAcylation site on alphaB-crystallins from rat heart also appears to be at Thr 170. Metabolic pulse-chase labeling studies of U373-MG astroglioma cells indicated that turnover of the carbohydrate on alphaB-crystallin is not static but proceeds many-fold more rapidly than turnover of the protein backbone itself, consistent with a regulatory role for O-GlcNAc on this small heat shock protein.
Notes: Roquemore, E P
Chevrier, M R
Cotter, R J
Hart, G W
5T3 GM07445/GM/NIGMS NIH HHS/United States
R01 CA-42486/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1996 Mar 19;35(11):3578-86. doi: 10.1021/bi951918j.
Author Address: Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham 35294-0005, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 277
Author: Roquemore, E. P., Dell, A., Morris, H. R., Panico, M., Reason, A. J., Savoy, L. A., Wistow, G. J., Zigler, J. S., Jr., Earles, B. J. and Hart, G. W.
Year: 1992
Title: Vertebrate lens alpha-crystallins are modified by O-linked N-acetylglucosamine
Journal: J Biol Chem
Volume: 267
Issue: 1
Pages: 555-63
Epub Date: 1992/01/15
Date: Jan 5
Short Title: Vertebrate lens alpha-crystallins are modified by O-linked N-acetylglucosamine
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1730617
Keywords: Acetylglucosamine/*metabolism
Alkaline Phosphatase/chemistry
Amino Acid Sequence
Animals
Binding Sites
Birds
Blotting, Western
Cattle
Chromatography, High Pressure Liquid
Crystallins/isolation & purification/*metabolism
Cyanogen Bromide/chemistry
Electrophoresis, Polyacrylamide Gel
Humans
Isoelectric Focusing
Macaca mulatta
Molecular Sequence Data
Precipitin Tests
Rats
Serine/metabolism
Spectrometry, Mass, Fast Atom Bombardment
Trypsin/chemistry
Abstract: Crystallins are structural proteins responsible for establishing the remarkable optical properties of the lens. Yet many of these highly conserved proteins are also expressed in nonocular tissues, where they have alternative functions apparently unrelated to their structural role in the lens. Here we report that lens alpha-crystallins, some of which function as heat-shock proteins in other tissues, are modified with O-linked N-acetylglucosamine (O-GlcNAc). An in vitro enzymatic assay that transfers [3H]Gal to terminal GlcNAc moieties labels alpha A and alpha B crystallins in lens homogenates from man, rhesus monkey, rat, cow, and rhea (an ostrich-like bird). O-Linkage of the saccharide is demonstrated by sensitivity to base-catalyzed beta-elimination and resistance to peptide:N-glycosidase F treatment. Chromatographic analyses of the beta-elimination products and fast atom bombardment-mass spectrometry of [3H]Gal-labeled tryptic peptides confirm the saccharide structure. Isoelectric focusing of [3H]Gal-labeled bovine lens proteins reveals the presence of O-GlcNAc on all four alpha-crystallin subunits, A1, A2, B1, and B2. Electrospray mass spectrometry of bovine alpha-crystallin demonstrates the presence of a single O-GlcNAc substitution on alpha A2. Gas-phase protein sequencing and fast atom bombardment-mass spectrometry of the major radiolabeled tryptic peptide from bovine alpha-crystallin reveal that GlcNAc is attached to the alpha A subunits at serine 162. This post-translational modification may play an important role in the molecular organization of lens alpha-crystallin.
Notes: Roquemore, E P
Dell, A
Morris, H R
Panico, M
Reason, A J
Savoy, L A
Wistow, G J
Zigler, J S Jr
Earles, B J
Hart, G W
CA42486/CA/NCI NIH HHS/United States
HD13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Jan 5;267(1):555-63.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1380
Author: Rossmann, M., Sukumaran, M., Penn, A. C., Veprintsev, D. B., Babu, M. M. and Greger, I. H.
Year: 2011
Title: Subunit-selective N-terminal domain associations organize the formation of AMPA receptor heteromers
Journal: Embo j
Volume: 30
Issue: 5
Pages: 959-71
Epub Date: 2011/02/15
Date: Mar 2
Short Title: Subunit-selective N-terminal domain associations organize the formation of AMPA receptor heteromers
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2011.16
PMCID: PMC3049212
Accession Number: 21317873
Keywords: Calcium/*metabolism
Cell Membrane/*metabolism
Crystallography, X-Ray
Electrophysiology
Humans
Ion Channels
Protein Conformation
Protein Multimerization
Protein Subunits
Protein Transport
Receptors, AMPA/*chemistry/*metabolism
Synapses
Ultracentrifugation
alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid/*metabolism
Abstract: The assembly of AMPA-type glutamate receptors (AMPARs) into distinct ion channel tetramers ultimately governs the nature of information transfer at excitatory synapses. How cells regulate the formation of diverse homo- and heteromeric AMPARs is unknown. Using a sensitive biophysical approach, we show that the extracellular, membrane-distal AMPAR N-terminal domains (NTDs) orchestrate selective routes of heteromeric assembly via a surprisingly wide spectrum of subunit-specific association affinities. Heteromerization is dominant, occurs at the level of the dimer, and results in a preferential incorporation of the functionally critical GluA2 subunit. Using a combination of structure-guided mutagenesis and electrophysiology, we further map evolutionarily variable hotspots in the NTD dimer interface, which modulate heteromerization capacity. This 'flexibility' of the NTD not only explains why heteromers predominate but also how GluA2-lacking, Ca(2+)-permeable homomers could form, which are induced under specific physiological and pathological conditions. Our findings reveal that distinct NTD properties set the stage for the biogenesis of functionally diverse pools of homo- and heteromeric AMPAR tetramers.
Notes: 1460-2075
Rossmann, Maxim
Sukumaran, Madhav
Penn, Andrew C
Veprintsev, Dmitry B
Babu, M Madan
Greger, Ingo H
MC_U105174197/Medical Research Council/United Kingdom
MC_U105185859/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2011 Mar 2;30(5):959-71. doi: 10.1038/emboj.2011.16. Epub 2011 Feb 11.
Author Address: Neurobiology Division, MRC Laboratory of Molecular Biology, Cambridge, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1843
Author: Roth, C., Chan, S., Offen, W. A., Hemsworth, G. R., Willems, L. I., King, D. T., Varghese, V., Britton, R., Vocadlo, D. J. and Davies, G. J.
Year: 2017
Title: Structural and functional insight into human O-GlcNAcase
Journal: Nat Chem Biol
Volume: 13
Issue: 6
Pages: 610-612
Epub Date: 2017/03/28
Date: Jun
Short Title: Structural and functional insight into human O-GlcNAcase
Alternate Journal: Nature chemical biology
ISSN: 1552-4450 (Print)
1552-4450
DOI: 10.1038/nchembio.2358
PMCID: PMC5438047
NIHMSID: EMS71832 company Alectos Therapeutics. D.J.V. serves as CSO and Chair of the Scientific Advisory Board of Alectos Therapeutics of which G.J.D is a member.
Accession Number: 28346405
Keywords: Acetylglucosamine/metabolism
Binding Sites
Enzyme Activation/drug effects
Enzyme Inhibitors/pharmacology
HEK293 Cells
Humans
Ligands
*Models, Molecular
Protein Binding
Protein Isoforms/chemistry/genetics
Protein Structure, Tertiary
beta-N-Acetylhexosaminidases/*chemistry/genetics/metabolism
Abstract: O-GlcNAc hydrolase (OGA) removes O-linked N-acetylglucosamine (O-GlcNAc) from a myriad of nucleocytoplasmic proteins. Through co-expression and assembly of OGA fragments, we determined the three-dimensional structure of human OGA, revealing an unusual helix-exchanged dimer that lays a structural foundation for an improved understanding of substrate recognition and regulation of OGA. Structures of OGA in complex with a series of inhibitors define a precise blueprint for the design of inhibitors that have clinical value.
Notes: 1552-4469
Roth, Christian
Orcid: 0000-0001-5806-0987
Chan, Sherry
Offen, Wendy A
Hemsworth, Glyn R
Orcid: 0000-0002-8226-1380
Willems, Lianne I
King, Dustin T
Varghese, Vimal
Britton, Robert
Vocadlo, David J
Orcid: 0000-0001-6897-5558
Davies, Gideon J
Orcid: 0000-0002-7343-776x
BB/K003836/1/Biotechnology and Biological Sciences Research Council/United Kingdom
MOP-123341/CIHR/Canada
Journal Article
Research Support, Non-U.S. Gov't
Nat Chem Biol. 2017 Jun;13(6):610-612. doi: 10.1038/nchembio.2358. Epub 2017 Mar 27.
Author Address: York Structural Biology Laboratory, Department of Chemistry University of York, York, UK.
Department of Chemistry, Simon Fraser University, Burnaby, British Columbia, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1818
Author: Roth, S. and Khalaila, I.
Year: 2017
Title: The effect of O-GlcNAcylation on hnRNP A1 translocation and interaction with transportin1
Journal: Exp Cell Res
Volume: 350
Issue: 1
Pages: 210-217
Epub Date: 2016/12/04
Date: Jan 1
Short Title: The effect of O-GlcNAcylation on hnRNP A1 translocation and interaction with transportin1
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2016.11.023
Accession Number: 27913144
Keywords: Acetylation
Cell Nucleus/*metabolism
Cells, Cultured
Cytoplasm/*metabolism
Heterogeneous Nuclear Ribonucleoprotein A1
Heterogeneous-Nuclear Ribonucleoprotein Group A-B/*metabolism
Humans
Phosphorylation
Protein Transport
RNA Precursors/metabolism
beta Karyopherins/*metabolism
*O-GlcNAcylation
*Protein-protein interaction
*Translocation
*Trn1
*hnRNP A1
Abstract: The heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) is a major pre-mRNA binding protein involved in transcription and translation. Although predominantly nuclear, hnRNP A1 shuttles rapidly between the nucleus and the cytosol, delivering its anchored pre-mRNA for further processing. Translocation is important for hnRNP A1 to accomplish its transcriptional and translational roles. Transportin1 (Trn1), a translocation protein, facilitates the translocation of hnRNP A1 back to the nucleus. Moreover, phosphorylation of serine residues at hnRNP A1 C-terminal domain affects its translocation. In this study, we found that phosphorylation is not the only modification that hnRNP A1 undergoes, but also O-linked N-acetylglucosaminylation (O-GlcNAcylation) could occur. Several putative novel O-GlcNAcylation and phosphorylation sites in hnRNP A1 were mapped. Whereas enhanced O-GlcNAcylation increased hnRNP A1 interaction with Trn1, enhanced phosphorylation reduced the interaction between the proteins. In addition, elevated O-GlcNAcylation resulted in hnRNP A1 seclusion in the nucleus, whereas elevated phosphorylation resulted in its accumulation in the cytosol. These findings suggest that a new player, i.e., O-GlcNAcylation, regulates hnRNP A1 translocation and interaction with Trn1, possibly affecting its function. There is a need for further study, to elucidate the role of O-GlcNAcylation in the regulation of the specific activities of hnRNP A1 in transcription and translation.
Notes: 1090-2422
Roth, Shira
Khalaila, Isam
Journal Article
United States
Exp Cell Res. 2017 Jan 1;350(1):210-217. doi: 10.1016/j.yexcr.2016.11.023. Epub 2016 Nov 29.
Author Address: Avram and Stella Goldstein-Goren Department of Biotechnology Engineering, Ben-Gurion University of the Negev, P.O.Box 653, Beer-Sheva 84105, Israel.
Avram and Stella Goldstein-Goren Department of Biotechnology Engineering, Ben-Gurion University of the Negev, P.O.Box 653, Beer-Sheva 84105, Israel. Electronic address: isam@bgu.ac.il.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 797
Author: Rotonda, J., Garcia-Calvo, M., Bull, H. G., Geissler, W. M., McKeever, B. M., Willoughby, C. A., Thornberry, N. A. and Becker, J. W.
Year: 2001
Title: The three-dimensional structure of human granzyme B compared to caspase-3, key mediators of cell death with cleavage specificity for aspartic acid in P1
Journal: Chem Biol
Volume: 8
Issue: 4
Pages: 357-68
Epub Date: 2001/04/28
Date: Apr
Short Title: The three-dimensional structure of human granzyme B compared to caspase-3, key mediators of cell death with cleavage specificity for aspartic acid in P1
Alternate Journal: Chemistry & biology
ISSN: 1074-5521 (Print)
1074-5521
DOI: 10.1016/s1074-5521(01)00018-7
Accession Number: 11325591
Keywords: Amino Acid Sequence
Animals
*Apoptosis
Aspartic Acid/*metabolism
Binding Sites
Caspase 3
Caspase Inhibitors
Caspases/*chemistry/*metabolism
Computational Biology
Crystallography, X-Ray
Enzyme Inhibitors/metabolism/pharmacology
Granzymes
Humans
Hydrogen Bonding
Mice
Models, Molecular
Molecular Sequence Data
Protein Structure, Secondary
Protein Structure, Tertiary
Rats
Sequence Alignment
Sequence Homology, Amino Acid
Serine Endopeptidases/*chemistry/*metabolism
Static Electricity
Substrate Specificity
Abstract: BACKGROUND: Granzyme B, one of the most abundant granzymes in cytotoxic T-lymphocyte (CTL) granules, and members of the caspase (cysteine aspartyl proteinases) family have a unique cleavage specificity for aspartic acid in P1 and play critical roles in the biochemical events that culminate in cell death. RESULTS: We have determined the three-dimensional structure of the complex of the human granzyme B with a potent tetrapeptide aldehyde inhibitor. The Asp-specific S1 subsite of human granzyme B is significantly larger and less charged than the corresponding Asp-specific site in the apoptosis-promoting caspases, and also larger than the corresponding subsite in rat granzyme B. CONCLUSIONS: The above differences account for the variation in substrate specificity among granzyme B, other serine proteases and the caspases, and enable the design of specific inhibitors that can probe the physiological functions of these proteins and the disease states with which they are associated.
Notes: Rotonda, J
Garcia-Calvo, M
Bull, H G
Geissler, W M
McKeever, B M
Willoughby, C A
Thornberry, N A
Becker, J W
Comparative Study
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
United States
Chem Biol. 2001 Apr;8(4):357-68. doi: 10.1016/s1074-5521(01)00018-7.
Author Address: Department of Endocrinology and Chemical Biology, Merck Research Laboratories, Rahway, NJ 07065-0900, USA. rotonda@merck.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 351
Author: Roughley, P. J. and White, R. J.
Year: 1989
Title: Dermatan sulphate proteoglycans of human articular cartilage. The properties of dermatan sulphate proteoglycans I and II
Journal: Biochem J
Volume: 262
Issue: 3
Pages: 823-7
Epub Date: 1989/09/15
Date: Sep 15
Short Title: Dermatan sulphate proteoglycans of human articular cartilage. The properties of dermatan sulphate proteoglycans I and II
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2620823
PMCID: PMC1133347
Accession Number: 2590169
Keywords: Amino Acid Sequence
Amino Acids/analysis
Cartilage, Articular/*analysis
Chondroitin Sulfate Proteoglycans/*analysis
Dermatan Sulfate
Electrophoresis, Polyacrylamide Gel
Humans
Infant
Molecular Sequence Data
Molecular Weight
Proteoglycans/*analysis
Abstract: Dermatan sulphate proteoglycans were purified from juvenile human articular cartilage, with a yield of about 2 mg/g wet wt. of cartilage. Both dermatan sulphate proteoglycan I (DS-PGI) and dermatan sulphate proteoglycan II (DS-PGII) were identified and the former was present in greater abundance. The two proteoglycans could not be resolved by agarose/polyacrylamide-gel electrophoresis, but could be resolved by SDS/polyacrylamide-gel electrophoresis, which indicated average Mr values of 200,000 and 98,000 for DS-PGI and DS-PGII respectively. After digestion with chondroitin ABC lyase the Mr values of the core proteins were 44,000 for DS-PGI and 43,000 and 47,000 for DS-PGII, with the smaller core protein being predominant in DS-PGII. Sequence analysis of the N-terminal 20 amino acid residues reveals the presence of a single site for the potential substitution of dermatan sulphate at residue 4 of DS-PGII and two such sites at residues 5 and 10 for DS-PGI.
Notes: 1470-8728
Roughley, P J
White, R J
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1989 Sep 15;262(3):823-7. doi: 10.1042/bj2620823.
Author Address: Joint Diseases Laboratory, Shriners Hospital for Crippled Children, Montreal, Quebec, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 718
Author: Roussel, A., Canaan, S., Egloff, M. P., Rivière, M., Dupuis, L., Verger, R. and Cambillau, C.
Year: 1999
Title: Crystal structure of human gastric lipase and model of lysosomal acid lipase, two lipolytic enzymes of medical interest
Journal: J Biol Chem
Volume: 274
Issue: 24
Pages: 16995-7002
Epub Date: 1999/06/08
Date: Jun 11
Short Title: Crystal structure of human gastric lipase and model of lysosomal acid lipase, two lipolytic enzymes of medical interest
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.274.24.16995
Accession Number: 10358049
Keywords: Amino Acid Sequence
Carboxypeptidases/chemistry
Catalytic Domain
Cathepsin A
Computer Simulation
Crystallography, X-Ray
Humans
Lipase/antagonists & inhibitors/*chemistry/genetics
Lipolysis
Lysosomes/*enzymology
Models, Molecular
Molecular Sequence Data
Organophosphonates/pharmacology
Recombinant Proteins/chemistry
Sequence Homology, Amino Acid
Stomach/*enzymology
Abstract: Fat digestion in humans requires not only the classical pancreatic lipase but also gastric lipase, which is stable and active despite the highly acidic stomach environment. We report here the structure of recombinant human gastric lipase at 3.0-A resolution, the first structure to be described within the mammalian acid lipase family. This globular enzyme (379 residues) consists of a core domain belonging to the alpha/beta hydrolase-fold family and a "cap" domain, which is analogous to that present in serine carboxypeptidases. It possesses a classical catalytic triad (Ser-153, His-353, Asp-324) and an oxyanion hole (NH groups of Gln-154 and Leu-67). Four N-glycosylation sites were identified on the electron density maps. The catalytic serine is deeply buried under a segment consisting of 30 residues, which can be defined as a lid and belonging to the cap domain. The displacement of the lid is necessary for the substrates to have access to Ser-153. A phosphonate inhibitor was positioned in the active site that clearly suggests the location of the hydrophobic substrate binding site. The lysosomal acid lipase was modeled by homology, and possible explanations for some previously reported mutations leading to the cholesterol ester storage disease are given based on the present model.
Notes: Roussel, A
Canaan, S
Egloff, M P
Rivière, M
Dupuis, L
Verger, R
Cambillau, C
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1999 Jun 11;274(24):16995-7002. doi: 10.1074/jbc.274.24.16995.
Author Address: Architecture et Fonction des Macromolécules Biologiques, CNRS-IFR1 UPR 9039, 31 chemin Joseph Aiguier, 13402 Marseille cedex 20, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 693
Author: Roussel, A., Yang, Y., Ferrato, F., Verger, R., Cambillau, C. and Lowe, M.
Year: 1998
Title: Structure and activity of rat pancreatic lipase-related protein 2
Journal: J Biol Chem
Volume: 273
Issue: 48
Pages: 32121-8
Epub Date: 1998/11/21
Date: Nov 27
Short Title: Structure and activity of rat pancreatic lipase-related protein 2
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.48.32121
Accession Number: 9822688
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Humans
Kinetics
Lactones/pharmacology
Lipase/*chemistry/isolation & purification/*metabolism
Models, Molecular
Molecular Sequence Data
Orlistat
Paraoxon/pharmacology
Pressure
*Protein Conformation
Rats
Recombinant Proteins/chemistry/isolation & purification/metabolism
Spodoptera
Substrate Specificity
Surface Properties
Transfection
Abstract: The pancreas expresses several members of the lipase gene family including pancreatic triglyceride lipase (PTL) and two homologous proteins, pancreatic lipase-related proteins 1 and 2 (PLRP1 and PLRP2). Despite their similar amino acid sequences, PTL, PLRP1, and PLRP2 differ in important kinetic properties. PLRP1 has no known activity. PTL and PLRP2 differ in substrate specificity, bile acid inhibition, colipase requirement, and interfacial activation. To begin understanding the structural explanations for these functional differences, we solved the crystal structure of rat (r)PLRP2 and further characterized its kinetic properties. The 1.8 A structure of rPLRP2, like the tertiary structure of human PTL, has a globular N-terminal domain and a beta-sandwich C-terminal domain. The lid domain occupied the closed position, suggesting that rPLRP2 should show interfacial activation. When we reexamined this issue with tripropionin as substrate, rPLRP2 exhibited interfacial activation. Because the active site topology of rPLRP2 resembled that of human PTL, we predicted and demonstrated that the lipase inhibitors E600 and tetrahydrolipstatin inhibit rPLRP2. Although PTL and rPLRP2 have similar active sites, rPLRP2 has a broader substrate specificity that we confirmed using a monolayer technique. With this assay, we showed for the first time that rPLRP2 prefers phosphatidylglycerol and ethanolamine over phosphatidylcholine. In summary, we confirmed and extended the observation that PLRP2 lipases have a broader substrate specificity than PTL, we demonstrated that PLRP2 lipases show interfacial activation, and we solved the first crystal structure of a PLRP2 lipase that contains a lid domain.
Notes: Roussel, A
Yang, Y
Ferrato, F
Verger, R
Cambillau, C
Lowe, M
HD3306002/HD/NICHD NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1998 Nov 27;273(48):32121-8. doi: 10.1074/jbc.273.48.32121.
Author Address: Architecture et Fonction des Macromolécules Biologiques, CNRS-IFR1 UPR 9039, 31 Chemin Joseph Aiguier, 13402 Marseille cedex 20, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1415
Author: Roversi, P., Johnson, S., Caesar, J. J., McLean, F., Leath, K. J., Tsiftsoglou, S. A., Morgan, B. P., Harris, C. L., Sim, R. B. and Lea, S. M.
Year: 2011
Title: Structural basis for complement factor I control and its disease-associated sequence polymorphisms
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 31
Pages: 12839-44
Epub Date: 2011/07/20
Date: Aug 2
Short Title: Structural basis for complement factor I control and its disease-associated sequence polymorphisms
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1102167108
PMCID: PMC3150940
Accession Number: 21768352
Keywords: Allosteric Regulation
Binding Sites/genetics
Catalytic Domain
Complement C3b/chemistry/metabolism
Complement C4b/chemistry/metabolism
Complement Factor I/*chemistry/*genetics/metabolism
Crystallization
Crystallography, X-Ray
Enzyme Precursors/chemistry/genetics/metabolism
Genetic Predisposition to Disease/genetics
Glycosylation
Humans
Models, Molecular
Mutation
*Polymorphism, Genetic
Protein Binding
*Protein Structure, Tertiary
Abstract: The complement system is a key component of innate and adaptive immune responses. Complement regulation is critical for prevention and control of disease. We have determined the crystal structure of the complement regulatory enzyme human factor I (fI). FI is in a proteolytically inactive form, demonstrating that it circulates in a zymogen-like state despite being fully processed to the mature sequence. Mapping of functional data from mutants of fI onto the structure suggests that this inactive form is maintained by the noncatalytic heavy-chain allosterically modulating activity of the light chain. Once the ternary complex of fI, a cofactor and a substrate is formed, the allosteric inhibition is released, and fI is oriented for cleavage. In addition to explaining how circulating fI is limited to cleaving only C3b/C4b, our model explains the molecular basis of disease-associated polymorphisms in fI and its cofactors.
Notes: 1091-6490
Roversi, Pietro
Johnson, Steven
Caesar, Joseph J E
McLean, Florence
Leath, Kirstin J
Tsiftsoglou, Stefanos A
Morgan, B Paul
Harris, Claire L
Sim, Robert B
Lea, Susan M
083599/Wellcome Trust/United Kingdom
G0400775/Medical Research Council/United Kingdom
G0400775(71657)/Medical Research Council/United Kingdom
G0701298/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Aug 2;108(31):12839-44. doi: 10.1073/pnas.1102167108. Epub 2011 Jul 18.
Author Address: Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 904
Author: Rowlinson, S. W., Kiefer, J. R., Prusakiewicz, J. J., Pawlitz, J. L., Kozak, K. R., Kalgutkar, A. S., Stallings, W. C., Kurumbail, R. G. and Marnett, L. J.
Year: 2003
Title: A novel mechanism of cyclooxygenase-2 inhibition involving interactions with Ser-530 and Tyr-385
Journal: J Biol Chem
Volume: 278
Issue: 46
Pages: 45763-9
Epub Date: 2003/08/20
Date: Nov 14
Short Title: A novel mechanism of cyclooxygenase-2 inhibition involving interactions with Ser-530 and Tyr-385
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M305481200
Accession Number: 12925531
Keywords: Animals
Anti-Inflammatory Agents, Non-Steroidal/*pharmacology
Arachidonic Acid/chemistry
Arginine/chemistry
Binding, Competitive
Cell Line
Crystallography, X-Ray
Cyclooxygenase 2
Diclofenac/antagonists & inhibitors/chemistry
Dose-Response Relationship, Drug
Enzyme Inhibitors/*pharmacology
Insecta
Isoenzymes/*antagonists & inhibitors
Mice
Models, Chemical
Models, Molecular
Mutagenesis, Site-Directed
Mutation
Piroxicam/chemistry
Prostaglandin-Endoperoxide Synthases
Protein Binding
Serine/*chemistry
Sulfonamides/chemistry
Time Factors
Tyrosine/*chemistry
Abstract: A variety of drugs inhibit the conversion of arachidonic acid to prostaglandin G2 by the cyclooxygenase (COX) activity of prostaglandin endoperoxide synthases. Several modes of inhibitor binding in the COX active site have been described including ion pairing of carboxylic acid containing inhibitors with Arg-120 of COX-1 and COX-2 and insertion of arylsulfonamides and sulfones into the COX-2 side pocket. Recent crystallographic evidence suggests that Tyr-385 and Ser-530 chelate polar or negatively charged groups in arachidonic acid and aspirin. We tested the generality of this binding mode by analyzing the action of a series of COX inhibitors against site-directed mutants of COX-2 bearing changes in Arg-120, Tyr-355, Tyr-348, and Ser-530. Interestingly, diclofenac inhibition was unaffected by the mutation of Arg-120 to alanine but was dramatically attenuated by the S530A mutation. Determination of the crystal structure of a complex of diclofenac with murine COX-2 demonstrates that diclofenac binds to COX-2 in an inverted conformation with its carboxylate group hydrogen-bonded to Tyr-385 and Ser-530. This finding represents the first experimental demonstration that the carboxylate group of an acidic non-steroidal anti-inflammatory drug can bind to a COX enzyme in an orientation that precludes the formation of a salt bridge with Arg-120. Mutagenesis experiments suggest Ser-530 is also important in time-dependent inhibition by nimesulide and piroxicam.
Notes: Rowlinson, Scott W
Kiefer, James R
Prusakiewicz, Jeffery J
Pawlitz, Jennifer L
Kozak, Kevin R
Kalgutkar, Amit S
Stallings, William C
Kurumbail, Ravi G
Marnett, Lawrence J
CA68485/CA/NCI NIH HHS/United States
CA89450/CA/NCI NIH HHS/United States
ES00267/ES/NIEHS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2003 Nov 14;278(46):45763-9. doi: 10.1074/jbc.M305481200. Epub 2003 Aug 18.
Author Address: Department of Biochemistry, Vanderbilt Institute of Chemical Biology, Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0146, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1504
Author: Ruan, H. B., Han, X., Li, M. D., Singh, J. P., Qian, K., Azarhoush, S., Zhao, L., Bennett, A. M., Samuel, V. T., Wu, J., Yates, J. R., 3rd and Yang, X.
Year: 2012
Title: O-GlcNAc transferase/host cell factor C1 complex regulates gluconeogenesis by modulating PGC-1α stability
Journal: Cell Metab
Volume: 16
Issue: 2
Pages: 226-37
Epub Date: 2012/08/14
Date: Aug 8
Short Title: O-GlcNAc transferase/host cell factor C1 complex regulates gluconeogenesis by modulating PGC-1α stability
Alternate Journal: Cell metabolism
ISSN: 1550-4131 (Print)
1550-4131
DOI: 10.1016/j.cmet.2012.07.006
PMCID: PMC3480732
NIHMSID: NIHMS395801
Accession Number: 22883232
Keywords: Analysis of Variance
Animals
Blotting, Western
Chromatin Immunoprecipitation
Chromatography, High Pressure Liquid
Gluconeogenesis/*physiology
HEK293 Cells
Heat-Shock Proteins/*metabolism
Hep G2 Cells
Host Cell Factor C1/*metabolism
Humans
Hyperglycemia/*physiopathology
Immunoprecipitation
Liver/metabolism
Mice
Mice, Inbred C57BL
Multiprotein Complexes/*metabolism/physiology
N-Acetylglucosaminyltransferases/*metabolism
Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha
Proteomics
Real-Time Polymerase Chain Reaction
Tandem Mass Spectrometry
Transcription Factors/*metabolism
Abstract: A major cause of hyperglycemia in diabetic patients is inappropriate hepatic gluconeogenesis. PGC-1α is a master regulator of gluconeogenesis, and its activity is controlled by various posttranslational modifications. A small portion of glucose metabolizes through the hexosamine biosynthetic pathway, which leads to O-linked β-N-acetylglucosamine (O-GlcNAc) modification of cytoplasmic and nuclear proteins. Using a proteomic approach, we identified a broad variety of proteins associated with O-GlcNAc transferase (OGT), among which host cell factor C1 (HCF-1) is highly abundant. HCF-1 recruits OGT to O-GlcNAcylate PGC-1α, and O-GlcNAcylation facilitates the binding of the deubiquitinase BAP1, thus protecting PGC-1α from degradation and promoting gluconeogenesis. Glucose availability modulates gluconeogenesis through the regulation of PGC-1α O-GlcNAcylation and stability by the OGT/HCF-1 complex. Hepatic knockdown of OGT and HCF-1 improves glucose homeostasis in diabetic mice. These findings define the OGT/HCF-1 complex as a glucose sensor and key regulator of gluconeogenesis, shedding light on new strategies for treating diabetes.
Notes: 1932-7420
Ruan, Hai-Bin
Han, Xuemei
Li, Min-Dian
Singh, Jay Prakash
Qian, Kevin
Azarhoush, Sascha
Zhao, Lin
Bennett, Anton M
Samuel, Varman T
Wu, Jing
Yates, John R 3rd
Yang, Xiaoyong
R01 DK089098/DK/NIDDK NIH HHS/United States
P41 RR011823/RR/NCRR NIH HHS/United States
P30 AG021342/AG/NIA NIH HHS/United States
P01 DK057751/DK/NIDDK NIH HHS/United States
P30 DK045735/DK/NIDDK NIH HHS/United States
P30-DK34989/DK/NIDDK NIH HHS/United States
P30-DK045735/DK/NIDDK NIH HHS/United States
P30 DK034989/DK/NIDDK NIH HHS/United States
P01-DK057751/DK/NIDDK NIH HHS/United States
P41 GM103533/GM/NIGMS NIH HHS/United States
P30-AG021342/AG/NIA NIH HHS/United States
R01-DK089098/DK/NIDDK NIH HHS/United States
P41-RR011823/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell Metab. 2012 Aug 8;16(2):226-37. doi: 10.1016/j.cmet.2012.07.006.
Author Address: Program in Integrative Cell Signaling and Neurobiology of Metabolism, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06519, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 736
Author: Rudd, P. M., Downing, A. K., Cadene, M., Harvey, D. J., Wormald, M. R., Weir, I., Dwek, R. A., Rifkin, D. B. and Gleizes, P. E.
Year: 2000
Title: Hybrid and complex glycans are linked to the conserved N-glycosylation site of the third eight-cysteine domain of LTBP-1 in insect cells
Journal: Biochemistry
Volume: 39
Issue: 7
Pages: 1596-603
Epub Date: 2000/02/26
Date: Feb 22
Short Title: Hybrid and complex glycans are linked to the conserved N-glycosylation site of the third eight-cysteine domain of LTBP-1 in insect cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi9918285
Accession Number: 10677208
Keywords: Amino Acid Sequence
Animals
Baculoviridae/genetics
Carbohydrate Conformation
Carrier Proteins/chemistry/*genetics/*metabolism
Cell Line
Chromatography, High Pressure Liquid
Conserved Sequence
Cysteine/chemistry/genetics/*metabolism
Glycosylation
Humans
*Intracellular Signaling Peptides and Proteins
Latent TGF-beta Binding Proteins
Molecular Sequence Data
Oligosaccharides/chemistry
Peptide Fragments/biosynthesis/chemistry/genetics/metabolism
Polysaccharides/chemistry/*genetics/*metabolism
Protein Structure, Tertiary
Recombinant Fusion Proteins/chemistry/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Spodoptera/genetics
Transforming Growth Factor beta/metabolism
Abstract: Covalent association of LTBP-1 (latent TGF-beta binding protein-1) to latent TGF-beta is mediated by the third eight-cysteine (also referred to as TB) module of LTBP-1, a domain designated as CR3. Spodoptera frugiperda (Sf9) cells have proved a suitable cell system in which to study this association and to produce recombinant CR3, and we show here that another lepidopteran cell line, Trichoplusia niTN-5B1-4 (High-Five) cells, allows the recovery of large amounts of functional recombinant CR3. CR3 contains an N-glycosylation site, which is conserved in all forms of LTBP known to date. When we examined the status of this N-glycosylation using MALDI-TOF mass spectrometry and enzymatic analysis, we found that CR3 is one of the rare recombinant peptides modified with complex glycans in insect cells. Sf9 cells mainly processed the fucosylated paucomannosidic structure (GlcNAc)(2)(Mannose)(3)Fucose, although hybrid and complex N-glycosylations were also detected. In High-Five cells, the peptide was found to be modified with a wide variety of hybrid and complex sugars in addition to paucomanosidic oligosaccharides. Most glycans had one or two fucose residues bound through alpha1,3 and alpha1,6 linkages to the innermost GlcNAc. On the basis of these results and on the structure of an eight-cysteine domain from fibrillin-1, we present a model of glycosylated CR3 and discuss the role of glycosylation in eight-cysteine domain protein-protein interactions.
Notes: Rudd, P M
Downing, A K
Cadene, M
Harvey, D J
Wormald, M R
Weir, I
Dwek, R A
Rifkin, D B
Gleizes, P E
CA23753/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 2000 Feb 22;39(7):1596-603. doi: 10.1021/bi9918285.
Author Address: Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 714
Author: Rudd, P. M., Wormald, M. R., Harvey, D. J., Devasahayam, M., McAlister, M. S., Brown, M. H., Davis, S. J., Barclay, A. N. and Dwek, R. A.
Year: 1999
Title: Oligosaccharide analysis and molecular modeling of soluble forms of glycoproteins belonging to the Ly-6, scavenger receptor, and immunoglobulin superfamilies expressed in Chinese hamster ovary cells
Journal: Glycobiology
Volume: 9
Issue: 5
Pages: 443-58
Epub Date: 1999/04/20
Date: May
Short Title: Oligosaccharide analysis and molecular modeling of soluble forms of glycoproteins belonging to the Ly-6, scavenger receptor, and immunoglobulin superfamilies expressed in Chinese hamster ovary cells
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/9.5.443
Accession Number: 10207177
Keywords: Animals
Antigens, CD/chemistry
Antigens, Differentiation/*chemistry/metabolism
Antigens, Ly/chemistry
CD2 Antigens/chemistry
CD4 Antigens/chemistry
CD48 Antigen
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cricetinae
Glycoproteins/*chemistry/metabolism
Glycosylation
Humans
*Membrane Proteins
*Models, Molecular
Molecular Sequence Data
Oligosaccharides/*analysis
Protein Conformation
Protein Processing, Post-Translational
Protein Structure, Secondary
Rats
Receptors, Immunologic/chemistry
*Receptors, Lipoprotein
Receptors, Scavenger
Recombinant Proteins/chemistry/metabolism
Scavenger Receptors, Class B
Thy-1 Antigens/chemistry
Abstract: Most cell surface molecules are glycoproteins consisting of linear arrays of globular domains containing stretches of amino acid sequence with similarities to regions in other proteins. These conserved regions form the basis for the classification of proteins into superfamilies. Recombinant soluble forms of six leukocyte antigens belonging to the Ly-6 (CD59), scavenger receptor (CD5), and immunoglobulin (CD2, CD48, CD4, and Thy-1) superfamilies were expressed in the same Chinese hamster ovary cell line, thus providing an opportunity to examine the extent to which N-linked oligosaccharide processing might vary in a superfamily-, domain-, or protein-dependent manner in a given cell. While we found no evidence for superfamily-specific modifications of the glycans, marked differences were seen in the types of oligosaccharides attached to individual proteins within a given superfamily. The relative importance of local protein surface properties versus the overall tertiary structure of the molecules in directing this protein-specific variation was examined in the context of molecular models. These were constructed using the 3D structures of the proteins, glycan data from this study, and an oligosaccharide structural database. The results indicated that both the overall organization of the domains and the local protein structure can have a large bearing on site-specific glycan modification of cells in stasis. This level of control ensures that the surface of a single cell will display a diverse repertoire of glycans and precludes the presentation of multiple copies of a single oligosaccharide on the cell surface. The glycans invariably shield large regions of the protein surfaces although, for the glycoproteins examined here, these did not hinder the known active sites of the molecules. The models also indicated that sugars are likely to play a role in the packing of the native cell surface glycoproteins and to limit nonspecific protein-protein interactions. In addition, glycans located close to the cell membrane are likely to affect crucially the orientation of the glycoproteins to which they are attached.
Notes: Rudd, P M
Wormald, M R
Harvey, D J
Devasahayam, M
McAlister, M S
Brown, M H
Davis, S J
Barclay, A N
Dwek, R A
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1999 May;9(5):443-58. doi: 10.1093/glycob/9.5.443.
Author Address: Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1698
Author: Ruggiero, F. M., Vilcaes, A. A., Iglesias-Bartolomé, R. and Daniotti, J. L.
Year: 2015
Title: Critical role of evolutionarily conserved glycosylation at Asn211 in the intracellular trafficking and activity of sialyltransferase ST3Gal-II
Journal: Biochem J
Volume: 469
Issue: 1
Pages: 83-95
Epub Date: 2015/04/29
Date: Jul 1
Short Title: Critical role of evolutionarily conserved glycosylation at Asn211 in the intracellular trafficking and activity of sialyltransferase ST3Gal-II
Alternate Journal: The Biochemical journal
ISSN: 0264-6021
DOI: 10.1042/bj20150072
Accession Number: 25916169
Keywords: Animals
Asparagine/genetics/metabolism
CHO Cells
Cricetinae
Cricetulus
Endoplasmic Reticulum/*enzymology/genetics
Evolution, Molecular
Glycosylation
Golgi Apparatus/*enzymology/genetics
Humans
Protein Structure, Tertiary
Protein Transport/physiology
Sialyltransferases/genetics/*metabolism
Golgi complex
N-glycan trimming
N-glycosylation
ST3Gal-II
ganglioside
glycolipid
sialyltransferase
Abstract: ST3Gal-II, a type II transmembrane protein, is the main mammalian sialyltransferase responsible for GD1a and GT1b ganglioside biosynthesis in brain. It contains two putative N-glycosylation sites (Asn(92) and Asn(211)). Whereas Asn(92) is only conserved in mammalian species, Asn(211) is highly conserved in mammals, birds and fish. The present study explores the occupancy and relevance for intracellular trafficking and enzyme activity of these potential N-glycosylations in human ST3Gal-II. We found that ST3Gal-II distributes along the Golgi complex, mainly in proximal compartments. By pharmacological, biochemical and site-directed mutagenesis, we observed that ST3Gal-II is mostly N-glycosylated at Asn(211) and that this co-translational modification is critical for its exit from the endoplasmic reticulum and proper Golgi localization. The individual N-glycosylation sites had different effects on ST3Gal-II enzymatic activity. Whereas the N-glycan at position Asn(211) seems to negatively influence the activity of the enzyme using both glycolipid and glycoprotein as acceptor substrates, the single N-glycan mutant at Asn(92) had only a moderate effect. Lastly, we demonstrated that the N-terminal ST3Gal-II domain containing the cytosolic, transmembrane and stem region (amino acids 1-51) is able to drive a protein reporter out of the endoplasmic reticulum and to retain it in the Golgi complex. This suggests that the C-terminal domain of ST3Gal-II depends on N-glycosylation to attain an optimum conformation for proper exit from the endoplasmic reticulum, but it does not represent an absolute requirement for Golgi complex retention of the enzyme.
Notes: 1470-8728
Ruggiero, Fernando M
Vilcaes, Aldo A
Iglesias-Bartolomé, Ramiro
Daniotti, José L
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem J. 2015 Jul 1;469(1):83-95. doi: 10.1042/BJ20150072. Epub 2015 Apr 28.
Author Address: Centro de Investigaciones en Química Biológica de Córdoba (CIQUIBIC, UNC-CONICET), Departamento de Química Biológica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Córdoba, Argentina.
Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, U.S.A.
Centro de Investigaciones en Química Biológica de Córdoba (CIQUIBIC, UNC-CONICET), Departamento de Química Biológica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Córdoba, Argentina daniotti@dqb.fcq.unc.edu.ar.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1222
Author: Ruiz-Canada, C., Kelleher, D. J. and Gilmore, R.
Year: 2009
Title: Cotranslational and posttranslational N-glycosylation of polypeptides by distinct mammalian OST isoforms
Journal: Cell
Volume: 136
Issue: 2
Pages: 272-83
Epub Date: 2009/01/27
Date: Jan 23
Short Title: Cotranslational and posttranslational N-glycosylation of polypeptides by distinct mammalian OST isoforms
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2008.11.047
PMCID: PMC2859625
NIHMSID: NIHMS190454
Accession Number: 19167329
Keywords: Endoplasmic Reticulum/metabolism
Gene Knockdown Techniques
Glycoproteins/metabolism
Glycosylation
HeLa Cells
Hexosyltransferases/genetics/*metabolism
Humans
Membrane Proteins/genetics/*metabolism
Models, Molecular
Protein Folding
Protein Isoforms/chemistry/genetics/metabolism
Protein Processing, Post-Translational
Proteins/chemistry/*metabolism
Abstract: Asparagine-linked glycosylation of polypeptides in the lumen of the endoplasmic reticulum is catalyzed by the hetero-oligomeric oligosaccharyltransferase (OST). OST isoforms with different catalytic subunits (STT3A versus STT3B) and distinct enzymatic properties are coexpressed in mammalian cells. Using siRNA to achieve isoform-specific knockdowns, we show that the OST isoforms cooperate and act sequentially to mediate protein N-glycosylation. The STT3A OST isoform is primarily responsible for cotranslational glycosylation of the nascent polypeptide as it enters the lumen of the endoplasmic reticulum. The STT3B isoform is required for efficient cotranslational glycosylation of an acceptor site adjacent to the N-terminal signal sequence of a secreted protein. Unlike STT3A, STT3B efficiently mediates posttranslational glycosylation of a carboxyl-terminal glycosylation site in an unfolded protein. These distinct and complementary roles for the OST isoforms allow sequential scanning of polypeptides for acceptor sites to insure the maximal efficiency of N-glycosylation.
Notes: 1097-4172
Ruiz-Canada, Catalina
Kelleher, Daniel J
Gilmore, Reid
R01 GM043768/GM/NIGMS NIH HHS/United States
R01 GM043768-20/GM/NIGMS NIH HHS/United States
GM 43768/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell. 2009 Jan 23;136(2):272-83. doi: 10.1016/j.cell.2008.11.047.
Author Address: Department of Biochemistry and Molecular Pharmacology University of Massachusetts Medical School, Worcester, MA 01605, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1403
Author: Ruiz-Carrillo, D., Koch, B., Parthier, C., Wermann, M., Dambe, T., Buchholz, M., Ludwig, H. H., Heiser, U., Rahfeld, J. U., Stubbs, M. T., Schilling, S. and Demuth, H. U.
Year: 2011
Title: Structures of glycosylated mammalian glutaminyl cyclases reveal conformational variability near the active center
Journal: Biochemistry
Volume: 50
Issue: 28
Pages: 6280-8
Epub Date: 2011/06/16
Date: Jul 19
Short Title: Structures of glycosylated mammalian glutaminyl cyclases reveal conformational variability near the active center
Alternate Journal: Biochemistry
ISSN: 0006-2960
DOI: 10.1021/bi200249h
Accession Number: 21671571
Keywords: Amino Acid Sequence
Aminoacyltransferases/*chemistry/genetics/*metabolism
Animals
Cattle
Conserved Sequence
Crystallography, X-Ray
Enzyme Activation/genetics
Escherichia coli/enzymology/genetics
Glycosylation
Humans
Mice
Molecular Sequence Data
Pichia/enzymology/genetics
Protein Conformation
Protein Processing, Post-Translational/genetics
Rats
Sheep
Abstract: Formation of N-terminal pyroglutamate (pGlu or pE) from glutaminyl or glutamyl precursors is catalyzed by glutaminyl cyclases (QC). As the formation of pGlu-amyloid has been linked with Alzheimer's disease, inhibitors of QCs are currently the subject of intense development. Here, we report three crystal structures of N-glycosylated mammalian QC from humans (hQC) and mice (mQC). Whereas the overall structures of the enzymes are similar to those reported previously, two surface loops in the neighborhood of the active center exhibit conformational variability. Furthermore, two conserved cysteine residues form a disulfide bond at the base of the active center that was not present in previous reports of hQC structure. Site-directed mutagenesis suggests a structure-stabilizing role of the disulfide bond. At the entrance to the active center, the conserved tryptophan residue, W(207), which displayed multiple orientations in previous structure, shows a single conformation in both glycosylated human and murine QCs. Although mutagenesis of W(207) into leucine or glutamine altered substrate conversion significantly, the binding constants of inhibitors such as the highly potent PQ50 (PBD150) were minimally affected. The crystal structure of PQ50 bound to the active center of murine QC reveals principal binding determinants provided by the catalytic zinc ion and a hydrophobic funnel. This study presents a first comparison of two mammalian QCs containing typical, conserved post-translational modifications.
Notes: 1520-4995
Ruiz-Carrillo, David
Koch, Birgit
Parthier, Christoph
Wermann, Michael
Dambe, Tresfore
Buchholz, Mirko
Ludwig, Hans-Henning
Heiser, Ulrich
Rahfeld, Jens-Ulrich
Stubbs, Milton T
Schilling, Stephan
Demuth, Hans-Ulrich
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2011 Jul 19;50(28):6280-8. doi: 10.1021/bi200249h. Epub 2011 Jun 27.
Author Address: Probiodrug AG, Weinbergweg 22, D-06120 Halle, Saale, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1530
Author: Ruotsalainen, H., Risteli, M., Wang, C., Wang, Y., Karppinen, M., Bergmann, U., Kvist, A. P., Pospiech, H., Herzig, K. H. and Myllylä, R.
Year: 2012
Title: The activities of lysyl hydroxylase 3 (LH3) regulate the amount and oligomerization status of adiponectin
Journal: PLoS One
Volume: 7
Issue: 11
Pages: e50045
Epub Date: 2012/12/05
Short Title: The activities of lysyl hydroxylase 3 (LH3) regulate the amount and oligomerization status of adiponectin
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0050045
PMCID: PMC3510199 following conflicts: Accepted patents concerning lysyl hydroxylase 3: US7951562 Method for glycosylating hydroxylysine residues in polypeptides or peptides US7341853 Method for glycosylating hydroxylysine residues. Pending patent applications concerning lysyl hydroxylase 3 and proteins with collagenous domain: EP2349326 A New pharmaceutical product US2010124548 Pharmaceutical product. The authors have received research funding for the employment of a technician and for some mouse expenses during one year from a commercial funder, Glugal Research/IP Finland, due to their interest of pursuing pending patent application of Oulu University concerning LH3 and adiponectin. However, the company has since withdrawn from the project as a result of change in their business strategy and has no rights for the presented results. There are no further patents, products in development or marketed products to declare. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials.
Accession Number: 23209641
Keywords: Adiponectin/*chemistry/*metabolism
Amino Acid Sequence
Animals
Cell Line
Enzyme Activation
Female
Gene Expression Regulation
Gene Knockout Techniques
Genotype
Glycosylation
Hydroxylation
Lysine/chemistry/metabolism
Male
Mice
Molecular Sequence Data
Molecular Weight
Mutation
Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase/genetics/*metabolism
*Protein Multimerization
Recombinant Proteins/genetics/metabolism
Abstract: Lysyl hydroxylase 3 (LH3) has lysyl hydroxylase, galactosyltransferase, and glucosyltransferase activities, which are sequentially required for the formation of glucosylgalactosyl hydroxylysines in collagens. Here we demonstrate for the first time that LH3 also modifies the lysine residues in the collagenous domain of adiponectin, which has important roles in glucose and lipid metabolism and inflammation. Hydroxylation and, especially, glycosylation of the lysine residues of adiponectin have been shown to be essential for the formation of the more active high molecular weight adiponectin oligomers and thus for its function. In cells that totally lack LH3 enzyme, the galactosylhydroxylysine residues of adiponectin were not glucosylated to glucosylgalactosylhydroxylysine residues and the formation of high and middle molecular weight adiponectin oligomers was impaired. Circulating adiponectin levels in mutant mice lacking the lysyl hydroxylase activity of LH3 were significantly reduced, which indicates that LH3 is required for complete modification of lysine residues in adiponectin and the loss of some of the glycosylated hydroxylysine residues severely affects the secretion of adiponectin. LH mutant mice with reduced adiponectin level showed a high fat diet-induced increase in glucose, triglyceride, and LDL-cholesterol levels, hallmarks of the metabolic syndrome in humans. Our results reveal the first indication that LH3 is an important regulator of adiponectin biosynthesis, secretion and activity and thus might be a potential candidate for therapeutic applications in diseases associated with obesity and insulin resistance.
Notes: 1932-6203
Ruotsalainen, Heli
Risteli, Maija
Wang, Chunguang
Wang, Yu
Karppinen, Marjo
Bergmann, Ulrich
Kvist, Ari-Pekka
Pospiech, Helmut
Herzig, Karl-Heinz
Myllylä, Raili
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2012;7(11):e50045. doi: 10.1371/journal.pone.0050045. Epub 2012 Nov 29.
Author Address: Department of Biochemistry, University of Oulu, Oulu, Finland. heli.ruotsalainen@oulu.fi
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 548
Author: Rusiniak, M. E., Bedi, G. S. and Back, N.
Year: 1994
Title: The carbohydrate structure of the asparagine-linked oligosaccharides of rat plasma thiostatin
Journal: Biochim Biophys Acta
Volume: 1208
Issue: 2
Pages: 316-23
Epub Date: 1994/10/19
Date: Oct 19
Short Title: The carbohydrate structure of the asparagine-linked oligosaccharides of rat plasma thiostatin
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/0167-4838(94)90119-8
Accession Number: 7947964
Keywords: Amino Acids/analysis
Animals
Asparagine/*chemistry
Carbohydrate Sequence
Carbohydrates/analysis
Glycopeptides/isolation & purification
Glycoside Hydrolases
Kininogens/blood/*chemistry/isolation & purification
Molecular Sequence Data
Oligosaccharides/*chemistry
Pronase
Rats
Sugar Alcohols/analysis
Abstract: The complete carbohydrate structure of the asparagine-linked oligosaccharides of rat plasma thiostatin was elucidated through chemical and enzymatic methods including gas chromatography-mass spectrometry (GC-MS) and lectin affinity chromatography. Pronase digestion of thiostatin yielded a major glycopeptide fraction with asparagine the most abundant amino acid present. Based on one mole of aspartic acid, the following molar ratios obtained for the four major amino acids: aspartic acid (1.0), threonine (0.53), glycine (0.48) and serine (0.30). Neutral sugar analysis yielded a 3:2 molar ratio for mannose to galactose based on an assigned value to mannose of 3. On this basis, the fraction also contained 3 residues of sialic acid and, on average, 0 to 1 residue of fucose. GC-MS of partially methylated alditol acetates from the glycopeptide fraction identified the presence of biantennary and triantennary structure. Analyses of the neutral sugar and amino-acid composition, together with methylation data, support a biantennary N-linked structure for this major glycopeptide fraction and a triantennary N-linked structure as a lesser component. Sequencing of the desialyated 14C-labelled glycopeptide fraction by sequential exoglycosidase digestion and lectin affinity chromatography uncovered the following saccharide order: terminal galactose, N-acetylglucosamine and pentasaccharide inner core. This sequence is consistent with the N-linked glycan structures demonstrated by methylation and compositional analyses.
Notes: Rusiniak, M E
Bedi, G S
Back, N
DE-09376/DE/NIDCR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Netherlands
Biochim Biophys Acta. 1994 Oct 19;1208(2):316-23. doi: 10.1016/0167-4838(94)90119-8.
Author Address: Department of Biochemical Pharmacology, School of Pharmacy, State University of New York at Buffalo 14260.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1217
Author: Rutsch, F., Gailus, S., Miousse, I. R., Suormala, T., Sagné, C., Toliat, M. R., Nürnberg, G., Wittkampf, T., Buers, I., Sharifi, A., Stucki, M., Becker, C., Baumgartner, M., Robenek, H., Marquardt, T., Höhne, W., Gasnier, B., Rosenblatt, D. S., Fowler, B. and Nürnberg, P.
Year: 2009
Title: Identification of a putative lysosomal cobalamin exporter altered in the cblF defect of vitamin B12 metabolism
Journal: Nat Genet
Volume: 41
Issue: 2
Pages: 234-9
Epub Date: 2009/01/13
Date: Feb
Short Title: Identification of a putative lysosomal cobalamin exporter altered in the cblF defect of vitamin B12 metabolism
Alternate Journal: Nature genetics
ISSN: 1061-4036
DOI: 10.1038/ng.294
Accession Number: 19136951
Keywords: Child
Chromosome Deletion
Chromosome Mapping
Chromosomes, Human, Pair 6
Female
HeLa Cells
Humans
Hyperhomocysteinemia/*complications/genetics
Lysosome-Associated Membrane Glycoproteins/metabolism
Male
Membrane Transport Proteins/*deficiency/genetics/metabolism
Methylmalonic Acid/*metabolism/urine
Nucleocytoplasmic Transport Proteins/genetics/metabolism/physiology
Polymorphism, Genetic
Proteins/*genetics/isolation & purification/metabolism
Tissue Distribution
Transcobalamins/*genetics/isolation & purification/metabolism
Vitamin B 12/*metabolism
Vitamin B 12 Deficiency/etiology/*genetics/metabolism
Abstract: Vitamin B(12) (cobalamin) is essential in animals for metabolism of branched chain amino acids and odd chain fatty acids, and for remethylation of homocysteine to methionine. In the cblF inborn error of vitamin B(12) metabolism, free vitamin accumulates in lysosomes, thus hindering its conversion to cofactors. Using homozygosity mapping in 12 unrelated cblF individuals and microcell-mediated chromosome transfer, we identified a candidate gene on chromosome 6q13, LMBRD1, encoding LMBD1, a lysosomal membrane protein with homology to lipocalin membrane receptor LIMR. We identified five different frameshift mutations in LMBRD1 resulting in loss of LMBD1 function, with 18 of the 24 disease chromosomes carrying the same mutation embedded in a common 1.34-Mb haplotype. Transfection of fibroblasts of individuals with cblF with wild-type LMBD1 rescued cobalamin coenzyme synthesis and function. This work identifies LMBRD1 as the gene underlying the cblF defect of cobalamin metabolism and suggests that LMBD1 is a lysosomal membrane exporter for cobalamin.
Notes: 1546-1718
Rutsch, Frank
Gailus, Susann
Miousse, Isabelle R
Suormala, Terttu
Sagné, Corinne
Toliat, Mohammad Reza
Nürnberg, Gudrun
Wittkampf, Tanja
Buers, Insa
Sharifi, Azita
Stucki, Martin
Becker, Christian
Baumgartner, Matthias
Robenek, Horst
Marquardt, Thorsten
Höhne, Wolfgang
Gasnier, Bruno
Rosenblatt, David S
Fowler, Brian
Nürnberg, Peter
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Genet. 2009 Feb;41(2):234-9. doi: 10.1038/ng.294. Epub 2009 Jan 11.
Author Address: Department of General Pediatrics, Münster University Children's Hospital, Albert-Schweitzer-Strasse 33, D-48149 Münster, Germany. rutschf@mednet.uni-muenster.de
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1053
Author: Rutz, C., Renner, A., Alken, M., Schulz, K., Beyermann, M., Wiesner, B., Rosenthal, W. and Schülein, R.
Year: 2006
Title: The corticotropin-releasing factor receptor type 2a contains an N-terminal pseudo signal peptide
Journal: J Biol Chem
Volume: 281
Issue: 34
Pages: 24910-21
Epub Date: 2006/06/13
Date: Aug 25
Short Title: The corticotropin-releasing factor receptor type 2a contains an N-terminal pseudo signal peptide
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M601554200
Accession Number: 16766521
Keywords: Amino Acid Substitution
Animals
Protein Sorting Signals
Protein Structure, Tertiary
Rats
Receptors, Corticotropin-Releasing Hormone/*chemistry/genetics/metabolism
Structure-Activity Relationship
Abstract: The corticotropin-releasing factor receptor type 2a (CRF(2(a)) receptor) belongs to the family of G protein-coupled receptors. The receptor possesses a putative N-terminal signal peptide that is believed to be cleaved-off after mediating the endoplasmic reticulum targeting/insertion process, like the corresponding sequence of the homologous CRF(1) receptor. Here, we have assessed the functional significance of the putative signal peptide of the CRF(2(a)) receptor and show that it is surprisingly completely incapable of mediating endoplasmic reticulum targeting, despite meeting all sequence criteria for a functional signal by prediction algorithms. Moreover, it is uncleaved and forms part of the mature receptor protein. Replacement of residue Asn(13) by hydrophobic or positively charged residues converts the sequence into a fully functional and cleaved signal peptide demonstrating that conventional signal peptide functions are inhibited by a single amino acid residue. Deletion of the domain leads to an increase in the amount of immature, intracellularly retained receptors demonstrating that the sequence has adopted a new function in receptor trafficking through the early secretory pathway. Taken together, our results identify a novel hydrophobic receptor domain in the family of the heptahelical G protein-coupled receptors and the first pseudo signal peptide of a eukaryotic membrane protein. Our data also show that the extreme N termini of the individual CRF receptor subtypes differ substantially.
Notes: Rutz, Claudia
Renner, Armin
Alken, Martina
Schulz, Katharina
Beyermann, Michael
Wiesner, Burkhard
Rosenthal, Walter
Schülein, Ralf
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Aug 25;281(34):24910-21. doi: 10.1074/jbc.M601554200. Epub 2006 Jun 8.
Author Address: Leibniz-Institut für Molekulare Pharmakologie, Robert-Rössle-Strasse 10, 13125 Berlin, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2047
Author: Ryan, P., Xu, M. M., Davey, A. K., Kassiou, M., Mellick, G. D. and Rudrawar, S.
Year: 2020
Title: O-GlcNAcylation of truncated NAC segment alters peptide-dependent effects on α-synuclein aggregation
Journal: Bioorg Chem
Volume: 94
Pages: 103389
Epub Date: 2019/11/23
Date: Jan
Short Title: O-GlcNAcylation of truncated NAC segment alters peptide-dependent effects on α-synuclein aggregation
Alternate Journal: Bioorganic chemistry
ISSN: 0045-2068
DOI: 10.1016/j.bioorg.2019.103389
Accession Number: 31753312
Keywords: Acetylglucosamine/chemistry/*metabolism
Carbohydrate Conformation
Dose-Response Relationship, Drug
Glycosylation
Humans
Parkinson Disease/metabolism
Protein Aggregates/drug effects
Structure-Activity Relationship
alpha-Synuclein/chemistry/genetics/*metabolism
*Neurodegenerative diseases
*O-GlcNAcylation
*Parkinson’s disease
*α-Synuclein aggregation
Abstract: Numerous post-translational modifications (PTMs) of the Parkinson's disease (PD) associated α-synuclein (α-syn) protein have been recognised to play critical roles in disease aetiology. Indeed, dysregulated phosphorylation and proteolysis are thought to modulate α-syn aggregation and disease progression. Among the PTMs, enzymatic glycosylation with N-acetylglucosamine (GlcNAc) onto the protein's hydroxylated amino acid residues is reported to deliver protective effects against its pathogenic processing. This modification has been reported to alter its pathogenic self-assembly. As such, manipulation of the protein's O-GlcNAcylation status has been proposed to offer a PD therapeutic route. However, targeting upstream cellular processes can lead to mechanism-based toxicity as the enzymes governing O-GlcNAc cycling modify thousands of acceptor substrates. Small glycopeptides that couple the protective effects of O-GlcNAc with the selectivity of recognition sequences may prove useful tools to modulate protein aggregation. Here we discuss efforts to probe the effects of various O-GlcNAc modified peptides on wild-type α-synuclein aggregation.
Notes: 1090-2120
Ryan, Philip
Xu, Ming-Ming
Davey, Andrew K
Kassiou, Michael
Mellick, George D
Rudrawar, Santosh
Journal Article
United States
Bioorg Chem. 2020 Jan;94:103389. doi: 10.1016/j.bioorg.2019.103389. Epub 2019 Nov 9.
Author Address: School of Pharmacy and Pharmacology, Griffith University, Gold Coast, QLD 4222, Australia; Menzies Health Institute Queensland, Griffith University, Gold Coast, QLD 4222, Australia; Quality Use of Medicines Network, Griffith University, Gold Coast, QLD 4222, Australia.
Griffith Institute for Drug Discovery, Griffith University, Nathan, QLD 4111, Australia.
School of Chemistry, The University of Sydney, NSW 2006, Australia.
Griffith Institute for Drug Discovery, Griffith University, Nathan, QLD 4111, Australia. Electronic address: g.mellick@griffith.edu.au.
School of Pharmacy and Pharmacology, Griffith University, Gold Coast, QLD 4222, Australia; Menzies Health Institute Queensland, Griffith University, Gold Coast, QLD 4222, Australia; Quality Use of Medicines Network, Griffith University, Gold Coast, QLD 4222, Australia. Electronic address: s.rudrawar@griffith.edu.au.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 708
Author: Rydberg, E. H., Sidhu, G., Vo, H. C., Hewitt, J., Côte, H. C., Wang, Y., Numao, S., MacGillivray, R. T., Overall, C. M., Brayer, G. D. and Withers, S. G.
Year: 1999
Title: Cloning, mutagenesis, and structural analysis of human pancreatic alpha-amylase expressed in Pichia pastoris
Journal: Protein Sci
Volume: 8
Issue: 3
Pages: 635-43
Epub Date: 1999/03/26
Date: Mar
Short Title: Cloning, mutagenesis, and structural analysis of human pancreatic alpha-amylase expressed in Pichia pastoris
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.8.3.635
PMCID: PMC2144294
Accession Number: 10091666
Keywords: Amino Acid Sequence
Base Sequence
Cloning, Molecular
DNA Primers
Glycosylation
Humans
Kinetics
Mass Spectrometry
Molecular Sequence Data
Mutagenesis, Site-Directed
Pancreas/*enzymology
Pichia/*genetics
Protein Conformation
alpha-Amylases/chemistry/*genetics/metabolism
Abstract: Human pancreatic alpha-amylase (HPA) was expressed in the methylotrophic yeast Pichia pastoris and two mutants (D197A and D197N) of a completely conserved active site carboxylic acid were generated. All recombinant proteins were shown by electrospray ionization mass spectrometry (ESI-MS) to be glycosylated and the site of attachment was shown to be Asn461 by peptide mapping in conjunction with ESI-MS. Treatment of these proteins with endoglycosidase F demonstrated that they contained a single N-linked oligosaccharide and yielded a protein product with a single N-acetyl glucosamine (GlcNAc), which could be crystallized. Solution of the crystal structure to a resolution of 2.0 A confirmed the location of the glycosyl group as Asn461 and showed that the recombinant protein had essentially the same conformation as the native enzyme. The kinetic parameters of the glycosylated and deglycosylated wild-type proteins were the same while the k(cat)/Km values for D197A and D197N were 10(6)-10(7) times lower than the wild-type enzyme. The decreased k(cat)/Km values for the mutants confirm that D197 plays a crucial role in the hydrolytic activity of HPA, presumably as the catalytic nucleophile.
Notes: 1469-896x
Rydberg, E H
Sidhu, G
Vo, H C
Hewitt, J
Côte, H C
Wang, Y
Numao, S
MacGillivray, R T
Overall, C M
Brayer, G D
Withers, S G
Journal Article
Research Support, Non-U.S. Gov't
Protein Sci. 1999 Mar;8(3):635-43. doi: 10.1110/ps.8.3.635.
Author Address: Department of Biochemistry & Molecular Biology, University of British Columbia, Vancouver, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1759
Author: Ryu, I. H., Lee, K. Y. and Do, S. I.
Year: 2016
Title: Aβ-affected pathogenic induction of S-nitrosylation of OGT and identification of Cys-NO linkage triplet
Journal: Biochim Biophys Acta
Volume: 1864
Issue: 5
Pages: 609-21
Epub Date: 2016/02/09
Date: May
Short Title: Aβ-affected pathogenic induction of S-nitrosylation of OGT and identification of Cys-NO linkage triplet
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbapap.2016.02.003
Accession Number: 26854602
Keywords: Acylation
Alzheimer Disease/*metabolism/pathology
Amyloid beta-Peptides/chemistry/*metabolism
Calcium/metabolism
Cell Line
Cysteine/chemistry
Glycogen Synthase Kinase 3/chemistry/metabolism
Glycogen Synthase Kinase 3 beta
Humans
N-Acetylglucosaminyltransferases/*chemistry/metabolism
Nitric Oxide/chemistry
Phosphorylation
Protein Processing, Post-Translational/*genetics
Reactive Oxygen Species
Signal Transduction/genetics
Aβ neurotoxicity
Cys-NO linkages
Hypo-O-GlcNAcylation
O-GlcNAc
Ogt
Sno-ogt
Abstract: Mechanistic link of protein hypo-O-GlcNAcylation to the pathogenesis of Alzheimer's disease (AD) remains unclear. Here, we found that S-nitrosylation of O-linked N-acetylglucosaminyltransferase (SNO-OGT) was induced by β-amyloid peptide (Aβ) exposure to SK-N-MC and SK-N-SH human neuroblastoma cells. Subsequently, Aβ-induced SNO-OGT led to protein hypo-O-GlcNAcylation globally including tau hypo-O-GlcNAcylation. Our results support that underlying mechanism for induction of SNO-OGT comprises the concerted action of Aβ-triggered Ca2+ entry into cells and nNOS-catalyzed NO production. Intriguingly, OGT was found to be associated with nNOS and its association was enhanced during Aβ treatment. In parallel with SNO-OGT-mediated tau hypo-O-GlcNAcylation, Aβ led to SNO-Akt-mediated GSK3β activation for tau phosphorylation, suggesting that tau hyperphosphorylation is established by synergistic connection between SNO-OGT and GSK3β activation. We also observed that Aβ-neurotoxicity including both reactive oxygen species (ROS) production and cell death was amplified with DON treatment, whereas it was restored by PUGNAc treatment, GlcNH2 treatment or OGT overexpression. Early time-course Aβ-monitoring assay revealed that premaintained hyper-O-GlcNAcylation inside cells blocked not only Aβ-triggered Ca2+ entry into cells but also induction of SNO-OGT and SNO-Akt. Together, these findings suggest that induction of SNO-OGT by Aβ exposure is a pathogenic mechanism to cause cellular hypo-O-GlcNAcylation by which Aβ neurotoxicity is executed, and conversely, hyper-O-GlcNAcylation within cells can defend against Aβ neurotoxicity. Furthermore, our Cys mapping demonstrates that cysteine-nitric oxide (Cys-NO) linkages in SNO-OGT occur at triple Cys845, Cys921, and Cys965 residues in C-terminal catalytic domain (C-CAT), suggesting that Cys-NO linkage triplet in SNO-OGT is associated with null OGT activity.
Notes: Ryu, In-Hyun
Lee, Ki-Young
Do, Su-Il
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2016 May;1864(5):609-21. doi: 10.1016/j.bbapap.2016.02.003. Epub 2016 Feb 5.
Author Address: Department of Life Science, Ajou University, Suwon City, Republic of Korea.
GG Pharmaceutical Institute, Seoul, Republic of Korea.
Department of Life Science, Ajou University, Suwon City, Republic of Korea. Electronic address: sido@ajou.ac.kr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1908
Author: Saad, A., Liet, B., Joucla, G., Santarelli, X., Charpentier, J., Claverol, S., Grosset, C. F. and Trézéguet, V.
Year: 2018
Title: Role of Glycanation and Convertase Maturation of Soluble Glypican-3 in Inhibiting Proliferation of Hepatocellular Carcinoma Cells
Journal: Biochemistry
Volume: 57
Issue: 7
Pages: 1201-1211
Epub Date: 2018/01/19
Date: Feb 20
Short Title: Role of Glycanation and Convertase Maturation of Soluble Glypican-3 in Inhibiting Proliferation of Hepatocellular Carcinoma Cells
Alternate Journal: Biochemistry
ISSN: 0006-2960
DOI: 10.1021/acs.biochem.7b01208
Accession Number: 29345911
Keywords: Apoptosis
Carcinoma, Hepatocellular/genetics/*metabolism
Cell Cycle
Cell Division
Cell Line, Tumor
Cell Proliferation
Glycosaminoglycans/analysis/genetics/metabolism
Glypicans/chemistry/genetics/*metabolism
Hep G2 Cells
Humans
Liver Neoplasms/genetics/*metabolism
Mutation
Protein Processing, Post-Translational
Wnt Signaling Pathway
beta Catenin/metabolism
Abstract: Glypican 3 (GPC3) is a complex heparan sulfate proteoglycan associated with the outer surface of the plasma membrane by a glycosylphosphatidylinositol (GPI) anchor. It is also N-glycosylated and processed by a furin-like convertase. GPC3 has numerous biological functions. Although GPC3 is undetectable in normal liver tissue, it is abnormally and highly overexpressed in hepatocellular carcinoma (HCC). Interestingly, proliferation of HCC cells such as HepG2 and HuH7 is inhibited when they express a soluble form of GPC3 after lentiviral transduction. To obtain more insight into the role of some of its post-translational modifications, we designed a mutant GPC3, sGPC3m, without its GPI anchor, convertase cleavage site, and glycosaminoglycan chains. The highly pure sGPC3m protein strongly inhibited HuH7 and HepG2 cell proliferation in vitro and induced a significant increase in their cell doubling time. It changed the morphology of HuH7 cells but not that of HepG2. It induced the enlargement of HuH7 cell nuclear area and the restructuration of adherent cell junctions. Unexpectedly, for both cell types, the levels of apoptosis, cell division, and β-catenin were not altered by sGPC3m, although growth inhibition was very efficient. Overall, our data show that glycanation and convertase maturation are not required for sGPC3m to inhibit HCC cell proliferation.
Notes: 1520-4995
Saad, Ahmad
Liet, Benjamin
Joucla, Gilles
Santarelli, Xavier
Charpentier, Justine
Claverol, Stéphane
Grosset, Christophe F
Trézéguet, Véronique
Orcid: 0000-0003-2487-7571
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2018 Feb 20;57(7):1201-1211. doi: 10.1021/acs.biochem.7b01208. Epub 2018 Feb 2.
Author Address: Univ. Bordeaux, CBMN, UMR 5248 , F-33615 Pessac, France.
CNRS, CBMN, UMR 5248 , F-33615 Pessac, France.
Bordeaux INP, CBMN, UMR 5248 , F-33615 Pessac, France.
Univ. Bordeaux, Inserm, BMGIC, U1035 , 33076 Bordeaux, France.
Univ. Bordeaux, Plateforme Protéome, CGFB , F-33076 Bordeaux, France.
Univ. Bordeaux, Inserm, GREF, U1053 , 33076 Bordeaux, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 709
Author: Saarinen, J., Welgus, H. G., Flizar, C. A., Kalkkinen, N. and Helin, J.
Year: 1999
Title: N-glycan structures of matrix metalloproteinase-1 derived from human fibroblasts and from HT-1080 fibrosarcoma cells
Journal: Eur J Biochem
Volume: 259
Issue: 3
Pages: 829-40
Epub Date: 1999/03/27
Date: Feb
Short Title: N-glycan structures of matrix metalloproteinase-1 derived from human fibroblasts and from HT-1080 fibrosarcoma cells
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.1999.00105.x
Accession Number: 10092871
Keywords: Carbohydrate Sequence
Cells, Cultured
Collagenases/*chemistry
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Mass Spectrometry
Matrix Metalloproteinase 1
Molecular Sequence Data
Oligosaccharides/chemistry
Polysaccharides/*chemistry
Abstract: Matrix metalloproteinase-1 (MMP-1) is a collagenolytic metalloproteinase capable of cleaving native triple-helical forms of several collagen subtypes, as well as a number of non-collagenous substrates. The role of MMP-1 in various diseases affecting the connective tissue is well characterized. MMP-1 is secreted as both glycosylated and unglycosylated species, and the two forms have been shown to be identical with respect to substrate specificity, specific activity and inhibitory profile. No function for the glycan moiety of the enzyme has been ascribed to date. In the present study, we report on the detailed characterization of MMP-1-derived oligosaccharides. Using strategies based on sequential exoglycosidase digestion combined with matrix-assisted laser desorption ionization-time of flight MS and electrospray tandem MS, we have characterized the N-glycan structures of MMP-1, derived from human dermal fibroblasts and from the HT-1080 fibrosarcoma cell line. MMP-1 derived from fibroblasts was found to carry mainly alpha 2,3-sialylated complex-type diantennary glycans. On the other hand, HT-1080 cells produce MMP-1 that has a heterogeneous glycosylation pattern, comprising diantennary glycans carrying Lewis X, LacdiNAc, sialylated LacdiNAc and GalNAc beta 1,4 (Fuc alpha 1,3)GlcNAc (LacdiNAc analogue of Lewis X) as terminal elements. We also show that, of the two potential glycosylation sites in the MMP-1 sequence, only Asn120 is used.
Notes: Saarinen, J
Welgus, H G
Flizar, C A
Kalkkinen, N
Helin, J
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1999 Feb;259(3):829-40. doi: 10.1046/j.1432-1327.1999.00105.x.
Author Address: Institute of Biotechnology, University of Helsinki, Finland. juhani.saarinen@helsinki.fi
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2001
Author: Sager, R. A., Woodford, M. R., Backe, S. J., Makedon, A. M., Baker-Williams, A. J., DiGregorio, B. T., Loiselle, D. R., Haystead, T. A., Zachara, N. E., Prodromou, C., Bourboulia, D., Schmidt, L. S., Linehan, W. M., Bratslavsky, G. and Mollapour, M.
Year: 2019
Title: Post-translational Regulation of FNIP1 Creates a Rheostat for the Molecular Chaperone Hsp90
Journal: Cell Rep
Volume: 26
Issue: 5
Pages: 1344-1356.e5
Epub Date: 2019/01/31
Date: Jan 29
Short Title: Post-translational Regulation of FNIP1 Creates a Rheostat for the Molecular Chaperone Hsp90
Alternate Journal: Cell reports
DOI: 10.1016/j.celrep.2019.01.018
PMCID: PMC6370319
NIHMSID: NIHMS1519903
Accession Number: 30699359
Keywords: Carrier Proteins/*metabolism
Casein Kinase II/metabolism
Glycosylation
HEK293 Cells
HSP90 Heat-Shock Proteins/*metabolism
Humans
Models, Biological
Nuclear Proteins/metabolism
Phosphoprotein Phosphatases/metabolism
Phosphorylation
Phosphoserine/metabolism
Proteasome Endopeptidase Complex/metabolism
Protein Binding
*Protein Processing, Post-Translational
Ubiquitination
*Bhd
*Birt-Hogg-Dubé syndrome
*Fnip1
*Hsp90
*O-GlcNAcylation
*Pp5
*co-chaperone
*folliculin-interacting protein 1
*heat shock protein 90
*serine/threonine protein phosphatase 5
Abstract: The molecular chaperone Hsp90 stabilizes and activates client proteins. Co-chaperones and post-translational modifications tightly regulate Hsp90 function and consequently lead to activation of clients. However, it is unclear whether this process occurs abruptly or gradually in the cellular context. We show that casein kinase-2 phosphorylation of the co-chaperone folliculin-interacting protein 1 (FNIP1) on priming serine-938 and subsequent relay phosphorylation on serine-939, 941, 946, and 948 promotes its gradual interaction with Hsp90. This leads to incremental inhibition of Hsp90 ATPase activity and gradual activation of both kinase and non-kinase clients. We further demonstrate that serine/threonine protein phosphatase 5 (PP5) dephosphorylates FNIP1, allowing the addition of O-GlcNAc (O-linked N-acetylglucosamine) to the priming serine-938. This process antagonizes phosphorylation of FNIP1, preventing its interaction with Hsp90, and consequently promotes FNIP1 lysine-1119 ubiquitination and proteasomal degradation. These findings provide a mechanism for gradual activation of the client proteins through intricate crosstalk of post-translational modifications of the co-chaperone FNIP1.
Notes: 2211-1247
Sager, Rebecca A
Woodford, Mark R
Backe, Sarah J
Makedon, Alan M
Baker-Williams, Alexander J
DiGregorio, Bryanna T
Loiselle, David R
Haystead, Timothy A
Zachara, Natasha E
Prodromou, Chrisostomos
Bourboulia, Dimitra
Schmidt, Laura S
Linehan, W Marston
Bratslavsky, Gennady
Mollapour, Mehdi
HHSN261200800001C/RC/CCR NIH HHS/United States
HHSN261200800001E/CA/NCI NIH HHS/United States
R01 GM124256/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Cell Rep. 2019 Jan 29;26(5):1344-1356.e5. doi: 10.1016/j.celrep.2019.01.018.
Author Address: Department of Urology, SUNY Upstate Medical University, Syracuse, NY 13210, USA; Department of Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, NY 13210, USA; Upstate Cancer Center, SUNY Upstate Medical University, Syracuse, NY 13210, USA.
Department of Urology, SUNY Upstate Medical University, Syracuse, NY 13210, USA; Upstate Cancer Center, SUNY Upstate Medical University, Syracuse, NY 13210, USA.
Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710, USA.
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Genome Damage and Stability Centre, University of Sussex, Brighton BN1 9RQ, UK.
Basic Science Program, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, MD 21702, USA; Urologic Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
Urologic Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
Department of Urology, SUNY Upstate Medical University, Syracuse, NY 13210, USA; Department of Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, NY 13210, USA; Upstate Cancer Center, SUNY Upstate Medical University, Syracuse, NY 13210, USA. Electronic address: mollapom@upstate.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 604
Author: Saharinen, J., Taipale, J. and Keski-Oja, J.
Year: 1996
Title: Association of the small latent transforming growth factor-beta with an eight cysteine repeat of its binding protein LTBP-1
Journal: Embo j
Volume: 15
Issue: 2
Pages: 245-53
Epub Date: 1996/01/15
Date: Jan 15
Short Title: Association of the small latent transforming growth factor-beta with an eight cysteine repeat of its binding protein LTBP-1
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
PMCID: PMC449939
Accession Number: 8617200
Keywords: Amino Acid Sequence
Animals
Binding Sites
CHO Cells
Carrier Proteins/biosynthesis/chemistry/*metabolism
Cell Line
Chlorocebus aethiops
Cricetinae
*Cysteine
Disulfides
Electrophoresis, Polyacrylamide Gel
Epidermal Growth Factor/metabolism
Extracellular Matrix/physiology
Extracellular Matrix Proteins/chemistry/metabolism
Humans
*Intracellular Signaling Peptides and Proteins
Kidney
Latent TGF-beta Binding Proteins
Models, Structural
Molecular Sequence Data
Polymerase Chain Reaction
Recombinant Proteins/biosynthesis/isolation & purification/metabolism
Repetitive Sequences, Nucleic Acid
Transfection
Transforming Growth Factor beta/biosynthesis/isolation & purification/*metabolism
Abstract: Transforming growth factor-betas (TGF-betas) are produced by most cells in large latent complexes of TGF-beta and its propeptide (LAP) associated with a binding protein. The latent TGF-beta binding proteins (LTBPs-1, -2 and -3) mediate the secretion and, subsequently, the association of latent TGF-beta complexes with the extracellular matrix (ECM). The association of beta1-LAP with LTBP-1 was characterized at the molecular level with an expression system in mammalian cells, where TGF-beta1 and various fragments of LTBP-1 were co-expressed and secreted with the aid of a signal peptide synthesized to the LTBP-1 constructs. Immunoblotting of the fusion protein complexes indicated that the third 8-Cys repeat of LTBP-1 bound covalently to the LAP region of TGF-beta1. The cysteine required for the association between LTBP-1 and beta1-LAP was mapped to Cys33 of beta1-LAP. The N-terminal region of LTBP-1 consisting of the first 400 amino acids was found to associate covalently with the ECM. The data indicate that an 8-Cys repeat of LTBP is capable of covalent and specific protein-protein interactions. These interactions are mediated by exchanging cysteine disulfide bonds between the core 8-Cys repeat and an optionally associated protein during the secretion. This is, to our knowledge, the first demonstration of an extracellular protein module that is able to exchange cysteine disulfide bonds with heterologous ligand proteins.
Notes: 1460-2075
Saharinen, J
Taipale, J
Keski-Oja, J
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1996 Jan 15;15(2):245-53.
Author Address: Department of Virology, University of Helinski, Finland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 534
Author: Saito, F., Tani, A., Miyatake, T. and Yanagisawa, K.
Year: 1995
Title: N-linked oligosaccharide of beta-amyloid precursor protein (beta APP) of C6 glioma cells: putative regulatory role in beta APP processing
Journal: Biochem Biophys Res Commun
Volume: 210
Issue: 3
Pages: 703-10
Epub Date: 1995/05/25
Date: May 25
Short Title: N-linked oligosaccharide of beta-amyloid precursor protein (beta APP) of C6 glioma cells: putative regulatory role in beta APP processing
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1006/bbrc.1995.1716
Accession Number: 7763244
Keywords: Amyloid beta-Protein Precursor/*biosynthesis/*chemistry/isolation & purification
Animals
Base Sequence
Blotting, Western
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
DNA, Complementary
Glioma
Humans
Lectins
Molecular Sequence Data
Molecular Weight
Oligodeoxyribonucleotides
Oligosaccharides/*chemistry/isolation & purification/*metabolism
*Protein Processing, Post-Translational
Recombinant Proteins/biosynthesis/chemistry/isolation & purification
Transfection
Tumor Cells, Cultured
Abstract: To determine the N-linked oligosaccharide structure of beta-amyloid precursor protein (beta APP), soluble derivative of beta APP (APPs) was purified from the conditioned medium of beta APP cDNA-transfected C6 glioma cells. Two types of APPs with different molecular weight (larger APPs, L-APPs; smaller APPs, S-APPs) were obtained. The antibody against the N-terminal half of amyloid beta-protein showed no immunoreactivity with S-APPs, suggesting extensive truncation at the carboxyl terminus. From lectin blot analysis, the main structure of the N-linked oligosaccharide shared by L- and S-APPs was deduced to be of bi- or triantennary complex type with a fucosylated trimannosyl core and a bisecting GlcNAc residue. Additionally L-APPs was deduced to have Gal beta 1-->4GlcNAc, Fuc alpha 1-->2Gal beta and Sia alpha 2-->6Gal beta structures on its outer chains. However, lectins which recognize Fuc alpha 1-->2Gal beta and Sia alpha 2-->6Gal beta structures showed no reactivity with S-APPs. The present results suggest that the processing of beta APP may be regulated via the heterogeneity in the fine structure of its sugar chains.
Notes: Saito, F
Tani, A
Miyatake, T
Yanagisawa, K
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 1995 May 25;210(3):703-10. doi: 10.1006/bbrc.1995.1716.
Author Address: Department of Neurology, School of Medicine, Tokyo Medical and Dental University, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1155
Author: Saito, S., Yano, K., Sharma, S., McMahon, H. E. and Shimasaki, S.
Year: 2008
Title: Characterization of the post-translational modification of recombinant human BMP-15 mature protein
Journal: Protein Sci
Volume: 17
Issue: 2
Pages: 362-70
Epub Date: 2008/01/30
Date: Feb
Short Title: Characterization of the post-translational modification of recombinant human BMP-15 mature protein
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1110/ps.073232608
PMCID: PMC2222730
Accession Number: 18227435
Keywords: Amino Acid Sequence
Animals
Bone Morphogenetic Protein 15
Glycosylation
Growth Differentiation Factor 9
Humans
Intercellular Signaling Peptides and Proteins/*chemistry/*metabolism
Molecular Sequence Data
Phosphorylation
Protein Processing, Post-Translational
Recombinant Proteins/chemistry/metabolism
Sequence Alignment
Abstract: Bone morphogenetic protein-15 (BMP-15) is an oocyte-secreted factor critical for the regulation of ovarian physiology. When recombinant human BMP-15 (rhBMP-15) produced in human embryonic kidney 293 cells was subjected to SDS-PAGE analysis, two mature protein forms corresponding to 16 kDa (P16) and 17 kDa (P17) were observed. Despite the physiological relevance and critical function of BMP-15 in female reproduction, little is known about the structure of rhBMP-15. Here, we have analyzed the structure of the rhBMP-15 mature proteins (P16 and P17) using state-of-the-art proteomics technology. Our findings are as follows: (1) the N-terminal amino acid of P16 and P17 is pyroglutamic acid; (2) the Ser residue at the sixth position of P16 is phosphorylated; (3) P17 is O-glycosylated at Thr10; and (4) the C-terminal amino acid of P16 and P17 is truncated. These findings are the first knowledge of the structure of rhBMP-15 mature protein toward understanding the molecular basis of BMP-15 function and could provide an important contribution to the rapidly progressing research area involving oocyte-specific growth factors in modulation of female fertility.
Notes: 1469-896x
Saito, Seiji
Yano, Keiichi
Sharma, Shweta
McMahon, Heather E
Shimasaki, Shunichi
R01 HD041494/HD/NICHD NIH HHS/United States
R01 HD41494/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Protein Sci. 2008 Feb;17(2):362-70. doi: 10.1110/ps.073232608.
Author Address: Antibody Research Laboratories, Pharmaceutical Research Center, Kyowa Hakko Kogyo Co., Ltd., Tokyo 194-8533, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 388
Author: Saito, T., Itoh, T. and Adachi, S.
Year: 1988
Title: Chemical structure of neutral sugar chains isolated from human mature milk kappa-casein
Journal: Biochim Biophys Acta
Volume: 964
Issue: 2
Pages: 213-20
Epub Date: 1988/02/17
Date: Feb 17
Short Title: Chemical structure of neutral sugar chains isolated from human mature milk kappa-casein
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/0304-4165(88)90169-9
Accession Number: 3342257
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
*Caseins
Female
Humans
Mass Spectrometry
Milk, Human/*analysis
Molecular Sequence Data
Oligosaccharides/*isolation & purification
Abstract: The carbohydrate chains linked to human kappa-casein from mature milk were released by alkaline borohydride treatment as reduced oligosaccharides. The neutral oligosaccharides of lower molecular weight were fractionated and purified by gel filtration and preparative thin layer chromatographies. Seven neutral oligosaccharides (a di- (0.5%), two tetra- (30.5%), two penta- (5.4%) and two hexasaccharide alditols (10.9%] were obtained in homogeneity, and followed by methylation analysis with gas-liquid chromatography-mass spectrometry and by anomer analysis with 13C nuclear magnetic resonance. Their chemical structures were identified to be Gal beta 1----3GalNAc-ol (I), Gal beta 1----3[Gal beta 1----4GlcNAc beta 1----6]GalNAc-ol (II), Gal beta 1----3[Fuc alpha 1----4GlcNAc beta 1----6]GalNAc-ol (III), GlcNAc beta 1----3/6Gal beta 1----3[Gal beta 1----4GlcNAc beta 1----6]GalNAc-ol (IV), GlcNAc beta 1----3/6Gal beta 1----3[Fuc alpha 1----4GlcNAc beta 1----6]GalNAc-ol (V), Fuc alpha 1----4GlcNAc beta 1----3/6Gal beta 1----3[Gal beta 1----4GlcNAc beta 1----6]GalNAc-ol (VI) and Fuc alpha 1----4GlcNAc beta 1----3/6Gal beta 1----3[Fuc alpha 1----4GlcNAc beta 1----6]GalNAc-ol (VII). Five oligosaccharide alditols (III-VII) were the novel carbohydrate chains of kappa-casein from mammalian milk.
Notes: Saito, T
Itoh, T
Adachi, S
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 1988 Feb 17;964(2):213-20. doi: 10.1016/0304-4165(88)90169-9.
Author Address: Laboratory of Food Hygiene, Faculty of Social Welfare, Tohoku Fukushi University, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 585
Author: Saitoh, A., Aoyagi, Y. and Asakura, H.
Year: 1993
Title: Structural analysis on the sugar chains of human alpha 1-antitrypsin: presence of fucosylated biantennary glycan in hepatocellular carcinoma
Journal: Arch Biochem Biophys
Volume: 303
Issue: 2
Pages: 281-7
Epub Date: 1993/06/01
Date: Jun
Short Title: Structural analysis on the sugar chains of human alpha 1-antitrypsin: presence of fucosylated biantennary glycan in hepatocellular carcinoma
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1993.1284
Accession Number: 8390218
Keywords: Carbohydrate Metabolism
Carbohydrate Sequence
Carbohydrates/*chemistry
Carcinoma, Hepatocellular/*chemistry
Chromatography, High Pressure Liquid
Fucose/*metabolism
Humans
Lectins/metabolism
Liver Neoplasms/*chemistry
Molecular Sequence Data
*Plant Lectins
Polysaccharides/*analysis
alpha 1-Antitrypsin/*chemistry/metabolism
alpha-L-Fucosidase/metabolism
Abstract: Chemical structures of the sugar chains of alpha 1-antitrypsin (AAT) from patients with hepatocellular carcinoma (HCC) and from healthy individuals with a different affinity for Lens culinaris agglutinin (LCA) were examined by pyridylamination of their oligosaccharides and stepwise exoglycosidase digestion in combination with reversed-phase and size-fractionation high-performance liquid chromatography. We found that the LCA-reactive species of AAT from patients with HCC carried both the biantennary sugar chain with a fucose residue at the innermost N-acetylglucosamine residue, Gal beta 1-4GlcNAc beta 1-2Man alpha 1-6(Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4(Fuc alpha 1-6)GlcNAc-PA, and the biantennary chain without a fucose residue, Gal beta 1-4GlcNAc beta 1-2Man alpha 1-6(Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4GlcNAc-PA, at a ratio of about 1:0.6. The LCA-nonreactive species of AAT contained the biantennary sugar chain Gal beta 1-4GlcNAc beta 1-2Man alpha 1-6(Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4GlcNAc as a major component. These results indicate that a characteristic feature of the carbohydrate chains of AAT from patients with HCC is an increment in fucosylation.
Notes: Saitoh, A
Aoyagi, Y
Asakura, H
Journal Article
United States
Arch Biochem Biophys. 1993 Jun;303(2):281-7. doi: 10.1006/abbi.1993.1284.
Author Address: Department of Internal Medicine, Niigata University School of Medicine, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1337
Author: Sakabe, K. and Hart, G. W.
Year: 2010
Title: O-GlcNAc transferase regulates mitotic chromatin dynamics
Journal: J Biol Chem
Volume: 285
Issue: 45
Pages: 34460-8
Epub Date: 2010/09/02
Date: Nov 5
Short Title: O-GlcNAc transferase regulates mitotic chromatin dynamics
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.158170
PMCID: PMC2966060
Accession Number: 20805223
Keywords: Chromatin/*enzymology/genetics
Chromatin Assembly and Disassembly/*physiology
Chromosome Segregation/*physiology
Chromosomes, Human/genetics/metabolism
DNA Breaks
Gene Expression Regulation, Enzymologic/physiology
Genome, Human/physiology
HeLa Cells
Histones/genetics/metabolism
Humans
Mitosis/*physiology
N-Acetylglucosaminyltransferases/genetics/*metabolism
Protein Processing, Post-Translational/*physiology
Protein Transport/physiology
Protein-Arginine N-Methyltransferases
Spindle Apparatus/enzymology/genetics
Abstract: Mitosis must faithfully divide the genome such that each progeny inherits the same genetic material. DNA condensation is crucial in ensuring that chromosomes are correctly attached to the mitotic spindle for segregation, preventing DNA breaks or constrictions from the contractile ring. Histones form an octameric complex of basic proteins important in regulating DNA organization and accessibility. Histone post-translational modifications are altered during mitosis, although the roles of these post-translational modifications remain poorly characterized. Here, we report that N-acetylglucosamine (O-GlcNAc) transferase (OGT), the enzyme catalyzing the addition of O-GlcNAc moieties to nuclear and cytoplasmic proteins at serine and threonine residues, regulates some aspects of mitotic chromatin dynamics. OGT protein amounts decrease during M phase. Modest overexpression of OGT alters mitotic histone post-translational modifications at Lys-9, Ser-10, Arg-17, and Lys-27 of histone H3. Overexpression of OGT also prevents mitotic phosphorylation of coactivator-associated arginine methyltransferase 1 (CARM1) and prevents its correct cellular localization during mitosis. Moreover, OGT overexpression results in an increase in abnormal chromosomal bridge formation. Together, these results show that regulating the amount of OGT during mitosis is important in ensuring correct chromosomal segregation during mitosis.
Notes: 1083-351x
Sakabe, Kaoru
Hart, Gerald W
P30 DK079637/DK/NIDDK NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
R01 CA2486/CA/NCI NIH HHS/United States
R01 DK61671/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2010 Nov 5;285(45):34460-8. doi: 10.1074/jbc.M110.158170. Epub 2010 Aug 30.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1354
Author: Sakabe, K., Wang, Z. and Hart, G. W.
Year: 2010
Title: Beta-N-acetylglucosamine (O-GlcNAc) is part of the histone code
Journal: Proc Natl Acad Sci U S A
Volume: 107
Issue: 46
Pages: 19915-20
Epub Date: 2010/11/04
Date: Nov 16
Short Title: Beta-N-acetylglucosamine (O-GlcNAc) is part of the histone code
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1009023107
PMCID: PMC2993388
Accession Number: 21045127
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Chromatin/metabolism
Glycosylation
HeLa Cells
Heat-Shock Response
*Histone Code
Histones/chemistry/metabolism
Humans
Mass Spectrometry
Models, Molecular
Molecular Sequence Data
N-Acetylglucosaminyltransferases/metabolism
Reproducibility of Results
Abstract: Dynamic posttranslational modification of serine and threonine residues of nucleocytoplasmic proteins by β-N-acetylglucosamine (O-GlcNAc) is a regulator of cellular processes such as transcription, signaling, and protein-protein interactions. Like phosphorylation, O-GlcNAc cycles in response to a wide variety of stimuli. Although cycling of O-GlcNAc is catalyzed by only two highly conserved enzymes, O-GlcNAc transferase (OGT), which adds the sugar, and β-N-acetylglucosaminidase (O-GlcNAcase), which hydrolyzes it, the targeting of these enzymes is highly specific and is controlled by myriad interacting subunits. Here, we demonstrate by multiple specific immunological and enzymatic approaches that histones, the proteins that package DNA within the nucleus, are O-GlcNAcylated in vivo. Histones also are substrates for OGT in vitro. We identify O-GlcNAc sites on histones H2A, H2B, and H4 using mass spectrometry. Finally, we show that histone O-GlcNAcylation changes during mitosis and with heat shock. Taken together, these data show that O-GlcNAc cycles dynamically on histones and can be considered part of the histone code.
Notes: 1091-6490
Sakabe, Kaoru
Wang, Zihao
Hart, Gerald W
R01 DK61671/DK/NIDDK NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
R01 CA42486/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Proc Natl Acad Sci U S A. 2010 Nov 16;107(46):19915-20. doi: 10.1073/pnas.1009023107. Epub 2010 Nov 2.
Author Address: Department of Biological Chemistry, The Johns Hopkins University, Baltimore, MD 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 800
Author: Salaün, C., Rodrigues, P. and Heard, J. M.
Year: 2001
Title: Transmembrane topology of PiT-2, a phosphate transporter-retrovirus receptor
Journal: J Virol
Volume: 75
Issue: 12
Pages: 5584-92
Epub Date: 2001/05/18
Date: Jun
Short Title: Transmembrane topology of PiT-2, a phosphate transporter-retrovirus receptor
Alternate Journal: Journal of virology
ISSN: 0022-538X (Print)
0022-538x
DOI: 10.1128/jvi.75.12.5584-5592.2001
PMCID: PMC114271
Accession Number: 11356966
Keywords: Amino Acid Sequence
Animals
Asparagine/metabolism
Carrier Proteins/*chemistry/*metabolism
Computational Biology
Epitopes
Glycosylation
Humans
Mice
Models, Biological
Molecular Sequence Data
Mutation
Precipitin Tests
Protein Biosynthesis
Rats
Receptors, Virus/*metabolism
Retroviridae/*metabolism
Sodium-Phosphate Cotransporter Proteins
Sodium-Phosphate Cotransporter Proteins, Type III
*Symporters
Abstract: PiT-1 and PiT-2 are related multiple transmembrane proteins which function as sodium-dependent phosphate transporters and as the cell receptors of several oncoretroviruses. Two copies of a homology domain that is found in distantly related species assign these proteins to a large family of phosphate transporters. A current membrane topology model of PiT-1 and PiT-2 predicts 10 transmembrane domains. However, the validity of this model has not been addressed experimentally. We addressed this issue by a comprehensive study of human PiT-2. Evidence was obtained for glycosylation of asparagine 81. Epitope tagging showed that the N- and C-terminal extremities are extracellular. The orientation of C-terminal-truncation mutants expressed in cell-free translation assays and incorporated into microsomal membranes was examined by immunoprecipitation. Data were interpreted with respect to previous knowledge about retrovirus binding sites, to the existence of repeated homology domains, and to predictions made in family members. A model in which PiT-2 has 12 transmembrane domains and extracellular N- and C-terminal extremities is proposed. This model, which differs significantly from previous predictions about PiT-2 topology, may be useful for further investigations of PiT-2 interactions with other proteins and for the understanding of PiT-2 transporter and virus receptor functions.
Notes: 1098-5514
Salaün, C
Rodrigues, P
Heard, J M
Journal Article
Research Support, Non-U.S. Gov't
J Virol. 2001 Jun;75(12):5584-92. doi: 10.1128/JVI.75.12.5584-5592.2001.
Author Address: Laboratoire Rétrovirus et Transfert Génétique, CNRS URA 1930, Institut Pasteur, 75724 Paris, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 597
Author: Sambasivam, H., Rassouli, M., Murray, R. K., Nagpurkar, A., Mookerjea, S., Azadi, P., Dell, A. and Morris, H. R.
Year: 1993
Title: Studies on the carbohydrate moiety and on the biosynthesis of rat C-reactive protein
Journal: J Biol Chem
Volume: 268
Issue: 14
Pages: 10007-16
Epub Date: 1993/05/15
Date: May 15
Short Title: Studies on the carbohydrate moiety and on the biosynthesis of rat C-reactive protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8486673
Keywords: Animals
Asparagine
C-Reactive Protein/*biosynthesis/*chemistry/genetics
Carbohydrate Conformation
Carbohydrate Sequence
Cell-Free System
Electrophoresis, Polyacrylamide Gel
Glycoproteins/*biosynthesis/*chemistry
Macromolecular Substances
Molecular Sequence Data
Molecular Weight
Oligosaccharides/chemical synthesis/isolation & purification
Protein Biosynthesis
Protein Processing, Post-Translational
Rabbits
Rats
Reticulocytes/metabolism
Sialic Acids/analysis
Spectrometry, Mass, Fast Atom Bombardment
Abstract: Rat C-reactive protein (CRP) is a pentameric glycoprotein composed of five apparently identical monomers, two of which form a disulfide-linked dimer (Rasosouli, M., Sambasivam, H., Azadi, P., Dell, A., Morris, H. R., Nagpurkar, A., Mookerjea, S., and Murray, R. K. (1992) J. Biol. Chem. 267, 2947-2954). In this study, the nature of the oligosaccharide chain of rat CRP was investigated by fast atom bombardment-mass spectrometry (FAB-MS), and general features of its biosynthetic pathway were also analyzed. FAB-MS, electrospray-mass spectrometry, and linkage analysis demonstrated that each monomer of rat CRP contained one oligosaccharide chain, predominantly a disialylated biantennary structure, attached to Asn-128. The biosynthesis of rat CRP was studied by immunoprecipitation of CRP synthesized in vitro and by cultured hepatocytes. The results revealed that each monomer of rat CRP was synthesized individually as a single-chain precursor with a cleavable signal sequence. The translocated species was sensitive to digestion by endoglycosidase H (endo H), indicating that it possessed a high mannose oligosaccharide. Rat CRP acquired the ability to bind to phosphorylcholine-Sepharose and to form the dimeric and oligomeric species prior to acquiring resistance to endo H. Studies using tunicamycin revealed that the N-linked oligosaccharide present in rat CRP was not required for formation of its dimeric component, oligomerization, ability to bind to phosphorylcholine, or secretion. The non-glycosylated rat CRP, however, was still able to bind to phosphorylcholine-Sepharose and to be secreted by hepatocytes.
Notes: Sambasivam, H
Rassouli, M
Murray, R K
Nagpurkar, A
Mookerjea, S
Azadi, P
Dell, A
Morris, H R
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1993 May 15;268(14):10007-16.
Author Address: Department of Biochemistry, University of Toronto, Ontario, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 347
Author: Samor, B., Michalski, J. C., Mazurier, C., Goudemand, M., De Waard, P., Vliegenthart, J. F., Strecker, G. and Montreuil, J.
Year: 1989
Title: Primary structure of the major O-glycosidically linked carbohydrate unit of human von Willebrand factor
Journal: Glycoconj J
Volume: 6
Issue: 3
Pages: 263-70
Epub Date: 1989/01/01
Short Title: Primary structure of the major O-glycosidically linked carbohydrate unit of human von Willebrand factor
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/bf01047846
Accession Number: 2535488
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Chromatography, Gel
Chromatography, Thin Layer
Glycoproteins/*chemistry/isolation & purification
Glycosides
Humans
Lectins
Magnetic Resonance Spectroscopy
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
von Willebrand Factor/*chemistry/isolation & purification
Abstract: A reduced tetrasaccharide chain was obtained from human von Willebrand factor (vWF) by mild alkaline borohydride treatment. The purification of this O-glycosidically-linked oligosaccharide was achieved by serial affinity chromatography on immobilized concanavalin A and Lens culinaris agglutinin and finally gel filtration. Its structure was determined by a combination of methylation studies and 500 MHz 1H-NMR spectroscopy to be: NeuAc(alpha 2-3)Gal(beta 1-3)[NeuAc(alpha 2-6)]GalNAc-ol.
Notes: Samor, B
Michalski, J C
Mazurier, C
Goudemand, M
De Waard, P
Vliegenthart, J F
Strecker, G
Montreuil, J
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 1989;6(3):263-70. doi: 10.1007/BF01047846.
Author Address: Centre Régional de Transfusion Sanguine, laboratoire d'Hémostase, Lille, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 845
Author: Samyn-Petit, B., Gruber, V., Flahaut, C., Wajda-Dubos, J. P., Farrer, S., Pons, A., Desmaizieres, G., Slomianny, M. C., Theisen, M. and Delannoy, P.
Year: 2001
Title: N-glycosylation potential of maize: the human lactoferrin used as a model
Journal: Glycoconj J
Volume: 18
Issue: 7
Pages: 519-27
Epub Date: 2002/08/02
Date: Jul
Short Title: N-glycosylation potential of maize: the human lactoferrin used as a model
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1023/a:1019640312730
Accession Number: 12151713
Keywords: Blotting, Western
Glycosylation
Humans
Lactoferrin/*metabolism
Models, Chemical
Recombinant Proteins/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Zea mays/*metabolism
Abstract: In order to determine the N-glycosylation potential of maize, a monocotyledon expression system for the production of recombinant glycoproteins, human lactoferrin was used as a model. The human lactoferrin coding sequence was inserted into the pUC18 plasmid under control of the wheat glutenin promoter. Maize was stably transformed and recombinant lactoferrin was purified from the fourth generation seeds. Glycosylation was analysed by gas chromatography, lectin detection, glycosidase digestions and mass spectrometry. The results indicated that both N-glycosylation sites of recombinant lactoferrin are mainly substituted by typical plant paucimannose-type glycans, with beta1,2-xylose and alpha1,3-linked fucose at the proximal N-acetylglucosamine, and that complex-type glycans with Lewis(a) determinants are not present in maize recombinant lactoferrin.
Notes: Samyn-Petit, B
Gruber, V
Flahaut, C
Wajda-Dubos, J P
Farrer, S
Pons, A
Desmaizieres, G
Slomianny, M C
Theisen, M
Delannoy, P
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 2001 Jul;18(7):519-27. doi: 10.1023/a:1019640312730.
Author Address: Meristem Therapeutics, 8 rue des Frères Lumière, 63100 Clermont-Ferrand, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 712
Author: Sánchez, L. M., Chirino, A. J. and Bjorkman, Pj
Year: 1999
Title: Crystal structure of human ZAG, a fat-depleting factor related to MHC molecules
Journal: Science
Volume: 283
Issue: 5409
Pages: 1914-9
Epub Date: 1999/04/17
Date: Mar 19
Short Title: Crystal structure of human ZAG, a fat-depleting factor related to MHC molecules
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.283.5409.1914
Accession Number: 10206894
Keywords: Binding Sites
Crystallography, X-Ray
Glycoproteins/blood/*chemistry/isolation & purification/metabolism
Glycosylation
HLA-A2 Antigen/chemistry/metabolism
Histocompatibility Antigens Class I/*chemistry
Humans
Hydrogen Bonding
Ligands
Lipid Metabolism
Models, Molecular
Peptides/metabolism
Protein Binding
Protein Conformation
Protein Folding
Protein Structure, Secondary
Protein Structure, Tertiary
*Seminal Plasma Proteins
beta 2-Microglobulin/metabolism
Abstract: Zn-alpha2-glycoprotein (ZAG) is a soluble protein that is present in serum and other body fluids. ZAG stimulates lipid degradation in adipocytes and causes the extensive fat losses associated with some advanced cancers. The 2.8 angstrom crystal structure of ZAG resembles a class I major histocompatibility complex (MHC) heavy chain, but ZAG does not bind the class I light chain beta2-microglobulin. The ZAG structure includes a large groove analogous to class I MHC peptide binding grooves. Instead of a peptide, the ZAG groove contains a nonpeptidic compound that may be implicated in lipid catabolism under normal or pathological conditions.
Notes: Sánchez, L M
Chirino, A J
Bjorkman, P j
Journal Article
United States
Science. 1999 Mar 19;283(5409):1914-9. doi: 10.1126/science.283.5409.1914.
Author Address: Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1948
Author: Sanchez-Pupo, R. E., Johnston, D. and Penuela, S.
Year: 2018
Title: N-Glycosylation Regulates Pannexin 2 Localization but Is Not Required for Interacting with Pannexin 1
Journal: Int J Mol Sci
Volume: 19
Issue: 7
Epub Date: 2018/06/23
Date: Jun 22
Short Title: N-Glycosylation Regulates Pannexin 2 Localization but Is Not Required for Interacting with Pannexin 1
Alternate Journal: International journal of molecular sciences
ISSN: 1422-0067
DOI: 10.3390/ijms19071837
PMCID: PMC6073767
Accession Number: 29932112
Keywords: Animals
Asparagine/genetics/metabolism
Binding Sites/genetics
Cell Line
Connexins/genetics/*metabolism
Glycosylation
HEK293 Cells
Humans
Microscopy, Confocal
Mutation
Nerve Tissue Proteins/*metabolism
Protein Binding
*Protein Processing, Post-Translational
Protein Transport/genetics
Rats
N-glycosylation
Panx1
Panx2
channels
pannexin
post-translational modification
subcellular localization
traffic
Abstract: Pannexins (Panx1, 2, 3) are channel-forming glycoproteins expressed in mammalian tissues. We previously reported that N-glycosylation acts as a regulator of the localization and intermixing of Panx1 and Panx3, but its effects on Panx2 are currently unknown. Panx1 and Panx2 intermixing can regulate channel properties, and both pannexins have been implicated in neuronal cell death after ischemia. Our objectives were to validate the predicted N-glycosylation site of Panx2 and to study the effects of Panx2 glycosylation on localization and its capacity to interact with Panx1. We used site-directed mutagenesis, enzymatic de-glycosylation, cell-surface biotinylation, co-immunoprecipitation, and confocal microscopy. Our results showed that N86 is the only N-glycosylation site of Panx2. Panx2 and the N86Q mutant are predominantly localized to the endoplasmic reticulum (ER) and cis-Golgi matrix with limited cell surface localization was seen only in the presence of Panx1. The Panx2 N86Q mutant is glycosylation-deficient and tends to aggregate in the ER reducing its cell surface trafficking but it can still interact with Panx1. Our study indicates that N-glycosylation may be important for folding and trafficking of Panx2. We found that the un-glycosylated forms of Panx1 and 2 can readily interact, regulating their localization and potentially their channel function in cells where they are co-expressed.
Notes: 1422-0067
Sanchez-Pupo, Rafael E
Orcid: 0000-0002-6970-2137
Johnston, Danielle
Penuela, Silvia
Orcid: 0000-0003-4829-5517
Journal Article
Int J Mol Sci. 2018 Jun 22;19(7):1837. doi: 10.3390/ijms19071837.
Author Address: Department of Anatomy and Cell Biology, Schulich School of Medicine & Dentistry, University of Western Ontario, London, ON N6A5C1, Canada. rsnchezp@uwo.ca.
Department of Anatomy and Cell Biology, Schulich School of Medicine & Dentistry, University of Western Ontario, London, ON N6A5C1, Canada. Danielle.Johnston@schulich.uwo.ca.
Department of Anatomy and Cell Biology, Schulich School of Medicine & Dentistry, University of Western Ontario, London, ON N6A5C1, Canada. spenuela@uwo.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1121
Author: Sánchez-Sánchez, F., Martínez-Redondo, F., Aroca-Aguilar, J. D., Coca-Prados, M. and Escribano, J.
Year: 2007
Title: Characterization of the intracellular proteolytic cleavage of myocilin and identification of calpain II as a myocilin-processing protease
Journal: J Biol Chem
Volume: 282
Issue: 38
Pages: 27810-24
Epub Date: 2007/07/26
Date: Sep 21
Short Title: Characterization of the intracellular proteolytic cleavage of myocilin and identification of calpain II as a myocilin-processing protease
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M609608200
Accession Number: 17650508
Keywords: Calcium-Binding Proteins/chemistry
Calpain/*chemistry
Cell Line
Cytoskeletal Proteins/*chemistry
Dipeptides/chemistry
Egtazic Acid/chemistry/pharmacology
Endopeptidase K/chemistry
Eye Proteins/*chemistry
Glaucoma/metabolism
Glycoproteins/*chemistry
Humans
Ionomycin/pharmacology
Mutation, Missense
Protein Conformation
Protein Structure, Tertiary
Transfection
Abstract: MYOC, a gene involved in different types of glaucoma, encodes myocilin, a secreted glycoprotein of unknown function, consisting of an N-terminal leucine-zipper-like domain, a central linker region, and a C-terminal olfactomedin-like domain. Recently, we have shown that myocilin undergoes an intracellular endoproteolytic processing. We show herein that the proteolytic cleavage in the linker region splits the two terminal domains. The C-terminal domain is secreted to the culture medium, whereas the N-terminal domain mainly remains intracellularly retained. In transiently transfected 293T cells, the cleavage was prevented by calpain inhibitors, such as calpeptin, calpain inhibitor IV, and calpastatin. Since calpains are calcium-activated proteases, we analyzed how changes in either intra- or extracellular calcium affected the cleavage of myocilin. Intracellular ionomycin-induced calcium uptake enhanced myocilin cleavage, whereas chelation of extracellular calcium by EGTA inhibited the proteolytic processing. Calpains I and II cleaved myocilin in vitro. However, in cells in culture, only RNA interference knockdown of calpain II reduced myocilin processing. Subcellular fractionation and digestion of the obtained fractions with proteinase K showed that full-length myocilin resides in the lumen of the endoplasmic reticulum together with a subpopulation of calpain II. These data revealed that calpain II is responsible for the intracellular processing of myocilin in the lumen of the endoplasmic reticulum. We propose that this cleavage might regulate extracellular interactions of myocilin, contributing to the control of intraocular pressure.
Notes: Sánchez-Sánchez, Francisco
Martínez-Redondo, Francisco
Aroca-Aguilar, J Daniel
Coca-Prados, Miguel
Escribano, Julio
EY001785/EY/NEI NIH HHS/United States
EY04873/EY/NEI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2007 Sep 21;282(38):27810-24. doi: 10.1074/jbc.M609608200. Epub 2007 Jul 24.
Author Address: Area de Genética, Facultad de Medicina/Centro Regional de Investigaciones Biomédicas, Universidad de Castilla-La Mancha, Avda. de Almansa, no. 14, 02006 Albacete, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1109
Author: Sanders, J., Chirgadze, D. Y., Sanders, P., Baker, S., Sullivan, A., Bhardwaja, A., Bolton, J., Reeve, M., Nakatake, N., Evans, M., Richards, T., Powell, M., Miguel, R. N., Blundell, T. L., Furmaniak, J. and Smith, B. R.
Year: 2007
Title: Crystal structure of the TSH receptor in complex with a thyroid-stimulating autoantibody
Journal: Thyroid
Volume: 17
Issue: 5
Pages: 395-410
Epub Date: 2007/06/05
Date: May
Short Title: Crystal structure of the TSH receptor in complex with a thyroid-stimulating autoantibody
Alternate Journal: Thyroid : official journal of the American Thyroid Association
ISSN: 1050-7256 (Print)
1050-7256
DOI: 10.1089/thy.2007.0034
Accession Number: 17542669
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cricetinae
Cricetulus
Crystallization
Follicle Stimulating Hormone/chemistry
Humans
Immunoglobulin Fab Fragments/chemistry
Immunoglobulins, Thyroid-Stimulating/*chemistry
Molecular Sequence Data
Mutation
Receptors, FSH/chemistry
Receptors, Thyrotropin/*chemistry/genetics
X-Ray Diffraction
Abstract: OBJECTIVE: To analyze interactions between the thyroid-stimulating hormone receptor (TSHR) and a thyroid-stimulating human monoclonal autoantibody (M22) at the molecular level. DESIGN: A complex of part of the TSHR extracellular domain (amino acids 1-260; TSHR260) bound to M22 Fab was prepared and purified. Crystals suitable for X-ray diffraction analysis were obtained and the structure solved at 2.55 A resolution. MAIN OUTCOME: TSHR260 comprises of a curved helical tube and M22 Fab clasps its concave surface at 90 degrees to the tube length axis. The interface buried in the complex is large (2,500 A(2)) and an extensive network of ionic, polar, and hydrophobic bonding is involved in the interaction. There is virtually no movement in the atoms of M22 residues on the binding interface compared to unbound M22 consistent with "lock and key" binding. Mutation of residues showing strong interactions in the structure influenced M22 activity, indicating that the binding detail observed in the complex reflects interactions of M22 with intact, functionally active TSHR. The receptor-binding arrangements of the autoantibody are very similar to those reported for follicle-stimulating hormone (FSH) binding to the FSH receptor (amino acids 1-268) and consequently to those of TSH itself. CONCLUSIONS: It is remarkable that the thyroid-stimulating autoantibody shows almost identical receptor-binding features to TSH although the structures and origins of these two ligands are very different. Furthermore, our structure of the TSHR and its complex with M22 provide foundations for developing new strategies to understand and control both glycoprotein hormone receptor activation and the autoimmune response to the TSHR.
Notes: Sanders, Jane
Chirgadze, Dimitri Y
Sanders, Paul
Baker, Stuart
Sullivan, Andrew
Bhardwaja, Anshu
Bolton, Jane
Reeve, Magnus
Nakatake, Nobuhiro
Evans, Michele
Richards, Tonya
Powell, Michael
Miguel, Ricardo Núñez
Blundell, Tom L
Furmaniak, Jadwiga
Smith, Bernard Rees
Journal Article
United States
Thyroid. 2007 May;17(5):395-410. doi: 10.1089/thy.2007.0034.
Author Address: FIRS Laboratories, RSR Ltd., Llanishen, Cardiff, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 262
Author: Sandy, J. D., Flannery, C. R., Neame, P. J. and Lohmander, L. S.
Year: 1992
Title: The structure of aggrecan fragments in human synovial fluid. Evidence for the involvement in osteoarthritis of a novel proteinase which cleaves the Glu 373-Ala 374 bond of the interglobular domain
Journal: J Clin Invest
Volume: 89
Issue: 5
Pages: 1512-6
Epub Date: 1992/05/01
Date: May
Short Title: The structure of aggrecan fragments in human synovial fluid. Evidence for the involvement in osteoarthritis of a novel proteinase which cleaves the Glu 373-Ala 374 bond of the interglobular domain
Alternate Journal: The Journal of clinical investigation
ISSN: 0021-9738 (Print)
0021-9738
DOI: 10.1172/jci115742
PMCID: PMC443022
Accession Number: 1569188
Keywords: Aggrecans
Amino Acid Sequence
Cartilage, Articular/*metabolism
Endopeptidases/*metabolism
*Extracellular Matrix Proteins
Humans
Lectins, C-Type
Molecular Sequence Data
Osteoarthritis/*enzymology
Proteoglycans/*metabolism
Substrate Specificity
Synovial Fluid/metabolism
Abstract: Synovial fluid was collected from patients with recent knee injury and from patients with early or late stage osteoarthritis. Chondroitin sulfate-substituted aggrecan fragments present in these fluids, and in normal bovine synovial fluid, were purified by cesium chloride gradient centrifugation, enzymically deglycosylated and fractionated by gel filtration on Superose-12. Each sample contained two major aggrecan core protein populations with apparent molecular masses of approximately 90 kD and 150 kD. For all samples, NH2-terminal analysis of both populations gave a single major sequence beginning ARGSV. This NH2 terminus results from cleavage of the human aggrecan core protein at the Glu 373-Ala 374 bond within the interglobular domain between the G1 and G2 domains. Cleavage at this site also occurs during control and interleukin-1 stimulated aggrecan catabolism in bovine cartilage explant cultures (Sandy, J., P. Neame, R. Boynton, and C. Flannery. 1991. J. Biol. Chem. 266:8683-8685). These results indicate that the major aggrecan fragments present in both osteoarthritic human synovial fluid and in normal bovine synovial fluid are large, being composed of a short NH2-terminal stretch of the interglobular domain, the G2 domain, the keratan sulfate domain, and variable lengths of the chondroitin sulfate domain(s). We conclude that the release of aggrecan fragments from articular cartilage into the synovial fluid seen at all stages of human osteoarthritis (Lohmander, L. S. 1991. Acta Orthop. Scand. 62:623-632) is promoted by the action of a normal cartilage proteinase which cleaves the Glu 373-Ala 374 bond of the interglobular domain.
Notes: Sandy, J D
Flannery, C R
Neame, P J
Lohmander, L S
AR 38580/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
J Clin Invest. 1992 May;89(5):1512-6. doi: 10.1172/JCI115742.
Author Address: Shriners Hospital for Crippled Children, Tampa, Florida 33612.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2018
Author: Sang, Q., Zhang, Z., Shi, J., Sun, X., Li, B., Yan, Z., Xue, S., Ai, A., Lyu, Q., Li, W., Zhang, J., Wu, L., Mao, X., Chen, B., Mu, J., Li, Q., Du, J., Sun, Q., Jin, L., He, L., Zhu, S., Kuang, Y. and Wang, L.
Year: 2019
Title: A pannexin 1 channelopathy causes human oocyte death
Journal: Sci Transl Med
Volume: 11
Issue: 485
Epub Date: 2019/03/29
Date: Mar 27
Short Title: A pannexin 1 channelopathy causes human oocyte death
Alternate Journal: Science translational medicine
ISSN: 1946-6234
DOI: 10.1126/scitranslmed.aav8731
Accession Number: 30918116
Keywords: Adenosine Triphosphate/metabolism
Adult
Animals
Cell Death/genetics
Cells, Cultured
Channelopathies/*genetics/metabolism/*pathology
Connexins/*genetics/metabolism
Female
Fertilization in Vitro
Glycosylation
Humans
Infertility, Female/*genetics/metabolism/*pathology
Male
Mice, Mutant Strains
Mice, Transgenic
*Mutation
Nerve Tissue Proteins/*genetics/metabolism
Oocytes/*metabolism/*pathology
Pedigree
Phenotype
Translational Medical Research
Abstract: Connexins and pannexins are two protein families that play an important role in cellular communication. Pannexin 1 (PANX1), one of the members of pannexin family, is a channel protein. It is glycosylated and forms three species, GLY0, GLY1, and GLY2. Here, we describe four independent families in which mutations in PANX1 cause familial or sporadic female infertility via a phenotype that we term "oocyte death." The mutations, which are associated with oocyte death, alter the PANX1 glycosylation pattern, influence the subcellular localization of PANX1 in cultured cells, and result in aberrant PANX1 channel activity, ATP release in oocytes, and mutant PANX1 GLY1. Overexpression of a patient-derived mutation in mice causes infertility, recapitulating the human oocyte death phenotype. Our findings demonstrate the critical role of PANX1 in human oocyte development, provide a genetic explanation for a subtype of infertility, and suggest a potential target for therapeutic intervention for this disease.
Notes: 1946-6242
Sang, Qing
Orcid: 0000-0002-9895-1394
Zhang, Zhihua
Orcid: 0000-0002-4725-883x
Shi, Juanzi
Orcid: 0000-0003-3918-820x
Sun, Xiaoxi
Orcid: 0000-0002-3366-5821
Li, Bin
Orcid: 0000-0002-6095-9219
Yan, Zheng
Orcid: 0000-0001-8966-6295
Xue, Songguo
Ai, Ai
Lyu, Qifeng
Li, Wei
Zhang, Jilin
Wu, Ling
Mao, Xiaoyan
Chen, Biaobang
Mu, Jian
Li, Qiaoli
Du, Jing
Sun, Qiang
Jin, Li
He, Lin
Zhu, Shujia
Kuang, Yanping
Orcid: 0000-0003-2985-5793
Wang, Lei
Orcid: 0000-0002-3400-0434
Case Reports
Journal Article
Research Support, Non-U.S. Gov't
United States
Sci Transl Med. 2019 Mar 27;11(485):eaav8731. doi: 10.1126/scitranslmed.aav8731.
Author Address: Zhongshan Hospital, State Key Laboratory of Genetic Engineering, Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, China. wangleiwanglei@fudan.edu.cn kuangyanp@126.com sangqing@fudan.edu.cn.
Zhongshan Hospital, State Key Laboratory of Genetic Engineering, Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, China.
Reproductive Medicine Center, Shaanxi Maternal and Child Care Service Center, Shaanxi 710069, China.
Shanghai Ji Ai Genetics and IVF Institute, Obstetrics and Gynecology Hospital, Fudan University, Shanghai 200011, China.
Reproductive Medicine Center, Shanghai Ninth Hospital, Shanghai Jiao Tong University, Shanghai 200011, China.
Reproductive Medicine Center, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai 2000120, China.
Institute of Neuroscience, Key Laboratory of Primate Neurobiology, CAS Center for Excellence in Brain Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China.
Shanghai Institute of Planned Parenthood Research, Shanghai 200011, China.
Bio-X Center, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders, Ministry of Education, Shanghai Jiao Tong University, Shanghai 200030, China.
Reproductive Medicine Center, Shanghai Ninth Hospital, Shanghai Jiao Tong University, Shanghai 200011, China. wangleiwanglei@fudan.edu.cn kuangyanp@126.com sangqing@fudan.edu.cn.
Shanghai Center for Women and Children's Health, Shanghai 200062, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 654
Author: Santoro, B., Grant, S. G., Bartsch, D. and Kandel, E. R.
Year: 1997
Title: Interactive cloning with the SH3 domain of N-src identifies a new brain specific ion channel protein, with homology to eag and cyclic nucleotide-gated channels
Journal: Proc Natl Acad Sci U S A
Volume: 94
Issue: 26
Pages: 14815-20
Epub Date: 1998/02/07
Date: Dec 23
Short Title: Interactive cloning with the SH3 domain of N-src identifies a new brain specific ion channel protein, with homology to eag and cyclic nucleotide-gated channels
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.94.26.14815
PMCID: PMC25120
Accession Number: 9405696
Keywords: Amino Acid Sequence
Animals
Brain/*metabolism
Cloning, Molecular
Cyclic Nucleotide-Gated Cation Channels
Ether-A-Go-Go Potassium Channels
*Genes, src
Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels
Ion Channel Gating
Ion Channels/*genetics/isolation & purification
Mice
Molecular Sequence Data
Nerve Tissue Proteins/*genetics/isolation & purification
Nucleotides, Cyclic
Organ Specificity
Potassium Channels/*genetics
Sequence Alignment
src Homology Domains/*genetics
Abstract: We have isolated a novel cDNA, that appears to represent a new class of ion channels, by using the yeast two-hybrid system and the SH3 domain of the neural form of Src (N-src) as a bait. The encoded polypeptide, BCNG-1, is distantly related to cyclic nucleotide-gated channels and the voltage-gated channels, Eag and H-erg. BCNG-1 is expressed exclusively in the brain, as a glycosylated protein of approximately 132 kDa. Immunohistochemical analysis indicates that BCNG-1 is preferentially expressed in specific subsets of neurons in the neocortex, hippocampus, and cerebellum, in particular pyramidal neurons and basket cells. Within individual neurons, the BCNG-1 protein is localized to either the dendrites or the axon terminals depending on the cell type. Southern blot analysis shows that several other BCNG-related sequences are present in the mouse genome, indicating the emergence of an entire subfamily of ion channel coding genes. These findings suggest the existence of a new type of ion channel, which is potentially able to modulate membrane excitability in the brain and could respond to regulation by cyclic nucleotides.
Notes: 1091-6490
Santoro, B
Grant, S G
Bartsch, D
Kandel, E R
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 1997 Dec 23;94(26):14815-20. doi: 10.1073/pnas.94.26.14815.
Author Address: Center for Neurobiology and Behavior, College of Physicians and Surgeons of Columbia University, and Howard Hughes Medical Institute, 722 West 168th Street, New York, NY 10032, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1679
Author: Sanyal, A., Chen, A. J., Nakayasu, E. S., Lazar, C. S., Zbornik, E. A., Worby, C. A., Koller, A. and Mattoo, S.
Year: 2015
Title: A novel link between Fic (filamentation induced by cAMP)-mediated adenylylation/AMPylation and the unfolded protein response
Journal: J Biol Chem
Volume: 290
Issue: 13
Pages: 8482-99
Epub Date: 2015/01/21
Date: Mar 27
Short Title: A novel link between Fic (filamentation induced by cAMP)-mediated adenylylation/AMPylation and the unfolded protein response
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M114.618348
PMCID: PMC4375499
Accession Number: 25601083
Keywords: Activating Transcription Factor 6/metabolism
Apoptosis
Carrier Proteins/*physiology
Cell Survival
Endoplasmic Reticulum/metabolism
Glycosylation
HEK293 Cells
HeLa Cells
Heat-Shock Proteins/metabolism
Humans
Hydrophobic and Hydrophilic Interactions
MCF-7 Cells
Membrane Proteins/*physiology
Nucleotidyltransferases/*physiology
*Protein Processing, Post-Translational
Protein Structure, Tertiary
Protein Transport
Signal Transduction
*Unfolded Protein Response
Up-Regulation
eIF-2 Kinase/metabolism
Amp
AMPylation
Adenylylation
BiP
Filamentation Induced by cAMP
HYPE/FicD
Post-translational Modification
Signaling
Unfolded Protein Response (UPR)
Abstract: The maintenance of endoplasmic reticulum (ER) homeostasis is a critical aspect of determining cell fate and requires a properly functioning unfolded protein response (UPR). We have discovered a previously unknown role of a post-translational modification termed adenylylation/AMPylation in regulating signal transduction events during UPR induction. A family of enzymes, defined by the presence of a Fic (filamentation induced by cAMP) domain, catalyzes this adenylylation reaction. The human genome encodes a single Fic protein, called HYPE (Huntingtin yeast interacting protein E), with adenylyltransferase activity but unknown physiological target(s). Here, we demonstrate that HYPE localizes to the lumen of the endoplasmic reticulum via its hydrophobic N terminus and adenylylates the ER molecular chaperone, BiP, at Ser-365 and Thr-366. BiP functions as a sentinel for protein misfolding and maintains ER homeostasis. We found that adenylylation enhances BiP's ATPase activity, which is required for refolding misfolded proteins while coping with ER stress. Accordingly, HYPE expression levels increase upon stress. Furthermore, siRNA-mediated knockdown of HYPE prevents the induction of an unfolded protein response. Thus, we identify HYPE as a new UPR regulator and provide the first functional data for Fic-mediated adenylylation in mammalian signaling.
Notes: 1083-351x
Sanyal, Anwesha
Chen, Andy J
Nakayasu, Ernesto S
Lazar, Cheri S
Zbornik, Erica A
Worby, Carolyn A
Koller, Antonius
Mattoo, Seema
R01 GM100092/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2015 Mar 27;290(13):8482-99. doi: 10.1074/jbc.M114.618348. Epub 2015 Jan 19.
Author Address: From the Department of Biological Sciences and.
Bindley Biosciences Center, Purdue University, West Lafayette, Indiana 47907.
the Department of Pharmacology, University of California at San Diego, La Jolla, California 92093, and.
the Department of Pathology, Stony Brook University, Stony Brook, New York 11794.
From the Department of Biological Sciences and smattoo@purdue.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1750
Author: Saraswat, M., Joenväärä, S., Tomar, A. K., Singh, S., Yadav, S. and Renkonen, R.
Year: 2016
Title: N-Glycoproteomics of Human Seminal Plasma Glycoproteins
Journal: J Proteome Res
Volume: 15
Issue: 3
Pages: 991-1001
Epub Date: 2016/01/23
Date: Mar 4
Short Title: N-Glycoproteomics of Human Seminal Plasma Glycoproteins
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/acs.jproteome.5b01069
Accession Number: 26791533
Keywords: Adult
Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Gene Ontology
Glycoproteins/chemistry/*metabolism
Glycosylation
Humans
Male
Polysaccharides/chemistry/metabolism
Proteome/chemistry/*metabolism
Proteomics
Semen/*metabolism
Young Adult
Cid-ms/ms
DC-SIGN ligands
GlycopeptideId
He-4
Lewis x
Lewis y
N-glycoproteomics
clusterin
heparin-affinity chromatography
human seminal plasma
Abstract: Seminal plasma aids sperm by inhibiting premature capacitation, helping in the intracervical transport and formation of an oviductal sperm reservoir, all of which appear to be important in the fertilization process. Epitopes such as Lewis x and y are known to be present on seminal plasma glycoproteins, which can modulate the maternal immune response. It is suggested by multiple studies that seminal plasma glycoproteins play, largely undiscovered, important roles in the process of fertilization. We have devised a strategy to analyze glycopeptides from a complex, unknown mixture of protease-digested proteins. This analysis provides identification of the glycoproteins, glycosylation sites, glycan compositions, and proposed structures from the original sample. This strategy has been applied to human seminal plasma total glycoproteins. We have elucidated glycan compositions and proposed structures for 243 glycopeptides belonging to 73 N-glycosylation sites on 50 glycoproteins. The majority of the proposed glycan structures were complex type (83%) followed by high-mannose (10%) and then hybrid (7%). Most of the glycoproteins were either sialylated, fucosylated, or both. Many Lewis x/a and y/b epitopes bearing glycans were found, suggesting immune-modulating epitopes on multiple seminal plasma glycoproteins. The study also shows that large scale N-glycosylation mapping is achievable with current techniques and the depth of the analysis is roughly proportional to the prefractionation and complexity of the sample.
Notes: 1535-3907
Saraswat, Mayank
Joenväärä, Sakari
Tomar, Anil Kumar
Singh, Sarman
Yadav, Savita
Renkonen, Risto
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2016 Mar 4;15(3):991-1001. doi: 10.1021/acs.jproteome.5b01069. Epub 2016 Feb 4.
Author Address: Transplantation Laboratory, Haartman Institute, University of Helsinki , Haartmaninkatu 3, P.O. Box 21, Helsinki FI-00014, Finland.
HUSLAB, Helsinki University Hospital, 00290 Helsinki, Finland.
Department of Biophysics, All India Institute of Medical Sciences , New Delhi 110029, India.
Division of Clinical Microbiology & Molecular Medicine, Department of Laboratory Medicine, All India Institute of Medical Sciences , New Delhi 110029, India.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1950
Author: Saraswat, M., Mäkitie, A., Tohmola, T., Dickinson, A., Saraswat, S., Joenväärä, S. and Renkonen, S.
Year: 2018
Title: Tongue Cancer Patients Can be Distinguished from Healthy Controls by Specific N-Glycopeptides Found in Serum
Journal: Proteomics Clin Appl
Volume: 12
Issue: 6
Pages: e1800061
Epub Date: 2018/07/12
Date: Nov
Short Title: Tongue Cancer Patients Can be Distinguished from Healthy Controls by Specific N-Glycopeptides Found in Serum
Alternate Journal: Proteomics. Clinical applications
ISSN: 1862-8346
DOI: 10.1002/prca.201800061
Accession Number: 29992770
Keywords: Adult
Aged
Biomarkers, Tumor/*blood
*Early Detection of Cancer
Female
Glycopeptides/*blood
Glycosylation
Haptoglobins/metabolism
Healthy Volunteers
Humans
Immunoglobulin G/blood
Male
Middle Aged
Neoplasm Staging
Polysaccharides/chemistry
Protein Disulfide-Isomerases/blood
Tongue Neoplasms/*blood
Transferrin/metabolism
*Hnscc
*IgG
*N-glycoproteomics
*Oscc
*transferrin
Abstract: PURPOSE: There are no blood biomarkers to detect early-stage oral cavity squamous cell carcinoma (OSCC) prior to clinical signs. Most OSCC incidence is associated with significant morbidity and poor survival. The authors aimed to use mass-spectrometry (MS) technology to find specific N-glycopeptides potentially serving as serum biomarkers for preclinical OSCC screening. EXPERIMENTAL DESIGN: Serum samples from 14 patients treated for OSCC (stage I or stage IV) with 12 age- and sex-matched controls are collected. Quantitative label-free N-glycoproteomics is performed, with MS/MS analysis of the statistically significantly different N-glycopeptides. RESULTS: Combined with a database search using web-based software (GlycopeptideID), MS/MS provided detailed N-glycopeptide information, including glycosylation site, glycan composition, and proposed structures. Thirty-eight tryptic N-glycopeptides are identified, having 19 unique N-glycosylation sites representing 14 glycoproteins. OSCC patients, including stage I tumors, can be differentiated from healthy controls based on the expression levels of these glycoforms. N-glycopeptides of IgG1, IgG4, haptoglobin, and transferrin have statistically significant different abundances between cases and controls. CONCLUSIONS AND CLINICAL RELEVANCE: The authors are the first to suggest specific N-glycopeptides to serve as potential serum biomarkers to detect preclinical OSCC in patients. These N-glycopeptides are the lead candidates for validation as future diagnostic modalities of OSCC as early as stage I.
Notes: 1862-8354
Saraswat, Mayank
Mäkitie, Antti
Tohmola, Tiialotta
Dickinson, Amy
Saraswat, Shruti
Joenväärä, Sakari
Renkonen, Suvi
Korvatautien tutkimussäätiö/International
Jane and Aatos Erkko foundation/International
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics Clin Appl. 2018 Nov;12(6):e1800061. doi: 10.1002/prca.201800061. Epub 2018 Jul 18.
Author Address: Transplantation Laboratory, University of Helsinki, Haartmaninkatu 3, P.O. Box 21, 00014, Helsinki, Finland.
Hospital District of Helsinki and Uusimaa Laboratory, Helsinki University Hospital, 00290, Helsinki, Finland.
Department of Otorhinolaryngology-Head and Neck Surgery, University of Helsinki and Helsinki University Hospital, 00130, Helsinki, Finland.
Division of Ear, Nose and Throat Diseases, Department of Clinical Sciences, Intervention and Technology, Karolinska Institutet and Karolinska Hospital, 11382, Stockholm, Sweden.
Department of Biosciences, University of Helsinki, P.O. Box 65, 00014, Helsinki, Finland.
Department of Biosciences and Nutrition, Karolinska Institutet, 11382, Stockholm, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1322
Author: Sarrazin, S., Lyon, M., Deakin, J. A., Guerrini, M., Lassalle, P., Delehedde, M. and Lortat-Jacob, H.
Year: 2010
Title: Characterization and binding activity of the chondroitin/dermatan sulfate chain from Endocan, a soluble endothelial proteoglycan
Journal: Glycobiology
Volume: 20
Issue: 11
Pages: 1380-8
Epub Date: 2010/06/29
Date: Nov
Short Title: Characterization and binding activity of the chondroitin/dermatan sulfate chain from Endocan, a soluble endothelial proteoglycan
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwq100
Accession Number: 20581009
Keywords: Binding Sites
Cells, Cultured
Chondroitin/*metabolism
Chromatography, Gel
Dermatan Sulfate/*metabolism
Humans
Magnetic Resonance Spectroscopy
Proteoglycans/*metabolism
Abstract: Endocan is a recently identified soluble chondroitin/dermatan sulfate (CS/DS) proteoglycan. Synthesized by endothelial cells, it has been found to be over-expressed in the vasculature surrounding a number of tumors, and by promoting growth factor mitogenic activities, hepatocyte growth factor/scatter factor (HGF/SF) in particular, it supports cellular proliferation. In this work, we characterized the glycosaminoglycan (GAG) chain of Endocan, purified either from the naturally producing human umbilical vein endothelial cells (HUVEC) or from a recombinant over-expression system in human embryonic kidney cells (HEK). Compositional analysis using different chondroitinases as well as nuclear magnetic resonance studies revealed that the GAG chains from both sources share many characteristics, with the exception of size (15 and 40 kDa, respectively, for HUVEC and HEK-293 cells). The DS-specific, IdoA-containing disaccharides contribute 30% of the chain (15% of which are 2-O-sulfated) and are mostly clustered in tetra- (35%), hexa- (12%), and octa- (5%) saccharide domains. Highly sulfated D, E, and B disaccharide units (HexA2S-GalNAc6S, HexA-GalNAc4S6S, and HexA2S-GalNAc4S) were also detected in significant amounts in both chains and may account for the HGF/SF-binding activity of the CS/DS. This work establishes that HEK-293 cells can be engineered to provide a valuable source of Endocan with authentic CS/DS chains, enabling the purification of sufficient amounts for structural and/or binding analysis and providing a possible model of Endocan CS/DS chain organization.
Notes: 1460-2423
Sarrazin, Stéphane
Lyon, Malcolm
Deakin, Jon A
Guerrini, Marco
Lassalle, Philippe
Delehedde, Maryse
Lortat-Jacob, Hugues
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2010 Nov;20(11):1380-8. doi: 10.1093/glycob/cwq100. Epub 2010 Jun 24.
Author Address: Institut de Biologie Structurale, UMR 5075 CNRS-CEA-UJF, 41 Rue Horowitz, 38027 Grenoble, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 662
Author: Sasaki, T., Brakebusch, C., Engel, J. and Timpl, R.
Year: 1998
Title: Mac-2 binding protein is a cell-adhesive protein of the extracellular matrix which self-assembles into ring-like structures and binds beta1 integrins, collagens and fibronectin
Journal: Embo j
Volume: 17
Issue: 6
Pages: 1606-13
Epub Date: 1998/05/02
Date: Mar 16
Short Title: Mac-2 binding protein is a cell-adhesive protein of the extracellular matrix which self-assembles into ring-like structures and binds beta1 integrins, collagens and fibronectin
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/17.6.1606
PMCID: PMC1170508
Accession Number: 9501082
Keywords: Animals
Antigens, Neoplasm
Biomarkers, Tumor
Carcinoma, Squamous Cell
Carrier Proteins/chemistry/isolation & purification/*metabolism/ultrastructure
Cell Adhesion/physiology
Collagen/*metabolism
Extracellular Matrix Proteins/chemistry/isolation &
purification/*metabolism/ultrastructure
Fibronectins/*metabolism
Glycoproteins/chemistry/isolation & purification/*metabolism/ultrastructure
Humans
Integrin beta1/*metabolism
Ligands
Membrane Glycoproteins/analysis
Mice
Organ Specificity
Protein Conformation
Recombinant Proteins/isolation & purification
Sequence Analysis
Tumor Cells, Cultured
Abstract: Human Mac-2 binding protein (M2BP) was prepared in recombinant form from the culture medium of 293 kidney cells and consisted of a 92 kDa subunit. The protein was obtained in a native state as indicated by CD spectroscopy, demonstrating alpha-helical and beta-type structure, and by protease resistance and immunological analysis. It was highly modified by N- and O-glycosylation but not by glycosaminoglycans. Ultracentrifugation showed non-covalent association into oligomers with molar masses of 1000-1500 kDa. Electron microscopy showed ring-like shapes with diameters of 30-40 nm. M2BP bound in solid-phase assays to collagens IV, V and VI, fibronectin and nidogen, but not to fibrillar collagens I and III or other basement membrane proteins. The protein also mediated adhesion of cell lines at comparable strength with laminin. Adhesion to M2BP was inhibited by antibodies to integrin beta1 subunits but not to alpha2 and alpha6 subunits, RGD peptide or lactose. This distinguishes cell adhesion of M2BP from that of laminin and excludes involvement of lactose-binding galectin-3. Immunological assays demonstrated variable secretion by cultured human cells of M2BP, which was detected in the extracellular matrix of several mouse tissues.
Notes: 1460-2075
Sasaki, T
Brakebusch, C
Engel, J
Timpl, R
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1998 Mar 16;17(6):1606-13. doi: 10.1093/emboj/17.6.1606.
Author Address: Max-Planck-Institut f¿r Biochemie, D-82152 Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 663
Author: Sasaki, T., Hohenester, E., Göhring, W. and Timpl, R.
Year: 1998
Title: Crystal structure and mapping by site-directed mutagenesis of the collagen-binding epitope of an activated form of BM-40/SPARC/osteonectin
Journal: Embo j
Volume: 17
Issue: 6
Pages: 1625-34
Epub Date: 1998/05/02
Date: Mar 16
Short Title: Crystal structure and mapping by site-directed mutagenesis of the collagen-binding epitope of an activated form of BM-40/SPARC/osteonectin
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/17.6.1625
PMCID: PMC1170510
Accession Number: 9501084
Keywords: Alanine
Amino Acid Substitution
Binding Sites
Biosensing Techniques
Collagen/*metabolism
Crystallography, X-Ray
*Epitope Mapping
Epitopes/analysis
Humans
Models, Molecular
Osteonectin/*chemistry/genetics/metabolism
Protein Structure, Secondary
Sequence Deletion
Abstract: The extracellular calcium-binding domain (positions 138-286) of the matrix protein BM-40 possesses a binding epitope of moderate affinity for several collagen types. This epitope was predicted to reside in helix alphaA and to be partially masked by helix alphaC. Here we show that deletion of helix alphaC produces a 10-fold increase in collagen affinity similar to that seen after proteolytic cleavage of this helix. The predicted removal of the steric constraint was clearly demonstrated by the crystal structure of the mutant at 2.8 A resolution. This constitutively activated mutant was used to map the collagen-binding site following alanine mutagenesis at 13 positions. Five residues were crucial for binding, R149 and N156 in helix alphaA, and L242, M245 and E246 in a loop region connecting the two EF hands of BM-40. These residues are spatially close and form a flat ring of 15 A diameter which matches the diameter of a triple-helical collagen domain. The mutations showed similar effects on binding to collagens I and IV, indicating nearly identical binding sites on both collagens. Selected mutations in the non-activated mutant DeltaI also reduced collagen binding, consistent with the same location of the epitope but in a more cryptic form in intact BM-40.
Notes: 1460-2075
Sasaki, T
Hohenester, E
Göhring, W
Timpl, R
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1998 Mar 16;17(6):1625-34. doi: 10.1093/emboj/17.6.1625.
Author Address: Max-Planck-Institut für Biochemie, D-82152, Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1021
Author: Sasaki, T., Knyazev, P. G., Clout, N. J., Cheburkin, Y., Göhring, W., Ullrich, A., Timpl, R. and Hohenester, E.
Year: 2006
Title: Structural basis for Gas6-Axl signalling
Journal: Embo j
Volume: 25
Issue: 1
Pages: 80-7
Epub Date: 2005/12/20
Date: Jan 11
Short Title: Structural basis for Gas6-Axl signalling
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7600912
PMCID: PMC1356355
Accession Number: 16362042
Keywords: Amino Acid Sequence
Binding Sites
Conserved Sequence
Crystallography, X-Ray
Dimerization
Humans
Intercellular Signaling Peptides and Proteins/*chemistry
Molecular Sequence Data
Oncogene Proteins/*chemistry
Protein Conformation
Proto-Oncogene Proteins
Receptor Protein-Tyrosine Kinases/*chemistry
Signal Transduction
Abstract: Receptor tyrosine kinases of the Axl family are activated by the vitamin K-dependent protein Gas6. Axl signalling plays important roles in cancer, spermatogenesis, immunity, and platelet function. The crystal structure at 3.3 A resolution of a minimal human Gas6/Axl complex reveals an assembly of 2:2 stoichiometry, in which the two immunoglobulin-like domains of the Axl ectodomain are crosslinked by the first laminin G-like domain of Gas6, with no direct Axl/Axl or Gas6/Gas6 contacts. There are two distinct Gas6/Axl contacts of very different size, both featuring interactions between edge beta-strands. Structure-based mutagenesis, protein binding assays and receptor activation experiments demonstrate that both the major and minor Gas6 binding sites are required for productive transmembrane signalling. Gas6-mediated Axl dimerisation is likely to occur in two steps, with a high-affinity 1:1 Gas6/Axl complex forming first. Only the minor Gas6 binding site is highly conserved in the other Axl family receptors, Sky/Tyro3 and Mer. Specificity at the major contact is suggested to result from the segregation of charged and apolar residues to opposite faces of the newly formed beta-sheet.
Notes: 1460-2075
Sasaki, Takako
Knyazev, Pjotr G
Clout, Naomi J
Cheburkin, Yuri
Göhring, Walter
Ullrich, Axel
Timpl, Rupert
Hohenester, Erhard
054334/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2006 Jan 11;25(1):80-7. doi: 10.1038/sj.emboj.7600912. Epub 2005 Dec 15.
Author Address: Max-Planck-Institut für Biochemie, Martinsried, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1499
Author: Sato, B., Katagiri, Y. U., Iijima, K., Yamada, H., Ito, S., Kawasaki, N., Okita, H., Fujimoto, J. and Kiyokawa, N.
Year: 2012
Title: The human CD10 lacking an N-glycan at Asn(628) is deficient in surface expression and neutral endopeptidase activity
Journal: Biochim Biophys Acta
Volume: 1820
Issue: 11
Pages: 1715-23
Epub Date: 2012/07/07
Date: Nov
Short Title: The human CD10 lacking an N-glycan at Asn(628) is deficient in surface expression and neutral endopeptidase activity
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbagen.2012.06.017
Accession Number: 22766194
Keywords: Glycoside Hydrolases/pharmacology
Glycosylation
HEK293 Cells
Humans
Membrane Microdomains/chemistry
Neprilysin/analysis/*chemistry/*physiology
Abstract: BACKGROUND: CD10, also known as neprilysin or enkephalinase exhibiting neutral endopeptidase (NEP) activity, is expressed by B-lineage hematopoietic cells as well as a variety of cells from normal tissues. It cleaves peptides such as cytokines to act for terminating inflammatory responses. Although CD10 molecules of the human pre-B-cell line NALM-6 have 6 consensus N-glycosylation sites, three of them are known to be N-glycosylated by X-ray crystallography. METHODS: In order to investigate the role of N-glycans in the full expression of NEP activity, we modified N-glycans by treatment of NALM6 cells with various glycosidases or alter each of the consensus N-glycosylation sites by generating site-directed mutagenesis and compared the NEP activities of the sugar-altered CD10 with those of intact CD10. RESULTS: CD10 of the human B-cell line NALM-6 was dominantly localized in raft microdomains and heterogeneously N-glycosylated. Although neither desialylation nor further degalactosylation caused defective NEP activity, removal of only a small part of N-glycans by treatment with glycopeptidase F under non-denaturing conditions decreased NEP activity completely. All of the three consensus sites of CD10 in HEK293 cells introduced with wild type-CD10 were confirmed to be N-glycosylated. Surface expression of N-glycan at Asn(628)-deleted CD10 by HEK293 cells was greatly decreased as well as it lost entire NEP activities. CONCLUSIONS: N-glycosylation at Asn(628) is essential not only for NEP activities, but also for surface expression. GENERAL SIGNIFICANCE: Quality control system does not allow dysfunctional ecto-type proteases to express on plasma membrane.
Notes: Sato, Ban
Katagiri, Yohko U
Iijima, Kazutoshi
Yamada, Hiroyuki
Ito, Satsuki
Kawasaki, Nana
Okita, Hajime
Fujimoto, Junichiro
Kiyokawa, Nobutaka
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2012 Nov;1820(11):1715-23. doi: 10.1016/j.bbagen.2012.06.017. Epub 2012 Jul 3.
Author Address: Department of Pediatric Hematology and Oncology Research, National Research Institute for Child Health and Development, Setagaya-ku, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 742
Author: Sato, C., Kim, J. H., Abe, Y., Saito, K., Yokoyama, S. and Kohda, D.
Year: 2000
Title: Characterization of the N-oligosaccharides attached to the atypical Asn-X-Cys sequence of recombinant human epidermal growth factor receptor
Journal: J Biochem
Volume: 127
Issue: 1
Pages: 65-72
Epub Date: 2000/03/25
Date: Jan
Short Title: Characterization of the N-oligosaccharides attached to the atypical Asn-X-Cys sequence of recombinant human epidermal growth factor receptor
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a022585
Accession Number: 10731668
Keywords: Amino Acid Sequence
Amino Acid Substitution/genetics
Animals
Asparagine/*chemistry/genetics/metabolism
CHO Cells
Carbohydrate Sequence
Cricetinae
Cysteine/*chemistry/genetics/metabolism
ErbB Receptors/biosynthesis/*chemistry/*genetics/metabolism
Glycosylation
Humans
Lysine/genetics/metabolism
Molecular Sequence Data
Oligosaccharides/*chemistry/metabolism
Recombinant Proteins/biosynthesis/*chemistry/metabolism
Transfection
Abstract: The extracellular domain of human EGF receptor (sEGFR) produced by CHO cells has been used in various biophysical studies to elucidate the molecular mechanism of EGF-induced receptor activation. We have found that the CHO sEGFR contains one oligosaccharide chain attached to an atypical N-glycosylation consensus sequence, Asn(32 )-X( 33 )-Cys(34 ). The oligosaccharide structure at Asn(32 ) is a mixture of the monosialo and asialo forms of a core fucosylated biantennary complex-type oligosaccharide. Deletion of this atypical glycosylation site by replacement of Asn(32 ) with lysine changed neither the expression nor function of the full length EGFR in CHO cells. The glycosylation at Asn(32 ) in CHO sEGFR was incomplete: 20% of Asn(32 ) remained unmodified. Thus, CHO sEGFR itself is heterogeneous with respect to the glycosylation at Asn(32 ), which may cause problems in biophysical studies. An attempt to remove the oligosaccharide at Asn(32 ) enzymatically did not succeed under nondenaturing conditions. Therefore, sEGFR with the mutation of Asn(32) -> Lys(32 )is useful for biophysical and biochemical studies, and, particularly, for X-ray crystallography.
Notes: Sato, C
Kim, J H
Abe, Y
Saito, K
Yokoyama, S
Kohda, D
Journal Article
England
J Biochem. 2000 Jan;127(1):65-72. doi: 10.1093/oxfordjournals.jbchem.a022585.
Author Address: Department of Structural Biology, Biomolecular Engineering Research Institute, Furuedai Suita, Osaka 565-0874, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 570
Author: Sato, T., Furukawa, K., Autero, M., Gahmberg, C. G. and Kobata, A.
Year: 1993
Title: Structural study of the sugar chains of human leukocyte common antigen CD45
Journal: Biochemistry
Volume: 32
Issue: 47
Pages: 12694-704
Epub Date: 1993/11/30
Date: Nov 30
Short Title: Structural study of the sugar chains of human leukocyte common antigen CD45
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00210a019
Accession Number: 8251489
Keywords: Asparagine/chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Electrophoresis, Paper
Glycoside Hydrolases/metabolism
Humans
Leukocyte Common Antigens/*chemistry
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification/metabolism
Sialic Acids
T-Lymphocytes/*immunology
beta-Galactosidase/metabolism
Abstract: The leukocyte cell surface glycoprotein CD45 is a protein tyrosine phosphatase and is involved in signal transduction mediated by the T cell antigen receptor. The asparagine-linked sugar chains were released as oligosaccharides from purified CD45 by hydrazinolysis. Approximately 6 mol of sugar chains was released from 1 mol of CD45. These sugar chains were converted to radioactive oligosaccharides by reduction with NaB3H4 and separated into neutral and acidic fractions by paper electrophoresis. All of the acidic oligosaccharides were converted to neutral ones by digestion with sialidase, indicating that they are sialyl derivatives. Binding of the sialylated oligosaccharides to an SNA-agarose column as well as methylation analysis revealed that the oligosaccharides have only alpha-2,6-linked sialic acid residues. The neutral and sialidase-treated acidic oligosaccharides were fractionated by serial lectin column chromatography followed by Bio-Gel P-4 column chromatography. Structural studies of each oligosaccharide by sequential exo- and endoglycosidase digestion and by methylation analysis revealed that CD45 contains mainly bi-, tri-, and tetraantennary complex-type sugar chains. About 46% of the tetraantennary complex-type sugar chains had the poly(N-acetyllactosamine) groups and 18% of the 2,4-branched triantennary complex-type sugar chains had the fucosyl N-acetyllactosamine group. A portion of the bi- and 2,4-branched triantennary complex-type sugar chains were bisected. In addition to these sugar chains, a small amount of high mannose-type and hybrid-type sugar chains were detected.
Notes: Sato, T
Furukawa, K
Autero, M
Gahmberg, C G
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1993 Nov 30;32(47):12694-704. doi: 10.1021/bi00210a019.
Author Address: Department of Biochemistry, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 965
Author: Satomi, Y., Shimonishi, Y., Hase, T. and Takao, T.
Year: 2004
Title: Site-specific carbohydrate profiling of human transferrin by nano-flow liquid chromatography/electrospray ionization mass spectrometry
Journal: Rapid Commun Mass Spectrom
Volume: 18
Issue: 24
Pages: 2983-8
Epub Date: 2004/11/13
Short Title: Site-specific carbohydrate profiling of human transferrin by nano-flow liquid chromatography/electrospray ionization mass spectrometry
Alternate Journal: Rapid communications in mass spectrometry : RCM
ISSN: 0951-4198 (Print)
0951-4198
DOI: 10.1002/rcm.1718
Accession Number: 15536627
Keywords: Amino Acid Sequence
Binding Sites
Carbohydrate Sequence
Chromatography, Liquid
Glycopeptides/analysis/chemistry
Glycosylation
Humans
Molecular Sequence Data
Spectrometry, Mass, Electrospray Ionization/*methods
Transferrin/*analysis/*chemistry
Abstract: Glycopeptides derived from a lysylendopeptidase digest of commercially available human transferrin were analyzed by nano-flow liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS), which permitted the carbohydrate profiles at Asn432 and Asn630 to be determined. Both are located in a well-known motif for N-glycosylation, Asn-Xaa-Ser/Thr. The contents of the carbohydrates at each site were significantly different from each other, and consisted of a variety of minor types of oligosaccharides in addition to the major one, a biantennary complex-type oligosaccharide. Nano-flow ESI tandem mass spectrometry (MS/MS) of the glycopeptides (Cys421-Lys433 and Ile619-Lys646) containing these two sites yielded predominantly ions originating from the non-reducing termini (oxonium ions) and reducing terminus, resulting from cleavage of the glycosidic bonds of the carbohydrate moieties; this permitted the structural read-out of a small minority of the carbohydrate moieties. In particular, the observation of oxonium ions at m/z 512.2 and 803.2 is useful for probing outer non-reducing terminal fucosylation, which represented carbohydrate structures consisting of Hex, dHex, and HexNAc, and NeuNAc, Hex, dHex, and HexNAc, respectively, from which the Lewis X structure (Galbeta1-4(Fucalpha1-3)GlcNAc) was readily deduced. Moreover, fucosylation at the reducing-terminal GlcNAc (Fucalpha1-6GlcNAc) specifically occurred at Asn630, as demonstrated by treatment of the glycopeptides with alpha1-3/4-L-fucosidase.
Notes: Satomi, Yoshinori
Shimonishi, Yasutsugu
Hase, Toshiharu
Takao, Toshifumi
Journal Article
Research Support, Non-U.S. Gov't
England
Rapid Commun Mass Spectrom. 2004;18(24):2983-8. doi: 10.1002/rcm.1718.
Author Address: Laboratory of Protein Profiling and Functional Proteomics, Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 962
Author: Saugstad, J. A., Roberts, J. A., Dong, J., Zeitouni, S. and Evans, R. J.
Year: 2004
Title: Analysis of the membrane topology of the acid-sensing ion channel 2a
Journal: J Biol Chem
Volume: 279
Issue: 53
Pages: 55514-9
Epub Date: 2004/10/27
Date: Dec 31
Short Title: Analysis of the membrane topology of the acid-sensing ion channel 2a
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M411849200
PMCID: PMC2928846
NIHMSID: NIHMS228519
Accession Number: 15504740
Keywords: Acid Sensing Ion Channels
Binding Sites
Cell Line
Cell Membrane/*metabolism
DNA, Complementary/metabolism
Electrophysiology
Glycosylation
Humans
Hydrogen-Ion Concentration
Immunoblotting
Immunohistochemistry
Ions
Membrane Proteins/*chemistry/metabolism
Models, Biological
Mutagenesis, Site-Directed
Mutation
Nerve Tissue Proteins/*chemistry/metabolism
Protein Conformation
Protein Structure, Tertiary
Sodium Channels/*chemistry/metabolism
Transfection
Abstract: Acid-sensing ion channels, or ASICs, are members of the amiloride-sensitive cationic channel superfamily that are predicted to have intracellular amino and carboxyl termini and two transmembrane domains connected by a large extracellular loop. This prediction comes from biochemical studies of the mammalian epithelial sodium channels where glycosylation mutants identified the extracellular regions of the channel and a combination of antibody sensitivity and protease action substantiated the intracellular nature of the amino and carboxyl termini. However, although there are highly conserved regions within the different cation channel family members, membrane topology prediction programs provide several alternative structures for the ASICs. Thus, we used glycosylation studies to define the actual membrane topology of the ASIC2a subtype. We deleted the five predicted endogenous asparagine-linked glycosylation sites (Asn-Xaa-(Ser/Thr)) at Asn-22, Asn-365, Asn-392, Asn-478, and Asn-487 to map the extracellular topology. We then introduced exogenous asparagine-linked glycosylation sites at Lys-4, Pro-37, Arg-63, Tyr-67, His-72, Ala-81, Tyr-414, Tyr-423, and Tyr-453 to define the transmembrane domain borders. Finally, we used cell permeabilization studies to confirm the intracellular amino termini of ASIC2a. The data show that Asn-365 and Asn-392 are extracellular and that the introduction of asparagine-linked glycosylation sites at His-72, Ala-81, Tyr-414, and Tyr-423 leads to an increase in molecular mass consistent with an extracellular apposition. In addition, heterologous expression of ASIC2a requires membrane permeabilization for antibody staining. These data confirm the membrane topology prediction that the ASIC2a subtype consists of intracellular amino and carboxyl termini and two transmembrane domains connected by a large extracellular loop.
Notes: 1083-351x
Saugstad, Julie A
Roberts, Jonathan A
Dong, Jin
Zeitouni, Suzanne
Evans, Richard J
WT_/Wellcome Trust/United Kingdom
R03 EY015267/EY/NEI NIH HHS/United States
R03 EY015267-01/EY/NEI NIH HHS/United States
R42 NS065515/NS/NINDS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2004 Dec 31;279(53):55514-9. doi: 10.1074/jbc.M411849200. Epub 2004 Oct 25.
Author Address: Robert S. Dow Neurobiology Laboratories, Legacy Research, 1225 N.E. 2nd Avenue, Portland, OR 97232, USA. JSaugstad@DowNeurobiology.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1407
Author: Savalas, L. R., Gasnier, B., Damme, M., Lübke, T., Wrocklage, C., Debacker, C., Jézégou, A., Reinheckel, T., Hasilik, A., Saftig, P. and Schröder, B.
Year: 2011
Title: Disrupted in renal carcinoma 2 (DIRC2), a novel transporter of the lysosomal membrane, is proteolytically processed by cathepsin L
Journal: Biochem J
Volume: 439
Issue: 1
Pages: 113-28
Epub Date: 2011/06/23
Date: Oct 1
Short Title: Disrupted in renal carcinoma 2 (DIRC2), a novel transporter of the lysosomal membrane, is proteolytically processed by cathepsin L
Alternate Journal: The Biochemical journal
ISSN: 0264-6021
DOI: 10.1042/bj20110166
Accession Number: 21692750
Keywords: Animals
Cathepsin L/genetics/*metabolism
Computational Biology
Electrophysiology
Fluorescent Antibody Technique, Indirect
HeLa Cells
Humans
Immunoblotting
Immunoprecipitation
Lysosome-Associated Membrane Glycoproteins/genetics/*metabolism
Membrane Transport Proteins/genetics/*metabolism
Mice
Neoplasm Proteins/genetics/*metabolism
Protein Binding
Xenopus
Abstract: DIRC2 (Disrupted in renal carcinoma 2) has been initially identified as a breakpoint-spanning gene in a chromosomal translocation putatively associated with the development of renal cancer. The DIRC2 protein belongs to the MFS (major facilitator superfamily) and has been previously detected by organellar proteomics as a tentative constituent of lysosomal membranes. In the present study, lysosomal residence of overexpressed as well as endogenous DIRC2 was shown by several approaches. DIRC2 is proteolytically processed into a N-glycosylated N-terminal and a non-glycosylated C-terminal fragment respectively. Proteolytic cleavage occurs in lysosomal compartments and critically depends on the activity of cathepsin L which was found to be indispensable for this process in murine embryonic fibroblasts. The cleavage site within DIRC2 was mapped between amino acid residues 214 and 261 using internal epitope tags, and is presumably located within the tentative fifth intralysosomal loop, assuming the typical MFS topology. Lysosomal targeting of DIRC2 was demonstrated to be mediated by a N-terminal dileucine motif. By disrupting this motif, DIRC2 can be redirected to the plasma membrane. Finally, in a whole-cell electrophysiological assay based on heterologous expression of the targeting mutant at the plasma membrane of Xenopus oocytes, the application of a complex metabolic mixture evokes an outward current associated with the surface expression of full-length DIRC2. Taken together, these data strongly support the idea that DIRC2 is an electrogenic lysosomal metabolite transporter which is subjected to and presumably modulated by limited proteolytic processing.
Notes: 1470-8728
Savalas, Lalu Rudyat Telly
Gasnier, Bruno
Damme, Markus
Lübke, Torben
Wrocklage, Christian
Debacker, Cécile
Jézégou, Adrien
Reinheckel, Thomas
Hasilik, Andrej
Saftig, Paul
Schröder, Bernd
Journal Article
Research Support, Non-U.S. Gov't
England
Biochem J. 2011 Oct 1;439(1):113-28. doi: 10.1042/BJ20110166.
Author Address: Biochemical Institute, Christian-Albrechts-University, Kiel, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1177
Author: Sayat, R., Leber, B., Grubac, V., Wiltshire, L. and Persad, S.
Year: 2008
Title: O-GlcNAc-glycosylation of beta-catenin regulates its nuclear localization and transcriptional activity
Journal: Exp Cell Res
Volume: 314
Issue: 15
Pages: 2774-87
Epub Date: 2008/07/01
Date: Sep 10
Short Title: O-GlcNAc-glycosylation of beta-catenin regulates its nuclear localization and transcriptional activity
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2008.05.017
Accession Number: 18586027
Keywords: Acetylglucosamine/*metabolism
Active Transport, Cell Nucleus/physiology
Carcinoma/metabolism
Cell Adhesion/genetics
Cell Line
Cell Line, Tumor
Cell Nucleus/*metabolism
Epithelial Cells/*metabolism
Gene Expression Regulation, Neoplastic
Glycosylation
Humans
Male
N-Acetylglucosaminyltransferases/metabolism
Prostate/*metabolism
Prostatic Neoplasms/metabolism
RNA, Messenger/metabolism
Signal Transduction
Transcriptional Activation/*genetics
Up-Regulation/physiology
beta Catenin/genetics/*metabolism
Abstract: Beta-catenin plays a role in intracellular adhesion and regulating gene expression. The latter role is associated with its oncogenic properties. Phosphorylation of beta-catenin controls its intracellular expression but mechanism/s that regulates the nuclear localization of beta-catenin is unknown. We demonstrate that O-GlcNAc glycosylation (O-GlcNAcylation) of beta-catenin negatively regulates its levels in the nucleus. We show that normal prostate cells (PNT1A) have significantly higher amounts of O-GlcNAcylated beta-catenin compared to prostate cancer (CaP) cells. The total nuclear levels of beta-catenin are higher in the CaP cells than PNT1A but only a minimal fraction of the nuclear beta-catenin in the CaP cells are O-GlcNAcylated. Increasing the levels of O-GlcNAcylated beta-catenin in the CaP cells with PUGNAc (O- (2-acetamido-2-deoxy-d-gluco-pyranosylidene) amino-N-phenylcarbamate) treatment is associated with a progressive decrease in the levels of beta-catenin in the nucleus. TOPFlash reporter assay and mRNA expressions of beta-catenin's target genes indicate that O-GlcNAcylation of beta-catenin results in a decrease in its transcriptional activity. We define a novel modification of beta-catenin that regulates its nuclear localization and transcriptional function.
Notes: 1090-2422
Sayat, Ria
Leber, Brian
Grubac, Vanja
Wiltshire, Lesley
Persad, Sujata
Journal Article
Research Support, Non-U.S. Gov't
United States
Exp Cell Res. 2008 Sep 10;314(15):2774-87. doi: 10.1016/j.yexcr.2008.05.017. Epub 2008 Jun 6.
Author Address: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, Canada L8N 3Z5.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1604
Author: Scally, S. W., Petersen, J., Law, S. C., Dudek, N. L., Nel, H. J., Loh, K. L., Wijeyewickrema, L. C., Eckle, S. B., van Heemst, J., Pike, R. N., McCluskey, J., Toes, R. E., La Gruta, N. L., Purcell, A. W., Reid, H. H., Thomas, R. and Rossjohn, J.
Year: 2013
Title: A molecular basis for the association of the HLA-DRB1 locus, citrullination, and rheumatoid arthritis
Journal: J Exp Med
Volume: 210
Issue: 12
Pages: 2569-82
Epub Date: 2013/11/06
Date: Nov 18
Short Title: A molecular basis for the association of the HLA-DRB1 locus, citrullination, and rheumatoid arthritis
Alternate Journal: The Journal of experimental medicine
ISSN: 0022-1007 (Print)
0022-1007
DOI: 10.1084/jem.20131241
PMCID: PMC3832918
Accession Number: 24190431
Keywords: Aggrecans/genetics/immunology/metabolism
Amino Acid Sequence
Animals
Antigen Presentation
Arthritis, Rheumatoid/*genetics/*immunology/metabolism
Autoantigens/chemistry/genetics/metabolism
CD4-Positive T-Lymphocytes/immunology/metabolism
Citrulline/metabolism
Epitopes/chemistry/genetics/metabolism
Genetic Association Studies
HLA-DR beta-Chains/chemistry/genetics/metabolism
HLA-DR4 Antigen/chemistry/genetics/metabolism
HLA-DRB1 Chains/chemistry/*genetics/*metabolism
Humans
Mice
Mice, Transgenic
Models, Molecular
Molecular Sequence Data
Polymorphism, Genetic
Vimentin/genetics/immunology/metabolism
Abstract: Rheumatoid arthritis (RA) is strongly associated with the human leukocyte antigen (HLA)-DRB1 locus that possesses the shared susceptibility epitope (SE) and the citrullination of self-antigens. We show how citrullinated aggrecan and vimentin epitopes bind to HLA-DRB1*04:01/04. Citrulline was accommodated within the electropositive P4 pocket of HLA-DRB1*04:01/04, whereas the electronegative P4 pocket of the RA-resistant HLA-DRB1*04:02 allomorph interacted with arginine or citrulline-containing epitopes. Peptide elution studies revealed P4 arginine-containing peptides from HLA-DRB1*04:02, but not from HLA-DRB1*04:01/04. Citrullination altered protease susceptibility of vimentin, thereby generating self-epitopes that are presented to T cells in HLA-DRB1*04:01(+) individuals. Using HLA-II tetramers, we observed citrullinated vimentin- and aggrecan-specific CD4(+) T cells in the peripheral blood of HLA-DRB1*04:01(+) RA-affected and healthy individuals. In RA patients, autoreactive T cell numbers correlated with disease activity and were deficient in regulatory T cells relative to healthy individuals. These findings reshape our understanding of the association between citrullination, the HLA-DRB1 locus, and T cell autoreactivity in RA.
Notes: 1540-9538
Scally, Stephen W
Petersen, Jan
Law, Soi Cheng
Dudek, Nadine L
Nel, Hendrik J
Loh, Khai Lee
Wijeyewickrema, Lakshmi C
Eckle, Sidonia B G
van Heemst, Jurgen
Pike, Robert N
McCluskey, James
Toes, Rene E
La Gruta, Nicole L
Purcell, Anthony W
Reid, Hugh H
Thomas, Ranjeny
Rossjohn, Jamie
Journal Article
Research Support, Non-U.S. Gov't
J Exp Med. 2013 Nov 18;210(12):2569-82. doi: 10.1084/jem.20131241. Epub 2013 Nov 4.
Author Address: Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 686
Author: Schäcke, H., Schumann, H., Hammami-Hauasli, N., Raghunath, M. and Bruckner-Tuderman, L.
Year: 1998
Title: Two forms of collagen XVII in keratinocytes. A full-length transmembrane protein and a soluble ectodomain
Journal: J Biol Chem
Volume: 273
Issue: 40
Pages: 25937-43
Epub Date: 1998/09/25
Date: Oct 2
Short Title: Two forms of collagen XVII in keratinocytes. A full-length transmembrane protein and a soluble ectodomain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.40.25937
Accession Number: 9748270
Keywords: Amino Acid Chloromethyl Ketones/pharmacology
Collagen/*chemistry
Collagenases/metabolism
Disulfides/chemistry
Endopeptidases/metabolism
Furin
Glycosylation
Keratinocytes/*chemistry
Membrane Proteins/chemistry
Mutation/genetics
Peptide Fragments/immunology
Protease Inhibitors/pharmacology
Protein Conformation
RNA, Messenger/metabolism
Recombinant Proteins/genetics
Skin/*chemistry
Subtilisins/antagonists & inhibitors
Abstract: The cDNA sequence of human collagen XVII predicts an unusual type II transmembrane protein, but a biochemical characterization of this structure has not been accomplished yet. Using domain-specific antibodies against recombinant collagen XVII fragments, we identified two molecular forms of the collagen in human skin and epithelial cells. Full-length collagen XVII appeared as a homotrimeric transmembrane molecule of three 180-kDa alpha1(XVII) chains. The globular intracellular domain was disulfide-linked, and the N-glycosylated extracellular domain of three 120-kDa polypeptides was triple-helical at physiological temperatures. A second, soluble form of collagen XVII in keratinocyte culture media was recognized with antibodies to the ectodomain, but not the endodomain. The soluble form exhibited molecular properties of the collagen XVII ectodomain: a triple-helical, N-glycosylated molecule of three 120-kDa polypeptides. Northern blot analysis with probes spanning either the distal 5'or the distal 3' end of the collagen XVII cDNA revealed an identical 6-kb mRNA, suggesting that both the 180- and 120-kDa polypeptides were translated from the same mRNA, and that the 120-kDa polypeptide was generated post-translationally. In concert, keratinocytes harboring a homozygous nonsense mutation in the COL17A1 gene synthesized neither the 180-kDa alpha1(XVII) chain nor the 120-kDa polypeptide. Finally, treatment of normal keratinocytes with a synthetic inhibitor of furin proprotein convertases, decanoyl-RVKR-chloromethyl ketone, prevented the generation of the 120-kDa polypeptide. These data strongly suggest that the soluble 120-kDa polypeptide represents a specifically cleaved ectodomain of collagen XVII, generated through furin-mediated proteolytic processing. Thus, collagen XVII is not only an unusual type II transmembrane collagen, but the first collagen with a specifically processed, soluble triple-helical ectodomain.
Notes: Schäcke, H
Schumann, H
Hammami-Hauasli, N
Raghunath, M
Bruckner-Tuderman, L
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1998 Oct 2;273(40):25937-43. doi: 10.1074/jbc.273.40.25937.
Author Address: Department of Dermatology, University of Muenster, D-48149 Muenster, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 249
Author: Schäffer, L. and Ljungqvist, L.
Year: 1992
Title: Identification of a disulfide bridge connecting the alpha-subunits of the extracellular domain of the insulin receptor
Journal: Biochem Biophys Res Commun
Volume: 189
Issue: 2
Pages: 650-3
Epub Date: 1992/12/15
Date: Dec 15
Short Title: Identification of a disulfide bridge connecting the alpha-subunits of the extracellular domain of the insulin receptor
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(92)92250-2
Accession Number: 1472036
Keywords: Amino Acid Sequence
Animals
Cell Line
Chromatography, High Pressure Liquid
Cricetinae
Cysteine/metabolism
Disulfides/analysis
Humans
Kidney
Macromolecular Substances
Molecular Sequence Data
Peptide Fragments/isolation & purification
Receptor, Insulin/biosynthesis/*chemistry/genetics
Recombinant Proteins/biosynthesis/chemistry
Transfection
Trypsin
Abstract: The alpha 2 beta 2 structure of the insulin receptor has previously been shown to involve one disulfide bridge between the alpha-subunits in the region containing Cys435, Cys468 and Cys524. We have digested the soluble extracellular domain of the insulin receptor with succinylated trypsin, partially separated the resulting peptides, and sequenced a number of fractions. The peptides containing Cys435 and Cys468 appeared in the same fraction, indicating that these two form a disulfide bond, and in another fraction we found the sequence of the peptide containing Cys524. Since it has been shown that the extracellular domain of the insulin receptor has no free thiols and since no other sequences containing cysteine were found in these fractions, we conclude that Cys524 forms a disulfide bond to the Cys524 in the other alpha-subunit.
Notes: Schäffer, L
Ljungqvist, L
Journal Article
United States
Biochem Biophys Res Commun. 1992 Dec 15;189(2):650-3. doi: 10.1016/0006-291x(92)92250-2.
Author Address: Novo Nordisk A/S, Novo Alle, Bagsvaerd, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 302
Author: Schaller, J., Akiyama, K., Kimura, H., Hess, D., Affolter, M. and Rickli, E. E.
Year: 1991
Title: Primary structure of a new actin-binding protein from human seminal plasma
Journal: Eur J Biochem
Volume: 196
Issue: 3
Pages: 743-50
Epub Date: 1991/03/28
Date: Mar 28
Short Title: Primary structure of a new actin-binding protein from human seminal plasma
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1991.tb15873.x
Accession Number: 2013294
Keywords: Actins/*metabolism
Amino Acid Sequence
Carbohydrates/analysis
Carrier Proteins/analysis/*chemistry
Disulfides
Humans
Male
Molecular Sequence Data
Semen/*chemistry
Abstract: Secretory actin-binding protein (SABP), a glycoprotein from human seminal plasma, was isolated according to Akiyama and Kimura [Akiyama, K. & Kimura, H. (1990) Biochim. Biophys. Acta 1040, 206-210]. The complete amino acid sequence of SABP was determined with the aid of fragments generated by trypsin, Staphylococcus aureus V8 protease and pepsin. The single polypeptide chain of SABP contains 118 amino acids with a calculated Mr of 13,506 and pyroglutamic acid as the N-terminal residue. A single N-glycosidic carbohydrate moiety is located at Asn77. The carbohydrate composition shows an unusually high amount of fucose. The arrangement of the two disulfide bonds is Cys37-Cys63 and Cys61-Cys95. Sequence comparison revealed a high degree of similarity with a 14-kDa submandibular gland protein from mouse (45% identity and 64% similarity). SABP is identical with a prolactin-inducible protein and a protein termed gross cystic disease fluid protein 15 (sequences translated from cDNA clones), both from human breast tissues. Although SABP was also detected in saliva, in extracts of the submandibular gland and seminal vesicles, little is known of its function.
Notes: Schaller, J
Akiyama, K
Kimura, H
Hess, D
Affolter, M
Rickli, E E
Journal Article
England
Eur J Biochem. 1991 Mar 28;196(3):743-50. doi: 10.1111/j.1432-1033.1991.tb15873.x.
Author Address: Institute of Biochemistry, University of Bern, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1363
Author: Scharf, L., Li, N. S., Hawk, A. J., Garzón, D., Zhang, T., Fox, L. M., Kazen, A. R., Shah, S., Haddadian, E. J., Gumperz, J. E., Saghatelian, A., Faraldo-Gómez, J. D., Meredith, S. C., Piccirilli, J. A. and Adams, E. J.
Year: 2010
Title: The 2.5 Å structure of CD1c in complex with a mycobacterial lipid reveals an open groove ideally suited for diverse antigen presentation
Journal: Immunity
Volume: 33
Issue: 6
Pages: 853-62
Epub Date: 2010/12/21
Date: Dec 14
Short Title: The 2.5 Å structure of CD1c in complex with a mycobacterial lipid reveals an open groove ideally suited for diverse antigen presentation
Alternate Journal: Immunity
ISSN: 1074-7613 (Print)
1074-7613
DOI: 10.1016/j.immuni.2010.11.026
PMCID: PMC3010391
NIHMSID: NIHMS256687
Accession Number: 21167756
Keywords: Antigen Presentation
Antigenic Variation
Antigens, Bacterial/*chemistry/immunology
Antigens, CD1/*chemistry/immunology
Cloning, Molecular
Computational Biology
Crystallization
Glycoproteins/*chemistry/immunology
Histocompatibility Antigens/metabolism
Humans
*Models, Immunological
Mycobacterium tuberculosis/*immunology
Peptide Fragments/metabolism
Protein Binding
*Protein Conformation
X-Rays
Abstract: CD1 molecules function to present lipid-based antigens to T cells. Here we present the crystal structure of CD1c at 2.5 Å resolution, in complex with the pathogenic Mycobacterium tuberculosis antigen mannosyl-β1-phosphomycoketide (MPM). CD1c accommodated MPM's methylated alkyl chain exclusively in the A' pocket, aided by a unique exit portal underneath the α1 helix. Most striking was an open F' pocket architecture lacking the closed cavity structure of other CD1 molecules, reminiscent of peptide binding grooves of classical major histocompatibility complex molecules. This feature, combined with tryptophan-fluorescence quenching during loading of a dodecameric lipopeptide antigen, provides a compelling model by which both the lipid and peptide moieties of the lipopeptide are involved in CD1c presentation of lipopeptides.
Notes: 1097-4180
Scharf, Louise
Li, Nan-Sheng
Hawk, Andrew J
Garzón, Diana
Zhang, Tejia
Fox, Lisa M
Kazen, Allison R
Shah, Sneha
Haddadian, Esmael J
Gumperz, Jenny E
Saghatelian, Alan
Faraldo-Gómez, José D
Meredith, Stephen C
Piccirilli, Joseph A
Adams, Erin J
R01 AI074940/AI/NIAID NIH HHS/United States
R01 GM081642/GM/NIGMS NIH HHS/United States
R01 GM081642-03/GM/NIGMS NIH HHS/United States
R01AI073922/AI/NIAID NIH HHS/United States
R01GM081642/GM/NIGMS NIH HHS/United States
R01 AI073922-01A1/AI/NIAID NIH HHS/United States
R01 AI073922/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Immunity. 2010 Dec 14;33(6):853-62. doi: 10.1016/j.immuni.2010.11.026.
Author Address: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, IL 60637, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1049
Author: Schellenberger, U., O'Rear, J., Guzzetta, A., Jue, R. A., Protter, A. A. and Pollitt, N. S.
Year: 2006
Title: The precursor to B-type natriuretic peptide is an O-linked glycoprotein
Journal: Arch Biochem Biophys
Volume: 451
Issue: 2
Pages: 160-6
Epub Date: 2006/06/06
Date: Jul 15
Short Title: The precursor to B-type natriuretic peptide is an O-linked glycoprotein
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1016/j.abb.2006.03.028
Accession Number: 16750161
Keywords: Amino Acid Sequence
Animals
CHO Cells
Carbohydrate Sequence
Cricetinae
Glycosylation
Heart Failure/blood
Humans
Mass Spectrometry
Molecular Sequence Data
Molecular Weight
Natriuretic Peptide, Brain/blood/chemistry/*metabolism
Neuraminidase/pharmacology
Peptide Fragments/blood/chemistry/isolation & purification/*metabolism
Peptide Mapping
Promoter Regions, Genetic
Protein Precursors/blood/*chemistry/genetics/*metabolism
Recombinant Proteins/chemistry/isolation & purification/metabolism
Sequence Analysis, Protein
Trypsin/pharmacology
Abstract: Human pro-B-type natriuretic peptide (proBNP), the precursor for B-type natriuretic peptide (BNP), was expressed in Chinese hamster ovary cells (CHO) and compared by Western blot analysis to BNP cross-reacting material immunoprecipitated from the plasma of heart failure patients. Both recombinant and native forms co-migrated as a diffuse band centered around 25 kDa and were reduced to a 12 kDa species by treatment with a mixture of O-link deglycosylation enzymes. The 108-amino acid CHO-expressed protein was examined by tryptic mapping and LC-MS and found to be an O-linked glycoprotein. Determination of the sites of O-glycosyl addition by blank cycle sequencing of tryptic and Glu-C (Staphylococcus aureus V8 protease) peptides showed that there are seven sites of glycosylation confined to a 36-amino acid residue stretch within the center of the propeptide region. This data is consistent with previous observations of higher molecular weight isoforms of BNP.
Notes: Schellenberger, Ute
O'Rear, Jessica
Guzzetta, Andrew
Jue, Rodney A
Protter, Andrew A
Pollitt, N Stephen
HL55096/HL/NHLBI NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
United States
Arch Biochem Biophys. 2006 Jul 15;451(2):160-6. doi: 10.1016/j.abb.2006.03.028. Epub 2006 Apr 19.
Author Address: Scios Inc., 6500 Paseo Padre Parkway, Fremont, CA 94555, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1462
Author: Schimpl, M., Borodkin, V. S., Gray, L. J. and van Aalten, D. M.
Year: 2012
Title: Synergy of peptide and sugar in O-GlcNAcase substrate recognition
Journal: Chem Biol
Volume: 19
Issue: 2
Pages: 173-8
Epub Date: 2012/03/01
Date: Feb 24
Short Title: Synergy of peptide and sugar in O-GlcNAcase substrate recognition
Alternate Journal: Chemistry & biology
ISSN: 1074-5521 (Print)
1074-5521
DOI: 10.1016/j.chembiol.2012.01.011
PMCID: PMC3476531
Accession Number: 22365600
Keywords: Amino Acid Sequence
Binding Sites
Carbohydrates/*chemistry
Crystallography, X-Ray
Enzyme Inhibitors/chemistry
Glycosylation
Humans
Hydrogen Bonding
Hydrophobic and Hydrophilic Interactions
Kinetics
N-Acetylglucosaminyltransferases/*chemistry/metabolism
Peptides/*chemistry/metabolism
Protein Binding
Protein Structure, Tertiary
Proteome/metabolism
Substrate Specificity
Abstract: Protein O-GlcNAcylation is an essential reversible posttranslational modification in higher eukaryotes. O-GlcNAc addition and removal is catalyzed by O-GlcNAc transferase and O-GlcNAcase, respectively. We report the molecular details of the interaction of a bacterial O-GlcNAcase homolog with three different synthetic glycopeptides derived from characterized O-GlcNAc sites in the human proteome. Strikingly, the peptides bind a conserved O-GlcNAcase substrate binding groove with similar orientation and conformation. In addition to extensive contacts with the sugar, O-GlcNAcase recognizes the peptide backbone through hydrophobic interactions and intramolecular hydrogen bonds, while avoiding interactions with the glycopeptide side chains. These findings elucidate the molecular basis of O-GlcNAcase substrate specificity, explaining how a single enzyme achieves cycling of the complete O-GlcNAc proteome. In addition, this work will aid development of O-GlcNAcase inhibitors that target the peptide binding site.
Notes: 1879-1301
Schimpl, Marianne
Borodkin, Vladimir S
Gray, Lindsey J
van Aalten, Daan M F
087590/Wellcome Trust/United Kingdom
G0900138/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Chem Biol. 2012 Feb 24;19(2):173-8. doi: 10.1016/j.chembiol.2012.01.011.
Author Address: Division of Cell Signalling & Immunology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1345
Author: Schimpl, M., Schüttelkopf, A. W., Borodkin, V. S. and van Aalten, D. M.
Year: 2010
Title: Human OGA binds substrates in a conserved peptide recognition groove
Journal: Biochem J
Volume: 432
Issue: 1
Pages: 1-7
Epub Date: 2010/09/25
Date: Nov 15
Short Title: Human OGA binds substrates in a conserved peptide recognition groove
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20101338
PMCID: PMC2973230
Accession Number: 20863279
Keywords: Acetylglucosamine/metabolism
Adaptor Proteins, Signal Transducing/metabolism
Amino Acid Sequence
Bacterial Proteins/chemistry/genetics/metabolism
Binding Sites/genetics
Cloning, Molecular
Conserved Sequence/genetics
Cyclic AMP Response Element-Binding Protein/metabolism
Enzyme Assays
Forkhead Box Protein O1
Forkhead Transcription Factors/metabolism
Glycosylation
HEK293 Cells
Humans
Kinetics
Models, Molecular
Molecular Sequence Data
Mutation
Peptides/*metabolism
Protein Binding
*Protein Structure, Tertiary
Recombinant Proteins/chemistry/metabolism
Rhodobacteraceae/enzymology/genetics
Sequence Homology, Amino Acid
Substrate Specificity
beta-N-Acetylhexosaminidases/*chemistry/genetics/*metabolism
Abstract: Modification of cellular proteins with O-GlcNAc (O-linked N-acetylglucosamine) competes with protein phosphorylation and regulates a plethora of cellular processes. O-GlcNAcylation is orchestrated by two opposing enzymes, O-GlcNAc transferase and OGA (O-GlcNAcase or β-N-acetylglucosaminidase), which recognize their target proteins via as yet unidentified mechanisms. In the present study, we uncovered the first insights into the mechanism of substrate recognition by human OGA. The structure of a novel bacterial OGA orthologue reveals a putative substrate-binding groove, conserved in metazoan OGAs. Guided by this structure, conserved amino acids lining this groove in human OGA were mutated and the activity on three different substrate proteins [TAB1 (transforming growth factor-β-activated protein kinase 1-binding protein 1), FoxO1 (forkhead box O1) and CREB (cAMP-response-element-binding protein)] was tested in an in vitro deglycosylation assay. The results provide the first evidence that human OGA may possess a substrate-recognition mechanism that involves interactions with O-GlcNAcylated proteins beyond the GlcNAc-binding site, with possible implications for differential regulation of cycling of O-GlcNAc on different proteins.
Notes: 1470-8728
Schimpl, Marianne
Schüttelkopf, Alexander W
Borodkin, Vladimir S
van Aalten, Daan M F
087590/Wellcome Trust/United Kingdom
G0500367/Medical Research Council/United Kingdom
G0900138/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2010 Nov 15;432(1):1-7. doi: 10.1042/BJ20101338.
Author Address: University of Dundee, Scotland, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1526
Author: Schimpl, M., Zheng, X., Borodkin, V. S., Blair, D. E., Ferenbach, A. T., Schüttelkopf, A. W., Navratilova, I., Aristotelous, T., Albarbarawi, O., Robinson, D. A., Macnaughtan, M. A. and van Aalten, D. M.
Year: 2012
Title: O-GlcNAc transferase invokes nucleotide sugar pyrophosphate participation in catalysis
Journal: Nat Chem Biol
Volume: 8
Issue: 12
Pages: 969-74
Epub Date: 2012/10/30
Date: Dec
Short Title: O-GlcNAc transferase invokes nucleotide sugar pyrophosphate participation in catalysis
Alternate Journal: Nature chemical biology
ISSN: 1552-4450 (Print)
1552-4450
DOI: 10.1038/nchembio.1108
PMCID: PMC3509171
NIHMSID: EMS50081
Accession Number: 23103942
Keywords: Adaptor Proteins, Signal Transducing/metabolism
Catalysis
Catalytic Domain
Crystallography, X-Ray
Diphosphates/*metabolism
Humans
Kinetics
Lysine/metabolism
Models, Molecular
Molecular Sequence Data
N-Acetylglucosaminyltransferases/*metabolism
Nucleotides/*metabolism
Phosphates/metabolism
Protein Binding
Protein Conformation
Protein Processing, Post-Translational
Stereoisomerism
Substrate Specificity
Surface Plasmon Resonance
Uridine Diphosphate Galactose/metabolism
Abstract: Protein O-GlcNAcylation is an essential post-translational modification on hundreds of intracellular proteins in metazoa, catalyzed by O-linked β-N-acetylglucosamine (O-GlcNAc) transferase (OGT) using unknown mechanisms of transfer and substrate recognition. Through crystallographic snapshots and mechanism-inspired chemical probes, we define how human OGT recognizes the sugar donor and acceptor peptide and uses a new catalytic mechanism of glycosyl transfer, involving the sugar donor α-phosphate as the catalytic base as well as an essential lysine. This mechanism seems to be a unique evolutionary solution to the spatial constraints imposed by a bulky protein acceptor substrate and explains the unexpected specificity of a recently reported metabolic OGT inhibitor.
Notes: 1552-4469
Schimpl, Marianne
Zheng, Xiaowei
Borodkin, Vladimir S
Blair, David E
Ferenbach, Andrew T
Schüttelkopf, Alexander W
Navratilova, Iva
Aristotelous, Tonia
Albarbarawi, Osama
Robinson, David A
Macnaughtan, Megan A
van Aalten, Daan M F
087590/Wellcome Trust/United Kingdom
G0900138/Medical Research Council/United Kingdom
WT087590MA/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Nat Chem Biol. 2012 Dec;8(12):969-74. doi: 10.1038/nchembio.1108. Epub 2012 Oct 28.
Author Address: College of Life Sciences, University of Dundee, Dundee, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 518
Author: Schindler, P. A., Settineri, C. A., Collet, X., Fielding, C. J. and Burlingame, A. L.
Year: 1995
Title: Site-specific detection and structural characterization of the glycosylation of human plasma proteins lecithin:cholesterol acyltransferase and apolipoprotein D using HPLC/electrospray mass spectrometry and sequential glycosidase digestion
Journal: Protein Sci
Volume: 4
Issue: 4
Pages: 791-803
Epub Date: 1995/04/01
Date: Apr
Short Title: Site-specific detection and structural characterization of the glycosylation of human plasma proteins lecithin:cholesterol acyltransferase and apolipoprotein D using HPLC/electrospray mass spectrometry and sequential glycosidase digestion
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1002/pro.5560040419
PMCID: PMC2143102
Accession Number: 7613477
Keywords: Amino Acid Sequence
Apolipoproteins/*chemistry/metabolism
Apolipoproteins D
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Glycopeptides/*analysis
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/analysis
Phosphatidylcholine-Sterol O-Acyltransferase/*chemistry/metabolism
Trypsin/metabolism
Abstract: Site-specific structural characterization of the glycosylation of human lecithin:cholesterol acyltransferase (LCAT) was carried out using microbore reversed-phase high performance liquid chromatography coupled with electrospray ionization mass spectrometry (HPLC/ESIMS). A recently described mass spectrometric technique involving monitoring of carbohydrate-specific fragment ions during HPLC/ESIMS was employed to locate eight different groups of glycopeptides in a digest of a human LCAT protein preparation. In addition to the four expected N-linked glycopeptides of LCAT, a di-O-linked glycopeptide was detected, as well as three additional glycopeptides. Structural information on the oligosaccharides from all eight glycopeptides was obtained by sequential glycosidase digestion of the glycopeptides followed by HPLC/ESIMS. All four potential N-linked glycosylation sites (Asn20, Asn84, Asn272, and Asn384) of LCAT were determined to contain sialylated triantennary and/or biantennary complex structures. Two unanticipated O-linked glycosylation sites were identified at Thr407 and Ser409 of the LCAT O-linked glycopeptide, each of which contain sialylated galactose beta 1-->3N-acetylgalactosamine structures. The three additional glycopeptides were determined to be from a copurifying protein, apolipoprotein D, which contains potential N-linked glycosylation sites at Asn45 and Asn78. These glycopeptides were determined to bear sialylated triantennary oligosaccharides or fucosylated sialylated biantennary oligosaccharides. Previous studies of LCAT indicated that removal of the glycosylation site at Asn272 converts this protein to a phospholipase (Francone OL, Evangelista L, Fielding CJ, 1993, Biochim Biophys Acta 1166:301-304). Our results indicate that the carbohydrate structures themselves are not the source of this functional discrimination; rather, it must be mediated by the structural environment around Asn272.
Notes: 1469-896x
Schindler, P A
Settineri, C A
Collet, X
Fielding, C J
Burlingame, A L
ES04704/ES/NIEHS NIH HHS/United States
H114237/PHS HHS/United States
RR01614/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Protein Sci. 1995 Apr;4(4):791-803. doi: 10.1002/pro.5560040419.
Author Address: Department of Pharmaceutical Chemistry, University of California, San Francisco 94143, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1342
Author: Schjoldager, K. T., Vester-Christensen, M. B., Bennett, E. P., Levery, S. B., Schwientek, T., Yin, W., Blixt, O. and Clausen, H.
Year: 2010
Title: O-glycosylation modulates proprotein convertase activation of angiopoietin-like protein 3: possible role of polypeptide GalNAc-transferase-2 in regulation of concentrations of plasma lipids
Journal: J Biol Chem
Volume: 285
Issue: 47
Pages: 36293-303
Epub Date: 2010/09/15
Date: Nov 19
Short Title: O-glycosylation modulates proprotein convertase activation of angiopoietin-like protein 3: possible role of polypeptide GalNAc-transferase-2 in regulation of concentrations of plasma lipids
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.156950
PMCID: PMC2978557
Accession Number: 20837471
Keywords: Angiopoietin-like Proteins
Angiopoietins/genetics/*metabolism
Animals
Blotting, Western
CHO Cells
Cricetinae
Cricetulus
Furin/metabolism
Glycopeptides/*metabolism
Glycosylation
Humans
Lipids/*blood
N-Acetylgalactosaminyltransferases/*metabolism
Proprotein Convertases/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: The angiopoietin-like protein 3 (ANGPTL3) is an important inhibitor of the endothelial and lipoprotein lipases and a promising drug target. ANGPTL3 undergoes proprotein convertase processing (RAPR(224)↓TT) for activation, and the processing site contains two potential GalNAc O-glycosylation sites immediately C-terminal (TT(226)). We developed an in vivo model system in CHO ldlD cells that was used to show that O-glycosylation in the processing site blocked processing of ANGPTL3. Genome-wide SNP association studies have identified the polypeptide GalNAc-transferase gene, GALNT2, as a candidate gene for low HDL and high triglyceride blood levels. We hypothesized that the GalNAc-T2 transferase performed critical O-glycosylation of proteins involved in lipid metabolism. Screening of a panel of proteins known to affect lipid metabolism for potential sites glycosylated by GalNAc-T2 led to identification of Thr(226) adjacent to the proprotein convertase processing site in ANGPTL3. We demonstrated that GalNAc-T2 glycosylation of Thr(226) in a peptide with the RAPR(224)↓TT processing site blocks in vitro furin cleavage. The study demonstrates that ANGPTL3 activation is modulated by O-glycosylation and that this step is probably controlled by GalNAc-T2.
Notes: 1083-351x
Schjoldager, Katrine T-B G
Vester-Christensen, Malene B
Bennett, Eric Paul
Levery, Steven B
Schwientek, Tilo
Yin, Wu
Blixt, Ola
Clausen, Henrik
Howard Hughes Medical Institute/United States
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Nov 19;285(47):36293-303. doi: 10.1074/jbc.M110.156950. Epub 2010 Sep 13.
Author Address: Center for Glycomics, Department of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 598
Author: Schmelzer, C. H., Harris, R. J., Butler, D., Yedinak, C. M., Wagner, K. L. and Burton, L. E.
Year: 1993
Title: Glycosylation pattern and disulfide assignments of recombinant human differentiation-stimulating factor
Journal: Arch Biochem Biophys
Volume: 302
Issue: 2
Pages: 484-9
Epub Date: 1993/05/01
Date: May
Short Title: Glycosylation pattern and disulfide assignments of recombinant human differentiation-stimulating factor
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1993.1243
Accession Number: 8489250
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cricetinae
Cystine/chemistry
Disulfides/chemistry
Glycosylation
Growth Inhibitors/*chemistry/*metabolism
Humans
*Interleukin-6
Leukemia Inhibitory Factor
Lymphokines/*chemistry/*metabolism
Molecular Sequence Data
Peptide Fragments/chemistry
*Protein Processing, Post-Translational
Sequence Analysis
Abstract: This report describes the post-translational modifications of recombinant human differentiation-stimulating factor, a 180-residue glycoprotein that is secreted from transfected Chinese hamster ovary cells. Peptide peptides containing six potential N-glycosylation sites were analyzed to determine that Asn residues 9, 34, 63, 73, 96, and 116 were utilized. Additional peptides, generated by tryptic digestion of peptic fragments, allowed the assignments of three intrachain disulfide bonds (Cys-18 to Cys-131, Cys-12 to Cys-134, and Cys-60 to Cys-163).
Notes: Schmelzer, C H
Harris, R J
Butler, D
Yedinak, C M
Wagner, K L
Burton, L E
Journal Article
United States
Arch Biochem Biophys. 1993 May;302(2):484-9. doi: 10.1006/abbi.1993.1243.
Author Address: Department of Recovery Process Research and Development, Genentech, Inc., South San Francisco, California 94080.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 445
Author: Schmid, K., Bürgi, W., Collins, J. H. and Nanno, S.
Year: 1974
Title: The disulfide bonds of alpha1-acid glycoprotein
Journal: Biochemistry
Volume: 13
Issue: 13
Pages: 2694-7
Epub Date: 1974/06/18
Date: Jun 18
Short Title: The disulfide bonds of alpha1-acid glycoprotein
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00710a006
Accession Number: 4603214
Keywords: Amino Acid Sequence
Amino Acids/analysis
Binding Sites
Chromatography, Gel
Chromatography, Ion Exchange
Chromatography, Thin Layer
Chymotrypsin
Disulfides/analysis
Electrophoresis, Paper
Glycoproteins/*blood
Humans
Oxidation-Reduction
Pepsin A
Peptide Fragments/analysis
Protein Binding
Protein Conformation
Tosyl Compounds
Trypsin
Notes: Schmid, K
Bürgi, W
Collins, J H
Nanno, S
Journal Article
United States
Biochemistry. 1974 Jun 18;13(13):2694-7. doi: 10.1021/bi00710a006.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 278
Author: Schmid, K., Hediger, M. A., Brossmer, R., Collins, J. H., Haupt, H., Marti, T., Offner, G. D., Schaller, J., Takagaki, K., Walsh, M. T. and et al.
Year: 1992
Title: Amino acid sequence of human plasma galactoglycoprotein: identity with the extracellular region of CD43 (sialophorin)
Journal: Proc Natl Acad Sci U S A
Volume: 89
Issue: 2
Pages: 663-7
Epub Date: 1992/01/15
Date: Jan 15
Short Title: Amino acid sequence of human plasma galactoglycoprotein: identity with the extracellular region of CD43 (sialophorin)
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.89.2.663
PMCID: PMC48299
Accession Number: 1731338
Keywords: Amino Acid Sequence
Antigens, CD/chemistry
Blood Proteins/*chemistry
Carbohydrate Sequence
Glycopeptides/chemistry
Glycoproteins/*chemistry
Humans
Leukosialin
Molecular Sequence Data
Molecular Weight
Peptide Fragments/chemistry
Protein Conformation
Sialoglycoproteins/*chemistry
Abstract: The amino acid sequence of galactoglycoprotein purified from human plasma was elucidated to 75% completeness by using chemical degradation of peptides and glycopeptides derived from digests of the protein with seven specific proteases. This sequence represents a polypeptide chain of approximately 220 amino acid residues including a high content of serine, threonine, alanine, and proline with one N-linked and multiple O-linked glycans. Comparison of peptide sequences from the native galactoglycoprotein and the deglycosylated derivative demonstrated the locations of 25 sites of O-glycosylation and three serine sites that are not glycosylated. The homogeneous N terminus was established as serine. C-terminal analysis revealed multiple C-terminal residues, suggesting that galactoglycoprotein molecules are of varying lengths. A search of a protein data base revealed that the galactoglycoprotein polypeptide is identical to the N-terminal (extracellular) polypeptide region of the blood-cell surface molecule CD43 (sialophorin, leukosialin). Further support of the relatedness of these molecules was obtained by immunoprecipitation of 125I-labeled galactoglycoprotein by monoclonal anti-CD43 antibodies. The composition and properties of the molecules together with the known structure of the gene encoding CD43 suggest that galactoprotein is derived by proteolytic cleavage from transmembrane "hexasaccharide CD43," known to be expressed on neutrophils, activated T lymphocytes, and platelets.
Notes: 1091-6490
Schmid, K
Hediger, M A
Brossmer, R
Collins, J H
Haupt, H
Marti, T
Offner, G D
Schaller, J
Takagaki, K
Walsh, M T
AI29880/AI/NIAID NIH HHS/United States
DK-43171/DK/NIDDK NIH HHS/United States
GM-10374/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1992 Jan 15;89(2):663-7. doi: 10.1073/pnas.89.2.663.
Author Address: Department of Biochemistry, Boston University School of Medicine, MA 02118.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 576
Author: Schneider, G., Lindqvist, Y. and Vihko, P.
Year: 1993
Title: Three-dimensional structure of rat acid phosphatase
Journal: Embo j
Volume: 12
Issue: 7
Pages: 2609-15
Epub Date: 1993/07/01
Date: Jul
Short Title: Three-dimensional structure of rat acid phosphatase
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
PMCID: PMC413507
Accession Number: 8334986
Keywords: Acid Phosphatase/*chemistry/genetics/metabolism
Animals
Baculoviridae/genetics
Binding Sites
Bisphosphoglycerate Mutase/chemistry
Cells, Cultured
Electrons
Glycosylation
Male
Models, Molecular
Moths
Prostate/enzymology
Protein Conformation
Rats
Recombinant Proteins/chemistry/genetics/metabolism
Substrate Specificity
X-Ray Diffraction
Abstract: The crystal structure of recombinant rat prostatic acid phosphatase was determined to 3 A resolution with protein crystallographic methods. The enzyme subunit is built up of two domains, an alpha/beta domain consisting of a seven-stranded mixed beta-sheet with helices on both sides of the sheet and a smaller alpha domain. Two disulfide bridges between residues 129-340 and 315-319 were found. Electron density at two of the glycosylation sites for parts of the carbohydrate moieties was observed. The dimer of acid phosphatase is formed through two-fold interactions of edge strand 3 from one subunit with strand 3 from the second subunit, thus extending the beta-sheet from seven to 14 strands. Other subunit-subunit interactions involve conserved residues from loops between helices and beta-strands. The fold of the alpha/beta domain is similar to the fold observed in phosphoglycerate mutase. The active site is at the carboxy end of the parallel strands of the alpha/beta domain. There is a strong residual electron density at the phosphate binding site which probably represents a bound chloride ion. Biochemical properties and results from site-directed mutagenesis experiments of acid phosphatase are correlated to the three-dimensional structure.
Notes: 1460-2075
Schneider, G
Lindqvist, Y
Vihko, P
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1993 Jul;12(7):2609-15.
Author Address: Department of Molecular Biology, Swedish University of Agricultural Sciences, Uppsala Biomedical Center.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1247
Author: Scholz, C. J., Sauer, G. and Deissler, H.
Year: 2009
Title: Glycosylation of tetraspanin Tspan-1 at four distinct sites promotes its transition through the endoplasmic reticulum
Journal: Protein Pept Lett
Volume: 16
Issue: 10
Pages: 1244-8
Epub Date: 2009/06/11
Short Title: Glycosylation of tetraspanin Tspan-1 at four distinct sites promotes its transition through the endoplasmic reticulum
Alternate Journal: Protein and peptide letters
ISSN: 0929-8665
DOI: 10.2174/092986609789071234
Accession Number: 19508227
Keywords: Binding Sites/genetics
Cell Line, Tumor
Cell Membrane/metabolism
Endoplasmic Reticulum/*metabolism
Female
Glycosylation
Green Fluorescent Proteins/genetics/metabolism
Humans
Immunoblotting
Lysosomes/metabolism
Mannose/metabolism
Membrane Proteins/chemistry/genetics/*metabolism
Microscopy, Fluorescence
Mutation
Oligosaccharides/metabolism
Ovarian Neoplasms/genetics/metabolism/pathology
Protein Folding
Protein Transport
Tetraspanins
Transfection
Abstract: We showed that Tspan-1, a tetraspanin overexpressed in many human cancers, harbours oligosaccharides at all four potential N-glycosylation sites. Its most abundant form contained only mannose-rich sugar chains but two distinct glycosylation sites could also contain complex carbohydrates. Glycosylation seemed to be required for correct folding and subsequent transition through the endoplasmic reticulum.
Notes: 1875-5305
Scholz, Claus-Jürgen
Sauer, Georg
Deissler, Helmut
Journal Article
Netherlands
Protein Pept Lett. 2009;16(10):1244-8. doi: 10.2174/092986609789071234.
Author Address: Department of Obstetrics and Gynaecology, University of Ulm, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1274
Author: Schröder, S., Matthes, F., Hyden, P., Andersson, C., Fogh, J., Müller-Loennies, S., Braulke, T., Gieselmann, V. and Matzner, U.
Year: 2010
Title: Site-specific analysis of N-linked oligosaccharides of recombinant lysosomal arylsulfatase A produced in different cell lines
Journal: Glycobiology
Volume: 20
Issue: 2
Pages: 248-59
Epub Date: 2009/10/30
Date: Feb
Short Title: Site-specific analysis of N-linked oligosaccharides of recombinant lysosomal arylsulfatase A produced in different cell lines
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwp171
Accession Number: 19864504
Keywords: Animals
CHO Cells
Cells, Cultured
Cerebroside-Sulfatase/*biosynthesis/isolation & purification/*metabolism
Chromatography, High Pressure Liquid
Cricetinae
Cricetulus
Humans
Lysosomes/*enzymology
Oligosaccharides/*analysis/*chemistry
Recombinant Proteins/biosynthesis/isolation & purification/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Metachromatic leukodystrophy (MLD) is a lysosomal storage disease caused by a deficiency of the lysosomal enzyme arylsulfatase A (ASA). Enzyme replacement therapy (ERT) is a therapeutic option for MLD and other lysosomal disorders. This therapy depends on N-linked oligosaccharide-mediated delivery of intravenously injected recombinant enzyme to the lysosomes of patient cells. Because of the importance of N-linked oligosaccharide side chains in ERT, we examined the composition of the three N-linked glycans of four different recombinant ASAs in a site-specific manner. Depending on the culture conditions and the cell line expressing the enzyme, we detected a high variability of the high-mannose-type N-glycans which prevail at all glycosylation sites. Our data show that the composition of the glycans is largely determined by substantial trimming in the medium. The susceptibility for trimming is different for the glycans at the three N-glycosylation sites. Interestingly, which of the glycans is most susceptible to trimming also depends on production conditions. CHO cells cultured under bioreactor conditions yielded recombinant ASA with the most preserved N-glycan structures, the highest mannose-6-phosphate content and the highest similarity to non-recombinant enzyme. Notably, roughly one-third of the N-glycans released from the three glycosylation sites were fucosylated. In the last years, numerous recombinant lysosomal enzymes were used for preclinical ERT trials. Our data show that the oligosaccharide structures were very different in these trials making it difficult to draw common conclusions from the various investigations.
Notes: 1460-2423
Schröder, Stephan
Matthes, Frank
Hyden, Pia
Andersson, Claes
Fogh, Jens
Müller-Loennies, Sven
Braulke, Thomas
Gieselmann, Volkmar
Matzner, Ulrich
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2010 Feb;20(2):248-59. doi: 10.1093/glycob/cwp171. Epub 2009 Oct 28.
Author Address: Rheinische Friedrich-Wilhelms Universität, Institut für Biochemie und Molekularbiologie, Nussallee 11, D-53115 Bonn, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 805
Author: Schuette, C. G., Weisgerber, J. and Sandhoff, K.
Year: 2001
Title: Complete analysis of the glycosylation and disulfide bond pattern of human beta-hexosaminidase B by MALDI-MS
Journal: Glycobiology
Volume: 11
Issue: 7
Pages: 549-56
Epub Date: 2001/07/12
Date: Jul
Short Title: Complete analysis of the glycosylation and disulfide bond pattern of human beta-hexosaminidase B by MALDI-MS
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/11.7.549
Accession Number: 11447134
Keywords: Amino Acid Sequence
Animals
Cell Line
Chromatography, High Pressure Liquid/methods
Chromatography, Liquid/methods
Disulfides/*metabolism
Glycosylation
Hexosaminidase B
Humans
Molecular Sequence Data
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/*methods
Spodoptera
beta-N-Acetylhexosaminidases/*blood/chemistry/genetics/isolation & purification
Abstract: beta-hexosaminidase B is an enzyme that is involved in the degradation of glycolipids and glycans in the lysosome. Mutation in the HEXB gene lead to Sandhoff disease, a glycolipid storage disorder characterized by severe neurodegeneration. So far, little structural information on the protein is available. Here, the complete analysis of the disulfide bond pattern of the protein is described for the first time. Additionally, the structures of the N-glycans are analyzed for the native human protein and for recombinant protein expressed in SF21 cells. For the analysis of the disulfide bond structure, the protein was proteolytically digested and the resulting peptides were analyzed by MALDI-MS. The analysis revealed three disulfide bonds (C91-C137; C309-C360; C534-C551) and a free cysteine (C487). The analysis of the N-glycosylation was performed by tryptic digestion of the protein, isolation of glycopeptides by lectin chromatography and mass measurement before and after enzymatic deglycosylation. Carbohydrate structures were calculated from the mass difference between glycosylated and deglycosylated peptide. For beta-hexosaminidase B from human placenta, four N-glycans were identified and analyzed, whereas the recombinant protein expressed in SF21 cells carried only three glycans. In both cases the glycosylation belongs to the mannose-core- or high-mannose-type, and some carbohydrate structures are fucosylated.
Notes: Schuette, C G
Weisgerber, J
Sandhoff, K
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2001 Jul;11(7):549-56. doi: 10.1093/glycob/11.7.549.
Author Address: Kekulé-Institut für Organische Chemie und Biochemie, Universität Bonn, Gerhard-Domagk-Str. 1, D-53121 Bonn, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 529
Author: Schulte, S. and Stoffel, W.
Year: 1993
Title: Ceramide UDPgalactosyltransferase from myelinating rat brain: purification, cloning, and expression
Journal: Proc Natl Acad Sci U S A
Volume: 90
Issue: 21
Pages: 10265-9
Epub Date: 1993/11/01
Date: Nov 1
Short Title: Ceramide UDPgalactosyltransferase from myelinating rat brain: purification, cloning, and expression
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.90.21.10265
PMCID: PMC47755
Accession Number: 7694285
Keywords: Amino Acid Sequence
Animals
Base Sequence
Brain/*enzymology
Chromatography, Affinity
Chromatography, Ion Exchange
Cloning, Molecular
DNA, Complementary/chemistry/metabolism
Galactosyltransferases/*biosynthesis/genetics/*isolation & purification
In Situ Hybridization
Kinetics
Microsomes/enzymology
Molecular Sequence Data
Myelin Sheath/*physiology
N-Acylsphingosine Galactosyltransferase
Oligodeoxyribonucleotides
Organ Specificity
Poly A/isolation & purification/metabolism
RNA/isolation & purification/metabolism
RNA, Messenger/analysis/biosynthesis
Rats
Rats, Wistar
Transcription, Genetic
Abstract: Cerebrosides and sulfatides are major glycosphingolipids of the lipid bilayer of the myelin sheath assembled by oligodendrocytes and Schwann cells during myelination. Cerebrosides are synthesized by ceramide UDPgalactosyltransferase [CGT; 2-hydroxyacylsphinogosine 1-beta-galactosyl-transferase; UDPgalactose:2-(2-hydroxyacyl)sphingosine 1-beta-D-galactosyltransferase; UDPgalactose:2-(2-hydroxyacyl)sphingosine 1-beta-D-galactosyltransferase, EC 2.4.1.45] with UDPgalactose and ceramide as substrates. Here we describe a purification method from microsomes of myelinating rat brains that includes ion exchange, dye ligand, and lectin affinity chromatography. The enzyme was identified as a 64-kDa high-mannose glycoprotein. A CGT-specific cDNA clone was isolated from a rat brain cDNA library using CGT oligonucleotides derived from peptide sequences. The cDNA insert encodes a polypeptide of 541 amino acid residues with a molecular weight of 61,126. The polypeptide has three putative glycosylation sites and one hydrophobic domain at the C terminus. A 20-residue N-terminal signal sequence is lost during cotranslational translocation. Northern blot analysis demonstrates that CGT expression is restricted to brain tissue and is time dependent, correlating with myelin basic protein expression. In situ hybridization reveals that CGT expression is restricted to the oligodendrocyte-containing cell layers of cerebrum and cerebellum, which also express myelin basic protein. The amino acid sequence of CGT shows significant homology to mammalian UDPglucuronyltransferases, which suggests a common evolutionary origin of these enzymes.
Notes: 1091-6490
Schulte, S
Stoffel, W
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):10265-9. doi: 10.1073/pnas.90.21.10265.
Author Address: Institute of Biochemistry, Medical Faculty of the University of Cologne, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1482
Author: Schürpf, T., Chen, Q., Liu, J. H., Wang, R., Springer, T. A. and Wang, J. H.
Year: 2012
Title: The RGD finger of Del-1 is a unique structural feature critical for integrin binding
Journal: Faseb j
Volume: 26
Issue: 8
Pages: 3412-20
Epub Date: 2012/05/19
Date: Aug
Short Title: The RGD finger of Del-1 is a unique structural feature critical for integrin binding
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638 (Print)
0892-6638
DOI: 10.1096/fj.11-202036
PMCID: PMC3405271
Accession Number: 22601780
Keywords: *Amino Acid Motifs
Calcium-Binding Proteins
Carrier Proteins/*chemistry/metabolism
Cell Adhesion Molecules
Epidermal Growth Factor/chemistry
Fucose/metabolism
Glycosylation
HEK293 Cells
Humans
Integrins/*metabolism
Models, Molecular
Oligopeptides/*chemistry
Protein Structure, Tertiary
Abstract: Developmental endothelial cell locus-1 (Del-1) glycoprotein is secreted by endothelial cells and a subset of macrophages. Del-1 plays a regulatory role in vascular remodeling and functions in innate immunity through interaction with integrin α(V)β(3). Del-1 contains 3 epidermal growth factor (EGF)-like repeats and 2 discoidin-like domains. An Arg-Gly-Asp (RGD) motif in the second EGF domain (EGF2) mediates adhesion by endothelial cells and phagocytes. We report the crystal structure of its 3 EGF domains. The RGD motif of EGF2 forms a type II' β turn at the tip of a long protruding loop, dubbed the RGD finger. Whereas EGF2 and EGF3 constitute a rigid rod via an interdomain calcium ion binding site, the long linker between EGF1 and EGF2 lends considerable flexibility to EGF1. Two unique O-linked glycans and 1 N-linked glycan locate to the opposite side of EGF2 from the RGD motif. These structural features favor integrin binding of the RGD finger. Mutagenesis data confirm the importance of having the RGD motif at the tip of the RGD finger. A database search for EGF domain sequences shows that this RGD finger is likely an evolutionary insertion and unique to the EGF domain of Del-1 and its homologue milk fat globule-EGF 8.
Notes: 1530-6860
Schürpf, Thomas
Chen, Qiang
Liu, Jin-Huan
Wang, Rui
Springer, Timothy A
Wang, Jia-Huai
P01 HL103526/HL/NHLBI NIH HHS/United States
HL103526/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
FASEB J. 2012 Aug;26(8):3412-20. doi: 10.1096/fj.11-202036. Epub 2012 May 17.
Author Address: Immune Disease Institute, Harvard Medical School, Boston, MA, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1038
Author: Schüttelkopf, A. W., Hamilton, G., Watts, C. and van Aalten, D. M.
Year: 2006
Title: Structural basis of reduction-dependent activation of human cystatin F
Journal: J Biol Chem
Volume: 281
Issue: 24
Pages: 16570-5
Epub Date: 2006/04/08
Date: Jun 16
Short Title: Structural basis of reduction-dependent activation of human cystatin F
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M601033200
Accession Number: 16601115
Keywords: Amino Acid Sequence
Animals
Biomarkers, Tumor/*chemistry
CHO Cells
Cell Line
Cricetinae
Crystallography, X-Ray/methods
Cystatins/*chemistry
Cysteine Endopeptidases/chemistry
Dimerization
Glycosylation
Histidine/chemistry
Humans
Models, Molecular
Molecular Conformation
Molecular Sequence Data
Protein Binding
Protein Conformation
Sequence Homology, Amino Acid
Abstract: Cystatins are important natural cysteine protease inhibitors targeting primarily papain-like cysteine proteases, including cathepsins and parasitic proteases like cruzipain, but also mammalian asparaginyl endopeptidase. Mammalian cystatin F, which is expressed almost exclusively in hematopoietic cells and accumulates in lysosome-like organelles, has been implicated in the regulation of antigen presentation and other immune processes. It is an unusual cystatin superfamily member with a redox-regulated activation mechanism and a restricted specificity profile. We describe the 2.1A crystal structure of human cystatin F in its dimeric "off" state. The two monomers interact in a fashion not seen before for cystatins or cystatin-like proteins that is crucially dependent on an unusual intermolecular disulfide bridge, suggesting how reduction leads to monomer formation and activation. Strikingly, core sugars for one of the two N-linked glycosylation sites of cystatin F are well ordered, and their conformation and interactions with the protein indicate that this unique feature of cystatin F may modulate its inhibitory properties, in particular its reduced affinity toward asparaginyl endopeptidase compared with other cystatins.
Notes: Schüttelkopf, Alexander W
Hamilton, Garth
Watts, Colin
van Aalten, Daan M F
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2006 Jun 16;281(24):16570-5. doi: 10.1074/jbc.M601033200. Epub 2006 Apr 6.
Author Address: Division of Biological Chemistry and Molecular Microbiology, Wellcome Trust Biocentre, School of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, Scotland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1958
Author: Scietti, L., Chiapparino, A., De Giorgi, F., Fumagalli, M., Khoriauli, L., Nergadze, S., Basu, S., Olieric, V., Cucca, L., Banushi, B., Profumo, A., Giulotto, E., Gissen, P. and Forneris, F.
Year: 2018
Title: Molecular architecture of the multifunctional collagen lysyl hydroxylase and glycosyltransferase LH3
Journal: Nat Commun
Volume: 9
Issue: 1
Pages: 3163
Epub Date: 2018/08/10
Date: Aug 8
Short Title: Molecular architecture of the multifunctional collagen lysyl hydroxylase and glycosyltransferase LH3
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-018-05631-5
PMCID: PMC6082870
Accession Number: 30089812
Keywords: Amino Acid Sequence
Catalysis
Catalytic Domain/genetics/physiology
Collagen/metabolism
Crystallography, X-Ray
Dimerization
Enzyme Activation
Enzyme Assays
Glycosyltransferases/*chemistry/genetics/*physiology
HEK293 Cells
HeLa Cells
Humans
Hydroxylation
Lysine/metabolism
Models, Molecular
*Molecular Structure
Mutant Proteins/genetics
Mutation
Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase/*chemistry/genetics/*physiology
Protein Interaction Mapping
Protein Structure, Tertiary
Recombinant Proteins
Abstract: Lysyl hydroxylases catalyze hydroxylation of collagen lysines, and sustain essential roles in extracellular matrix (ECM) maturation and remodeling. Malfunctions in these enzymes cause severe connective tissue disorders. Human lysyl hydroxylase 3 (LH3/PLOD3) bears multiple enzymatic activities, as it catalyzes collagen lysine hydroxylation and also their subsequent glycosylation. Our understanding of LH3 functions is currently hampered by lack of molecular structure information. Here, we present high resolution crystal structures of full-length human LH3 in complex with cofactors and donor substrates. The elongated homodimeric LH3 architecture shows two distinct catalytic sites at the N- and C-terminal boundaries of each monomer, separated by an accessory domain. The glycosyltransferase domain displays distinguishing features compared to other known glycosyltransferases. Known disease-related mutations map in close proximity to the catalytic sites. Collectively, our results provide a structural framework characterizing the multiple functions of LH3, and the molecular mechanisms of collagen-related diseases involving human lysyl hydroxylases.
Notes: 2041-1723
Scietti, Luigi
Chiapparino, Antonella
Orcid: 0000-0001-6336-2736
De Giorgi, Francesca
Orcid: 0000-0002-5008-3305
Fumagalli, Marco
Khoriauli, Lela
Orcid: 0000-0002-3095-2104
Nergadze, Solomon
Orcid: 0000-0001-8121-0189
Basu, Shibom
Olieric, Vincent
Orcid: 0000-0002-0533-7222
Cucca, Lucia
Banushi, Blerida
Profumo, Antonella
Giulotto, Elena
Gissen, Paul
Orcid: 0000-0002-9712-6122
Forneris, Federico
Orcid: 0000-0002-7818-1804
Programma Dipartimenti di Eccellenza 2018-2022/Ministero dell'Istruzione, dell'Università e della Ricerca (Ministry of Education, University and Research)/International
Programma Giovani Ricercatori Rita Levi-Montalcini/Ministero dell'Istruzione, dell'Università e della Ricerca (Ministry of Education, University and Research)/International
MSCA-IF Grant agreement n. 745934 -/European Commission (EC)/International
FP7/2007-201 (Biostruct-X) Grant agreements 7551 and 10205/European Commission (EC)/International
14/21/45/DH | NIHR | Health Services and Delivery Research Programme (Health Services and Delivery Research (HS&amp;DR) Programme)/International
CDA 2013/Giovanni Armenise-Harvard Foundation/International
My First AIRC Grant (Grant id. 20075)/Fondazione Italiana per la Ricerca sul Cancro (Italian Foundation for Cancer Research)/International
"COME TRUE" (id. 2015-0768)/Fondazione Cariplo (Cariplo Foundation)/International
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2018 Aug 8;9(1):3163. doi: 10.1038/s41467-018-05631-5.
Author Address: The Armenise-Harvard Laboratory of Structural Biology, Department of Biology and Biotechnology, University of Pavia, Via Ferrata 9/A, 27100, Pavia, Italy.
Laboratory of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Via Taramelli 3/B, 27100, Pavia, Italy.
Laboratory of Molecular Biology, Department of Biology and Biotechnology, University of Pavia, Via Ferrata 9/A, 27100, Pavia, Italy.
Swiss Light Source, Paul Scherrer Institut, Villigen, 5232, Switzerland.
Laboratory of Analytical Chemistry, Department of Chemistry, University of Pavia, Via Taramelli 12, 27100, Pavia, Italy.
MRC Laboratory for Molecular Cell Biology, University College London, London, WC1E 6BT, UK.
Translational Research Institute, The University of Queensland Diamantina Institute, Princess Alexandra Hospital, 37 Kent Street, Brisbane, Australia.
UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London, WC1N 1EH, UK.
The Armenise-Harvard Laboratory of Structural Biology, Department of Biology and Biotechnology, University of Pavia, Via Ferrata 9/A, 27100, Pavia, Italy. federico.forneris@unipv.it.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1826
Author: Secco, B., Camiré, É, Brière, M. A., Caron, A., Billong, A., Gélinas, Y., Lemay, A. M., Tharp, K. M., Lee, P. L., Gobeil, S., Guimond, J. V., Patey, N., Guertin, D. A., Stahl, A., Haddad, É, Marsolais, D., Bossé, Y., Birsoy, K. and Laplante, M.
Year: 2017
Title: Amplification of Adipogenic Commitment by VSTM2A
Journal: Cell Rep
Volume: 18
Issue: 1
Pages: 93-106
Epub Date: 2017/01/05
Date: Jan 3
Short Title: Amplification of Adipogenic Commitment by VSTM2A
Alternate Journal: Cell reports
DOI: 10.1016/j.celrep.2016.12.015
PMCID: PMC5551894
NIHMSID: NIHMS888139
Accession Number: 28052263
Keywords: 3T3-L1 Cells
Adipocytes/metabolism
*Adipogenesis
Adipose Tissue, White/blood supply/cytology
Animals
Biomarkers/metabolism
Bone Morphogenetic Proteins/metabolism
Cell Differentiation
*Cell Lineage
Gene Knockdown Techniques
Humans
Male
Membrane Proteins/*metabolism
Mesenchymal Stem Cells/cytology/metabolism
Mice
Mice, Inbred C57BL
Middle Aged
NIH 3T3 Cells
Neovascularization, Physiologic
PPAR gamma/metabolism
Receptors, Immunologic/*metabolism
Signal Transduction
*3t3-l1
*adipogenic commitment
*adipose tissue
*cell differentiation
*cell signaling
*obesity
*preadipocyte
Abstract: Despite progress in our comprehension of the mechanisms regulating adipose tissue development, the nature of the factors that functionally characterize adipose precursors is still elusive. Defining the early steps regulating adipocyte development is needed for the generation of tools to control adipose tissue size and function. Here, we report the discovery of V-set and transmembrane domain containing 2A (VSTM2A) as a protein expressed and secreted by committed preadipocytes. VSTM2A expression is elevated in the early phases of adipogenesis in vitro and adipose tissue development in vivo. We show that VSTM2A-producing cells associate with the vasculature and express the common surface markers of adipocyte progenitors. Overexpression of VSTM2A induces adipogenesis, whereas its depletion impairs this process. VSTM2A controls preadipocyte determination at least in part by modulating BMP signaling and PPARγ2 activation. We propose a model in which VSTM2A is produced to preserve and amplify the adipogenic capability of adipose precursors.
Notes: 2211-1247
Secco, Blandine
Camiré, Étienne
Brière, Marc-Antoine
Caron, Alexandre
Billong, Armande
Gélinas, Yves
Lemay, Anne-Marie
Tharp, Kevin M
Lee, Peter L
Gobeil, Stéphane
Guimond, Jean V
Patey, Natacha
Guertin, David A
Stahl, Andreas
Haddad, Élie
Marsolais, David
Bossé, Yohan
Birsoy, Kivanc
Laplante, Mathieu
MOP123387/CIHR/Canada
R01 DK094004/DK/NIDDK NIH HHS/United States
R01 CA196986/CA/NCI NIH HHS/United States
R01 DK101293/DK/NIDDK NIH HHS/United States
NIH0010513932/Intramural NIH HHS/United States
R01 DK089202/DK/NIDDK NIH HHS/United States
Journal Article
Cell Rep. 2017 Jan 3;18(1):93-106. doi: 10.1016/j.celrep.2016.12.015.
Author Address: Centre de recherche de l'Institut universitaire de cardiologie et de pneumologie de Québec (CRIUCPQ), Université Laval, Faculté de médecine, 2725 Chemin Ste-Foy, QC G1V 4G5, Canada.
Program for Metabolic Biology, Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USA.
University of Massachusetts Medical School, Program in Molecular Medicine, Worcester, MA 01605, USA.
Centre hospitalier universitaire de Québec (CHU de Québec), Université Laval, Faculté de médecine, 2705 Boulevard Laurier, QC G1V 4G2, Canada.
CIUSSS du Centre-Sud-de-l'ile-de-Montréal, CLSC des Faubourgs, 66 rue Sainte-Catherine Est, Montréal, QC H2X 1K6, Canada.
Centre Hospitalier Universitaire de Sainte-Justine (CHU de Sainte-Justine), Faculté de Médecine, Département de pathologie et biologie cellulaire, Université de Montréal, 3175 Chemin Côte Ste-Catherine, Montréal, QC H3T 1C5, Canada.
Centre Hospitalier Universitaire de Sainte-Justine (CHU de Sainte-Justine), Faculté de Médecine, Département de pédiatrie et Département de microbiologie, infectiologie et immunologie, Université de Montréal, 3175 Chemin Côte Ste-Catherine, Montréal, QC H3T 1C5, Canada.
Laboratory of Metabolic Regulation and Genetics, The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA.
Centre de recherche de l'Institut universitaire de cardiologie et de pneumologie de Québec (CRIUCPQ), Université Laval, Faculté de médecine, 2725 Chemin Ste-Foy, QC G1V 4G5, Canada. Electronic address: mathieu.laplante@criucpq.ulaval.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1899
Author: Seegar, T. C. M., Killingsworth, L. B., Saha, N., Meyer, P. A., Patra, D., Zimmerman, B., Janes, P. W., Rubinstein, E., Nikolov, D. B., Skiniotis, G., Kruse, A. C. and Blacklow, S. C.
Year: 2017
Title: Structural Basis for Regulated Proteolysis by the α-Secretase ADAM10
Journal: Cell
Volume: 171
Issue: 7
Pages: 1638-1648.e7
Epub Date: 2017/12/12
Date: Dec 14
Short Title: Structural Basis for Regulated Proteolysis by the α-Secretase ADAM10
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2017.11.014
PMCID: PMC5773094
NIHMSID: NIHMS919617
Accession Number: 29224781
Keywords: ADAM10 Protein/*chemistry/metabolism
Amyloid Precursor Protein Secretases/*chemistry/metabolism
Crystallography, X-Ray
Humans
Membrane Proteins/*chemistry/metabolism
Models, Molecular
*Proteolysis
Receptors, Notch/metabolism
Signal Transduction
Adam10
Notch signaling
X-ray crystallography
amyloid precursor protein
Abstract: Cleavage of membrane-anchored proteins by ADAM (a disintegrin and metalloproteinase) endopeptidases plays a key role in a wide variety of biological signal transduction and protein turnover processes. Among ADAM family members, ADAM10 stands out as particularly important because it is both responsible for regulated proteolysis of Notch receptors and catalyzes the non-amyloidogenic α-secretase cleavage of the Alzheimer's precursor protein (APP). We present here the X-ray crystal structure of the ADAM10 ectodomain, which, together with biochemical and cellular studies, reveals how access to the enzyme active site is regulated. The enzyme adopts an unanticipated architecture in which the C-terminal cysteine-rich domain partially occludes the enzyme active site, preventing unfettered substrate access. Binding of a modulatory antibody to the cysteine-rich domain liberates the catalytic domain from autoinhibition, enhancing enzymatic activity toward a peptide substrate. Together, these studies reveal a mechanism for regulation of ADAM activity and offer a roadmap for its modulation.
Notes: 1097-4172
Seegar, Tom C M
Killingsworth, Lauren B
Saha, Nayanendu
Meyer, Peter A
Patra, Dhabaleswar
Zimmerman, Brandon
Janes, Peter W
Rubinstein, Eric
Nikolov, Dimitar B
Skiniotis, Georgios
Kruse, Andrew C
Blacklow, Stephen C
P41 GM103403/GM/NIGMS NIH HHS/United States
T32 HL007627/HL/NHLBI NIH HHS/United States
R01 CA092433/CA/NCI NIH HHS/United States
R01 NS038486/NS/NINDS NIH HHS/United States
R35 CA220340/CA/NCI NIH HHS/United States
P01 CA119070/CA/NCI NIH HHS/United States
Journal Article
Cell. 2017 Dec 14;171(7):1638-1648.e7. doi: 10.1016/j.cell.2017.11.014. Epub 2017 Dec 7.
Author Address: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
SBGrid Initiative, Harvard Medical School, Boston, MA 02115, USA.
Life Sciences Institute and Department of Biological Chemistry, University of Michigan, Ann Arbor, MI 48109, USA.
Department of Biochemistry and Molecular Biology, Monash University, VIC 3800, Australia.
Inserm and Université Paris-Sud, Institut André Lwoff, Villejuif, France.
Department of Molecular and Cellular Physiology, and Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA. Electronic address: stephen_blacklow@hms.harvard.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 353
Author: Seidah, N. G., Ladenheim, R., Mbikay, M., Hamelin, J., Lutfalla, G., Rougeon, F., Lazure, C. and Chrétien, M.
Year: 1989
Title: The cDNA structure of rat plasma kallikrein
Journal: Dna
Volume: 8
Issue: 8
Pages: 563-74
Epub Date: 1989/10/01
Date: Oct
Short Title: The cDNA structure of rat plasma kallikrein
Alternate Journal: DNA (Mary Ann Liebert, Inc.)
ISSN: 0198-0238 (Print)
0198-0238
DOI: 10.1089/dna.1989.8.563
Accession Number: 2598771
Keywords: Amino Acid Sequence
Animals
Base Sequence
Blotting, Northern
Blotting, Southern
Cloning, Molecular
Dna
Kallikreins/*genetics
Liver/enzymology
Mice
Molecular Sequence Data
Polymerase Chain Reaction
RNA, Messenger/metabolism
Rats
Rats, Inbred Strains
Restriction Mapping
Abstract: From a liver cDNA library we have isolated and characterized the cDNA encoding rat plasma kallikrein. The cDNA structure contains 2,456 nucleotides with a 2,082-nucleotide-long open reading frame. Protein sequence data suggest that the signal peptide is 19 amino acids long. This results in a mature plasma prekallikrein containing 619 amino acids. Determination of tissue distributions using Northern blot analysis (3.0-kb transcript) and the polymerase chain-reaction methodology on RNA preparations demonstrated that in the rat the liver is the main source of this enzyme. Southern blots suggested the presence of a single gene coding for rat plasma kallikrein. Finally, although Southern blots revealed a homologous gene in mouse, the mRNA corresponding to the mouse hepatic proteinase is barely detectable on Northern blots, suggesting inefficient transcription or high turnover of the mRNA in this species.
Notes: Seidah, N G
Ladenheim, R
Mbikay, M
Hamelin, J
Lutfalla, G
Rougeon, F
Lazure, C
Chrétien, M
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
DNA. 1989 Oct;8(8):563-74. doi: 10.1089/dna.1989.8.563.
Author Address: Laboratory of Biochemical Neuronendocrinology, Clinical Research Institute of Montreal, Quebec, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 456
Author: Seidah, N. G., Rochemont, J., Hamelin, J., Lis, M. and Chrétien, M.
Year: 1981
Title: Primary structure of the major human pituitary pro-opiomelanocortin NH2-terminal glycopeptide. Evidence for an aldosterone-stimulating activity
Journal: J Biol Chem
Volume: 256
Issue: 15
Pages: 7977-84
Epub Date: 1981/08/10
Date: Aug 10
Short Title: Primary structure of the major human pituitary pro-opiomelanocortin NH2-terminal glycopeptide. Evidence for an aldosterone-stimulating activity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6267033
Keywords: Adrenal Gland Neoplasms/metabolism
*Adrenocorticotropic Hormone/pharmacology
Aldosterone/*metabolism
Amino Acid Sequence
Animals
Base Sequence
Cyanogen Bromide
Dna
Genes
*Glycopeptides/pharmacology
Humans
Iodoacetamide
*Peptide Fragments
Pituitary Gland/*analysis
*Pituitary Hormones, Anterior/pharmacology
*Pro-Opiomelanocortin
Species Specificity
Swine
Abstract: The isolation and complete purification of a human glycopeptide representing the major immunoreactive form of the pituitary NH2-terminal segment of pro-opiomelanocortin is presented. The complete sequence of this peptide was determined following CNBr fragmentation and it is shown to be 76 amino acids long. It bears an O-glycosylation site at Thr 45 and an N-glycosidic linkage at Asn 65. Compared to the reported genomic DNA sequence (Chang, A. C. Y., Cochet, M., and Cohen, S. N. (1980) Proc. Natl. Acad. Sci. U. S. A. 77, 4890-4894), one variation exists, namely Arg 22 replacing Gly 22. Two disulfide bridges linking Cys 2 to 8 and Cys 20 to 24 have been determined. Based on the sequence and disulfide bridge localization, a large degree of homology exists between the NH2-terminal sequence of the human peptide and all calcitonins, especially porcine calcitonin. The human NH2-terminal peptide is shown to stimulate the release of aldosterone from isolated cells of a human adrenal tumor, in at least equipotency to adrenocorticotropic hormone and the porcine NH2-terminal analogue, which is 100 times more potent than angiotensin II. Finally, this reported sequence completes that of the human DNA which was lacking the first 19 amino acids due to the presence of a 2-kilobase intron.
Notes: Seidah, N G
Rochemont, J
Hamelin, J
Lis, M
Chrétien, M
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1981 Aug 10;256(15):7977-84.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1940
Author: Selvan, N., George, S., Serajee, F. J., Shaw, M., Hobson, L., Kalscheuer, V., Prasad, N., Levy, S. E., Taylor, J., Aftimos, S., Schwartz, C. E., Huq, A. M., Gecz, J. and Wells, L.
Year: 2018
Title: O-GlcNAc transferase missense mutations linked to X-linked intellectual disability deregulate genes involved in cell fate determination and signaling
Journal: J Biol Chem
Volume: 293
Issue: 27
Pages: 10810-10824
Epub Date: 2018/05/18
Date: Jul 6
Short Title: O-GlcNAc transferase missense mutations linked to X-linked intellectual disability deregulate genes involved in cell fate determination and signaling
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA118.002583
PMCID: PMC6036218
Accession Number: 29769320
Keywords: Cell Differentiation
*Cell Lineage
Child
Crystallography, X-Ray
Embryonic Stem Cells/*metabolism/pathology
Female
Gene Expression Profiling
Gene Expression Regulation, Developmental
*Genes, X-Linked
*Genetic Markers
Humans
Infant, Newborn
Intellectual Disability/enzymology/*genetics/pathology
Male
*Mutation, Missense
N-Acetylglucosaminyltransferases/chemistry/*genetics/metabolism
Pedigree
Protein Conformation
Signal Transduction
*RNA-seq
*Xlid
*enzyme kinetics
*genetic disease
*glycosyltransferase
O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
O-linked N-acetylglucosamine (O-GlcNAc)
*intellectual disability
*post-translational modification
*transcriptomics
this article
Abstract: It is estimated that ∼1% of the world's population has intellectual disability, with males affected more often than females. OGT is an X-linked gene encoding for the enzyme O-GlcNAc transferase (OGT), which carries out the reversible addition of N-acetylglucosamine (GlcNAc) to Ser/Thr residues of its intracellular substrates. Three missense mutations in the tetratricopeptide (TPR) repeats of OGT have recently been reported to cause X-linked intellectual disability (XLID). Here, we report the discovery of two additional novel missense mutations (c.775 G>A, p.A259T, and c.1016 A>G, p.E339G) in the TPR domain of OGT that segregate with XLID in affected families. Characterization of all five of these XLID missense variants of OGT demonstrates modest declines in thermodynamic stability and/or activities of the variants. We engineered each of the mutations into a male human embryonic stem cell line using CRISPR/Cas9. Investigation of the global O-GlcNAc profile as well as OGT and O-GlcNAc hydrolase levels by Western blotting showed no gross changes in steady-state levels in the engineered lines. However, analyses of the differential transcriptomes of the OGT variant-expressing stem cells revealed shared deregulation of genes involved in cell fate determination and liver X receptor/retinoid X receptor signaling, which has been implicated in neuronal development. Thus, here we reveal two additional mutations encoding residues in the TPR regions of OGT that appear causal for XLID and provide evidence that the relatively stable and active TPR variants may share a common, unelucidated mechanism of altering gene expression profiles in human embryonic stem cells.
Notes: 1083-351x
Selvan, Nithya
George, Stephan
Serajee, Fatema J
Shaw, Marie
Hobson, Lynne
Kalscheuer, Vera
Prasad, Nripesh
Levy, Shawn E
Taylor, Juliet
Aftimos, Salim
Schwartz, Charles E
Huq, Ahm M
Gecz, Jozef
Orcid: 0000-0002-7884-6861
Wells, Lance
P41 GM103490/GM/NIGMS NIH HHS/United States
Case Reports
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2018 Jul 6;293(27):10810-10824. doi: 10.1074/jbc.RA118.002583. Epub 2018 May 16.
Author Address: From the Department of Biochemistry and Molecular Biology, Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602.
the Departments of Pediatrics and of Neurology, Wayne State University, Detroit, Michigan 48201.
the Adelaide Medical School, Faculty of Health and Medical Sciences, University of Adelaide, Adelaide South Australia 5006, Australia.
the Women's and Children's Hospital, North Adelaide, South Australia 5006, Australia.
the Research Group Development and Disease, Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany.
the Genomic Services Laboratory, HudsonAlpha Institute for Biotechnology, Huntsville, Alabama 35806.
the Genetic Health Services New Zealand-Northern Hub, Auckland City Hospital, Auckland 1142, New Zealand.
the Greenwood Genetic Center, Greenwood, South Carolina 29646, and.
the South Australian Health and Medical Research Institute, Adelaide, South Australia 5006, Australia.
From the Department of Biochemistry and Molecular Biology, Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602, lwells@ccrc.uga.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1692
Author: Selvan, N., Mariappa, D., van den Toorn, H. W., Heck, A. J., Ferenbach, A. T. and van Aalten, D. M.
Year: 2015
Title: The Early Metazoan Trichoplax adhaerens Possesses a Functional O-GlcNAc System
Journal: J Biol Chem
Volume: 290
Issue: 19
Pages: 11969-82
Epub Date: 2015/03/18
Date: May 8
Short Title: The Early Metazoan Trichoplax adhaerens Possesses a Functional O-GlcNAc System
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M114.628750
PMCID: PMC4424335
Accession Number: 25778404
Keywords: Acetylglucosamine/chemistry
Animals
Animals, Genetically Modified
Cell Nucleus/enzymology
Crosses, Genetic
Cytoplasm/enzymology
Drosophila melanogaster
HEK293 Cells
Humans
Inhibitory Concentration 50
Molecular Sequence Data
N-Acetylglucosaminyltransferases/*metabolism
Placozoa/*enzymology
Protein Processing, Post-Translational
Protein Structure, Tertiary
RNA Interference
Signal Transduction
Animal Model
Drosophila Genetics
Evolution
O-GlcNAcylation
Post-translational Modification (PTM)
Trichoplax adhaerens
Abstract: Protein O-GlcNAcylation is a reversible post-translational signaling modification of nucleocytoplasmic proteins that is essential for embryonic development in bilateria. In a search for a reductionist model to study O-GlcNAc signaling, we discovered the presence of functional O-GlcNAc transferase (OGT), O-GlcNAcase (OGA), and nucleocytoplasmic protein O-GlcNAcylation in the most basal extant animal, the placozoan Trichoplax adhaerens. We show via enzymatic characterization of Trichoplax OGT/OGA and genetic rescue experiments in Drosophila melanogaster that these proteins possess activities/functions similar to their bilaterian counterparts. The acquisition of O-GlcNAc signaling by metazoa may have facilitated the rapid and complex signaling mechanisms required for the evolution of multicellular organisms.
Notes: 1083-351x
Selvan, Nithya
Mariappa, Daniel
van den Toorn, Henk W P
Heck, Albert J R
Ferenbach, Andrew T
van Aalten, Daan M F
087590/Wellcome Trust/United Kingdom
G0900138/Medical Research Council/United Kingdom
MR/M004139/1/Medical Research Council/United Kingdom
WT087590MA/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2015 May 8;290(19):11969-82. doi: 10.1074/jbc.M114.628750. Epub 2015 Mar 16.
Author Address: From the Division of Molecular Microbiology and.
MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee, DD1 5EH, United Kingdom and.
the Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
From the Division of Molecular Microbiology and MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee, DD1 5EH, United Kingdom and dmfvanaalten@dundee.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1378
Author: Semavina, M., Saha, N., Kolev, M. V., Goldgur, Y., Giger, R. J., Himanen, J. P. and Nikolov, D. B.
Year: 2011
Title: Crystal structure of the Nogo-receptor-2
Journal: Protein Sci
Volume: 20
Issue: 4
Pages: 684-9
Epub Date: 2011/02/11
Date: Apr
Short Title: Crystal structure of the Nogo-receptor-2
Alternate Journal: Protein science : a publication of the Protein Society
ISSN: 0961-8368 (Print)
0961-8368
DOI: 10.1002/pro.597
PMCID: PMC3081546
Accession Number: 21308849
Keywords: Animals
Crystallography, X-Ray
GPI-Linked Proteins
Models, Molecular
Molecular Sequence Data
Myelin Proteins
Myelin-Associated Glycoprotein/metabolism
Nogo Receptor 1
Protein Binding
Protein Isoforms/*genetics/metabolism
*Protein Structure, Tertiary
Rats
Receptors, Cell Surface
Receptors, Peptide/*chemistry/metabolism
Recombinant Proteins/genetics/metabolism
Abstract: The inhibition of axon regeneration upon mechanical injury is dependent on interactions between Nogo receptors (NgRs) and their myelin-derived ligands. NgRs are composed of a leucine-rich repeat (LRR) region, thought to be structurally similar among the different isoforms of the receptor, and a divergent "stalk" region. It has been shown by others that the LRR and stalk regions of NgR1 and NgR2 have distinct roles in conferring binding affinity to the myelin associated glycoprotein (MAG) in vivo. Here, we show that purified recombinant full length NgR1 and NgR2 maintain significantly higher binding affinity for purified MAG as compared to the isolated LRR region of either NgR1 or NgR2. We also present the crystal structure of the LRR and part of the stalk regions of NgR2 and compare it to the previously reported NgR1 structure with respect to the distinct signaling properties of the two receptor isoforms.
Notes: 1469-896x
Semavina, Mariya
Saha, Nayanendu
Kolev, Momchil V
Goldgur, Yehuda
Giger, Roman J
Himanen, Juha P
Nikolov, Dimitar B
NS047333/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Protein Sci. 2011 Apr;20(4):684-9. doi: 10.1002/pro.597.
Author Address: Structural Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York 10065, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1813
Author: Seo, H. G., Kim, H. B., Kang, M. J., Ryum, J. H., Yi, E. C. and Cho, J. W.
Year: 2016
Title: Identification of the nuclear localisation signal of O-GlcNAc transferase and its nuclear import regulation
Journal: Sci Rep
Volume: 6
Pages: 34614
Epub Date: 2016/10/08
Date: Oct 7
Short Title: Identification of the nuclear localisation signal of O-GlcNAc transferase and its nuclear import regulation
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/srep34614
PMCID: PMC5054401
Accession Number: 27713473
Keywords: Active Transport, Cell Nucleus/physiology
Cell Nucleus/*enzymology/genetics
HEK293 Cells
HeLa Cells
Humans
N-Acetylglucosaminyltransferases/genetics/*metabolism
Nuclear Localization Signals/genetics/*metabolism
Abstract: Nucleocytoplasmic O-GlcNAc transferase (OGT) attaches a single GlcNAc to hydroxyl groups of serine and threonine residues. Although the cellular localisation of OGT is important to regulate a variety of cellular processes, the molecular mechanisms regulating the nuclear localisation of OGT is unclear. Here, we characterised three amino acids (DFP; residues 451-453) as the nuclear localisation signal of OGT and demonstrated that this motif mediated the nuclear import of non-diffusible β-galactosidase. OGT bound the importin α5 protein, and this association was abolished when the DFP motif of OGT was mutated or deleted. We also revealed that O-GlcNAcylation of Ser389, which resides in the tetratricopeptide repeats, plays an important role in the nuclear localisation of OGT. Our findings may explain how OGT, which possesses a NLS, exists in the nucleus and cytosol simultaneously.
Notes: 2045-2322
Seo, Hyeon Gyu
Kim, Han Byeol
Kang, Min Jueng
Ryum, Joo Hwan
Yi, Eugene C
Cho, Jin Won
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2016 Oct 7;6:34614. doi: 10.1038/srep34614.
Author Address: Department of Integrated OMICS for Biomedical Science, Graduate School, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul 03722, Republic of Korea.
Department of Molecular Medicine and Biopharmaceutical Sciences, School of Convergence Science and Technology and College of Medicine or College of Pharmacy, Seoul National University, 28 Yeongeon-dong, Jongno-gu, Seoul 03080, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1083
Author: Sevigny, M. B., Li, C. F., Alas, M. and Hughes-Fulford, M.
Year: 2006
Title: Glycosylation regulates turnover of cyclooxygenase-2
Journal: FEBS Lett
Volume: 580
Issue: 28-29
Pages: 6533-6
Epub Date: 2006/11/23
Date: Dec 11
Short Title: Glycosylation regulates turnover of cyclooxygenase-2
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2006.10.073
Accession Number: 17113084
Keywords: Animals
Arachidonic Acid/pharmacology
Asparagine/genetics
COS Cells
Chlorocebus aethiops
Cyclooxygenase 2/*metabolism
Glycosylation/drug effects
Humans
Isoenzymes/metabolism
Membrane Proteins/*metabolism
Mutagenesis/drug effects
Abstract: Cyclooxygenase-2 (COX-2) catalyzes the rate-limiting step in the prostanoid biosynthesis pathway, converting arachidonic acid into prostaglandin H(2). COX-2 exists as 72 and 74kDa glycoforms, the latter resulting from an additional oligosaccharide chain at residue Asn(580). In this study, Asn(580) was mutated to determine the biological significance of this variable glycosylation. COS-1 cells transfected with the mutant gene were unable to express the 74kDa glycoform and were found to accumulate more COX-2 protein and have five times greater COX-2 activity than cells expressing both glycoforms. Thus, COX-2 turnover appears to depend upon glycosylation of the 72kDa glycoform.
Notes: Sevigny, Mary B
Li, Chai-Fei
Alas, Monika
Hughes-Fulford, Millie
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
England
FEBS Lett. 2006 Dec 11;580(28-29):6533-6. doi: 10.1016/j.febslet.2006.10.073. Epub 2006 Nov 9.
Author Address: Department of Natural Sciences and Mathematics, Dominican University of California, San Rafael, CA 94901, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 615
Author: Sgroi, D., Nocks, A. and Stamenkovic, I.
Year: 1996
Title: A single N-linked glycosylation site is implicated in the regulation of ligand recognition by the I-type lectins CD22 and CD33
Journal: J Biol Chem
Volume: 271
Issue: 31
Pages: 18803-9
Epub Date: 1996/08/02
Date: Aug 2
Short Title: A single N-linked glycosylation site is implicated in the regulation of ligand recognition by the I-type lectins CD22 and CD33
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.31.18803
Accession Number: 8702538
Keywords: Amino Acid Sequence
Animals
Antigens, CD/*chemistry/genetics/*metabolism
Antigens, Differentiation, B-Lymphocyte/*chemistry/genetics/*metabolism
Antigens, Differentiation, Myelomonocytic/*chemistry/genetics/*metabolism
Binding Sites/genetics
Cell Adhesion
*Cell Adhesion Molecules
Cell Line
Glycosylation
Humans
*Lectins
Ligands
Membrane Glycoproteins/genetics
Molecular Sequence Data
Molecular Structure
Mutagenesis, Site-Directed
Myelin-Associated Glycoprotein/genetics
Rats
Receptors, Immunologic/genetics
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Homology, Amino Acid
Sialic Acid Binding Ig-like Lectin 1
Sialic Acid Binding Ig-like Lectin 2
Sialic Acid Binding Ig-like Lectin 3
Transfection
Abstract: CD22 is an immunoglobulin superfamily B lymphocyte-specific adhesion receptor and a member of the recently identified I-type class of lectins. Recent work has shown that CD22 specifically recognizes sialic acid linked alpha2,6 to terminal N-linked oligosaccharides on selected cell surface glycoproteins. CD22-ligand interaction is regulated by the activity of a beta-galactoside alpha2, 6-sialyltransferase that can inactivate CD22-mediated binding by sialylating the CD22 receptor itself. These observations suggest that N-linked glycosylation sites on the CD22 molecule may play a role in the regulation of CD22-mediated adhesion. In this work we have performed site-specific mutagenesis of potential N-linked glycosylation sites on CD22 in an effort to determine whether they might be involved in ligand recognition. We show that mutation of a single potential N-linked glycosylation site in the first immunoglobulin domain of CD22 completely abrogates ligand recognition. Interestingly, this site is characterized by the sequence NCT, where the cysteine is thought to be involved in an intrachain disulfide bond. Site-directed mutagenesis of similar NC(T/S) motifs in the first or second Ig domains of the I-type lectins myelin-associated glycoprotein, and sialoadhesin did not disrupt their ability to mediate sialic acid binding. In contrast, mutation of a NCS motif in the first Ig domain of the I-type lectin CD33 unmasked its sialic acid binding activity. These observations suggest that a single N-linked glycosylation site located at a similar position in the CD22 and CD33 glycoproteins is critical for regulating ligand recognition by both receptors.
Notes: Sgroi, D
Nocks, A
Stamenkovic, I
AI/01252/AI/NIAID NIH HHS/United States
GM/AI 48614/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1996 Aug 2;271(31):18803-9. doi: 10.1074/jbc.271.31.18803.
Author Address: Department of Pathology, Harvard Medical School, Boston, Massachusetts 02129, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1816
Author: Shah, P., Cheasty, A., Foxton, C., Raynham, T., Farooq, M., Gutierrez, I. F., Lejeune, A., Pritchard, M., Turnbull, A., Pang, L., Owen, P., Boyd, S., Stowell, A., Jordan, A., Hamilton, N. M., Hitchin, J. R., Stockley, M., MacDonald, E., Quesada, M. J., Trivier, E., Skeete, J., Ovaa, H., Moolenaar, W. H. and Ryder, H.
Year: 2016
Title: Discovery of potent inhibitors of the lysophospholipase autotaxin
Journal: Bioorg Med Chem Lett
Volume: 26
Issue: 22
Pages: 5403-5410
Epub Date: 2016/10/27
Date: Nov 15
Short Title: Discovery of potent inhibitors of the lysophospholipase autotaxin
Alternate Journal: Bioorganic & medicinal chemistry letters
ISSN: 0960-894x
DOI: 10.1016/j.bmcl.2016.10.036
Accession Number: 27780639
Keywords: Animals
Antineoplastic Agents/chemistry/pharmacokinetics/pharmacology
Crystallography, X-Ray
Enzyme Inhibitors/*chemistry/pharmacokinetics/*pharmacology
Humans
Lysophospholipase/*antagonists & inhibitors/metabolism
Lysophospholipids/*metabolism
Mice
Molecular Docking Simulation
Molecular Targeted Therapy
Neoplasms/drug therapy/enzymology
Phosphoric Diester Hydrolases/*metabolism
Pyridines/chemistry/pharmacokinetics/pharmacology
*Autotaxin (ATX)
*Cancer
*Lysophosphatidic acid (LPA)
*Lysophosphatidylcholine (LPC)
Abstract: The autotaxin-lysophosphatidic acid (ATX-LPA) axis has been implicated in several disease conditions including inflammation, fibrosis and cancer. This makes ATX an attractive drug target and its inhibition may lead to useful therapeutic agents. Through a high throughput screen (HTS) we identified a series of small molecule inhibitors of ATX which have subsequently been optimized for potency, selectivity and developability properties. This has delivered drug-like compounds such as 9v (CRT0273750) which modulate LPA levels in plasma and are suitable for in vivo studies. X-ray crystallography has revealed that these compounds have an unexpected binding mode in that they do not interact with the active site zinc ions but instead occupy the hydrophobic LPC pocket extending from the active site of ATX together with occupying the LPA 'exit' channel.
Notes: 1464-3405
Shah, Pritom
Cheasty, Anne
Foxton, Caroline
Raynham, Tony
Farooq, Muddasar
Gutierrez, Irene Farre
Lejeune, Aurore
Pritchard, Michelle
Turnbull, Andrew
Pang, Leon
Owen, Paul
Boyd, Susan
Stowell, Alexandra
Jordan, Allan
Hamilton, Niall M
Hitchin, James R
Stockley, Martin
MacDonald, Ellen
Quesada, Mar Jimenez
Trivier, Elisabeth
Skeete, Jana
Ovaa, Huib
Moolenaar, Wouter H
Ryder, Hamish
Journal Article
England
Bioorg Med Chem Lett. 2016 Nov 15;26(22):5403-5410. doi: 10.1016/j.bmcl.2016.10.036. Epub 2016 Oct 14.
Author Address: Cancer Research Technology, Discovery Laboratories, Babraham Research Campus, Cambridge CB22 3AT, UK.
Drug Discovery Unit, Cancer Research UK Manchester Institute, The University of Manchester, Wilmslow Road, Manchester M20 4BX, UK.
Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 287
Author: Shailubhai, K., Pukazhenthi, B. S., Saxena, E. S., Varma, G. M. and Vijay, I. K.
Year: 1991
Title: Glucosidase I, a transmembrane endoplasmic reticular glycoprotein with a luminal catalytic domain
Journal: J Biol Chem
Volume: 266
Issue: 25
Pages: 16587-93
Epub Date: 1991/09/05
Date: Sep 5
Short Title: Glucosidase I, a transmembrane endoplasmic reticular glycoprotein with a luminal catalytic domain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1885588
Keywords: Amino Acid Sequence
Animals
Breast/enzymology
Electrophoresis, Polyacrylamide Gel
Endoplasmic Reticulum/*enzymology
Ethylmaleimide
Intracellular Membranes/*enzymology
Membrane Proteins/*metabolism
Molecular Sequence Data
Rats
Trypsin
alpha-Glucosidases/chemistry/*metabolism
Abstract: We have analyzed the functional domain structure of rat mammary glucosidase I, an enzyme involved in N-linked glycoprotein processing, using biochemical and immunological approaches. The enzyme contains a high mannose type sugar chain that can be cleaved by endo-beta-N-acetyl-D-glucosaminidase H without significantly affecting the catalytic activity. Based on trypsin digestion pattern and the data on membrane topography, glucosidase I constitutes a single polypeptide chain of 85 kDa with two contiguous domains: a membrane-bound domain that anchors the protein to the endoplasmic reticulum and a luminal domain. A catalytically active 39-kDa domain could be released from membranes by limited proteolysis of saponin-permeabilized membranes with trypsin. This domain appeared to contain the active site of the enzyme and had the ability to bind to glucosidase I-specific affinity gel. Phase partitioning with Triton X-114 indicated the amphiphilic nature of the native enzyme, consistent with its location as an integral membrane protein, whereas the 39-kDa fragment partitioned in the aqueous phase, a characteristic of soluble polypeptide. These results indicate that glucosidase I is a transmembrane protein with a luminally oriented catalytic domain. Such an orientation of the catalytic domain may facilitate the sequential processing of asparagine-linked oligosaccharide, soon after its transfer en bloc by the oligosaccharyl transferase complex in the lumen of endoplasmic reticulum.
Notes: Shailubhai, K
Pukazhenthi, B S
Saxena, E S
Varma, G M
Vijay, I K
DK-19682/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1991 Sep 5;266(25):16587-93.
Author Address: Department of Animal Sciences, University of Maryland, College Park 20742.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2078
Author: Shajahan, A., Archer-Hartmann, S., Supekar, N. T., Gleinich, A. S., Heiss, C. and Azadi, P.
Year: 2021
Title: Comprehensive characterization of N- and O- glycosylation of SARS-CoV-2 human receptor angiotensin converting enzyme 2
Journal: Glycobiology
Volume: 31
Issue: 4
Pages: 410-424
Epub Date: 2020/11/03
Date: May 3
Short Title: Comprehensive characterization of N- and O- glycosylation of SARS-CoV-2 human receptor angiotensin converting enzyme 2
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwaa101
PMCID: PMC7665489
Accession Number: 33135055
Keywords: Angiotensin-Converting Enzyme 2/*chemistry/*metabolism
COVID-19/*metabolism/virology
Glycomics
Glycosylation
HEK293 Cells
Humans
Polysaccharides/*metabolism
Protein Binding
Protein Conformation
SARS-CoV-2/*physiology
*ACE2 glycosylation
*ACE2 site mapping
*COVID-19 receptor
*SARS-CoV-2 receptor
*hACE2 N-glycosylation
Abstract: The emergence of the coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has created the need for development of new therapeutic strategies. Understanding the mode of viral attachment, entry and replication has become a key aspect of such interventions. The coronavirus surface features a trimeric spike (S) protein that is essential for viral attachment, entry and membrane fusion. The S protein of SARS-CoV-2 binds to human angiotensin converting enzyme 2 (hACE2) for entry. Herein, we describe glycomic and glycoproteomic analysis of hACE2 expressed in HEK293 cells. We observed high glycan occupancy (73.2 to 100%) at all seven possible N-glycosylation sites and surprisingly detected one novel O-glycosylation site. To deduce the detailed structure of glycan epitopes on hACE2 that may be involved in viral binding, we have characterized the terminal sialic acid linkages, the presence of bisecting GlcNAc and the pattern of N-glycan fucosylation. We have conducted extensive manual interpretation of each glycopeptide and glycan spectrum, in addition to using bioinformatics tools to validate the hACE2 glycosylation. Our elucidation of the site-specific glycosylation and its terminal orientations on the hACE2 receptor, along with the modeling of hACE2 glycosylation sites can aid in understanding the intriguing virus-receptor interactions and assist in the development of novel therapeutics to prevent viral entry. The relevance of studying the role of ACE2 is further increased due to some recent reports about the varying ACE2 dependent complications with regard to age, sex, race and pre-existing conditions of COVID-19 patients.
Notes: 1460-2423
Shajahan, Asif
Orcid: 0000-0002-4804-1130
Archer-Hartmann, Stephanie
Supekar, Nitin T
Gleinich, Anne S
Orcid: 0000-0001-9305-4339
Heiss, Christian
Azadi, Parastoo
Journal Article
Glycobiology. 2021 May 3;31(4):410-424. doi: 10.1093/glycob/cwaa101.
Author Address: Complex Carbohydrate Research Center, The University of Georgia, 315 Riverbend Rd, Athens, GA 30602, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2072
Author: Shajahan, A., Supekar, N. T., Gleinich, A. S. and Azadi, P.
Year: 2020
Title: Deducing the N- and O-glycosylation profile of the spike protein of novel coronavirus SARS-CoV-2
Journal: Glycobiology
Volume: 30
Issue: 12
Pages: 981-988
Epub Date: 2020/05/05
Date: Dec 9
Short Title: Deducing the N- and O-glycosylation profile of the spike protein of novel coronavirus SARS-CoV-2
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwaa042
PMCID: PMC7239183
Accession Number: 32363391
Keywords: COVID-19/*genetics/pathology/virology
Coronavirus Infections
Humans
Pandemics
Polysaccharides/*genetics/metabolism
Protein Binding/genetics
SARS-CoV-2/*genetics/pathogenicity
Spike Glycoprotein, Coronavirus/genetics/*isolation & purification
*Covid-19
*S1 S2 glycosylation
*SARS-CoV-2 glycosylation
*coronavirus vaccine
*spike protein
Abstract: The current emergence of the novel coronavirus pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) demands the development of new therapeutic strategies to prevent rapid progress of mortalities. The coronavirus spike (S) protein, which facilitates viral attachment, entry and membrane fusion is heavily glycosylated and plays a critical role in the elicitation of the host immune response. The spike protein is comprised of two protein subunits (S1 and S2), which together possess 22 potential N-glycosylation sites. Herein, we report the glycosylation mapping on spike protein subunits S1 and S2 expressed on human cells through high-resolution mass spectrometry. We have characterized the quantitative N-glycosylation profile on spike protein and interestingly, observed unexpected O-glycosylation modifications on the receptor-binding domain of spike protein subunit S1. Even though O-glycosylation has been predicted on the spike protein of SARS-CoV-2, this is the first report of experimental data for both the site of O-glycosylation and identity of the O-glycans attached on the subunit S1. Our data on the N- and O-glycosylation are strengthened by extensive manual interpretation of each glycopeptide spectra in addition to using bioinformatics tools to confirm the complexity of glycosylation in the spike protein. The elucidation of the glycan repertoire on the spike protein provides insights into the viral binding studies and more importantly, propels research toward the development of a suitable vaccine candidate.
Notes: 1460-2423
Shajahan, Asif
Supekar, Nitin T
Gleinich, Anne S
Azadi, Parastoo
S10 OD018530/OD/NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Glycobiology. 2020 Dec 9;30(12):981-988. doi: 10.1093/glycob/cwaa042.
Author Address: Complex Carbohydrate Research Center, The University of Georgia, 315 Riverbend Rd, Athens, GA 30602, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1933
Author: Shan, H., Sun, J., Shi, M., Liu, X., Shi, Z., Yu, W. and Gu, Y.
Year: 2018
Title: Generation and characterization of a site-specific antibody for SIRT1 O-GlcNAcylated at serine 549
Journal: Glycobiology
Volume: 28
Issue: 7
Pages: 482-487
Epub Date: 2018/04/25
Date: Jul 1
Short Title: Generation and characterization of a site-specific antibody for SIRT1 O-GlcNAcylated at serine 549
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwy040
Accession Number: 29688431
Keywords: 3T3 Cells
Animals
Antibodies/*immunology
Antibody Specificity
Female
Glycosylation
HEK293 Cells
Humans
Mice
Mice, Inbred BALB C
*Protein Processing, Post-Translational
Rabbits
Sirtuin 1/*immunology/metabolism
Abstract: O-linked N-acetyl-β-d-glucosamine (O-GlcNAc) is a dynamic post-translational modification that modifies thousands of proteins. However, the roles and mechanisms of O-GlcNAcylation have been clarified in only a few proteins, and one of the main reasons for this is the lack of site-specific anti-O-GlcNAc antibodies. Recently, we found that SIRT1, which is an NAD+-dependent deacetylase, is O-GlcNAcylated at the serine 549 site (S549) and plays a cytoprotective role under stress. However, the mechanism underlying the roles of SIRT1 O-GlcNAcylation remains unclear. Here, we describe a site-specific antibody for SIRT1 O-GlcNAcylated at S549, named SIRT1-549-O. This antibody can be used for immunoprecipitation and western blotting assays, and it can be used to recognize the endogenous levels of both human and mouse SIRT1 O-GlcNAcylation. Therefore, this antibody not only provides an effective method to further understand the roles of SIRT1 O-GlcNAcylation but also makes it possible to discover the genetic and pharmacological factors that could regulate SIRT1 activity by modulating its O-GlcNAcylation.
Notes: 1460-2423
Shan, Hui
Sun, Jiahui
Shi, Minghui
Liu, Xue
Shi, Zhu
Yu, Wengong
Gu, Yuchao
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2018 Jul 1;28(7):482-487. doi: 10.1093/glycob/cwy040.
Author Address: Key Laboratory of Marine Drugs, Ministry of Education; School of Medicine and Pharmacy, Ocean University of China, 5 Yushan Road, Qingdao, China.
Laboratory for Marine Drugs and Bioproducts of Qingdao National Laboratory for Marine Science and Technology, Qingdao, China.
Key Laboratory of Glycoscience & Glycotechnology of Shandong Province, Qingdao, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1951
Author: Shan, S., Chatterjee, A., Qiu, Y., Hammes, H. P., Wieland, T. and Feng, Y.
Year: 2018
Title: O-GlcNAcylation of FoxO1 mediates nucleoside diphosphate kinase B deficiency induced endothelial damage
Journal: Sci Rep
Volume: 8
Issue: 1
Pages: 10581
Epub Date: 2018/07/14
Date: Jul 12
Short Title: O-GlcNAcylation of FoxO1 mediates nucleoside diphosphate kinase B deficiency induced endothelial damage
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/s41598-018-28892-y
PMCID: PMC6043576
Accession Number: 30002415
Keywords: Acetylglucosamine/metabolism
Angiopoietin-2/*genetics/metabolism
Animals
Cell Nucleus/metabolism
Diabetic Retinopathy/genetics/*pathology
Disease Models, Animal
Endothelium, Vascular/cytology/enzymology/pathology
Forkhead Box Protein O1/genetics/*metabolism
Gene Knockdown Techniques
Human Umbilical Vein Endothelial Cells
Humans
Mice
Mice, Knockout
NM23 Nucleoside Diphosphate Kinases/*deficiency/genetics/*metabolism
Primary Cell Culture
RNA, Small Interfering/metabolism
Retina/cytology/enzymology/*pathology
Retinal Vessels/cytology/enzymology/pathology
Up-Regulation
Abstract: Nucleoside diphosphate kinase B (NDPK-B) acts as a protective factor in the retinal vasculature. NDPK-B deficiency leads to retinal vasoregression mimicking diabetic retinopathy (DR). Angiopoetin 2 (Ang-2), an initiator of retinal vasoregression in DR, is upregulated in NDPK-B deficient retinas and in NDPK-B depleted endothelial cells (ECs) in vitro. We therefore investigated the importance of Ang-2 in NDPK-B deficient retinas and characterized the mechanisms of Ang-2 upregulation upon NDPK-B depletion in cultured ECs. The crucial role of retinal Ang-2 in the initiation of vasoregression was verified by crossing NDPK-B deficient with Ang-2 haplodeficient mice. On the molecular level, FoxO1, a transcription factor regulating Ang-2, was upregulated in NDPK-B depleted ECs. Knockdown of FoxO1 abolished the elevation of Ang-2 induced by NDPK-B depletion. Furthermore O-GlcNAcylated FoxO1 was found preferentially in the nucleus. An increased O-GlcNAcylation of FoxO1 was revealed upon NDPK-B depletion. In accordance, the inhibition of protein O-GlcNAcylation normalized NDPK-B depletion induced Ang-2 upregulation. In summary, we demonstrated that the upregulation of Ang-2 upon NDPK-B deficiency is driven by O-GlcNAcylation of FoxO1. Our data provide evidence for a central role of protein O-GlcNAcylation in NDPK-B associated vascular damage and point to the hexosamine pathway as an important target in retinal vasoregression.
Notes: 2045-2322
Shan, Shenliang
Chatterjee, Anupriya
Qiu, Yi
Hammes, Hans-Peter
Wieland, Thomas
Feng, Yuxi
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Sci Rep. 2018 Jul 12;8(1):10581. doi: 10.1038/s41598-018-28892-y.
Author Address: Experimental Pharmacology Mannheim (EPM), European Center of Angioscience, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany.
5th Medical Clinic, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany.
DZHK (German Centre for Cardiovascular Research), partner site Heidelberg/Mannheim, Mannheim, Germany.
Experimental Pharmacology Mannheim (EPM), European Center of Angioscience, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany. yuxi.feng@medma.uni-heidelberg.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1857
Author: Sharif, S., Shi, J., Bourakba, M., Ruijtenbeek, R. and Pieters, R. J.
Year: 2017
Title: Measuring O-GlcNAc cleavage by OGA and cell lysates on a peptide microarray
Journal: Anal Biochem
Volume: 532
Pages: 12-18
Epub Date: 2017/06/03
Date: Sep 1
Short Title: Measuring O-GlcNAc cleavage by OGA and cell lysates on a peptide microarray
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697
DOI: 10.1016/j.ab.2017.05.027
Accession Number: 28571750
Keywords: Acetylglucosamine/*metabolism
Caco-2 Cells
HT29 Cells
Humans
MCF-7 Cells
Microarray Analysis
Neoplasms/*metabolism/pathology
Peptide Fragments/*analysis/metabolism
Protein Processing, Post-Translational
Signal Transduction
beta-N-Acetylhexosaminidases/*metabolism
*Cell lysate
*O-GlcNAcase
*O-GlcNAcylated peptide
*O-GlcNAcylation
*Peptide microarray
Abstract: O-GlcNAcylation is a post-translational modification resulting from the addition of an N-acetylglucosamine moiety to the hydroxyl groups of serine and threonine residues of nuclear and cytoplasmic proteins. In addition, O-GlcNAcylated proteins can be phosphorylated, which suggests the possibility for crosstalk between O-GlcNAcylation and phosphorylation. Dysregulation of O-GlcNAcylation affects cell signaling, transcriptional regulation, cell cycle control and can e.g. lead to tumorigenesis and tumor metastasis. There is a strong demand for efficient analytical techniques to better detect and investigate this abundant modification and its role in cancer. Herein we demonstrated the utility of an O-GlcNAcylated peptide array to examine O-GlcNAcase (OGA) activity and substrate specificity of both purified protein as well cell lysates of different cancer cell lines. Using this microarray, we clearly observed OGA activity and also inhibition thereof by OGA inhibitor thiamet G. Interestingly, different levels of OGA activity were observed of lysates derived from different cancer cell lines. This suggests that the tool may be useful in cancer research and biomarker development.
Notes: 1096-0309
Sharif, Suhela
Shi, Jie
Bourakba, Mostafa
Ruijtenbeek, Rob
Pieters, Roland J
Journal Article
United States
Anal Biochem. 2017 Sep 1;532:12-18. doi: 10.1016/j.ab.2017.05.027. Epub 2017 May 29.
Author Address: Department of Chemical Biology and Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, P. O. Box 80082, 3508 TB Utrecht, The Netherlands.
PamGene International BV, 's-Hertogenbosch, The Netherlands.
Department of Chemical Biology and Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, P. O. Box 80082, 3508 TB Utrecht, The Netherlands; PamGene International BV, 's-Hertogenbosch, The Netherlands.
Department of Chemical Biology and Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, P. O. Box 80082, 3508 TB Utrecht, The Netherlands. Electronic address: r.j.pieters@uu.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2004
Author: Sharif, S., Shi, J., Ruijtenbeek, R. and Pieters, R. J.
Year: 2019
Title: Study of cross talk between phosphatases and OGA on a ZO-3-derived peptide
Journal: Amino Acids
Volume: 51
Issue: 4
Pages: 739-743
Epub Date: 2019/02/07
Date: Apr
Short Title: Study of cross talk between phosphatases and OGA on a ZO-3-derived peptide
Alternate Journal: Amino acids
ISSN: 0939-4451
DOI: 10.1007/s00726-019-02699-1
Accession Number: 30725225
Keywords: Humans
N-Acetylglucosaminyltransferases/*metabolism
Peptide Fragments/*metabolism
Phosphoric Monoester Hydrolases/*metabolism
Phosphorylation
Protein Array Analysis
*Protein Processing, Post-Translational
Tyrosine/*metabolism
Zonula Occludens Proteins/*metabolism
Cross talk
O-GlcNAc
O-GlcNAcase
Peptide microarray
Phosphatase
Zonula-occludens-3 (ZO-3)
Abstract: O-GlcNAcylation, like phosphorylation, is a dynamic and rapid posttranslational modification which regulates many cellular processes. Phosphorylation on tyrosine and O-GlcNAcylation on nearby serine or threonine residues may modulate each other. Indeed, by using a microarray with a peptide model system based on the ZO-3 protein, extensive cross talk between O-GlcNAcylation by OGT and phosphorylation by kinases was observed. However, studying the effects of kinases and OGT without the reverse processes catalyzed by phosphatases and O-GlcNAcase (OGA) does not provide a complete picture of the cross talk. The study of the missing part showed that nearby phosphorylation affects the de-O-GlcNAcylation by OGA, but not to the same extent as it affects the O-GlcNAcylation by OGT. Both the phosphorylation and de-phosphorylation processes were only slightly affected by the presence of an O-GlcNAc residue on a nearby serine.
Notes: 1438-2199
Sharif, Suhela
Shi, Jie
Ruijtenbeek, Rob
Pieters, Roland J
Orcid: 0000-0003-4723-3584
201306180061/China Scholarship Council/
Journal Article
Austria
Amino Acids. 2019 Apr;51(4):739-743. doi: 10.1007/s00726-019-02699-1. Epub 2019 Feb 6.
Author Address: Department of Chemical Biology and Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, P. O. Box 80082, 3508 TB, Utrecht, The Netherlands.
PamGene International BV, 's-Hertogenbosch, The Netherlands.
Department of Chemical Biology and Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, P. O. Box 80082, 3508 TB, Utrecht, The Netherlands. R.J.Pieters@uu.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2030
Author: Sharma, N. S., Gupta, V. K., Dauer, P., Kesh, K., Hadad, R., Giri, B., Chandra, A., Dudeja, V., Slawson, C., Banerjee, S., Vickers, S. M., Saluja, A. and Banerjee, S.
Year: 2019
Title: O-GlcNAc modification of Sox2 regulates self-renewal in pancreatic cancer by promoting its stability
Journal: Theranostics
Volume: 9
Issue: 12
Pages: 3410-3424
Epub Date: 2019/07/10
Short Title: O-GlcNAc modification of Sox2 regulates self-renewal in pancreatic cancer by promoting its stability
Alternate Journal: Theranostics
ISSN: 1838-7640
DOI: 10.7150/thno.32615
PMCID: PMC6587167
Accession Number: 31281487
Keywords: Adenocarcinoma/pathology
Animals
Clustered Regularly Interspaced Short Palindromic Repeats
Gene Expression Regulation, Neoplastic
Glycosylation
Humans
Mice
Mutagenesis, Site-Directed
N-Acetylglucosaminyltransferases/metabolism
Neoplasm Recurrence, Local/*genetics/metabolism
Octamer Transcription Factor-3/metabolism
Pancreatic Neoplasms/*pathology
RNA-Seq
SOXB1 Transcription Factors/*genetics/metabolism
*O-GlcNAc
*Ogt
*Pancreatic cancer
*Sox2
*metabolism
*self-renewal
been licensed to Minneamrita Therapeutics, LLC. AKS is the co-founder and the Chief
Scientific Officer of this company. SB is a consultant with Minneamrita Therapeutics
LLC and this relationship is managed by University of Miami. The remaining authors
declare no conflict of interest.
Abstract: Pancreatic adenocarcinoma (PDAC) claims more than 90% of the patients diagnosed with the disease owing to its aggressive biology that is manifested by high rate of tumor recurrence. Aberrant upregulation in the transcriptional activity of proteins involved in self-renewal like Sox2, Oct4 and Nanog is instrumental in these recurrence phenomena. In cancer, Sox2 is aberrantly "turned-on" leading to activation of downstream genes those results in relapse of the tumor. Molecular mechanisms that regulate the activity of Sox2 in PDAC are not known. In the current study, we have studied the how glycosylation of Sox2 by O-GlcNAc transferase (OGT) can affect its transcriptional activity and thus regulate self-renewal in cancer. Methods: RNA-Seq analysis of CRISPR-OGTi PDAC cells indicated a deregulation of differentiation and self-renewal pathways in PDAC. Pancreatic tumor burden following inhibition of OGT in vivo was done by using small molecule inhibitor, OSMI, on subcutaneous implantation of PDAC cells. Sox2 activity assay was performed by Dual Luciferase Reporter Assay kit. Results: Our study shows for the first time that in PDAC, glycosylation of Sox2 by OGT stabilizes it in the nucleus. Site directed mutagenesis of this site (S246A) prevents this modification. We further show that inhibition of OGT delayed initiation of pancreatic tumors by inhibition of Sox2. We also show that targeting OGT in vivo with a small molecule-inhibitor OSMI, results in decreased tumor burden in PDAC. Conclusion: Understanding this mechanism of SOX2 regulation by its glycosylation is expected to pave the way for development of novel therapy that has the potential to eradicate the cells responsible for tumor-recurrence.
Notes: 1838-7640
Sharma, Nikita S
Gupta, Vineet K
Dauer, Patricia
Kesh, Kousik
Hadad, Roey
Giri, Bhuwan
Chandra, Anjali
Dudeja, Vikas
Slawson, Chad
Banerjee, Santanu
Vickers, Selwyn M
Saluja, Ashok
Banerjee, Sulagna
R01 CA124723/CA/NCI NIH HHS/United States
R01 CA170946/CA/NCI NIH HHS/United States
R01 CA184274/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Theranostics. 2019 May 24;9(12):3410-3424. doi: 10.7150/thno.32615. eCollection 2019.
Author Address: Department of Surgery, University of Miami, Miami, FL.
Department of Pharmacology, University of Minnesota, Minneapolis Minnesota.
Department of Psychology, Harvard University.
Sylvester Comprehensive Cancer Center, Miami, FL.
University of Kansas Medical Center, Kansas City, KS.
School of Medicine Dean's Office, University of Alabama at Birmingham.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2024
Author: Sharma, P., Reichert, M., Lu, Y., Markello, T. C., Adams, D. R., Steinbach, P. J., Fuqua, B. K., Parisi, X., Kaler, S. G., Vulpe, C. D., Anderson, G. J., Gahl, W. A. and Malicdan, M. C. V.
Year: 2019
Title: Biallelic HEPHL1 variants impair ferroxidase activity and cause an abnormal hair phenotype
Journal: PLoS Genet
Volume: 15
Issue: 5
Pages: e1008143
Epub Date: 2019/05/28
Date: May
Short Title: Biallelic HEPHL1 variants impair ferroxidase activity and cause an abnormal hair phenotype
Alternate Journal: PLoS genetics
ISSN: 1553-7390 (Print)
1553-7390
DOI: 10.1371/journal.pgen.1008143
PMCID: PMC6534290
Accession Number: 31125343
Keywords: Adult
Alleles
Animals
Binding Sites
Ceruloplasmin/metabolism
Child, Preschool
Copper/metabolism
Female
Gene Expression Regulation/genetics
Genetic Variation/genetics
HEK293 Cells
Hair
Humans
Iron/metabolism
Male
Membrane Proteins/genetics/metabolism
Mice
Mice, Knockout
Oxidation-Reduction
Oxidoreductases/*genetics/*metabolism
Phenotype
Abstract: Maintenance of the correct redox status of iron is functionally important for critical biological processes. Multicopper ferroxidases play an important role in oxidizing ferrous iron, released from the cells, into ferric iron, which is subsequently distributed by transferrin. Two well-characterized ferroxidases, ceruloplasmin (CP) and hephaestin (HEPH) facilitate this reaction in different tissues. Recently, a novel ferroxidase, Hephaestin like 1 (HEPHL1), also known as zyklopen, was identified. Here we report a child with compound heterozygous mutations in HEPHL1 (NM_001098672) who presented with abnormal hair (pili torti and trichorrhexis nodosa) and cognitive dysfunction. The maternal missense mutation affected mRNA splicing, leading to skipping of exon 5 and causing an in-frame deletion of 85 amino acids (c.809_1063del; p.Leu271_ala355del). The paternal mutation (c.3176T>C; p.Met1059Thr) changed a highly conserved methionine that is part of a typical type I copper binding site in HEPHL1. We demonstrated that HEPHL1 has ferroxidase activity and that the patient's two mutations exhibited loss of this ferroxidase activity. Consistent with these findings, the patient's fibroblasts accumulated intracellular iron and exhibited reduced activity of the copper-dependent enzyme, lysyl oxidase. These results suggest that the patient's biallelic variants are loss-of-function mutations. Hence, we generated a Hephl1 knockout mouse model that was viable and had curly whiskers, consistent with the hair phenotype in our patient. These results enhance our understanding of the function of HEPHL1 and implicate altered ferroxidase activity in hair growth and hair disorders.
Notes: 1553-7404
Sharma, Prashant
Orcid: 0000-0002-1165-980x
Reichert, Marie
Lu, Yan
Orcid: 0000-0002-7522-9454
Markello, Thomas C
Orcid: 0000-0002-2298-9759
Adams, David R
Steinbach, Peter J
Orcid: 0000-0002-6710-3472
Fuqua, Brie K
Orcid: 0000-0002-4875-6523
Parisi, Xenia
Kaler, Stephen G
Vulpe, Christopher D
Anderson, Gregory J
Gahl, William A
Orcid: 0000-0002-2494-6752
Malicdan, May Christine V
R01 DK056376/DK/NIDDK NIH HHS/United States
T32 HL069766/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, N.I.H., Intramural
PLoS Genet. 2019 May 24;15(5):e1008143. doi: 10.1371/journal.pgen.1008143. eCollection 2019 May.
Author Address: NIH Undiagnosed Diseases Program, Common Fund, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
Office of the Clinical Director, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
Iron Metabolism Laboratory, QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia.
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland Bethesda, Maryland, United States of America.
Center for Molecular Modeling, Center for Information Technology, National Institutes of Health, Bethesda, Maryland, United States of America.
Department of Medicine, University of California, Los Angeles, United States of America.
Section on Translational Neuroscience, Molecular Medicine Branch, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, United States of America.
Center for Environmental and Human Toxicology, Department of Physiological Sciences, University of Florida, Gainesville, Florida, United States of America.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1846
Author: Shen, B., Zhang, W., Shi, Z., Tian, F., Deng, Y., Sun, C., Wang, G., Qin, W. and Qian, X.
Year: 2017
Title: A novel strategy for global mapping of O-GlcNAc proteins and peptides using selective enzymatic deglycosylation, HILIC enrichment and mass spectrometry identification
Journal: Talanta
Volume: 169
Pages: 195-202
Epub Date: 2017/04/17
Date: Jul 1
Short Title: A novel strategy for global mapping of O-GlcNAc proteins and peptides using selective enzymatic deglycosylation, HILIC enrichment and mass spectrometry identification
Alternate Journal: Talanta
ISSN: 0039-9140
DOI: 10.1016/j.talanta.2017.03.049
Accession Number: 28411811
Keywords: Acetylglucosamine/*chemistry
Adult
Biomarkers/urine
Chromatography, High Pressure Liquid/*methods
Female
Glycosylation
Humans
Male
Peptide Fragments/chemistry/*urine
*Protein Processing, Post-Translational
Proteins/*analysis/chemistry
Proteome/*analysis
Proteomics
Tandem Mass Spectrometry/*methods
HILIC enrichment
O-GlcNAcylation
Selective enzymatic deglycosylation
Urinary protein
Abstract: O-GlcNAcylation is a kind of dynamic O-linked glycosylation of nucleocytoplasmic and mitochondrial proteins. It serves as a major nutrient sensor to regulate numerous biological processes including transcriptional regulation, cell metabolism, cellular signaling, and protein degradation. Dysregulation of cellular O-GlcNAcylated levels contributes to the etiologies of many diseases such as diabetes, neurodegenerative disease and cancer. However, deeper insight into the biological mechanism of O-GlcNAcylation is hampered by its extremely low stoichiometry and the lack of efficient enrichment approaches for large-scale identification by mass spectrometry. Herein, we developed a novel strategy for the global identification of O-GlcNAc proteins and peptides using selective enzymatic deglycosylation, HILIC enrichment and mass spectrometry analysis. Standard O-GlcNAc peptides can be efficiently enriched even in the presence of 500-fold more abundant non-O-GlcNAc peptides and identified by mass spectrometry with a low nanogram detection sensitivity. This strategy successfully achieved the first large-scale enrichment and characterization of O-GlcNAc proteins and peptides in human urine. A total of 474 O-GlcNAc peptides corresponding to 457 O-GlcNAc proteins were identified by mass spectrometry analysis, which is at least three times more than that obtained by commonly used enrichment methods. A large number of unreported O-GlcNAc proteins related to cell cycle, biological regulation, metabolic and developmental process were found in our data. The above results demonstrated that this novel strategy is highly efficient in the global enrichment and identification of O-GlcNAc peptides. These data provide new insights into the biological function of O-GlcNAcylation in human urine, which is correlated with the physiological states and pathological changes of human body and therefore indicate the potential of this strategy for biomarker discovery from human urine.
Notes: 1873-3573
Shen, Bingquan
Zhang, Wanjun
Shi, Zhaomei
Tian, Fang
Deng, Yulin
Sun, Changqing
Wang, Guangshun
Qin, Weijie
Qian, Xiaohong
Journal Article
Netherlands
Talanta. 2017 Jul 1;169:195-202. doi: 10.1016/j.talanta.2017.03.049. Epub 2017 Mar 18.
Author Address: School of Life Science, Beijing Institute of Technology, Beijing 100081, PR China; National Center for Protein Sciences Beijing, State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, BPRC-Tianjin Baodi Hospital Joint Center, Beijing 102206, PR China.
National Center for Protein Sciences Beijing, State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, BPRC-Tianjin Baodi Hospital Joint Center, Beijing 102206, PR China.
School of Life Science, Beijing Institute of Technology, Beijing 100081, PR China.
Tianjin Baodi Hospital, Tianjin 301800, PR China.
Tianjin Baodi Hospital, Tianjin 301800, PR China. Electronic address: wgs@bddhospital.com.
National Center for Protein Sciences Beijing, State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, BPRC-Tianjin Baodi Hospital Joint Center, Beijing 102206, PR China. Electronic address: aunp_dna@126.com.
School of Life Science, Beijing Institute of Technology, Beijing 100081, PR China; National Center for Protein Sciences Beijing, State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, BPRC-Tianjin Baodi Hospital Joint Center, Beijing 102206, PR China. Electronic address: qianxh1@163.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1455
Author: Shen, D. L., Gloster, T. M., Yuzwa, S. A. and Vocadlo, D. J.
Year: 2012
Title: Insights into O-linked N-acetylglucosamine ([0-9]O-GlcNAc) processing and dynamics through kinetic analysis of O-GlcNAc transferase and O-GlcNAcase activity on protein substrates
Journal: J Biol Chem
Volume: 287
Issue: 19
Pages: 15395-408
Epub Date: 2012/02/09
Date: May 4
Short Title: Insights into O-linked N-acetylglucosamine ([0-9]O-GlcNAc) processing and dynamics through kinetic analysis of O-GlcNAc transferase and O-GlcNAcase activity on protein substrates
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.310664
PMCID: PMC3346082
Accession Number: 22311971
Keywords: Acetylglucosamine/*metabolism
Adaptor Proteins, Signal Transducing/genetics/metabolism
Blotting, Western
Calcium-Calmodulin-Dependent Protein Kinase Type 4/genetics/metabolism
Glycosylation
Humans
Kinetics
Mass Spectrometry
Membrane Glycoproteins/genetics/metabolism
Mutation
N-Acetylglucosaminyltransferases/genetics/*metabolism
Nuclear Pore Complex Proteins/genetics/metabolism
Phosphorylation
Protein Processing, Post-Translational
Protein-Arginine N-Methyltransferases/genetics/metabolism
Proteins/genetics/*metabolism
Recombinant Proteins/genetics/metabolism
Serine/genetics/metabolism
Substrate Specificity
beta-N-Acetylhexosaminidases/genetics/*metabolism
tau Proteins/genetics/metabolism
Abstract: Cellular O-linked N-acetylglucosamine (O-GlcNAc) levels are modulated by two enzymes: uridine diphosphate-N-acetyl-D-glucosamine:polypeptidyltransferase (OGT) and O-GlcNAcase (OGA). To quantitatively address the activity of these enzymes on protein substrates, we generated five structurally diverse proteins in both unmodified and O-GlcNAc-modified states. We found a remarkably invariant upper limit for k(cat)/K(m) values for human OGA (hOGA)-catalyzed processing of these modified proteins, which suggests that hOGA processing is driven by the GlcNAc moiety and is independent of the protein. Human OGT (hOGT) activity ranged more widely, by up to 15-fold, suggesting that hOGT is the senior partner in fine tuning protein O-GlcNAc levels. This was supported by the observation that K(m,app) values for UDP-GlcNAc varied considerably (from 1 μM to over 20 μM), depending on the protein substrate, suggesting that some OGT substrates will be nutrient-responsive, whereas others are constitutively modified. The ratios of k(cat)/K(m) values obtained from hOGT and hOGA kinetic studies enable a prediction of the dynamic equilibrium position of O-GlcNAc levels that can be recapitulated in vitro and suggest the relative O-GlcNAc stoichiometries of target proteins in the absence of other factors. We show that changes in the specific activities of hOGT and hOGA measured in vitro on calcium/calmodulin-dependent kinase IV (CaMKIV) and its pseudophosphorylated form can account for previously reported changes in CaMKIV O-GlcNAc levels observed in cells. These studies provide kinetic evidence for the interplay between O-GlcNAc and phosphorylation on proteins and indicate that these effects can be mediated by changes in hOGT and hOGA kinetic activity.
Notes: 1083-351x
Shen, David L
Gloster, Tracey M
Yuzwa, Scott A
Vocadlo, David J
082572/Wellcome Trust/United Kingdom
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 May 4;287(19):15395-408. doi: 10.1074/jbc.M111.310664. Epub 2012 Feb 6.
Author Address: Department of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1820
Author: Shen, D. L., Liu, T. W., Zandberg, W., Clark, T., Eskandari, R., Alteen, M. G., Tan, H. Y., Zhu, Y., Cecioni, S. and Vocadlo, D.
Year: 2017
Title: Catalytic Promiscuity of O-GlcNAc Transferase Enables Unexpected Metabolic Engineering of Cytoplasmic Proteins with 2-Azido-2-deoxy-glucose
Journal: ACS Chem Biol
Volume: 12
Issue: 1
Pages: 206-213
Epub Date: 2016/12/10
Date: Jan 20
Short Title: Catalytic Promiscuity of O-GlcNAc Transferase Enables Unexpected Metabolic Engineering of Cytoplasmic Proteins with 2-Azido-2-deoxy-glucose
Alternate Journal: ACS chemical biology
ISSN: 1554-8929
DOI: 10.1021/acschembio.6b00876
Accession Number: 27935279
Keywords: Adaptor Proteins, Signal Transducing/metabolism
Animals
Azides/*chemistry/metabolism
COS Cells
Calcium-Calmodulin-Dependent Protein Kinase Type 4/metabolism
Chlorocebus aethiops
Deoxyglucose/*analogs & derivatives/chemistry
Glucose/analogs & derivatives/*chemistry/metabolism
Glycosylation
Humans
Membrane Glycoproteins/metabolism
Metabolic Engineering
Mice
N-Acetylglucosaminyltransferases/genetics/*metabolism
Nuclear Pore Complex Proteins/metabolism
Substrate Specificity
Tritium
Uridine Diphosphate Glucose/analogs & derivatives/chemistry/metabolism
beta-N-Acetylhexosaminidases/chemistry
tau Proteins/metabolism
Abstract: O-GlcNAc transferase (OGT) catalyzes the installation of N-acetylglucosamine (GlcNAc) O-linked to nucleocytoplasmic proteins (O-GlcNAc) within multicellular eukaryotes. OGT shows surprising tolerance for structural changes in the sugar component of its nucleotide sugar donor substrate, uridine diphosphate N-acetylglucosamine (UDP-GlcNAc). Here, we find that OGT uses UDP-glucose to install O-linked glucose (O-Glc) onto proteins only 25-fold less efficiently than O-GlcNAc. Spurred by this observation, we show that OGT transfers 2-azido-2-deoxy-d-glucose (GlcAz) in vitro from UDP-GlcAz to proteins. Further, feeding cells with per-O-acetyl GlcAz (AcGlcAz), in combination with inhibition or inducible knockout of OGT, shows OGT-dependent modification of nuclear and cytoplasmic proteins with O-GlcAz as detected using microscopy, immunoblot, and proteomics. We find that O-GlcAz is reversible within cells, and an unidentified cellular enzyme exists to cleave O-Glc that can also process O-GlcAz. We anticipate that AcGlcAz will prove to be a useful tool to study the O-GlcNAc modification. We also speculate that, given the high concentration of UDP-Glc within certain mammalian tissues, O-Glc may exist within mammals and serve as a physiologically relevant modification.
Notes: 1554-8937
Shen, David L
Liu, Ta-Wei
Zandberg, Wesley
Clark, Tom
Eskandari, Razieh
Alteen, Matthew G
Tan, Hong Yee
Zhu, Yanping
Cecioni, Samy
Vocadlo, David
Orcid: 0000-0001-6897-5558
MOP-102756/CIHR/Canada
Journal Article
United States
ACS Chem Biol. 2017 Jan 20;12(1):206-213. doi: 10.1021/acschembio.6b00876. Epub 2016 Dec 9.
Author Address: Department of Chemistry, Simon Fraser University , Burnaby, British Columbia V5A 1S6, Canada.
Department of Molecular Biology and Biochemistry, Simon Fraser University , Burnaby, British Columbia V5A 1S6, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2008
Author: Shen, H., Liu, D., Wu, K., Lei, J. and Yan, N.
Year: 2019
Title: Structures of human Na(v)1.7 channel in complex with auxiliary subunits and animal toxins
Journal: Science
Volume: 363
Issue: 6433
Pages: 1303-1308
Epub Date: 2019/02/16
Date: Mar 22
Short Title: Structures of human Na(v)1.7 channel in complex with auxiliary subunits and animal toxins
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.aaw2493
Accession Number: 30765606
Keywords: Amino Acid Sequence
Animals
Binding Sites
Cryoelectron Microscopy
HEK293 Cells
Humans
NAV1.7 Voltage-Gated Sodium Channel/*chemistry
Peptides/*chemistry
Protein Conformation
Saxitoxin/*chemistry
Spider Venoms/*chemistry
Tetrodotoxin/*chemistry
Voltage-Gated Sodium Channel Blockers/*chemistry
Voltage-Gated Sodium Channel beta-1 Subunit/*chemistry
Voltage-Gated Sodium Channel beta-2 Subunit/*chemistry
Abstract: Voltage-gated sodium channel Na(v)1.7 represents a promising target for pain relief. Here we report the cryo-electron microscopy structures of the human Na(v)1.7-β1-β2 complex bound to two combinations of pore blockers and gating modifier toxins (GMTs), tetrodotoxin with protoxin-II and saxitoxin with huwentoxin-IV, both determined at overall resolutions of 3.2 angstroms. The two structures are nearly identical except for minor shifts of voltage-sensing domain II (VSD(II)), whose S3-S4 linker accommodates the two GMTs in a similar manner. One additional protoxin-II sits on top of the S3-S4 linker in VSD(IV) The structures may represent an inactivated state with all four VSDs "up" and the intracellular gate closed. The structures illuminate the path toward mechanistic understanding of the function and disease of Na(v)1.7 and establish the foundation for structure-aided development of analgesics.
Notes: 1095-9203
Shen, Huaizong
Orcid: 0000-0002-1216-5393
Liu, Dongliang
Orcid: 0000-0001-9618-7671
Wu, Kun
Orcid: 0000-0002-3075-8272
Lei, Jianlin
Orcid: 0000-0002-9384-8742
Yan, Nieng
Orcid: 0000-0003-4829-7416
Journal Article
United States
Science. 2019 Mar 22;363(6433):1303-1308. doi: 10.1126/science.aaw2493. Epub 2019 Feb 14.
Author Address: State Key Laboratory of Membrane Biology, Tsinghua University, Beijing 100084, China.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua University, Beijing 100084, China.
Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China.
Medical Research Center, Beijing Key Laboratory of Cardiopulmonary Cerebral Resuscitation, Beijing Chao-Yang Hospital, Capital Medical University, Beijing 100020, China.
Technology Center for Protein Sciences, Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China.
State Key Laboratory of Membrane Biology, Tsinghua University, Beijing 100084, China. nyan@princeton.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1815
Author: Shen, P. S., Yang, X., DeCaen, P. G., Liu, X., Bulkley, D., Clapham, D. E. and Cao, E.
Year: 2016
Title: The Structure of the Polycystic Kidney Disease Channel PKD2 in Lipid Nanodiscs
Journal: Cell
Volume: 167
Issue: 3
Pages: 763-773.e11
Epub Date: 2016/10/22
Date: Oct 20
Short Title: The Structure of the Polycystic Kidney Disease Channel PKD2 in Lipid Nanodiscs
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2016.09.048
PMCID: PMC6055481
NIHMSID: NIHMS975692
Accession Number: 27768895
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cricetulus
Cryoelectron Microscopy
HEK293 Cells
Humans
Lipid Bilayers/chemistry
Mutation, Missense
Nanostructures/chemistry
Polycystic Kidney, Autosomal Dominant/genetics/*metabolism
Protein Conformation, alpha-Helical
Protein Domains
TRPP Cation Channels/*chemistry/genetics
Adpkd
Pkd2
TRP channel
Trpp2
cryo-EM
ion channel
polycystic kidney disease
polycystin
single particle electron cryo-microscopy
Abstract: The Polycystic Kidney Disease 2 (Pkd2) gene is mutated in autosomal dominant polycystic kidney disease (ADPKD), one of the most common human monogenic disorders. Here, we present the cryo-EM structure of PKD2 in lipid bilayers at 3.0 Å resolution, which establishes PKD2 as a homotetrameric ion channel and provides insight into potential mechanisms for its activation. The PKD2 voltage-sensor domain retains two of four gating charges commonly found in those of voltage-gated ion channels. The PKD2 ion permeation pathway is constricted at the selectivity filter and near the cytoplasmic end of S6, suggesting that two gates regulate ion conduction. The extracellular domain of PKD2, a hotspot for ADPKD pathogenic mutations, contributes to channel assembly and strategically interacts with the transmembrane core, likely serving as a physical substrate for extracellular stimuli to allosterically gate the channel. Finally, our structure establishes the molecular basis for the majority of pathogenic mutations in Pkd2-related ADPKD.
Notes: 1097-4172
Shen, Peter S
Yang, Xiaoyong
DeCaen, Paul G
Liu, Xiaowen
Bulkley, David
Clapham, David E
Cao, Erhu
P30 DK090868/DK/NIDDK NIH HHS/United States
R01 DK110575/DK/NIDDK NIH HHS/United States
T32 HL007572/HL/NHLBI NIH HHS/United States
R01 DK105175/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Cell. 2016 Oct 20;167(3):763-773.e11. doi: 10.1016/j.cell.2016.09.048.
Author Address: Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84112-5650, USA.
Department of Pharmacology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA; Howard Hughes Medical Institute, Boston, 02115 MA, USA; Cardiology, Boston Children's Hospital, Boston, 02115 MA, USA; Neurobiology, Harvard Medical School, Boston, MA 02115, USA.
Howard Hughes Medical Institute, Boston, 02115 MA, USA; Cardiology, Boston Children's Hospital, Boston, 02115 MA, USA; Neurobiology, Harvard Medical School, Boston, MA 02115, USA.
Keck Advanced Microscopy Laboratory and Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94143, USA.
Howard Hughes Medical Institute, Boston, 02115 MA, USA; Cardiology, Boston Children's Hospital, Boston, 02115 MA, USA; Neurobiology, Harvard Medical School, Boston, MA 02115, USA. Electronic address: dclapham@enders.tch.harvard.edu.
Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84112-5650, USA. Electronic address: erhu.cao@biochem.utah.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1737
Author: Shi, J., Gu, J. H., Dai, C. L., Gu, J., Jin, X., Sun, J., Iqbal, K., Liu, F. and Gong, C. X.
Year: 2015
Title: O-GlcNAcylation regulates ischemia-induced neuronal apoptosis through AKT signaling
Journal: Sci Rep
Volume: 5
Pages: 14500
Epub Date: 2015/09/29
Date: Sep 28
Short Title: O-GlcNAcylation regulates ischemia-induced neuronal apoptosis through AKT signaling
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/srep14500
PMCID: PMC4585968
Accession Number: 26412745
Keywords: Animals
*Apoptosis/genetics
Brain Ischemia/genetics/*metabolism
Disease Models, Animal
Gene Expression
Glycosylation
Mice
Neurons/*metabolism
Phosphorylation
Proto-Oncogene Proteins c-akt/genetics/*metabolism
*Signal Transduction
bcl-Associated Death Protein/metabolism
Abstract: Apoptosis plays an important role in neural development and neurological disorders. In this study, we found that O-GlcNAcylation, a unique protein posttranslational modification with O-linked β-N-acetylglucosamine (GlcNAc), promoted apoptosis through attenuating phosphorylation/activation of AKT and Bad. By using co-immunoprecipitation and mutagenesis techniques, we identified O-GlcNAc modification at both Thr308 and Ser473 of AKT. O-GlcNAcylation-induced apoptosis was attenuated by over-expression of AKT. We also found a dynamic elevation of protein O-GlcNAcylation during the first four hours of cerebral ischemia, followed by continuous decline after middle cerebral artery occlusion (MCAO) in the mouse brain. The elevation of O-GlcNAcylation coincided with activation of cell apoptosis. Finally, we found a negative correlation between AKT phosphorylation and O-GlcNAcylation in ischemic brain tissue. These results indicate that cerebral ischemia induces a rapid increase of O-GlcNAcylation that promotes apoptosis through down-regulation of AKT activity. These findings provide a novel mechanism through which O-GlcNAcylation regulates ischemia-induced neuronal apoptosis through AKT signaling.
Notes: 2045-2322
Shi, Jianhua
Gu, Jin-hua
Dai, Chun-ling
Gu, Jianlan
Jin, Xiaoxia
Sun, Jianming
Iqbal, Khalid
Liu, Fei
Gong, Cheng-Xin
R01 AG027429/AG/NIA NIH HHS/United States
R03 TW008123/TW/FIC NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Sci Rep. 2015 Sep 28;5:14500. doi: 10.1038/srep14500.
Author Address: Jiangsu Key Laboratory of Neuroregeneration, Co-Innovation Center of Neuroregeneration, Nantong University, Nantong, Jiangsu 226001, China.
Department of Biochemistry, Nantong University Medical School, Nantong, Jiangsu 226001, China.
Department of Neurochemistry, Inge Grundke-Iqbal Research Floor, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York 10314, United States of America.
Department of Pathophysiology, Nantong University Medical School, Nantong, Jiangsu 226001, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1767
Author: Shi, J., Sharif, S., Ruijtenbeek, R. and Pieters, R. J.
Year: 2016
Title: Activity Based High-Throughput Screening for Novel O-GlcNAc Transferase Substrates Using a Dynamic Peptide Microarray
Journal: PLoS One
Volume: 11
Issue: 3
Pages: e0151085
Epub Date: 2016/03/10
Short Title: Activity Based High-Throughput Screening for Novel O-GlcNAc Transferase Substrates Using a Dynamic Peptide Microarray
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0151085
PMCID: PMC4784888 commercial company PamGene International BV. There are no patents, products in development or marketed products to declare. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials.
Accession Number: 26960196
Keywords: Humans
N-Acetylglucosaminyltransferases/*metabolism
Peptides/*metabolism
*Protein Array Analysis
Retinoblastoma-Like Protein p130/metabolism
Substrate Specificity
Abstract: O-GlcNAcylation is a reversible and dynamic protein post-translational modification in mammalian cells. The O-GlcNAc cycle is catalyzed by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). O-GlcNAcylation plays important role in many vital cellular events including transcription, cell cycle regulation, stress response and protein degradation, and altered O-GlcNAcylation has long been implicated in cancer, diabetes and neurodegenerative diseases. Recently, numerous approaches have been developed to identify OGT substrates and study their function, but there is still a strong demand for highly efficient techniques. Here we demonstrated the utility of the peptide microarray approach to discover novel OGT substrates and study its specificity. Interestingly, the protein RBL-2, which is a key regulator of entry into cell division and may function as a tumor suppressor, was identified as a substrate for three isoforms of OGT. Using peptide Ala scanning, we found Ser 420 is one possible O-GlcNAc site in RBL-2. Moreover, substitution of Ser 420, on its own, inhibited OGT activity, raising the possibility of mechanism-based development for selective OGT inhibitors. This approach will prove useful for both discovery of novel OGT substrates and studying OGT specificity.
Notes: 1932-6203
Shi, Jie
Sharif, Suhela
Ruijtenbeek, Rob
Pieters, Roland J
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2016 Mar 9;11(3):e0151085. doi: 10.1371/journal.pone.0151085. eCollection 2016.
Author Address: Department of Medicinal Chemistry and Chemical Biology, Utrecht University, Utrecht, The Netherlands.
PamGene International BV, 's-Hertogenbosch, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1858
Author: Shi, J., Tomašič, T., Sharif, S., Brouwer, A. J., Anderluh, M., Ruijtenbeek, R. and Pieters, R. J.
Year: 2017
Title: Peptide microarray analysis of the cross-talk between O-GlcNAcylation and tyrosine phosphorylation
Journal: FEBS Lett
Volume: 591
Issue: 13
Pages: 1872-1883
Epub Date: 2017/06/06
Date: Jul
Short Title: Peptide microarray analysis of the cross-talk between O-GlcNAcylation and tyrosine phosphorylation
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1002/1873-3468.12708
Accession Number: 28580691
Keywords: Acetylglucosamine/*metabolism
HeLa Cells
Humans
Janus Kinase 2/metabolism
Molecular Dynamics Simulation
Peptide Fragments/chemistry/metabolism
Phosphorylation
*Protein Array Analysis
Protein Conformation
Signal Transduction
Tyrosine/*metabolism
Zonula Occludens Proteins/chemistry
*Jak2
*O-GlcNAcylation
*Zo-3
*cross-talk
*peptide microarray
*tyrosine phosphorylation
Abstract: O-GlcNAcylation of proteins regulates important cellular processes. A few reports noted that O-GlcNAcylation exhibits cross-talk with tyrosine phosphorylation. With an activity-based microarray analysis of 256 tyrosine kinase peptide substrates, we found that phosphorylation of six peptides by Jak2 inhibits their subsequent O-GlcNAcylation. However, O-GlcNAcylation has no detectable effect on their subsequent phosphorylation. A specific peptide (ZO3_357_371), derived from the ZO-3 protein, was studied in detail. Kinetic results show that the presence of a phosphate at Tyr364 of ZO3_357_371 slows the O-GlcNAcylation of nearby Ser369, while the presence of a GlcNAc at Ser369 has no significant effect on the phosphorylation of this peptide at Tyr364. These findings provide a glimpse into the new paradigm for cellular signaling control by cross-talk.
Notes: 1873-3468
Shi, Jie
Tomašič, Tihomir
Sharif, Suhela
Brouwer, Arwin J
Anderluh, Marko
Ruijtenbeek, Rob
Pieters, Roland J
Orcid: 0000-0003-4723-3584
Letter
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2017 Jul;591(13):1872-1883. doi: 10.1002/1873-3468.12708. Epub 2017 Jun 27.
Author Address: Department of Chemical Biology and Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands.
Faculty of Pharmacy, University of Ljubljana, Slovenia.
PamGene International BV, 's-Hertogenbosch, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1492
Author: Shi, J., Wu, S., Dai, C. L., Li, Y., Grundke-Iqbal, I., Iqbal, K., Liu, F. and Gong, C. X.
Year: 2012
Title: Diverse regulation of AKT and GSK-3β by O-GlcNAcylation in various types of cells
Journal: FEBS Lett
Volume: 586
Issue: 16
Pages: 2443-50
Epub Date: 2012/06/13
Date: Jul 30
Short Title: Diverse regulation of AKT and GSK-3β by O-GlcNAcylation in various types of cells
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2012.05.063
PMCID: PMC3407308
NIHMSID: NIHMS384357
Accession Number: 22687243
Keywords: Acetylglucosamine/*chemistry
Animals
*Gene Expression Regulation, Enzymologic
Glycogen Synthase Kinase 3/*metabolism
Glycogen Synthase Kinase 3 beta
HEK293 Cells
Hep G2 Cells
Hippocampus/metabolism
Humans
Insulin/metabolism
Mice
Neurons/metabolism
Phosphorylation
Protein Processing, Post-Translational
Proto-Oncogene Proteins c-akt/*metabolism
RNA Interference
Tissue Distribution
Abstract: Protein kinase B (AKT) and glycogen synthase kinase-3β (GSK-3β) are major components of insulin-AKT signaling that plays crucial roles in various types of tissue. Recent studies found that these two kinases are modified posttranslationally by O-GlcNAcylation. Here, we demonstrate that O-GlcNAcylation regulated phosphorylation/activation of AKT and GSK-3β in different manners in kidney HEK-293FT cells, but did not affect these two kinases in hepatic HepG2 cells. In neuronal cells, O-GlcNAcylation regulated phosphorylation of AKT negatively, but had no effect on GSK-3β. These results suggest protein-specific and cell type-specific regulation of AKT and GSK-3β by O-GlcNAcylation. Therefore, studies on the roles of AKT and GSK-3β O-GlcNAcylation should be done in a tissue- and cell type-specific manner.
Notes: 1873-3468
Shi, Jianhua
Wu, Shiliang
Dai, Chun-ling
Li, Yi
Grundke-Iqbal, Inge
Iqbal, Khalid
Liu, Fei
Gong, Cheng-Xin
R01 AG031969/AG/NIA NIH HHS/United States
R01 AG027429/AG/NIA NIH HHS/United States
R21 AG031969/AG/NIA NIH HHS/United States
R01 AG019158/AG/NIA NIH HHS/United States
R03 TW008123/TW/FIC NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
FEBS Lett. 2012 Jul 30;586(16):2443-50. doi: 10.1016/j.febslet.2012.05.063. Epub 2012 Jun 8.
Author Address: Department of Biochemistry, Soochow University Medical School, Soochow, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1427
Author: Shi, Q., Padmanabhan, R., Villegas, C. J., Gu, S. and Jiang, J. X.
Year: 2011
Title: Membrane topological structure of neutral system N/A amino acid transporter 4 (SNAT4) protein
Journal: J Biol Chem
Volume: 286
Issue: 44
Pages: 38086-38094
Epub Date: 2011/09/16
Date: Nov 4
Short Title: Membrane topological structure of neutral system N/A amino acid transporter 4 (SNAT4) protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.220277
PMCID: PMC3207422
Accession Number: 21917917
Keywords: Amino Acid Transport System A/*chemistry/metabolism
Animals
Asparagine/chemistry
CHO Cells
Cell Line
Cricetinae
Cricetulus
Glycosylation
Humans
Membrane Proteins/chemistry
Membrane Transport Proteins/*chemistry
Mice
Mutagenesis, Site-Directed
Open Reading Frames
Protein Conformation
Protein Structure, Tertiary
Abstract: Members of system N/A amino acid transporter (SNAT) family mediate transport of neutral amino acids, including l-alanine, l-glutamine, and l-histidine, across the plasma membrane and are involved in a variety of cellular functions. By using chemical labeling, glycosylation, immunofluorescence combined with molecular modeling approaches, we resolved the membrane topological structure of SNAT4, a transporter expressed predominantly in liver. To analyze the orientation using the chemical labeling and biotinylation approach, the "Cys-null" mutant of SNAT4 was first generated by mutating all five endogenous cysteine residues. Based on predicted topological structures, a single cysteine residue was introduced individually into all possible nontransmembrane domains of the Cys-null mutant. The cells expressing these mutants were labeled with N-biotinylaminoethyl methanethiosulfonate, a membrane-impermeable cysteine-directed reagent. We mapped the orientations of N- and C-terminal domains. There are three extracellular loop domains, and among them, the second loop domain is the largest that spans from amino acid residue ∼242 to ∼335. The orientation of this domain was further confirmed by the identification of two N-glycosylated residues, Asn-260 and Asn-264. Together, we showed that SNAT4 contains 10 transmembrane domains with extracellular N and C termini and a large N-glycosylated, extracellular loop domain. This is the first report concerning membrane topological structure of mammalian SNAT transporters, which will provide important implications for our understanding of structure-function of the members in this amino acid transporter family.
Notes: 1083-351x
Shi, Qian
Padmanabhan, Rugmani
Villegas, Carla J
Gu, Sumin
Jiang, Jean X
R01 EY012085/EY/NEI NIH HHS/United States
R29 EY012085/EY/NEI NIH HHS/United States
EY12085/EY/NEI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 Nov 4;286(44):38086-38094. doi: 10.1074/jbc.M111.220277. Epub 2011 Sep 14.
Author Address: Department of Biochemistry, University of Texas Health Science Center, San Antonio, Texas 78229-3900.
Department of Biochemistry, University of Texas Health Science Center, San Antonio, Texas 78229-3900. Electronic address: jiangj@uthscsa.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1326
Author: Shi, Y., Tomic, J., Wen, F., Shaha, S., Bahlo, A., Harrison, R., Dennis, J. W., Williams, R., Gross, B. J., Walker, S., Zuccolo, J., Deans, J. P., Hart, G. W. and Spaner, D. E.
Year: 2010
Title: Aberrant O-GlcNAcylation characterizes chronic lymphocytic leukemia
Journal: Leukemia
Volume: 24
Issue: 9
Pages: 1588-98
Epub Date: 2010/07/30
Date: Sep
Short Title: Aberrant O-GlcNAcylation characterizes chronic lymphocytic leukemia
Alternate Journal: Leukemia
ISSN: 0887-6924 (Print)
0887-6924
DOI: 10.1038/leu.2010.152
PMCID: PMC4361888
NIHMSID: NIHMS656046
Accession Number: 20668475
Keywords: Acetylglucosamine/*metabolism
Acylation
Adult
Aged
Aged, 80 and over
Base Sequence
Cytokines/metabolism
DNA Primers
Female
Humans
Immunophenotyping
Leukemia, Lymphocytic, Chronic, B-Cell/*metabolism/pathology
Male
Middle Aged
Polymerase Chain Reaction
Toll-Like Receptors/metabolism
Tumor Cells, Cultured
Abstract: O-linked N-Acetylglucosamine (O-GlcNAc) post-translational modifications originate from the activity of the hexosamine pathway, and are known to affect intracellular signaling processes. As aberrant responses to microenvironmental signals are a feature of chronic lymphocytic leukemia (CLL), O-GlcNAcylated protein levels were measured in primary CLL cells. In contrast to normal circulating and tonsillar B cells, CLL cells expressed high levels of O-GlcNAcylated proteins, including p53, c-myc and Akt. O-GlcNAcylation in CLL cells increased following activation with cytokines and through toll-like receptors (TLRs), or after loading with hexosamine pathway substrates. However, high baseline O-GlcNAc levels were associated with impaired signaling responses to TLR agonists, chemotherapeutic agents, B cell receptor crosslinking and mitogens. Indolent and aggressive clinical behavior of CLL cells were found to correlate with higher and lower O-GlcNAc levels, respectively. These findings suggest that intracellular O-GlcNAcylation is associated with the pathogenesis of CLL, which could potentially have therapeutic implications.
Notes: 1476-5551
Shi, Y
Tomic, J
Wen, F
Shaha, S
Bahlo, A
Harrison, R
Dennis, J W
Williams, R
Gross, B J
Walker, S
Zuccolo, J
Deans, J P
Hart, G W
Spaner, D E
190633/Canadian Institutes of Health Research/Canada
R01CA42486/CA/NCI NIH HHS/United States
R03 MH076518/MH/NIMH NIH HHS/United States
R01 DK61671/DK/NIDDK NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
MOP-79405/Canadian Institutes of Health Research/Canada
MOP-43938/Canadian Institutes of Health Research/Canada
R01 DK061671/DK/NIDDK NIH HHS/United States
15095/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Leukemia. 2010 Sep;24(9):1588-98. doi: 10.1038/leu.2010.152. Epub 2010 Jul 29.
Author Address: Division of Molecular and Cellular Biology, Research Institute, Sunnybrook Health Sciences Center, Toronto, Ontario, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 395
Author: Shibata, S., Takeda, T. and Natori, Y.
Year: 1988
Title: The structure of nephritogenoside. A nephritogenic glycopeptide with alpha-N-glycosidic linkage
Journal: J Biol Chem
Volume: 263
Issue: 25
Pages: 12483-5
Epub Date: 1988/09/05
Date: Sep 5
Short Title: The structure of nephritogenoside. A nephritogenic glycopeptide with alpha-N-glycosidic linkage
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3410849
Keywords: Amino Acid Sequence
Amino Acids/analysis
Animals
Basement Membrane/analysis
Carbohydrate Conformation
Chromatography, Gel
Collagen
*Glycoproteins/analysis
Kidney Glomerulus/*analysis
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Rats
Receptors, Concanavalin A
Repetitive Sequences, Nucleic Acid
Abstract: Nephritogenoside is a glycopeptide with the ability to induce chronic progressive glomerulonephritis (end stage kidney) in homologous animals, by a single footpad injection. This substance contains a novel carbohydrate-peptide linkage, i.e. the trisaccharide (3 glucose residues) chain is alpha-N-glycosidically linked to the polypeptide chain through the amido nitrogen of an asparagine residue at the N-terminal. It was found that the peptide portion of nephritogenoside is composed of twenty-one amino acids (Asn1-Pro-Leu-Phe-Gly5-Ile-Ala-Gly-Glu-Asp10-Gly-Pro-Thr-Gly-Pr o15-Ser-Gly-Ile- Val-Gly20-Gln21) and that the peptide portion has a repeated Gly-X-Y structure.
Notes: Shibata, S
Takeda, T
Natori, Y
Comparative Study
Journal Article
United States
J Biol Chem. 1988 Sep 5;263(25):12483-5.
Author Address: Clinical Research Institute, National Medical Center, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 930
Author: Shieh, J. J., Pan, C. J., Mansfield, B. C. and Chou, J. Y.
Year: 2004
Title: The islet-specific glucose-6-phosphatase-related protein, implicated in diabetes, is a glycoprotein embedded in the endoplasmic reticulum membrane
Journal: FEBS Lett
Volume: 562
Issue: 1-3
Pages: 160-4
Epub Date: 2004/03/27
Date: Mar 26
Short Title: The islet-specific glucose-6-phosphatase-related protein, implicated in diabetes, is a glycoprotein embedded in the endoplasmic reticulum membrane
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/s0014-5793(04)00223-6
Accession Number: 15044018
Keywords: Acetylcysteine/*analogs & derivatives/metabolism
Amino Acid Sequence
Animals
COS Cells
Calreticulin/metabolism
Cysteine Endopeptidases/metabolism
Diabetes Mellitus/*metabolism
Endoplasmic Reticulum/chemistry/*metabolism
Enzyme Inhibitors/metabolism
*Glucose-6-Phosphatase
Glycoproteins/chemistry/genetics/*metabolism
Humans
Membrane Proteins/chemistry/genetics/*metabolism
Mice
Multienzyme Complexes/antagonists & inhibitors/metabolism
Proteasome Endopeptidase Complex
Protein Structure, Secondary
Proteins/chemistry/genetics/*metabolism
Recombinant Fusion Proteins/genetics/metabolism
Abstract: The islet-specific glucose-6-phosphatase-related protein (IGRP) has no known catalytic activity, but is of interest because it is the source of the peptide autoantigen targeted by a prevalent population of pathogenic CD8(+) T cells in non-obese diabetic mice. To better understand the potential roles of this protein in diabetes mellitus, we examine the subcellular localization and membrane topography of human IGRP. We show that IGRP is a glycoprotein, held in the endoplasmic reticulum by nine transmembrane domains, which is degraded in cells predominantly through the proteasome pathway that generates the major histocompatibility complex class I-presented peptides.
Notes: Shieh, Jeng-Jer
Pan, Chi-Jiunn
Mansfield, Brian C
Chou, Janice Yang
Journal Article
England
FEBS Lett. 2004 Mar 26;562(1-3):160-4. doi: 10.1016/S0014-5793(04)00223-6.
Author Address: Section on Cellular Differentiation, Heritable Disorders Branch, National Institute of Child Health and Human Development, Building 10, Room 9S241, National Institutes of Health, Bethesda, MD 20892-1830, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 599
Author: Shikata, Y., Hayashi, Y., Yoshimatsu, K., Ohya, Y., Seto, T., Fukushima, K. and Yoshida, Y.
Year: 1993
Title: Pro-major basic protein has three types of sugar chains at the pro-portion
Journal: Biochim Biophys Acta
Volume: 1163
Issue: 3
Pages: 243-9
Epub Date: 1993/06/04
Date: Jun 4
Short Title: Pro-major basic protein has three types of sugar chains at the pro-portion
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/0167-4838(93)90158-n
Accession Number: 8507662
Keywords: Amino Acid Sequence
Animals
Asparagine/chemistry
Blood Proteins/*chemistry/metabolism
Cricetinae
Eosinophil Granule Proteins
Glycopeptides/chemistry
Glycoproteins/*chemistry/metabolism
Glycosylation
Molecular Sequence Data
Monosaccharides/*analysis
Peptide Mapping
Protein Precursors/*chemistry/metabolism
Protein Processing, Post-Translational
Protein Sorting Signals/chemistry/metabolism
*Ribonucleases
Sequence Analysis
Serine/chemistry
Serine Endopeptidases/metabolism
Threonine/chemistry
Abstract: The amino-acid sequence of purified recombinant pro-major basic protein from Chinese hamster kidney cells was determined to verify the primary structure and glycosylation sites. Reduced and S-carboxamidemethylated protein was first digested with Achromobacter proteinase I. Each peptide was characterized by amino-acid analysis and amino-acid sequence analysis. We could identify all the peptides which were expected from the pro-major basic protein cDNA sequence. Sequence analysis and deglycosylation study revealed that Ser-8, Thr-9, Ser-46 and Asn-70 were glycosylated. The results indicated that proMBP has three types of sugar chains, O-glycoside, N-glycoside and glycosaminoglycan, in the pro-portion.
Notes: Shikata, Y
Hayashi, Y
Yoshimatsu, K
Ohya, Y
Seto, T
Fukushima, K
Yoshida, Y
Journal Article
Netherlands
Biochim Biophys Acta. 1993 Jun 4;1163(3):243-9. doi: 10.1016/0167-4838(93)90158-n.
Author Address: Tsukuba Research Laboratories, Eisai Company Ltd., Ibaraki, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1318
Author: Shim, A. H., Liu, H., Focia, P. J., Chen, X., Lin, P. C. and He, X.
Year: 2010
Title: Structures of a platelet-derived growth factor/propeptide complex and a platelet-derived growth factor/receptor complex
Journal: Proc Natl Acad Sci U S A
Volume: 107
Issue: 25
Pages: 11307-12
Epub Date: 2010/06/11
Date: Jun 22
Short Title: Structures of a platelet-derived growth factor/propeptide complex and a platelet-derived growth factor/receptor complex
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1000806107
PMCID: PMC2895058
Accession Number: 20534510
Keywords: Animals
Crystallography, X-Ray/methods
Humans
Hydrophobic and Hydrophilic Interactions
Molecular Conformation
Peptides/chemistry
Platelet-Derived Growth Factor/*chemistry/metabolism
Protein Binding
Protein Conformation
Receptor Protein-Tyrosine Kinases/metabolism
Receptor, Platelet-Derived Growth Factor beta/chemistry
Recombinant Proteins/chemistry
Signal Transduction
Thermodynamics
Abstract: Platelet-derived growth factors (PDGFs) and their receptors (PDGFRs) are prototypic growth factors and receptor tyrosine kinases which have critical functions in development. We show that PDGFs share a conserved region in their prodomain sequences which can remain noncovalently associated with the mature cystine-knot growth factor domain after processing. The structure of the PDGF-A/propeptide complex reveals this conserved, hydrophobic association mode. We also present the structure of the complex between PDGF-B and the first three Ig domains of PDGFRbeta, showing that two PDGF-B protomers clamp PDGFRbeta at their dimerization seam. The PDGF-B:PDGFRbeta interface is predominantly hydrophobic, and PDGFRs and the PDGF propeptides occupy overlapping positions on mature PDGFs, rationalizing the need of propeptides by PDGFs to cover functionally important hydrophobic surfaces during secretion. A large-scale structural organization and rearrangement is observed for PDGF-B upon receptor binding, in which the PDGF-B L1 loop, disordered in the structure of the free form, adopts a highly specific conformation to form hydrophobic interactions with the third Ig domain of PDGFRbeta. Calorimetric data also shows that the membrane-proximal homotypic PDGFRalpha interaction, albeit required for activation, contributes negatively to ligand binding. The structural and biochemical data together offer insights into PDGF-PDGFR signaling, as well as strategies for PDGF-antagonism.
Notes: 1091-6490
Shim, Ann Hye-Ryong
Liu, Heli
Focia, Pamela J
Chen, Xiaoyan
Lin, P Charles
He, Xiaolin
R01 GM078055/GM/NIGMS NIH HHS/United States
GM047477/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2010 Jun 22;107(25):11307-12. doi: 10.1073/pnas.1000806107. Epub 2010 Jun 2.
Author Address: Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine, Searle 8-417, 303 East Chicago Avenue, Chicago, IL 60611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 861
Author: Shimaoka, M., Xiao, T., Liu, J. H., Yang, Y., Dong, Y., Jun, C. D., McCormack, A., Zhang, R., Joachimiak, A., Takagi, J., Wang, J. H. and Springer, T. A.
Year: 2003
Title: Structures of the alpha L I domain and its complex with ICAM-1 reveal a shape-shifting pathway for integrin regulation
Journal: Cell
Volume: 112
Issue: 1
Pages: 99-111
Epub Date: 2003/01/16
Date: Jan 10
Short Title: Structures of the alpha L I domain and its complex with ICAM-1 reveal a shape-shifting pathway for integrin regulation
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/s0092-8674(02)01257-6
PMCID: PMC4372089
NIHMSID: NIHMS671056
Accession Number: 12526797
Keywords: Amino Acid Substitution
Animals
Cells, Cultured
Crystallography, X-Ray
Disulfides/chemistry
Escherichia coli/genetics
Glutamine/chemistry/metabolism
Integrins/*metabolism
Intercellular Adhesion Molecule-1/chemistry/isolation & purification/*metabolism
Ligands
Lymphocyte Function-Associated Antigen-1/*chemistry/genetics/isolation &
purification/*metabolism
Magnesium/chemistry/metabolism
Models, Molecular
Molecular Conformation
Mutation
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Spectrum Analysis, Raman
Surface Plasmon Resonance
Abstract: The structure of the I domain of integrin alpha L beta 2 bound to the Ig superfamily ligand ICAM-1 reveals the open ligand binding conformation and the first example of an integrin-IgSF interface. The I domain Mg2+ directly coordinates Glu-34 of ICAM-1, and a dramatic swing of I domain residue Glu-241 enables a critical salt bridge. Liganded and unliganded structures for both high- and intermediate-affinity mutant I domains reveal that ligand binding can induce conformational change in the alpha L I domain and that allosteric signals can convert the closed conformation to intermediate or open conformations without ligand binding. Pulling down on the C-terminal alpha 7 helix with introduced disulfide bonds ratchets the beta 6-alpha 7 loop into three different positions in the closed, intermediate, and open conformations, with a progressive increase in affinity.
Notes: 1097-4172
Shimaoka, Motomu
Xiao, Tsan
Liu, Jin-Huan
Yang, Yuting
Dong, Yicheng
Jun, Chang-Duk
McCormack, Alison
Zhang, Rongguang
Joachimiak, Andrzej
Takagi, Junichi
Wang, Jia-Huai
Springer, Timothy A
HL48675/HL/NHLBI NIH HHS/United States
CA31798/CA/NCI NIH HHS/United States
P01 HL048675/HL/NHLBI NIH HHS/United States
R01 CA031798/CA/NCI NIH HHS/United States
R37 CA031798/CA/NCI NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Cell. 2003 Jan 10;112(1):99-111. doi: 10.1016/s0092-8674(02)01257-6.
Author Address: The Center for Blood Research, Department of Pathology, Department of Anesthesia, Department of Pediatrics, Boston, Massachusetts 02115.
Dana-Farber Cancer Institute, Department of Pediatrics, Department of Medicine, Department of Biological Chemistry, Department of Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115.
Biosciences Division, Argonne National Laboratory, Argonne, Illinois 60439.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 250
Author: Shimizu, N., Hara, H., Sogabe, T., Sakai, H., Ihara, I., Inoue, H., Nakamura, T. and Shimizu, S.
Year: 1992
Title: Hepatocyte growth factor is linked by O-glycosylated oligosaccharide on the alpha chain
Journal: Biochem Biophys Res Commun
Volume: 189
Issue: 3
Pages: 1329-35
Epub Date: 1992/12/30
Date: Dec 30
Short Title: Hepatocyte growth factor is linked by O-glycosylated oligosaccharide on the alpha chain
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(92)90219-b
Accession Number: 1482348
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Cysteine
Glycosylation
Hepatocyte Growth Factor/*chemistry
Humans
Lysine
Macromolecular Substances
Molecular Sequence Data
Monosaccharides/analysis
Oligosaccharides/*chemistry
Peptide Fragments/chemistry/isolation & purification
Recombinant Proteins/chemistry
Abstract: The glycosylation site and the structure of O-glycosylated oligosaccharide of recombinant human HGF were investigated. N-acetylgalactosamine (GalNAc) in the alpha chain suggested the presence of O-glycosylated oligosaccharide. Sugar analysis and amino acid sequence analysis of peptide fragments produced by limited degradation revealed that O-glycosylated oligosaccharide linked to Thr445 of the alpha chain. The molecular weight of the oligosaccharide was determined with ion spray mass spectrometry. From these studies, the structure of the O-glycosylated oligosaccharide on the alpha chain of HGF was concluded as [formula: see text].
Notes: Shimizu, N
Hara, H
Sogabe, T
Sakai, H
Ihara, I
Inoue, H
Nakamura, T
Shimizu, S
Journal Article
United States
Biochem Biophys Res Commun. 1992 Dec 30;189(3):1329-35. doi: 10.1016/0006-291x(92)90219-b.
Author Address: Pharmaceuticals Research Center, Toyobo Co. Ltd., Otsu, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1905
Author: Shin, H., Cha, H. J., Na, K., Lee, M. J., Cho, J. Y., Kim, C. Y., Kim, E. K., Kang, C. M., Kim, H. and Paik, Y. K.
Year: 2018
Title: O-GlcNAcylation of the Tumor Suppressor FOXO3 Triggers Aberrant Cancer Cell Growth
Journal: Cancer Res
Volume: 78
Issue: 5
Pages: 1214-1224
Epub Date: 2018/01/06
Date: Mar 1
Short Title: O-GlcNAcylation of the Tumor Suppressor FOXO3 Triggers Aberrant Cancer Cell Growth
Alternate Journal: Cancer research
ISSN: 0008-5472
DOI: 10.1158/0008-5472.can-17-3512
Accession Number: 29301793
Keywords: Apoptosis
Biomarkers, Tumor/genetics/*metabolism
Case-Control Studies
Cell Proliferation
Cyclin-Dependent Kinase Inhibitor p21/genetics/metabolism
Forkhead Box Protein O3/genetics/*metabolism
Glycosylation
Humans
Neoplasms/genetics/metabolism/*pathology
Pancreatic Neoplasms/genetics/metabolism/*pathology
Prognosis
Protein Processing, Post-Translational
Proto-Oncogene Proteins c-mdm2/genetics/metabolism
Serine/genetics/metabolism
Tumor Cells, Cultured
Tumor Suppressor Protein p53/genetics/metabolism
Abstract: Posttranslational modifications of tumor suppressors can induce abnormal cell growth. Here, we identify site-specific O-GlcNAcylation as a critical block of FOXO3 that may abrogate a part of the p53 pathway, resulting in aberrant cancer cell growth. Of seven O-GlcNAcylation sites identified within the FOXO3 transactivation domain, we found that changes in O-GlcNAcylation at Ser284 modulated p21-mediated cancer cell growth. Overexpression of either O-GlcNAcylated FOXO3 (FOX-OV) or a Ser-to-Ala mutant (S284A) in PANC-1 cells indicated that S284 O-GlcNAc acts as a critical block of the FOXO tumor suppressor and induces proliferation in PANC-1 cancer cells by stimulating the MDM2-p53-p21 axis. Furthermore, S284A mutant cells lacking S284 O-GlcNAc and FOX-OV cells exhibited opposing MDM2-p53-p21 axis expression patterns at both the mRNA and protein levels. Thus, our study provides evidence to support a role for S284 O-GlcNAc as a critical block of FOXO3 to induce subsequent cancer cell growth via abrogation of the p53 regulatory circuit.Significance: These findings highlight a posttranslational mechanism for indirect abrogation of the p53 pathway, one that may occur with some frequency in human cancer cells. Cancer Res; 78(5); 1214-24. ©2018 AACR.
Notes: 1538-7445
Shin, Heon
Cha, Hyun-Jeong
Na, Keun
Lee, Min Jung
Cho, Jin-Young
Kim, Chae-Yeon
Kim, Eun Kyung
Kang, Chang Moo
Kim, Hoguen
Paik, Young-Ki
Journal Article
Research Support, Non-U.S. Gov't
United States
Cancer Res. 2018 Mar 1;78(5):1214-1224. doi: 10.1158/0008-5472.CAN-17-3512. Epub 2018 Jan 4.
Author Address: Yonsei Proteome Research Center, Yonsei University, Seoul, Republic of Korea.
Graduate program in Integrated OMICS for Biomedical Science, Yonsei University, Seoul, Republic of Korea.
Department of Pathology, Yonsei University College of Medicine, Seoul, Republic of Korea.
Department of Surgery, Yonsei University College of Medicine, Seoul, Republic of Korea.
Yonsei Proteome Research Center, Yonsei University, Seoul, Republic of Korea. paikyk@yonsei.ac.kr.
Department of Biochemistry, Yonsei University, Seoul, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 477
Author: Shinoda, T., Takahashi, N., Takayasu, T., Okuyama, T. and Shimizu, A.
Year: 1981
Title: Complete amino acid sequence of the Fc region of a human delta chain
Journal: Proc Natl Acad Sci U S A
Volume: 78
Issue: 2
Pages: 785-9
Epub Date: 1981/02/01
Date: Feb
Short Title: Complete amino acid sequence of the Fc region of a human delta chain
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.78.2.785
PMCID: PMC319887
Accession Number: 6785754
Keywords: Amino Acid Sequence
Biological Evolution
Disulfides
Humans
Immunoglobulin D
*Immunoglobulin Fc Fragments
*Immunoglobulin Heavy Chains
*Immunoglobulin delta-Chains
Myeloma Proteins
Peptide Fragments
Protein Conformation
Abstract: The complete amino acid sequence of an Fc-like fragment designated Fc delta (t) and obtained by limited proteolysis with trypsin of an intact myeloma IgD protein (NIG-65) has been determined. The fragment contains 226 amino acid residues and has a molecular weight of 32,000 per monomeric unit. It has three glucosamine oligosaccharides at asparagine residues 68, 159, and 210. Of these, glucosamine-159 is characteristic of the delta chain and has no counterpart position in any of the other classes. On the other hand, glucosamine-68 is shared by gamma, mu, and epsilon, and glucosamine-210 is shared by alpha and mu. Although the Fc delta (t) has the common framework structure of immunoglobulins, its sequence has many individual characteristics when its two domains are compared separately with the counterpart domain of other heavy chains. Such comparison has shown that the two Fc domains of the delta chain should be placed in an independent branch in topology; for all the other classes, the Fc domains are paired well with their counterparts. The comparison has also shown that there are three prominent gaps by which each domain can be divided into two homologous halves. For each class of immunoglobulin, a moderate degree of internal homology exists between the first half and the second half of each domain of the Fc, suggesting that the primordial gene may have coded for a unit about the size of a half domain. Based on this observation together with sequence comparisons, a possible genetic mechanism is proposed for the origin and evolution of the genes for immunoglobulin domains.
Notes: 1091-6490
Shinoda, T
Takahashi, N
Takayasu, T
Okuyama, T
Shimizu, A
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 1981 Feb;78(2):785-9. doi: 10.1073/pnas.78.2.785.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1051
Author: Shintani, Y., Takashima, S., Asano, Y., Kato, H., Liao, Y., Yamazaki, S., Tsukamoto, O., Seguchi, O., Yamamoto, H., Fukushima, T., Sugahara, K., Kitakaze, M. and Hori, M.
Year: 2006
Title: Glycosaminoglycan modification of neuropilin-1 modulates VEGFR2 signaling
Journal: Embo j
Volume: 25
Issue: 13
Pages: 3045-55
Epub Date: 2006/06/10
Date: Jul 12
Short Title: Glycosaminoglycan modification of neuropilin-1 modulates VEGFR2 signaling
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7601188
PMCID: PMC1500974
Accession Number: 16763549
Keywords: Amino Acid Sequence
Cell Movement/physiology
Cell Survival/physiology
Cells, Cultured
Chondroitin Sulfates/metabolism
Endothelium, Vascular/cytology/metabolism
Gene Expression Regulation
Glycosaminoglycans/*metabolism
Heparitin Sulfate/metabolism
Humans
Molecular Sequence Data
Myocytes, Smooth Muscle/metabolism
Neuropilin-1/metabolism/*physiology
Protein Processing, Post-Translational
Sequence Homology, Amino Acid
Serine/metabolism
Signal Transduction
Vascular Endothelial Growth Factor A/metabolism
Vascular Endothelial Growth Factor Receptor-2/biosynthesis/*physiology
Abstract: Neuropilin-1 (NRP1) is a co-receptor for vascular endothelial growth factor (VEGF) that enhances the angiogenic signals cooperatively with VEGFR2. VEGF signaling is essential for physiological and pathological angiogenesis through its effects on vascular endothelial cells (ECs) and smooth muscle cells (SMCs), but the mechanisms coordinating this response are not well understood. Here we show that a substantial fraction of NRP1 is proteoglycan modified with either heparan sulfate or chondroitin sulfate on a single conserved Ser. The composition of the NRP1 glycosaminoglycan (GAG) chains differs between ECs and SMCs. Glycosylation increased VEGF binding in both cell types, but the differential GAG composition of NRP1 mediates opposite responsiveness to VEGF in ECs and SMCs. Finally, NRP1 expression and its GAG modification post-transcriptionally regulate VEGFR2 protein expression. These findings indicate that GAG modification of NRP1 plays a critical role in modulating VEGF signaling, and may provide new insights into physiological and pathological angiogenesis.
Notes: 1460-2075
Shintani, Yasunori
Takashima, Seiji
Asano, Yoshihiro
Kato, Hisakazu
Liao, Yulin
Yamazaki, Satoru
Tsukamoto, Osamu
Seguchi, Osamu
Yamamoto, Hiroyuki
Fukushima, Tomi
Sugahara, Kazuyuki
Kitakaze, Masafumi
Hori, Masatsugu
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2006 Jul 12;25(13):3045-55. doi: 10.1038/sj.emboj.7601188. Epub 2006 Jun 8.
Author Address: Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Suita, Osaka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1007
Author: Shiokawa, D., Shika, Y., Saito, K., Yamazaki, K. and Tanuma, S.
Year: 2005
Title: Physical and biochemical properties of mammalian DNase X proteins: non-AUG translation initiation of porcine and bovine mRNAs for DNase X
Journal: Biochem J
Volume: 392
Issue: Pt 3
Pages: 511-7
Epub Date: 2005/08/19
Date: Dec 15
Short Title: Physical and biochemical properties of mammalian DNase X proteins: non-AUG translation initiation of porcine and bovine mRNAs for DNase X
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20051114
PMCID: PMC1316290
Accession Number: 16107205
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cattle
Codon, Initiator/*genetics
Consensus Sequence
Deoxyribonucleases/biosynthesis/*chemistry/*genetics/metabolism
Gene Expression Profiling
Gene Expression Regulation, Enzymologic
HeLa Cells
Humans
Mice
Molecular Sequence Data
*Peptide Chain Initiation, Translational
Protein Biosynthesis
RNA, Messenger/genetics/*metabolism
Sequence Homology, Amino Acid
Swine/*genetics
Abstract: DNase X is the first human DNase protein identified as being homologous with DNase I. In the present study we describe the isolation of several mammalian DNase X cDNAs and the molecular characterization of their coding proteins. A sequence comparison reveals some conserved characteristics: all the mammalian DNase X proteins have an N-terminal signal peptide, a potential N-linked glycosylation site and a C-terminal hydrophobic domain. Human DNase X, ectopically expressed in HeLa S3 cells, is located in the ER (endoplasmic reticulum) and is modified by an N-linked glycosylation at Asn-243. Gene expression analyses show that the high expression level in muscular tissues, a known feature of human DNASE X, is also observed in mouse DNase X. Interestingly, the translation of porcine and bovine DNase X proteins occurs in the absence of an in-frame AUG initiation codon. We show that their mRNAs utilize a conserved CUG triplet for translation initiation.
Notes: 1470-8728
Shiokawa, Daisuke
Shika, Yukari
Saito, Kazuki
Yamazaki, Kosuke
Tanuma, Sei-ichi
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2005 Dec 15;392(Pt 3):511-7. doi: 10.1042/BJ20051114.
Author Address: Department of Biochemistry, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1595
Author: Shnitsar, V., Li, J., Li, X., Calmettes, C., Basu, A., Casey, J. R., Moraes, T. F. and Reithmeier, R. A. F.
Year: 2013
Title: A substrate access tunnel in the cytosolic domain is not an essential feature of the solute carrier 4 (SLC4) family of bicarbonate transporters
Journal: J Biol Chem
Volume: 288
Issue: 47
Pages: 33848-33860
Epub Date: 2013/10/15
Date: Nov 22
Short Title: A substrate access tunnel in the cytosolic domain is not an essential feature of the solute carrier 4 (SLC4) family of bicarbonate transporters
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.511865
PMCID: PMC3837127
Accession Number: 24121512
Keywords: Amino Acid Substitution
Anion Exchange Protein 1, Erythrocyte/*chemistry/genetics/*metabolism
Crystallography, X-Ray
Gene Expression Regulation
HEK293 Cells
Humans
Hydrogen-Ion Concentration
Ion Transport/physiology
Mutation, Missense
Protein Structure, Tertiary
Sodium-Bicarbonate Symporters/*chemistry/genetics/*metabolism
Anion Exchanger 1 (AE1)
Anion Transport
Band 3
Membrane Proteins
Protein Folding
Slc4
Site-directed Mutagenesis
Trafficking
Transporters
X-ray Crystallography
Abstract: Anion exchanger 1 (AE1; Band 3; SLC4A1) is the founding member of the solute carrier 4 (SLC4) family of bicarbonate transporters that includes chloride/bicarbonate AEs and Na(+)-bicarbonate co-transporters (NBCs). These membrane proteins consist of an amino-terminal cytosolic domain involved in protein interactions and a carboxyl-terminal membrane domain that carries out the transport function. Mutation of a conserved arginine residue (R298S) in the cytosolic domain of NBCe1 (SLC4A4) is linked to proximal renal tubular acidosis and results in impaired transport function, suggesting that the cytosolic domain plays a role in substrate permeation. Introduction of single and double mutations at the equivalent arginine (Arg(283)) and at an interacting glutamate (Glu(85)) in the cytosolic domain of human AE1 (cdAE1) had no effect on the cell surface expression or the transport activity of AE1 expressed in HEK-293 cells. In addition, the membrane domain of AE1 (mdAE1) efficiently mediated anion transport. A 2.1-Å resolution crystal structure of cdΔ54AE1 (residues 55-356 of cdAE1) lacking the amino-terminal and carboxyl-terminal disordered regions, produced at physiological pH, revealed an extensive hydrogen-bonded network involving Arg(283) and Glu(85). Mutations at these residues affected the pH-dependent conformational changes and stability of cdΔ54AE1. As these structural alterations did not impair functional expression of AE1, the cytosolic and membrane domains operate independently. A substrate access tunnel within the cytosolic domain is not present in AE1 and therefore is not an essential feature of the SLC4 family of bicarbonate transporters.
Notes: 1083-351x
Shnitsar, Volodymyr
Li, Jing
Li, Xuyao
Calmettes, Charles
Basu, Arghya
Casey, Joseph R
Moraes, Trevor F
Reithmeier, Reinhart A F
MOP102493/Canadian Institutes of Health Research/Canada
P41 RR015301/RR/NCRR NIH HHS/United States
RR-15301/RR/NCRR NIH HHS/United States
MOP115182/Canadian Institutes of Health Research/Canada
MOP13388/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2013 Nov 22;288(47):33848-33860. doi: 10.1074/jbc.M113.511865. Epub 2013 Oct 11.
Author Address: Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
Department of Biochemistry and Membrane Protein Disease Research Group, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada. Electronic address: r.reithmeier@utoronto.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 447
Author: Shome, B. and Parlow, A. F.
Year: 1974
Title: Human follicle stimulating hormone: first proposal for the amino acid sequence of the hormone-specific, beta subunit (hFSHb)
Journal: J Clin Endocrinol Metab
Volume: 39
Issue: 1
Pages: 203-5
Epub Date: 1974/07/01
Date: Jul
Short Title: Human follicle stimulating hormone: first proposal for the amino acid sequence of the hormone-specific, beta subunit (hFSHb)
Alternate Journal: The Journal of clinical endocrinology and metabolism
ISSN: 0021-972X (Print)
0021-972x
DOI: 10.1210/jcem-39-1-203
Accession Number: 4835136
Keywords: *Amino Acid Sequence
Chromatography, Paper
Chromatography, Thin Layer
Electrophoresis, Paper
Follicle Stimulating Hormone/*analysis
Humans
Notes: Shome, B
Parlow, A F
Journal Article
United States
J Clin Endocrinol Metab. 1974 Jul;39(1):203-5. doi: 10.1210/jcem-39-1-203.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1203
Author: Shore, D. A., Issafras, H., Landais, E., Teyton, L. and Wilson, I. A.
Year: 2008
Title: The crystal structure of CD8 in complex with YTS156.7.7 Fab and interaction with other CD8 antibodies define the binding mode of CD8 alphabeta to MHC class I
Journal: J Mol Biol
Volume: 384
Issue: 5
Pages: 1190-202
Epub Date: 2008/10/22
Date: Dec 31
Short Title: The crystal structure of CD8 in complex with YTS156.7.7 Fab and interaction with other CD8 antibodies define the binding mode of CD8 alphabeta to MHC class I
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2008.09.069
PMCID: PMC2631232
NIHMSID: NIHMS84532
Accession Number: 18929574
Keywords: Animals
Antibodies, Monoclonal/*immunology
CD8 Antigens/*chemistry/*metabolism
Crystallography, X-Ray
Dimerization
Epitopes/immunology
Histocompatibility Antigens Class I/*metabolism
Immunoglobulin Fab Fragments/*chemistry/immunology
Lymphocyte Activation
Mice
Models, Molecular
Protein Binding
Protein Structure, Quaternary
Protein Structure, Secondary
Abstract: The CD8alphabeta heterodimer interacts with class I pMHC on antigen-presenting cells as a co-receptor for TCR-mediated activation of cytotoxic T cells. To characterize this immunologically important interaction, we used monoclonal antibodies (mAbs) specific to either CD8alpha or CD8beta to probe the mechanism of CD8alphabeta binding to pMHCI. The YTS156.7 mAb inhibits this interaction and blocks T cell activation. To elucidate the molecular basis for this inhibition, the crystal structure of the CD8alphabeta immunoglobulin-like ectodomains were determined in complex with mAb YTS156.7 Fab at 2.7 A resolution. The YTS156.7 epitope on CD8beta was identified and implies that residues in the CDR1 and CDR2-equivalent loops of CD8beta are occluded upon binding to class I pMHC. To further characterize the pMHCI/CD8alphabeta interaction, binding of class I tetramers to CD8alphabeta on the surface of T cells was assessed in the presence of anti-CD8 mAbs. In contrast to YTS156.7, mAb YTS105.18, which is specific for CD8alpha, does not inhibit binding of CD8alphabeta to class I tetramers, indicating the YTS105.18 epitope is not occluded in the pMHCI/CD8alphabeta complex. Together, these data indicate a model for the pMHCI/CD8alphabeta interaction similar to that observed for CD8alphaalpha in the CD8alphaalpha/pMHCI complex, but in which CD8alpha occupies the lower orientation (membrane proximal to the antigen presenting cell), and CD8beta occupies the upper position (membrane distal). The implication of this molecular assembly for the function of CD8alphabeta in T cell activation is discussed.
Notes: 1089-8638
Shore, D A
Issafras, H
Landais, E
Teyton, L
Wilson, I A
R01 CA058896/CA/NCI NIH HHS/United States
R01 AI042267-10/AI/NIAID NIH HHS/United States
R01 AI042266/AI/NIAID NIH HHS/United States
AI042267/AI/NIAID NIH HHS/United States
R37 AI042266/AI/NIAID NIH HHS/United States
CA58896/CA/NCI NIH HHS/United States
AI042266/AI/NIAID NIH HHS/United States
R01 AI042267/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Mol Biol. 2008 Dec 31;384(5):1190-202. doi: 10.1016/j.jmb.2008.09.069. Epub 2008 Oct 7.
Author Address: Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1643
Author: Sidhu, N. S., Schreiber, K., Pröpper, K., Becker, S., Usón, I., Sheldrick, G. M., Gärtner, J., Krätzner, R. and Steinfeld, R.
Year: 2014
Title: Structure of sulfamidase provides insight into the molecular pathology of mucopolysaccharidosis IIIA
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 70
Issue: Pt 5
Pages: 1321-35
Epub Date: 2014/05/13
Date: May
Short Title: Structure of sulfamidase provides insight into the molecular pathology of mucopolysaccharidosis IIIA
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s1399004714002739
PMCID: PMC4014121
Accession Number: 24816101
Keywords: Binding Sites
Catalytic Domain
Crystallography, X-Ray
Humans
Hydrolases/*chemistry/genetics/*metabolism
Models, Molecular
Mucopolysaccharidosis III/physiopathology
Phosphates/metabolism
Protein Conformation
Structural Homology, Protein
Structure-Activity Relationship
Sulfates/metabolism
mucopolysaccharidosis IIIA
sulfamidase
Abstract: Mucopolysaccharidosis type IIIA (Sanfilippo A syndrome), a fatal childhood-onset neurodegenerative disease with mild facial, visceral and skeletal abnormalities, is caused by an inherited deficiency of the enzyme N-sulfoglucosamine sulfohydrolase (SGSH; sulfamidase). More than 100 mutations in the SGSH gene have been found to reduce or eliminate its enzymatic activity. However, the molecular understanding of the effect of these mutations has been confined by a lack of structural data for this enzyme. Here, the crystal structure of glycosylated SGSH is presented at 2 Å resolution. Despite the low sequence identity between this unique N-sulfatase and the group of O-sulfatases, they share a similar overall fold and active-site architecture, including a catalytic formylglycine, a divalent metal-binding site and a sulfate-binding site. However, a highly conserved lysine in O-sulfatases is replaced in SGSH by an arginine (Arg282) that is positioned to bind the N-linked sulfate substrate. The structure also provides insight into the diverse effects of pathogenic mutations on SGSH function in mucopolysaccharidosis type IIIA and convincing evidence for the molecular consequences of many missense mutations. Further, the molecular characterization of SGSH mutations will lay the groundwork for the development of structure-based drug design for this devastating neurodegenerative disorder.
Notes: 1399-0047
Sidhu, Navdeep S
Schreiber, Kathrin
Pröpper, Kevin
Becker, Stefan
Usón, Isabel
Sheldrick, George M
Gärtner, Jutta
Krätzner, Ralph
Steinfeld, Robert
Journal Article
Research Support, Non-U.S. Gov't
Acta Crystallogr D Biol Crystallogr. 2014 May;70(Pt 5):1321-35. doi: 10.1107/S1399004714002739. Epub 2014 Apr 30.
Author Address: Department of Neuropediatrics, Faculty of Medicine, University of Göttingen, Robert-Koch-Strasse 40, 37075 Göttingen, Germany.
Department of Structural Chemistry, Institute of Inorganic Chemistry, University of Göttingen, Tammannstrasse 4, 37077 Göttingen, Germany.
Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany.
Instituto de Biologia Molecular de Barcelona (IBMB-CSIC), Barcelona Science Park, Baldiri Reixach 15, 08028 Barcelona, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 752
Author: Silverman, G. L., Qin, K., Moore, R. C., Yang, Y., Mastrangelo, P., Tremblay, P., Prusiner, S. B., Cohen, F. E. and Westaway, D.
Year: 2000
Title: Doppel is an N-glycosylated, glycosylphosphatidylinositol-anchored protein. Expression in testis and ectopic production in the brains of Prnp(0/0) mice predisposed to Purkinje cell loss
Journal: J Biol Chem
Volume: 275
Issue: 35
Pages: 26834-41
Epub Date: 2000/06/08
Date: Sep 1
Short Title: Doppel is an N-glycosylated, glycosylphosphatidylinositol-anchored protein. Expression in testis and ectopic production in the brains of Prnp(0/0) mice predisposed to Purkinje cell loss
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M003888200
Accession Number: 10842180
Keywords: Amino Acid Sequence
Animals
Base Sequence
Brain/*metabolism
DNA Primers
GPI-Linked Proteins
Glycosylation
Glycosylphosphatidylinositols/chemistry/genetics/*metabolism
Mice
Mice, Knockout
Molecular Sequence Data
Peptide Mapping
Prions/chemistry/genetics/*metabolism
Protein Structure, Secondary
Purkinje Cells/*cytology
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Transgenes
Abstract: The Prnd gene encodes a homolog of the cellular prion protein (PrP(C)) called doppel (Dpl). Up-regulation of Prnd mRNA in two distinct lines of PrP gene ablated (Prnp(0/0)) mice, designated Rcm0 and Ngsk, is associated with death of Purkinje cells. Using recombinant Dpl expressed in Escherichia coli and mouse neuroblastoma cells we demonstrate that wild type (wt) Dpl, like PrP(C), adopts a predominantly alpha-helical conformation, forms intramolecular disulfide bonds, has two N-linked oligosaccharides, and is presented on the cell surface via a glycosylphosphatidylinositol anchor. Dpl protein was detected in testis of wt mice. Using Triton X-114 phase partitioning to enrich for glycosylphosphatidylinositol-anchored proteins, Dpl was detected in brain samples from Rcm0 Prnp(0/0) mice but was absent in equivalent samples from wt mice and ZrchI Prnp(0/0) mice, indicating that ectopic expression of this protein may cause cerebellar pathology in Rcm0 mice. Biochemical and structural similarities between PrP(C) and Dpl documented here parallel the observation that ataxic Ngsk Prnp(0/0) mice can be rescued by overexpression of wild-type PrP transgenes, and suggest that cell surface PrP(C) can antagonize the toxic effect of Dpl expressed in the central nervous system.
Notes: Silverman, G L
Qin, K
Moore, R C
Yang, Y
Mastrangelo, P
Tremblay, P
Prusiner, S B
Cohen, F E
Westaway, D
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2000 Sep 1;275(35):26834-41. doi: 10.1074/jbc.M003888200.
Author Address: Centre for Research in Neurodegenerative Diseases, Department of Laboratory Medicine and Pathobiology, and Mass Spectrometry Laboratory, Molecular Medicine Research Centre, University of Toronto, Toronto, Ontario M5S 3H2, Canada and.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 804
Author: Silverman, H. S., Parry, S., Sutton-Smith, M., Burdick, M. D., McDermott, K., Reid, C. J., Batra, S. K., Morris, H. R., Hollingsworth, M. A., Dell, A. and Harris, A.
Year: 2001
Title: In vivo glycosylation of mucin tandem repeats
Journal: Glycobiology
Volume: 11
Issue: 6
Pages: 459-71
Epub Date: 2001/07/11
Date: Jun
Short Title: In vivo glycosylation of mucin tandem repeats
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/11.6.459
Accession Number: 11445551
Keywords: Amino Acid Sequence
Antigens, Tumor-Associated, Carbohydrate/biosynthesis
Blood Group Antigens
Carbohydrate Sequence
Glycosylation
Humans
Molecular Sequence Data
Mucin-1/metabolism
Mucins/genetics/*metabolism
Oligosaccharides/chemistry
*Protein Processing, Post-Translational
Recombinant Fusion Proteins/metabolism
*Repetitive Sequences, Amino Acid
Spectrometry, Mass, Fast Atom Bombardment
Abstract: The biochemical and biophysical properties of mucins are largely determined by extensive O-glycosylation of serine- and threonine-rich tandem repeat (TR) domains. In a number of human diseases aberrant O-glycosylation is associated with variations in the properties of the cell surface-associated and secreted mucins. To evaluate in vivo the O-glycosylation of mucin TR domains, we generated recombinant chimeric mucins with TR sequences from MUC2, MUC4, MUC5AC, or MUC5B, which were substituted for the native TRs of epitope-tagged MUC1 protein (MUC1F). These hybrid mucins were extensively O-glycosylated and showed the expected association with the cell surface and release into culture media. The presence of different TR domains within the chimeric mucins appears to have limited influence on their posttranslational processing. Alterations in glycosylation were detailed by fast atom bombardment mass spectrometry and reactivity with antibodies against particular blood-group and tumor-associated carbohydrate antigens. Future applications of these chimeras will include investigations of mucin posttranslational modification in the context of disease.
Notes: Silverman, H S
Parry, S
Sutton-Smith, M
Burdick, M D
McDermott, K
Reid, C J
Batra, S K
Morris, H R
Hollingsworth, M A
Dell, A
Harris, A
P30 CA 36727/CA/NCI NIH HHS/United States
R01 CA57632/CA/NCI NIH HHS/United States
R01 CA78590/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 2001 Jun;11(6):459-71. doi: 10.1093/glycob/11.6.459.
Author Address: Paediatric Molecular Genetics, Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, OX3 9DS, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1141
Author: Sim, L., Quezada-Calvillo, R., Sterchi, E. E., Nichols, B. L. and Rose, D. R.
Year: 2008
Title: Human intestinal maltase-glucoamylase: crystal structure of the N-terminal catalytic subunit and basis of inhibition and substrate specificity
Journal: J Mol Biol
Volume: 375
Issue: 3
Pages: 782-92
Epub Date: 2007/11/27
Date: Jan 18
Short Title: Human intestinal maltase-glucoamylase: crystal structure of the N-terminal catalytic subunit and basis of inhibition and substrate specificity
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2007.10.069
Accession Number: 18036614
Keywords: Acarbose/chemistry/metabolism
Amino Acid Sequence
Apoenzymes/chemistry/metabolism
Binding Sites
Catalytic Domain
Crystallography, X-Ray
Cysteine/chemistry
Disulfides/chemistry
Enzyme Inhibitors/*metabolism
Humans
Hydrogen Bonding
Intestinal Mucosa/*enzymology
Intestines/*enzymology
Kinetics
Models, Chemical
Models, Molecular
Molecular Sequence Data
Molecular Weight
Mutation
Protein Binding
Protein Structure, Secondary
Protein Subunits/*chemistry
Recombinant Proteins/chemistry/isolation & purification/metabolism
Sequence Homology, Amino Acid
Substrate Specificity
alpha-Glucosidases/*chemistry/genetics/metabolism
Abstract: Human maltase-glucoamylase (MGAM) is one of the two enzymes responsible for catalyzing the last glucose-releasing step in starch digestion. MGAM is anchored to the small-intestinal brush-border epithelial cells and contains two homologous glycosyl hydrolase family 31 catalytic subunits: an N-terminal subunit (NtMGAM) found near the membrane-bound end and a C-terminal luminal subunit (CtMGAM). In this study, we report the crystal structure of the human NtMGAM subunit in its apo form (to 2.0 A) and in complex with acarbose (to 1.9 A). Structural analysis of the NtMGAM-acarbose complex reveals that acarbose is bound to the NtMGAM active site primarily through side-chain interactions with its acarvosine unit, and almost no interactions are made with its glycone rings. These observations, along with results from kinetic studies, suggest that the NtMGAM active site contains two primary sugar subsites and that NtMGAM and CtMGAM differ in their substrate specificities despite their structural relationship. Additional sequence analysis of the CtMGAM subunit suggests several features that could explain the higher affinity of the CtMGAM subunit for longer maltose oligosaccharides. The results provide a structural basis for the complementary roles of these glycosyl hydrolase family 31 subunits in the bioprocessing of complex starch structures into glucose.
Notes: 1089-8638
Sim, Lyann
Quezada-Calvillo, Roberto
Sterchi, Erwin E
Nichols, Buford L
Rose, David R
RR-01646/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2008 Jan 18;375(3):782-92. doi: 10.1016/j.jmb.2007.10.069. Epub 2007 Nov 1.
Author Address: Division of Cancer Genomics and Proteomics, Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, 101 College Street, Toronto, ON, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1302
Author: Sim, L., Willemsma, C., Mohan, S., Naim, H. Y., Pinto, B. M. and Rose, D. R.
Year: 2010
Title: Structural basis for substrate selectivity in human maltase-glucoamylase and sucrase-isomaltase N-terminal domains
Journal: J Biol Chem
Volume: 285
Issue: 23
Pages: 17763-70
Epub Date: 2010/04/02
Date: Jun 4
Short Title: Structural basis for substrate selectivity in human maltase-glucoamylase and sucrase-isomaltase N-terminal domains
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.078980
PMCID: PMC2878540
Accession Number: 20356844
Keywords: Animals
Carbohydrates/chemistry
Crystallography, X-Ray/methods
Diabetes Mellitus/metabolism
Drosophila/metabolism
Humans
Hydrolysis
Hydrophobic and Hydrophilic Interactions
Kinetics
Oligo-1,6-Glucosidase/*chemistry
Polysaccharides/chemistry
Protein Structure, Tertiary
Substrate Specificity
Sucrase/*chemistry
alpha-Glucosidases/*chemistry
Abstract: Human maltase-glucoamylase (MGAM) and sucrase-isomaltase (SI) are small intestinal enzymes that work concurrently to hydrolyze the mixture of linear alpha-1,4- and branched alpha-1,6-oligosaccharide substrates that typically make up terminal starch digestion products. MGAM and SI are each composed of duplicated catalytic domains, N- and C-terminal, which display overlapping substrate specificities. The N-terminal catalytic domain of human MGAM (ntMGAM) has a preference for short linear alpha-1,4-oligosaccharides, whereas N-terminal SI (ntSI) has a broader specificity for both alpha-1,4- and alpha-1,6-oligosaccharides. Here we present the crystal structure of the human ntSI, in apo form to 3.2 A and in complex with the inhibitor kotalanol to 2.15 A resolution. Structural comparison with the previously solved structure of ntMGAM reveals key active site differences in ntSI, including a narrow hydrophobic +1 subsite, which may account for its additional substrate specificity for alpha-1,6 substrates.
Notes: 1083-351x
Sim, Lyann
Willemsma, Carly
Mohan, Sankar
Naim, Hassan Y
Pinto, B Mario
Rose, David R
DMR0225180/GM/NIGMS NIH HHS/United States
P41 RR001646/RR/NCRR NIH HHS/United States
FRN79400/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Jun 4;285(23):17763-70. doi: 10.1074/jbc.M109.078980. Epub 2010 Mar 31.
Author Address: Department of Medical Biophysics, University of Toronto, Ontario Cancer Institute, Toronto, Ontario M56 2M9, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 910
Author: Simizu, S., Ishida, K., Wierzba, M. K. and Osada, H.
Year: 2004
Title: Secretion of heparanase protein is regulated by glycosylation in human tumor cell lines
Journal: J Biol Chem
Volume: 279
Issue: 4
Pages: 2697-703
Epub Date: 2003/10/24
Date: Jan 23
Short Title: Secretion of heparanase protein is regulated by glycosylation in human tumor cell lines
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M300541200
Accession Number: 14573609
Keywords: Cell Line, Tumor
Glucuronidase/*metabolism
Glycosylation
Humans
Neoplasms/*enzymology/metabolism
Protein Transport
Abstract: The endo-beta-d-glucuronidase, heparanase, is capable of specifically degrading heparan sulfate, and this activity is associated with the metastatic potential of tumor cells. The predicted amino acid sequence of heparanase includes six putative N-glycosylation sites; however, the precise biochemical role of glycosylated heparanase remains unknown. In this study, we examined the link between glycosylation and the function of heparanase in human tumor cell lines. Heparanase protein was glycosylated at six Asn residues in human tumor cell lines. Treatment with a glycosylation inhibitor demonstrated that glycosylation was not required for the activity of heparanase. However, glycosylation affected the kinetics of endoplasmic reticulum-to-Golgi transport and of secretion of the enzyme.
Notes: Simizu, Siro
Ishida, Keisuke
Wierzba, Michal K
Osada, Hiroyuki
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2004 Jan 23;279(4):2697-703. doi: 10.1074/jbc.M300541200. Epub 2003 Oct 22.
Author Address: Antibiotics Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1942
Author: Simon, D. N., Wriston, A., Fan, Q., Shabanowitz, J., Florwick, A., Dharmaraj, T., Peterson, S. B., Gruenbaum, Y., Carlson, C. R., Grønning-Wang, L. M., Hunt, D. F. and Wilson, K. L.
Year: 2018
Title: OGT (O-GlcNAc Transferase) Selectively Modifies Multiple Residues Unique to Lamin A
Journal: Cells
Volume: 7
Issue: 5
Epub Date: 2018/05/19
Date: May 17
Short Title: OGT (O-GlcNAc Transferase) Selectively Modifies Multiple Residues Unique to Lamin A
Alternate Journal: Cells
ISSN: 2073-4409 (Print)
2073-4409
DOI: 10.3390/cells7050044
PMCID: PMC5981268
Accession Number: 29772801
Keywords: O-GlcNAcylation
O-linked N-acetylglucosamine (O-GlcNAc) Transferase (OGT)
lamin
nuclear lamina
Abstract: The LMNA gene encodes lamins A and C with key roles in nuclear structure, signaling, gene regulation, and genome integrity. Mutations in LMNA cause over 12 diseases ('laminopathies'). Lamins A and C are identical for their first 566 residues. However, they form separate filaments in vivo, with apparently distinct roles. We report that lamin A is β-O-linked N-acetylglucosamine-(O-GlcNAc)-modified in human hepatoma (Huh7) cells and in mouse liver. In vitro assays with purified O-GlcNAc transferase (OGT) enzyme showed robust O-GlcNAcylation of recombinant mature lamin A tails (residues 385⁻646), with no detectable modification of lamin B1, lamin C, or 'progerin' (Δ50) tails. Using mass spectrometry, we identified 11 O-GlcNAc sites in a 'sweet spot' unique to lamin A, with up to seven sugars per peptide. Most sites were unpredicted by current algorithms. Double-mutant (S612A/T643A) lamin A tails were still robustly O-GlcNAc-modified at seven sites. By contrast, O-GlcNAcylation was undetectable on tails bearing deletion Δ50, which causes Hutchinson⁻Gilford progeria syndrome, and greatly reduced by deletion Δ35. We conclude that residues deleted in progeria are required for substrate recognition and/or modification by OGT in vitro. Interestingly, deletion Δ35, which does not remove the majority of identified O-GlcNAc sites, does remove potential OGT-association motifs (lamin A residues 622⁻625 and 639⁻645) homologous to that in mouse Tet1. These biochemical results are significant because they identify a novel molecular pathway that may profoundly influence lamin A function. The hypothesis that lamin A is selectively regulated by OGT warrants future testing in vivo, along with two predictions: genetic variants may contribute to disease by perturbing OGT-dependent regulation, and nutrient or other stresses might cause OGT to misregulate wildtype lamin A.
Notes: 2073-4409
Simon, Dan N
Wriston, Amanda
Fan, Qiong
Orcid: 0000-0002-5131-2341
Shabanowitz, Jeffrey
Florwick, Alyssa
Dharmaraj, Tejas
Peterson, Sherket B
Gruenbaum, Yosef
Carlson, Cathrine R
Grønning-Wang, Line M
Hunt, Donald F
Wilson, Katherine L
R01 GM037537/GM/NIGMS NIH HHS/United States
Journal Article
Cells. 2018 May 17;7(5):44. doi: 10.3390/cells7050044.
Author Address: Department of Cell Biology, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205, USA. dsimon01@rockefeller.edu.
Department of Chemistry, University of Virginia, Charlottesville, VA 22904, USA. asw5kg@virginia.edu.
Department of Nutrition, Institute of Basic Medical Sciences, University of Oslo, 0317 Oslo, Norway. qiong.fan@medisin.uio.no.
Department of Chemistry, University of Virginia, Charlottesville, VA 22904, USA. js4c@virginia.edu.
Department of Cell Biology, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205, USA. alyssa.florwick@duke.edu.
Department of Cell Biology, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205, USA. tdharma1@jhu.edu.
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. speter54@its.jnj.com.
Department of Genetics, Institute of Life Sciences, Hebrew University of Jerusalem, Givat Ram Jerusalem 91904, Israel. gru@mail.huji.ac.il.
Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway. c.r.carlson@medisin.uio.no.
Department of Nutrition, Institute of Basic Medical Sciences, University of Oslo, 0317 Oslo, Norway. lmgronningwang@gmail.com.
Department of Chemistry, University of Virginia, Charlottesville, VA 22904, USA. dfh@virginia.edu.
Department of Pathology, University of Virginia, Charlottesville, VA 22904, USA. dfh@virginia.edu.
Department of Cell Biology, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205, USA. klwilson@jhmi.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 815
Author: Simonovic, M., Gettins, P. G. and Volz, K.
Year: 2001
Title: Crystal structure of human PEDF, a potent anti-angiogenic and neurite growth-promoting factor
Journal: Proc Natl Acad Sci U S A
Volume: 98
Issue: 20
Pages: 11131-5
Epub Date: 2001/09/20
Date: Sep 25
Short Title: Crystal structure of human PEDF, a potent anti-angiogenic and neurite growth-promoting factor
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.211268598
PMCID: PMC58695
Accession Number: 11562499
Keywords: Amino Acid Sequence
Angiogenesis Inhibitors
Animals
Cell Line
Cricetinae
Crystallography, X-Ray
*Eye Proteins
Glycosylation
Humans
Mammals
Models, Molecular
Molecular Sequence Data
Nerve Growth Factor/*chemistry
*Nerve Growth Factors
Protein Conformation
Protein Structure, Secondary
Proteins/*chemistry/isolation & purification
Recombinant Proteins/chemistry/isolation & purification
Sequence Alignment
Sequence Homology, Amino Acid
Serpins/*chemistry/isolation & purification
Transfection
alpha 1-Antitrypsin/chemistry
Abstract: Pigment epithelium-derived factor (PEDF), a noninhibitory member of the serpin superfamily, is the most potent inhibitor of angiogenesis in the mammalian ocular compartment. It also has neurotrophic activity, both in the retina and in the central nervous system, and is highly up-regulated in young versus senescent fibroblasts. To provide a structural basis for understanding its many biological roles, we have solved the crystal structure of glycosylated human PEDF to 2.85 A. The structure revealed the organization of possible receptor and heparin-binding sites, and showed that, unlike any other previously characterized serpin, PEDF has a striking asymmetric charge distribution that might be of functional importance. These results provide a starting point for future detailed structure/function analyses into possible mechanisms of PEDF action that could lead to development of therapeutics against uncontrolled angiogenesis.
Notes: 1091-6490
Simonovic, M
Gettins, P G
Volz, K
HL64013/HL/NHLBI NIH HHS/United States
GM47522/GM/NIGMS NIH HHS/United States
P01 HL064013/HL/NHLBI NIH HHS/United States
R37 HL049234/HL/NHLBI NIH HHS/United States
HL49234/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2001 Sep 25;98(20):11131-5. doi: 10.1073/pnas.211268598. Epub 2001 Sep 18.
Author Address: Department of Biochemistry, College of Medicine, University of Illinois, Chicago, IL 60612-7334, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 346
Author: Simpson, R. J., Moritz, R. L., Rubira, M. R., Gorman, J. J. and Van Snick, J.
Year: 1989
Title: Complete amino acid sequence of a new murine T-cell growth factor P40
Journal: Eur J Biochem
Volume: 183
Issue: 3
Pages: 715-22
Epub Date: 1989/08/15
Date: Aug 15
Short Title: Complete amino acid sequence of a new murine T-cell growth factor P40
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1989.tb21103.x
Accession Number: 2528451
Keywords: Amino Acid Sequence
Animals
Cell Line
Chromatography, High Pressure Liquid
Clone Cells
Cyanogen Bromide
*Glycoproteins/isolation & purification
*Growth Substances/isolation & purification
Interleukin-9
Mass Spectrometry
Mice
Molecular Sequence Data
Molecular Weight
Peptide Fragments/isolation & purification
Peptide Hydrolases
Peptides/chemical synthesis
T-Lymphocytes, Helper-Inducer/*immunology
Trypsin
Abstract: A new murine T-cell growth factor, designated P40, which supports growth of helper T-cells in the absence of interleukin-2, interleukin-4 and antigen has been isolated from helper T-cell lines in sufficient quantities (100 micrograms) to permit its complete amino acid sequence determination. This was achieved by a combination of sensitive peptide mapping using microbore reversed-phase high performance liquid chromatography and automated microsequence analysis. Attempts to obtain N-terminal sequence data on P40 were unsuccessful due to N-terminal blockage of the native molecule. The nature of this N-terminal blocking was established using a combination of amino acid analysis, fast-atom-bombardment mass spectrometry and peptide synthesis. The P40 molecule, a single polypeptide chain comprising 126 amino acid residues, is structurally distinct from other known T-cell growth factors. No similarity was revealed when the amino acid sequence of P40 was compared with other proteins whose biochemical structure is known. The protein sequence data reported here predict four N-linked glycosylation sites in the P40 molecule.
Notes: Simpson, R J
Moritz, R L
Rubira, M R
Gorman, J J
Van Snick, J
Journal Article
England
Eur J Biochem. 1989 Aug 15;183(3):715-22. doi: 10.1111/j.1432-1033.1989.tb21103.x.
Author Address: Joint Protein Structure Laboratory, Ludwig Institute for Cancer Research (Melbourne Branch), Royal Melbourne Hospital, Parkville, Victoria, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2037
Author: Singh, J. P., Qian, K., Lee, J. S., Zhou, J., Han, X., Zhang, B., Ong, Q., Ni, W., Jiang, M., Ruan, H. B., Li, M. D., Zhang, K., Ding, Z., Lee, P., Singh, K., Wu, J., Herzog, R. I., Kaech, S., Wendel, H. G., Yates, J. R., 3rd, Han, W., Sherwin, R. S., Nie, Y. and Yang, X.
Year: 2020
Title: O-GlcNAcase targets pyruvate kinase M2 to regulate tumor growth
Journal: Oncogene
Volume: 39
Issue: 3
Pages: 560-573
Epub Date: 2019/09/11
Date: Jan
Short Title: O-GlcNAcase targets pyruvate kinase M2 to regulate tumor growth
Alternate Journal: Oncogene
ISSN: 0950-9232 (Print)
0950-9232
DOI: 10.1038/s41388-019-0975-3
PMCID: PMC7107572
NIHMSID: NIHMS1532844 interest with the contents of this article.
Accession Number: 31501520
Keywords: Acetylation
Acetylglucosamine/metabolism
Animals
Antigens, Neoplasm/*metabolism
Carrier Proteins/*metabolism
Cell Line, Tumor
Datasets as Topic
Disease Progression
Female
Gene Expression Profiling
Glycolysis
HEK293 Cells
Histone Acetyltransferases/*metabolism
Humans
Hyaluronoglucosaminidase/*metabolism
Male
Membrane Proteins/*metabolism
Mice
N-Acetylglucosaminyltransferases/*metabolism
Neoplasm Grading
Neoplasm Staging
Neoplasms/metabolism/*pathology
Protein Processing, Post-Translational
Thyroid Hormones/*metabolism
Tissue Array Analysis
Up-Regulation
Xenograft Model Antitumor Assays
Abstract: Cancer cells are known to adopt aerobic glycolysis in order to fuel tumor growth, but the molecular basis of this metabolic shift remains largely undefined. O-GlcNAcase (OGA) is an enzyme harboring O-linked β-N-acetylglucosamine (O-GlcNAc) hydrolase and cryptic lysine acetyltransferase activities. Here, we report that OGA is upregulated in a wide range of human cancers and drives aerobic glycolysis and tumor growth by inhibiting pyruvate kinase M2 (PKM2). PKM2 is dynamically O-GlcNAcylated in response to changes in glucose availability. Under high glucose conditions, PKM2 is a target of OGA-associated acetyltransferase activity, which facilitates O-GlcNAcylation of PKM2 by O-GlcNAc transferase (OGT). O-GlcNAcylation inhibits PKM2 catalytic activity and thereby promotes aerobic glycolysis and tumor growth. These studies define a causative role for OGA in tumor progression and reveal PKM2 O-GlcNAcylation as a metabolic rheostat that mediates exquisite control of aerobic glycolysis.
Notes: 1476-5594
Singh, Jay Prakash
Qian, Kevin
Lee, Jeong-Sang
Zhou, Jinfeng
Han, Xuemei
Zhang, Bichen
Ong, Qunxiang
Ni, Weiming
Jiang, Mingzuo
Ruan, Hai-Bin
Orcid: 0000-0002-3858-1272
Li, Min-Dian
Zhang, Kaisi
Ding, Zhaobing
Lee, Philip
Singh, Kamini
Orcid: 0000-0002-4608-222x
Wu, Jing
Herzog, Raimund I
Kaech, Susan
Wendel, Hans-Guido
Yates, John R 3rd
Orcid: 0000-0001-5267-1672
Han, Weiping
Orcid: 0000-0002-5023-2104
Sherwin, Robert S
Nie, Yongzhan
Yang, Xiaoyong
Orcid: 0000-0002-5315-7285
P01 DK057751/DK/NIDDK NIH HHS/United States
P30 CA008748/CA/NCI NIH HHS/United States
R01 DK089098/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Oncogene. 2020 Jan;39(3):560-573. doi: 10.1038/s41388-019-0975-3. Epub 2019 Sep 9.
Author Address: Program in Integrative Cell Signaling and Neurobiology of Metabolism, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA.
Department of Comparative Medicine, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA.
Department of Cellular and Molecular Physiology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA.
State Key Laboratory of Cancer Biology, Xijing Hospital of Digestive Diseases, Fourth Military Medical University, 127 West Changle Road, Xi'an, 710032, Shaanxi, China.
Department of Chemical Physiology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA, 92037, USA.
Singapore Bioimaging Consortium, Singapore, Singapore.
Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, 10065, USA.
School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, 710049, Shaanxi, China.
Department of Medicine, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA.
Department of Immunobiology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA.
Program in Integrative Cell Signaling and Neurobiology of Metabolism, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA. xiaoyong.yang@yale.edu.
Department of Comparative Medicine, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA. xiaoyong.yang@yale.edu.
Department of Cellular and Molecular Physiology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT, 06519, USA. xiaoyong.yang@yale.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 409
Author: Sinha, S., Watorek, W., Karr, S., Giles, J., Bode, W. and Travis, J.
Year: 1987
Title: Primary structure of human neutrophil elastase
Journal: Proc Natl Acad Sci U S A
Volume: 84
Issue: 8
Pages: 2228-32
Epub Date: 1987/04/01
Date: Apr
Short Title: Primary structure of human neutrophil elastase
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.84.8.2228
PMCID: PMC304622
Accession Number: 3550808
Keywords: Amino Acid Sequence
Cyanogen Bromide
Humans
Neutrophils/*enzymology
Pancreatic Elastase/*blood
Peptide Fragments/analysis
Peptide Hydrolases
Abstract: The complete amino acid sequence of human neutrophil elastase has been determined. The protein consists of 218 amino acid residues, contains two asparagine-linked carbohydrate side chains, and is joined together by four disulfide bonds. Comparison of the sequence to other serine proteinases indicates only moderate homology with porcine pancreatic elastase (43.0%) or neutrophil cathepsin G (37.2%). In particular, many of the residues suggested to play important roles in the mechanism by which the pancreatic elastase functions are significantly changed in the neutrophil enzyme, indicating alternative types of binding with the human proteinase.
Notes: 1091-6490
Sinha, S
Watorek, W
Karr, S
Giles, J
Bode, W
Travis, J
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1987 Apr;84(8):2228-32. doi: 10.1073/pnas.84.8.2228.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 900
Author: Sinkó, E., Iliás, A., Ujhelly, O., Homolya, L., Scheffer, G. L., Bergen, A. A., Sarkadi, B. and Váradi, A.
Year: 2003
Title: Subcellular localization and N-glycosylation of human ABCC6, expressed in MDCKII cells
Journal: Biochem Biophys Res Commun
Volume: 308
Issue: 2
Pages: 263-9
Epub Date: 2003/08/07
Date: Aug 22
Short Title: Subcellular localization and N-glycosylation of human ABCC6, expressed in MDCKII cells
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/s0006-291x(03)01349-4
Accession Number: 12901863
Keywords: Animals
Binding Sites
Cell Line
Cell Polarity
Dogs
Glycosylation
Humans
Models, Molecular
Multidrug Resistance-Associated Proteins/*chemistry/genetics/*metabolism
Mutation
Pseudoxanthoma Elasticum/genetics
Recombinant Proteins/chemistry/genetics/metabolism
Spodoptera
Subcellular Fractions/metabolism
Transduction, Genetic
Trypsin
Tunicamycin/pharmacology
Abstract: Mutations in the gene coding for a human ABC transporter protein, ABCC6 (MRP6), are responsible for the development of pseudoxanthoma elasticum. Here, we demonstrate that human ABCC6, when expressed by retroviral transduction in polarized mammalian (MDCKII) cells, is exclusively localized to the basolateral membrane. The human ABCC6 in MDCKII cells was found to be glycosylated, in contrast to the underglycosylated form of the protein, as expressed in Sf9 cells. In order to localize the major glycosylation site(s) in ABCC6, we applied limited proteolysis on the fully glycosylated and underglycosylated forms, followed by immunodetection with region-specific antibodies for ABCC6. Our results indicate that Asn15, which is located in the extracellular N-terminal region of human ABCC6, is the only N-glycosylation site in this protein. The polarized mammalian expression system characterized here provides a useful tool for further examination of routing, glycosylation, and function of the normal and pathological variants of human ABCC6.
Notes: Sinkó, Emese
Iliás, Attila
Ujhelly, Olga
Homolya, László
Scheffer, George L
Bergen, Arthur A B
Sarkadi, Balázs
Váradi, András
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2003 Aug 22;308(2):263-9. doi: 10.1016/s0006-291x(03)01349-4.
Author Address: Institute of Enzymology, Hungarian Academy of Sciences, Budapest, Hungary.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1690
Author: Skálová, T., Bláha, J., Harlos, K., Dušková, J., Koval, T., Stránský, J., Hašek, J., Vaněk, O. and Dohnálek, J.
Year: 2015
Title: Four crystal structures of human LLT1, a ligand of human NKR-P1, in varied glycosylation and oligomerization states
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 71
Issue: Pt 3
Pages: 578-91
Epub Date: 2015/03/12
Date: Mar
Short Title: Four crystal structures of human LLT1, a ligand of human NKR-P1, in varied glycosylation and oligomerization states
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s1399004714027928
PMCID: PMC4356368
Accession Number: 25760607
Keywords: Glycosylation
Humans
Lectins, C-Type/*chemistry/genetics/metabolism
NK Cell Lectin-Like Receptor Subfamily B/chemistry/genetics/metabolism
*Protein Multimerization
Protein Structure, Quaternary
Receptors, Cell Surface/*chemistry/genetics/metabolism
C-type lectin-like ligand
Llt1
Abstract: Human LLT1 is a C-type lectin-like ligand of NKR-P1 (CD161, gene KLRB1), a C-type lectin-like receptor of natural killer cells. Using X-ray diffraction, the first experimental structures of human LLT1 were determined. Four structures of LLT1 under various conditions were determined: monomeric, dimeric deglycosylated after the first N-acetylglucosamine unit in two forms and hexameric with homogeneous GlcNAc2Man5 glycosylation. The dimeric form follows the classical dimerization mode of human CD69. The monomeric form keeps the same fold with the exception of the position of an outer part of the long loop region. The hexamer of glycosylated LLT1 consists of three classical dimers. The hexameric packing may indicate a possible mode of interaction of C-type lectin-like proteins in the glycosylated form.
Notes: 1399-0047
Skálová, Tereza
Bláha, Jan
Harlos, Karl
Dušková, Jarmila
Koval', Tomáš
Stránský, Jan
Hašek, Jindřich
Vaněk, Ondřej
Dohnálek, Jan
Journal Article
Research Support, Non-U.S. Gov't
Acta Crystallogr D Biol Crystallogr. 2015 Mar;71(Pt 3):578-91. doi: 10.1107/S1399004714027928. Epub 2015 Feb 26.
Author Address: Institute of Biotechnology, Academy of Sciences of the Czech Republic, v.v.i., Vídeňská 1083, 142 20 Praha 4, Czech Republic.
Department of Biochemistry, Faculty of Science, Charles University Prague, Hlavova 8, 128 40 Praha, Czech Republic.
Division of Structural Biology, The Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, England.
Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic, v.v.i., Heyrovského nám. 2, 162 06 Praha 6, Czech Republic.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 266
Author: Skelton, T. P., Kumar, S., Smith, P. L., Beranek, M. C. and Baenziger, J. U.
Year: 1992
Title: Pro-opiomelanocortin synthesized by corticotrophs bears asparagine-linked oligosaccharides terminating with SO4-4GalNAc beta 1,4GlcNAc beta 1,2Man alpha
Journal: J Biol Chem
Volume: 267
Issue: 18
Pages: 12998-3006
Epub Date: 1992/06/25
Date: Jun 25
Short Title: Pro-opiomelanocortin synthesized by corticotrophs bears asparagine-linked oligosaccharides terminating with SO4-4GalNAc beta 1,4GlcNAc beta 1,2Man alpha
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1618797
Keywords: Amino Acid Sequence
Asparagine/*chemistry
Carbohydrate Sequence
Cells, Cultured
Chromatography, High Pressure Liquid
Fluorescent Antibody Technique
Molecular Sequence Data
N-Acetylgalactosaminyltransferases/metabolism
Oligosaccharides/*biosynthesis/chemistry
Pituitary Gland/cytology/*metabolism
Precipitin Tests
Pro-Opiomelanocortin/*biosynthesis/chemistry
Sulfotransferases/metabolism
Abstract: We have determined that greater than or equal to 80% of the Asn-linked oligosaccharides on the glycosylated form of mouse adrenocorticotropin (15-kDa adrenocorticotropin (ACTH)) bear one or more branches terminating with the sequence SO4-4GalNAc beta 1,4GlcNAc beta 1,2Man alpha (S4GGnM). Proopiomelanocortin (POMC), the precursor of ACTH, is the first example of a glycoprotein that is not a member of the glycoprotein hormone family to bear such sulfated structures. Like lutropin and thyrotropin, 15-kDa ACTH bears dibranched oligosaccharides terminating with SO4-4-GalNAc; however, at least half of the oligosaccharides on 15-kDa ACTH terminating with SO4-4-GalNAc consist of more highly branched structures that have not previously been described. Both the GalNAc beta 1,4GlcNAc beta 1,2Man-4-sulfotransferase and the glycoprotein hormone-specific GalNAc-transferase are expressed in the corticotroph-derived AtT-20 cell line. A tripeptide recognition sequence, Pro-Val-Lys, similar to the Pro-Leu-Arg sequence required for recognition of glycoprotein hormone alpha- and beta-subunits by the glycoprotein hormone-specific GalNAc-transferase, is present 8 residues amino-terminal to the glycosylated Asn of 15-kDa ACTH. Thus, POMC has the features expected for specific addition of the S4GGnM sequence to its oligosaccharides. The recent discovery of a receptor in hepatic endothelial cells that recognizes oligosaccharides terminating with S4GGnM suggests these sulfated oligosaccharides will regulate the circulatory half-life of glycosylated POMC cleavage products.
Notes: Skelton, T P
Kumar, S
Smith, P L
Beranek, M C
Baenziger, J U
R01-DK-41738/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Jun 25;267(18):12998-3006.
Author Address: Department of Pathology, Washington University Medical School, St. Louis, Missouri 63110.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 255
Author: Skerka, C., Timmann, C., Horstmann, R. D. and Zipfel, P. F.
Year: 1992
Title: Two additional human serum proteins structurally related to complement factor H. Evidence for a family of factor H-related genes
Journal: J Immunol
Volume: 148
Issue: 10
Pages: 3313-8
Epub Date: 1992/05/15
Date: May 15
Short Title: Two additional human serum proteins structurally related to complement factor H. Evidence for a family of factor H-related genes
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
Accession Number: 1533657
Keywords: Amino Acid Sequence
Base Sequence
Blood Proteins/genetics/*isolation & purification
Complement C3b Inactivator Proteins/*genetics
Complement Factor H
DNA/isolation & purification
Gene Expression
Humans
Immunoblotting
Molecular Sequence Data
Repetitive Sequences, Nucleic Acid
Abstract: We identify and characterize two human serum proteins with an apparent molecular mass of 24 and 29 kDa, which are antigenically related to complement factor H. These proteins represent differently glycosylated forms and are encoded by the same mRNA. The corresponding cDNA clone is 1051 bp in size and hybridized to a 1.4-kb mRNA derived from human liver. The predicted translation product represents a protein of 270 amino acids, which displays a hydrophobic leader sequence, indicative of a secreted protein. The secreted part is organized in four short consensus repeats (SCR) and has a single putative N-linked glycosylation site. This predicted sequence is closely related to that of the previously described factor H-related proteins h37 and h42, which are also derived from a 1.4-kb mRNA. Amino acid comparison of these factor H-related proteins showed identical leader sequences, an exchange of three amino acids in SCR1, identical sequences of SCR2, and a lower degree of homology between SCR3-4 (h24 and h29) and SCR4-5 (h37 and h42). In addition, SCR3-4 of h24 and h29 display homology to SCR19-20 of human complement factor H. The relatedness of structural elements of the factor H-related proteins h24, h29, h37, and h42 and of factor H, suggests a function common to these proteins and indicates the existence of a gene family consisting of factor H and at least two factor H-related genes.
Notes: Skerka, C
Timmann, C
Horstmann, R D
Zipfel, P F
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 1992 May 15;148(10):3313-8.
Author Address: Bernhard Nocht Institute for Tropical Medicine, Hamburg, FRG.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1195
Author: Skiniotis, G., Lupardus, P. J., Martick, M., Walz, T. and Garcia, K. C.
Year: 2008
Title: Structural organization of a full-length gp130/LIF-R cytokine receptor transmembrane complex
Journal: Mol Cell
Volume: 31
Issue: 5
Pages: 737-48
Epub Date: 2008/09/09
Date: Sep 5
Short Title: Structural organization of a full-length gp130/LIF-R cytokine receptor transmembrane complex
Alternate Journal: Molecular cell
ISSN: 1097-2765 (Print)
1097-2765
DOI: 10.1016/j.molcel.2008.08.011
PMCID: PMC2607196
NIHMSID: HHMIMS69382
Accession Number: 18775332
Keywords: Animals
Ciliary Neurotrophic Factor/chemistry/genetics/metabolism
Crystallography, X-Ray
Cytokine Receptor gp130/*chemistry/genetics/metabolism
Humans
Models, Molecular
Molecular Sequence Data
Multiprotein Complexes/*chemistry/ultrastructure
*Protein Structure, Quaternary
Receptor, Ciliary Neurotrophic Factor/chemistry/genetics/metabolism
Receptors, OSM-LIF/*chemistry/genetics/metabolism
Signal Transduction/*physiology
Thermodynamics
Abstract: gp130 is a shared receptor for at least nine cytokines and can signal either as a homodimer or as a heterodimer with Leukemia Inhibitory Factor Receptor (LIF-R). Here, we biophysically and structurally characterize the full-length, transmembrane form of a quaternary cytokine receptor complex consisting of gp130, LIF-R, the cytokine Ciliary Neurotrophic Factor (CNTF), and its alpha receptor (CNTF-Ralpha). Thermodynamic analysis indicates that, unlike the cooperative assembly of the symmetric gp130/Interleukin-6/IL-6Ralpha hexameric complex, CNTF/CNTF-Ralpha heterodimerizes gp130 and LIF-R via noncooperative energetics to form an asymmetric 1:1:1:1 complex. Single particle electron microscopic analysis of the full-length gp130/LIF-R/CNTF-Ralpha/CNTF quaternary complex elucidates an asymmetric structural arrangement, in which the receptor extracellular and transmembrane segments join as a continuous, rigid unit, poised to sensitively transduce ligand engagement to the membrane-proximal intracellular signaling regions. These studies also enumerate the organizing principles for assembly of the "tall" class of gp130 family cytokine receptor complexes including LIF, IL-27, IL-12, and others.
Notes: 1097-4164
Skiniotis, Georgios
Lupardus, Patrick J
Martick, Monika
Walz, Thomas
Garcia, K Christopher
P01 GM062580/GM/NIGMS NIH HHS/United States
R01 AI051321-06A1/AI/NIAID NIH HHS/United States
R37 AI051321/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
AI51321/AI/NIAID NIH HHS/United States
GM62580/GM/NIGMS NIH HHS/United States
R01 AI051321/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cell. 2008 Sep 5;31(5):737-48. doi: 10.1016/j.molcel.2008.08.011.
Author Address: Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1320
Author: Skorobogatko, Y. V., Deuso, J., Adolf-Bryfogle, J., Nowak, M. G., Gong, Y., Lippa, C. F. and Vosseller, K.
Year: 2011
Title: Human Alzheimer's disease synaptic O-GlcNAc site mapping and iTRAQ expression proteomics with ion trap mass spectrometry
Journal: Amino Acids
Volume: 40
Issue: 3
Pages: 765-79
Epub Date: 2010/06/22
Date: Mar
Short Title: Human Alzheimer's disease synaptic O-GlcNAc site mapping and iTRAQ expression proteomics with ion trap mass spectrometry
Alternate Journal: Amino acids
ISSN: 0939-4451
DOI: 10.1007/s00726-010-0645-9
Accession Number: 20563614
Keywords: Acetylglucosamine/*metabolism
Alzheimer Disease/*genetics/*metabolism
Amino Acid Sequence
Animals
Cerebral Cortex/chemistry/metabolism
Glycosylation
Humans
Mass Spectrometry
Mice
Molecular Sequence Data
Neuronal Plasticity
Peptide Mapping
Phosphorylation
Proteins/chemistry/genetics/*metabolism
Proteomics/*methods
Sequence Alignment
Synapses/*chemistry/*metabolism
Abstract: Neuronal synaptic functional deficits are linked to impaired learning and memory in Alzheimer's disease (AD). We recently demonstrated that O-GlcNAc, a novel cytosolic and nuclear carbohydrate post-translational modification, is enriched at neuronal synapses and positively regulates synaptic plasticity linked to learning and memory in mice. Reduced levels of O-GlcNAc have been observed in AD, suggesting a possible link to deficits in synaptic plasticity. Using lectin enrichment and mass spectrometry, we mapped several human cortical synaptic O-GlcNAc modification sites. Overlap in patterns of O-GlcNAcation between mouse and human appears to be high, as previously mapped mouse synaptic O-GlcNAc sites in Bassoon, Piccolo, and tubulin polymerization promoting protein p25 were identified in human. Novel O-GlcNAc modification sites were identified on Mek2 and RPN13/ADRM1. Mek2 is a signaling component of the Erk 1/2 pathway involved in synaptic plasticity. RPN13 is a component of the proteasomal degradation pathway. The potential interplay of phosphorylation with mapped O-GlcNAc sites, and possible implication of those sites in synaptic plasticity in normal versus AD states is discussed. iTRAQ is a powerful differential isotopic quantitative approach in proteomics. Pulsed Q dissociation (PQD) is a recently introduced fragmentation strategy that enables detection of low mass iTRAQ reporter ions in ion trap mass spectrometry. We optimized LTQ ion trap settings for PQD-based iTRAQ quantitation and demonstrated its utility in O-GlcNAc site mapping. Using iTRAQ, abnormal synaptic expression levels of several proteins previously implicated in AD pathology were observed in addition to novel changes in synaptic specific protein expression including Synapsin II.
Notes: 1438-2199
Skorobogatko, Yuliya V
Deuso, John
Adolf-Bryfogle, Jared
Nowak, Matthew G
Gong, Yuesong
Lippa, Carol Frances
Vosseller, Keith
Journal Article
Austria
Amino Acids. 2011 Mar;40(3):765-79. doi: 10.1007/s00726-010-0645-9. Epub 2010 Jun 19.
Author Address: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1180
Author: Slawson, C., Lakshmanan, T., Knapp, S. and Hart, G. W.
Year: 2008
Title: A mitotic GlcNAcylation/phosphorylation signaling complex alters the posttranslational state of the cytoskeletal protein vimentin
Journal: Mol Biol Cell
Volume: 19
Issue: 10
Pages: 4130-40
Epub Date: 2008/07/26
Date: Oct
Short Title: A mitotic GlcNAcylation/phosphorylation signaling complex alters the posttranslational state of the cytoskeletal protein vimentin
Alternate Journal: Molecular biology of the cell
ISSN: 1059-1524 (Print)
1059-1524
DOI: 10.1091/mbc.e07-11-1146
PMCID: PMC2555957
Accession Number: 18653473
Keywords: Acetylglucosamine/*metabolism
Aurora Kinase B
Aurora Kinases
Cell Division
Cytokinesis
HeLa Cells
Humans
Microscopy, Confocal
Miosis
*Mitosis
Models, Biological
N-Acetylglucosaminyltransferases/*metabolism
Phosphorylation
*Protein Processing, Post-Translational
Protein-Serine-Threonine Kinases/metabolism
Signal Transduction
Vimentin/*metabolism
Abstract: O-linked beta-N-acetylglucosamine (O-GlcNAc) is a highly dynamic intracellular protein modification responsive to stress, hormones, nutrients, and cell cycle stage. Alterations in O-GlcNAc addition or removal (cycling) impair cell cycle progression and cytokinesis, but the mechanisms are not well understood. Here, we demonstrate that the enzymes responsible for O-GlcNAc cycling, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) are in a transient complex at M phase with the mitotic kinase Aurora B and protein phosphatase 1. OGT colocalized to the midbody during telophase with Aurora B. Furthermore, these proteins coprecipitated with each other in a late mitotic extract. The complex was stable under Aurora inhibition; however, the total cellular levels of O-GlcNAc were increased and the localization of OGT was decreased at the midbody after Aurora inhibition. Vimentin, an intermediate filament protein, is an M phase substrate for both Aurora B and OGT. Overexpression of OGT or OGA led to defects in mitotic phosphorylation on multiple sites, whereas OGT overexpression increased mitotic GlcNAcylation of vimentin. OGA inhibition caused a decrease in vimentin late mitotic phosphorylation but increased GlcNAcylation. Together, these data demonstrate that the O-GlcNAc cycling enzymes associate with kinases and phosphatases at M phase to regulate the posttranslational status of vimentin.
Notes: 1939-4586
Slawson, Chad
Lakshmanan, T
Knapp, Spencer
Hart, Gerald W
R37 HD013563/HD/NICHD NIH HHS/United States
CA42496/CA/NCI NIH HHS/United States
HD13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Mol Biol Cell. 2008 Oct;19(10):4130-40. doi: 10.1091/mbc.e07-11-1146. Epub 2008 Jul 23.
Author Address: Department of Biological Chemistry, The Johns Hopkins School of Medicine, Baltimore, MD 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 999
Author: Slawson, C., Zachara, N. E., Vosseller, K., Cheung, W. D., Lane, M. D. and Hart, G. W.
Year: 2005
Title: Perturbations in O-linked beta-N-acetylglucosamine protein modification cause severe defects in mitotic progression and cytokinesis
Journal: J Biol Chem
Volume: 280
Issue: 38
Pages: 32944-56
Epub Date: 2005/07/20
Date: Sep 23
Short Title: Perturbations in O-linked beta-N-acetylglucosamine protein modification cause severe defects in mitotic progression and cytokinesis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M503396200
Accession Number: 16027160
Keywords: 3T3-L1 Cells
Acetylglucosamine/*chemistry
Adenoviridae/genetics
Animals
Blotting, Western
Cell Cycle
Cell Division
Cell Nucleus/metabolism
Cell Proliferation
Cytokinesis
Cytoplasm/metabolism
Flow Cytometry
Glycosylation
HeLa Cells
Humans
Mice
Microscopy, Confocal
Microscopy, Fluorescence
*Mitosis
NIH 3T3 Cells
Phenotype
Phosphorylation
Thymidine/chemistry
Time Factors
Abstract: The dynamic modification of nuclear and cytoplasmic proteins with O-linked beta-N-acetylglucosamine (O-GlcNAc) is a regulatory post-translational modification that is rapidly responsive to morphogens, hormones, nutrients, and cellular stress. Here we show that O-GlcNAc is an important regulator of the cell cycle. Increased O-GlcNAc (pharmacologically or genetically) results in growth defects linked to delays in G2/M progression, altered mitotic phosphorylation, and cyclin expression. Overexpression of O-GlcNAcase, the enzyme that removes O-GlcNAc, induces a mitotic exit phenotype accompanied by a delay in mitotic phosphorylation, altered cyclin expression, and pronounced disruption in nuclear organization. Overexpression of the O-GlcNAc transferase, the enzyme that adds O-GlcNAc, results in a polyploid phenotype with faulty cytokinesis. Notably, O-GlcNAc transferase is concentrated at the mitotic spindle and midbody at M phase. These data suggest that dynamic O-GlcNAc processing is a pivotal regulatory component of the cell cycle, controlling cell cycle progression by regulating mitotic phosphorylation, cyclin expression, and cell division.
Notes: Slawson, Chad
Zachara, Natasha E
Vosseller, Keith
Cheung, Win D
Lane, M Daniel
Hart, Gerald W
CA42486/CA/NCI NIH HHS/United States
DK61671/DK/NIDDK NIH HHS/United States
HD13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2005 Sep 23;280(38):32944-56. doi: 10.1074/jbc.M503396200. Epub 2005 Jul 18.
Author Address: Department of Biological Chemistry, the Johns Hopkins School of Medicine, Baltimore, Maryland 21205-2185, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 593
Author: Slomiany, B. L., Murty, V. L. and Slomiany, A.
Year: 1993
Title: Structural features of carbohydrate chains in human salivary mucins
Journal: Int J Biochem
Volume: 25
Issue: 2
Pages: 259-65
Epub Date: 1993/02/01
Date: Feb
Short Title: Structural features of carbohydrate chains in human salivary mucins
Alternate Journal: The International journal of biochemistry
ISSN: 0020-711X (Print)
0020-711x
DOI: 10.1016/0020-711x(93)90015-7
Accession Number: 8444322
Keywords: ABO Blood-Group System
Adult
Carbohydrate Sequence
Carbohydrates/*chemistry
Glycoproteins/chemistry
Humans
Molecular Sequence Data
Molecular Weight
Mucins/*chemistry
N-Acetylneuraminic Acid
Oligosaccharides/analysis/chemistry
Saliva/*chemistry
Sialic Acids/analysis
Submandibular Gland/metabolism
Sugar Alcohols/chemistry
beta-Galactosidase/metabolism
beta-N-Acetylhexosaminidases/metabolism
Abstract: 1. The structure of carbohydrate chains in the low and high molecular weight mucus glycoprotein forms from submandibular-sublingual saliva of individuals with blood group B was investigated. 2. Alkaline borohydride reductive cleavage of the glycoproteins yielded in each case a population of neutral (55%) and acidic (45%) oligosaccharide alditols ranging in size from 3 to 16 sugar units. 3. The predominant neutral oligosaccharides in both glycoprotein forms consisted of 16 and 15 sugar units arranged in triantennary fashion, and carried blood group B and I antigenic determinants. 4. Three of the oligosaccharides in each glycoprotein contained sialic acid and ranged in size from 3 to 12 sugar units. In two oligosaccharides sialic acid was linked to C3 of galactose and in one to C6 of N-acetylgalactosamine. The sulfated oligosaccharide in both glycoproteins was identified as a pentasaccharide with the sulfate ester group at C6 of N-acetylglucosamine. 5. The results demonstrate that contrary to the earlier view the low and high molecular weight mucus glycoprotein forms of human saliva contain identical carbohydrate chains.
Notes: Slomiany, B L
Murty, V L
Slomiany, A
Journal Article
England
Int J Biochem. 1993 Feb;25(2):259-65. doi: 10.1016/0020-711x(93)90015-7.
Author Address: Research Center, New Jersey Dental School, University of Medicine and Dentistry of New Jersey, Newark 07103-2400.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 317
Author: Smagula, R. M., Van Halbeek, H., Decker, J. M., Muchmore, A. V., Moody, C. E. and Sherblom, A. P.
Year: 1990
Title: Pregnancy-associated changes in oligomannose oligosaccharides of human and bovine uromodulin (Tamm-Horsfall glycoprotein)
Journal: Glycoconj J
Volume: 7
Issue: 6
Pages: 609-24
Epub Date: 1990/01/01
Short Title: Pregnancy-associated changes in oligomannose oligosaccharides of human and bovine uromodulin (Tamm-Horsfall glycoprotein)
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/bf01189081
Accession Number: 2136357
Keywords: Animals
Carbohydrate Sequence
Cattle
Cell Division
Chromatography, Gel
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
Female
Humans
Mannose/*metabolism
Mice
Molecular Sequence Data
Mucoproteins/*metabolism/urine
Oligosaccharides/*metabolism
Pregnancy
Pregnancy Proteins/*metabolism
T-Lymphocytes/immunology
Uromodulin
Abstract: The urinary glycoprotein uromodulin (Tamm-Horsfall glycoprotein) exhibits a pregnancy-associated ability to inhibit antigen-specific T cell proliferation, and the activity is associated with a carbohydrate moiety [Muchmore and Decker (1985) Science 229:479-81; Hession et al., (1987) Science 237:1479-84; Muchmore, Shifrin and Decker (1987) J Immunol 138:2547-53]. We report here that the Man6(7)GlcNAc2-R glycopeptides derived from uromodulin inhibit antigen-specific T cell proliferation by 50% at 0.2-2 microM, and further studies, reported elsewhere, confirm that oligomannose glycopeptides from other sources are also inhibitory, with Man9GlcNAc2-R the most inhibitory of those tested [Muchmore et al., J Leukocyte Biol (in press)]. In this work, we have extended the observation of pregnancy-associated inhibitory activity to a second species, and have compared the oligomannose profile of Tamm-Horsfall glycoprotein (nonpregnant) with that of uromodulin (pregnant) derived from both human and bovine sources. Surprisingly, there was a pregnancy-associated decrease in the total content of oligomannose chains due predominantly to a reduction in Man5GlcNAc2-R and Man6GlcNAc2-R. Man7GlcNAc2-R, which did not decrease with pregnancy, comprised a significantly greater proportion of the total oligomannose chains in pregnant vs. nonpregnant samples from both species (human; 34.6% vs. 25.9%: bovine; 14.4% vs. 7.2%).
Notes: Smagula, R M
Van Halbeek, H
Decker, J M
Muchmore, A V
Moody, C E
Sherblom, A P
P41-RR-05351/RR/NCRR NIH HHS/United States
S07 RR07161/RR/NCRR NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Glycoconj J. 1990;7(6):609-24. doi: 10.1007/BF01189081.
Author Address: Department of Biochemistry, University of Maine, Orono 04469.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1381
Author: Smet-Nocca, C., Broncel, M., Wieruszeski, J. M., Tokarski, C., Hanoulle, X., Leroy, A., Landrieu, I., Rolando, C., Lippens, G. and Hackenberger, C. P.
Year: 2011
Title: Identification of O-GlcNAc sites within peptides of the Tau protein and their impact on phosphorylation
Journal: Mol Biosyst
Volume: 7
Issue: 5
Pages: 1420-9
Epub Date: 2011/02/18
Date: May
Short Title: Identification of O-GlcNAc sites within peptides of the Tau protein and their impact on phosphorylation
Alternate Journal: Molecular bioSystems
ISSN: 1742-2051
DOI: 10.1039/c0mb00337a
Accession Number: 21327254
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Binding Sites
Cyclin A/genetics/metabolism
Cyclin-Dependent Kinase 2/genetics/metabolism
Glycogen Synthase Kinase 3/genetics/metabolism
Glycogen Synthase Kinase 3 beta
Glycosylation
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
N-Acetylglucosaminyltransferases/genetics/*metabolism
Peptides/*metabolism
Phosphorylation
Recombinant Proteins/metabolism
Serine/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Substrate Specificity
tau Proteins/chemistry/*metabolism
Abstract: Phosphorylation of the microtubule-associated Tau protein plays a major role in the regulation of its activity of tubulin polymerization and/or stabilization of microtubule assembly. A dysregulation of the phosphorylation/dephosphorylation balance leading to the hyperphosphorylation of Tau proteins in neurons is thought to favor their aggregation into insoluble filaments. This in turn might underlie neuronal death as encountered in many neurodegenerative disorders, including Alzheimer's disease. Another post-translational modification, the O-linked β-N-acetylglucosaminylation (O-GlcNAcylation), controls the phosphorylation state of Tau, although the precise mechanism is not known. Moreover, analytical difficulties have hampered the precise localization of the O-GlcNAc sites on Tau, except for the S400 site that was very recently identified on the basis of ETD-FT-MS. Here, we identify three O-GlcNAc sites by screening a library of small peptides sampling the proline-rich, the microtubule-associated repeats and the carboxy-terminal domains of Tau as potential substrates for the O-β-N-acetylglucosaminyltransferase (OGT). The in vitro activity of the nucleocytoplasmic OGT was assessed by tandem mass spectrometry and NMR spectroscopy. Using phosphorylated peptides, we establish the relationship between phosphate and O-GlcNAc incorporation at these sites. Phosphorylation of neighboring residues S396 and S404 was found to decrease significantly S400 O-GlcNAcylation. Reciprocally, S400 O-GlcNAcylation reduces S404 phosphorylation by the CDK2/cyclinA3 kinase and interrupts the GSK3β-mediated sequential phosphorylation process.
Notes: 1742-2051
Smet-Nocca, Caroline
Broncel, Malgorzata
Wieruszeski, Jean-Michel
Tokarski, Caroline
Hanoulle, Xavier
Leroy, Arnaud
Landrieu, Isabelle
Rolando, Christian
Lippens, Guy
Hackenberger, Christian P R
Journal Article
Research Support, Non-U.S. Gov't
England
Mol Biosyst. 2011 May;7(5):1420-9. doi: 10.1039/c0mb00337a. Epub 2011 Feb 16.
Author Address: Unité de Glycobiologie Structurale et Fonctionnelle, CNRS UMR8576, IFR 147, Université de Lille1, 59655 Villeneuve d'Ascq Cedex, France. caroline.smet@univ-lille1.fr
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 265
Author: Smith, C. A., Anderson, B. F., Baker, H. M. and Baker, E. N.
Year: 1992
Title: Metal substitution in transferrins: the crystal structure of human copper-lactoferrin at 2.1-A resolution
Journal: Biochemistry
Volume: 31
Issue: 18
Pages: 4527-33
Epub Date: 1992/05/12
Date: May 12
Short Title: Metal substitution in transferrins: the crystal structure of human copper-lactoferrin at 2.1-A resolution
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00133a020
Accession Number: 1581307
Keywords: Binding Sites
Copper/*chemistry
Humans
Iron/*chemistry/metabolism
Lactoferrin/*chemistry
Models, Molecular
Particle Size
Peptides/chemistry
Protein Binding
Protein Conformation
Stereoisomerism
Transferrin/*chemistry/metabolism
X-Ray Diffraction
Abstract: The structural consequences of binding a metal other than iron to a transferrin have been examined by crystallographic analysis of human copper-lactoferrin, Cu2Lf. X-ray diffraction data were collected from crystals of Cu2Lf, using a diffractometer, to 2.6-A resolution, and oscillation photography on a synchrotron source, to 2.1-A resolution. The structure was refined crystallographically, by restrained least-squares methods, starting with a model based on the isomorphous diferric structure from which the ligands, metal ions, anions, and solvent molecules had been deleted. The final model, comprising 5321 protein atoms (691 residues), 2 Cu2+ ions, 2 (bi)carbonate ions, and 308 solvent molecules has good stereochemistry (rms deviation of bond lengths from standard values of 0.018 A) and gives a crystallographic R value of 0.196 for 43,525 reflections in the range 7.5-2.1-A resolution. The copper coordination is different in the two binding sites. In the N-terminal site, the geometry is square pyramidal, with equatorial bonds to Asp 60, Tyr 192, His 253, and a monodentate anion and a longer apical bond to Tyr 92. In the C-terminal site, the geometry is distorted octahedral, with bonds to Asp 395, Tyr 435, Tyr 528, and His 597 and an asymmetrically bidentate anion. The protein structure is the same as for the diferric protein, Fe2Lf, demonstrating that the closure of the protein domains over the metal is the same in each case irrespective of whether Fe3+ or Cu2+ is bound and that copper could be transported and delivered to cells equally well as iron. The differences in metal coordination are achieved by small movements of the metal ion and anion within each binding site, which do not affect the protein structure.
Notes: Smith, C A
Anderson, B F
Baker, H M
Baker, E N
HD-20859/HD/NICHD NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1992 May 12;31(18):4527-33. doi: 10.1021/bi00133a020.
Author Address: Department of Chemistry and Biochemistry, Massey University, Palmerston North, New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 945
Author: Smith, C. A., Anderson, B. F., Baker, H. M. and Baker, E. N.
Year: 1994
Title: Structure of copper- and oxalate-substituted human lactoferrin at 2.0 A resolution
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 50
Issue: Pt 3
Pages: 302-16
Epub Date: 1994/05/01
Date: May 1
Short Title: Structure of copper- and oxalate-substituted human lactoferrin at 2.0 A resolution
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s0907444994000491
Accession Number: 15299444
Abstract: The three-dimensional structure of human dicupric monooxalate lactoferrin, Cu(2)oxLf, has been determined to 2.0 A resolution, using X-ray diffraction data collected by diffractometry to 2.5 A resolution, and oscillation photography on a synchrotron source to 2.0 A resolution. Difference electron-density maps calculated between Cu(2)oxLf and both dicupric lactoferrin, Cu(2)Lf, and diferric lactoferrin, Fe(2)Lf, showed that the oxalate had replaced a carbonate in the C-terminal binding site, and that, relative to Cu(2)Lf, there were no significant differences in the N-terminal site. The structure was then refined crystallographically by restrained least-squares methods. The final model, in which the r.m.s. deviation in bond distances is 0.017 A, contains 5314 protein atoms (691 residues), two Cu(2+) ions, one bicarbonate ion, one oxalate ion, 325 solvent molecules and one sugar residue. The crystallographic R factor of 0.193 is for 46 134 reflections in the range 8.0 to 2.0 A resolution. The oxalate ion is coordinated to copper in a 1,2-bidentate fashion, and the added bulk of the anion results in the rearrangement of the side chains of nearby arginine and tyrosine residues. No other major alterations in the molecule can be observed, the overall protein structure being the same as that for Cu(2)Lf and Fe(2)Lf.
Notes: Smith, C A
Anderson, B F
Baker, H M
Baker, E N
Journal Article
United States
Acta Crystallogr D Biol Crystallogr. 1994 May 1;50(Pt 3):302-16. doi: 10.1107/S0907444994000491.
Author Address: Department of Chemistry and Biochemistry, Massey University, Palmerston North, New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 628
Author: Smith, K. D., Davies, M. J., Bailey, D., Renouf, D. V. and Hounsell, E. F.
Year: 1996
Title: Analysis of the glycosylation patterns of the extracellular domain of the epidermal growth factor receptor expressed in Chinese hamster ovary fibroblasts
Journal: Growth Factors
Volume: 13
Issue: 1-2
Pages: 121-32
Epub Date: 1996/01/01
Short Title: Analysis of the glycosylation patterns of the extracellular domain of the epidermal growth factor receptor expressed in Chinese hamster ovary fibroblasts
Alternate Journal: Growth factors (Chur, Switzerland)
ISSN: 0897-7194 (Print)
0897-7194
DOI: 10.3109/08977199609034572
Accession Number: 8962717
Keywords: Amidohydrolases/metabolism
Amino Acid Sequence
Animals
Binding Sites/genetics
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Cricetinae
Electrophoresis, Polyacrylamide Gel
Epidermal Growth Factor/metabolism
ErbB Receptors/chemistry/*metabolism
Gas Chromatography-Mass Spectrometry
Gene Expression Regulation/genetics
Glycosylation
Molecular Sequence Data
Monosaccharides/analysis
Oligosaccharides/chemistry
Peptide Mapping
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Peptides/chemistry
Protein Binding
Recombinant Proteins/genetics
Sequence Alignment
Sequence Analysis
Trypsin/metabolism
Abstract: The extracellular domain (621 N-terminal amino acids) of the p170 epidermal growth factor (EGF) receptor has eleven consensus N-linked glycosylation sites. When expressed in Chinese hamster ovary cells this was glycosylated with a combination of high mannose and complex chains. The latter chains were shown by chromatographic separation and mass spectrometric analysis of tryptic digests to be clustered in the EGF-binding domain. Treatment with the endoglycosidase, peptide-N-glycosidase F (PNGase F), reduced the molecular weight from 110 kDa to 75 kDa. Released oligosaccharides were characterised at high sensitivity by high pH anion exchange chromatography with pulsed amperometric detection and gas-liquid chromatography/mass spectrometry. The data were consistent with the complex chains being trisialylated tetra-antennary oligosaccharides fucosylated on the reducing terminal GlcNAc. The large hydrodynamic mass of these oligosaccharides could influence ligand binding, an effect which is likely to vary with the difference in consensus glycosylation sites of proteins related to p170 i.e. p185erbB2/neu, p180erbB3 and p180erbB4.
Notes: Smith, K D
Davies, M J
Bailey, D
Renouf, D V
Hounsell, E F
Journal Article
Research Support, Non-U.S. Gov't
England
Growth Factors. 1996;13(1-2):121-32. doi: 10.3109/08977199609034572.
Author Address: Department of Pharmaceutical Sciences, University of Strathclyde, Glasgow, Scotland, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 235
Author: Smith, P. L., Skelton, T. P., Fiete, D., Dharmesh, S. M., Beranek, M. C., MacPhail, L., Broze, G. J., Jr. and Baenziger, J. U.
Year: 1992
Title: The asparagine-linked oligosaccharides on tissue factor pathway inhibitor terminate with SO4-4GalNAc beta 1, 4GlcNAc beta 1,2 Mana alpha
Journal: J Biol Chem
Volume: 267
Issue: 27
Pages: 19140-6
Epub Date: 1992/10/05
Date: Sep 25
Short Title: The asparagine-linked oligosaccharides on tissue factor pathway inhibitor terminate with SO4-4GalNAc beta 1, 4GlcNAc beta 1,2 Mana alpha
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1388166
Keywords: Acetates/chemistry
Acetic Acid
Animals
Asparagine/chemistry
CHO Cells
Carbohydrate Sequence
Cricetinae
Endothelium, Vascular/metabolism
Galactosyltransferases/metabolism
Humans
Lipoproteins/*chemistry/metabolism
Metabolic Clearance Rate
Molecular Sequence Data
N-Acetylgalactosaminyltransferases/metabolism
Oligosaccharides/chemistry/metabolism
Recombinant Proteins/chemistry/metabolism
Sulfotransferases/metabolism
beta-N-Acetylhexosaminidases/metabolism
Abstract: Tissue factor pathway inhibitor (TFPI) produced by endothelial cells contains sulfated Asn-linked oligosaccharides. We have determined that greater than 70% of the oligosaccharides on recombinant TFPI expressed in 293 cells terminate with the sequence SO4-4GalNAc beta 1, 4GlcNAc beta 1, 2Man alpha. Oligosaccharides terminating with this sequence have previously been described on lutropin, thyrotropin, and pro-opiomelanocortin: glycoproteins synthesized in the anterior pituitary. A GalNAc-transferase that recognizes the tripeptide motif Pro-Xaa-Arg/Lys 6-9 residues N-terminal to Asn glycosylation sites accounts for the specific addition of GalNAc to the oligosaccharide acceptor on these glycoproteins, whereas a GalNAc beta 1,4GlcNAc beta 1, 2Man alpha-4-sulfotransferase accounts for the addition of sulfate. The sulfated oligosaccharides present on these hormones are responsible for their rapid clearance from plasma by a receptor in hepatic reticuloendothelial cells. GalNAc- and sulfotransferase activities with the same properties as those expressed in the pituitary are detected at high levels in 293 cells and at lower levels in endothelial cells. Chinese hamster ovary (CHO) cells do not contain detectable levels of either transferase and rTFPI expressed in CHO cells does not contain sulfated Asn-linked oligosaccharides. TFPI contains the sequence Pro-Phe-Lys, 9 residues N-terminal to the glycosylation site at position 228; this tripeptide may act as the recognition sequence for the GalNAc-transferase. rTFPI produced by 293 cells, but not that produced by CHO cells, is bound by the receptor on hepatic reticuloendothelial cells suggesting the sulfated structures play a role in the biologic behavior of TFPI.
Notes: Smith, P L
Skelton, T P
Fiete, D
Dharmesh, S M
Beranek, M C
MacPhail, L
Broze, G J Jr
Baenziger, J U
R37-CA21923/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Sep 25;267(27):19140-6.
Author Address: Department of Pathology, Washington University School of Medicine, St. Louis, Missouri.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 545
Author: Snyder, P. M., McDonald, F. J., Stokes, J. B. and Welsh, M. J.
Year: 1994
Title: Membrane topology of the amiloride-sensitive epithelial sodium channel
Journal: J Biol Chem
Volume: 269
Issue: 39
Pages: 24379-83
Epub Date: 1994/09/30
Date: Sep 30
Short Title: Membrane topology of the amiloride-sensitive epithelial sodium channel
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7929098
Keywords: Amiloride/*pharmacology
Amino Acid Sequence
Animals
Cells, Cultured
Epithelium/drug effects/physiology
Female
Glycosylation
Intracellular Membranes/chemistry/physiology
Membrane Potentials
Molecular Sequence Data
Oocytes
Protein Conformation
Recombinant Proteins
Sodium Channels/*chemistry/drug effects/physiology
Xenopus laevis
Abstract: The amiloride-sensitive epithelial sodium channel (ENaC) is involved in fluid and electrolyte absorption across a number of epithelia, and cloning of several ENaC subunits has begun to facilitate investigation of the structure, function, and regulation of this channel. Analysis of the amino acid sequence has revealed two potential membrane-spanning domains, but little else is known about the structure of ENaC. To investigate the membrane topology of one subunit, alpha rENaC, we used in vitro transcription, translation, and translocation into microsomal membranes. This generated a glycosylated protein of 93 kDa. Sequence analysis also revealed eight potential sites for N-glycosylation, six of which were found to be glycosylated (Asn190, Asn259, Asn320, Asn339, Asn424, and Asn538), indicating that they are extracellular. The C terminus was localized as intracellular based on antibody recognition and protease sensitivity of a tagged epitope at the C terminus. The N terminus was also found to be intracellular, based on its protease sensitivity. Similar results were obtained by expression in Xenopus oocytes. Together, these results support a model of alpha rENaC consisting of an intracellular N terminus and C terminus, a large N-glycosylated extracellular domain, and two membrane-spanning domains that each pass once through the plasma membrane. Because of their sequence similarity, it is likely that this structure is shared by other ENaC subunits and possibly the degenerins of Caenorhabditis elegans as well.
Notes: Snyder, P M
McDonald, F J
Stokes, J B
Welsh, M J
HL07344/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1994 Sep 30;269(39):24379-83.
Author Address: Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1282
Author: Sobolevsky, A. I., Rosconi, M. P. and Gouaux, E.
Year: 2009
Title: X-ray structure, symmetry and mechanism of an AMPA-subtype glutamate receptor
Journal: Nature
Volume: 462
Issue: 7274
Pages: 745-56
Epub Date: 2009/12/01
Date: Dec 10
Short Title: X-ray structure, symmetry and mechanism of an AMPA-subtype glutamate receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature08624
PMCID: PMC2861655
NIHMSID: NIHMS165986
Accession Number: 19946266
Keywords: Animals
Cell Line
Crystallization
Crystallography, X-Ray
Ion Channel Gating
Models, Molecular
Potassium Channels/chemistry/metabolism
Protein Conformation
Protein Subunits/chemistry/metabolism
Rats
Receptors, AMPA/antagonists & inhibitors/*chemistry/*metabolism
alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid/metabolism
Abstract: Ionotropic glutamate receptors mediate most excitatory neurotransmission in the central nervous system and function by opening a transmembrane ion channel upon binding of glutamate. Despite their crucial role in neurobiology, the architecture and atomic structure of an intact ionotropic glutamate receptor are unknown. Here we report the crystal structure of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)-sensitive, homotetrameric, rat GluA2 receptor at 3.6 A resolution in complex with a competitive antagonist. The receptor harbours an overall axis of two-fold symmetry with the extracellular domains organized as pairs of local dimers and with the ion channel domain exhibiting four-fold symmetry. A symmetry mismatch between the extracellular and ion channel domains is mediated by two pairs of conformationally distinct subunits, A/C and B/D. Therefore, the stereochemical manner in which the A/C subunits are coupled to the ion channel gate is different from the B/D subunits. Guided by the GluA2 structure and site-directed cysteine mutagenesis, we suggest that GluN1 and GluN2A NMDA (N-methyl-d-aspartate) receptors have a similar architecture, with subunits arranged in a 1-2-1-2 pattern. We exploit the GluA2 structure to develop mechanisms of ion channel activation, desensitization and inhibition by non-competitive antagonists and pore blockers.
Notes: 1476-4687
Sobolevsky, Alexander I
Rosconi, Michael P
Gouaux, Eric
F32 NS049767-05/NS/NINDS NIH HHS/United States
R01 NS038631/NS/NINDS NIH HHS/United States
R01 NS038631-06/NS/NINDS NIH HHS/United States
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2009 Dec 10;462(7274):745-56. doi: 10.1038/nature08624.
Author Address: Vollum Institute, Oregon Health and Science University, 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1190
Author: Soesanto, Y. A., Luo, B., Jones, D., Taylor, R., Gabrielsen, J. S., Parker, G. and McClain, D. A.
Year: 2008
Title: Regulation of Akt signaling by O-GlcNAc in euglycemia
Journal: Am J Physiol Endocrinol Metab
Volume: 295
Issue: 4
Pages: E974-80
Epub Date: 2008/08/30
Date: Oct
Short Title: Regulation of Akt signaling by O-GlcNAc in euglycemia
Alternate Journal: American journal of physiology. Endocrinology and metabolism
ISSN: 0193-1849 (Print)
0193-1849
DOI: 10.1152/ajpendo.90366.2008
PMCID: PMC2575895
Accession Number: 18728220
Keywords: Adenoviridae/genetics
Animals
Blood Glucose/*physiology
Cell Line
Genetic Vectors/physiology
Liver/metabolism
Mice
Mice, Inbred C57BL
N-Acetylglucosaminyltransferases/*physiology
Oncogene Protein v-akt/*physiology
Phosphatidylinositol 3-Kinases/metabolism
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction/*physiology
Wheat Germ Agglutinins
Abstract: The hexosamine biosynthesis pathway (HBP) regulates the posttranslational modification of nuclear and cytoplasmic protein by O-linked N-acetylglucosamine (O-GlcNAc). Numerous studies have demonstrated that, in hyperglycemic conditions, excessive glucose flux through this pathway contributes to the development of insulin resistance. The role of the HBP in euglycemia, however, remains largely unknown. Here we investigated the effect of O-GlcNAc on hepatic Akt signaling at physiological concentrations of glucose. In HepG2 cells cultured in 5 mM glucose, removal of O-GlcNAc by adenoviral-mediated overexpression of O-GlcNAcase increased Akt activity and phosphorylation. We also observed that Akt was recognized by succinylated wheat germ agglutinin (sWGA), which specifically binds O-GlcNAc. Overexpression of O-GlcNAcase in HepG2 cells reduced the levels of Akt in sWGA precipitates. The increased Akt activity was accompanied by increased phosphorylation of Akt substrates and reduced mRNA for glucose-6-phosphatase and phosphoenolpyruvate carboxykinase (PEPCK). The increased Akt activity was not a result of activation of its upstream activator phosphoinositide 3-kinase (PI 3-kinase). Further demonstrating Akt regulation by O-GlcNAc, we found that overexpression of O-GlcNAcase in the livers of euglycemic mice also significantly increased Akt activity, resulting in increased phosphorylation of downstream targets and decreased mRNA for glucose-6-phosphatase. Together, these data suggest that O-GlcNAc regulates Akt signaling in hepatic models under euglycemic conditions.
Notes: 1522-1555
Soesanto, Yudi A
Luo, Bai
Jones, Deborah
Taylor, Rodrick
Gabrielsen, J Scott
Parker, Glendon
McClain, Donald A
5-T32-DK-007115/DK/NIDDK NIH HHS/United States
R01-DK-43526/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Am J Physiol Endocrinol Metab. 2008 Oct;295(4):E974-80. doi: 10.1152/ajpendo.90366.2008. Epub 2008 Aug 26.
Author Address: Division of Endocrinology, University of Utah School of Medicine, Salt Lake City, Utah 84132, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1642
Author: Sohn, K. C., Lee, E. J., Shin, J. M., Lim, E. H., No, Y., Lee, J. Y., Yoon, T. Y., Lee, Y. H., Im, M., Lee, Y., Seo, Y. J., Lee, J. H. and Kim, C. D.
Year: 2014
Title: Regulation of keratinocyte differentiation by O-GlcNAcylation
Journal: J Dermatol Sci
Volume: 75
Issue: 1
Pages: 10-5
Epub Date: 2014/05/08
Date: Jul
Short Title: Regulation of keratinocyte differentiation by O-GlcNAcylation
Alternate Journal: Journal of dermatological science
ISSN: 0923-1811
DOI: 10.1016/j.jdermsci.2014.04.010
Accession Number: 24802710
Keywords: Acetylglucosamine/*metabolism
Calcium/metabolism
*Cell Differentiation
Cells, Cultured
Glycosylation
Humans
Keratinocytes/*metabolism
Membrane Proteins/genetics/metabolism
Promoter Regions, Genetic
Protein Precursors/genetics/metabolism
Protein Processing, Post-Translational
Sp1 Transcription Factor/genetics/metabolism
Time Factors
Transcription, Genetic
Transfection
beta-N-Acetylhexosaminidases/genetics/metabolism
Keratinocyte differentiation
O-GlcNAcylation
Sp-1
Abstract: BACKGROUND: O-linked β-N-acetylglucosamine (O-GlcNAc) modification is one of the posttranslational modification, emerging as an important regulatory mechanism in various cellular events. OBJECTIVE: We attempted to investigate whether O-GlcNAcylation is involved in keratinocyte differentiation. METHODS: Immunohistochemistry and Western blot were performed to demonstrate O-GlcNAcylation in keratinocyte differentiation. RESULTS: During calcium-induced keratinocyte differentiation, overall O-GlcNAcylation was decreased in a temporal manner. We focused our attention on transcription factor Sp-1, which is implicated in keratinocyte differentiation. Total Sp-1 level did not change during keratinocyte differentiation. However, O-GlcNAcylated Sp-1 was decreased in a keratinocyte differentiation-dependent manner. Interestingly, transcriptional activity of Sp-1, in terms of involucrin and loricrin promoter activities, was markedly increased by overexpression of O-GlcNAcase (OGA). In addition, membrane permeable non-O-GlcNAcylated Sp-1 did show transcriptional activity, while membrane permeable O-GlcNAcylated Sp-1 did not, suggesting O-GlcNAcylated Sp-1 is an inactive form in keratinocyte differentiation. CONCLUSION: Our results reveal that O-GlcNAcylation is a dynamic regulatory mechanism for keratinocyte differentiation.
Notes: 1873-569x
Sohn, Kyung-Cheol
Lee, Eun Jin
Shin, Jung-Min
Lim, Eun-Hwa
No, Yoonoo
Lee, Ji Yeoun
Yoon, Tae Young
Lee, Young Ho
Im, Myung
Lee, Young
Seo, Young-Joon
Lee, Jeung-Hoon
Kim, Chang Deok
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Dermatol Sci. 2014 Jul;75(1):10-5. doi: 10.1016/j.jdermsci.2014.04.010. Epub 2014 Apr 30.
Author Address: Department of Dermatology and Research Institute for Medical Sciences, School of Medicine, Chungnam National University, Daejeon, Republic of Korea.
Department of Dermatology, School of Medicine and Medical Research Institute, Chungbuk National University, Cheongju, Republic of Korea.
Department of Anatomy, School of Medicine, Chungnam National University, Daejeon, Republic of Korea.
Department of Dermatology and Research Institute for Medical Sciences, School of Medicine, Chungnam National University, Daejeon, Republic of Korea. Electronic address: cdkimd@gmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1319
Author: Soisson, S. M., Patel, S. B., Abeywickrema, P. D., Byrne, N. J., Diehl, R. E., Hall, D. L., Ford, R. E., Reid, J. C., Rickert, K. W., Shipman, J. M., Sharma, S. and Lumb, K. J.
Year: 2010
Title: Structural definition and substrate specificity of the S28 protease family: the crystal structure of human prolylcarboxypeptidase
Journal: BMC Struct Biol
Volume: 10
Pages: 16
Epub Date: 2010/06/15
Date: Jun 11
Short Title: Structural definition and substrate specificity of the S28 protease family: the crystal structure of human prolylcarboxypeptidase
Alternate Journal: BMC structural biology
ISSN: 1472-6807
DOI: 10.1186/1472-6807-10-16
PMCID: PMC2893456
Accession Number: 20540760
Keywords: Amino Acid Sequence
Binding Sites
Carboxypeptidases/*chemistry/genetics/metabolism
Catalytic Domain
Crystallography, X-Ray
Humans
Molecular Sequence Data
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Substrate Specificity
Abstract: BACKGROUND: The unique S28 family of proteases is comprised of the carboxypeptidase PRCP and the aminopeptidase DPP7. The structural basis of the different substrate specificities of the two enzymes is not understood nor has the structure of the S28 fold been described. RESULTS: The experimentally phased 2.8 A crystal structure is presented for human PRCP. PRCP contains an alpha/beta hydrolase domain harboring the catalytic Asp-His-Ser triad and a novel helical structural domain that caps the active site. Structural comparisons with prolylendopeptidase and DPP4 identify the S1 proline binding site of PRCP. A structure-based alignment with the previously undescribed structure of DPP7 illuminates the mechanism of orthogonal substrate specificity of PRCP and DPP7. PRCP has an extended active-site cleft that can accommodate proline substrates with multiple N-terminal residues. In contrast, the substrate binding groove of DPP7 is occluded by a short amino-acid insertion unique to DPP7 that creates a truncated active site selective for dipeptidyl proteolysis of N-terminal substrates. CONCLUSION: The results define the structure of the S28 family of proteases, provide the structural basis of PRCP and DPP7 substrate specificity and enable the rational design of selective PRCP modulators.
Notes: 1472-6807
Soisson, Stephen M
Patel, Sangita B
Abeywickrema, Pravien D
Byrne, Noel J
Diehl, Ronald E
Hall, Dawn L
Ford, Rachael E
Reid, John C
Rickert, Keith W
Shipman, Jennifer M
Sharma, Sujata
Lumb, Kevin J
Journal Article
BMC Struct Biol. 2010 Jun 11;10:16. doi: 10.1186/1472-6807-10-16.
Author Address: Global Structural Biology, Merck Research Laboratories, P,O, Box 4, West Point, PA 19486, USA. stephen_soisson@merck.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 778
Author: Somers, W. S., Tang, J., Shaw, G. D. and Camphausen, R. T.
Year: 2000
Title: Insights into the molecular basis of leukocyte tethering and rolling revealed by structures of P- and E-selectin bound to SLe(X) and PSGL-1
Journal: Cell
Volume: 103
Issue: 3
Pages: 467-79
Epub Date: 2000/11/18
Date: Oct 27
Short Title: Insights into the molecular basis of leukocyte tethering and rolling revealed by structures of P- and E-selectin bound to SLe(X) and PSGL-1
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/s0092-8674(00)00138-0
Accession Number: 11081633
Keywords: Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
E-Selectin/*chemistry/*metabolism
Epidermal Growth Factor/chemistry
Humans
Lectins/chemistry
Leukocytes/chemistry/*metabolism
Membrane Glycoproteins/chemistry/*metabolism
Models, Molecular
Molecular Sequence Data
Oligosaccharides/*metabolism
P-Selectin/*chemistry/*metabolism
Peptide Fragments/chemistry/metabolism
Protein Binding
Protein Structure, Tertiary
Static Electricity
Structure-Activity Relationship
Sulfur/metabolism
Tyrosine/metabolism
Abstract: P-, E- and L-selectin constitute a family of cell adhesion receptors that mediate the initial tethering and rolling of leukocytes on inflamed endothelium as a prelude to their firm attachment and extravasation into tissues. The selectins bind weakly to sialyl Lewisx (SLe(X))-like glycans, but with high-affinity to specific glycoprotein counterreceptors, including PSGL-1. Here, we report crystal structures of human P- and E-selectin constructs containing the lectin and EGF (LE) domains co-complexed with SLe(X). We also present the crystal structure of P-selectin LE co-complexed with the N-terminal domain of human PSGL-1 modified by both tyrosine sulfation and SLe(X). These structures reveal differences in how E- and P-selectin bind SLe(X) and the molecular basis of the high-affinity interaction between P-selectin and PSGL-1.
Notes: Somers, W S
Tang, J
Shaw, G D
Camphausen, R T
Journal Article
United States
Cell. 2000 Oct 27;103(3):467-79. doi: 10.1016/s0092-8674(00)00138-0.
Author Address: Genetics Institute, Wyeth Research, Cambridge, Massachusetts 02140, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1666
Author: Song, E., Mayampurath, A., Yu, C. Y., Tang, H. and Mechref, Y.
Year: 2014
Title: Glycoproteomics: identifying the glycosylation of prostate specific antigen at normal and high isoelectric points by LC-MS/MS
Journal: J Proteome Res
Volume: 13
Issue: 12
Pages: 5570-80
Epub Date: 2014/10/21
Date: Dec 5
Short Title: Glycoproteomics: identifying the glycosylation of prostate specific antigen at normal and high isoelectric points by LC-MS/MS
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr500575r
PMCID: PMC4261947
Accession Number: 25327667
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Gel
Glycopeptides/chemistry
Glycosylation
Humans
Isoelectric Point
Kallikreins/*metabolism
Molecular Sequence Data
Prostate-Specific Antigen/*metabolism
*Protein Processing, Post-Translational
Proteome/metabolism
Proteomics
Tandem Mass Spectrometry
Lc−ms/ms
N-linked glycosylation
Psa
Prostate specific antigen
glycopeptide
glycoproteomics
Abstract: Prostate specific antigen (PSA) is currently used as a biomarker to diagnose prostate cancer. PSA testing has been widely used to detect and screen prostate cancer. However, in the diagnostic gray zone, the PSA test does not clearly distinguish between benign prostate hypertrophy and prostate cancer due to their overlap. To develop more specific and sensitive candidate biomarkers for prostate cancer, an in-depth understanding of the biochemical characteristics of PSA (such as glycosylation) is needed. PSA has a single glycosylation site at Asn69, with glycans constituting approximately 8% of the protein by weight. Here, we report the comprehensive identification and quantitation of N-glycans from two PSA isoforms using LC-MS/MS. There were 56 N-glycans associated with PSA, whereas 57 N-glycans were observed in the case of the PSA-high isoelectric point (pI) isoform (PSAH). Three sulfated/phosphorylated glycopeptides were detected, the identification of which was supported by tandem MS data. One of these sulfated/phosphorylated N-glycans, HexNAc5Hex4dHex1s/p1 was identified in both PSA and PSAH at relative intensities of 0.52 and 0.28%, respectively. Quantitatively, the variations were monitored between these two isoforms. Because we were one of the laboratories participating in the 2012 ABRF Glycoprotein Research Group (gPRG) study, those results were compared to that presented in this study. Our qualitative and quantitative results summarized here were comparable to those that were summarized in the interlaboratory study.
Notes: 1535-3907
Song, Ehwang
Mayampurath, Anoop
Yu, Chuan-Yih
Tang, Haixu
Mechref, Yehia
R01 GM093322/GM/NIGMS NIH HHS/United States
R01 GM112490/GM/NIGMS NIH HHS/United States
1R01GM093322-04/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Proteome Res. 2014 Dec 5;13(12):5570-80. doi: 10.1021/pr500575r. Epub 2014 Nov 10.
Author Address: Department of Chemistry and Biochemistry, Texas Tech University , Lubbock, Texas 79409, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 978
Author: Song, G., Yang, Y., Liu, J. H., Casasnovas, J. M., Shimaoka, M., Springer, T. A. and Wang, J. H.
Year: 2005
Title: An atomic resolution view of ICAM recognition in a complex between the binding domains of ICAM-3 and integrin alphaLbeta2
Journal: Proc Natl Acad Sci U S A
Volume: 102
Issue: 9
Pages: 3366-71
Epub Date: 2005/02/25
Date: Mar 1
Short Title: An atomic resolution view of ICAM recognition in a complex between the binding domains of ICAM-3 and integrin alphaLbeta2
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0500200102
PMCID: PMC552929
Accession Number: 15728350
Keywords: Amino Acid Sequence
Antigens, CD/chemistry/*metabolism
Binding Sites
Cell Adhesion Molecules
Lymphocyte Function-Associated Antigen-1/chemistry/*metabolism
Models, Molecular
Molecular Sequence Data
Sequence Homology, Amino Acid
Surface Plasmon Resonance
Abstract: Within the Ig superfamily (IgSF), intercellular adhesion molecules (ICAMs) form a subfamily that binds the leukocyte integrin alphaLbeta2. We report a 1.65-A-resolution crystal structure of the ICAM-3 N-terminal domain (D1) in complex with the inserted domain, the ligand-binding domain of alphaLbeta2. This high-resolution structure and comparisons among ICAM subfamily members establish that the binding of ICAM-3 D1 onto the inserted domain represents a common docking mode for ICAM subfamily members. The markedly different off-rates of ICAM-1, -2, and -3 appear to be determined by the hydrophobicity of residues that surround a metal coordination bond in the alphaLbeta2-binding interfaces. Variation in composition of glycans on the periphery of the interfaces influences on-rate.
Notes: 1091-6490
Song, Gang
Yang, Yuting
Liu, Jin-Huan
Casasnovas, Jose M
Shimaoka, Motomu
Springer, Timothy A
Wang, Jia-Huai
HL48675/HL/NHLBI NIH HHS/United States
CA31798/CA/NCI NIH HHS/United States
P01 HL048675/HL/NHLBI NIH HHS/United States
R01 CA031798/CA/NCI NIH HHS/United States
R37 CA031798/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2005 Mar 1;102(9):3366-71. doi: 10.1073/pnas.0500200102. Epub 2005 Feb 22.
Author Address: CBR Institute for Biomedical Research, Department of Pathology, Harvard Medical School, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2042
Author: Song, H., Ma, J., Bian, Z., Chen, S., Zhu, J., Wang, J., Huang, N., Yin, M., Sun, F., Xu, M. and Pan, Q.
Year: 2019
Title: Global profiling of O-GlcNAcylated and/or phosphorylated proteins in hepatoblastoma
Journal: Signal Transduct Target Ther
Volume: 4
Pages: 40
Epub Date: 2019/10/23
Short Title: Global profiling of O-GlcNAcylated and/or phosphorylated proteins in hepatoblastoma
Alternate Journal: Signal transduction and targeted therapy
ISSN: 2095-9907 (Print)
2059-3635
DOI: 10.1038/s41392-019-0067-4
PMCID: PMC6799812
Accession Number: 31637018
Keywords: *Epigenetics
*Molecular medicine
Abstract: O-linked-β-N-acetylglucosamine (O-GlcNAc) glycosylation (O-GlcNAcylation) and phosphorylation are critical posttranslational modifications that are involved in regulating the functions of proteins involved in tumorigenesis and the development of various solid tumors. However, a detailed characterization of the patterns of these modifications at the peptide or protein level in hepatoblastoma (HB), a highly malignant primary hepatic tumor with an extremely low incidence in children, has not been performed. Here, we examined O-GlcNAc-modified or phospho-modified peptides and proteins in HB through quantitative proteomic analysis of HB tissues and paired normal liver tissues. Our results identified 114 O-GlcNAcylated peptides belonging to 78 proteins and 3494 phosphorylated peptides in 2088 proteins. Interestingly, 41 proteins were modified by both O-GlcNAcylation and phosphorylation. These proteins are involved in multiple molecular and cellular processes, including chromatin remodeling, transcription, translation, transportation, and organelle organization. In addition, we verified the accuracy of the proteomics results and found a competitive inhibitory effect between O-GlcNAcylation and phosphorylation of HSPB1. Further, O-GlcNAcylation modification of HSPB1 promoted proliferation and enhanced the chemotherapeutic resistance of HB cell lines in vitro. Collectively, our research suggests that O-GlcNAc-modified and/or phospho-modified proteins may play a crucial role in the pathogenesis of HB.
Notes: 2059-3635
Song, Hang
Ma, Ji
Bian, Zhixuan
Chen, Shuhua
Zhu, Jiabei
Wang, Jing
Huang, Nan
Yin, Minzhi
Sun, Fenyong
Xu, Min
Pan, Qiuhui
Journal Article
Research Support, Non-U.S. Gov't
Signal Transduct Target Ther. 2019 Oct 11;4:40. doi: 10.1038/s41392-019-0067-4. eCollection 2019.
Author Address: 1Department of Laboratory Medicine, Shanghai Children's Medical Center, School of Medicine, Shanghai Jiaotong University, 200127 Shanghai, China.
Department of Laboratory Medicine, Yunfu People's Hospital, 527300 Guangdong, China.
3Department of Surgery, Shanghai Children's Medical Center, School of Medicine, Shanghai Jiaotong University, 200127 Shanghai, China.
4Department of Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, 200072 Shanghai, China.
5Department of Pathology, Shanghai Children's Medical Center, School of Medicine, Shanghai Jiaotong University, 200127 Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1297
Author: Songsrirote, K., Li, Z., Ashford, D., Bateman, A. and Thomas-Oates, J.
Year: 2010
Title: Development and application of mass spectrometric methods for the analysis of progranulin N-glycosylation
Journal: J Proteomics
Volume: 73
Issue: 8
Pages: 1479-90
Epub Date: 2010/03/02
Date: Jun 16
Short Title: Development and application of mass spectrometric methods for the analysis of progranulin N-glycosylation
Alternate Journal: Journal of proteomics
ISSN: 1874-3919
DOI: 10.1016/j.jprot.2010.02.013
Accession Number: 20188224
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Glycosylation
Humans
Intercellular Signaling Peptides and Proteins/chemistry/*metabolism
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase/metabolism
Mass Spectrometry/*methods
Molecular Sequence Data
Polysaccharides/chemistry/isolation & purification
Progranulins
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods
Abstract: PGRN is a modular protein with 7 1/2 repeats of the granulin domain separated by short spacer sequences. Elevated expression of PGRN is associated with cancer growth, while mutations of PGRN cause frontotemporal lobar degeneration (FTLD), an early onset form of dementia. PGRN is a glycoprotein, containing five N-glycosylation consensus sequons, three of which fall within granulin domains. A method tailored to enable detailed analysis of the PGRN oligosaccharides and glycopeptides has been developed. The approach involves in-gel deglycosylation using peptide-N-glycosidase F (PNGase F) followed by permethylation of the released oligosaccharides. Permethylation was applied for rapid sample clean-up and to improve sensitivity of MS detection and mass spectrometric fragmentation. Reversed-phase monolithic LC-ESI-MS/MS was used for analysis of permethylated oligosaccharides, enabling structural characterization of released N-linked glycans in one chromatographic run. In-gel tryptic digestion was further applied to the gel pieces containing deglycosylated protein, for N-glycosylation site determination. In addition, glycopeptides were produced using in-solution pronase digestion to identify species of N-glycan attached at particular sites. The method developed was applied to progranulin (PGRN) to characterize the structures of the released glycans and to identify the sites of glycosylation. Glycosylation of four out of five potential PGRN N-glycosylation consensus sites was demonstrated (the final one remains undetermined), with one of the four observed to be partially occupied. Two of the observed glycosylation sites occur within granulin domains, which may have important implications for understanding the structural basis of PGRN action.
Notes: 1876-7737
Songsrirote, Kriangsak
Li, Zhi
Ashford, David
Bateman, Andrew
Thomas-Oates, Jane
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Proteomics. 2010 Jun 16;73(8):1479-90. doi: 10.1016/j.jprot.2010.02.013. Epub 2010 Feb 24.
Author Address: Department of Chemistry, University of York, York, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1650
Author: Sorvillo, N., Kaijen, P. H., Matsumoto, M., Fujimura, Y., van der Zwaan, C., Verbij, F. C., Pos, W., Fijnheer, R., Voorberg, J. and Meijer, A. B.
Year: 2014
Title: Identification of N-linked glycosylation and putative O-fucosylation, C-mannosylation sites in plasma derived ADAMTS13
Journal: J Thromb Haemost
Volume: 12
Issue: 5
Pages: 670-9
Epub Date: 2014/07/01
Date: May
Short Title: Identification of N-linked glycosylation and putative O-fucosylation, C-mannosylation sites in plasma derived ADAMTS13
Alternate Journal: Journal of thrombosis and haemostasis : JTH
ISSN: 1538-7836
DOI: 10.1111/jth.12535
Accession Number: 24977290
Keywords: ADAM Proteins/*blood/genetics
ADAMTS13 Protein
Amino Acid Sequence
Autoimmune Diseases/immunology
Fucose/chemistry
Glycosylation
HEK293 Cells
Hexoses/chemistry
Humans
Mannose/chemistry
Molecular Sequence Data
Protein Structure, Tertiary
Purpura, Thrombotic Thrombocytopenic/*blood/genetics
Sequence Homology, Amino Acid
Tandem Mass Spectrometry
Thrombospondins/blood
Abstract: BACKGROUND: Acquired deficiency of ADAMTS13 causes a rare and life-threatening disorder called thrombotic thrombocytopenic purpura (TTP). Several studies have shown that aberrant glycosylation can play an important role in the pathogenesis of autoimmune diseases.N-linked glycosylation and putative O-fucosylation sites have been predicted or identified in recombinant ADAMTS13. However, it is not known which of these sites are glycosylated in plasma derived ADAMTS13. OBJECTIVES: Here we investigated the presence of putative O-fucosylation, C-mannosylation and N-linked glycosylation sites on plasma derived ADAMTS13. METHODS/RESULTS: Sites of N-linked glycosylation were determined by the use of peptide N-glycosidase-F (PNGase F), which removes the entire carbohydrate from the side chain of asparagines. Nine of the 10 predicted N-linked glycosylation sites were identified in or near the metalloproteinase,spacer, thrombospondin type 1 repeat (TSR1) and the CUB domain of plasma ADAMTS13. Moreover, six putative O-fucosylated sites were identified in the TSR domains of plasma ADAMTS13 by performing searches of the tandem mass spectrometry (MS/MS) data for loss of hexose (162 Da), deoxyhexose (146 Da), or hexose deoxyhexose(308 Da). The use of electron transfer dissociation (ETD) allowed for unambiguous identification of the modified sites. In addition to putative O-fucosylation and N-linked glycosylation, two putative C-mannosylation sites were identified within the TSR1 and TSR4 domains of ADAMTS13. CONCLUSIONS: Our data identify several glycosylation sites on plasma derived ADAMTS13. We anticipate that our findings may be relevant for the initiation of autoimmune reactivity against ADAMTS13 in patients with acquired TTP.
Notes: 1538-7836
Sorvillo, N
Kaijen, P H
Matsumoto, M
Fujimura, Y
van der Zwaan, C
Verbij, F C
Pos, W
Fijnheer, R
Voorberg, J
Meijer, A B
Journal Article
Research Support, Non-U.S. Gov't
England
J Thromb Haemost. 2014 May;12(5):670-9. doi: 10.1111/jth.12535.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 454
Author: Sottrup-Jensen, L., Stepanik, T. M., Kristensen, T., Wierzbicki, D. M., Jones, C. M., Lønblad, P. B., Magnusson, S. and Petersen, T. E.
Year: 1984
Title: Primary structure of human alpha 2-macroglobulin. V. The complete structure
Journal: J Biol Chem
Volume: 259
Issue: 13
Pages: 8318-27
Epub Date: 1984/07/10
Date: Jul 10
Short Title: Primary structure of human alpha 2-macroglobulin. V. The complete structure
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6203908
Keywords: Amino Acid Sequence
Animals
Chickens
Cyanogen Bromide
Humans
Macromolecular Substances
Mice
Molecular Weight
Rats
Species Specificity
*alpha-Macroglobulins
Abstract: The primary structure of the tetrameric plasma glycoprotein human alpha 2-macroglobulin has been determined. The identical subunits contain 1451 amino acid residues. Glucosamine-based oligosaccharide groups are attached to asparagine residues 32, 47, 224, 373, 387, 846, 968, and 1401. Eleven intrachain disulfide bridges have been placed (Cys25-Cys63, Cys228-Cys276, Cys246-Cys264, Cys255-Cys408, Cys572-Cys748, Cys619-Cys666, Cys798-Cys826, Cys824-Cys860, Cys898-Cys1298, Cys1056-Cys1104, and Cys1329-Cys1444). Cys-447 probably forms an interchain bridge with Cys-447 from another subunit. The beta-SH group of Cys-949 is thiol esterified to the gamma-carbonyl group of Glx-952, thus forming an activatable reactive site which can mediate covalent binding of nucleophiles. A putative transglutaminase cross-linking site is constituted by Gln-670 and Gln-671. The primary sites of proteolytic cleavage in the activation cleavage area (the "bait" region) are located in the sequence: -Arg681-Val-Gly-Phe-Tyr-Glu-. The molecular weight of the unmodified alpha 2-macroglobulin subunit is 160,837 and approximately 179,000, including the carbohydrate groups. The presence of possible internal homologies within the alpha 2-macroglobulin subunit is discussed. A comparison of stretches of sequences from alpha 2-macroglobulin with partial sequence data for complement components C3 and C4 indicates that these proteins are evolutionary related. The properties of alpha 2-macroglobulin are discussed within the context of proteolytically regulated systems with particular reference to the complement components C3 and C4.
Notes: Sottrup-Jensen, L
Stepanik, T M
Kristensen, T
Wierzbicki, D M
Jones, C M
Lønblad, P B
Magnusson, S
Petersen, T E
HL 16238/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1984 Jul 10;259(13):8318-27.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 301
Author: Spellman, M. W., Leonard, C. K., Basa, L. J., Gelineo, I. and van Halbeek, H.
Year: 1991
Title: Carbohydrate structures of recombinant soluble human CD4 expressed in Chinese hamster ovary cells
Journal: Biochemistry
Volume: 30
Issue: 9
Pages: 2395-406
Epub Date: 1991/03/05
Date: Mar 5
Short Title: Carbohydrate structures of recombinant soluble human CD4 expressed in Chinese hamster ovary cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00223a015
Accession Number: 2001369
Keywords: Amidohydrolases
Amino Acid Sequence
Animals
*CD4 Antigens/genetics
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Cricetulus
Female
Glycopeptides/isolation & purification
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Ovary
Peptide Mapping
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Recombinant Proteins/chemistry
Transfection
Trypsin
Abstract: Infection of T-lymphocytes and macrophages by human immunodeficiency virus (HIV) is mediated by the binding of the HIV envelope glycoprotein to the cell-surface receptor glycoprotein CD4. A soluble, recombinant CD4 molecule (rCD4), produced by expression of a truncated CD4 gene in Chinese hamster ovary (CHO) cells [Smith et al. (1987) Science 238, 1704-1707], is in clinical trials as a potential therapeutic agent in the treatment of acquired immunodeficiency syndrome (AIDS). In the present study, the structures of the Asn-linked oligosaccharides of soluble rCD4 have been elucidated. The rCD4 molecule has two potential sites for N-glycosylation, Asn-271 and Asn-300. Tryptic glycopeptides containing either of the sites were purified by reversed-phase HPLC, and their oligosaccharides were released enzymatically. The structures of the oligosaccharides were determined by methylation analysis, high-pH anion-exchange chromatography, fast-atom bombardment mass spectrometry, and 1H NMR spectroscopy at 500 MHz. Asn-271 was found to carry diantennary N-acetyllactosamine-type ("complex") oligosaccharides, of which 8% were asialo, 55% were monosialyl, and 37% were disialyl. Approximately 18% of these structures contained fucose alpha(1-->6) linked to the reducing GlcNAc residue. Two different hybrid structures were found to account for 34% of the oligosaccharides attached to Asn-300. The remainder of the oligosaccharides attached to Asn-300 were diantennary N-acetyllactosamine-type, of which 10% were asialo, 61% were monosialyl, and 29% were disialyl. Approximately 9% of the hybrid structures and 40% of the N-acetyllactosamine structures at Asn-300 were found to contain fucose alpha(1-->6) linked to the innermost GlcNAc residue.
Notes: Spellman, M W
Leonard, C K
Basa, L J
Gelineo, I
van Halbeek, H
P01-AI-27135/AI/NIAID NIH HHS/United States
P41-RR-05351/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1991 Mar 5;30(9):2395-406. doi: 10.1021/bi00223a015.
Author Address: Department of Medicinal and Analytical Chemistry, Genentech, Inc., South San Francisco, California 94080.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 299
Author: Spik, G., Coddeville, B., Strecker, G., Montreuil, J., Regoeczi, E., Chindemi, P. A. and Rudolph, J. R.
Year: 1991
Title: Carbohydrate microheterogeneity of rat serotransferrin. Determination of glycan primary structures and characterization of a new type of trisialylated diantennary glycan
Journal: Eur J Biochem
Volume: 195
Issue: 2
Pages: 397-405
Epub Date: 1991/01/30
Date: Jan 30
Short Title: Carbohydrate microheterogeneity of rat serotransferrin. Determination of glycan primary structures and characterization of a new type of trisialylated diantennary glycan
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1991.tb15719.x
Accession Number: 1997323
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, DEAE-Cellulose
Concanavalin A/metabolism
Glycopeptides/analysis
Hydrogen
Magnetic Resonance Spectroscopy
Methylation
Molecular Sequence Data
Polysaccharides/*chemistry
Rats
Transferrin/*chemistry/metabolism
Abstract: A previously established procedure [Regoeczi, E., Chindemi, P.A., Rudolph, J. R., Spik, G. & Montreuil, J. (1987) Biochem. Cell Biol. 65, 948-954] was used to isolate from three DEAE-cellulose chromatographic fractions of diferric rat serotransferrin (rTf) subpopulations having discernible affinities for concanavalin A (ConA). These entities are designated rTf-1 (not retarded by ConA column), rTf-2 (retarded) and rTf-3 (bound). Each rTf type was found to be endowed with carbohydrate sufficient to account for a single diantennary glycan/protein molecule. Glycan structures were determined on the glycopeptides by employing GLC/MS and 400-MHz 1H-NMR spectroscopy. All glycans possessed a common, trimannosyl-N,N'-diacetylchitobiose core with or without one L-fucose alpha-1,6-linked to the Asn-linked GlcNAc. However, there were differences in the antennae. Thus, in rTf-3, both antennae were of the disialylated diantennary N-acetyllactosamine type which is frequently encountered in other plasma glycoproteins. However, the alpha-1,3-Man-linked antenna in rTf-1 as well as rTf-2 had the sequence: Neu5Ac(alpha 2-3)Gal(beta 1-3)[Neu5Ac(alpha 2-6)]GlcNAc(beta 1-2)Man. In addition, the alpha-1,6-Man-linked antenna deviated in rTf-2 from the standard structure by having the sequence: Neu5Ac(alpha 2-3)Gal(beta 1-3)GlcNAc(beta 1-2)Man. The possible relevance of the above structures to the ConA binding of rTf is discussed. A further preparation, obtained from the most anionic DEAE-cellulose fraction (peak V) or rTf contained several tetrasialylated diantennary glycans whose precise structures remain to be established in future studies.
Notes: Spik, G
Coddeville, B
Strecker, G
Montreuil, J
Regoeczi, E
Chindemi, P A
Rudolph, J R
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1991 Jan 30;195(2):397-405. doi: 10.1111/j.1432-1033.1991.tb15719.x.
Author Address: Laboratoire de Chimie Biologique, Université des Sciences et Techniques de Lille Flandres-Artois, Villeneuve d'Ascq, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 484
Author: Spik, G., Strecker, G., Fournet, B., Bouquelet, S., Montreuil, J., Dorland, L., van Halbeek, H. and Vliegenthart, J. F.
Year: 1982
Title: Primary structure of the glycans from human lactotransferrin
Journal: Eur J Biochem
Volume: 121
Issue: 2
Pages: 413-9
Epub Date: 1982/01/01
Date: Jan
Short Title: Primary structure of the glycans from human lactotransferrin
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1982.tb05803.x
Accession Number: 7060557
Keywords: Chemical Phenomena
Chemistry
Glycopeptides/isolation & purification
Humans
Lactoferrin/*analysis
Lactoglobulins/*analysis
Magnetic Resonance Spectroscopy
Polysaccharides/*isolation & purification
Abstract: The polypeptide chain of human lactotransferrin possesses two glycoslyation sites to which glycans are linked through an N-(beta-aspartyl)-N-acetylglucosaminylamine bond and which are structurally heterogenous. After chymotryptic or pronase digestions, glycopeptides with five different glycan structures could be isolated. For three of them, the structure has been determined by the application of methanolysis, methylation analysis, hydrazinolysis/nitrous deamination, enzymatic cleavage and 1H-NMR spectroscopy at 360 MHz: Glycopeptides A and B: NeuAc(alpha 2-6)Gal(beta 1-4)GlcNAc(beta 1-2)Man(alpha 1-3)[NeuAc(alpha 2-6)Gal(beta 1-4)GlcNAc(beta 1-2)Man(alpha 1-6)]Man(beta 1-4)GlcNAc(beta-1-4)[Fuc(alpha 1-6)]GlcNAc(beta 1-)Asn; Glycopeptide C: NeuAc(alpha 2-6)(Gal(beta 1-4)GlcNAc(beta 1-2)Man(alpha 1-3)(Gal(beta 1-4)[Fuc(alpha 1-3)]GlcNAc(beta 1-2)Man(alpha 1-6))Man(beta 1-4)GlcNAc(beta 1-4)[Fuc(alpha 1-6)]GlcNAc(beta 1-)Asn. Glycopeptide D: NeuAc(alpha 2-6)Gal(beta 1-4)GlcNAc(beta 1-2)Man(alpha 1-3)[Gal(beta 1-4)GlcNAc(beta 1-2)Man(alpha 1-6)]Man(beta 1-4)Glc-NAc(beta 1-4)[Fuc(alpha 1-6)]GlcNAc(beta 1-)Asn. Two other glycopeptides were obtained in very low amount and possess more complex structures.
Notes: Spik, G
Strecker, G
Fournet, B
Bouquelet, S
Montreuil, J
Dorland, L
van Halbeek, H
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1982 Jan;121(2):413-9. doi: 10.1111/j.1432-1033.1982.tb05803.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 680
Author: Spraggon, G., Applegate, D., Everse, S. J., Zhang, J. Z., Veerapandian, L., Redman, C., Doolittle, R. F. and Grieninger, G.
Year: 1998
Title: Crystal structure of a recombinant alphaEC domain from human fibrinogen-420
Journal: Proc Natl Acad Sci U S A
Volume: 95
Issue: 16
Pages: 9099-104
Epub Date: 1998/08/05
Date: Aug 4
Short Title: Crystal structure of a recombinant alphaEC domain from human fibrinogen-420
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.95.16.9099
PMCID: PMC21298
Accession Number: 9689040
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Fibrinogen/*chemistry/genetics
Humans
Molecular Sequence Data
Pichia/genetics
Protein Conformation
Recombinant Proteins/chemistry/genetics
Sequence Homology, Amino Acid
Abstract: The crystal structure of a recombinant alphaEC domain from human fibrinogen-420 has been determined at a resolution of 2.1 A. The protein, which corresponds to the carboxyl domain of the alphaE chain, was expressed in and purified from Pichia pastoris cells. Felicitously, during crystallization an amino-terminal segment was removed, apparently by a contaminating protease, allowing the 201-residue remaining parent body to crystallize. An x-ray structure was determined by molecular replacement. The electron density was clearly defined, partly as a result of averaging made possible by there being eight molecules in the asymmetric unit related by noncrystallographic symmetry (P1 space group). Virtually all of an asparagine-linked sugar cluster is present. Comparison with structures of the beta- and gamma-chain carboxyl domains of human fibrinogen revealed that the binding cleft is essentially neutral and should not bind Gly-Pro-Arg or Gly-His-Arg peptides of the sort bound by those other domains. Nonetheless, the cleft is clearly evident, and the possibility of binding a carbohydrate ligand like sialic acid has been considered.
Notes: 1091-6490
Spraggon, G
Applegate, D
Everse, S J
Zhang, J Z
Veerapandian, L
Redman, C
Doolittle, R F
Grieninger, G
R01 HL026873/HL/NHLBI NIH HHS/United States
HL26873/HL/NHLBI NIH HHS/United States
HL37457/HL/NHLBI NIH HHS/United States
HL51050/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1998 Aug 4;95(16):9099-104. doi: 10.1073/pnas.95.16.9099.
Author Address: Center for Molecular Genetics, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0634, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1334
Author: Srikanth, B., Vaidya, M. M. and Kalraiya, R. D.
Year: 2010
Title: O-GlcNAcylation determines the solubility, filament organization, and stability of keratins 8 and 18
Journal: J Biol Chem
Volume: 285
Issue: 44
Pages: 34062-71
Epub Date: 2010/08/24
Date: Oct 29
Short Title: O-GlcNAcylation determines the solubility, filament organization, and stability of keratins 8 and 18
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.098996
PMCID: PMC2962505
Accession Number: 20729549
Keywords: Acetylglucosamine/*chemistry
Cell Line, Transformed
Cell Line, Tumor
Glycosylation
Hepatocytes/metabolism
Humans
Intermediate Filaments/chemistry
Keratin-18/*chemistry
Keratin-8/*chemistry
Mutation
Phosphorylation
Proteasome Endopeptidase Complex/chemistry
Protein Conformation
Reverse Transcriptase Polymerase Chain Reaction
Solubility
Abstract: Keratins 8 and 18 (K8/18) are intermediate filament proteins expressed specifically in simple epithelial tissues. Dynamic equilibrium of these phosphoglycoproteins in the soluble and filament pool is an important determinant of their cellular functions, and it is known to be regulated by site-specific phosphorylation. However, little is known about the role of dynamic O-GlcNAcylation on this keratin pair. Here, by comparing immortalized (Chang) and transformed hepatocyte (HepG2) cell lines, we have demonstrated that O-GlcNAcylation of K8/18 exhibits a positive correlation with their solubility (Nonidet P-40 extractability). Heat stress, which increases K8/18 solubility, resulted in a simultaneous increase in O-GlcNAc on these proteins. Conversely, increasing O-GlcNAc levels were associated with a concurrent increase in their solubility. This was also associated with a notable decrease in total cellular levels of K8/18. Unaltered levels of transcripts and the reduced half-life of K8 and K18 indicated their decreased stability on increasing O-GlcNAcylation. On the contrary, the K18 glycosylation mutant (K18 S29A/S30A/S48A) was notably more stable than the wild type K18 in Chang cells. The K18-O-GlcNAc mutant accumulated as aggregates upon stable expression, which possibly altered endogenous filament architecture. These results strongly indicate the involvement of O-GlcNAc on K8/18 in regulating their solubility and stability, which may have a bearing on the functions of these keratins.
Notes: 1083-351x
Srikanth, Budnar
Vaidya, Milind M
Kalraiya, Rajiv D
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Oct 29;285(44):34062-71. doi: 10.1074/jbc.M109.098996. Epub 2010 Aug 21.
Author Address: Advanced Centre for Treatment Research and Education in Cancer, Tata Memorial Centre, Kharghar, Navi Mumbai 410210, India.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 558
Author: Staab, J. F., Ginkel, D. L., Rosenberg, G. B. and Munford, R. S.
Year: 1994
Title: A saposin-like domain influences the intracellular localization, stability, and catalytic activity of human acyloxyacyl hydrolase
Journal: J Biol Chem
Volume: 269
Issue: 38
Pages: 23736-42
Epub Date: 1994/09/23
Date: Sep 23
Short Title: A saposin-like domain influences the intracellular localization, stability, and catalytic activity of human acyloxyacyl hydrolase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8089145
Keywords: Animals
Base Sequence
Carboxylic Ester Hydrolases/chemistry/*metabolism
Cell Compartmentation
Cell Line
Chymotrypsin/metabolism
Cricetinae
DNA Primers/chemistry
Glycoproteins/*chemistry
Humans
In Vitro Techniques
Molecular Sequence Data
Protein Precursors/metabolism
Protein Processing, Post-Translational
Saposins
Sphingolipid Activator Proteins
Substrate Specificity
Transfection
Trypsin/metabolism
Abstract: Acyloxyacyl hydrolase, a leukocyte enzyme that acts on bacterial lipopolysaccharides (LPSs) and many glycerolipids, is synthesized as a precursor polypeptide that undergoes internal disulfide linkage before being proteolytically processed into two subunits. The larger subunit contains an amino acid sequence (Gly-X-Ser-X-Gly) that is found at the active sites of many lipases, while the smaller subunit has amino acid sequence similarity to saposins (sphingolipid activator proteins), cofactors for sphingolipid glycohydrolases. We show here that both acyloxyacyl hydrolase subunits are required for catalytic activity toward LPS and glycerophosphatidylcholine. In addition, mutations that truncate or delete the small subunit have profound effects on the intracellular localization, proteolytic processing, and stability of the enzyme in baby hamster kidney cells. Remarkably, proteolytic cleavage of the precursor protein increases the activity of the enzyme toward LPS by 10-20-fold without altering its activity toward glycerophosphatidylcholine. Proper orientation of the two subunits thus seems very important for the substrate specificity of this unusual enzyme.
Notes: Staab, J F
Ginkel, D L
Rosenberg, G B
Munford, R S
AI18188/AI/NIAID NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1994 Sep 23;269(38):23736-42.
Author Address: Department of Internal Medicine, University of Texas Southwestern Medical School, Dallas 75235-9113.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 532
Author: Stadie, T. R., Chai, W., Lawson, A. M., Byfield, P. G. and Hanisch, F. G.
Year: 1995
Title: Studies on the order and site specificity of GalNAc transfer to MUC1 tandem repeats by UDP-GalNAc: polypeptide N-acetylgalactosaminyltransferase from milk or mammary carcinoma cells
Journal: Eur J Biochem
Volume: 229
Issue: 1
Pages: 140-7
Epub Date: 1995/04/01
Date: Apr 1
Short Title: Studies on the order and site specificity of GalNAc transfer to MUC1 tandem repeats by UDP-GalNAc: polypeptide N-acetylgalactosaminyltransferase from milk or mammary carcinoma cells
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
Accession Number: 7744025
Keywords: Amino Acid Sequence
Animals
Binding Sites
Breast Neoplasms/*enzymology
Glycosylation
Membrane Glycoproteins/*chemistry/metabolism
Milk, Human/*enzymology
Molecular Sequence Data
Mucin-1
Mucins/*chemistry/metabolism
N-Acetylgalactosaminyltransferases/isolation & purification/*metabolism
Peptides/*chemical synthesis/metabolism
Repetitive Sequences, Nucleic Acid
Tumor Cells, Cultured/enzymology
Abstract: A synthetic peptide [TAP25, (T1aAPPAHGVT9S10APDT14RPAPGS20)T1bAPPA5b] corresponding to one repeat (T1a-S20) and five overlapping amino acids (T1b-A5b) of the MUC1 core protein served as an acceptor substrate for in vitro glycosylation. TAP25 was glycosylated using the detergent-solubilized UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferases from the breast carcinoma cell line T47D, the colon carcinoma cell line HT29 and from human premature skim milk. The glycosylated peptides were isolated by ultrafiltration, purified by reverse-phase HPLC and further analysed by liquid secondary ion mass spectrometry (LSIMS). Three different glycosylation species, mono-, di- and triglycosylated peptides were identified. Automated Edman sequencing and LSIMS of proteolytic fragments independently revealed the sites of GalNAc incorporation and confirmed that the threonine residues Thr9 and Thr1b are the preferred sites of glycosylation independent of the enzyme source, while Thr14 remained non-glycosylated even with the enzyme preparation from milk. In addition, evidence was obtained that at least 20% of the glycosylated peptides exhibited GalNAc incorporation at Ser20. On the basis of kinetic studies a preferred sequence of GalNAc addition to the three acceptor sites has been concluded (Thr9-->Thr1b-->Ser20). Although Thr14 within the PDTRP motif of the tandem repeats remained non-glycosylated, the introduction of GalNAc into adjacent positions significantly decreased the immunoreactivity of antibodies SM-3, HMFG-1 and HMFG-2 defining overlapping epitopes of this motif. It is assumed that glycosylation at Thr9, Thr1b and Ser20 distorts the peptide conformation of the binding epitope.
Notes: Stadie, T R
Chai, W
Lawson, A M
Byfield, P G
Hanisch, F G
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1995 Apr 1;229(1):140-7.
Author Address: Institute of Immunobiology, University Clinic, Cologne, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 808
Author: Stallcup, W. B. and Dahlin-Huppe, K.
Year: 2001
Title: Chondroitin sulfate and cytoplasmic domain-dependent membrane targeting of the NG2 proteoglycan promotes retraction fiber formation and cell polarization
Journal: J Cell Sci
Volume: 114
Issue: Pt 12
Pages: 2315-25
Epub Date: 2001/08/09
Date: Jun
Short Title: Chondroitin sulfate and cytoplasmic domain-dependent membrane targeting of the NG2 proteoglycan promotes retraction fiber formation and cell polarization
Alternate Journal: Journal of cell science
ISSN: 0021-9533 (Print)
0021-9533
Accession Number: 11493670
Keywords: Animals
Antigens/*chemistry/genetics/*metabolism
Blotting, Western
Cell Membrane/*metabolism
*Cell Polarity
Chondroitin Sulfates/*metabolism
Cytoplasm/*metabolism
Cytoskeleton/*metabolism
Humans
Microscopy, Fluorescence
Point Mutation
Protein Transport
Proteoglycans/*chemistry/genetics/*metabolism
Rats
Serine/genetics/metabolism
Transfection
Tumor Cells, Cultured
rho GTP-Binding Proteins/metabolism
Abstract: Targeting of the NG2 proteoglycan to cellular retraction fibers was studied by expressing mutant NG2 molecules lacking specific structural elements of the proteoglycan. Both the cytoplasmic domain and the chondroitin sulfate chain of NG2 appear to have roles in sorting NG2 to subcellular microdomains destined to become retraction fibers. Neither of these structural features alone is sufficient to allow optimal targeting of NG2 to retraction fibers, but together they promote efficient localization of the proteoglycan to these sites. This pattern of NG2 sorting seems to be necessary for optimal retraction fiber formation, as cells expressing poorly targeted NG2 mutants are noticeably deficient in their ability to extend retraction fibers. Furthermore, retraction fiber formation correlates strongly with the tendency of cells to assume a polarized morphology with NG2-positive retraction fibers at one pole of the cell and actin-rich lamellipodia at the other. This polarization can be triggered either through engagement of NG2 by the substratum or by exposure to lysophosphatidic acid, a potent activator of the rho GTPase. These results suggest a possible role for NG2 in regulating rho-dependent mechanisms in the trailing processes of motile cells.
Notes: Stallcup, W B
Dahlin-Huppe, K
P01 HD25938/HD/NICHD NIH HHS/United States
R01 AR44400/AR/NIAMS NIH HHS/United States
R01 NS21990/NS/NINDS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
J Cell Sci. 2001 Jun;114(Pt 12):2315-25.
Author Address: The Burnham Institute, La Jolla Cancer Research Center, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1193
Author: Stamler, R., Keutmann, H. T., Sidis, Y., Kattamuri, C., Schneyer, A. and Thompson, T. B.
Year: 2008
Title: The structure of FSTL3.activin A complex. Differential binding of N-terminal domains influences follistatin-type antagonist specificity
Journal: J Biol Chem
Volume: 283
Issue: 47
Pages: 32831-8
Epub Date: 2008/09/05
Date: Nov 21
Short Title: The structure of FSTL3.activin A complex. Differential binding of N-terminal domains influences follistatin-type antagonist specificity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M801266200
PMCID: PMC2583307
Accession Number: 18768470
Keywords: Activin Receptors/chemistry
Activins/*chemistry
Binding Sites
Cell Line
Crystallography, X-Ray
Electrons
Follistatin/*chemistry
Follistatin-Related Proteins/*chemistry
Humans
Ligands
Models, Molecular
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Surface Properties
Abstract: Transforming growth factor beta family ligands are neutralized by a number of structurally divergent antagonists. Follistatin-type antagonists, which include splice variants of follistatin (FS288 and FS315) and follistatin-like 3 (FSTL3), have high affinity for activin A but differ in their affinity for other ligands, particularly bone morphogenetic proteins. To understand the structural basis for ligand specificity within FS-type antagonists, we determined the x-ray structure of activin A in complex with FSTL3 to a resolution of 2.5 A. Similar to the previously resolved FS.activin A structures, the ligand is encircled by two antagonist molecules blocking all ligand receptor-binding sites. Recently, the significance of the FS N-terminal domain interaction at the ligand type I receptor site has been questioned; however, our data show that for FSTL3, the N-terminal domain forms a more intimate contact with activin A, implying that this interaction is stronger than that for FS. Furthermore, binding studies revealed that replacing the FSTL3 N-terminal domain with the corresponding FS domain considerably lowers activin A affinity. Therefore, both structural and biochemical evidence support a significant interaction of the N-terminal domain of FSTL3 with activin A. In addition, structural comparisons with bone morphogenetic proteins suggest that the interface where the N-terminal domain binds may be the key site for determining FS-type antagonist specificity.
Notes: 1083-351x
Stamler, Robin
Keutmann, Henry T
Sidis, Yisrael
Kattamuri, Chandramohan
Schneyer, Alan
Thompson, Thomas B
DK053828/DK/NIDDK NIH HHS/United States
GM084186/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2008 Nov 21;283(47):32831-8. doi: 10.1074/jbc.M801266200. Epub 2008 Sep 2.
Author Address: Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati Medical Sciences Building, Cincinnati, Ohio 45267, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 792
Author: Stamper, C. C., Zhang, Y., Tobin, J. F., Erbe, D. V., Ikemizu, S., Davis, S. J., Stahl, M. L., Seehra, J., Somers, W. S. and Mosyak, L.
Year: 2001
Title: Crystal structure of the B7-1/CTLA-4 complex that inhibits human immune responses
Journal: Nature
Volume: 410
Issue: 6828
Pages: 608-11
Epub Date: 2001/03/30
Date: Mar 29
Short Title: Crystal structure of the B7-1/CTLA-4 complex that inhibits human immune responses
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/35069118
Accession Number: 11279502
Keywords: Abatacept
Animals
Antigens, CD
Antigens, Differentiation/*chemistry/genetics/immunology
B7-1 Antigen/*chemistry/genetics/immunology
CHO Cells
CTLA-4 Antigen
Cricetinae
Cricetulus
Crystallography, X-Ray
Humans
Immunity/physiology
*Immunoconjugates
Macromolecular Substances
Mice
Models, Molecular
Mutation
Protein Conformation
Recombinant Proteins/chemistry
T-Lymphocytes/chemistry/*immunology
Abstract: Optimal immune responses require both an antigen-specific and a co-stimulatory signal. The shared ligands B7-1 and B7-2 on antigen-presenting cells deliver the co-stimulatory signal through CD28 and CTLA-4 on T cells. Signalling through CD28 augments the T-cell response, whereas CTLA-4 signalling attenuates it. Numerous animal studies and recent clinical trials indicate that manipulating these interactions holds considerable promise for immunotherapy. With the consequences of these signals well established, and details of the downstream signalling events emerging, understanding the molecular nature of these extracellular interactions becomes crucial. Here we report the crystal structure of the human CTLA-4/B7-1 co-stimulatory complex at 3.0 A resolution. In contrast to other interacting cell-surface molecules, the relatively small CTLA-4/B7-1 binding interface exhibits an unusually high degree of shape complementarity. CTLA-4 forms homodimers through a newly defined interface of highly conserved residues. In the crystal lattice, CTLA-4 and B7-1 pack in a strikingly periodic arrangement in which bivalent CTLA-4 homodimers bridge bivalent B7-1 homodimers. This zipper-like oligomerization provides the structural basis for forming unusually stable signalling complexes at the T-cell surface, underscoring the importance of potent inhibitory signalling in human immune responses.
Notes: Stamper, C C
Zhang, Y
Tobin, J F
Erbe, D V
Ikemizu, S
Davis, S J
Stahl, M L
Seehra, J
Somers, W S
Mosyak, L
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2001 Mar 29;410(6828):608-11. doi: 10.1038/35069118.
Author Address: Departments of Biological Chemistry and Musculoskeletal Sciences, Wyeth Research, 87 Cambridge Park Drive, Cambridge, Massachusetts 02140, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 697
Author: Ständker, L., Wobst, P., Mark, S. and Forssmann, W. G.
Year: 1998
Title: Isolation and characterization of circulating 13-kDa C-terminal fragments of human insulin-like growth factor binding protein-5
Journal: FEBS Lett
Volume: 441
Issue: 2
Pages: 281-6
Epub Date: 1999/01/12
Date: Dec 18
Short Title: Isolation and characterization of circulating 13-kDa C-terminal fragments of human insulin-like growth factor binding protein-5
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/s0014-5793(98)01497-5
Accession Number: 9883900
Keywords: Amino Acid Sequence
Glycosylation
Humans
Hydrolysis
Insulin-Like Growth Factor Binding Protein 5/blood/*chemistry
Molecular Sequence Data
Peptide Fragments/blood/chemistry/*isolation & purification
Abstract: The insulin-like growth factor binding proteins (IGFBPs) are responsible for regulation of the effects and the bioavailability of the insulin-like growth factors (IGFs). We screened for circulating fragments of human IGFBP-5 in human hemofiltrate. Identification of IGFBP-5 peptides in the fractions of our peptide bank generated from hemofiltrate was performed by their immunoreactivity and their capacity to bind IGF-I. Different fragments of IGFBP-5 with molecular sizes from 12 to 25 kDa were identified. C-terminal peptides of IGFBP-5 with molecular masses of 13.3 and 13.5 kDa were purified by consecutive chromatographic steps and sequenced. Sequence analysis of the peptides revealed the (double) sequences (K)FVGGAENXAHPRII and MVPRAVYLPNXDRKG. In addition, a smaller fragment with Mr 2722 of the central IGFBP-5 region was purified and showed the sequence HTRISELKAEAVKKDRRKKLTQS (residues 121-143) indicating plasma proteolysis of IGFBP-5 C-terminal to amino acids Lys-120, Ser-143, Lys-144, and Arg-188. According to mass spectrometric and sequence analysis, Thr-152 was shown to be O-glycosylated. Fractions containing C-terminal IGFBP-5 fragments revealed significant IGF-I binding properties. Our results indicate that plasma proteolysis of IGFBP-5 preferentially occurs C-terminally to basic residues and generates different C-terminal fragments, possibly acting in an IGF-dependent manner and bearing intrinsic biological functions.
Notes: Ständker, L
Wobst, P
Mark, S
Forssmann, W G
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1998 Dec 18;441(2):281-6. doi: 10.1016/s0014-5793(98)01497-5.
Author Address: Lower Saxony Institute for Peptide Research, Hannover, Germany. 106535.324@compuserve.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1707
Author: Stateva, S. R. and Villalobo, A.
Year: 2015
Title: O-GlcNAcylation of the human epidermal growth factor receptor
Journal: Org Biomol Chem
Volume: 13
Issue: 30
Pages: 8196-204
Epub Date: 2015/06/26
Date: Aug 14
Short Title: O-GlcNAcylation of the human epidermal growth factor receptor
Alternate Journal: Organic & biomolecular chemistry
ISSN: 1477-0520
DOI: 10.1039/c5ob00443h
Accession Number: 26108188
Keywords: Acetylglucosamine/*metabolism
Acetylglucosaminidase/antagonists & inhibitors/metabolism
Animals
Cell Line, Tumor
Enzyme Inhibitors/pharmacology
ErbB Receptors/*metabolism
Glycosylation/drug effects
Humans
Immunoblotting
Immunoprecipitation
Mice
Staining and Labeling
Abstract: The reversible O-linked attachment of single β-D-N-acetylglucosamine (GlcNAc) moieties to serine/threonine residues in target proteins is a frequently occurring post-translational modification affecting the functionality of many cellular systems. In this report we present experimental evidence suggesting that the epidermal growth factor receptor (EGFR) is subjected to O-GlcNAcylation in human carcinoma epidermoid A431 cells and human lung carcinoma A549 cells. However, no signal was detected in human cervix adenocarcinoma HeLa cells or in mouse EGFR-T17 fibroblasts ectopically expressing the human EGFR. We detected a positive O-GlcNAcylation signal in the immunoprecipitated EGFR by Western blotting using two distinct specific anti-O-GlcNAc antibodies even after N-deglycosylation of the receptor using peptide-N-glycosidase F (PNGase F). Conversely, the presence of EGFR was detected by Western blotting using an anti-EGFR antibody in the immunocomplex of O-GlcNAcylated proteins immunoprecipitated with an anti-O-GlcNAc antibody. These signals were enhanced when the O-linked β-N-acetylglucosaminidase (OGA) inhibitor Thiamet G was added to prevent the deglycosylation of the GlcNAc moiety(ies). Moreover, we also detected a positive signal in the immunoprecipitated and N-deglycosylated EGFR using PNGase F, and tunicamycin when the cells were metabolically labeled with azido-GlcNAc (GlcNAz), biotinylated and probed with a streptavidin-labeled peroxidase. Finally, EGFR and O-linked β-N-acetylglucosamine transferase (OGT) co-immunoprecipitate, and incubation of the immunoprecipitated EGFR with the immunoprecipitated OGT in the presence of uridine 5'-diphospho-N-acetylglucosamine (UDP-GlcNAc) resulted in a significant enhancement of the EGFR O-GlcNAcylation signal as detected by Western blotting using an anti-O-GlcNAc antibody. We conclude that the human EGFR is subjected to O-GlcNAcylation in the A431 and A549 tumor cell lines.
Notes: 1477-0539
Stateva, Silviya R
Villalobo, Antonio
Journal Article
Research Support, Non-U.S. Gov't
England
Org Biomol Chem. 2015 Aug 14;13(30):8196-204. doi: 10.1039/c5ob00443h.
Author Address: Department of Cancer Biology, Instituto de Investigaciones Biomédicas, Consejo Superior de Investigaciones Científicas and Universidad Autónoma de Madrid, c/Arturo Duperier 4, E-28029 Madrid, Spain. antonio.villalobo@iib.uam.es.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1034
Author: Stauber, D. J., Debler, E. W., Horton, P. A., Smith, K. A. and Wilson, I. A.
Year: 2006
Title: Crystal structure of the IL-2 signaling complex: paradigm for a heterotrimeric cytokine receptor
Journal: Proc Natl Acad Sci U S A
Volume: 103
Issue: 8
Pages: 2788-93
Epub Date: 2006/02/16
Date: Feb 21
Short Title: Crystal structure of the IL-2 signaling complex: paradigm for a heterotrimeric cytokine receptor
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0511161103
PMCID: PMC1413841
Accession Number: 16477002
Keywords: Crystallography
Humans
Interleukin-2/*chemistry
Protein Structure, Quaternary
Receptors, Interleukin-2/*chemistry
Signal Transduction
Abstract: IL-2 is a cytokine that functions as a growth factor and central regulator in the immune system and mediates its effects through ligand-induced hetero-trimerization of the receptor subunits IL-2R alpha, IL-2R beta, and gamma(c). Here, we describe the crystal structure of the trimeric assembly of the human IL-2 receptor ectodomains in complex with IL-2 at 3.0 A resolution. The quaternary structure is consistent with a stepwise assembly from IL-2/IL-2R alpha to IL-2/IL-2R alpha/IL-2R beta to IL-2/IL-2R alpha/IL-2R beta/gamma(c). The IL-2R alpha subunit forms the largest of the three IL-2/IL-2R interfaces, which, together with the high abundance of charge-charge interactions, correlates well with the rapid association rate and high-affinity interaction of IL-2R alpha with IL-2 at the cell surface. Surprisingly, IL-2R alpha makes no contacts with IL-2R beta or gamma(c), and only minor changes are observed in the IL-2 structure in response to receptor binding. These findings support the principal role of IL-2R alpha to deliver IL-2 to the signaling complex and act as regulator of signal transduction. Cooperativity in assembly of the final quaternary complex is easily explained by the extraordinarily extensive set of interfaces found within the fully assembled IL-2 signaling complex, which nearly span the entire length of the IL-2R beta and gamma(c) subunits. Helix A of IL-2 wedges tightly between IL-2R beta and gamma(c) to form a three-way junction that coalesces into a composite binding site for the final gamma(c) recruitment. The IL-2/gamma(c) interface itself exhibits the smallest buried surface and the fewest hydrogen bonds in the complex, which is consistent with its promiscuous use in other cytokine receptor complexes.
Notes: 1091-6490
Stauber, Deborah J
Debler, Erik W
Horton, Patricia A
Smith, Kendall A
Wilson, Ian A
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2006 Feb 21;103(8):2788-93. doi: 10.1073/pnas.0511161103. Epub 2006 Feb 13.
Author Address: Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1745
Author: Stavenhagen, K., Plomp, R. and Wuhrer, M.
Year: 2015
Title: Site-Specific Protein N- and O-Glycosylation Analysis by a C18-Porous Graphitized Carbon-Liquid Chromatography-Electrospray Ionization Mass Spectrometry Approach Using Pronase Treated Glycopeptides
Journal: Anal Chem
Volume: 87
Issue: 23
Pages: 11691-9
Epub Date: 2015/11/05
Date: Dec 1
Short Title: Site-Specific Protein N- and O-Glycosylation Analysis by a C18-Porous Graphitized Carbon-Liquid Chromatography-Electrospray Ionization Mass Spectrometry Approach Using Pronase Treated Glycopeptides
Alternate Journal: Analytical chemistry
ISSN: 0003-2700
DOI: 10.1021/acs.analchem.5b02366
Accession Number: 26536155
Keywords: Carbon/*chemistry
Chromatography, Liquid
Glycopeptides/*analysis/*chemistry/metabolism
Glycosylation
Humans
Particle Size
Porosity
Pronase/*metabolism
*Spectrometry, Mass, Electrospray Ionization
Surface Properties
Abstract: The analysis of N- and O-glycopeptides remains challenging due to the microheterogeneity (different glycoforms attached to one glycosylation site) and macroheterogeneity (site occupancy) of the glycoprotein. Trypsin is by far the most commonly used protease in glycoproteomic studies; however, it often results in long peptides that can harbor more than one glycan which may hamper site identification. The use of unspecific proteases such as Pronase can largely overcome this problem by generating glycopeptides with a small peptide portion. While the resulting glycopeptides are very useful for tandem mass spectrometric investigation, the analysis with conventional 1D-LC-ESI-MS/MS approaches can lead to incomplete glycosylation coverage because of the very heterogeneous physicochemical properties of the glycopeptides depending on the peptide sequence as well as the size and charges of the glycan moiety. Here, we describe a universal workflow for site-specific N- and O-glycopeptide analysis of Pronase treated glycoproteins with integrated, sequential C18 reverse phase and porous graphitized carbon-LC-ESI-QTOF-MS/MS employing a combination of lower- and enhanced-energy collision-induced dissociation. The approach was evaluated on glycoprotein standards and also applied to investigate the glycosylation of human IgG3 providing details on the hitherto uncharacterized glycosylation site Asn392 of the CH3 domain. This analytical tool can be applied to a variety of glycoproteins for site-specific N- and O-glycopeptide analysis, resulting in a good glycopeptide coverage within a single sample run and, thus, requiring only small amounts of sample.
Notes: 1520-6882
Stavenhagen, Kathrin
Plomp, Rosina
Wuhrer, Manfred
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Chem. 2015 Dec 1;87(23):11691-9. doi: 10.1021/acs.analchem.5b02366. Epub 2015 Nov 12.
Author Address: Division of BioAnalytical Chemistry, VU University Amsterdam , De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands.
Center for Proteomics and Metabolomics, Leiden University Medical Center , P.O. Box 9600, 2300 RC Leiden, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 329
Author: Stearns, N. A., Dong, J. M., Pan, J. X., Brenner, D. A. and Sahagian, G. G.
Year: 1990
Title: Comparison of cathepsin L synthesized by normal and transformed cells at the gene, message, protein, and oligosaccharide levels
Journal: Arch Biochem Biophys
Volume: 283
Issue: 2
Pages: 447-57
Epub Date: 1990/12/01
Date: Dec
Short Title: Comparison of cathepsin L synthesized by normal and transformed cells at the gene, message, protein, and oligosaccharide levels
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1016/0003-9861(90)90666-m
Accession Number: 2275556
Keywords: Amino Acid Sequence
Animals
Base Sequence
Cathepsin L
Cathepsins/biosynthesis/*genetics
Cell Line
*Cell Transformation, Neoplastic
Cloning, Molecular
Cysteine Endopeptidases
*Endopeptidases
Gene Library
*Genes
Glycosylation
Kirsten murine sarcoma virus/*genetics
Mice
Molecular Sequence Data
Mutagenesis, Site-Directed
Oligonucleotide Probes
RNA, Messenger/*genetics/isolation & purification
Abstract: The major excreted protein of transformed mouse fibroblasts (MEP) has recently been identified as the lysosomal cysteine protease, cathepsin L. The synthesis and intracellular trafficking of this protein in mouse fibroblasts are regulated by growth factors and malignant transformation. To further define the basis for this regulation, a cDNA encoding MEP/cathepsin L was isolated from a mouse liver cDNA library and used to compare cathepsin L of normal and Kirsten sarcoma virus-transformed NIH 3T3 fibroblasts. Although cathepsin L message levels were elevated 20-fold in the transformed fibroblasts, normal and transformed cells displayed similar cathepsin L genomic DNA digest patterns and gene copy numbers, and cathepsin L mRNA sequences appeared identical by RNase protection analysis. These findings indicate that (i) cathepsin L is synthesized from the same gene in normal and transformed cells and (ii) cathepsin L polypeptides made by these cells are translated with the same primary sequence. Cathepsin L polypeptides synthesized by quiescent, growing, and transformed cells displayed similar isoelectric focusing patterns, suggesting similar post-translational modification. Site-directed mutagenesis of the mouse liver cDNA and expression in COS monkey cells was used to examine the glycosylation of mouse cathepsin L. The results indicated that only one of the two potential N-linked glycosylation sites (the one at Asn221) is glycosylated. Analysis by ion exchange chromatography on QAE-Sephadex, and affinity chromatography on mannose 6-phosphate receptor-Affi-Gel 10, indicated that the cathepsin L oligosaccharide was phosphorylated similarly in normal and transformed cells. Although several phosphorylated oligosaccharide species were observed, the major species contained two phosphomonoester moieties and bound efficiently to the receptor. These findings suggest that cathepsin L made by normal and transformed mouse fibroblasts are identical and substantiate the hypothesis that trafficking of cathepsin L in these cells is regulated by growth-induced changes in the lysosomal protein transport system.
Notes: Stearns, N A
Dong, J M
Pan, J X
Brenner, D A
Sahagian, G G
DK36632/DK/NIDDK NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Arch Biochem Biophys. 1990 Dec;283(2):447-57. doi: 10.1016/0003-9861(90)90666-m.
Author Address: Department of Physiology, School of Medicine, Tufts University, Boston, Massachusetts 02111.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1329
Author: Stec, B., Cheltsov, A. and Millán, J. L.
Year: 2010
Title: Refined structures of placental alkaline phosphatase show a consistent pattern of interactions at the peripheral site
Journal: Acta Crystallogr Sect F Struct Biol Cryst Commun
Volume: 66
Issue: Pt 8
Pages: 866-70
Epub Date: 2010/08/10
Date: Aug 1
Short Title: Refined structures of placental alkaline phosphatase show a consistent pattern of interactions at the peripheral site
Alternate Journal: Acta crystallographica. Section F, Structural biology and crystallization communications
ISSN: 1744-3091
DOI: 10.1107/s1744309110019767
PMCID: PMC2917279
Accession Number: 20693656
Keywords: Alkaline Phosphatase/*chemistry
Crystallography, X-Ray
Female
Humans
Models, Molecular
Placenta/*enzymology
Pregnancy
*Protein Interaction Domains and Motifs
Protein Structure, Quaternary
Abstract: In order to gain deeper insights into the functional sites of human placental alkaline phosphatase, the structures of the enzyme with the putative regulators L-Phe, pNPP and 5'-AMP [Llinas et al. (2005), J. Mol. Biol. 350, 441-451] were re-refined. Significant variations in ligand positioning and identity were found compared with the previous report. The multiple corrections to the model improved the phases and the electron-density maps, allowing the modeling of omitted side chains and multiple disordered residues. These improvements led to a change in the position of L-Phe at the peripheral binding site, which appeared to be reversed. The structure with pNPP contained only p-nitrophenol in three distinct sites, while the structure with 5'-AMP contained the p-nitrophenyl group in two of the sites instead of 5'-AMP. Comparison of the re-refined models shows a consistent pattern of interactions at the peripheral site.
Notes: 1744-3091
Stec, Boguslaw
Cheltsov, Anton
Millán, José Luis
Journal Article
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2010 Aug 1;66(Pt 8):866-70. doi: 10.1107/S1744309110019767. Epub 2010 Jul 27.
Author Address: Sanford-Burnham Medical Research Institute, La Jolla, CA 92037, USA. bstec@burnham.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1305
Author: Steenhuis, P., Herder, S., Gelis, S., Braulke, T. and Storch, S.
Year: 2010
Title: Lysosomal targeting of the CLN7 membrane glycoprotein and transport via the plasma membrane require a dileucine motif
Journal: Traffic
Volume: 11
Issue: 7
Pages: 987-1000
Epub Date: 2010/04/22
Date: Jul 1
Short Title: Lysosomal targeting of the CLN7 membrane glycoprotein and transport via the plasma membrane require a dileucine motif
Alternate Journal: Traffic (Copenhagen, Denmark)
ISSN: 1398-9219
DOI: 10.1111/j.1600-0854.2010.01073.x
Accession Number: 20406422
Keywords: *Amino Acid Motifs/genetics
Animals
Biological Transport/genetics
Biotinylation
COS Cells
Cell Line
Cell Membrane/genetics/*metabolism
Chlorocebus aethiops
Clathrin/genetics/metabolism
Cytosol/metabolism
Endocytosis/genetics/physiology
HeLa Cells
Humans
*Leucine
Lysosomal Storage Diseases/genetics/metabolism
Lysosome-Associated Membrane Glycoproteins/genetics/metabolism
Lysosomes/genetics/*metabolism
Membrane Glycoproteins/genetics/*metabolism
*Membrane Transport Proteins/genetics/metabolism
Protein Structure, Tertiary/genetics
Protein Transport/genetics
Recombinant Proteins/metabolism
Abstract: CLN7 is a polytopic lysosomal membrane protein deficient in variant late infantile neuronal ceroid lipofuscinosis, a neurodegenerative lysosomal storage disorder. In this study fluorescence protease protection assays and mutational analyses revealed the N- and C-terminal tails of CLN7 in the cytosol and two N-glycosylation sites at N371 and N376. Both partially and non-glycosylated CLN7 were correctly transported to lysosomes. To identify lysosomal targeting motifs, we generated CD4-chimera fused to the N- and C-terminal domains of CLN7. Lysosomal localization of the chimeric proteins requires a consensus acidic dileucine-based motif in the N-terminus and two tandem tyrosine-based signals in the C-terminus. Mutation of these sorting motifs resulted in cell surface redistribution of CD4 chimeras. However, the dileucine-based motif is of critical importance for lysosomal localization of the full-length CLN7 in different cell lines. Cell surface biotinylation revealed that at equilibrium 22% of total CLN7 is localized at the plasma membrane. Mutation of the dileucine motif or the co-expression of dominant-negative mutant dynamin K44A led to a further increase of CLN7 at the plasma membrane. Our data demonstrate that CLN7 contains several cytoplasmic lysosomal targeting signals of which the N-terminal dileucine-based motif is required for the predominant lysosomal targeting along the indirect pathway and clathrin-mediated endocytosis of CLN7.
Notes: 1600-0854
Steenhuis, Pieter
Herder, Stephanie
Gelis, Suyin
Braulke, Thomas
Storch, Stephan
Journal Article
Research Support, Non-U.S. Gov't
England
Traffic. 2010 Jul 1;11(7):987-1000. doi: 10.1111/j.1600-0854.2010.01073.x. Epub 2010 Apr 16.
Author Address: Department of Biochemistry, Children's Hospital, University Medical Center Hamburg-Eppendorf, Martinistr. 52, D-20246 Hamburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1552
Author: Steentoft, C., Vakhrushev, S. Y., Joshi, H. J., Kong, Y., Vester-Christensen, M. B., Schjoldager, K. T., Lavrsen, K., Dabelsteen, S., Pedersen, N. B., Marcos-Silva, L., Gupta, R., Bennett, E. P., Mandel, U., Brunak, S., Wandall, H. H., Levery, S. B. and Clausen, H.
Year: 2013
Title: Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology
Journal: Embo j
Volume: 32
Issue: 10
Pages: 1478-88
Epub Date: 2013/04/16
Date: May 15
Short Title: Precision mapping of the human O-GalNAc glycoproteome through SimpleCell technology
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2013.79
PMCID: PMC3655468
Accession Number: 23584533
Keywords: Algorithms
Amino Acid Motifs
Cell Line, Tumor
Genetic Engineering/methods
Glycoproteins/*analysis/metabolism
Glycosylation
Humans
N-Acetylgalactosaminyltransferases/genetics/*metabolism
Proteomics/*methods
Abstract: Glycosylation is the most abundant and diverse posttranslational modification of proteins. While several types of glycosylation can be predicted by the protein sequence context, and substantial knowledge of these glycoproteomes is available, our knowledge of the GalNAc-type O-glycosylation is highly limited. This type of glycosylation is unique in being regulated by 20 polypeptide GalNAc-transferases attaching the initiating GalNAc monosaccharides to Ser and Thr (and likely some Tyr) residues. We have developed a genetic engineering approach using human cell lines to simplify O-glycosylation (SimpleCells) that enables proteome-wide discovery of O-glycan sites using 'bottom-up' ETD-based mass spectrometric analysis. We implemented this on 12 human cell lines from different organs, and present a first map of the human O-glycoproteome with almost 3000 glycosites in over 600 O-glycoproteins as well as an improved NetOGlyc4.0 model for prediction of O-glycosylation. The finding of unique subsets of O-glycoproteins in each cell line provides evidence that the O-glycoproteome is differentially regulated and dynamic. The greatly expanded view of the O-glycoproteome should facilitate the exploration of how site-specific O-glycosylation regulates protein function.
Notes: 1460-2075
Steentoft, Catharina
Vakhrushev, Sergey Y
Joshi, Hiren J
Kong, Yun
Vester-Christensen, Malene B
Schjoldager, Katrine T-B G
Lavrsen, Kirstine
Dabelsteen, Sally
Pedersen, Nis B
Marcos-Silva, Lara
Gupta, Ramneek
Bennett, Eric Paul
Mandel, Ulla
Brunak, Søren
Wandall, Hans H
Levery, Steven B
Clausen, Henrik
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2013 May 15;32(10):1478-88. doi: 10.1038/emboj.2013.79. Epub 2013 Apr 12.
Author Address: Copenhagen Center for Glycomics, Department of Cellular and Molecular Medicine and School of Dentistry, University of Copenhagen, Copenhagen N, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1733
Author: Stein, A. J., Bain, G., Prodanovich, P., Santini, A. M., Darlington, J., Stelzer, N. M., Sidhu, R. S., Schaub, J., Goulet, L., Lonergan, D., Calderon, I., Evans, J. F. and Hutchinson, J. H.
Year: 2015
Title: Structural Basis for Inhibition of Human Autotaxin by Four Potent Compounds with Distinct Modes of Binding
Journal: Mol Pharmacol
Volume: 88
Issue: 6
Pages: 982-92
Epub Date: 2015/09/16
Date: Dec
Short Title: Structural Basis for Inhibition of Human Autotaxin by Four Potent Compounds with Distinct Modes of Binding
Alternate Journal: Molecular pharmacology
ISSN: 0026-895x
DOI: 10.1124/mol.115.100404
Accession Number: 26371182
Keywords: Animals
Cell Line, Tumor
Crystallization
Enzyme Inhibitors/*chemistry/*metabolism
HEK293 Cells
Humans
Mice
Phosphoric Diester Hydrolases/*chemistry/*metabolism
Protein Binding/physiology
Protein Structure, Secondary
Protein Structure, Tertiary
Structure-Activity Relationship
Abstract: Autotaxin (ATX) is a secreted enzyme that hydrolyzes lysophosphatidylcholine to lysophosphatidic acid (LPA). LPA is a bioactive phospholipid that regulates diverse biological processes, including cell proliferation, migration, and survival/apoptosis, through the activation of a family of G protein-coupled receptors. The ATX-LPA pathway has been implicated in many pathologic conditions, including cancer, fibrosis, inflammation, cholestatic pruritus, and pain. Therefore, ATX inhibitors represent an attractive strategy for the development of therapeutics to treat a variety of diseases. Mouse and rat ATX have been crystallized previously with LPA or small-molecule inhibitors bound. Here, we present the crystal structures of human ATX in complex with four previously unpublished, structurally distinct ATX inhibitors. We demonstrate that the mechanism of inhibition of each compound reflects its unique interactions with human ATX. Our studies may provide a basis for the rational design of novel ATX inhibitors.
Notes: 1521-0111
Stein, Adam J
Bain, Gretchen
Prodanovich, Pat
Santini, Angelina M
Darlington, Janice
Stelzer, Nina M P
Sidhu, Ranjinder S
Schaub, Jeffrey
Goulet, Lance
Lonergan, Dave
Calderon, Imelda
Evans, Jilly F
Hutchinson, John H
Journal Article
Research Support, Non-U.S. Gov't
United States
Mol Pharmacol. 2015 Dec;88(6):982-92. doi: 10.1124/mol.115.100404. Epub 2015 Sep 14.
Author Address: Cayman Chemical Company, Ann Arbor, Michigan (A.J.S., N.M.P.S., R.S.S., J.S.); and PharmAkea, San Diego, California (G.B., P.P., A.M.S., J.D., L.G., D.L., I.C., J.F.E., J.H.H.).
Cayman Chemical Company, Ann Arbor, Michigan (A.J.S., N.M.P.S., R.S.S., J.S.); and PharmAkea, San Diego, California (G.B., P.P., A.M.S., J.D., L.G., D.L., I.C., J.F.E., J.H.H.) gbain@pharmakea.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 356
Author: Stein, C., Hille, A., Seidel, J., Rijnbout, S., Waheed, A., Schmidt, B., Geuze, H. and von Figura, K.
Year: 1989
Title: Cloning and expression of human steroid-sulfatase. Membrane topology, glycosylation, and subcellular distribution in BHK-21 cells
Journal: J Biol Chem
Volume: 264
Issue: 23
Pages: 13865-72
Epub Date: 1989/08/15
Date: Aug 15
Short Title: Cloning and expression of human steroid-sulfatase. Membrane topology, glycosylation, and subcellular distribution in BHK-21 cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2668275
Keywords: Amino Acid Sequence
Animals
Arylsulfatases/biosynthesis/*genetics
Base Sequence
Cell Line
Cell Membrane/enzymology
*Cloning, Molecular
Cricetinae
DNA/genetics/isolation & purification
Female
Fluorescent Antibody Technique
Glycosylation
Humans
Kidney
Molecular Sequence Data
Placenta/enzymology
Plasmids
Pregnancy
Steryl-Sulfatase
Sulfatases/*genetics
Transfection
Abstract: A 2.4-kilobase cDNA clone for human steroid-sulfatase (STS) was isolated and sequenced, which encoded an enzymatically active protein. The deduced amino acid sequence comprises 583 amino acids with an N-terminal signal peptide of 21 or 23 residues and four potential N-glycosylation sites. Two of the N-glycosylation sites are utilized and were localized to the asparagine residues 47 and 259. STS has the solubility properties of an integral membrane protein. The resistance of STS toward proteinase K after translocation into microsomes suggests that most, if not all, sequences of STS are exposed at the luminal side of microsomes. The deduced amino acid sequence predicts two membrane-spanning domains (amino acids 185-211 and 213-237) separated by a helix-breaking proline residue. We propose for STS a three-domain model. Two glycosylated luminally oriented domains of 161 and 346 residues are separated by a hydrophobic domain spanning the membrane twice in opposite directions. STS expressed in BHK-21 cells is located predominantly in the endoplasmic reticulum; smaller fractions are found in the Golgi, at the cell surface, multivesicular endosomes, as well as in lysosomes. The stability of STS in lysosomes may be related to the high homology of the two luminal domains of STS with the lysosomal sulfatases, arylsulfatase A, and arylsulfatase B. In spite of its similarity with these two lysosomal sulfatases, STS does not contain mannose 6-phosphate residues and is transported to lysosomes by a mannose 6-phosphate receptor-independent mechanism.
Notes: Stein, C
Hille, A
Seidel, J
Rijnbout, S
Waheed, A
Schmidt, B
Geuze, H
von Figura, K
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1989 Aug 15;264(23):13865-72.
Author Address: Universität Göttingen, Abteilung Biochemie II, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1609
Author: Steiner, N. K., Dakshanamurthy, S., Nguyen, N. and Hurley, C. K.
Year: 2014
Title: Allelic variation of killer cell immunoglobulin-like receptor 2DS5 impacts glycosylation altering cell surface expression levels
Journal: Hum Immunol
Volume: 75
Issue: 2
Pages: 124-8
Epub Date: 2013/11/26
Date: Feb
Short Title: Allelic variation of killer cell immunoglobulin-like receptor 2DS5 impacts glycosylation altering cell surface expression levels
Alternate Journal: Human immunology
ISSN: 0198-8859 (Print)
0198-8859
DOI: 10.1016/j.humimm.2013.11.012
PMCID: PMC3924325
NIHMSID: NIHMS550482
Accession Number: 24269691
Keywords: Alleles
Computational Biology
Cytokines/metabolism
Cytotoxicity, Immunologic/genetics
*Gene Expression Regulation/genetics
*Glycosylation
Humans
Jurkat Cells
Killer Cells, Natural/*immunology
Lymphocyte Activation/genetics
Models, Molecular
Mutagenesis, Site-Directed
Mutation/genetics
Polymorphism, Genetic
Protein Isoforms/genetics/*metabolism
Receptors, KIR/genetics/*metabolism
Transgenes/genetics
2dl
Cell surface molecules
Human
Kir
Killer cell immunoglobulin-like receptors
Natural killer cells
Polymorphism
killer cell Immunoglobulin-like receptor
two extracellular domains, long cytoplasmic tail
Abstract: Natural killer cell stimulatory receptor gene, KIR2DS5, is polymorphic. While KIR2DS5*002 is most frequently observed, other alleles have also been found. The proteins encoded by these alleles (KIR2DS5*002-*009) are expressed at varying levels on the surface of NKL and Jurkat transfectants. Gel electrophoresis of all allelic products showed two isoforms which differ in the extent of maturation of N-linked glycosylation. These isoforms differed in intensity and molecular weight among the allelic products. Site-directed mutagenesis was used to identify polymorphic variation at residues 123 and 157 as key in altering glycosylation and levels of surface expression.
Notes: 1879-1166
Steiner, Noriko K
Dakshanamurthy, Sivanesan
Nguyen, Nicholas
Hurley, Carolyn Katovich
P30 CA051008/CA/NCI NIH HHS/United States
CA051008/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Hum Immunol. 2014 Feb;75(2):124-8. doi: 10.1016/j.humimm.2013.11.012. Epub 2013 Nov 20.
Author Address: Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057, USA.
Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057, USA. Electronic address: hurleyc@georgetown.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1842
Author: Stelzl, T., Geillinger-Kästle, K. E., Stolz, J. and Daniel, H.
Year: 2017
Title: Glycans in the intestinal peptide transporter PEPT1 contribute to function and protect from proteolysis
Journal: Am J Physiol Gastrointest Liver Physiol
Volume: 312
Issue: 6
Pages: G580-g591
Epub Date: 2017/03/25
Date: Jun 1
Short Title: Glycans in the intestinal peptide transporter PEPT1 contribute to function and protect from proteolysis
Alternate Journal: American journal of physiology. Gastrointestinal and liver physiology
ISSN: 0193-1857
DOI: 10.1152/ajpgi.00343.2016
Accession Number: 28336547
Keywords: Animals
Biological Transport
Biotinylation
Cell Line
Dipeptides/*metabolism
Endopeptidase K/*metabolism
Epithelial Cells/*metabolism
Glycosylation
Intestinal Mucosa/*metabolism
Kinetics
Membrane Potentials
Mice
Mutation
Peptide Transporter 1
*Protein Processing, Post-Translational
Protein Stability
*Proteolysis
Symporters/genetics/*metabolism
Transfection
Trypsin/metabolism
Xenopus laevis
N-linked glycosylation
glycoprotein
peptide transport
Abstract: Despite the fact that many membrane proteins carry extracellular glycans, little is known about whether the glycan chains also affect protein function. We recently demonstrated that the proton-coupled oligopeptide transporter 1 (PEPT1) in the intestine is glycosylated at six asparagine residues (N50, N406, N439, N510, N515, and N532). Mutagenesis-induced disruption of the individual N-glycosylation site N50, which is highly conserved among mammals, was detected to significantly enhance the PEPT1-mediated inward transport of peptides. Here, we show that for the murine protein the inhibition of glycosylation at sequon N50 by substituting N50 with glutamine, lysine, or cysteine or by replacing S52 with alanine equally altered PEPT1 transport kinetics in oocytes. Furthermore, we provide evidence that the uptake of [(14)C]glycyl-sarcosine in immortalized murine small intestinal (MODE-K) or colonic epithelial (PTK-6) cells stably expressing the PEPT1 transporter N50Q is also significantly increased relative to the wild-type protein. By using electrophysiological recordings and tracer flux studies, we further demonstrate that the rise in transport velocity observed for PEPT1 N50Q is bidirectional. In line with these findings, we show that attachment of biotin derivatives, comparable in weight with two to four monosaccharides, to the PEPT1 N50C transporter slows down the transport velocity. In addition, our experiments provide strong evidence that glycosylation of PEPT1 confers resistance against proteolytic cleavage by proteinase K, whereas a remarkable intrinsic stability against trypsin, even in the absence of N-linked glycans, was detected.NEW & NOTEWORTHY This study highlights the role of N50-linked glycans in modulating the bidirectional transport activity of the murine peptide transporter PEPT1. Electrophysiological and tracer flux measurements in Xenopus oocytes have shown that removal of the N50 glycans increases the maximal peptide transport rate in the inward and outward directions. This effect could be largely reversed by replacement of N50 glycans with structurally dissimilar biotin derivatives. In addition, N-glycans were detected to stabilize PEPT1 against proteolytic cleavage.
Notes: 1522-1547
Stelzl, Tamara
Geillinger-Kästle, Kerstin E
Stolz, Jürgen
Daniel, Hannelore
Journal Article
United States
Am J Physiol Gastrointest Liver Physiol. 2017 Jun 1;312(6):G580-G591. doi: 10.1152/ajpgi.00343.2016. Epub 2017 Mar 23.
Author Address: Nutritional Physiology, Technische Universität München, Freising, Germany.
Nutritional Physiology, Technische Universität München, Freising, Germany hannelore.daniel@tum.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1259
Author: Stiegler, A. L., Burden, S. J. and Hubbard, S. R.
Year: 2009
Title: Crystal structure of the frizzled-like cysteine-rich domain of the receptor tyrosine kinase MuSK
Journal: J Mol Biol
Volume: 393
Issue: 1
Pages: 1-9
Epub Date: 2009/08/12
Date: Oct 16
Short Title: Crystal structure of the frizzled-like cysteine-rich domain of the receptor tyrosine kinase MuSK
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2009.07.091
PMCID: PMC2754272
NIHMSID: NIHMS143106
Accession Number: 19664639
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Dimerization
Disulfides
Models, Molecular
Molecular Sequence Data
Protein Structure, Quaternary
Protein Structure, Tertiary
Rats
Receptor Protein-Tyrosine Kinases/*chemistry
Receptors, Cholinergic/*chemistry
Sequence Alignment
Abstract: Muscle-specific kinase (MuSK) is an essential receptor tyrosine kinase for the establishment and maintenance of the neuromuscular junction (NMJ). Activation of MuSK by agrin, a neuronally derived heparan-sulfate proteoglycan, and LRP4 (low-density lipoprotein receptor-related protein-4), the agrin receptor, leads to clustering of acetylcholine receptors on the postsynaptic side of the NMJ. The ectodomain of MuSK comprises three immunoglobulin-like domains and a cysteine-rich domain (Fz-CRD) related to those in Frizzled proteins, the receptors for Wnts. Here, we report the crystal structure of the MuSK Fz-CRD at 2.1 A resolution. The structure reveals a five-disulfide-bridged domain similar to CRDs of Frizzled proteins but with a divergent C-terminal region. An asymmetric dimer present in the crystal structure implicates surface hydrophobic residues that may function in homotypic or heterotypic interactions to mediate co-clustering of MuSK, rapsyn, and acetylcholine receptors at the NMJ.
Notes: 1089-8638
Stiegler, Amy L
Burden, Steven J
Hubbard, Stevan R
R01 NS027963-11/NS/NINDS NIH HHS/United States
R01 NS036193/NS/NINDS NIH HHS/United States
R01 NS053414-04/NS/NINDS NIH HHS/United States
T32 GM066704/GM/NIGMS NIH HHS/United States
R01 NS053414/NS/NINDS NIH HHS/United States
R01 NS027963/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Mol Biol. 2009 Oct 16;393(1):1-9. doi: 10.1016/j.jmb.2009.07.091. Epub 2009 Aug 4.
Author Address: Structural Biology Program, Kimmel Center for Biology and Medicine of the Skirball Institute, New York University School of Medicine, New York, NY 10016, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 710
Author: Stimson, E., Hope, J., Chong, A. and Burlingame, A. L.
Year: 1999
Title: Site-specific characterization of the N-linked glycans of murine prion protein by high-performance liquid chromatography/electrospray mass spectrometry and exoglycosidase digestions
Journal: Biochemistry
Volume: 38
Issue: 15
Pages: 4885-95
Epub Date: 1999/04/14
Date: Apr 13
Short Title: Site-specific characterization of the N-linked glycans of murine prion protein by high-performance liquid chromatography/electrospray mass spectrometry and exoglycosidase digestions
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi982330q
Accession Number: 10200178
Keywords: Amino Acid Sequence
Animals
Asparagine/metabolism
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Glycoside Hydrolases/*metabolism
Hydrolysis
Mass Spectrometry
Mice
Molecular Sequence Data
Polysaccharides/chemistry/*metabolism
Prions/chemistry/*metabolism
Abstract: The murine prion protein PrP gene encodes a protein of 254 amino acids with two consensus sites for Asn-linked glycosylation at codons 180 and 196. A partial site-specific study of the N-linked glycans from hamster PrP has previously been carried out by mass spectrometry [Stahl, N., Baldwin, M. A., Teplow, D. B., Hood, L., Gibson, B. W., Burlingame, A. L., and Prusiner, S. B. (1993) Biochemistry 32, 1991-2002] and revealed that the glycosylation at Asn-181 (equivalent to mouse 180) is heterogeneous, comprising over 30 glycoforms. The identification of the glycosylated peptide spanning Asn-197 was not reported. Recent technical advances in electrospray mass spectrometry now provide the sensitivity to detect low femtomole quantities of glycopeptides with >5000 mass resolution and 30 ppm mass measurement [Medzihradszky, K. F., Besman, M. J., and Burlingame, A. L. (1998) Rapid Commun. Mass Spectrom. 12, 472-478]. This performance coupled with stepwise exoglycosidase digestion has been employed to establish the differential nature of the structural complexity (glycoforms) of the glycans at Asn-180 and Asn-196 from a single strain infected with the ME7 strain. Some sixty structures have been found characterized by neutral and sialylated bi-, tri-, and tetraantennary complex-type bearing outer-arm alpha(1-3)-fucosylation (the Lewisx and sialyl-Lewisx epitopes), core alpha(1,6) fucosylation, and the presence of terminal HexNAc residues. The Lewisx trisaccharide is the major nonreducing structure at Asn-180, and significant amounts of both Lewisx and sialyl Lewisx epitopes are observed at Asn-196. The abundance of the Lewisx and sialyl Lewisx epitopes on murine PrPSc may indicate a role for these structures in the normal function of PrPC or the pathophysiology of PrPSc.
Notes: Stimson, E
Hope, J
Chong, A
Burlingame, A L
RR 01614/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1999 Apr 13;38(15):4885-95. doi: 10.1021/bi982330q.
Author Address: Ludwig Institute for Cancer Research, 91 Riding House Street, London, U.K.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1092
Author: Storch, S., Pohl, S., Quitsch, A., Falley, K. and Braulke, T.
Year: 2007
Title: C-terminal prenylation of the CLN3 membrane glycoprotein is required for efficient endosomal sorting to lysosomes
Journal: Traffic
Volume: 8
Issue: 4
Pages: 431-44
Epub Date: 2007/02/09
Date: Apr
Short Title: C-terminal prenylation of the CLN3 membrane glycoprotein is required for efficient endosomal sorting to lysosomes
Alternate Journal: Traffic (Copenhagen, Denmark)
ISSN: 1398-9219 (Print)
1398-9219
DOI: 10.1111/j.1600-0854.2007.00537.x
Accession Number: 17286803
Keywords: Animals
COS Cells
Chlorocebus aethiops
Endosomes/*physiology
Humans
Lysosomes/*metabolism
Membrane Glycoproteins/chemistry/*metabolism/physiology
Molecular Chaperones/*metabolism/physiology
Peptide Fragments/chemistry/*metabolism/physiology
Protein Prenylation/*physiology
Protein Sorting Signals/*physiology
Protein Structure, Tertiary/genetics
Protein Transport/physiology
Abstract: Mutations in the polytopic lysosomal membrane glycoprotein CLN3 result in a severe neurodegenerative disorder. Previous studies identified two cytosolic signal structures contributing to lysosomal targeting. We now examined the role of glycosylation and the C-terminal CAAX motif in lysosomal transport of CLN3 in non-neuronal and neuronal cells. Mutational analysis revealed that in COS7 cells, CLN3 is glycosylated at asparagine residues 71 and 85. Both partially and non-glycosylated CLN3 were transported correctly to lysosomes. Mevalonate incorporation and farnesyltransferase inhibitor studies indicate that CLN3 is prenylated most likely at cysteine 435. Substitution of cysteine 435 reduced the steady-state level of CLN3 in lysosomes most likely because of impaired sorting in early endosomal structures, particularly in neuronal cells. Additionally, the cell surface expression of CLN3 was increased in the presence of farnesyltransferase inhibitors. Alteration of the spacing between the transmembrane domain and the CAAX motif or the substitution of the entire C-terminal domain of CLN3 with cytoplasmic tails of mannose 6-phosphate receptors have demonstrated the importance of the C-terminal domain of proper length and composition for exit of the endoplasmic reticulum. The data suggest that co-operative signal structures in different cytoplasmic domains of CLN3 are required for efficient sorting and for transport to the lysosome.
Notes: Storch, Stephan
Pohl, Sandra
Quitsch, Arne
Falley, Katrin
Braulke, Thomas
Journal Article
Research Support, Non-U.S. Gov't
England
Traffic. 2007 Apr;8(4):431-44. doi: 10.1111/j.1600-0854.2007.00537.x. Epub 2007 Feb 7.
Author Address: Department of Biochemistry, Children's Hospital, University Hospital Hamburg, Martinistr. 52, Bldg. W 23, 20246 Hamburg, Germany. storch@uke.uni-hamburg.de
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 994
Author: Sträter, N., Jasper, B., Scholte, M., Krebs, B., Duff, A. P., Langley, D. B., Han, R., Averill, B. A., Freeman, H. C. and Guss, J. M.
Year: 2005
Title: Crystal structures of recombinant human purple Acid phosphatase with and without an inhibitory conformation of the repression loop
Journal: J Mol Biol
Volume: 351
Issue: 1
Pages: 233-46
Epub Date: 2005/07/05
Date: Aug 5
Short Title: Crystal structures of recombinant human purple Acid phosphatase with and without an inhibitory conformation of the repression loop
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2005.04.014
Accession Number: 15993892
Keywords: Acid Phosphatase/*chemistry
Cloning, Molecular
Crystallization
*Crystallography, X-Ray
Glycoproteins/*chemistry
Humans
Protein Binding
Protein Conformation
Recombinant Proteins
Abstract: The crystal structure of human purple acid phosphatase recombinantly expressed in Escherichia coli (rHPAP(Ec)) and Pichia pastoris (rHPAP(Pp)) has been determined in two different crystal forms, both at 2.2A resolution. In both cases, the enzyme crystallized in its oxidized (inactive) state, in which both Fe atoms in the dinuclear active site are Fe(III). The main difference between the two structures is the conformation of the enzyme "repression loop". Proteolytic cleavage of this loop in vivo or in vitro results in significant activation of the mammalian PAPs. In the crystals obtained from rHPAP(Ec), the carboxylate side-chain of Asp145 of this loop acts as a bidentate ligand that bridges the two metal atoms, in a manner analogous to a possible binding mode for a phosphate ester substrate in the enzyme-substrate complex. The carboxylate side-chain of Asp145 and the neighboring Phe146 side-chain thus block the active site, thereby inactivating the enzyme. In the crystal structure of rHPAP(Pp), the enzyme "repression loop" has an open conformation similar to that observed in other mammalian PAP structures. The present structures demonstrate that the repression loop exhibits significant conformational flexibility, and the observed alternate binding mode suggests a possible inhibitory role for this loop.
Notes: Sträter, Norbert
Jasper, Beate
Scholte, Marcel
Krebs, Bernt
Duff, Anthony P
Langley, David B
Han, Runlin
Averill, Bruce A
Freeman, Hans C
Guss, J Mitchell
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2005 Aug 5;351(1):233-46. doi: 10.1016/j.jmb.2005.04.014. Epub 2005 Apr 26.
Author Address: Biotechnologisch-Biomedizinisches Zentrum, Fakultät für Chemie und Mineralogie der Universität Leipzig, Deutscher Platz 5, 04103 Leipzig, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 771
Author: Stroop, C. J., Weber, W., Gerwig, G. J., Nimtz, M., Kamerling, J. P. and Vliegenthart, J. F.
Year: 2000
Title: Characterization of the carbohydrate chains of the secreted form of the human epidermal growth factor receptor
Journal: Glycobiology
Volume: 10
Issue: 9
Pages: 901-17
Epub Date: 2000/09/16
Date: Sep
Short Title: Characterization of the carbohydrate chains of the secreted form of the human epidermal growth factor receptor
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/10.9.901
Accession Number: 10988252
Keywords: Amidohydrolases/metabolism
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
ErbB Receptors/*chemistry/genetics/isolation & purification/*metabolism
Glycosylation
Humans
Magnetic Resonance Spectroscopy
Membrane Glycoproteins/*chemistry/genetics/isolation & purification/*metabolism
Methylation
Molecular Sequence Data
Oligosaccharides/*analysis/*chemistry/metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Solubility
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tumor Cells, Cultured
Abstract: The human epidermal growth factor receptor (EGFR) is a transmembrane glycoprotein having 11 potential N-glycosylation sites in its extracellular domain. N-Glycosylation is needed for proper membrane insertion, EGF binding and receptor functioning. The human epidermoid carcinoma A431 cell line secretes a soluble 105 kDa glycoprotein (sEGFR) that represents the extracellular domain of the membrane-bound form, and its glycosylation pattern has been investigated. After liberation of the oligosaccharides from sEGFR with PNGase F, the glycans were fractionated along different routes, including Concanavalin A affinity chromatography, anion-exchange chromatography, HPLC and high-pH anion-exchange chromatography. The oligosaccharide fractions were characterized by 500- and 600-MHz 1H-NMR spectroscopy and mass spectrometry (FAB, ESI, and MALDI-TOF). The oligomannose-type glycans range from Man5GlcNAc2 to Man8GlcNAc2 and account for 17% of the total carbohydrate moiety. Furthermore, di-, tri'- and tetraantennary complex-type structures are present, both neutral and (alpha2-3)-sialylated (up to tetrasialo), comprising 24 and 59%, respectively, of the total carbohydrate moiety. In this study, 32 new complex-type glycans are characterized containing the Le(x), Le(Y), and sialyl-Le(x) determinants, the bloodgroup A and H antigens, as well as the ALe(Y) determinant. This first comprehensive glycosylation study on a human nonrecombinant receptor shows the immense heterogeneity of the glycosylation of sEGFR.
Notes: Stroop, C J
Weber, W
Gerwig, G J
Nimtz, M
Kamerling, J P
Vliegenthart, J F
Journal Article
England
Glycobiology. 2000 Sep;10(9):901-17. doi: 10.1093/glycob/10.9.901.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 958
Author: Strop, P., Bankovich, A. J., Hansen, K. C., Garcia, K. C. and Brunger, A. T.
Year: 2004
Title: Structure of a human A-type potassium channel interacting protein DPPX, a member of the dipeptidyl aminopeptidase family
Journal: J Mol Biol
Volume: 343
Issue: 4
Pages: 1055-65
Epub Date: 2004/10/13
Date: Oct 29
Short Title: Structure of a human A-type potassium channel interacting protein DPPX, a member of the dipeptidyl aminopeptidase family
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/j.jmb.2004.09.003
Accession Number: 15476821
Keywords: Alternative Splicing
Catalytic Domain
Cystine/chemistry/genetics/metabolism
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases/*chemistry/metabolism
Glycosylation
Humans
Isoenzymes/chemistry/metabolism
Potassium Channels, Voltage-Gated/*metabolism
Protein Structure, Quaternary
Protein Structure, Tertiary
Shal Potassium Channels
Static Electricity
Abstract: It has recently been reported that dipeptidyl aminopeptidase X (DPPX) interacts with the voltage-gated potassium channel Kv4 and that co-expression of DPPX together with Kv4 pore forming alpha-subunits, and potassium channel interacting proteins (KChIPs), reconstitutes properties of native A-type potassium channels in vitro. Here we report the X-ray crystal structure of the extracellular domain of human DPPX determined at 3.0A resolution. This structure reveals the potential for a surface electrostatic change based on the protonation state of histidine. Subtle changes in extracellular pH might modulate the interaction of DPPX with Kv4.2 and possibly with other proteins. We propose models of DPPX interaction with the voltage-gated potassium channel complex. The dimeric structure of DPPX is highly homologous to the related protein DPP-IV. Comparison of the active sites of DPPX and DPP-IV reveals loss of the catalytic serine residue but the presence of an additional serine near the "active" site. However, the arrangement of residues is inconsistent with that of canonical serine proteases and DPPX is unlikely to function as a protease (dipeptidyl aminopeptidase).
Notes: Strop, Pavel
Bankovich, Alexander J
Hansen, Kirk C
Garcia, K Christopher
Brunger, Axel T
T32 AI007290/AI/NIAID NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
England
J Mol Biol. 2004 Oct 29;343(4):1055-65. doi: 10.1016/j.jmb.2004.09.003.
Author Address: Howard Hughes Medical Institute and Department of Molecular and Cellular Physiology, Stanford Synchrotron Radiation Laboratory, Stanford University, James H. Clark Center E300, 318 Campus Drive, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1557
Author: Strum, J. S., Nwosu, C. C., Hua, S., Kronewitter, S. R., Seipert, R. R., Bachelor, R. J., An, H. J. and Lebrilla, C. B.
Year: 2013
Title: Automated assignments of N- and O-site specific glycosylation with extensive glycan heterogeneity of glycoprotein mixtures
Journal: Anal Chem
Volume: 85
Issue: 12
Pages: 5666-75
Epub Date: 2013/05/15
Date: Jun 18
Short Title: Automated assignments of N- and O-site specific glycosylation with extensive glycan heterogeneity of glycoprotein mixtures
Alternate Journal: Analytical chemistry
ISSN: 0003-2700 (Print)
0003-2700
DOI: 10.1021/ac4006556
PMCID: PMC3692395
NIHMSID: NIHMS485606
Accession Number: 23662732
Keywords: Amino Acid Sequence
Animals
Binding Sites/physiology
Cattle
Chromatography, Liquid/methods
Glycoproteins/analysis/genetics/*metabolism
Glycosylation
Humans
Molecular Sequence Data
Nitrogen/analysis/*metabolism
Oxygen/analysis/*metabolism
Polysaccharides/analysis/genetics/*metabolism
Random Allocation
Tandem Mass Spectrometry/*methods
Abstract: Site-specific glycosylation (SSG) of glycoproteins remains a considerable challenge and limits further progress in the areas of proteomics and glycomics. Effective methods require new approaches in sample preparation, detection, and data analysis. While the field has advanced in sample preparation and detection, automated data analysis remains an important goal. A new bioinformatics approach implemented in software called GP Finder automatically distinguishes correct assignments from random matches and complements experimental techniques that are optimal for glycopeptides, including nonspecific proteolysis and high mass resolution liquid chromatography/tandem mass spectrometry (LC/MS/MS). SSG for multiple N- and O-glycosylation sites, including extensive glycan heterogeneity, was annotated for single proteins and protein mixtures with a 5% false-discovery rate, generating hundreds of nonrandom glycopeptide matches and demonstrating the proof-of-concept for a self-consistency scoring algorithm shown to be compliant with the target-decoy approach (TDA). The approach was further applied to a mixture of N-glycoproteins from unprocessed human milk and O-glycoproteins from very-low-density-lipoprotein (vLDL) particles.
Notes: 1520-6882
Strum, John S
Nwosu, Charles C
Hua, Serenus
Kronewitter, Scott R
Seipert, Richard R
Bachelor, Robert J
An, Hyun Joo
Lebrilla, Carlito B
R01 GM049077/GM/NIGMS NIH HHS/United States
S10 RR027639/RR/NCRR NIH HHS/United States
R01GM049077/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Anal Chem. 2013 Jun 18;85(12):5666-75. doi: 10.1021/ac4006556. Epub 2013 May 24.
Author Address: Department of Chemistry, University of California, Davis, California 95616, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 530
Author: Stults, J. T., O'Connell, K. L., Garcia, C., Wong, S., Engel, A. M., Garbers, D. L. and Lowe, D. G.
Year: 1994
Title: The disulfide linkages and glycosylation sites of the human natriuretic peptide receptor-C homodimer
Journal: Biochemistry
Volume: 33
Issue: 37
Pages: 11372-81
Epub Date: 1994/09/20
Date: Sep 20
Short Title: The disulfide linkages and glycosylation sites of the human natriuretic peptide receptor-C homodimer
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00203a036
Accession Number: 7727388
Keywords: Amino Acid Sequence
Animals
Baculoviridae
CHO Cells
Carbohydrate Sequence
Cell Line
Cricetinae
Cytoplasm/metabolism
Disulfides
Glycosylation
Humans
Macromolecular Substances
Mass Spectrometry
Molecular Sequence Data
Mutagenesis, Site-Directed
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/chemistry/isolation & purification
Point Mutation
Protein Conformation
Receptors, Atrial Natriuretic Factor/*chemistry/*metabolism
Recombinant Fusion Proteins/chemistry/metabolism
Sequence Homology, Amino Acid
Spodoptera
Transfection
Trypsin
Abstract: The natriuretic peptide receptor-C (NPR-C) constitutes greater than 95% of the natriuretic peptide binding sites in vivo. This cell surface glycoprotein is a disulfide-linked homodimer with a subunit molecular weight of 68,000. Two sources and types of ANP affinity-purified human NPR-C were used to map disulfide linkages and glycosylation sites of this receptor by mass spectrometry: the extracellular domain obtained by papain cleavage of a receptor-IgG fusion protein expressed in Chinese hamster ovary cells, and a baculovirus/Sf9-expressed cytoplasmic truncation mutant in which 34 of 37 cytoplasmic domain amino acids were deleted. Two intramolecular disulfide bonded loops were found in the 435 amino acid extracellular domain (C63-C91, C168-C216). The juxtamembrane residues C428 and C431 are involved in homodimer formation, confirmed by site-directed mutagenesis of full-length NPR. Three of the four potential Asn-linked glycosylation sites are occupied: N41 (complex), N248 (high mannose), and N349 (complex; partial occupancy). These data describe the intra- and intermolecular linkages in NPR-C, providing a model for the homologous guanylyl cyclase receptors, NPR-A and NPR-B; both of the cyclase receptors likely contain the first amino-terminal 29 amino acid loop, but only NPR-A possesses the second 49 amino acid loop in common with NPR-C.
Notes: Stults, J T
O'Connell, K L
Garcia, C
Wong, S
Engel, A M
Garbers, D L
Lowe, D G
Comparative Study
Journal Article
United States
Biochemistry. 1994 Sep 20;33(37):11372-81. doi: 10.1021/bi00203a036.
Author Address: Department of Protein Chemistry, Genentech, Inc., South San Francisco, California 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1924
Author: Su, Q., Hu, F., Liu, Y., Ge, X., Mei, C., Yu, S., Shen, A., Zhou, Q., Yan, C., Lei, J., Zhang, Y., Liu, X. and Wang, T.
Year: 2018
Title: Cryo-EM structure of the polycystic kidney disease-like channel PKD2L1
Journal: Nat Commun
Volume: 9
Issue: 1
Pages: 1192
Epub Date: 2018/03/24
Date: Mar 22
Short Title: Cryo-EM structure of the polycystic kidney disease-like channel PKD2L1
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/s41467-018-03606-0
PMCID: PMC5864754
Accession Number: 29567962
Keywords: Animals
Calcium Channels/*chemistry/genetics/metabolism/ultrastructure
Cryoelectron Microscopy
Humans
Mice
Protein Conformation, alpha-Helical
Protein Domains
Receptors, Cell Surface/*chemistry/genetics/metabolism/ultrastructure
TRPP Cation Channels/chemistry/genetics/metabolism
Abstract: PKD2L1, also termed TRPP3 from the TRPP subfamily (polycystic TRP channels), is involved in the sour sensation and other pH-dependent processes. PKD2L1 is believed to be a nonselective cation channel that can be regulated by voltage, protons, and calcium. Despite its considerable importance, the molecular mechanisms underlying PKD2L1 regulations are largely unknown. Here, we determine the PKD2L1 atomic structure at 3.38 Å resolution by cryo-electron microscopy, whereby side chains of nearly all residues are assigned. Unlike its ortholog PKD2, the pore helix (PH) and transmembrane segment 6 (S6) of PKD2L1, which are involved in upper and lower-gate opening, adopt an open conformation. Structural comparisons of PKD2L1 with a PKD2-based homologous model indicate that the pore domain dilation is coupled to conformational changes of voltage-sensing domains (VSDs) via a series of π-π interactions, suggesting a potential PKD2L1 gating mechanism.
Notes: 2041-1723
Su, Qiang
Hu, Feizhuo
Liu, Yuxia
Ge, Xiaofei
Mei, Changlin
Yu, Shengqiang
Shen, Aiwen
Zhou, Qiang
Yan, Chuangye
Lei, Jianlin
Orcid: 0000-0002-9384-8742
Zhang, Yanqing
Liu, Xiaodong
Wang, Tingliang
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2018 Mar 22;9(1):1192. doi: 10.1038/s41467-018-03606-0.
Author Address: Ministry of Education Key Laboratory of Protein Science, Tsinghua University, Beijing, 100084, China.
School of Life Sciences, Tsinghua University, Beijing, 100084, China.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua University, Beijing, 100084, China.
School of Medicine, Tsinghua University, Beijing, 100084, China.
X-Lab for Transmembrane Signaling Research, Department of Biomedical Engineering and McGovern Institute for Brain Research, Tsinghua University, Beijing, 100084, China.
School of Biological Science and Medical Engineering, Beihang University, Beijing, 100083, China.
Beijing Advanced Innovation Center for Biomedical Engineering, Beihang University, Beijing, China, 102402.
Department of Nephrology, Changzheng Hospital, Second Military Medical University, Shanghai, 200433, China.
Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing, 100084, China.
Ministry of Education Key Laboratory of Protein Science, Tsinghua University, Beijing, 100084, China. zhangyanqing15@mail.tsinghua.edu.cn.
School of Life Sciences, Tsinghua University, Beijing, 100084, China. zhangyanqing15@mail.tsinghua.edu.cn.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua University, Beijing, 100084, China. zhangyanqing15@mail.tsinghua.edu.cn.
Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing, 100084, China. zhangyanqing15@mail.tsinghua.edu.cn.
School of Life Sciences, Tsinghua University, Beijing, 100084, China. liu-lab@vip.163.com.
School of Medicine, Tsinghua University, Beijing, 100084, China. liu-lab@vip.163.com.
X-Lab for Transmembrane Signaling Research, Department of Biomedical Engineering and McGovern Institute for Brain Research, Tsinghua University, Beijing, 100084, China. liu-lab@vip.163.com.
School of Biological Science and Medical Engineering, Beihang University, Beijing, 100083, China. liu-lab@vip.163.com.
Beijing Advanced Innovation Center for Biomedical Engineering, Beihang University, Beijing, China, 102402. liu-lab@vip.163.com.
Ministry of Education Key Laboratory of Protein Science, Tsinghua University, Beijing, 100084, China. wangtingliang@mail.tsinghua.edu.cn.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua University, Beijing, 100084, China. wangtingliang@mail.tsinghua.edu.cn.
School of Medicine, Tsinghua University, Beijing, 100084, China. wangtingliang@mail.tsinghua.edu.cn.
Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing, 100084, China. wangtingliang@mail.tsinghua.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1945
Author: Sudom, A., Talreja, S., Danao, J., Bragg, E., Kegel, R., Min, X., Richardson, J., Zhang, Z., Sharkov, N., Marcora, E., Thibault, S., Bradley, J., Wood, S., Lim, A. C., Chen, H., Wang, S., Foltz, I. N., Sambashivan, S. and Wang, Z.
Year: 2018
Title: Molecular basis for the loss-of-function effects of the Alzheimer's disease-associated R47H variant of the immune receptor TREM2
Journal: J Biol Chem
Volume: 293
Issue: 32
Pages: 12634-12646
Epub Date: 2018/05/26
Date: Aug 10
Short Title: Molecular basis for the loss-of-function effects of the Alzheimer's disease-associated R47H variant of the immune receptor TREM2
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA118.002352
PMCID: PMC6093241
Accession Number: 29794134
Keywords: Alzheimer Disease/*genetics/pathology
Crystallography, X-Ray
Dendritic Cells/chemistry/pathology
*Genetic Predisposition to Disease
Genetic Variation
Humans
Ligands
Macrophages/chemistry/pathology
Membrane Glycoproteins/*chemistry/genetics
Microglia/chemistry/pathology
Mutant Proteins/*chemistry/genetics
Nerve Tissue Proteins/chemistry/genetics
Osteoclasts/chemistry/pathology
Protein Conformation
Protein Domains/genetics
Receptors, Immunologic/*chemistry/genetics
*Alzheimer disease
*R47h
*Trem2
*X-ray crystallography
*antibody
*lipid-binding protein
*microglia
*myeloid cell
*neurodegeneration
*neuroinflammation
*scavenger receptor
*structure
*triggering receptor expressed on myeloid cells 2
time the research was conducted
Abstract: Triggering receptor expressed on myeloid cells 2 (TREM2) is an immune receptor expressed on the surface of microglia, macrophages, dendritic cells, and osteoclasts. The R47H TREM2 variant is a significant risk factor for late-onset Alzheimer's disease (AD), and the molecular basis of R47H TREM2 loss of function is an emerging area of TREM2 biology. Here, we report three high-resolution structures of the extracellular ligand-binding domains (ECDs) of R47H TREM2, apo-WT, and phosphatidylserine (PS)-bound WT TREM2 at 1.8, 2.2, and 2.2 Å, respectively. The structures reveal that Arg(47) plays a critical role in maintaining the structural features of the complementarity-determining region 2 (CDR2) loop and the putative positive ligand-interacting surface (PLIS), stabilizing conformations capable of ligand interaction. This is exemplified in the PS-bound structure, in which the CDR2 loop and PLIS drive critical interactions with PS via surfaces that are disrupted in the variant. Together with in vitro and in vivo characterization, our structural findings elucidate the molecular mechanism underlying loss of ligand binding, putative oligomerization, and functional activity of R47H TREM2. They also help unravel how decreased in vitro and in vivo stability of TREM2 contribute to loss of function in disease.
Notes: 1083-351x
Sudom, Athena
Talreja, Santosh
Danao, Jean
Bragg, Eric
Kegel, Rob
Min, Xiaoshan
Richardson, Jason
Zhang, Zhongqi
Sharkov, Nikolai
Marcora, Edoardo
Orcid: 0000-0002-3829-4927
Thibault, Steve
Bradley, Jodi
Orcid: 0000-0002-4887-778x
Wood, Steve
Lim, Ai-Ching
Chen, Hang
Wang, Songli
Foltz, Ian N
Sambashivan, Shilpa
Wang, Zhulun
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2018 Aug 10;293(32):12634-12646. doi: 10.1074/jbc.RA118.002352. Epub 2018 May 24.
Author Address: From Amgen Discovery Research, Amgen Inc., San Francisco, California 94080, asudom@amgen.com.
From Amgen Discovery Research, Amgen Inc., San Francisco, California 94080.
Amgen Discovery Research, Amgen Inc., Thousand Oaks, California 91320, and.
Amgen Discovery Research, Amgen Inc., Burnaby, British Columbia V5A 1V7, Canada.
From Amgen Discovery Research, Amgen Inc., San Francisco, California 94080, shilpashivan@gmail.com.
From Amgen Discovery Research, Amgen Inc., San Francisco, California 94080, zwang@amgen.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1421
Author: Sugi, Y., Takahashi, K., Nakano, K., Hosono, A. and Kaminogawa, S.
Year: 2011
Title: Transcription of the Tollip gene is elevated in intestinal epithelial cells through impaired O-GlcNAcylation-dependent nuclear translocation of the negative regulator Elf-1
Journal: Biochem Biophys Res Commun
Volume: 412
Issue: 4
Pages: 704-9
Epub Date: 2011/08/27
Date: Sep 9
Short Title: Transcription of the Tollip gene is elevated in intestinal epithelial cells through impaired O-GlcNAcylation-dependent nuclear translocation of the negative regulator Elf-1
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2011.08.035
Accession Number: 21867680
Keywords: Acetylglucosamine/*metabolism
Active Transport, Cell Nucleus
Acylation
Animals
Caco-2 Cells
Cell Nucleus/*metabolism
Ephrin-A2/*metabolism
Female
*Gene Expression Regulation
Humans
Intestinal Mucosa/*metabolism
Intracellular Signaling Peptides and Proteins/*genetics
Mice
Mice, Inbred BALB C
Response Elements
*Transcription, Genetic
Abstract: Intestinal epithelial cells (IECs) must be tolerant of the large number of commensal bacteria inhabiting the intestinal tract to avoid excessive inflammatory reactions. Toll-interacting protein (Tollip), a negative regulator of Toll-like receptor signaling, is known to be expressed at high levels in IECs, and to thereby contribute to the hyporesponsiveness of IECs to commensals. In this study, we analyzed the underlying mechanisms for elevated transcription of the Tollip gene in IECs using a human IEC line, Caco-2, and a human monocyte line, THP-1, as a control. Elf-1 was identified as a transcription factor that negatively regulates Tollip gene expression. The transcription factor Elf-1 was localized in the nucleus by O-linked N-acetylglucosamine (O-GlcNAc) modification, whereas the unmodified form was detected only in the cytoplasm. Comparison of Caco-2 and THP-1 cells revealed that O-GlcNAc modification of Elf-1 was significantly lower in IECs than in monocytes. Collectively, the results indicate that insufficient O-GlcNAc modification prevents Elf-1-mediated transcriptional repression and thereby upregulates Tollip gene expression in IECs.
Notes: 1090-2104
Sugi, Yutaka
Takahashi, Kyoko
Nakano, Kou
Hosono, Akira
Kaminogawa, Shuichi
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2011 Sep 9;412(4):704-9. doi: 10.1016/j.bbrc.2011.08.035. Epub 2011 Aug 16.
Author Address: College of Bioresource Sciences, Nihon University, Fujisawa-shi, Kanagawa, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 522
Author: Sugiura, T., Takamatsu, H., Kudo, A. and Amann, E.
Year: 1995
Title: Expression and characterization of murine osteoblast-specific factor 2 (OSF-2) in a baculovirus expression system
Journal: Protein Expr Purif
Volume: 6
Issue: 3
Pages: 305-11
Epub Date: 1995/06/01
Date: Jun
Short Title: Expression and characterization of murine osteoblast-specific factor 2 (OSF-2) in a baculovirus expression system
Alternate Journal: Protein expression and purification
ISSN: 1046-5928 (Print)
1046-5928
DOI: 10.1006/prep.1995.1040
Accession Number: 7663166
Keywords: Amidohydrolases/metabolism
Animals
Baculoviridae/genetics
Blotting, Western
Cell Adhesion Molecules/*biosynthesis/genetics/metabolism
Cells, Cultured
Concanavalin A/metabolism
Glycosylation
Heparin/metabolism
Mice
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Protein Binding
Recombinant Proteins/biosynthesis/metabolism
Spodoptera/cytology/virology
Tunicamycin/pharmacology
Abstract: Osteoblast-specific factor 2 (OSF-2) is a approximately 90-kDa protein selectively expressed in bone. OSF-2 cDNA was recently isolated from mouse and human cDNA libraries and shows limited sequence homology with fasciclin I, a cell adhesion protein expressed in insect nerve cells. Here we describe the expression of recombinant murine OSF-2 (rmOSF-2) in a baculovirus/insect cell system. Western blotting analysis employing polyclonal antiserum raised against a C-terminal synthetic OSF-2 peptide detected a protein of approximately 90-kDa as early as 2 days after infection of Sf9 cells with the recombinant virus. Tunicamycin treatment of infected cells resulted in a mobility shift of OSF-2 (approximately 90-kDa band) on Western blots. N-Glycanase digestion resulted in the same mobility shift of OSF-2, indicating that rmOSF-2 expressed in insect cells is N-glycosylated. However, OSF-2 was insensitive to endoglycosidase H digestion while a major fraction of this protein had affinity for concanavalin A. Finally, it was demonstrated that rmOSF-2 was able to bind to heparin. This finding suggests that OSF-2 might be associated with the bone extracellular matrix after secretion by osteoblasts and participate in cell adhesion and/or cell communication. The establishment of the baculovirus expression system with a high productivity of recombinant OSF-2 (around 40 micrograms/ml at maximum) and its heparin binding properties should allow us to obtain large amounts of rmOSF-2.
Notes: Sugiura, T
Takamatsu, H
Kudo, A
Amann, E
Journal Article
United States
Protein Expr Purif. 1995 Jun;6(3):305-11. doi: 10.1006/prep.1995.1040.
Author Address: Laboratory of Molecular Biology, Hoechst Japan Limited, Saitama.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 587
Author: Sugo, T., Mas, E., Abouakil, N., Endo, T., Escribano, M. J., Kobata, A. and Lombardo, D.
Year: 1993
Title: The structure of N-linked oligosaccharides of human pancreatic bile-salt-dependent lipase
Journal: Eur J Biochem
Volume: 216
Issue: 3
Pages: 799-805
Epub Date: 1993/09/15
Date: Sep 15
Short Title: The structure of N-linked oligosaccharides of human pancreatic bile-salt-dependent lipase
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb18200.x
Accession Number: 8404898
Keywords: Bile Acids and Salts/*chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
Chromatography, Gel
Humans
Lipase/*chemistry
Molecular Sequence Data
Oligosaccharides/analysis/*chemistry
Pancreatic Juice/*enzymology
Abstract: This study describes the structure of the N-linked oligosaccharide chains of bile-salt-dependent lipase isolated from the pancreatic juice of a normal donor. After hydrazinolysis, neutral sugar chains were separated from acidic chains by paper electrophoresis and were fractionated using serial column chromatography with immobilized lectins and Bio-Gel P-4 filtration. Structural analysis was performed by means of sequential glycosidase digestion and revealed that the neutral sugar chains are mainly of the biantennary complex type. Fucose residues were identified for some trimannosyl core structures and were alpha(1-6) or alpha(1-2) linked to the innermost GlcNAc residue and a terminal Gal residue, respectively. Sialyl residues were also involved in the oligosaccharide structures. Most of these structures have no linear N-acetyllactosamine repeats. Evidence from several approaches suggests that the sugar chains of the human pancreatic bile-salt-dependent lipase possess a blood-group-related antigenic determinant.
Notes: Sugo, T
Mas, E
Abouakil, N
Endo, T
Escribano, M J
Kobata, A
Lombardo, D
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1993 Sep 15;216(3):799-805. doi: 10.1111/j.1432-1033.1993.tb18200.x.
Author Address: Institute of Medical Science, University of Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1379
Author: Sukumaran, M., Rossmann, M., Shrivastava, I., Dutta, A., Bahar, I. and Greger, I. H.
Year: 2011
Title: Dynamics and allosteric potential of the AMPA receptor N-terminal domain
Journal: Embo j
Volume: 30
Issue: 5
Pages: 972-82
Epub Date: 2011/02/15
Date: Mar 2
Short Title: Dynamics and allosteric potential of the AMPA receptor N-terminal domain
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2011.17
PMCID: PMC3049213
Accession Number: 21317871
Keywords: *Allosteric Regulation
Amino Acid Sequence
Calcium/*metabolism
Cell Membrane/*metabolism
Crystallography, X-Ray
Electrophysiology
Humans
Ion Channels
Molecular Sequence Data
Protein Conformation
Protein Multimerization
Protein Subunits
Protein Transport
Receptors, AMPA/*chemistry/*metabolism
Sequence Homology, Amino Acid
Ultracentrifugation
alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid/*metabolism
Abstract: Glutamate-gated ion channels (ionotropic glutamate receptors, iGluRs) sense the extracellular milieu via an extensive extracellular portion, comprised of two clamshell-shaped segments. The distal, N-terminal domain (NTD) has allosteric potential in NMDA-type iGluRs, which has not been ascribed to the analogous domain in AMPA receptors (AMPARs). In this study, we present new structural data uncovering dynamic properties of the GluA2 and GluA3 AMPAR NTDs. GluA3 features a zipped-open dimer interface with unconstrained lower clamshell lobes, reminiscent of metabotropic GluRs (mGluRs). The resulting labile interface supports interprotomer rotations, which can be transmitted to downstream receptor segments. Normal mode analysis reveals two dominant mechanisms of AMPAR NTD motion: intraprotomer clamshell motions and interprotomer counter-rotations, as well as accessible interconversion between AMPAR and mGluR conformations. In addition, we detect electron density for a potential ligand in the GluA2 interlobe cleft, which may trigger lobe motions. Together, these data support a dynamic role for the AMPAR NTDs, which widens the allosteric landscape of the receptor and could provide a novel target for ligand development.
Notes: 1460-2075
Sukumaran, Madhav
Rossmann, Maxim
Shrivastava, Indira
Dutta, Anindita
Bahar, Ivet
Greger, Ingo H
R01 GM086238/GM/NIGMS NIH HHS/United States
U54GM087519/GM/NIGMS NIH HHS/United States
U54 GM087519/GM/NIGMS NIH HHS/United States
MC_U105174197/Medical Research Council/United Kingdom
R01 GM086238-04/GM/NIGMS NIH HHS/United States
5R01GM086238-02/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
EMBO J. 2011 Mar 2;30(5):972-82. doi: 10.1038/emboj.2011.17. Epub 2011 Feb 11.
Author Address: Neurobiology Division, MRC Laboratory of Molecular Biology, Cambridge, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1516
Author: Sumer-Bayraktar, Z., Nguyen-Khuong, T., Jayo, R., Chen, D. D., Ali, S., Packer, N. H. and Thaysen-Andersen, M.
Year: 2012
Title: Micro- and macroheterogeneity of N-glycosylation yields size and charge isoforms of human sex hormone binding globulin circulating in serum
Journal: Proteomics
Volume: 12
Issue: 22
Pages: 3315-27
Epub Date: 2012/09/25
Date: Nov
Short Title: Micro- and macroheterogeneity of N-glycosylation yields size and charge isoforms of human sex hormone binding globulin circulating in serum
Alternate Journal: Proteomics
ISSN: 1615-9853
DOI: 10.1002/pmic.201200354
Accession Number: 23001782
Keywords: Amino Acid Sequence
Electrophoresis, Gel, Two-Dimensional
Glycomics/methods
Glycoproteins/blood/*chemistry/*metabolism
Glycosylation
Humans
Molecular Sequence Data
Polysaccharides/analysis/*chemistry/*metabolism
Protein Isoforms
Proteomics/methods
Sex Hormone-Binding Globulin/*chemistry/*metabolism
Abstract: Human sex hormone binding globulin (hSHBG) is a serum glycoprotein central to the transport and targeted delivery of sex hormones to steroid-sensitive tissues. Several molecular mechanisms of action of hSHBG, including the function of its attached glycans remain unknown. Here, we perform a detailed site-specific characterization of the N- and O-linked glycosylation of serum-derived hSHBG. MS-driven glycoproteomics and glycomics combined with exoglycosidase treatment were used in a bottom-up and top-down manner to determine glycosylation sites, site-specific occupancies and monosaccharide compositions, detailed glycan structures, and the higher level arrangement of glycans on intact hSHBG. It was found that serum-derived hSHBG is N-glycosylated at Asn(351) and Asn(367) with average molar occupancies of 85.1 and 95.3%, respectively. Both sites are occupied by the same six sialylated and partly core fucosylated bi- and triantennary N-Glycoforms with lactosamine-type antennas of the form (±NeuAcα6)Galβ4GlcNAc. N-Glycoforms of Asn(367) were slightly more branched and core fucosylated than Asn(351) N-glycoforms due probably to a more surface-exposed glycosylation site. The N-terminal Thr(7) was fully occupied by the two O-linked glycans NeuAcα3Galβ3(NeuAcα6)GalNAc (where NeuAc is N-acetylneuraminic acid and GalNAc is N-acetylgalactosamine) and NeuAcα3Galβ3GalNAc in a 1:6 molar ratio. Electrophoretic analysis of intact hSHBG revealed size and charge heterogeneity of the isoforms circulating in blood serum. Interestingly, the size and charge heterogeneity were shown to originate predominantly from differential Asn(351) glycan occupancies and N-glycan sialylation that may modulate the hSHBG activity. To date, this work represents the most detailed structural map of the heterogeneous hSHBG glycosylation, which is a prerequisite for investigating the functional aspects of the hSHBG glycans.
Notes: 1615-9861
Sumer-Bayraktar, Zeynep
Nguyen-Khuong, Terry
Jayo, Roxana
Chen, David D Y
Ali, Sinan
Packer, Nicolle H
Thaysen-Andersen, Morten
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2012 Nov;12(22):3315-27. doi: 10.1002/pmic.201200354. Epub 2012 Oct 29.
Author Address: Department of Chemistry and Biomolecular Sciences, Macquarie University, Sydney, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2048
Author: Sun, C., Lan, W., Li, B., Zuo, R., Xing, H., Liu, M., Li, J., Yao, Y., Wu, J., Tang, Y., Liu, H. and Zhou, Y.
Year: 2019
Title: Glucose regulates tissue-specific chondro-osteogenic differentiation of human cartilage endplate stem cells via O-GlcNAcylation of Sox9 and Runx2
Journal: Stem Cell Res Ther
Volume: 10
Issue: 1
Pages: 357
Epub Date: 2019/11/30
Date: Nov 28
Short Title: Glucose regulates tissue-specific chondro-osteogenic differentiation of human cartilage endplate stem cells via O-GlcNAcylation of Sox9 and Runx2
Alternate Journal: Stem cell research & therapy
ISSN: 1757-6512
DOI: 10.1186/s13287-019-1440-5
PMCID: PMC6883626
Accession Number: 31779679
Keywords: Adult
Cartilage/cytology/*metabolism
Cell Differentiation/*drug effects
Chondrogenesis/*drug effects
Core Binding Factor Alpha 1 Subunit/*metabolism
Female
Glucose/*pharmacology
Glycosylation
Growth Plate/cytology/*metabolism
Humans
Male
Middle Aged
Osteogenesis/*drug effects
SOX9 Transcription Factor/*metabolism
Stem Cells/cytology/*metabolism
*Cartilage endplate stem cells
*Chondrogenic differentiation
*Glucose
*O-GlcNAcylation
*Osteogenic differentiation
Abstract: BACKGROUND: The degenerative disc disease (DDD) is a major cause of low back pain. The physiological low-glucose microenvironment of the cartilage endplate (CEP) is disrupted in DDD. Glucose influences protein O-GlcNAcylation via the hexosamine biosynthetic pathway (HBP), which is the key to stem cell fate. Thiamet-G is an inhibitor of O-GlcNAcase for accumulating O-GlcNAcylated proteins while 6-diazo-5-oxo-L-norleucine (DON) inhibits HBP. Mechanisms of DDD are incompletely understood but include CEP degeneration and calcification. We aimed to identify the molecular mechanisms of glucose in CEP calcification in DDD. METHODS: We assessed normal and degenerated CEP tissues from patients, and the effects of chondrogenesis and osteogenesis of the CEP were determined by western blot and immunohistochemical staining. Cartilage endplate stem cells (CESCs) were induced with low-, normal-, and high-glucose medium for 21 days, and chondrogenic and osteogenic differentiations were measured by Q-PCR, western blot, and immunohistochemical staining. CESCs were induced with low-glucose and high-glucose medium with or without Thiamet-G or DON for 21 days, and chondrogenic and osteogenic differentiations were measured by Q-PCR, western blot, and immunohistochemical staining. Sox9 and Runx2 O-GlcNAcylation were measured by immunofluorescence. The effects of O-GlcNAcylation on the downstream genes of Sox9 and Runx2 were determined by Q-PCR and western blot. RESULTS: Degenerated CEPs from DDD patients lost chondrogenesis, acquired osteogenesis, and had higher protein O-GlcNAcylation level compared to normal CEPs from LVF patients. CESC chondrogenic differentiation gradually decreased while osteogenic differentiation gradually increased from low- to high-glucose differentiation medium. Furthermore, Thiamet-G promoted CESC osteogenic differentiation and inhibited chondrogenic differentiation in low-glucose differentiation medium; however, DON acted opposite role in high-glucose differentiation medium. Interestingly, we found that Sox9 and Runx2 were O-GlcNAcylated in differentiated CESCs. Finally, O-GlcNAcylation of Sox9 and Runx2 decreased chondrogenesis and increased osteogenesis in CESCs. CONCLUSIONS: Our findings demonstrate the effect of glucose concentration on regulating the chondrogenic and osteogenic differentiation potential of CESCs and provide insight into the mechanism of how glucose concentration regulates Sox9 and Runx2 O-GlcNAcylation to affect the differentiation of CESCs, which may represent a target for CEP degeneration therapy.
Notes: 1757-6512
Sun, Chao
Orcid: 0000-0002-7238-9698
Lan, Weiren
Li, Bin
Zuo, Rui
Xing, Hui
Liu, Minghan
Li, Jie
Yao, Yuan
Wu, Junlong
Tang, Yu
Liu, Huan
Zhou, Yue
Journal Article
Research Support, Non-U.S. Gov't
Stem Cell Res Ther. 2019 Nov 28;10(1):357. doi: 10.1186/s13287-019-1440-5.
Author Address: Department of Orthopedics, Xinqiao Hospital, Army Medical University, Chongqing, 400038, People's Republic of China.
Department of Orthopedics, Xinqiao Hospital, Army Medical University, Chongqing, 400038, People's Republic of China. tangyu628@sina.com.
Department of Orthopaedics, The Second Affiliated Hospital of Southwest Medical University, Lu Zhou, 646000, Sichuan, People's Republic of China. 20016040@163.com.
Department of Orthopedics, Xinqiao Hospital, Army Medical University, Chongqing, 400038, People's Republic of China. happyzhou@vip.163.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2062
Author: Sun, Q. H., Wang, Y. S., Liu, G., Zhou, H. L., Jian, Y. P., Liu, M. D., Zhang, D., Ding, Q., Zhao, R. X., Chen, J. F., Li, Y. N., Liang, J., Li, Y. L., Quan, C. S. and Xu, Z. X.
Year: 2020
Title: Enhanced O-linked Glcnacylation in Crohn's disease promotes intestinal inflammation
Journal: EBioMedicine
Volume: 53
Pages: 102693
Epub Date: 2020/03/03
Date: Mar
Short Title: Enhanced O-linked Glcnacylation in Crohn's disease promotes intestinal inflammation
Alternate Journal: EBioMedicine
ISSN: 2352-3964
DOI: 10.1016/j.ebiom.2020.102693
PMCID: PMC7047186
Accession Number: 32114385
Keywords: Acetylation
Acetylglucosamine/*metabolism
Animals
Autophagy
Crohn Disease/*metabolism
Female
HCT116 Cells
HT29 Cells
Humans
Intestinal Mucosa/metabolism
Male
Mice
Mice, Inbred C57BL
NF-kappa B/*metabolism
*Protein Processing, Post-Translational
Crohn's disease (CD)
Nf-κb
O-Linked β-N-acetylglucosamine (O-GlcNAc)
UDP-N-acetylglucosamine (UDP-GlcNAc)
adherent-invasive Escherichia coli (AIEC) LF82
inflammatory bowel disease
interest exists.
Abstract: BACKGROUND: Treatment of Crohn's disease (CD) remains to be a challenge due to limited insights for its pathogenesis. We aimed to determine the role of O-Linked β-N-acetylglucosamine (O-GlcNAc) in the development of CD and evaluate therapeutic effects of O-GlcNAc inhibitors on CD. METHODS: O-GlcNAc in intestinal epithelial tissues of CD, adherent-invasive Escherichia coli (AIEC) LF82-infected cells and mice was determined by immunoblot and immunohistochemistry. AIEC LF82 and dextran sulfate sodium were administrated into C57BL/6 mice for estabolishing inflammatory bowel disease model and for therapeutic study. FINDINGS: O-GlcNAc was increased in intestinal epithelial tissues of CD patients and AIEC LF82-infected mice. Infection of AIEC LF82 up-regulated the level of UDP-GlcNAc and increased O-GlcNAc in human colon epithelial HCT116 and HT-29 cells. We identified that IKKβ and NF-κB were O-Glycosylated in AIEC LF82-treated cells. Mutations of IKKβ (S733A) and p65 (T352A) abrogated the O-GlcNAc in IKKβ and NF-κB and inhibited AIEC LF82-induced activation of NF-κB. Application of 6-diazO-5-oxO-L-norleucine, an agent that blocks the production of UDP-GlcNAc and inhibits O-GlcNAc, inactivated NF-κB in AIEC LF82-infected cells, enhanced the formation of autophagy, promoted the removal of cell-associated AIEC LF82, alleviated intestinal epithelial inflammation, and improved the survival of the colitis mice. INTERPRETATION: Intestinal inflammation in CD is associated with increased O-GlcNAc modification, which is required for NF-κB activation and suppression of autophagy. Targeting O-GlcNAc could be an effective therapy for inflammatory bowel disease. FUNDING: National Natural Science Foundation of China (Nos. 81573087 and 81772924) and International Cooperation Foundation of Jilin Province (20190701006GH).
Notes: 2352-3964
Sun, Qian-Hui
Wang, Yi-Shu
Liu, Guolong
Zhou, Hong-Lan
Jian, Yong-Ping
Liu, Ming-Di
Zhang, Dan
Ding, Qiang
Zhao, Rui-Xun
Chen, Jian-Feng
Li, Yi-Ning
Liang, Jiyong
Li, Yu-Lin
Quan, Cheng-Shi
Xu, Zhi-Xiang
Journal Article
EBioMedicine. 2020 Mar;53:102693. doi: 10.1016/j.ebiom.2020.102693. Epub 2020 Feb 27.
Author Address: Key Laboratory of Pathobiology, Ministry of Education, Norman Bethune College of Medicine, Jilin University, Changchun, China; Department of Medicine, University of Alabama at Birmingham, Birmingham, AL 35294, United States.
Key Laboratory of Pathobiology, Ministry of Education, Norman Bethune College of Medicine, Jilin University, Changchun, China.
Department of Medical Oncology, Guangzhou First People's Hospital, School of Medicine, South China University of Technology, Guangzhou, Guangdong, China.
Department of Urology, The First Hospital of Jilin University, Changchun, China.
Department of Medicine, University of Alabama at Birmingham, Birmingham, AL 35294, United States.
Department of Neurosurgery, UT MD Anderson Cancer Center, Houston, TX 77030, United States.
Key Laboratory of Pathobiology, Ministry of Education, Norman Bethune College of Medicine, Jilin University, Changchun, China. Electronic address: ylli@jlu.edu.cn.
Key Laboratory of Pathobiology, Ministry of Education, Norman Bethune College of Medicine, Jilin University, Changchun, China. Electronic address: quancs@jlu.edu.cn.
Key Laboratory of Pathobiology, Ministry of Education, Norman Bethune College of Medicine, Jilin University, Changchun, China; College of Biological Sciences, Henan University, Kaifeng, China; Department of Medicine, University of Alabama at Birmingham, Birmingham, AL 35294, United States. Electronic address: zhixiangxu08@gmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1930
Author: Sun, S., Hu, Y., Jia, L., Eshghi, S. T., Liu, Y., Shah, P. and Zhang, H.
Year: 2018
Title: Site-Specific Profiling of Serum Glycoproteins Using N-Linked Glycan and Glycosite Analysis Revealing Atypical N-Glycosylation Sites on Albumin and α-1B-Glycoprotein
Journal: Anal Chem
Volume: 90
Issue: 10
Pages: 6292-6299
Epub Date: 2018/04/20
Date: May 15
Short Title: Site-Specific Profiling of Serum Glycoproteins Using N-Linked Glycan and Glycosite Analysis Revealing Atypical N-Glycosylation Sites on Albumin and α-1B-Glycoprotein
Alternate Journal: Analytical chemistry
ISSN: 0003-2700 (Print)
0003-2700
DOI: 10.1021/acs.analchem.8b01051
PMCID: PMC6467210
NIHMSID: NIHMS1014643
Accession Number: 29671580
Keywords: Albumins/*chemistry
Glycoproteins/*blood
Glycosylation
Humans
Peptides/*analysis
Polysaccharides/*analysis
Abstract: Most serum proteins are N-linked glycosylated, and therefore the glycoproteomic profiling of serum is essential for characterization of serum proteins. In this study, we profiled serum N-glycoproteome by our recently developed N-glycoproteomic method using solid-phase extraction of N-linked glycans and glycosite-containing peptides (NGAG) coupled with LC-MS/MS and site-specific glycosylation analysis using GPQuest software. Our data indicated that half of identified N-glycosites were modified by at least two glycans, with a majority of them being sialylated. Specifically, 3/4 of glycosites were modified by biantennary N-glycans and 1/3 of glycosites were modified by triantennary sialylated N-glycans. In addition, two novel atypical glycosites (with N-X-V motif) were identified and validated from albumin and α-1B-glycoprotein. The widespread presence of these two glycosites among individuals was further confirmed by individual serum analyses.
Notes: 1520-6882
Sun, Shisheng
Orcid: 0000-0002-7242-7164
Hu, Yingwei
Orcid: 0000-0002-4629-0985
Jia, Li
Eshghi, Shadi Toghi
Liu, Yang
Shah, Punit
Zhang, Hui
Orcid: 0000-0001-8726-7098
P01 HL107153/HL/NHLBI NIH HHS/United States
R21 AI122382/AI/NIAID NIH HHS/United States
U01 CA152813/CA/NCI NIH HHS/United States
U24 CA210985/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Anal Chem. 2018 May 15;90(10):6292-6299. doi: 10.1021/acs.analchem.8b01051. Epub 2018 May 1.
Author Address: College of Life Science , Northwest University , Xi'an , Shaanxi Province 710069 , China.
Department of Pathology , Johns Hopkins University , Baltimore , Maryland 21287 , United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1166
Author: Sun, W., Parry, S., Panico, M., Morris, H. R., Kjellberg, M., Engström, A., Dell, A. and Schedin-Weiss, S.
Year: 2008
Title: N-glycans and the N terminus of protein C inhibitor affect the cofactor-enhanced rates of thrombin inhibition
Journal: J Biol Chem
Volume: 283
Issue: 27
Pages: 18601-11
Epub Date: 2008/05/10
Date: Jul 4
Short Title: N-glycans and the N terminus of protein C inhibitor affect the cofactor-enhanced rates of thrombin inhibition
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M800608200
Accession Number: 18467335
Keywords: Atherosclerosis/metabolism
Glycosylation
Heparin/chemistry/metabolism
Humans
Polysaccharides/*chemistry/metabolism
Protein C Inhibitor/*chemistry/metabolism
Protein Structure, Tertiary/physiology
Substrate Specificity/physiology
Thrombin/*chemistry/metabolism
Thrombomodulin/chemistry/metabolism
Thrombophilia/metabolism
Abstract: Protein C inhibitor (PCI) is a serine protease inhibitor, displaying broad protease specificity, found in blood and other tissues. In blood, it is capable of inhibiting both procoagulant and anticoagulant proteases. Mechanisms that provide specificity to PCI remain largely unrevealed. In this study we have for the first time provided a full explanation for the marked size heterogeneity of blood-derived PCI and identified functional differences between naturally occurring PCI variants. The heterogeneity was caused by differences in N-glycan structures, N-glycosylation occupancy, and the presence of a Delta6-N-cleaved form. Bi-, tri-, and tetra-antennary complex N-glycans were identified. Fucose residues were identified both on the core GlcNAc and as parts of sialyl-Le(a/x) epitopes. Moreover, a glycan with a composition that implied a di-sialyl antenna was observed. PCI was N-glycosylated at all three potential N-glycosylation sites, Asn-230, Asn-243, and Asn-319, but a small fraction of PCI lacked the N-glycan at Asn-243. The overall removal of N-glycans affected the maximal heparin- and thrombomodulin-enhanced rates of thrombin inhibition differently in different solution conditions. In contrast, the Delta6-N-region increased both the heparin- and the thrombomodulin-enhanced rates of thrombin inhibition at all conditions examined. These results thus demonstrate that the N-linked glycans and the N-terminal region of blood-derived PCI in different ways affect the cofactor-enhanced rates of thrombin inhibition and provide information on the mechanisms by which this may be achieved. The findings are medically important, in view of the documented association of PCI with atherosclerotic plaques and the promising effect of PCI on reducing hypercoagulability states.
Notes: Sun, Wei
Parry, Simon
Panico, Maria
Morris, Howard R
Kjellberg, Margareta
Engström, Ake
Dell, Anne
Schedin-Weiss, Sophia
Biotechnology and Biological Sciences Research Council/United Kingdom
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2008 Jul 4;283(27):18601-11. doi: 10.1074/jbc.M800608200. Epub 2008 May 8.
Author Address: Department of Medical Biochemistry and Microbiology, Uppsala University, Box 582, Uppsala SE-751 23, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1357
Author: Sun, W., Parry, S., Ubhayasekera, W., Engström, A., Dell, A. and Schedin-Weiss, S.
Year: 2010
Title: Further insight into the roles of the glycans attached to human blood protein C inhibitor
Journal: Biochem Biophys Res Commun
Volume: 403
Issue: 2
Pages: 198-202
Epub Date: 2010/11/09
Date: Dec 10
Short Title: Further insight into the roles of the glycans attached to human blood protein C inhibitor
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2010.11.005
Accession Number: 21056543
Keywords: Amino Acid Sequence
Blood Donors
Glycosylation
Humans
Molecular Sequence Data
Polysaccharides/*chemistry
Protein C Inhibitor/*chemistry
Protein Conformation
Spectrometry, Mass, Electrospray Ionization
Abstract: Protein C inhibitor (PCI) is a 57-kDa glycoprotein that exists in many tissues and secretions in human. As a member of the serpin superfamily of proteins it displays unusually broad protease specificity. PCI is implicated in the regulation of a wide range of processes, including blood coagulation, fertilization, prevention of tumors and pathogen defence. It has been reported that PCI isolated from human blood plasma is highly heterogeneous, and that this heterogeneity is caused by differences in N-glycan structures, N-glycosylation occupancy, and the presence of two forms that differ by the presence or absence of 6 amino acids at the amino-terminus. In this study we have verified that such heterogeneity exists in PCI purified from single individuals, and that individuals of two different ethnicities possess a similar PCI pattern, verifying that the micro-heterogeneity is conserved among humans. Furthermore, we have provided experimental evidence that PCI in both individuals is O-glycosylated on Thr20 with a core type 1 O-glycan, which is mostly NeuAcGalGalNAc. Modeling suggested that the O-glycan attachment site is located in proximity to several ligand-binding sites of the inhibitor.
Notes: 1090-2104
Sun, Wei
Parry, Simon
Ubhayasekera, Wimal
Engström, Ake
Dell, Anne
Schedin-Weiss, Sophia
B19088/Biotechnology and Biological Sciences Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2010 Dec 10;403(2):198-202. doi: 10.1016/j.bbrc.2010.11.005. Epub 2010 Nov 5.
Author Address: Department of Medical Biochemistry and Microbiology, Uppsala University, Box 582, Uppsala SE-751 23, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 554
Author: Sutton, C. W., O'Neill, J. A. and Cottrell, J. S.
Year: 1994
Title: Site-specific characterization of glycoprotein carbohydrates by exoglycosidase digestion and laser desorption mass spectrometry
Journal: Anal Biochem
Volume: 218
Issue: 1
Pages: 34-46
Epub Date: 1994/04/01
Date: Apr
Short Title: Site-specific characterization of glycoprotein carbohydrates by exoglycosidase digestion and laser desorption mass spectrometry
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1006/abio.1994.1138
Accession Number: 8053566
Keywords: Amidohydrolases
Amino Acid Sequence
Carbohydrate Sequence
Glycopeptides/*analysis
Glycoproteins/*analysis
*Glycoside Hydrolases
Lasers
Mass Spectrometry/*methods
Metalloendopeptidases/*antagonists & inhibitors
Molecular Sequence Data
Molecular Weight
Oligosaccharides/analysis
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Sensitivity and Specificity
Substrate Specificity
Tissue Inhibitor of Metalloproteinases
Abstract: A rapid and sensitive method for sequencing oligosaccharides has been developed, using matrix-assisted laser desorption mass spectrometry to monitor the digestion of glycopeptides by specific exoglycosidases. Recombinant human tissue inhibitor of metalloproteinases (TIMP), which has two glycosylation sites, has been characterized to illustrate this new approach for obtaining site-specific information. Glycopeptides which span residues Asn30 and Asn78 were generated by tryptic digestion of 1 nmol of TIMP and separated by reverse-phase high-performance liquid chromatography. The oligosaccharide composition of the glycoforms was inferred from the observed mass shifts following digestion by peptide-N-glycosidase F. Composition and sequence were then elucidated by digestion with specific exoglycosidases, using a total of 200 pmol of each glycopeptide. Glycopeptides from well-characterized proteins, fetuin, alpha 1-acid glycoprotein, and tissue plasminogen activator were also analyzed to confirm exoglycosidase specificity for glycopeptides and establish the quantitative significance of the relative intensities of peaks in the mass spectra. Both TIMP glycosylation sites exhibited extensive heterogeneity comprising mainly fucosylated complex oligosaccharides, but in different proportions. The Asn78 site also contained 4.4% nonfucosylated mannose (Man4) oligosaccharide. The merits and limitations of this approach as a universal method for oligosaccharide analysis are discussed.
Notes: Sutton, C W
O'Neill, J A
Cottrell, J S
Journal Article
United States
Anal Biochem. 1994 Apr;218(1):34-46. doi: 10.1006/abio.1994.1138.
Author Address: Finnigan MAT Ltd., Paradise, Hemel Hempstead, Hertfordshire, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1637
Author: Suzuki, H., Ohto, U., Higaki, K., Mena-Barragán, T., Aguilar-Moncayo, M., Ortiz Mellet, C., Nanba, E., Garcia Fernandez, J. M., Suzuki, Y. and Shimizu, T.
Year: 2014
Title: Structural basis of pharmacological chaperoning for human β-galactosidase
Journal: J Biol Chem
Volume: 289
Issue: 21
Pages: 14560-8
Epub Date: 2014/04/17
Date: May 23
Short Title: Structural basis of pharmacological chaperoning for human β-galactosidase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.529529
PMCID: PMC4031513
Accession Number: 24737316
Keywords: 1-Deoxynojirimycin/analogs & derivatives/chemistry/pharmacology
Catalytic Domain
Crystallography, X-Ray
Cyclohexenes/chemistry/pharmacology
Enzyme Inhibitors/chemistry/*pharmacology
Enzyme Stability/drug effects
Gangliosidosis, GM1/*enzymology/genetics
Hexosamines/chemistry/pharmacology
Humans
Hydrogen Bonding
Hydrogen-Ion Concentration
Imino Sugars/chemistry/pharmacology
Inositol/analogs & derivatives/chemistry/pharmacology
Kinetics
Models, Molecular
Molecular Structure
Mucopolysaccharidosis IV/*enzymology/genetics
Mutation
Protein Structure, Tertiary
Static Electricity
Structure-Activity Relationship
beta-Galactosidase/*antagonists & inhibitors/chemistry/genetics
Crystal Structure
Galactose
Glycosidases
Lysosomal Storage Disease
X-ray Crystallography
Abstract: GM1 gangliosidosis and Morquio B disease are autosomal recessive diseases caused by the defect in the lysosomal β-galactosidase (β-Gal), frequently related to misfolding and subsequent endoplasmic reticulum-associated degradation. Pharmacological chaperone (PC) therapy is a newly developed molecular therapeutic approach by using small molecule ligands of the mutant enzyme that are able to promote the correct folding and prevent endoplasmic reticulum-associated degradation and promote trafficking to the lysosome. In this report, we describe the enzymological properties of purified recombinant human β-Gal(WT) and two representative mutations in GM1 gangliosidosis Japanese patients, β-Gal(R201C) and β-Gal(I51T). We have also evaluated the PC effect of two competitive inhibitors of β-Gal. Moreover, we provide a detailed atomic view of the recognition mechanism of these compounds in comparison with two structurally related analogues. All compounds bind to the active site of β-Gal with the sugar-mimicking moiety making hydrogen bonds to active site residues. Moreover, the binding affinity, the enzyme selectivity, and the PC potential are strongly affected by the mono- or bicyclic structure of the core as well as the orientation, nature, and length of the exocyclic substituent. These results provide understanding on the mechanism of action of β-Gal selective chaperoning by newly developed PC compounds.
Notes: 1083-351x
Suzuki, Hironori
Ohto, Umeharu
Higaki, Katsumi
Mena-Barragán, Teresa
Aguilar-Moncayo, Matilde
Ortiz Mellet, Carmen
Nanba, Eiji
Garcia Fernandez, Jose M
Suzuki, Yoshiyuki
Shimizu, Toshiyuki
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2014 May 23;289(21):14560-8. doi: 10.1074/jbc.M113.529529. Epub 2014 Apr 15.
Author Address: From the Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
the Division of Functional Genomics, Research Center for Bioscience and Technology, Tottori University, 86 Nishi-cho, Yonago, Tottori 683-8503, Japan.
the Department of Organic Chemistry, Faculty of Chemistry, University of Seville, Profesor García González 1, E-41012 Seville, Spain.
the Institute for Chemical Research (IIQ), CSIC, University of Sevilla, Americo Vespucio 49, Isla de la Cartuja, E-41092 Sevilla, Spain.
the International University of Health and Welfare Graduate School, Kita Kanemaru, Otawara, Tochigi 324-8501, Japan, and.
From the Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan, CREST, Japan Science and Technology Agency, 4-1-8 Honcho Kawaguchi, Saitama 332-0012, Japan shimizu@mol.f.u-tokyo.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 928
Author: Suzuki, M. and Mizuno, A.
Year: 2004
Title: A novel human Cl(-) channel family related to Drosophila flightless locus
Journal: J Biol Chem
Volume: 279
Issue: 21
Pages: 22461-8
Epub Date: 2004/03/11
Date: May 21
Short Title: A novel human Cl(-) channel family related to Drosophila flightless locus
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M313813200
Accession Number: 15010458
Keywords: Amino Acid Sequence
Amino Acids/chemistry
Animals
Anions
CHO Cells
Calcium/pharmacology
Cell Line
Chelating Agents/pharmacology
Chloride Channels/chemistry/*physiology
Cricetinae
DNA, Complementary/metabolism
Drosophila/*genetics/metabolism
Drosophila Proteins/genetics/*physiology
Electrophysiology
Gelsolin/genetics/*physiology
Humans
Immunohistochemistry
Ionomycin/pharmacology
Kinetics
Membrane Proteins/chemistry
Mice
Microscopy, Fluorescence
Molecular Sequence Data
Mutation
Neoplasm Proteins/chemistry
RNA, Messenger/metabolism
Sequence Homology, Amino Acid
Abstract: Large conductance chloride (maxi-Cl(-)) currents have been recorded in some cells, but there is still little information on the molecular nature of the channel underlying this conductance. We report here that tweety, a gene located in Drosophila flightless, has a structure similar to those of known channels and that human homologues of tweety (hTTYH1-3) are novel maxi-Cl(-) channels. hTTYH3 mRNA was found to be distributed in excitable tissues. The whole cell current of hTTYH3 was large enough to be discriminated from the control but emerged only after treatment with ionomycin. Analysis of pore mutants suggested that positively charged amino acids contributed to anion selectivity. Like a maxi-Cl(-) channel in situ, the hTTYH3 single channel showed 26-picosiemen linear current voltage, complex kinetics, 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid sensitivity, subconductance, and the permeability order of I(-) > Br(-) > Cl(-). Similarly, hTTYH2 encoded an ionomycin-induced maxi-Cl(-) channel, but TTYH1 encoded a Ca(2+)-independent and swelling-activated maxi-Cl(-) channel. Therefore, the hTTYH family encoded maxi-Cl(-) channels of mammals. Further studies on the hTTYH family should lead to the elucidation of physiological and pathophysiological roles of novel Cl(-) channel molecules.
Notes: Suzuki, Makoto
Mizuno, Atsuko
Journal Article
United States
J Biol Chem. 2004 May 21;279(21):22461-8. doi: 10.1074/jbc.M313813200. Epub 2004 Mar 8.
Author Address: Department of Pharmacology, Jichi Medical School 3311-1, Yakushiji, Minamikawachi, Tochigi 329-0498, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1460
Author: Svensson, G., Awad, W., Håkansson, M., Mani, K. and Logan, D. T.
Year: 2012
Title: Crystal structure of N-glycosylated human glypican-1 core protein: structure of two loops evolutionarily conserved in vertebrate glypican-1
Journal: J Biol Chem
Volume: 287
Issue: 17
Pages: 14040-51
Epub Date: 2012/02/22
Date: Apr 20
Short Title: Crystal structure of N-glycosylated human glypican-1 core protein: structure of two loops evolutionarily conserved in vertebrate glypican-1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.322487
PMCID: PMC3340175
Accession Number: 22351761
Keywords: Animals
Cell Membrane/metabolism
Circular Dichroism
Cloning, Molecular
Conserved Sequence
Crystallography, X-Ray/*methods
Disulfides/chemistry
Glycoproteins/chemistry
Glycosylation
Glypicans/*chemistry
HEK293 Cells
Heparitin Sulfate/chemistry
Humans
Models, Molecular
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Proteoglycans/chemistry
Abstract: Glypicans are a family of cell-surface proteoglycans that regulate Wnt, hedgehog, bone morphogenetic protein, and fibroblast growth factor signaling. Loss-of-function mutations in glypican core proteins and in glycosaminoglycan-synthesizing enzymes have revealed that glypican core proteins and their glycosaminoglycan chains are important in shaping animal development. Glypican core proteins consist of a stable α-helical domain containing 14 conserved Cys residues followed by a glycosaminoglycan attachment domain that becomes exclusively substituted with heparan sulfate (HS) and presumably adopts a random coil conformation. Removal of the α-helical domain results in almost exclusive addition of the glycosaminoglycan chondroitin sulfate, suggesting that factors in the α-helical domain promote assembly of HS. Glypican-1 is involved in brain development and is one of six members of the vertebrate family of glypicans. We expressed and crystallized N-glycosylated human glypican-1 lacking HS and N-glycosylated glypican-1 lacking the HS attachment domain. The crystal structure of glypican-1 was solved using crystals of selenomethionine-labeled glypican-1 core protein lacking the HS domain. No additional electron density was observed for crystals of glypican-1 containing the HS attachment domain, and CD spectra of the two protein species were highly similar. The crystal structure of N-glycosylated human glypican-1 core protein at 2.5 Å, the first crystal structure of a vertebrate glypican, reveals the complete disulfide bond arrangement of the conserved Cys residues, and it also extends the structural knowledge of glypicans for one α-helix and two long loops. Importantly, the loops are evolutionarily conserved in vertebrate glypican-1, and one of them is involved in glycosaminoglycan class determination.
Notes: 1083-351x
Svensson, Gabriel
Awad, Wael
Håkansson, Maria
Mani, Katrin
Logan, Derek T
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Apr 20;287(17):14040-51. doi: 10.1074/jbc.M111.322487. Epub 2012 Feb 20.
Author Address: Department of Experimental Medical Science, Division of Neuroscience, Glycobiology Group, Lund University, Biomedical Center A13, SE-221 84 Lund, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1428
Author: Svensson, G., Hyrenius Wittsten, A., Linse, S. and Mani, K.
Year: 2011
Title: The structural role of N-linked glycans on human glypican-1
Journal: Biochemistry
Volume: 50
Issue: 43
Pages: 9377-87
Epub Date: 2011/09/22
Date: Nov 1
Short Title: The structural role of N-linked glycans on human glypican-1
Alternate Journal: Biochemistry
ISSN: 0006-2960
DOI: 10.1021/bi200218s
Accession Number: 21932778
Keywords: Cell Line
Glycosylation
Glypicans/*chemistry/*genetics/metabolism
Humans
Mutation
Polysaccharides/*analysis
Protein Biosynthesis
Protein Folding
Protein Stability
Recombinant Proteins/chemistry/genetics
Abstract: Glypicans are cell-surface heparan sulfate proteoglycans that regulate developmental signaling pathways by binding growth factors to their heparan sulfate chains. The primary structures of glypican core proteins contain potential N-glycosylation sites, but the importance of N-glycosylation in glypicans has never been investigated in detail. Here, we studied the role of the possible N-glycosylation sites at Asn-79 and Asn-116 in recombinant anchorless glypican-1 expressed in eukaryotic cells. Mutagenesis and enzymatic cleavage indicated that the potential N-glycosylation sites are invariably occupied. Experiments using the drug tunicamycin to inhibit the N-linked glycosylation of glypican-1 showed that secretion of anchorless glypican-1 was reduced and that the protein did not accumulate inside the cells. Heparan sulfate substitution of N-glycosylation mutant N116Q was similar to wild-type glypican-1 while the N79Q mutant and also the double mutant N79Q,N116Q were mostly secreted as high-molecular-weight heparan sulfate proteoglycan. N-Glycosylation mutants and N-deglycosylated glypican-1 had far-UV circular dichroism and fluorescence emission spectra that were highly similar to those of N-glycosylated glypican-1. A single unfolding transition at high concentrations of urea was found for both N-deglycosylated glypican-1 and glypican-1 in which the N-glycosylation sites had been removed by mutagenesis when chemical denaturation was monitored by circular dichroism and fluorescence emission spectroscopy. In summary, we have found that the potential N-glycosylation sites in glypican-1 are invariably occupied and that the N-linked glycans on glypican-1 affect protein expression and heparan sulfate substitution but that correct folding can be obtained in the absence of N-linked glycans.
Notes: 1520-4995
Svensson, Gabriel
Hyrenius Wittsten, Axel
Linse, Sara
Mani, Katrin
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2011 Nov 1;50(43):9377-87. doi: 10.1021/bi200218s. Epub 2011 Oct 5.
Author Address: Department of Experimental Medical Science, Division of Neuroscience, Glycobiology Group, Lund University, Biomedical Center A13, SE-221 84, Lund, Sweden. gabriel.svensson@med.lu.se
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 430
Author: Swiedler, S. J., Freed, J. H., Tarentino, A. L., Plummer, T. H., Jr. and Hart, G. W.
Year: 1985
Title: Oligosaccharide microheterogeneity of the murine major histocompatibility antigens. Reproducible site-specific patterns of sialylation and branching in asparagine-linked oligosaccharides
Journal: J Biol Chem
Volume: 260
Issue: 7
Pages: 4046-54
Epub Date: 1985/04/10
Date: Apr 10
Short Title: Oligosaccharide microheterogeneity of the murine major histocompatibility antigens. Reproducible site-specific patterns of sialylation and branching in asparagine-linked oligosaccharides
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3980466
Keywords: Animals
Binding Sites
Chromatography, Gel
Chromatography, High Pressure Liquid
Electrophoresis, Polyacrylamide Gel
H-2 Antigens/*analysis
Lymphoma/immunology
Mice
Oligosaccharides/*analysis
Abstract: The influence of peptide structure of endogenous cell-surface glycoproteins on the branching and sialylation of their asparagine-linked oligosaccharides was evaluated in a murine B cell lymphoma, AKTB-1b. This cell line simultaneously synthesizes two classes of major histocompatibility antigens that, within each class, share a high degree of amino acid sequence homology and possess potential N-linked glycosylation sites at invariant positions. [3H]Mannose-labeled oligosaccharides were released from each of 11 purified glycosylation sites by the almond peptide:N-glycosidase and analyzed by a variety of chromatographic procedures and glycosidase treatments. The data indicate: 1) a unique distribution of oligosaccharide structures is present at each glycosylation site; 2) each site-specific oligosaccharide pattern is highly reproducible, independent of the number of in vivo tumor passages. The heavy chain of the class I antigens, H-2Kk and H-2Dk contain two and three sites, respectively, in which biantennary structures predominate. However, each site varies with respect to the extent of sialylation and the proportions of more highly branched structures present. The class II antigens, I-Ak and I-Ek, each contain an alpha-chain site toward the N terminus and a single beta-chain site where the overall extent of sialylation is similar, yet the distributions of antennary structures are dramatically different for each. The alpha-chains of each class II antigen also contain a more C-terminal underglycosylated site where sialylation and branching are reduced to differing degrees depending upon the site. The influence of peptide structure on oligosaccharide microheterogeneity is manifest at two levels. First, the overall distributions of oligosaccharides at corresponding sites on structurally related glycoproteins are similar. Second, the specific "fingerprint" of sialylation and branching patterns at a particular site are reproducibly unique. These data suggest that subtle changes in peptide structure are reflected in the extent of sialylation and branching of oligosaccharides found at corresponding glycosylation sites of structurally related glycoproteins.
Notes: Swiedler, S J
Freed, J H
Tarentino, A L
Plummer, T H Jr
Hart, G W
30471/PHS HHS/United States
CA-20820/CA/NCI NIH HHS/United States
HD-13563/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1985 Apr 10;260(7):4046-54.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1705
Author: Tagliabracci, V. S., Wiley, S. E., Guo, X., Kinch, L. N., Durrant, E., Wen, J., Xiao, J., Cui, J., Nguyen, K. B., Engel, J. L., Coon, J. J., Grishin, N., Pinna, L. A., Pagliarini, D. J. and Dixon, J. E.
Year: 2015
Title: A Single Kinase Generates the Majority of the Secreted Phosphoproteome
Journal: Cell
Volume: 161
Issue: 7
Pages: 1619-32
Epub Date: 2015/06/20
Date: Jun 18
Short Title: A Single Kinase Generates the Majority of the Secreted Phosphoproteome
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2015.05.028
PMCID: PMC4963185
NIHMSID: NIHMS699715
Accession Number: 26091039
Keywords: Amino Acid Sequence
Blood Proteins/metabolism
Casein Kinase I/*chemistry/genetics/*metabolism
Cell Adhesion
Cell Movement
Cerebrospinal Fluid Proteins/metabolism
Extracellular Matrix Proteins/*chemistry/genetics/*metabolism
Gene Knockout Techniques
Gene Ontology
Humans
Molecular Sequence Data
Phosphoproteins/analysis
Secretory Pathway
Substrate Specificity
Abstract: The existence of extracellular phosphoproteins has been acknowledged for over a century. However, research in this area has been undeveloped largely because the kinases that phosphorylate secreted proteins have escaped identification. Fam20C is a kinase that phosphorylates S-x-E/pS motifs on proteins in milk and in the extracellular matrix of bones and teeth. Here, we show that Fam20C generates the majority of the extracellular phosphoproteome. Using CRISPR/Cas9 genome editing, mass spectrometry, and biochemistry, we identify more than 100 secreted phosphoproteins as genuine Fam20C substrates. Further, we show that Fam20C exhibits broader substrate specificity than previously appreciated. Functional annotations of Fam20C substrates suggest roles for the kinase beyond biomineralization, including lipid homeostasis, wound healing, and cell migration and adhesion. Our results establish Fam20C as the major secretory pathway protein kinase and serve as a foundation for new areas of investigation into the role of secreted protein phosphorylation in human biology and disease.
Notes: 1097-4172
Tagliabracci, Vincent S
Wiley, Sandra E
Guo, Xiao
Kinch, Lisa N
Durrant, Eric
Wen, Jianzhong
Xiao, Junyu
Cui, Jixin
Nguyen, Kim B
Engel, James L
Coon, Joshua J
Grishin, Nick
Pinna, Lorenzo A
Pagliarini, David J
Dixon, Jack E
K99 DK099254/DK/NIDDK NIH HHS/United States
K99DK099254/DK/NIDDK NIH HHS/United States
R01 DK018024/DK/NIDDK NIH HHS/United States
R01 DK098672/DK/NIDDK NIH HHS/United States
DK018024-37/DK/NIDDK NIH HHS/United States
R01DK098672/DK/NIDDK NIH HHS/United States
DK018849-36/DK/NIDDK NIH HHS/United States
T32 GM007752/GM/NIGMS NIH HHS/United States
R01 GM094575/GM/NIGMS NIH HHS/United States
R01 DK018849/DK/NIDDK NIH HHS/United States
R37 DK018024/DK/NIDDK NIH HHS/United States
GM094575/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell. 2015 Jun 18;161(7):1619-32. doi: 10.1016/j.cell.2015.05.028.
Author Address: Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.
Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA; Department of Chemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
Department of Biophysics, University of Texas, Southwestern Medical Center, Dallas, TX 75390-9050, USA.
Department of Chemistry, University of Wisconsin-Madison, Madison, WI 53706, USA; Department of Biomolecular Chemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
Department of Biophysics, University of Texas, Southwestern Medical Center, Dallas, TX 75390-9050, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
Department of Biomedical Sciences, University of Padova, 35121 Padova, Italy; Venetian Institute of Molecular Medicine, 35129 Padova, Italy.
Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA.
Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA; Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA; Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: jedixon@mail.ucsd.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 949
Author: Tai, H. C., Khidekel, N., Ficarro, S. B., Peters, E. C. and Hsieh-Wilson, L. C.
Year: 2004
Title: Parallel identification of O-GlcNAc-modified proteins from cell lysates
Journal: J Am Chem Soc
Volume: 126
Issue: 34
Pages: 10500-1
Epub Date: 2004/08/26
Date: Sep 1
Short Title: Parallel identification of O-GlcNAc-modified proteins from cell lysates
Alternate Journal: Journal of the American Chemical Society
ISSN: 0002-7863 (Print)
0002-7863
DOI: 10.1021/ja047872b
Accession Number: 15327282
Keywords: Acetylglucosamine/chemistry/*metabolism
Activating Transcription Factor 1
Amino Acid Sequence
CREB-Binding Protein
DNA-Binding Proteins/chemistry/metabolism
Glycosylation
HeLa Cells
Humans
Mass Spectrometry
Molecular Sequence Data
N-Acetylglucosaminyltransferases/chemistry/metabolism
Nuclear Proteins/chemistry/metabolism
Proto-Oncogene Proteins c-fos/chemistry/metabolism
Proto-Oncogene Proteins c-jun/chemistry/metabolism
Trans-Activators/chemistry/metabolism
Transcription Factors/chemistry/*metabolism
Abstract: We report a new strategy for the parallel identification of O-GlcNAc-glycosylated proteins from cell lysates. The approach permits specific proteins of interest to be rapidly interrogated for the modification in any tissue or cell type and can be extended to peptides to facilitate the mapping of glycosylation sites. As an illustration of the approach, we identified four new O-GlcNAc-glycosylated proteins of low cellular abundance (c-Fos, c-Jun, ATF-1, and CBP) and two short regions of glycosylation in the enzyme O-GlcNAc transferase (OGT). The ability to target specific proteins across various tissue or cell types complements emerging proteomic technologies and should advance our understanding of this important posttranslational modification.
Notes: Tai, Hwan-Ching
Khidekel, Nelly
Ficarro, Scott B
Peters, Eric C
Hsieh-Wilson, Linda C
GM07616-21/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Am Chem Soc. 2004 Sep 1;126(34):10500-1. doi: 10.1021/ja047872b.
Author Address: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1137
Author: Tajiri, M., Ohyama, C. and Wada, Y.
Year: 2008
Title: Oligosaccharide profiles of the prostate specific antigen in free and complexed forms from the prostate cancer patient serum and in seminal plasma: a glycopeptide approach
Journal: Glycobiology
Volume: 18
Issue: 1
Pages: 2-8
Epub Date: 2007/10/25
Date: Jan
Short Title: Oligosaccharide profiles of the prostate specific antigen in free and complexed forms from the prostate cancer patient serum and in seminal plasma: a glycopeptide approach
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cwm117
Accession Number: 17956937
Keywords: Amino Acid Sequence
Biomarkers, Tumor/blood/*chemistry
Cell Line, Tumor
Fucose/analysis
Glycopeptides/*chemistry
Humans
Male
Molecular Sequence Data
Oligosaccharides/blood/*chemistry
Prostate-Specific Antigen/blood/*chemistry/isolation & purification
Prostatic Neoplasms/blood/*chemistry/diagnosis
Semen/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: The oligosaccharide structures of prostate specific antigen (PSA) are expected to be useful in discriminating prostate cancer from benign conditions both accompanied by increased serum PSA levels. A large proportion of PSA forms a covalent complex with a glycoprotein, alpha(1)-antichymotrypsin, in human blood. In the present study, the glycan profiles of free and complexed forms of PSA from cancer patient serum and of seminal plasma PSA were compared by analyzing the glycopeptides obtained by lysylendopeptidase digestion of the electrophoretically separated PSA with mass spectrometry. The profiles of the PSA N-glycans from the free and complexed molecules were quite similar to each other and consisted of fucosylated biantennary oligosaccharides as the major class. They were mostly sialylated, and a considerable sialic acid fraction was alpha2,3-linked as determined by Streptococcus pneumoniae neuraminidase digestion of the glycopeptides. In the seminal plasma PSA, high-mannose and hybrid types of oligosaccharides were predominant, and the sialic acids attached to the latter as well as to biantennary oligosaccahrides were exclusively alpha2,6-linked because they were removed by Arthrobacter ureafaciens neuraminidase but resistant to S. pneumoniae neuraminidase. Complex-type oligosaccharides from other sources were found in the seminal plasma sample, indicating that analysis of released glycans carries a risk of being misleading. The results suggest that identification of alpha2,3-linked sialic acids on PSA potentially discriminates malignant from benign conditions, if the analysis is applied to oligosaccharides specifically attached to the N-glycosylation site of PSA in either a free or a complexed form in the serum.
Notes: 1460-2423
Tajiri, Michiko
Ohyama, Chikara
Wada, Yoshinao
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2008 Jan;18(1):2-8. doi: 10.1093/glycob/cwm117. Epub 2007 Oct 23.
Author Address: Department of Molecular Medicine, Osaka Medical Center and Research Institute for Maternal and Child Health, 840 Murodo-cho Izumi, Osaka 594-1101, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1001
Author: Tajiri, M., Yoshida, S. and Wada, Y.
Year: 2005
Title: Differential analysis of site-specific glycans on plasma and cellular fibronectins: application of a hydrophilic affinity method for glycopeptide enrichment
Journal: Glycobiology
Volume: 15
Issue: 12
Pages: 1332-40
Epub Date: 2005/07/23
Date: Dec
Short Title: Differential analysis of site-specific glycans on plasma and cellular fibronectins: application of a hydrophilic affinity method for glycopeptide enrichment
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwj019
Accession Number: 16037490
Keywords: Agglutinins/chemistry
Amino Acid Sequence
Apolipoprotein B-100
Apolipoproteins B/chemistry
Binding Sites
Cations
Chromatography
Chromatography, Affinity/*methods
Chromatography, High Pressure Liquid
Databases, Protein
Disulfides/chemistry
Fibrin/chemistry
Fibroblasts/metabolism
Fibronectins/*blood/*chemistry
Glycopeptides/*chemistry
Glycosylation
Heparin/chemistry
Humans
Hydrogen Bonding
Lectins/chemistry
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/chemistry
Polysaccharides/*chemistry
Protein Binding
Protein Structure, Tertiary
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Threonine/chemistry
Time Factors
Transferrin/chemistry
Trypsin/chemistry
Abstract: Isolation of glycopeptides utilizing hydrogen bonding between glycopeptide glycans and a carbohydrate-gel matrix in the organic phase is useful for site-specific characterization of oligosaccharides of glycoproteins, when combined with mass spectrometry. In this study, recovery of glycopeptides was improved by including divalent cations or increasing the organic solvent in the binding solution, without losing specificity, whereas it was still less effective for those with a long peptide backbone exceeding 50 amino acid residues. The method was then applied to the analysis of glycan heterogeneities at seven N-glycosylation sites in each of the plasma and cellular fibronectins (FNs). There was a remarkable site-specific difference in fucosylation between these isoforms; Asn1244 selectively escaped the global fucosylation of cellular FN, whereas only Asn1007 and Asn2108 of the plasma isoform underwent modification. In addition, a new O-glycosylation site was identified at Thr279 in the connecting segment between the fibrin- and heparin-binding domain and the collagen-binding domain, and the glycopeptide was reactive to a peanut agglutinin lectin. Considering that another mucin-type O-glycosylation site lies within a different connecting segment, the O-glycosylation of FN was suggested to play a significant role in segregating the neighboring domains and thus maintaining the topology of FN and the domain functions. In addition, the method was applied to apolipoprotein B-100 (apoB100) whose N-glycan structures at 17 of 19 potential sites have been reported, and characterized the remaining sites. The results also demonstrated that the enriched glycopeptide provides resources for site-specific analysis of oligosaccharides in glycoproteomics.
Notes: Tajiri, Michiko
Yoshida, Shumi
Wada, Yoshinao
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 2005 Dec;15(12):1332-40. doi: 10.1093/glycob/cwj019. Epub 2005 Jul 21.
Author Address: CREST, Japan Science and Technology Agency, 4-1-8 Honcho Kawaguchi, Saitama 332-0012, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1341
Author: Takahashi, K., Wall, S. B., Suzuki, H., Smith, A. D. th, Hall, S., Poulsen, K., Kilian, M., Mobley, J. A., Julian, B. A., Mestecky, J., Novak, J. and Renfrow, M. B.
Year: 2010
Title: Clustered O-glycans of IgA1: defining macro- and microheterogeneity by use of electron capture/transfer dissociation
Journal: Mol Cell Proteomics
Volume: 9
Issue: 11
Pages: 2545-57
Epub Date: 2010/09/09
Date: Nov
Short Title: Clustered O-glycans of IgA1: defining macro- and microheterogeneity by use of electron capture/transfer dissociation
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M110.001834
PMCID: PMC2984237
Accession Number: 20823119
Keywords: Amino Acid Sequence
*Electrons
Fourier Analysis
Glomerulonephritis, IGA/metabolism
Glycopeptides/analysis
Glycosylation
Humans
Immunoglobulin A/*chemistry
Mass Spectrometry
Molecular Sequence Data
Polysaccharides/*chemistry
Abstract: IgA nephropathy (IgAN) is the most common primary glomerulonephritis in the world. Aberrantly glycosylated IgA1, with galactose (Gal)-deficient hinge region (HR) O-glycans, plays a pivotal role in the pathogenesis of the disease. It is not known whether the glycosylation defect occurs randomly or preferentially at specific sites. We have described the utility of activated ion-electron capture dissociation (AI-ECD) mass spectrometric analysis of IgA1 O-glycosylation. However, locating and characterizing the entire range of O-glycan attachment sites are analytically challenging due to the clustered serine and threonine residues in the HR of IgA1 heavy chain. To address this problem, we analyzed all glycoforms of the HR glycopeptides of a Gal-deficient IgA1 myeloma protein, mimicking the aberrant IgA1 in patients with IgAN, by use of a combination of IgA-specific proteases + trypsin and AI-ECD Fourier transform ion cyclotron resonance (FT-ICR) tandem mass spectrometry (MS/MS). The IgA-specific proteases provided a variety of IgA1 HR fragments that allowed unambiguous localization of all O-glycosylation sites in the six most abundant glycoforms, including the sites deficient in Gal. Additionally, this protocol was adapted for on-line liquid chromatography (LC)-AI-ECD MS/MS and LC-electron transfer dissociation MS/MS analysis. Our results thus represent a new clinically relevant approach that requires ECD/electron transfer dissociation-type fragmentation to define the molecular events leading to pathogenesis of a chronic kidney disease. Furthermore, this work offers generally applicable principles for the analysis of clustered sites of O-glycosylation.
Notes: 1535-9484
Takahashi, Kazuo
Wall, Stephanie B
Suzuki, Hitoshi
Smith, Archer D 4th
Hall, Stacy
Poulsen, Knud
Kilian, Mogens
Mobley, James A
Julian, Bruce A
Mestecky, Jiri
Novak, Jan
Renfrow, Matthew B
DK082753/DK/NIDDK NIH HHS/United States
R01 GM098539/GM/NIGMS NIH HHS/United States
R21 DK077279-01A2/DK/NIDDK NIH HHS/United States
R21 DK075868/DK/NIDDK NIH HHS/United States
R21 DK077279-02/DK/NIDDK NIH HHS/United States
RR17261/RR/NCRR NIH HHS/United States
DK080301/DK/NIDDK NIH HHS/United States
DK077279/DK/NIDDK NIH HHS/United States
R21 DK080301/DK/NIDDK NIH HHS/United States
R01 DK078244/DK/NIDDK NIH HHS/United States
DK071802/DK/NIDDK NIH HHS/United States
R21 DK083663/DK/NIDDK NIH HHS/United States
S10 RR017261/RR/NCRR NIH HHS/United States
R01 DK071802/DK/NIDDK NIH HHS/United States
DK083663/DK/NIDDK NIH HHS/United States
R01 DK082753/DK/NIDDK NIH HHS/United States
DK078244/DK/NIDDK NIH HHS/United States
R56 DK078244/DK/NIDDK NIH HHS/United States
R21 DK077279/DK/NIDDK NIH HHS/United States
DK075868/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2010 Nov;9(11):2545-57. doi: 10.1074/mcp.M110.001834. Epub 2010 Sep 7.
Author Address: Biomedical FT-ICR MS Laboratory, Department of Biochemistry and Molecular Genetics, University of Alabama, Birmingham, Alabama 35294-0005, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 411
Author: Takahashi, N., Ishii, I., Ishihara, H., Mori, M., Tejima, S., Jefferis, R., Endo, S. and Arata, Y.
Year: 1987
Title: Comparative structural study of the N-linked oligosaccharides of human normal and pathological immunoglobulin G
Journal: Biochemistry
Volume: 26
Issue: 4
Pages: 1137-44
Epub Date: 1987/02/24
Date: Feb 24
Short Title: Comparative structural study of the N-linked oligosaccharides of human normal and pathological immunoglobulin G
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00378a023
Accession Number: 3567160
Keywords: Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Humans
*Immunoglobulin G/isolation & purification
Magnetic Resonance Spectroscopy
Multiple Myeloma/*immunology
*Oligosaccharides/isolation & purification
Abstract: The structures of oligosaccharides of normal and pathological immunoglobulin G (IgG) are reported. Asparagine-linked neutral oligosaccharides were released by N-oligosaccharide glycopeptidase (almond) digestion. The reducing ends of the oligosaccharide chains thus obtained were aminated with a fluorescent reagent, 2-aminopyridine, and the mixture of pyridylamino derivatives of the oligosaccharides was separated by reverse-phase high-performance liquid chromatography. It was possible to separate 15 out of the 16 kinds of oligosaccharides that have been suggested to exist in normal human IgG. High-resolution proton nuclear magnetic resonance spectroscopy was used along with chemical methods to determine the structures of the separated oligosaccharides. It has been shown that in normal IgG a biantennary complex-type oligosaccharide with a fucose residue (formula; see text) is predominant and four kinds of oligosaccharides, which are biantennary with bisecting N-acetylglucosamine and without fucose residues, exist only in a very small quantity. The results obtained for normal IgG were compared with those obtained for three myeloma IgG proteins. It has been found that the most abundant species that exist in the pathological proteins analyzed in the present work lack one or two galactose residues at the nonreducing terminal. We show that the fractions of fucose-containing oligosaccharides are markedly decreased in the heavy-chain disease protein Per. It is of particular interest that in this paraprotein the major component is a biantennary complex-type oligosaccharide that lacks a fucose residue and an oligosaccharide with the structure (Formula: see text) exists as one of the most abundant components.
Notes: Takahashi, N
Ishii, I
Ishihara, H
Mori, M
Tejima, S
Jefferis, R
Endo, S
Arata, Y
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1987 Feb 24;26(4):1137-44. doi: 10.1021/bi00378a023.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 512
Author: Takahashi, N., Nakagawa, H., Fujikawa, K., Kawamura, Y. and Tomiya, N.
Year: 1995
Title: Three-dimensional elution mapping of pyridylaminated N-linked neutral and sialyl oligosaccharides
Journal: Anal Biochem
Volume: 226
Issue: 1
Pages: 139-46
Epub Date: 1995/03/20
Date: Mar 20
Short Title: Three-dimensional elution mapping of pyridylaminated N-linked neutral and sialyl oligosaccharides
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1006/abio.1995.1201
Accession Number: 7540366
Keywords: Aminopyridines/chemistry
Animals
Blood Proteins/*chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Cattle
Chromatography, High Pressure Liquid
Humans
Immunoglobulin G/chemistry
Molecular Sequence Data
Neuraminidase/metabolism
Oligosaccharides/*analysis/chemistry
alpha-Fetoproteins/chemistry
alpha-L-Fucosidase/metabolism
beta-Galactosidase/metabolism
beta-N-Acetylhexosaminidases/metabolism
Abstract: We propose a three-dimensional (3-D) sugar-mapping technique for pyridylaminated (PA) neutral and sialyl oligosaccharides as a powerful structural characterization of N-linked oligosaccharides using only picomoles of samples. The new map consists of the elution data from 42 different sialyl oligosaccharides, 26 of which are mono-, 7 of which are di-, 7 of which are tri-, and 2 of which are tetra-sialylated oligosaccharides. The 20 standard sialyl oligosaccharides were released from human serum and calf fetuin by digestion with glycoamidase A. The other 22 standard sialyl oligosaccharides were obtained by subsequent digestion of the above 20 sialyl oligosaccharides with beta-galactosidase, beta-N-acetylhexosaminidase, alpha-fucosidase, and alpha 2-->3 specific sialidase. The present 3-D mapping method involves the following four steps: First, a neutral and sialyl PA-oligosaccharide mixture is separated by HPLC on the diethylaminoethyl (DEAE) column according to the sialic acid content, and the elution data are considered as one of the three dimensions (Z-axis). Then, neutral, mono-, di-, tri-, and tetra-sialyl oligosaccharides are individually separated on the octadecylsilyl (ODS)-silica (X-axis) and amide-silica (Y-axis) columns. The fourth step is to plot the coordinates on a two-dimensional (2-D) map. Thus, for each of the groups separated on the DEAE column, a 2-D map can be achieved. By repeating the whole process for each group of different sialylation, the layers of the 2-D map lined up on the Z-axis form a 3-D map.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Takahashi, N
Nakagawa, H
Fujikawa, K
Kawamura, Y
Tomiya, N
Journal Article
United States
Anal Biochem. 1995 Mar 20;226(1):139-46. doi: 10.1006/abio.1995.1201.
Author Address: GlycoLab, Nakano Vinegar Company Ltd., Handa-City, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 462
Author: Takahashi, N., Takahashi, Y. and Putnam, F. W.
Year: 1984
Title: Structure of human hemopexin: O-glycosyl and N-glycosyl sites and unusual clustering of tryptophan residues
Journal: Proc Natl Acad Sci U S A
Volume: 81
Issue: 7
Pages: 2021-5
Epub Date: 1984/04/01
Date: Apr
Short Title: Structure of human hemopexin: O-glycosyl and N-glycosyl sites and unusual clustering of tryptophan residues
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.81.7.2021
PMCID: PMC345428
Accession Number: 6371807
Keywords: Amino Acid Sequence
Cyanogen Bromide
Endopeptidases
Glycosides/*analysis
Hemin
*Hemopexin
Humans
Oligosaccharides/analysis
Peptide Fragments/analysis
Tryptophan/*analysis
Abstract: The primary structure of human hemopexin is being deduced from sequence analysis of a series of peptides obtained from chemical and enzymatic digests of the protein. Human hemopexin consists of about 440 amino acid residues. It has five sites of attachment of glucosamine oligosaccharides at the signal sequence of Asn-X-Thr/Ser. A unique structural feature is the virtual blocking of the amino-terminal threonine residue, which is O-linked to a galactosamine oligosaccharide that has not previously been identified in this protein. The galactosamine oligosaccharide and one glucosamine oligosaccharide are located in the amino-terminal region, three of the glucosamine oligosaccharides are in the middle region, and one glucosamine oligosaccharide is in the carboxyl-terminal region of the protein. Two of the five glucosamine oligosaccharides are present in a histidine-rich sequence of the middle region of the protein, in which the histidines flank beta-turns presumably at the surface of hemopexin. Clusters of tryptophan residues occur in four regions, each of which contains three or four tryptophan residues separated by 0-12 other residues. This clustering is significant because both histidine and tryptophan have been implicated in the binding of heme. A computer analysis did not identify significant matches of human hemopexin to any protein, including cytochromes and other heme-binding proteins, which suggests that the human hemopexin gene evolved from a unique primordial gene differing from those of other heme-binding proteins.
Notes: 1091-6490
Takahashi, N
Takahashi, Y
Putnam, F W
AM 19221/AM/NIADDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1984 Apr;81(7):2021-5. doi: 10.1073/pnas.81.7.2021.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 704
Author: Takahashi, N., Yamada, W., Masuda, K., Araki, H., Tsukamoto, Y., Galinha, A., Sautès, C., Kato, K. and Shimada, I.
Year: 1998
Title: N-glycan structures of a recombinant mouse soluble Fcgamma receptor II
Journal: Glycoconj J
Volume: 15
Issue: 9
Pages: 905-14
Epub Date: 1999/03/03
Date: Sep
Short Title: N-glycan structures of a recombinant mouse soluble Fcgamma receptor II
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1023/a:1006915200989
Accession Number: 10052594
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Cells, Cultured
Chromatography, High Pressure Liquid
Glycoproteins/chemistry
Mice
Molecular Sequence Data
Oligosaccharides/chemistry
Polysaccharides/*chemistry
Receptors, IgG/*chemistry/genetics
Recombinant Proteins/chemistry
Solubility
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: N-glycans of a recombinant mouse soluble Fcgamma receptor II (sFcgammaRII) expressed in baby hamster kidney cells were released from glycopeptides by digestion with glycoamidase A (from sweet almond), and the reducing ends of the oligosaccharides were reductively aminated with 2-aminopyridine. The derivatized N-glycans were separated and structurally identified by a three-dimensional high-performance liquid chromatography (HPLC) mapping technique on three kinds of HPLC columns [Takahashi, et al. (1995) Anal. Biochem. 226:139-46]. Eighteen different major N-glycan structures were identified, of which six were neutral (45%), five mono-sialyl (49%), one di-sialyl (4.6%), five tri-sialyl (1.1%), and one tetra-sialyl (0.3%). All N-glycan structures determined were complex type with fucosylation at the N-acetylglucosamine residue of the reducing end, and N-acetylneuraminic acid, when present, was alpha-(2,3)-linked. The existence of a unique structure containing both N-acetylgalactosamine and alpha-(2,3)-N-acetylneuraminic acid residues at the reducing ends, as below, was confirmed by MALDI-TOF mass spectrometry. Carbohydrate sequence [see text]
Notes: Takahashi, N
Yamada, W
Masuda, K
Araki, H
Tsukamoto, Y
Galinha, A
Sautès, C
Kato, K
Shimada, I
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 1998 Sep;15(9):905-14. doi: 10.1023/a:1006915200989.
Author Address: GlycoLab, Nakano Vinegar Co., Ltd., Handa-City Aichi, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 324
Author: Takahashi, T., Yamaguchi, N., Mita, S., Yamaguchi, Y., Suda, T., Tominaga, A., Kikuchi, Y., Miura, Y. and Takatsu, K.
Year: 1990
Title: Structural comparison of murine T-cell (B151K12)-derived T-cell-replacing factor (IL-5) with rIL-5: dimer formation is essential for the expression of biological activity
Journal: Mol Immunol
Volume: 27
Issue: 9
Pages: 911-20
Epub Date: 1990/09/01
Date: Sep
Short Title: Structural comparison of murine T-cell (B151K12)-derived T-cell-replacing factor (IL-5) with rIL-5: dimer formation is essential for the expression of biological activity
Alternate Journal: Molecular immunology
ISSN: 0161-5890 (Print)
0161-5890
DOI: 10.1016/0161-5890(90)90158-v
Accession Number: 2215480
Keywords: Amino Acid Sequence
Animals
Cell Differentiation
Cell Line
Chromatography, Affinity
Disulfides
Eosinophils/cytology
Female
Interleukin-5/*chemistry/isolation & purification/*physiology
Macromolecular Substances
Mice
Mice, Inbred BALB C
Molecular Sequence Data
Protein Binding
Recombinant Proteins/chemistry
Structure-Activity Relationship
T-Lymphocytes/metabolism
Abstract: T-cell-replacing factor (TRF)/IL-5 is a T-cell-derived glycoprotein which has pleiotropic activity on lymphoid and myeloid cells. IL-5 polypeptide translated into Xenopus oocytes are heterogeneous in molecular size (40,000 to 60,000 under nonreducing conditions) and yields a monomeric form (Mr of 25,000 to 30,000) under reducing conditions (J. Immun., 140, 1175-1181, 1988). We purified T-cell-derived TRF and rIL-5 using anti-TRF/IL-5 antibody-coupled affinity column from supernatants of a T-cell hybridoma B151K12 and supernatants of HeLa cells, respectively, which had been transfected with murine IL-5 cDNA, and determined their partial N-terminal amino acid sequence (27 residues for B151-TRF and 13 residues for rIL-5). A single amino acid sequence of each sample obtained beginning from methionine that was identical to that predicted from IL-5 cDNA. This finding supports the notion that secreted B151-TRF polypeptide consists of 113 amino acids. Purified B151-TRF supported eosinophilopoiesis of human bone marrow cells as effective as mouse rIL-5 and human rIL-5. B151-TRF competitively inhibited 35S-labeled rIL-5 binding to target cells to the same extent at rIL-5. Treatment of purified rIL-5 and B151-TRF with reducing reagents such as 2-ME, sodium borohydride or dithiothreitol produced a monomeric form of IL-5 which did not exert a biological activity. Reduction and alkylation of rIL-5 caused the loss of binding to its target cells. These results strongly suggest that B151-TRF exists as a homodimer and its primary structure and secondary structures are identical to those of rIL-5. Moreover, the formation of inter-molecular disulfide bond(s) linked by two pairs of cystein residues is essential for the expression of the biological activity of mouse IL-5.
Notes: Takahashi, T
Yamaguchi, N
Mita, S
Yamaguchi, Y
Suda, T
Tominaga, A
Kikuchi, Y
Miura, Y
Takatsu, K
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
England
Mol Immunol. 1990 Sep;27(9):911-20. doi: 10.1016/0161-5890(90)90158-v.
Author Address: Department of Biology, Kumamoto University Medical School, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1627
Author: Takakura, D., Harazono, A., Hashii, N. and Kawasaki, N.
Year: 2014
Title: Selective glycopeptide profiling by acetone enrichment and LC/MS
Journal: J Proteomics
Volume: 101
Pages: 17-30
Epub Date: 2014/02/18
Date: Apr 14
Short Title: Selective glycopeptide profiling by acetone enrichment and LC/MS
Alternate Journal: Journal of proteomics
ISSN: 1874-3919
DOI: 10.1016/j.jprot.2014.02.005
Accession Number: 24530628
Keywords: Acetone/*pharmacology
Amino Acid Sequence
Blood Chemical Analysis/methods
Blood Proteins/*isolation & purification/*metabolism
Carbohydrate Sequence
Chromatography, Liquid/methods
Glycopeptides/*isolation & purification/*metabolism
Humans
Mass Spectrometry/*methods
Metabolome
Molecular Sequence Data
Tandem Mass Spectrometry/methods
Acetone
Glycopeptide enrichment
Lc/ms
Abstract: LC/MS is commonly used for site-specific glycosylation analysis of glycoproteins in cells and tissues. A limitation of this technique is the difficulty in acquiring reliable mass spectra for glycopeptides, mainly due to their high heterogeneity and poor hydrophobicity. Here, we establish a versatile method for efficient glycopeptide enrichment to acquire reliable mass spectra. Several lines of evidence using model glycoproteins suggest that our method is based on the different solubility between non-glycosylated and glycosylated peptides in acetone. We also provide data showing that the acetone-precipitated glycopeptide enrichment was successful in acquiring a more comprehensive MS/MS data set for the various glycoforms of each glycopeptide in crude human serum. We propose that this method is a powerful tool for the acquisition of reliable mass spectra from trace amounts of glycopeptides and an alternative to lectin affinity enrichment. BIOLOGICAL SIGNIFICANCE: In this study, we established a versatile method for glycopeptide enrichment to acquire reliable mass spectra because of the limitation of conventional enrichment methods, such as lectin affinity chromatography. Our enrichment method is capable of isolating glycopeptides from complex peptide mixtures such as crude serum.
Notes: 1876-7737
Takakura, Daisuke
Harazono, Akira
Hashii, Noritaka
Kawasaki, Nana
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
J Proteomics. 2014 Apr 14;101:17-30. doi: 10.1016/j.jprot.2014.02.005. Epub 2014 Feb 12.
Author Address: Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan.
Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan. Electronic address: nana@nihs.go.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 607
Author: Takakura-Yamamoto, R., Yamamoto, S., Fukuda, S. and Kurimoto, M.
Year: 1996
Title: O-glycosylated species of natural human tumor-necrosis factor-alpha
Journal: Eur J Biochem
Volume: 235
Issue: 1-2
Pages: 431-7
Epub Date: 1996/01/15
Date: Jan 15
Short Title: O-glycosylated species of natural human tumor-necrosis factor-alpha
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1996.00431.x
Accession Number: 8631363
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Cell Line
Glycosylation
Humans
Molecular Sequence Data
Molecular Structure
Oligosaccharides/chemistry/isolation & purification
Peptides
Sugar Alcohols/chemistry/isolation & purification
Tumor Necrosis Factor-alpha/*chemistry/genetics/isolation & purification
Abstract: Tumor-necrosis factor-alpha, produced by human B-cell lymphoblastoid cell line BALL-1, was expressed as four protein bands on SDS/PAGE analysis. It may have been glycosylated, based on the fact that the heavier two of the four bands disappeared after neuraminidase treatment. Sugar composition analyses revealed that the tumor necrosis factor-alpha contained galactose, N-acetylgalactosamine and N-acetylneuraminic acid as sugar components. To prepare sugar chains, tumor necrosis factor-alpha was treated with alkaline borodeuteride and the oligosaccharide-alditols liberated were fractionated by gel-filtration chromatography on a Bio-Gel P-4 column, followed by normal-phase HPLC. Three oligosaccharide-alditols were obtained, and the structures of two of them were identified by methylation analysis and exoglycosidase digestion. The structures of these oligosaccharide-alditols were Gal beta 1-3(NeuAc alpha 2-6)GalNAcol and Gal beta 1-3GalNAcol (GalNAcol, N-acetylgalactosaminitol). The structure of the remaining oligosaccharide-alditol was determined to be NeuAc alpha 2-3Gal1-3GalNAcol by composition and methylation analyses. About 20% of tumor necrosis factor-alpha was found to be 0-glycosylated, based on the results of the sugar composition and structure analyses. An amino acid sequence analysis of the glycosylated peptides was performed after Staphylococcus aureus V8 protease digestion of tumor necrosis factor-alpha had been completed, and it was proved that the 0-glycosylation site of tumor necrosis factor-alpha was Ser 4.
Notes: Takakura-Yamamoto, R
Yamamoto, S
Fukuda, S
Kurimoto, M
Journal Article
England
Eur J Biochem. 1996 Jan 15;235(1-2):431-7. doi: 10.1111/j.1432-1033.1996.00431.x.
Author Address: Fujisaki Institute, Hayashibara Biochemical Laboratories, Inc., Okayama, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 948
Author: Takamatsu, S., Katsumata, T., Inoue, N., Watanabe, T., Fujibayashi, Y. and Takeuchi, M.
Year: 2004
Title: Abnormal biantennary sugar chains are expressed in human chorionic gonadotropin produced in the choriocarcinoma cell line, JEG-3
Journal: Glycoconj J
Volume: 20
Issue: 7-8
Pages: 473-81
Epub Date: 2004/08/19
Short Title: Abnormal biantennary sugar chains are expressed in human chorionic gonadotropin produced in the choriocarcinoma cell line, JEG-3
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1023/B:GLYC.0000038293.37376.9f
Accession Number: 15316280
Keywords: Carbohydrate Sequence
Cell Line, Tumor
Choriocarcinoma/chemistry/*metabolism
Chorionic Gonadotropin/*chemistry/metabolism
Female
Humans
Molecular Sequence Data
N-Acetylglucosaminyltransferases/*metabolism
Oligosaccharides/analysis/*biosynthesis
Pregnancy
Abstract: Over the past two decades, sugar chain structures of human chorionic gonadotropin (hCG) produced in healthy people, three types of trophoblastic disease, and some types of cell lines have been analyzed. The abnormal biantennary structure of hCG is a good marker for the diagnosis of malignant choriocarcinoma. In spite of much research, hCG with an abnormal biantennary structure is only detected in the urine of choriocarcinoma or pregnant diabetic patients. We hypothesized that the formation mechanism of the abnormal biantennary sugar chain structure is mainly caused by high GnT-IV activity. To confirm this, we measured the N-acetylglucosaminyltransferase (GnT)-IV activity and hCG productivity in three choriocarcinoma cell lines, and selected JEG-3 cells. hCG samples were purified from medium conditioned by JEG-3 cells, and their sugar chain structures were analyzed. We detected an abnormal biantennary structure, and the proportions were different from those previously reported in the urine samples of choriocarcinoma patients. These findings proved our hypothesis and suggest the usefulness of JEG-3 cells for further analyses of abnormal biantennary structure formation.
Notes: Takamatsu, Shinji
Katsumata, Toshiyuki
Inoue, Noboru
Watanabe, Toshinori
Fujibayashi, Yasuhisa
Takeuchi, Makoto
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 2004;20(7-8):473-81. doi: 10.1023/B:GLYC.0000038293.37376.9f.
Author Address: Biomedical Imaging Research Center, University of Fukui, 23-3 Shimoaizuki, Matsuoka, Yoshida, Fukui 910-1193, Japan. shinjit@fmsrsa.fukui-med.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 501
Author: Takasaki, S., Yamashita, K., Suzuki, K. and Kobata, A.
Year: 1980
Title: Structural studies of the sugar chains of cold-insoluble globulin isolated from human plasma
Journal: J Biochem
Volume: 88
Issue: 6
Pages: 1587-94
Epub Date: 1980/12/01
Date: Dec
Short Title: Structural studies of the sugar chains of cold-insoluble globulin isolated from human plasma
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a133133
Accession Number: 7462199
Keywords: Carbohydrate Sequence
Chemical Phenomena
Chemistry
Electrophoresis, Paper
Fibronectins/*blood
Humans
Hydrazines
Hydrogen-Ion Concentration
Methylation
Oligosaccharides/analysis
Sialic Acids/analysis
Abstract: The asparagine-linked sugar chains of cold-insoluble globulin isolated from human plasma were released as oligosaccharides from the polypeptide moiety by hydrazinolysis. These oligosaccharides were N-acetylated and could be labeled by reduction with NaB[3H]4. The yield of radioactive oligosaccharides indicated that the glycoprotein has four asparagine-linked sugar chains in one molecule. More than 90% of the radioactive oligosaccharides contain N-acetylneuraminic acid, and could be separated into two acidic oligosaccharides, A-1 and A-2. By sequential exoglycosidase digestion in combination with methylation studies, their structures were elucidated as Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3) Man beta 1 leads to 4GlcNAc beta 1 leads to 4GlcNAc and NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6 (NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2 Man alpha 1 leads to 3)-Man beta 1 leads to 4GlcNAc beta 1 leads to 4GlcNAc.
Notes: Takasaki, S
Yamashita, K
Suzuki, K
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
England
J Biochem. 1980 Dec;88(6):1587-94. doi: 10.1093/oxfordjournals.jbchem.a133133.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 486
Author: Takayasu, T., Suzuki, S., Kametani, F., Takahashi, N., Shinoda, T., Okuyama, T. and Munekata, E.
Year: 1982
Title: Amino acid sequence of galactosamine-containing glycopeptides in the hinge region of a human immunoglobulin D
Journal: Biochem Biophys Res Commun
Volume: 105
Issue: 3
Pages: 1066-71
Epub Date: 1982/04/14
Date: Apr 14
Short Title: Amino acid sequence of galactosamine-containing glycopeptides in the hinge region of a human immunoglobulin D
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(82)91078-6
Accession Number: 7092891
Keywords: Amino Acid Sequence
Galactosamine/*analysis
Glycopeptides/analysis
Humans
*Immunoglobulin D/isolation & purification
Multiple Myeloma/immunology
Protein Conformation
Notes: Takayasu, T
Suzuki, S
Kametani, F
Takahashi, N
Shinoda, T
Okuyama, T
Munekata, E
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 1982 Apr 14;105(3):1066-71. doi: 10.1016/0006-291x(82)91078-6.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 481
Author: Takayasu, T., Takahashi, N. and Shinoda, T.
Year: 1980
Title: Amino acid sequence and location of the three glycopeptides in the Fc region of human immunoglobulin D
Journal: Biochem Biophys Res Commun
Volume: 97
Issue: 2
Pages: 635-41
Epub Date: 1980/11/28
Date: Nov 28
Short Title: Amino acid sequence and location of the three glycopeptides in the Fc region of human immunoglobulin D
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(80)90311-3
Accession Number: 7008791
Keywords: Amino Acid Sequence
Glucosamine/analysis
Glycopeptides/*analysis
Humans
*Immunoglobulin D
*Immunoglobulin Fc Fragments
Peptide Hydrolases
Staphylococcus aureus/enzymology
Notes: Takayasu, T
Takahashi, N
Shinoda, T
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 1980 Nov 28;97(2):635-41. doi: 10.1016/0006-291x(80)90311-3.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 504
Author: Taketa, K., Fujii, Y. and Taga, H.
Year: 1993
Title: Characterization of E-PHA-reactive alpha-fetoprotein isoforms by two-dimensional lectin affinity electrophoresis
Journal: Electrophoresis
Volume: 14
Issue: 12
Pages: 1333-7
Epub Date: 1993/12/01
Date: Dec
Short Title: Characterization of E-PHA-reactive alpha-fetoprotein isoforms by two-dimensional lectin affinity electrophoresis
Alternate Journal: Electrophoresis
ISSN: 0173-0835 (Print)
0173-0835
DOI: 10.1002/elps.11501401205
Accession Number: 7511099
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Concanavalin A
Electrophoresis, Agar Gel
*Electrophoresis, Gel, Two-Dimensional
Fetal Blood/*chemistry
Molecular Sequence Data
Molecular Structure
Neuraminidase/pharmacology
*Phytohemagglutinins
alpha-Fetoproteins/*analysis/chemistry
beta-Galactosidase/pharmacology
Abstract: Erythroagglutinating phytohemagglutinin (E-PHA)-dependent isoforms of human alpha-fetoprotein (AFP) from cord blood were analyzed for their carbohydrate structures by two-dimensional electrophoresis with E-PHA combined with extended agarose gel electrophoresis or with affinity electrophoresis with concanavalin A or Allomyrina dichtoma lectin. By means of neuraminidase and/or beta-galactosidase treatment, AFP-P2 was identified as alpha 2-->6 disialo-AFP, AFP-P3 as having biantennary structures with alpha 2-->6 monosialylated galactose of the Mannose (Man) alpha 1-->6 arm, AFP-P4 as having alpha 2-->6 monosialylated galactose of the Man alpha 1-->3 arm, and AFP-P5 as disialo-AFP with alpha 2-->3 sialylated galactose of the Man alpha 1-->6 antenna with the alpha 2-->6 sialylated galactose of the other antenna. Desialylated AFP with the terminal galactose of the Man alpha 1-->6 antenna with or without the galactose of the other arm also had a migration of AFP-P4, and other hydrolytic intermediates without the terminal galactose of the Man alpha 1-->6 arm with and without the galactose of the other antenna had mobilities of AFP-P3s and AFP-P3, respectively. Thus, the present system of two-dimensional lectin affinity electrophoreses would provide a model for the determination of the sugar chain structure of glycoproteins.
Notes: Taketa, K
Fujii, Y
Taga, H
Journal Article
Research Support, Non-U.S. Gov't
Germany
Electrophoresis. 1993 Dec;14(12):1333-7. doi: 10.1002/elps.11501401205.
Author Address: Department of Public Health, Okayama University Medical School, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1429
Author: Takeuchi, H., Fernández-Valdivia, R. C., Caswell, D. S., Nita-Lazar, A., Rana, N. A., Garner, T. P., Weldeghiorghis, T. K., Macnaughtan, M. A., Jafar-Nejad, H. and Haltiwanger, R. S.
Year: 2011
Title: Rumi functions as both a protein O-glucosyltransferase and a protein O-xylosyltransferase
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 40
Pages: 16600-5
Epub Date: 2011/09/29
Date: Oct 4
Short Title: Rumi functions as both a protein O-glucosyltransferase and a protein O-xylosyltransferase
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1109696108
PMCID: PMC3189016
Accession Number: 21949356
Keywords: Animals
Drosophila/genetics/*physiology
Drosophila Proteins/genetics/*metabolism
Factor VII/genetics/*metabolism
Glucosyltransferases/genetics/*metabolism
Humans
Mass Spectrometry
Mice
Mutation/genetics
Pentosyltransferases/*metabolism
Signal Transduction/genetics/*physiology
Substrate Specificity
Abstract: Mutations in rumi result in a temperature-sensitive loss of Notch signaling in Drosophila. Drosophila Rumi is a soluble, endoplasmic reticulum-retained protein with a CAP10 domain that functions as a protein O-glucosyltransferase. In human and mouse genomes, three potential Rumi homologues exist: one with a high degree of identity to Drosophila Rumi (52%), and two others with lower degrees of identity but including a CAP10 domain (KDELC1 and KDELC2). Here we show that both mouse and human Rumi, but not KDELC1 or KDELC2, catalyze transfer of glucose from UDP-glucose to an EGF repeat from human factor VII. Similarly, human Rumi, but not KDELC1 or KDELC2, rescues the Notch phenotypes in Drosophila rumi clones. During characterization of the Rumi enzymes, we noted that, in addition to protein O-glucosyltransferase activity, both mammalian and Drosophila Rumi also showed significant protein O-xylosyltransferase activity. Rumi transfers Xyl or glucose to serine 52 in the O-glucose consensus sequence ( ) of factor VII EGF repeat. Surprisingly, the second serine (S53) facilitates transfer of Xyl, but not glucose, to the EGF repeat by Rumi. EGF16 of mouse Notch2, which has a diserine motif in the consensus sequence ( ), is also modified with either O-Xyl or O-glucose glycans in cells. Mutation of the second serine (S590A) causes a loss of O-Xyl but not O-glucose at this site. Altogether, our data establish dual substrate specificity for the glycosyltransferase Rumi and provide evidence that amino acid sequences of the recipient EGF repeat significantly influence which donor substrate (UDP-glucose or UDP-Xyl) is used.
Notes: 1091-6490
Takeuchi, Hideyuki
Fernández-Valdivia, Rodrigo C
Caswell, Devin S
Nita-Lazar, Aleksandra
Rana, Nadia A
Garner, Thomas P
Weldeghiorghis, Thomas K
Macnaughtan, Megan A
Jafar-Nejad, Hamed
Haltiwanger, Robert S
R01 GM061126/GM/NIGMS NIH HHS/United States
R01 GM084135/GM/NIGMS NIH HHS/United States
GM084135/GM/NIGMS NIH HHS/United States
GM61126/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 Oct 4;108(40):16600-5. doi: 10.1073/pnas.1109696108. Epub 2011 Sep 26.
Author Address: Department of Biochemistry and Cell Biology, Institute of Cell and Developmental Biology, Stony Brook University, Stony Brook, NY 11794, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1962
Author: Takeuchi, H., Schneider, M., Williamson, D. B., Ito, A., Takeuchi, M., Handford, P. A. and Haltiwanger, R. S.
Year: 2018
Title: Two novel protein O-glucosyltransferases that modify sites distinct from POGLUT1 and affect Notch trafficking and signaling
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 36
Pages: E8395-e8402
Epub Date: 2018/08/22
Date: Sep 4
Short Title: Two novel protein O-glucosyltransferases that modify sites distinct from POGLUT1 and affect Notch trafficking and signaling
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1804005115
PMCID: PMC6130362
Accession Number: 30127001
Keywords: Animals
Glucosyltransferases/genetics/*metabolism
Glycosylation
HEK293 Cells
Humans
Mice
NIH 3T3 Cells
Protein Transport/physiology
Receptor, Notch1/genetics/*metabolism
Receptor, Notch2/genetics/*metabolism
Receptor, Notch3/genetics/*metabolism
Repetitive Sequences, Amino Acid
Signal Transduction/*physiology
*Notch
*O-glucose
*development
*glycosyltransferases
*signal transduction
Abstract: The Notch-signaling pathway is normally activated by Notch-ligand interactions. A recent structural analysis suggested that a novel O-linked hexose modification on serine 435 of the mammalian NOTCH1 core ligand-binding domain lies at the interface with its ligands. This serine occurs between conserved cysteines 3 and 4 of Epidermal Growth Factor-like (EGF) repeat 11 of NOTCH1, a site distinct from those modified by protein O-glucosyltransferase 1 (POGLUT1), suggesting that a different enzyme is responsible. Here, we identify two novel protein O-glucosyltransferases, POGLUT2 and POGLUT3 (formerly KDELC1 and KDELC2, respectively), which transfer O-glucose (O-Glc) from UDP-Glc to serine 435. Mass spectrometric analysis of NOTCH1 produced in HEK293T cells lacking POGLUT2, POGLUT3, or both genes showed that either POGLUT2 or POGLUT3 can add this novel O-Glc modification. EGF11 of NOTCH2 does not have a serine residue in the same location for this O-glucosylation, but EGF10 of NOTCH3 (homologous to EGF11 in NOTCH1 and -2) is also modified at the same position. Comparison of the sites suggests a consensus sequence for modification. In vitro assays with POGLUT2 and POGLUT3 showed that both enzymes modified only properly folded EGF repeats and displayed distinct acceptor specificities toward NOTCH1 EGF11 and NOTCH3 EGF10. Mutation of the O-Glc modification site on EGF11 (serine 435) in combination with sensitizing O-fucose mutations in EGF8 or EGF12 affected cell-surface presentation of NOTCH1 or reduced activation of NOTCH1 by Delta-like1, respectively. This study identifies a previously undescribed mechanism for fine-tuning the Notch-signaling pathway in mammals.
Notes: 1091-6490
Takeuchi, Hideyuki
Schneider, Michael
Williamson, Daniel B
Ito, Atsuko
Takeuchi, Megumi
Handford, Penny A
Haltiwanger, Robert S
R01 GM061126/GM/NIGMS NIH HHS/United States
T32 GM107004/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Proc Natl Acad Sci U S A. 2018 Sep 4;115(36):E8395-E8402. doi: 10.1073/pnas.1804005115. Epub 2018 Aug 20.
Author Address: Complex Carbohydrate Research Center, The University of Georgia, Athens, GA 30602.
Department of Biochemistry and Cell Biology, Stony Brook University, Stony Brook, NY 11795.
Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom.
Complex Carbohydrate Research Center, The University of Georgia, Athens, GA 30602; rhalti@uga.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 364
Author: Takeuchi, M., Inoue, N., Strickland, T. W., Kubota, M., Wada, M., Shimizu, R., Hoshi, S., Kozutsumi, H., Takasaki, S. and Kobata, A.
Year: 1989
Title: Relationship between sugar chain structure and biological activity of recombinant human erythropoietin produced in Chinese hamster ovary cells
Journal: Proc Natl Acad Sci U S A
Volume: 86
Issue: 20
Pages: 7819-22
Epub Date: 1989/10/01
Date: Oct
Short Title: Relationship between sugar chain structure and biological activity of recombinant human erythropoietin produced in Chinese hamster ovary cells
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.86.20.7819
PMCID: PMC298162
Accession Number: 2813359
Keywords: Animals
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Cricetulus
Erythropoietin/genetics/isolation & purification/*pharmacology
Female
Genes
Humans
Molecular Sequence Data
Oligosaccharides/*isolation & purification
Ovary
Recombinant Proteins/isolation & purification/pharmacology
Transfection
Abstract: Two forms of erythropoietin, EPO-bi and EPO-tetra, with different biological activities were isolated from the culture medium of a recombinant Chinese hamster ovary cell line, B8-300, into which the human erythropoietin gene had been introduced. EPO-bi, an unusual form, showed only one-seventh the in vivo activity and 3 times higher in vitro activity of the previously described recombinant human EPO (standard EPO). In contrast, EPO-tetra showed both in vivo and in vitro activities comparable to those of the standard EPO. EPO-bi, EPO-tetra, and the standard EPO had the same amino acid composition and immunoreactivity. However, structural analyses of their N-linked sugar chains revealed that EPO-bi contains the biantennary complex type as the major sugar chain, while EPO-tetra and the standard EPO contain the tetraantennary complex type as the major sugar chain. From examination of various preparations of recombinant human EPO, we found a positive correlation between the in vivo activity of EPO and the ratio of tetraantennary to biantennary oligosaccharides. These results suggest that higher branching of the N-linked sugar chains is essential for effective expression of in vivo biological activity of EPO.
Notes: 1091-6490
Takeuchi, M
Inoue, N
Strickland, T W
Kubota, M
Wada, M
Shimizu, R
Hoshi, S
Kozutsumi, H
Takasaki, S
Kobata, A
Journal Article
Proc Natl Acad Sci U S A. 1989 Oct;86(20):7819-22. doi: 10.1073/pnas.86.20.7819.
Author Address: Pharmaceutical Laboratory, Kirin Brewery, Gunma, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 391
Author: Takeuchi, M., Takasaki, S., Miyazaki, H., Kato, T., Hoshi, S., Kochibe, N. and Kobata, A.
Year: 1988
Title: Comparative study of the asparagine-linked sugar chains of human erythropoietins purified from urine and the culture medium of recombinant Chinese hamster ovary cells
Journal: J Biol Chem
Volume: 263
Issue: 8
Pages: 3657-63
Epub Date: 1988/03/15
Date: Mar 15
Short Title: Comparative study of the asparagine-linked sugar chains of human erythropoietins purified from urine and the culture medium of recombinant Chinese hamster ovary cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 3346214
Keywords: Animals
*Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Cricetinae
Cricetulus
*Erythropoietin/isolation & purification/urine
Female
Humans
Methylation
Molecular Sequence Data
Oligosaccharides/*analysis
Ovary
Recombination, Genetic
Abstract: The asparagine-linked sugar chains of human erythropoietin produced by recombinant Chinese hamster ovary cells and naturally occurring human urinary erythropoietin were liberated by hydrazinolysis and fractionated by paper electrophoresis, lectin affinity chromatography, and Bio-Gel P-4 column chromatography. Both erythropoietins had three asparagine-linked sugar chains in one molecule, all of which were acidic complex type. Structural analysis of them revealed that the sugar chains from both erythropoietins are quite similar except for sialyl linkage. All sugar chains of erythropoietin produced by recombinant Chinese hamster ovary cells contain only the NeuAc alpha 2----3Gal linkage, while those of human urinary erythropoietin contain the NeuAc alpha 2----6Gal linkage together with the NeuAc alpha 2----3Gal linkage. The major sugar chains were of fucosylated tetraantennary complex type with and without N-acetyllactosamine repeating units in their outer chain moieties in common, and small amounts of 2,4- and 2,6-branched triantennary and biantennary sugar chains were detected. This paper proved, for the first time, that recombinant technique can produce glycoprotein hormone whose carbohydrate structures are common to the major sugar chains of the native one.
Notes: Takeuchi, M
Takasaki, S
Miyazaki, H
Kato, T
Hoshi, S
Kochibe, N
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1988 Mar 15;263(8):3657-63.
Author Address: Department of Biochemistry, University of Tokyo, KIRIN, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1721
Author: Takeuchi, Y., Morise, J., Morita, I., Takematsu, H. and Oka, S.
Year: 2015
Title: Role of Site-Specific N-Glycans Expressed on GluA2 in the Regulation of Cell Surface Expression of AMPA-Type Glutamate Receptors
Journal: PLoS One
Volume: 10
Issue: 8
Pages: e0135644
Epub Date: 2015/08/14
Short Title: Role of Site-Specific N-Glycans Expressed on GluA2 in the Regulation of Cell Surface Expression of AMPA-Type Glutamate Receptors
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0135644
PMCID: PMC4535760
Accession Number: 26271046
Keywords: CD57 Antigens/metabolism
Cell Line
Electrophoresis, Polyacrylamide Gel
*Gene Expression Regulation
Humans
Immunoprecipitation
Polysaccharides/*genetics/metabolism
Receptors, AMPA/*genetics/metabolism
Receptors, Glutamate/*genetics/metabolism
Abstract: The AMPA-type glutamate receptor (AMPAR), which is a tetrameric complex composed of four subunits (GluA1-4) with several combinations, mediates the majority of rapid excitatory synaptic transmissions in the nervous system. Cell surface expression levels of AMPAR modulate synaptic plasticity, which is considered one of the molecular bases for learning and memory formation. To date, a unique trisaccharide (HSO3-3GlcAβ1-3Galβ1-4GlcNAc), human natural killer-1 (HNK-1) carbohydrate, was found expressed specifically on N-linked glycans of GluA2 and regulated the cell surface expression of AMPAR and the spine maturation process. However, evidence that the HNK-1 epitope on N-glycans of GluA2 directly affects these phenomena is lacking. Moreover, it is thought that other N-glycans on GluA2 also have potential roles in the regulation of AMPAR functions. In the present study, using a series of mutants lacking potential N-glycosylation sites (N256, N370, N406, and N413) within GluA2, we demonstrated that the mutant lacking the N-glycan at N370 strongly suppressed the intracellular trafficking of GluA2 from the endoplasmic reticulum (ER) in HEK293 cells. Cell surface expression of GluA1, which is a major subunit of AMPAR in neurons, was also suppressed by co-expression of the GluA2 N370S mutant. The N370S mutant and wild-type GluA2 were co-immunoprecipitated with GluA1, suggesting that N370S was properly associated with GluA1. Moreover, we found that N413 was the main potential site of the HNK-1 epitope that promoted the interaction of GluA2 with N-cadherin, resulting in enhanced cell surface expression of GluA2. The HNK-1 epitope on N-glycan at the N413 of GluA2 was also involved in the cell surface expression of GluA1. Thus, our data suggested that site-specific N-glycans on GluA2 regulate the intracellular trafficking and cell surface expression of AMPAR.
Notes: 1932-6203
Takeuchi, Yusuke
Morise, Jyoji
Morita, Ippei
Takematsu, Hiromu
Oka, Shogo
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2015 Aug 13;10(8):e0135644. doi: 10.1371/journal.pone.0135644. eCollection 2015.
Author Address: Department of Biological Chemistry, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
Department of Biological Chemistry, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 468
Author: Takio, K., Towatari, T., Katunuma, N., Teller, D. C. and Titani, K.
Year: 1983
Title: Homology of amino acid sequences of rat liver cathepsins B and H with that of papain
Journal: Proc Natl Acad Sci U S A
Volume: 80
Issue: 12
Pages: 3666-70
Epub Date: 1983/06/01
Date: Jun
Short Title: Homology of amino acid sequences of rat liver cathepsins B and H with that of papain
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.80.12.3666
PMCID: PMC394111
Accession Number: 6574504
Keywords: Amino Acid Sequence
Animals
Cathepsin B
Cathepsin H
*Cathepsins
Cyanogen Bromide
*Cysteine Endopeptidases
Liver/*enzymology
Models, Molecular
*Papain
Peptide Fragments/analysis
Protein Conformation
Rats
Abstract: The amino acid sequences of rat liver lysosomal thiol endopeptidases, cathepsins B and H, are presented and compared with that of the plant thiol protease papain. The 252-residue sequence of cathepsin B and the 220-residue sequence of cathepsin H were determined largely by automated Edman degradation of their intact polypeptide chains and of the two chains of each enzyme generated by limited proteolysis. Subfragments of the chains were produced by enzymatic digestion and by chemical cleavage of methionyl and tryptophanyl bonds. Comparison of the amino acid sequences of cathepsins B and H with each other and with that of papain demonstrates a striking homology among their primary structures. Sequence identity is extremely high in regions which, according to the three-dimensional structure of papain, constitute the catalytic site. The results not only reveal the first structural features of mammalian thiol endopeptidases but also provide insight into the evolutionary relationships among plant and mammalian thiol proteases.
Notes: 1091-6490
Takio, K
Towatari, T
Katunuma, N
Teller, D C
Titani, K
GM 13401/GM/NIGMS NIH HHS/United States
GM 15731/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1983 Jun;80(12):3666-70. doi: 10.1073/pnas.80.12.3666.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1035
Author: Tamada, T., Honjo, E., Maeda, Y., Okamoto, T., Ishibashi, M., Tokunaga, M. and Kuroki, R.
Year: 2006
Title: Homodimeric cross-over structure of the human granulocyte colony-stimulating factor (GCSF) receptor signaling complex
Journal: Proc Natl Acad Sci U S A
Volume: 103
Issue: 9
Pages: 3135-40
Epub Date: 2006/02/24
Date: Feb 28
Short Title: Homodimeric cross-over structure of the human granulocyte colony-stimulating factor (GCSF) receptor signaling complex
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0511264103
PMCID: PMC1413920
Accession Number: 16492764
Keywords: Amino Acid Sequence
Binding Sites
Conserved Sequence
Crystallography, X-Ray
Cytokine Receptor gp130/chemistry/metabolism
Dimerization
Granulocyte Colony-Stimulating Factor/*chemistry/genetics/*metabolism
Humans
Ligands
Models, Molecular
Molecular Sequence Data
Protein Structure, Quaternary
Receptors, Granulocyte Colony-Stimulating Factor/*chemistry/genetics/*metabolism
Recombinant Proteins
Sequence Alignment
*Signal Transduction
Structural Homology, Protein
Abstract: A crystal structure of the signaling complex between human granulocyte colony-stimulating factor (GCSF) and a ligand binding region of GCSF receptor (GCSF-R), has been determined to 2.8 A resolution. The GCSF:GCSF-R complex formed a 2:2 stoichiometry by means of a cross-over interaction between the Ig-like domains of GCSF-R and GCSF. The conformation of the complex is quite different from that between human GCSF and the cytokine receptor homologous domain of mouse GCSF-R, but similar to that of the IL-6/gp130 signaling complex. The Ig-like domain cross-over structure necessary for GCSF-R activation is consistent with previously reported thermodynamic and mutational analyses.
Notes: 1091-6490
Tamada, Taro
Honjo, Eijiro
Maeda, Yoshitake
Okamoto, Tomoyuki
Ishibashi, Matsujiro
Tokunaga, Masao
Kuroki, Ryota
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2006 Feb 28;103(9):3135-40. doi: 10.1073/pnas.0511264103. Epub 2006 Feb 21.
Author Address: Research Group for Molecular Structural Biology, Quantum Beam Science Directorate, Japan Atomic Energy Agency, Tokai, Ibaraki.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1368
Author: Tamborero, S., Vilar, M., Martínez-Gil, L., Johnson, A. E. and Mingarro, I.
Year: 2011
Title: Membrane insertion and topology of the translocating chain-associating membrane protein (TRAM)
Journal: J Mol Biol
Volume: 406
Issue: 4
Pages: 571-82
Epub Date: 2011/01/18
Date: Mar 4
Short Title: Membrane insertion and topology of the translocating chain-associating membrane protein (TRAM)
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2011.01.009
Accession Number: 21237175
Keywords: Animals
Endoplasmic Reticulum/chemistry/metabolism
Intracellular Membranes/*chemistry/*metabolism
Membrane Glycoproteins/*chemistry/*metabolism
Mice
Models, Biological
Models, Molecular
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Abstract: The translocating chain-associating membrane protein (TRAM) is a glycoprotein involved in the translocation of secreted proteins into the endoplasmic reticulum (ER) lumen and in the insertion of integral membrane proteins into the lipid bilayer. As a major step toward elucidating the structure of the functional ER translocation/insertion machinery, we have characterized the membrane integration mechanism and the transmembrane topology of TRAM using two approaches: photocross-linking and truncated C-terminal reporter tag fusions. Our data indicate that TRAM is recognized by the signal recognition particle and translocon components, and suggest a membrane topology with eight transmembrane segments, including several poorly hydrophobic segments. Furthermore, we studied the membrane insertion capacity of these poorly hydrophobic segments into the ER membrane by themselves. Finally, we confirmed the main features of the proposed membrane topology in mammalian cells expressing full-length TRAM.
Notes: 1089-8638
Tamborero, Silvia
Vilar, Marçal
Martínez-Gil, Luis
Johnson, Arthur E
Mingarro, Ismael
R01 GM026494/GM/NIGMS NIH HHS/United States
GM26494/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2011 Mar 4;406(4):571-82. doi: 10.1016/j.jmb.2011.01.009. Epub 2011 Jan 13.
Author Address: Departament de Bioquímica i Biologia Molecular, Universitat de València, E-46 100 Burjassot, Spain.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1874
Author: Tan, E. P., McGreal, S. R., Graw, S., Tessman, R., Koppel, S. J., Dhakal, P., Zhang, Z., Machacek, M., Zachara, N. E., Koestler, D. C., Peterson, K. R., Thyfault, J. P., Swerdlow, R. H., Krishnamurthy, P., DiTacchio, L., Apte, U. and Slawson, C.
Year: 2017
Title: Sustained O-GlcNAcylation reprograms mitochondrial function to regulate energy metabolism
Journal: J Biol Chem
Volume: 292
Issue: 36
Pages: 14940-14962
Epub Date: 2017/07/26
Date: Sep 8
Short Title: Sustained O-GlcNAcylation reprograms mitochondrial function to regulate energy metabolism
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M117.797944
PMCID: PMC5592672
Accession Number: 28739801
Keywords: Acetylglucosamine/*metabolism
Animals
*Energy Metabolism
Glycosylation
Humans
Male
Mice
Mice, Inbred C57BL
Mice, Knockout
Mitochondria/*metabolism
N-Acetylglucosaminyltransferases/deficiency/genetics/*metabolism
Tumor Cells, Cultured
beta-N-Acetylhexosaminidases/genetics/*metabolism
*O-GlcNAcase
*O-GlcNAcylation
*Ogt
*bioenergetics
*mitochondria
*nuclear factor 2 (erythroid-derived 2-like factor) (NFE2L2) (Nrf2)
*reactive oxygen species (ROS)
this article
Abstract: Dysfunctional mitochondria and generation of reactive oxygen species (ROS) promote chronic diseases, which have spurred interest in the molecular mechanisms underlying these conditions. Previously, we have demonstrated that disruption of post-translational modification of proteins with β-linked N-acetylglucosamine (O-GlcNAcylation) via overexpression of the O-GlcNAc-regulating enzymes O-GlcNAc transferase (OGT) or O-GlcNAcase (OGA) impairs mitochondrial function. Here, we report that sustained alterations in O-GlcNAcylation either by pharmacological or genetic manipulation also alter metabolic function. Sustained O-GlcNAc elevation in SH-SY5Y neuroblastoma cells increased OGA expression and reduced cellular respiration and ROS generation. Cells with elevated O-GlcNAc levels had elongated mitochondria and increased mitochondrial membrane potential, and RNA-sequencing analysis indicated transcriptome reprogramming and down-regulation of the NRF2-mediated antioxidant response. Sustained O-GlcNAcylation in mouse brain and liver validated the metabolic phenotypes observed in the cells, and OGT knockdown in the liver elevated ROS levels, impaired respiration, and increased the NRF2 antioxidant response. Moreover, elevated O-GlcNAc levels promoted weight loss and lowered respiration in mice and skewed the mice toward carbohydrate-dependent metabolism as determined by indirect calorimetry. In summary, sustained elevation in O-GlcNAcylation coupled with increased OGA expression reprograms energy metabolism, a finding that has potential implications for the etiology, development, and management of metabolic diseases.
Notes: 1083-351x
Tan, Ee Phie
McGreal, Steven R
Graw, Stefan
Tessman, Robert
Koppel, Scott J
Dhakal, Pramod
Zhang, Zhen
Machacek, Miranda
Zachara, Natasha E
Koestler, Devin C
Peterson, Kenneth R
Thyfault, John P
Swerdlow, Russell H
Krishnamurthy, Partha
DiTacchio, Luciano
Apte, Udayan
Slawson, Chad
R01 DK100595/DK/NIDDK NIH HHS/United States
S10 RR027564/RR/NCRR NIH HHS/United States
P20 GM104936/GM/NIGMS NIH HHS/United States
R01 DK098414/DK/NIDDK NIH HHS/United States
P30 AG035982/AG/NIA NIH HHS/United States
I01 BX002567/BX/BLRD VA/United States
P30 GM103326/GM/NIGMS NIH HHS/United States
T32 HD057850/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2017 Sep 8;292(36):14940-14962. doi: 10.1074/jbc.M117.797944. Epub 2017 Jul 24.
Author Address: From the Departments of Biochemistry and Molecular Biology.
Pharmacology, Toxicology and Therapeutics.
Biostatistics.
Molecular and Integrative Physiology.
Pathology and Laboratory Medicine, and.
the Department of Biological Chemistry, The Johns Hopkins University of Medicine, Baltimore, Maryland 21205.
Neurology, University of Kansas Medical Center and.
University of Kansas Alzheimer's Disease Center, Kansas City, Kansas 64108 and.
From the Departments of Biochemistry and Molecular Biology, cslawson@kumc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1971
Author: Tan, H. Y., Eskandari, R., Shen, D., Zhu, Y., Liu, T. W., Willems, L. I., Alteen, M. G., Madden, Z. and Vocadlo, D. J.
Year: 2018
Title: Direct One-Step Fluorescent Labeling of O-GlcNAc-Modified Proteins in Live Cells Using Metabolic Intermediates
Journal: J Am Chem Soc
Volume: 140
Issue: 45
Pages: 15300-15308
Epub Date: 2018/10/09
Date: Nov 14
Short Title: Direct One-Step Fluorescent Labeling of O-GlcNAc-Modified Proteins in Live Cells Using Metabolic Intermediates
Alternate Journal: Journal of the American Chemical Society
ISSN: 0002-7863
DOI: 10.1021/jacs.8b08260
Accession Number: 30296064
Keywords: Acetylglucosamine/*chemistry/metabolism
*Fluorescence
Glycosylation
HeLa Cells
Humans
Molecular Structure
N-Acetylglucosaminyltransferases/*chemistry/metabolism
Abstract: The modification of proteins with O-linked N-acetylglucosamine ( O-GlcNAc) by the enzyme O-GlcNAc transferase (OGT) has emerged as an important regulator of cellular physiology. Metabolic labeling strategies to monitor O-GlcNAcylation in cells have proven of great value for uncovering the molecular roles of O-GlcNAc. These strategies rely on two-step labeling procedures, which limits the scope of experiments that can be performed. Here, we report on the creation of fluorescent uridine 5'-diphospho- N-acetylglucosamine (UDP-GlcNAc) analogues in which the N-acyl group of glucosamine is modified with a suitable linker and fluorophore. Using human OGT, we show these donor sugar substrates permit direct monitoring of OGT activity on protein substrates in vitro. We show that feeding cells with a corresponding fluorescent metabolic precursor for the last step of the hexosamine biosynthetic pathway (HBP) leads to its metabolic assimilation and labeling of O-GlcNAcylated proteins within live cells. This one-step metabolic feeding strategy permits labeling of O-GlcNAcylated proteins with a fluorescent glucosamine-nitrobenzoxadiazole (GlcN-NBD) conjugate that accumulates in a time- and dose-dependent manner. Because no genetic engineering of cells is required, we anticipate this strategy should be generally amenable to studying the roles of O-GlcNAc in cellular physiology as well as to gain an improved understanding of the regulation of OGT within cells. The further expansion of this one-step in-cell labeling strategy should enable performing a range of experiments including two-color pulse chase experiments and monitoring OGT activity on specific protein substrates in live cells.
Notes: 1520-5126
Tan, Hong Yee
Eskandari, Razieh
Shen, David
Zhu, Yanping
Liu, Ta-Wei
Willems, Lianne I
Alteen, Matthew G
Madden, Zarina
Vocadlo, David J
Orcid: 0000-0001-6897-5558
Journal Article
Research Support, Non-U.S. Gov't
United States
J Am Chem Soc. 2018 Nov 14;140(45):15300-15308. doi: 10.1021/jacs.8b08260. Epub 2018 Oct 22.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1229
Author: Tan, K., Duquette, M., Joachimiak, A. and Lawler, J.
Year: 2009
Title: The crystal structure of the signature domain of cartilage oligomeric matrix protein: implications for collagen, glycosaminoglycan and integrin binding
Journal: Faseb j
Volume: 23
Issue: 8
Pages: 2490-501
Epub Date: 2009/03/12
Date: Aug
Short Title: The crystal structure of the signature domain of cartilage oligomeric matrix protein: implications for collagen, glycosaminoglycan and integrin binding
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638 (Print)
0892-6638
DOI: 10.1096/fj.08-128090
PMCID: PMC2717772
Accession Number: 19276170
Keywords: Base Sequence
Binding Sites
Cartilage Oligomeric Matrix Protein
Collagen/metabolism
Crystallography, X-Ray
Cysteine/chemistry
DNA, Complementary/genetics
Extracellular Matrix Proteins/*chemistry/genetics/*metabolism
Glycoproteins/*chemistry/genetics/*metabolism
Glycosaminoglycans/metabolism
Humans
In Vitro Techniques
Integrins/metabolism
Ligands
Matrilin Proteins
Models, Molecular
Mutation
Oligopeptides/chemistry
Protein Binding
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Static Electricity
Thrombospondin 1/chemistry
Thrombospondins/chemistry
Abstract: Cartilage oligomeric matrix protein (COMP), or thrombospondin-5 (TSP-5), is a secreted glycoprotein that is important for growth plate organization and function. Mutations in COMP cause two skeletal dysplasias, pseudoachondroplasia (PSACH) and multiple epiphyseal dysplasia (EDM1). In this study, we determined the structure of a recombinant protein that contains the last epidermal growth factor repeat, the type 3 repeats and the C-terminal domain (CTD) of COMP to 3.15-A resolution limit by X-ray crystallography. The CTD is a beta-sandwich that is composed of 15 antiparallel beta-strands, and the type 3 repeats are a contiguous series of calcium binding sites that associate with the CTD at multiple points. The crystal packing reveals an exposed potential metal-ion-dependent adhesion site (MIDAS) on one edge of the beta-sandwich that is common to all TSPs and may serve as a binding site for collagens and other ligands. Disease-causing mutations in COMP disrupt calcium binding, disulfide bond formation, intramolecular interactions, or sites for potential ligand binding. The structure presented here and its unique molecular packing in the crystal identify potential interactive sites for glycosaminoglycans, integrins, and collagens, which are key to cartilage structure and function.
Notes: 1530-6860
Tan, Kemin
Duquette, Mark
Joachimiak, Andrzej
Lawler, Jack
R01 HL049081/HL/NHLBI NIH HHS/United States
HL49081/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
FASEB J. 2009 Aug;23(8):2490-501. doi: 10.1096/fj.08-128090. Epub 2009 Mar 10.
Author Address: Midwest Center for Structural Genomics, Biosciences Division, Argonne National Laboratory, Argonne, Illinois, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 855
Author: Tan, K., Duquette, M., Liu, J. H., Dong, Y., Zhang, R., Joachimiak, A., Lawler, J. and Wang, J. H.
Year: 2002
Title: Crystal structure of the TSP-1 type 1 repeats: a novel layered fold and its biological implication
Journal: J Cell Biol
Volume: 159
Issue: 2
Pages: 373-82
Epub Date: 2002/10/23
Date: Oct 28
Short Title: Crystal structure of the TSP-1 type 1 repeats: a novel layered fold and its biological implication
Alternate Journal: The Journal of cell biology
ISSN: 0021-9525 (Print)
0021-9525
DOI: 10.1083/jcb.200206062
PMCID: PMC2173040
Accession Number: 12391027
Keywords: Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Humans
Molecular Sequence Data
Protein Folding
Protein Structure, Tertiary
Structure-Activity Relationship
Thrombospondin 1/*chemistry
Abstract: Thrombospondin-1 (TSP-1) contains three type 1 repeats (TSRs), which mediate cell attachment, glycosaminoglycan binding, inhibition of angiogenesis, activation of TGFbeta, and inhibition of matrix metalloproteinases. The crystal structure of the TSRs reported in this article reveals a novel, antiparallel, three-stranded fold that consists of alternating stacked layers of tryptophan and arginine residues from respective strands, capped by disulfide bonds on each end. The front face of the TSR contains a right-handed spiral, positively charged groove that might be the "recognition" face, mediating interactions with various ligands. This is the first high-resolution crystal structure of a TSR domain that provides a prototypic architecture for structural and functional exploration of the diverse members of the TSR superfamily.
Notes: 1540-8140
Tan, Kemin
Duquette, Mark
Liu, Jin-huan
Dong, Yicheng
Zhang, Rongguang
Joachimiak, Andrzej
Lawler, Jack
Wang, Jia-huai
P01 CA092644/CA/NCI NIH HHS/United States
HL 68003/HL/NHLBI NIH HHS/United States
R01 HL049081/HL/NHLBI NIH HHS/United States
CA 92644/CA/NCI NIH HHS/United States
GM 56008/GM/NIGMS NIH HHS/United States
R01 HL068003/HL/NHLBI NIH HHS/United States
HL 49081/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
J Cell Biol. 2002 Oct 28;159(2):373-82. doi: 10.1083/jcb.200206062. Epub 2002 Oct 21.
Author Address: Dana-Farber Cancer Institute, Harvard Medical School, 44 Binney Street, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1145
Author: Tan, K., Duquette, M., Liu, J. H., Shanmugasundaram, K., Joachimiak, A., Gallagher, J. T., Rigby, A. C., Wang, J. H. and Lawler, J.
Year: 2008
Title: Heparin-induced cis- and trans-dimerization modes of the thrombospondin-1 N-terminal domain
Journal: J Biol Chem
Volume: 283
Issue: 7
Pages: 3932-41
Epub Date: 2007/12/11
Date: Feb 15
Short Title: Heparin-induced cis- and trans-dimerization modes of the thrombospondin-1 N-terminal domain
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M705203200
PMCID: PMC2918642
NIHMSID: NIHMS211188
Accession Number: 18065761
Keywords: Chromatography, Gel
Crystallization
Crystallography, X-Ray
Dimerization
Heparin/*chemistry
Humans
Models, Molecular
Protein Conformation
Recombinant Proteins/chemistry/metabolism
Thrombospondin 1/chemistry/*metabolism
Abstract: Through its interactions with proteins and proteoglycans, thrombospondin-1 (TSP-1) functions at the interface of the cell membrane and the extracellular matrix to regulate matrix structure and cellular phenotype. We have previously determined the structure of the high affinity heparin-binding domain of TSP-1, designated TSPN-1, in association with the synthetic heparin, Arixtra. To establish that the binding of TSPN-1 to Arixtra is representative of the association with naturally occurring heparins, we have determined the structures of TSPN-1 in complex with heparin oligosaccharides containing eight (dp8) and ten (dp10) subunits, by x-ray crystallography. We have found that dp8 and dp10 bind to TSPN-1 in a manner similar to Arixtra and that dp8 and dp10 induce the formation of trans and cis TSPN-1 dimers, respectively. In silico docking calculations partnered with our crystal structures support the importance of arginine residues in positions 29, 42, and 77 in binding sulfate groups of the dp8 and dp10 forms of heparin. The ability of several TSPN-1 domains to bind to glycosaminoglycans simultaneously probably increases the affinity of binding through multivalent interactions. The formation of cis and trans dimers of the TSPN-1 domain with relatively short segments of heparin further enhances the ability of TSP-1 to participate in high affinity binding to glycosaminoglycans. Dimer formation may also involve TSPN-1 domains from two separate TSP-1 molecules. This association would enable glycosaminoglycans to cluster TSP-1.
Notes: 1083-351x
Tan, Kemin
Duquette, Mark
Liu, Jin-Huan
Shanmugasundaram, Kumaran
Joachimiak, Andrzej
Gallagher, John T
Rigby, Alan C
Wang, Jia-huai
Lawler, Jack
HL48675/HL/NHLBI NIH HHS/United States
R01 HL049081/HL/NHLBI NIH HHS/United States
HL49081/HL/NHLBI NIH HHS/United States
P01 HL048675/HL/NHLBI NIH HHS/United States
R01 HL049081-10/HL/NHLBI NIH HHS/United States
R01 HL068003/HL/NHLBI NIH HHS/United States
HL68003/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2008 Feb 15;283(7):3932-41. doi: 10.1074/jbc.M705203200. Epub 2007 Dec 7.
Author Address: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1395
Author: Tan, K. and Lawler, J.
Year: 2011
Title: The structure of the Ca²+-binding, glycosylated F-spondin domain of F-spondin - A C2-domain variant in an extracellular matrix protein
Journal: BMC Struct Biol
Volume: 11
Pages: 22
Epub Date: 2011/05/17
Date: May 10
Short Title: The structure of the Ca²+-binding, glycosylated F-spondin domain of F-spondin - A C2-domain variant in an extracellular matrix protein
Alternate Journal: BMC structural biology
ISSN: 1472-6807
DOI: 10.1186/1472-6807-11-22
PMCID: PMC3117680
Accession Number: 21569239
Keywords: Amino Acid Sequence
Binding Sites
Calcium/*chemistry
Crystallography, X-Ray
Disulfides
Extracellular Matrix Proteins/*chemistry
Glycosylation
Humans
Molecular Sequence Data
Protein Binding
Protein Structure, Tertiary
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: BACKGROUND: F-spondin is a multi-domain extracellular matrix (ECM) protein and a contact-repellent molecule that directs axon outgrowth and cell migration during development. The reelin_N domain and the F-spondin domain (FS domain) comprise a proteolytic fragment that interacts with the cell membrane and guides the projection of commissural axons to floor plate. The FS domain is found in F-spondins, mindins, M-spondin and amphiF-spondin. RESULTS: We present the crystal structure of human F-spondin FS domain at 1.95Å resolution. The structure reveals a Ca2+-binding C2 domain variant with an 8-stranded antiparallel β-sandwich fold. Though the primary sequences of the FS domains of F-spondin and mindin are less than 36% identical, their overall structures are very similar. The unique feature of F-spondin FS domain is the presence of three disulfide bonds associated with the N- and C-termini of the domain and a highly conserved N-linked glycosylation site. The integrin-binding motif found in mindin is not conserved in the F-spondin FS domain. CONCLUSION: The structure of the F-spondin FS domain completes the structural studies of the multiple-domain ECM molecule. The homology of its core structure to a common Ca2+- and lipid-binding C2 domain suggests that the F-spondin FS domain may be responsible for part of the membrane targeting of F-spondin in its regulation of axon development. The structural properties of the FS domain revealed in this study pave the way for further exploration into the functions of F-spondin.
Notes: 1472-6807
Tan, Kemin
Lawler, Jack
HL49081/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
BMC Struct Biol. 2011 May 10;11:22. doi: 10.1186/1472-6807-11-22.
Author Address: Midwest Center for Structural Genomics and Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, IL 60439, USA. ktan@anl.gov
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 835
Author: Tan, K., Zelus, B. D., Meijers, R., Liu, J. H., Bergelson, J. M., Duke, N., Zhang, R., Joachimiak, A., Holmes, K. V. and Wang, J. H.
Year: 2002
Title: Crystal structure of murine sCEACAM1a[1,4]: a coronavirus receptor in the CEA family
Journal: Embo j
Volume: 21
Issue: 9
Pages: 2076-86
Epub Date: 2002/05/01
Date: May 1
Short Title: Crystal structure of murine sCEACAM1a[1,4]: a coronavirus receptor in the CEA family
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/21.9.2076
PMCID: PMC125375
Accession Number: 11980704
Keywords: Amino Acid Sequence
Animals
Antigens, CD
Carcinoembryonic Antigen/chemistry
Cell Adhesion Molecules
Conserved Sequence
Coronavirus/chemistry/physiology
Crystallography, X-Ray
Glycoproteins/*chemistry
Mice
Molecular Sequence Data
Multigene Family
Protein Binding/physiology
Protein Isoforms/chemistry
Protein Structure, Tertiary
Receptors, Coronavirus
Receptors, Virus/*chemistry
Sequence Alignment
Abstract: CEACAM1 is a member of the carcinoembryonic antigen (CEA) family. Isoforms of murine CEACAM1 serve as receptors for mouse hepatitis virus (MHV), a murine coronavirus. Here we report the crystal structure of soluble murine sCEACAM1a[1,4], which is composed of two Ig-like domains and has MHV neutralizing activity. Its N-terminal domain has a uniquely folded CC' loop that encompasses key virus-binding residues. This is the first atomic structure of any member of the CEA family, and provides a prototypic architecture for functional exploration of CEA family members. We discuss the structural basis of virus receptor activities of murine CEACAM1 proteins, binding of Neisseria to human CEACAM1, and other homophilic and heterophilic interactions of CEA family members.
Notes: 1460-2075
Tan, Kemin
Zelus, Bruce D
Meijers, Rob
Liu, Jin-huan
Bergelson, Jeffrey M
Duke, Norma
Zhang, Rongguang
Joachimiak, Andrzej
Holmes, Kathryn V
Wang, Jia-huai
R01 AI026075/AI/NIAID NIH HHS/United States
HL48675/HL/NHLBI NIH HHS/United States
AI 26075/AI/NIAID NIH HHS/United States
AI 25231/AI/NIAID NIH HHS/United States
GM56008/GM/NIGMS NIH HHS/United States
HL 54734/HL/NHLBI NIH HHS/United States
R01 AI025231/AI/NIAID NIH HHS/United States
P01 HL048675/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
EMBO J. 2002 May 1;21(9):2076-86. doi: 10.1093/emboj/21.9.2076.
Author Address: Dana-Farber Cancer Institute and Department of Medicine, Harvard Medical School, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1834
Author: Tan, S., Zhang, H., Chai, Y., Song, H., Tong, Z., Wang, Q., Qi, J., Wong, G., Zhu, X., Liu, W. J., Gao, S., Wang, Z., Shi, Y., Yang, F., Gao, G. F. and Yan, J.
Year: 2017
Title: An unexpected N-terminal loop in PD-1 dominates binding by nivolumab
Journal: Nat Commun
Volume: 8
Pages: 14369
Epub Date: 2017/02/07
Date: Feb 6
Short Title: An unexpected N-terminal loop in PD-1 dominates binding by nivolumab
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms14369
PMCID: PMC5303876
Accession Number: 28165004
Keywords: Antibodies, Monoclonal/*pharmacology
Antibodies, Monoclonal, Humanized/chemistry/pharmacology
Antineoplastic Agents, Immunological/*pharmacology
B7-H1 Antigen/*metabolism
Crystallography, X-Ray
Drug Design
Glycosylation
Humans
Immunotherapy/methods
Neoplasms/*drug therapy
Nivolumab
Programmed Cell Death 1 Receptor/*antagonists & inhibitors/chemistry/metabolism
Protein Binding
Protein Domains
Abstract: Cancer immunotherapy by targeting of immune checkpoint molecules has been a research 'hot-spot' in recent years. Nivolumab, a human monoclonal antibody targeting PD-1, has been widely used clinically since 2014. However, the binding mechanism of nivolumab to PD-1 has not yet been shown, despite a recent report describing the complex structure of pembrolizumab/PD-1. It has previously been speculated that PD-1 glycosylation is involved in nivolumab recognition. Here we report the complex structure of nivolumab with PD-1 and evaluate the effects of PD-1 N-glycosylation on the interactions with nivolumab. Structural and functional analyses unexpectedly reveal an N-terminal loop outside the IgV domain of PD-1. This loop is not involved in recognition of PD-L1 but dominates binding to nivolumab, whereas N-glycosylation is not involved in binding at all. Nivolumab binds to a completely different area than pembrolizumab. These results provide the basis for the design of future inhibitory molecules targeting PD-1.
Notes: 2041-1723
Tan, Shuguang
Zhang, Hao
Chai, Yan
Song, Hao
Tong, Zhou
Wang, Qihui
Qi, Jianxun
Wong, Gary
Zhu, Xiaodong
Liu, William J
Gao, Shan
Wang, Zhongfu
Shi, Yi
Yang, Fuquan
Gao, George F
Yan, Jinghua
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2017 Feb 6;8:14369. doi: 10.1038/ncomms14369.
Author Address: CAS Key Laboratory of Microbial Physiological and Metabolic engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
School of Life Sciences, Anhui University, Hefei 230601, China.
Research Network of Immunity and Health (RNIH), Beijing Institutes of Life Science, Chinese Academy of Sciences, Beijing 100101, China.
Beijing Combio Co., Ltd., Beijing 100111, China.
National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention (China CDC), Beijing 102206, China.
CAS Key Laboratory of Bio-medical Diagnostics, Suzhou Institute of Biomedical Engineering and Technology, Chinese Academy of Sciences, Suzhou 215163, China.
Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education and Provincial Key Laboratory of Biotechnology, College of Life Sciences, Northwest University, Xi'an 710069, China.
Laboratory of Protein and Peptide Pharmaceuticals &Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 716
Author: Tanaka, A., Iwase, H., Hiki, Y., Kokubo, T., Ishii-Karakasa, I., Toma, K., Kobayashi, Y. and Hotta, K.
Year: 1998
Title: Evidence for a site-specific fucosylation of N-linked oligosaccharide of immunoglobulin A1 from normal human serum
Journal: Glycoconj J
Volume: 15
Issue: 10
Pages: 995-1000
Epub Date: 1999/04/22
Date: Oct
Short Title: Evidence for a site-specific fucosylation of N-linked oligosaccharide of immunoglobulin A1 from normal human serum
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1023/a:1006989910120
Accession Number: 10211705
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Fucose/*metabolism
Humans
Immunoglobulin A/*blood/chemistry
Molecular Sequence Data
Oligosaccharides/chemistry/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Glycopeptides containing the N-linked oligosaccharide from human serum IgA1 were analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOFMS). Two glycopeptides, GP1 and GP2, prepared from the endoproteinase Asp-N digest of the IgA1 heavy chain, were derived from the CH2 domain (N-glycan site at Asn263) and the tailpiece portion (N-glycan site at Asn459), respectively. The structure of the attached sugar chain was deduced from the mass number of the glycopeptide and confirmed by a two-dimensional mapping technique for a pyridylaminated oligosaccharide. GP1 was composed of two major components having a fully galactosylated bianntena sugar chain with or without a bisecting N-acetylglucosamine (GlcNAc) residue. On the other hand, the GP2 fraction corresponded to the glycopeptides having a fully galactosylated and fucosylated bianntena sugar chain partly bearing a bisecting GlcNAc residue. Thus, the site-specific fucosylation of the N-linked oligosaccharide on the tailpiece of the alpha1 chain became evident for normal human serum IgA1.
Notes: Tanaka, A
Iwase, H
Hiki, Y
Kokubo, T
Ishii-Karakasa, I
Toma, K
Kobayashi, Y
Hotta, K
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 1998 Oct;15(10):995-1000. doi: 10.1023/a:1006989910120.
Author Address: Analytical Research Laboratory, Asahi Chemical Industry Co., Ltd., Fuji, Shizuoka, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 921
Author: Tanaka, K., Xu, W., Zhou, F. and You, G.
Year: 2004
Title: Role of glycosylation in the organic anion transporter OAT1
Journal: J Biol Chem
Volume: 279
Issue: 15
Pages: 14961-6
Epub Date: 2004/01/30
Date: Apr 9
Short Title: Role of glycosylation in the organic anion transporter OAT1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M400197200
Accession Number: 14749323
Keywords: Animals
Anions
Asparagine/chemistry
Binding Sites
Biological Transport
Biotinylation
Blotting, Western
COS Cells
Cell Line
Cell Membrane/chemistry/metabolism
Cloning, Molecular
Electrophoresis, Polyacrylamide Gel
Glutamine/chemistry
Glycosylation
HeLa Cells
Humans
Immunoblotting
Microscopy, Fluorescence
Microscopy, Phase-Contrast
Mutagenesis, Site-Directed
Mutation
Organic Anion Transport Protein 1/*chemistry
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/pharmacology
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Structure-Activity Relationship
Transfection
Abstract: Organic anion transporters (OAT) play essential roles in the body disposition of clinically important anionic drugs, including antiviral drugs, antitumor drugs, antibiotics, antihypertensives, and anti-inflammatories. We reported previously (Kuze, K., Graves, P., Leahy, A., Wilson, P., Stuhlmann, H., and You, G. (1999) J. Biol. Chem. 274, 1519-1524) that tunicamycin, an inhibitor of asparagine-linked glycosylation, significantly inhibited organic anion transport in COS-7 cells expressing a mouse organic anion transporter (mOAT1), suggesting an important role of glycosylation in mOAT1 function. In the present study, we investigated the effect of disrupting putative glycosylation sites in mOAT1 as well as its human counterpart, hOAT1, by mutating asparagine to glutamine and assessing mutant transporters in HeLa cells. We showed that the putative glycosylation site Asp-39 in mOAT1 was not glycosylated but the corresponding site (Asp-39) in hOAT1 was glycosylated. Disrupting Asp-39 resulted in a complete loss of transport activity in both mOAT1 and hOAT1 without affecting their cell surface expression, suggesting that the loss of function is not because of deglycosylation of Asp-39 per se but rather is likely because of the change of this important amino acid critically involved in the substrate binding. Single replacement of asparagines at other sites had no effect on transport activity indicating that glycosylation at individual sites is not essential for OAT function. In contrast, a simultaneous replacement of all asparagines in both mOAT1 and hOAT1 impaired the trafficking of the transporters to the plasma membrane. In summary, we provided the evidence that 1) Asp-39 is crucially involved in substrate recognition of OAT1, 2) glycosylation at individual sites is not required for OAT1 function, and 3) glycosylation plays an important role in the targeting of OAT1 onto the plasma membrane. This study is the first molecular identification and characterization of glycosylation of OAT1 and may provide important insights into the structure-function relationships of the organic anion transporter family.
Notes: Tanaka, Kunihiko
Xu, Wen
Zhou, Fanfan
You, Guofeng
R01-DK 60034/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2004 Apr 9;279(15):14961-6. doi: 10.1074/jbc.M400197200. Epub 2004 Jan 28.
Author Address: Department of Pharmaceutics, Rutgers, The State University of New Jersey, Piscataway, New Jersey 08854, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1555
Author: Tang, C. and Welty, D. F.
Year: 2013
Title: A dynamic view to the modulation of phosphorylation and O-GlcNAcylation by inhibition of O-GlcNAcase
Journal: Comput Biol Chem
Volume: 45
Pages: 9-16
Epub Date: 2013/04/24
Date: Aug
Short Title: A dynamic view to the modulation of phosphorylation and O-GlcNAcylation by inhibition of O-GlcNAcase
Alternate Journal: Computational biology and chemistry
ISSN: 1476-9271
DOI: 10.1016/j.compbiolchem.2013.03.001
Accession Number: 23608187
Keywords: Animals
Computational Biology
Glycosylation
Kinetics
Mice
Mice, Transgenic
N-Acetylglucosaminyltransferases/*antagonists & inhibitors/metabolism
Phosphorylation
*Thermodynamics
Abstract: Protein phosphorylation and O-GlcNAcylation are reciprocally regulated. As hyperphosphorylation is implicated in tau pathology, approaches have been exploited to reduce the magnitude of tau phosphorylation by increasing the level of tau O-GlcNAcylation. With mathematic models constructed to describe different kinetic scenarios, we analyzed the temporal change of an O-GlcNAcylated protein in contrast to that of the phosphorylated form upon inhibition of O-GlcNAcase (OGA). The analyses indicate that when degradation of the modified protein is negligible relative to the naked one, the magnitude of O-GlcNAcylated protein increase is proportional to the level of inhibition, while the extent of phosphorylated protein decline varies due to other factors. Furthermore, the increase of O-GlcNAcylated protein parallels with the decrease of phosphorylated form upon acute or short-term inhibition of OGA, as observed in many in vitro and short term in vivo studies. However, phosphorylated protein is predicted to return to its initial level while O-GlcNAcylated protein to achieve a higher steady level under sustained inhibition. This simulated result is in line with a recent report on long-term inhibition of OGA in transgenic mice. Noticeably, inhibition withdrawal is anticipated to cause a transient rise of phosphorylated protein. If degradation of modified proteins proceeds in addition to the naked one, the characteristic temporal profiles of each form in response to OGA inhibition would depend on the relative importance of individual degradation pathways. The models described herein may serve as a useful investigational tool that will provide insight into pharmacological intervention for tauopathies in particular and for reciprocally modulated reactions in general.
Notes: 1476-928x
Tang, Cuyue
Welty, Devin F
Journal Article
England
Comput Biol Chem. 2013 Aug;45:9-16. doi: 10.1016/j.compbiolchem.2013.03.001. Epub 2013 Mar 23.
Author Address: EnVivo Pharmaceuticals, 500 Arsenal Street, Watertown, MA 02472, USA. ctang@envivopharma.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1413
Author: Tanimoto, K., Suzuki, K., Jokitalo, E., Sakai, N., Sakaguchi, T., Tamura, D., Fujii, G., Aoki, K., Takada, S., Ishida, R., Tanabe, M., Itoh, H., Yoneda, Y., Sohda, M., Misumi, Y. and Nakamura, N.
Year: 2011
Title: Characterization of YIPF3 and YIPF4, cis-Golgi Localizing Yip domain family proteins
Journal: Cell Struct Funct
Volume: 36
Issue: 2
Pages: 171-85
Epub Date: 2011/07/16
Short Title: Characterization of YIPF3 and YIPF4, cis-Golgi Localizing Yip domain family proteins
Alternate Journal: Cell structure and function
ISSN: 0386-7196
DOI: 10.1247/csf.11002
Accession Number: 21757827
Keywords: Adaptor Proteins, Vesicular Transport
Amino Acid Sequence
Endoplasmic Reticulum/chemistry/metabolism
Glycosylation
Golgi Apparatus/chemistry/*metabolism
HeLa Cells
Humans
Membrane Proteins/*genetics/*metabolism
Molecular Sequence Data
Mutation
Protein Structure, Tertiary
RNA Interference
RNA, Small Interfering/metabolism
Recombinant Proteins/analysis/genetics/metabolism
Saccharomyces cerevisiae Proteins/chemistry/metabolism
Abstract: The Yip1 domain family (YIPF) proteins are homologues of yeast Yip1p and Yif1p, which are proposed to function in ER to Golgi transport. Here, we report the characterization of YIPF3 and YIPF4, homologues of human Yif1p and Yip1p, respectively. Immunofluorescence and immuno-electron microscopy showed that both YIPF3 and YIPF4 are clearly concentrated in the cis-Golgi. While YIPF4 was detected as a single mobility form consistent with its predicted molecular weight, three different mobility forms of YIPF3 were detected by western blotting. Biochemical and immunofluorescence experiments strongly indicated that YIPF3 is synthesized in the ER as a N-glycosylated form (40 kDa), is then O-glycosylated in the Golgi apparatus to become a lower mobility form (46 kDa) and finally becomes a higher mobility form cleaved at its C-terminal luminal domain (36 kDa). YIPF3 and YIPF4 form a complex in the Golgi apparatus, and this was suggested to be important for their proper localization and function. The knockdown of YIPF3 or YIPF4 in HeLa cells induced fragmentation of the Golgi apparatus, suggesting their involvement in the maintenance of the Golgi structure.
Notes: 1347-3700
Tanimoto, Kouji
Suzuki, Kurumi
Jokitalo, Eija
Sakai, Noriko
Sakaguchi, Tomoaki
Tamura, Daisuke
Fujii, Gourou
Aoki, Kenji
Takada, Saya
Ishida, Ryuichi
Tanabe, Masako
Itoh, Hideaki
Yoneda, Yukio
Sohda, Miwa
Misumi, Yoshio
Nakamura, Nobuhiro
Journal Article
Research Support, Non-U.S. Gov't
Japan
Cell Struct Funct. 2011;36(2):171-85. doi: 10.1247/csf.11002. Epub 2011 Jul 14.
Author Address: Graduate School of Natural Science and Technology and School of Pharmacy, Kanazawa University, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1547
Author: Tanji, H., Ohto, U., Shibata, T., Miyake, K. and Shimizu, T.
Year: 2013
Title: Structural reorganization of the Toll-like receptor 8 dimer induced by agonistic ligands
Journal: Science
Volume: 339
Issue: 6126
Pages: 1426-9
Epub Date: 2013/03/23
Date: Mar 22
Short Title: Structural reorganization of the Toll-like receptor 8 dimer induced by agonistic ligands
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1229159
Accession Number: 23520111
Keywords: Amino Acid Sequence
Crystallography, X-Ray
Humans
Hydrogen Bonding
Imidazoles/chemistry/*metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Mutant Proteins/chemistry/metabolism
Protein Binding
Protein Conformation
Protein Multimerization
Protein Structure, Secondary
Protein Structure, Tertiary
Quinolines/chemistry/*metabolism
Signal Transduction
Thiazoles/chemistry/*metabolism
Toll-Like Receptor 8/*agonists/*chemistry/metabolism
Abstract: Toll-like receptor 7 (TLR7) and TLR8 recognize single-stranded RNA and initiate innate immune responses. Several synthetic agonists of TLR7-TLR8 display novel therapeutic potential; however, the molecular basis for ligand recognition and activation of signaling by TLR7 or TLR8 is largely unknown. In this study, the crystal structures of unliganded and ligand-induced activated human TLR8 dimers were elucidated. Ligand recognition was mediated by a dimerization interface formed by two protomers. Upon ligand stimulation, the TLR8 dimer was reorganized such that the two C termini were brought into proximity. The loop between leucine-rich repeat 14 (LRR14) and LRR15 was cleaved; however, the N- and C-terminal halves remained associated and contributed to ligand recognition and dimerization. Thus, ligand binding induces reorganization of the TLR8 dimer, which enables downstream signaling processes.
Notes: 1095-9203
Tanji, Hiromi
Ohto, Umeharu
Shibata, Takuma
Miyake, Kensuke
Shimizu, Toshiyuki
Journal Article
Research Support, Non-U.S. Gov't
United States
Science. 2013 Mar 22;339(6126):1426-9. doi: 10.1126/science.1229159.
Author Address: Graduate School of Pharmaceutical Sciences, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1112
Author: Tansky, M. F., Pothoulakis, C. and Leeman, S. E.
Year: 2007
Title: Functional consequences of alteration of N-linked glycosylation sites on the neurokinin 1 receptor
Journal: Proc Natl Acad Sci U S A
Volume: 104
Issue: 25
Pages: 10691-6
Epub Date: 2007/06/15
Date: Jun 19
Short Title: Functional consequences of alteration of N-linked glycosylation sites on the neurokinin 1 receptor
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0703394104
PMCID: PMC1965574
Accession Number: 17563389
Keywords: Amino Acid Substitution
Cell Line
Enzyme Activation/genetics
Genetic Vectors
Glycine/*metabolism
Glycosylation
Humans
Immunohistochemistry
Interleukin-8/metabolism
JNK Mitogen-Activated Protein Kinases/metabolism
Kinetics
Lentivirus/genetics
Ligands
Mitogen-Activated Protein Kinase 1/metabolism
Mitogen-Activated Protein Kinase 3/metabolism
Neurokinin A/metabolism
Receptors, Neurokinin-1/genetics/*metabolism
Substance P/metabolism
p38 Mitogen-Activated Protein Kinases/metabolism
Abstract: The neurokinin 1 receptor (NK1R), a G protein-coupled receptor involved in diverse functions including pain and inflammation, has two putative N-linked glycosylation sites, Asn-14 and Asn-18. We studied the role of N-linked glycosylation in the functioning of the NK1R by constructing three receptor mutants: two single mutants (Asn --> Gln-14 and Asn --> Gln-18) and a double mutant, lacking both glycosylation sites. Using a lentiviral transfection system, the mutants were stably transfected into NCM 460 cells, a nontransformed human colonic epithelial cell line. We observed that the magnitude of glycosylation as estimated by changes in gel migration depends on the number of glycosylation sites available, with the wild-type receptor containing the greatest amount of glycosylation. All mutant receptors were able to bind to substance P and neurokinin A ligand with similar affinities; however, the double mutant, nonglycosylated NK1R showed only half the B(max) of the wild-type NK1R. In terms of receptor function, the ablation of both N-linked glycosylation sites did not have a profound effect on the receptors' abilities to activate the MAP kinase families (p42/p44, JNK, and p38), but did affect SP-induced IL-8 secretion. All mutants were able to internalize, but the kinetics of internalization of the double mutant receptor was more rapid, when compared with wild-type NK1R. Therefore, glycosylation of NK1R may stabilize the receptor in the plasma membrane. These results contribute to the ongoing elucidation of the role of glycosylation in G protein-coupled receptors and the study of the neurokinin receptors in particular.
Notes: 1091-6490
Tansky, Morris F
Pothoulakis, Charalabos
Leeman, Susan E
R01 DK047343/DK/NIDDK NIH HHS/United States
5R21NS04322-02/NS/NINDS NIH HHS/United States
R0-1 DK 47343/DK/NIDDK NIH HHS/United States
Comparative Study
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2007 Jun 19;104(25):10691-6. doi: 10.1073/pnas.0703394104. Epub 2007 Jun 11.
Author Address: Department of Pharmacology, Boston University School of Medicine, Boston, MA 02118, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1081
Author: Tao, G. Z., Kirby, C., Whelan, S. A., Rossi, F., Bi, X., MacLaren, M., Gentalen, E., O'Neill, R. A., Hart, G. W. and Omary, M. B.
Year: 2006
Title: Reciprocal keratin 18 Ser48 O-GlcNAcylation and Ser52 phosphorylation using peptide analysis
Journal: Biochem Biophys Res Commun
Volume: 351
Issue: 3
Pages: 708-12
Epub Date: 2006/11/07
Date: Dec 22
Short Title: Reciprocal keratin 18 Ser48 O-GlcNAcylation and Ser52 phosphorylation using peptide analysis
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/j.bbrc.2006.10.092
PMCID: PMC2692749
NIHMSID: NIHMS14089
Accession Number: 17084817
Keywords: Acylation
Amino Acid Substitution
Glycine/*chemistry
Keratin-18/*chemistry
Peptides/chemistry
Phosphorylation
Serine/*chemistry
Structure-Activity Relationship
Abstract: Phosphorylation and O-GlcNAcylation of keratin 18 (K18) are highly dynamic and involve primarily independent K18 populations. We used in vitro phosphorylation and O-GlcNAcylation of wild-type, phospho-Ser52, glyco-Ser48, and Ser-to-Ala mutant 17mer peptides (K18 amino acids 40-56), which include the major K18 glycosylation (Ser48) and phosphorylation (Ser52) sites, to address whether each modification blocks the other. The glyco-K18 peptide blocks Ser52 phosphorylation by protein kinase C, an in vivo K18 kinase, while the phospho-K18 peptide blocks its O-GlcNAcylation. Our findings support the reciprocity of these two post-translational modifications. Therefore, regulation of protein Ser/Thr phosphorylation and glycosylation at proximal sites can be interdependent and provides a potential mechanism of counter regulation.
Notes: 1090-2104
Tao, Guo-Zhong
Kirby, Celeste
Whelan, Stephen A
Rossi, Frank
Bi, Xiahui
MacLaren, Michael
Gentalen, Erik
O'Neill, Roger A
Hart, Gerald W
Omary, M Bishr
DK61671/DK/NIDDK NIH HHS/United States
R56 DK052951/DK/NIDDK NIH HHS/United States
DK52951/DK/NIDDK NIH HHS/United States
R37 HD013563/HD/NICHD NIH HHS/United States
HD13563/HD/NICHD NIH HHS/United States
R01 DK052951/DK/NIDDK NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
R01 DK052951-10/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Biochem Biophys Res Commun. 2006 Dec 22;351(3):708-12. doi: 10.1016/j.bbrc.2006.10.092. Epub 2006 Oct 27.
Author Address: Palo Alto VA Medical Center, Stanford University, Palo Alto, CA 94304, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1769
Author: Tao, T., He, Z., Shao, Z. and Lu, H.
Year: 2016
Title: TAB3 O-GlcNAcylation promotes metastasis of triple negative breast cancer
Journal: Oncotarget
Volume: 7
Issue: 16
Pages: 22807-18
Epub Date: 2016/03/25
Date: Apr 19
Short Title: TAB3 O-GlcNAcylation promotes metastasis of triple negative breast cancer
Alternate Journal: Oncotarget
ISSN: 1949-2553
DOI: 10.18632/oncotarget.8182
PMCID: PMC5008402
Accession Number: 27009840
Keywords: Acylation
Adaptor Proteins, Signal Transducing/*metabolism
Animals
Female
Heterografts
Humans
Kaplan-Meier Estimate
Mice
Mice, Nude
N-Acetylglucosaminyltransferases/metabolism
Neoplasm Invasiveness/pathology
Protein Processing, Post-Translational
Triple Negative Breast Neoplasms/metabolism/mortality/*pathology
O-glcNAcylation
Tab3
metastasis
triple negative breast cancer
Abstract: O-GlcNAcylation is a post-translational modification that regulates a broad range of nuclear and cytoplasmic proteins and is emerging as a key regulator of various biological processes. Although previous studies have shown that increased levels of global O-GlcNAcylation and O-GlcNActransferase are linked to the incidence of metastasis in triple negative breast cancer (TNBC) patients, the molecular basis behind this is not fully understood. In this study, we have determined that the TAK1 binding protein 3 (TAB3) was O-GlcNAcylated at Ser408 by OGT in the TNBC, which was required for its Thr404 phosphorylation, TAK1 activation and downstream nuclear factor kappa B (NF-κB) activation in TNBC. O-GlcNAcylation of TAB3 was induced by p38 MAPK and it in turn enhances the TAK1 mediated p38MAPK activation, which forms the positive feedback loop in TAB3mediated NF-κB activation. In TNBC, TAB3O-GlcNAcylationmediated cell migration and invasion by activating its downstream NF-κB. The expression of TAB3 O-GlcNAcylation increased in TNBC patients, and it was significantly correlated with poor prognoses of the patients. Our study provides insights into the mechanism of TAB3 regulating activity and suggests its important implications in TNBC metastasis.
Notes: 1949-2553
Tao, Tao
He, Zhixian
Shao, Zhiming
Lu, Haojie
R01 AA020203/AA/NIAAA NIH HHS/United States
Journal Article
Oncotarget. 2016 Apr 19;7(16):22807-18. doi: 10.18632/oncotarget.8182.
Author Address: Shanghai Cancer Center and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, P.R. China.
Department of Chemistry, Fudan University, Shanghai 200433, P.R. China.
Department of General Surgery, Affiliated Hospital of Nantong University, Nantong 226001, P.R. China.
Key Laboratory of Glycoconjugates Research Ministry of Public Health, Fudan University, Shanghai 200032, P.R. China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1963
Author: Taparra, K., Wang, H., Malek, R., Lafargue, A., Barbhuiya, M. A., Wang, X., Simons, B. W., Ballew, M., Nugent, K., Groves, J., Williams, R. D., Shiraishi, T., Verdone, J., Yildirir, G., Henry, R., Zhang, B., Wong, J., Wang, K. K., Nelkin, B. D., Pienta, K. J., Felsher, D., Zachara, N. E. and Tran, P. T.
Year: 2018
Title: O-GlcNAcylation is required for mutant KRAS-induced lung tumorigenesis
Journal: J Clin Invest
Volume: 128
Issue: 11
Pages: 4924-4937
Epub Date: 2018/08/22
Date: Nov 1
Short Title: O-GlcNAcylation is required for mutant KRAS-induced lung tumorigenesis
Alternate Journal: The Journal of clinical investigation
ISSN: 0021-9738 (Print)
0021-9738
DOI: 10.1172/jci94844
PMCID: PMC6205381
Accession Number: 30130254
Keywords: A549 Cells
Acylation
Amino Acid Substitution
Animals
Cell Transformation, Neoplastic/genetics/*metabolism/pathology
*Epithelial-Mesenchymal Transition
Female
Glucose/genetics/metabolism
HEK293 Cells
Hexosamines/genetics/metabolism
Humans
Lung Neoplasms/*enzymology/genetics/pathology
Mice
Mice, Nude
Mice, Transgenic
*Mutation, Missense
*Protein Processing, Post-Translational
Proto-Oncogene Proteins p21(ras)/genetics/*metabolism
*Cancer
*Cellular senescence
*Glycobiology
*Oncology
Abstract: Mutant KRAS drives glycolytic flux in lung cancer, potentially impacting aberrant protein glycosylation. Recent evidence suggests aberrant KRAS drives flux of glucose into the hexosamine biosynthetic pathway (HBP). HBP is required for various glycosylation processes, such as protein N- or O-glycosylation and glycolipid synthesis. However, its function during tumorigenesis is poorly understood. One contributor and proposed target of KRAS-driven cancers is a developmentally conserved epithelial plasticity program called epithelial-mesenchymal transition (EMT). Here we showed in novel autochthonous mouse models that EMT accelerated KrasG12D lung tumorigenesis by upregulating expression of key enzymes of the HBP pathway. We demonstrated that HBP was required for suppressing KrasG12D-induced senescence, and targeting HBP significantly delayed KrasG12D lung tumorigenesis. To explore the mechanism, we investigated protein glycosylation downstream of HBP and found elevated levels of O-linked β-N-acetylglucosamine (O-GlcNAcylation) posttranslational modification on intracellular proteins. O-GlcNAcylation suppressed KrasG12D oncogene-induced senescence (OIS) and accelerated lung tumorigenesis. Conversely, loss of O-GlcNAcylation delayed lung tumorigenesis. O-GlcNAcylation of proteins SNAI1 and c-MYC correlated with the EMT-HBP axis and accelerated lung tumorigenesis. Our results demonstrated that O-GlcNAcylation was sufficient and required to accelerate KrasG12D lung tumorigenesis in vivo, which was reinforced by epithelial plasticity programs.
Notes: 1558-8238
Taparra, Kekoa
Wang, Hailun
Malek, Reem
Lafargue, Audrey
Barbhuiya, Mustafa A
Wang, Xing
Simons, Brian W
Ballew, Matthew
Nugent, Katriana
Groves, Jennifer
Williams, Russell D
Shiraishi, Takumi
Verdone, James
Yildirir, Gokben
Henry, Roger
Zhang, Bin
Wong, John
Wang, Ken Kang-Hsin
Nelkin, Barry D
Pienta, Kenneth J
Felsher, Dean
Zachara, Natasha E
Tran, Phuoc T
R21 DK108782/DK/NIDDK NIH HHS/United States
R01 CA166348/CA/NCI NIH HHS/United States
F31 CA189588/CA/NCI NIH HHS/United States
U01 CA212007/CA/NCI NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Clin Invest. 2018 Nov 1;128(11):4924-4937. doi: 10.1172/JCI94844. Epub 2018 Sep 24.
Author Address: Department of Radiation Oncology and Molecular Radiation Sciences, Sidney Kimmel Comprehensive Cancer Center.
Program in Cellular and Molecular Medicine.
Department of Urology, James Buchanan Brady Urological Institute.
Department of Biological Chemistry, and.
Department of Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Division of Medical Oncology, Departments of Medicine and Pathology, Stanford University School of Medicine, Stanford, California, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1921
Author: Tarbet, H. J., Dolat, L., Smith, T. J., Condon, B. M., O'Brien, E. T., 3rd, Valdivia, R. H. and Boyce, M.
Year: 2018
Title: Site-specific glycosylation regulates the form and function of the intermediate filament cytoskeleton
Journal: Elife
Volume: 7
Epub Date: 2018/03/08
Date: Mar 7
Short Title: Site-specific glycosylation regulates the form and function of the intermediate filament cytoskeleton
Alternate Journal: eLife
ISSN: 2050-084x
DOI: 10.7554/eLife.31807
PMCID: PMC5841932
Accession Number: 29513221
Keywords: Acetylglucosamine/*genetics/metabolism
Animals
Cell Movement/*genetics
Cytoskeleton/*genetics
Glycosylation
Humans
Intermediate Filaments/*genetics
N-Acetylglucosaminyltransferases/genetics
Phosphorylation
Protein Processing, Post-Translational/genetics
Signal Transduction
Vimentin/genetics
*Chlamydia trachomatis
*O-GlcNAc
*cell biology
*cell migration
*chemical biology
*glycosylation
*human
*intermediate filaments
Abstract: Intermediate filaments (IF) are a major component of the metazoan cytoskeleton and are essential for normal cell morphology, motility, and signal transduction. Dysregulation of IFs causes a wide range of human diseases, including skin disorders, cardiomyopathies, lipodystrophy, and neuropathy. Despite this pathophysiological significance, how cells regulate IF structure, dynamics, and function remains poorly understood. Here, we show that site-specific modification of the prototypical IF protein vimentin with O-linked β-N-acetylglucosamine (O-GlcNAc) mediates its homotypic protein-protein interactions and is required in human cells for IF morphology and cell migration. In addition, we show that the intracellular pathogen Chlamydia trachomatis, which remodels the host IF cytoskeleton during infection, requires specific vimentin glycosylation sites and O-GlcNAc transferase activity to maintain its replicative niche. Our results provide new insight into the biochemical and cell biological functions of vimentin O-GlcNAcylation, and may have broad implications for our understanding of the regulation of IF proteins in general.
Notes: 2050-084x
Tarbet, Heather J
Dolat, Lee
Smith, Timothy J
Condon, Brett M
O'Brien, E Timothy 3rd
Valdivia, Raphael H
Boyce, Michael
Orcid: 0000-0002-2729-4876
R56 AI107951/AI/NIAID NIH HHS/United States
T32 GM007184/GM/NIGMS NIH HHS/United States
R01 AI107951/AI/NIAID NIH HHS/United States
R01 AI123083/AI/NIAID NIH HHS/United States
P41 EB002025/EB/NIBIB NIH HHS/United States
1R01AI107951/AI/NIAID NIH HHS/United States
1R01AI123083/AI/NIAID NIH HHS/United States
P41-EB002025/EB/NIBIB NIH HHS/United States
R01 GM118847/GM/NIGMS NIH HHS/United States
S10 OD018164/OD/NIH HHS/United States
1R01GM118847-01/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Elife. 2018 Mar 7;7:e31807. doi: 10.7554/eLife.31807.
Author Address: Department of Biochemistry, Duke University School of Medicine, Durham, United States.
Department of Molecular Genetics and Microbiology, Duke University School of Medicine, Durham, United States.
Center for Host-Microbial Interactions, Duke University School of Medicine, Durham, United States.
Department of Physics and Astronomy, University of North Carolina, Chapel Hill, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1856
Author: Taylor, N. M. I., Manolaridis, I., Jackson, S. M., Kowal, J., Stahlberg, H. and Locher, K. P.
Year: 2017
Title: Structure of the human multidrug transporter ABCG2
Journal: Nature
Volume: 546
Issue: 7659
Pages: 504-509
Epub Date: 2017/05/30
Date: Jun 22
Short Title: Structure of the human multidrug transporter ABCG2
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature22345
Accession Number: 28554189
Keywords: ATP Binding Cassette Transporter, Subfamily G, Member 2/antagonists &
inhibitors/*chemistry/metabolism/*ultrastructure
Adenosine Triphosphatases/chemistry/genetics/metabolism/ultrastructure
Amino Acid Sequence
Antibodies/chemistry/immunology/ultrastructure
Binding Sites
Biological Transport
Cholesterol/chemistry/metabolism
*Cryoelectron Microscopy
Humans
Immunoglobulin Fab Fragments/chemistry/immunology/ultrastructure
Models, Molecular
Neoplasm Proteins/antagonists & inhibitors/*chemistry/metabolism/*ultrastructure
Polymorphism, Single Nucleotide/genetics
Protein Domains
Abstract: ABCG2 is a constitutively expressed ATP-binding cassette (ABC) transporter that protects many tissues against xenobiotic molecules. Its activity affects the pharmacokinetics of commonly used drugs and limits the delivery of therapeutics into tumour cells, thus contributing to multidrug resistance. Here we present the structure of human ABCG2 determined by cryo-electron microscopy, providing the first high-resolution insight into a human multidrug transporter. We visualize ABCG2 in complex with two antigen-binding fragments of the human-specific, inhibitory antibody 5D3 that recognizes extracellular loops of the transporter. We observe two cholesterol molecules bound in the multidrug-binding pocket that is located in a central, hydrophobic, inward-facing translocation pathway between the transmembrane domains. Combined with functional in vitro analyses, our results suggest a multidrug recognition and transport mechanism of ABCG2, rationalize disease-causing single nucleotide polymorphisms and the allosteric inhibition by the 5D3 antibody, and provide the structural basis of cholesterol recognition by other G-subfamily ABC transporters.
Notes: 1476-4687
Taylor, Nicholas M I
Manolaridis, Ioannis
Jackson, Scott M
Kowal, Julia
Stahlberg, Henning
Locher, Kaspar P
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2017 Jun 22;546(7659):504-509. doi: 10.1038/nature22345. Epub 2017 May 29.
Author Address: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Mattenstrasse 26, 4058 Basel, Switzerland.
Institute of Molecular Biology and Biophysics, ETH Zürich, Otto-Stern-Weg 5, 8093 Zürich, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1151
Author: Taylor, R. P., Parker, G. J., Hazel, M. W., Soesanto, Y., Fuller, W., Yazzie, M. J. and McClain, D. A.
Year: 2008
Title: Glucose deprivation stimulates O-GlcNAc modification of proteins through up-regulation of O-linked N-acetylglucosaminyltransferase
Journal: J Biol Chem
Volume: 283
Issue: 10
Pages: 6050-7
Epub Date: 2008/01/05
Date: Mar 7
Short Title: Glucose deprivation stimulates O-GlcNAc modification of proteins through up-regulation of O-linked N-acetylglucosaminyltransferase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M707328200
Accession Number: 18174169
Keywords: Acetylglucosamine/*metabolism
Cell Line, Tumor
Glucosamine/metabolism
Glucose/deficiency/*metabolism
Glycogen Synthase/metabolism
Glycosylation
Humans
N-Acetylglucosaminyltransferases/*metabolism
Protein Processing, Post-Translational/*physiology
Time Factors
Abstract: O-Linked N-acetylglucosamine (O-GlcNAc) is a post-translational modification of proteins that functions as a nutrient sensing mechanism. Here we report on regulation of O-GlcNAcylation over a broad range of glucose concentrations. We have discovered a significant induction of O-GlcNAc modification of a limited number of proteins under conditions of glucose deprivation. Beginning 12 h after treatment, glucose-deprived human hepatocellular carcinoma (HepG2) cells demonstrate a 7.8-fold increase in total O-GlcNAc modification compared with cells cultured in normal glucose (5 mm; p = 0.008). Some of the targets of glucose deprivation-induced O-GlcNAcylation are distinct from those modified in response to high glucose (20 mm) or glucosamine (10 mm) treatment, suggesting differential targeting with glucose deprivation and glucose excess. O-GlcNAcylation of glycogen synthase is significantly increased with glucose deprivation, and this O-GlcNAc increase contributes to a 60% decrease (p = 0.004) in glycogen synthase activity. Increased O-GlcNAc modification is not mediated by increased UDP-GlcNAc, the rate-limiting substrate for O-GlcNAcylation. Rather, the mRNA for nucleocytoplasmic O-linked N-acetylglucosaminyltransferase (OGT) increases 3.4-fold within 6 h of glucose deprivation (p = 0.006). Within 12 h, OGT protein increases 1.7-fold (p = 0.01) compared with normal glucose-treated cells. In addition, 12-h glucose deprivation leads to a 49% decrease in O-GlcNAcase protein levels (p = 0.03). We conclude that increased O-GlcNAc modification stimulated by glucose deprivation results from increased OGT and decreased O-GlcNAcase levels and that these changes affect cell metabolism, thus inactivating glycogen synthase.
Notes: Taylor, Rodrick P
Parker, Glendon J
Hazel, Mark W
Soesanto, Yudi
Fuller, William
Yazzie, Marla J
McClain, Donald A
R01 DK 43526/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
J Biol Chem. 2008 Mar 7;283(10):6050-7. doi: 10.1074/jbc.M707328200. Epub 2008 Jan 3.
Author Address: Departments of Biochemistry and Medicine, University of Utah School of Medicine, 30 N. 2030 East, Salt Lake City, UT 84132, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1178
Author: Teillet, F., Gaboriaud, C., Lacroix, M., Martin, L., Arlaud, G. J. and Thielens, N. M.
Year: 2008
Title: Crystal structure of the CUB1-EGF-CUB2 domain of human MASP-1/3 and identification of its interaction sites with mannan-binding lectin and ficolins
Journal: J Biol Chem
Volume: 283
Issue: 37
Pages: 25715-25724
Epub Date: 2008/07/04
Date: Sep 12
Short Title: Crystal structure of the CUB1-EGF-CUB2 domain of human MASP-1/3 and identification of its interaction sites with mannan-binding lectin and ficolins
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M803551200
Accession Number: 18596036
Keywords: Amino Acid Sequence
Complement C1r/*chemistry
Crystallography, X-Ray/methods
Epidermal Growth Factor/*chemistry
Humans
Kinetics
Lectins/*chemistry
Mannose-Binding Lectin/*chemistry
Mannose-Binding Protein-Associated Serine Proteases/*chemistry
Molecular Conformation
Molecular Sequence Data
Protein Binding
Protein Structure, Tertiary
Sequence Homology, Amino Acid
Surface Plasmon Resonance
Abstract: MASP-1 and MASP-3 are homologous proteases arising from alternative splicing of the MASP1/3 gene. They include an identical CUB(1)-EGF-CUB(2)-CCP(1)-CCP(2) module array prolonged by different serine protease domains at the C-terminal end. The x-ray structure of the CUB(1)-EGF-CUB(2) domain of human MASP-1/3, responsible for interaction of MASP-1 and -3 with their partner proteins mannan-binding lectin (MBL) and ficolins, was solved to a resolution of 2.3A(.) The structure shows a head-to-tail homodimer mainly stabilized by hydrophobic interactions between the CUB(1) module of one monomer and the epidermal growth factor (EGF) module of its counterpart. A Ca(2+) ion bound primarily to both EGF modules stabilizes the intra- and inter-monomer CUB(1)-EGF interfaces. Additional Ca(2+) ions are bound to each CUB(1) and CUB(2) module through six ligands contributed by Glu(49), Asp(57), Asp(102), and Ser(104) (CUB(1)) and their counterparts Glu(216), Asp(226), Asp(263), and Ser(265) (CUB(2)), plus one and two water molecules, respectively. To identify the residues involved in interaction of MASP-1 and -3 with MBL and L- and H-ficolins, 27 point mutants of human MASP-3 were generated, and their binding properties were analyzed using surface plasmon resonance spectroscopy. These mutations map two homologous binding sites contributed by modules CUB(1) and CUB(2), located in close vicinity of their Ca(2+)-binding sites and stabilized by the Ca(2+) ion. This information allows us to propose a model of the MBL-MASP-1/3 interaction, involving a major electrostatic interaction between two acidic Ca(2+) ligands of MASP-1/3 and a conserved lysine of MBL. Based on these and other data, a schematic model of a MBL.MASP complex is proposed.
Notes: Teillet, Florence
Gaboriaud, Christine
Lacroix, Monique
Martin, Lydie
Arlaud, Gérard J
Thielens, Nicole M
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2008 Sep 12;283(37):25715-25724. doi: 10.1074/jbc.M803551200. Epub 2008 Jul 2.
Author Address: Laboratoire d'Enzymologie Moléculaire, 38027 Grenoble Cedex 1, France.
Laboratoire de Cristallographie et Cristallogenèse des Protéines, Institut de Biologie Structurale Jean-Pierre Ebel, CNRS-CEA-UJF, UMR 5075, 41 Rue Jules Horowitz, 38027 Grenoble Cedex 1, France.
Laboratoire d'Enzymologie Moléculaire, 38027 Grenoble Cedex 1, France. Electronic address: nicole.thielens@ibs.fr.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 936
Author: Tempel, W., Karaveg, K., Liu, Z. J., Rose, J., Wang, B. C. and Moremen, K. W.
Year: 2004
Title: Structure of mouse Golgi alpha-mannosidase IA reveals the molecular basis for substrate specificity among class 1 (family 47 glycosylhydrolase) alpha1,2-mannosidases
Journal: J Biol Chem
Volume: 279
Issue: 28
Pages: 29774-86
Epub Date: 2004/04/23
Date: Jul 9
Short Title: Structure of mouse Golgi alpha-mannosidase IA reveals the molecular basis for substrate specificity among class 1 (family 47 glycosylhydrolase) alpha1,2-mannosidases
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M403065200
Accession Number: 15102839
Keywords: Amino Acid Sequence
Animals
Binding Sites
Carbohydrate Conformation
Carbohydrate Sequence
Catalytic Domain
Crystallography, X-Ray
Endoplasmic Reticulum/metabolism
Golgi Apparatus/*enzymology
Humans
Mice
Molecular Sequence Data
Molecular Structure
Oligosaccharides/chemistry/*metabolism
*Protein Structure, Tertiary
Sequence Alignment
Substrate Specificity
alpha-Mannosidase/*chemistry/genetics/metabolism
Abstract: Three subfamilies of mammalian Class 1 processing alpha1,2-mannosidases (family 47 glycosidases) play critical roles in the maturation of Asn-linked glycoproteins in the endoplasmic reticulum (ER) and Golgi complex as well as influencing the timing and recognition for disposal of terminally unfolded proteins by ER-associated degradation. In an effort to define the structural basis for substrate recognition among Class 1 mannosidases, we have crystallized murine Golgi mannosidase IA (space group P2(1)2(1)2(1)), and the structure was solved to 1.5-A resolution by molecular replacement. The enzyme assumes an (alphaalpha)(7) barrel structure with a Ca(2+) ion coordinated at the base of the barrel similar to other Class 1 mannosidases. Critical residues within the barrel structure that coordinate the Ca(2+) ion or presumably bind and catalyze the hydrolysis of the glycone are also highly conserved. A Man(6)GlcNAc(2) oligosaccharide attached to Asn(515) in the murine enzyme was found to extend into the active site of an adjoining protein unit in the crystal lattice in a presumed enzyme-product complex. In contrast to an analogous complex previously isolated for Saccharomyces cerevisiae ER mannosidase I, the oligosaccharide in the active site of the murine Golgi enzyme assumes a different conformation to present an alternate oligosaccharide branch into the active site pocket. A comparison of the observed protein-carbohydrate interactions for the murine Golgi enzyme with the binding cleft topologies of the other family 47 glycosidases provides a framework for understanding the structural basis for substrate recognition among this class of enzymes.
Notes: Tempel, Wolfram
Karaveg, Khanita
Liu, Zhi-Jie
Rose, John
Wang, Bi-Cheng
Moremen, Kelley W
GM47533/GM/NIGMS NIH HHS/United States
RR05351/RR/NCRR NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2004 Jul 9;279(28):29774-86. doi: 10.1074/jbc.M403065200. Epub 2004 Apr 21.
Author Address: Department of Biochemistry and Molecular Biology, University of Georgia, Athens, Georgia 30602, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1300
Author: Teo, C. F., Ingale, S., Wolfert, M. A., Elsayed, G. A., Nöt, L. G., Chatham, J. C., Wells, L. and Boons, G. J.
Year: 2010
Title: Glycopeptide-specific monoclonal antibodies suggest new roles for O-GlcNAc
Journal: Nat Chem Biol
Volume: 6
Issue: 5
Pages: 338-43
Epub Date: 2010/03/23
Date: May
Short Title: Glycopeptide-specific monoclonal antibodies suggest new roles for O-GlcNAc
Alternate Journal: Nature chemical biology
ISSN: 1552-4450 (Print)
1552-4450
DOI: 10.1038/nchembio.338
PMCID: PMC2857662
NIHMSID: NIHMS179240
Accession Number: 20305658
Keywords: Acetylglucosamine/*chemistry/immunology
Antibodies, Monoclonal/*immunology
Enzyme-Linked Immunosorbent Assay
Glycopeptides/*immunology
Immunoprecipitation
Abstract: Studies of post-translational modification by beta-N-acetyl-D-glucosamine (O-GlcNAc) are hampered by a lack of efficient tools such as O-GlcNAc-specific antibodies that can be used for detection, isolation and site localization. We have obtained a large panel of O-GlcNAc-specific IgG monoclonal antibodies having a broad spectrum of binding partners by combining three-component immunogen methodology with hybridoma technology. Immunoprecipitation followed by large-scale shotgun proteomics led to the identification of more than 200 mammalian O-GlcNAc-modified proteins, including a large number of new glycoproteins. A substantial number of the glycoproteins were enriched by only one of the antibodies. This observation, combined with the results of inhibition ELISAs, suggests that the antibodies, in addition to their O-GlcNAc dependence, also appear to have different but overlapping local peptide determinants. The monoclonal antibodies made it possible to delineate differentially modified proteins of liver in response to trauma-hemorrhage and resuscitation in a rat model.
Notes: 1552-4469
Teo, Chin Fen
Ingale, Sampat
Wolfert, Margreet A
Elsayed, Galal A
Nöt, Laszlo G
Chatham, John C
Wells, Lance
Boons, Geert-Jan
R01 HL067464/HL/NHLBI NIH HHS/United States
R01 HL079364-04/HL/NHLBI NIH HHS/United States
P41 GM103390/GM/NIGMS NIH HHS/United States
R01 HL079364/HL/NHLBI NIH HHS/United States
R01CA088986/CA/NCI NIH HHS/United States
R01 CA088986-08/CA/NCI NIH HHS/United States
P41 RR005351-20/RR/NCRR NIH HHS/United States
HL067464/HL/NHLBI NIH HHS/United States
P41 RR005351/RR/NCRR NIH HHS/United States
R01 DK075069-04/DK/NIDDK NIH HHS/United States
R01 HL067464-08/HL/NHLBI NIH HHS/United States
R01 CA088986/CA/NCI NIH HHS/United States
R01 DK075069/DK/NIDDK NIH HHS/United States
P41R005351/PHS HHS/United States
HL079364/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Chem Biol. 2010 May;6(5):338-43. doi: 10.1038/nchembio.338. Epub 2010 Mar 21.
Author Address: Department of Biochemistry and Molecular Biology, University of Georgia, Athens, Georgia, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1651
Author: Teo, C. F. and Wells, L.
Year: 2014
Title: Monitoring protein O-linked β-N-acetylglucosamine status via metabolic labeling and copper-free click chemistry
Journal: Anal Biochem
Volume: 464
Pages: 70-2
Epub Date: 2014/07/06
Date: Nov 1
Short Title: Monitoring protein O-linked β-N-acetylglucosamine status via metabolic labeling and copper-free click chemistry
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/j.ab.2014.06.010
PMCID: PMC4172539
NIHMSID: NIHMS611310
Accession Number: 24995865
Keywords: Acetylglucosamine/*metabolism
*Click Chemistry
Copper/chemistry
Azido sugars
Cyclooctyne
O-GlcNAc stoichiometry
PEGylation
Spaac
Abstract: O-Linked β-N-acetylglucosamine (O-GlcNAc) modification found on the serine and threonine residues of intracellular proteins is an inducible post-translational modification that regulates numerous biological processes. In combination with other cell biological and biochemical approaches, a robust and streamlined strategy for detecting the number and stoichiometry of O-GlcNAc modification can provide valuable insights for decoding the functions of O-GlcNAc at the molecular level. Here, we report an optimized workflow for evaluating the O-GlcNAc status of proteins using a combination of metabolic labeling and click chemistry-based mass tagging. This method is strategically complementary to the chemoenzymatic-based mass-tagging method.
Notes: 1096-0309
Teo, Chin Fen
Wells, Lance
P01 GM107012/GM/NIGMS NIH HHS/United States
P41 GM103490/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Anal Biochem. 2014 Nov 1;464:70-2. doi: 10.1016/j.ab.2014.06.010. Epub 2014 Jul 1.
Author Address: Complex Carbohydrate Research Center, University of Georgia, Athens, GA 30602, USA.
Complex Carbohydrate Research Center, University of Georgia, Athens, GA 30602, USA. Electronic address: lwellsuf@uga.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1919
Author: Teplyakov, A., Obmolova, G., Luo, J. and Gilliland, G. L.
Year: 2018
Title: Crystal structure of B-cell co-receptor CD19 in complex with antibody B43 reveals an unexpected fold
Journal: Proteins
Volume: 86
Issue: 5
Pages: 495-500
Epub Date: 2018/03/01
Date: May
Short Title: Crystal structure of B-cell co-receptor CD19 in complex with antibody B43 reveals an unexpected fold
Alternate Journal: Proteins
ISSN: 0887-3585
DOI: 10.1002/prot.25485
Accession Number: 29490423
Keywords: Amino Acid Sequence
Animals
Antibodies, Monoclonal/*chemistry
Antigens, CD19/*chemistry
B-Lymphocytes/cytology
Binding Sites
Crystallography, X-Ray
HEK293 Cells
Humans
Mice
Models, Molecular
Protein Binding
Protein Folding
Protein Structure, Secondary
*blinatumomab
*epitope
*immunoglobulin fold
*specificity
*topology
Abstract: CD19 is a transmembrane protein expressed on malignant B cells, but not in other lineages or other tissues, which makes it an attractive target for monoclonal antibody-mediated immunotherapy. Anti-CD19 antibody B43 was utilized in a bispecific T-cell engager (BiTE) blinatumomab that demonstrated potency for the treatment of relapsed acute lymphoblastic leukemia. To gain insight into the mechanism of action of the antibody, the crystal structure of B43 Fab was determined in complex with CD19 and in the unbound form. The structure revealed the binding epitope, explained the lack of cross-reactivity toward non-human species, and suggested the key-and-lock mechanism of antigen recognition. Most unexpectedly, the structure revealed a unique molecular topology of CD19. Rather than a tandem of c-type immunoglobulin folds predicted from the amino acid sequence, the extracellular domain of CD19 exhibits an elongated β-sandwich formed by two immunoglobulin folds by swapping their C-terminal halves. This is the first structure of CD19, which has no sequence homologs.
Notes: 1097-0134
Teplyakov, Alexey
Orcid: 0000-0003-0296-0016
Obmolova, Galina
Luo, Jinquan
Gilliland, Gary L
Journal Article
United States
Proteins. 2018 May;86(5):495-500. doi: 10.1002/prot.25485. Epub 2018 Mar 10.
Author Address: Janssen Research and Development, LLC, 1400 McKean Road, Spring House, Pennsylvania, 19477.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 271
Author: Terrana, B., Tecce, M. F., Manetti, R., Ceccarini, C., Lamba, D. and Segre, A. L.
Year: 1990
Title: Unique structure of glycopeptide from alpha-fetoprotein produced in human hepatoma cell line, as determined by 1H-nuclear magnetic resonance spectroscopy
Journal: Clin Chem
Volume: 36
Issue: 6
Pages: 879-82
Epub Date: 1990/06/01
Date: Jun
Short Title: Unique structure of glycopeptide from alpha-fetoprotein produced in human hepatoma cell line, as determined by 1H-nuclear magnetic resonance spectroscopy
Alternate Journal: Clinical chemistry
ISSN: 0009-9147 (Print)
0009-9147
Accession Number: 1694115
Keywords: Carcinoma, Hepatocellular/*analysis
Cell Line
Glycopeptides/*analysis
Humans
Liver Neoplasms/*analysis
Magnetic Resonance Spectroscopy
Protons
alpha-Fetoproteins/*analysis
Abstract: Determination of alpha-fetoprotein is used in diagnosis of tumors and neural tube defects. A good reliable source of alpha-fetoprotein would be an obvious advantage to the preparation of diagnostic reagents and their standardization. We have recently developed a method for the production of alpha-fetoprotein from a human hepatoma cell line. This method, which is suitable for scaling up, allowed us to produce 40 g of alpha-fetoprotein from culture supernatant liquid through a simple purification procedure. We have previously shown this protein to be identical to alpha-fetoprotein produced from other sources. However, because the presence of different glycoforms has been reported in alpha-fetoprotein preparations, both from human sources and from other species, it was important to establish the type and extent of glycosylation of alpha-fetoprotein prepared by our method. By using 1H-NMR spectroscopy we were able to establish that our product contains a single N-linked biantennary, fully sialylated complex-type oligosaccharide, typical of human hepatomas.
Notes: Terrana, B
Tecce, M F
Manetti, R
Ceccarini, C
Lamba, D
Segre, A L
Journal Article
England
Clin Chem. 1990 Jun;36(6):879-82.
Author Address: Centro Ricerche Sclavo, Siena, Italy.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1703
Author: Terzyan, S. S., Burgett, A. W., Heroux, A., Smith, C. A., Mooers, B. H. and Hanigan, M. H.
Year: 2015
Title: Human γ-Glutamyl Transpeptidase 1: STRUCTURES OF THE FREE ENZYME, INHIBITOR-BOUND TETRAHEDRAL TRANSITION STATES, AND GLUTAMATE-BOUND ENZYME REVEAL NOVEL MOVEMENT WITHIN THE ACTIVE SITE DURING CATALYSIS
Journal: J Biol Chem
Volume: 290
Issue: 28
Pages: 17576-86
Epub Date: 2015/05/28
Date: Jul 10
Short Title: Human γ-Glutamyl Transpeptidase 1: STRUCTURES OF THE FREE ENZYME, INHIBITOR-BOUND TETRAHEDRAL TRANSITION STATES, AND GLUTAMATE-BOUND ENZYME REVEAL NOVEL MOVEMENT WITHIN THE ACTIVE SITE DURING CATALYSIS
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M115.659680
PMCID: PMC4498091
Accession Number: 26013825
Keywords: Aminobutyrates/chemistry/pharmacology
Apoenzymes/chemistry
Catalysis
Catalytic Domain
Crystallography, X-Ray
Enzyme Inhibitors/chemistry/pharmacology
Glutamic Acid/metabolism
Humans
Models, Molecular
Organophosphonates/chemistry/pharmacology
Protein Conformation
Recombinant Proteins/chemistry/genetics/metabolism
gamma-Glutamyltransferase/antagonists & inhibitors/*chemistry/genetics
N-terminal hydrolase
crystal structure
cysteine
enzyme inactivation
gamma-glutamyl transferase
gamma-glutamyl transpeptidase
glutathione
human
oxyanion hole
Abstract: γ-Glutamyl transpeptidase 1 (GGT1) is a cell surface, N-terminal nucleophile hydrolase that cleaves glutathione and other γ-glutamyl compounds. GGT1 expression is essential in cysteine homeostasis, and its induction has been implicated in the pathology of asthma, reperfusion injury, and cancer. In this study, we report four new crystal structures of human GGT1 (hGGT1) that show conformational changes within the active site as the enzyme progresses from the free enzyme to inhibitor-bound tetrahedral transition states and finally to the glutamate-bound structure prior to the release of this final product of the reaction. The structure of the apoenzyme shows flexibility within the active site. The serine-borate-bound hGGT1 crystal structure demonstrates that serine-borate occupies the active site of the enzyme, resulting in an enzyme-inhibitor complex that replicates the enzyme's tetrahedral intermediate/transition state. The structure of GGsTop-bound hGGT1 reveals its interactions with the enzyme and why neutral phosphonate diesters are more potent inhibitors than monoanionic phosphonates. These structures are the first structures for any eukaryotic GGT that include a molecule in the active site covalently bound to the catalytic Thr-381. The glutamate-bound structure shows the conformation of the enzyme prior to release of the final product and reveals novel information regarding the displacement of the main chain atoms that form the oxyanion hole and movement of the lid loop region when the active site is occupied. These data provide new insights into the mechanism of hGGT1-catalyzed reactions and will be invaluable in the development of new classes of hGGT1 inhibitors for therapeutic use.
Notes: 1083-351x
Terzyan, Simon S
Burgett, Anthony W G
Heroux, Annie
Smith, Clyde A
Mooers, Blaine H M
Hanigan, Marie H
P41 RR012408/RR/NCRR NIH HHS/United States
P41-GM111244/GM/NIGMS NIH HHS/United States
R01 AI088011/AI/NIAID NIH HHS/United States
P41RR012408/RR/NCRR NIH HHS/United States
P41 GM103473/GM/NIGMS NIH HHS/United States
P41GM103393/GM/NIGMS NIH HHS/United States
P41 GM111244/GM/NIGMS NIH HHS/United States
R01AI088011/AI/NIAID NIH HHS/United States
P20 GM103640/GM/NIGMS NIH HHS/United States
P20GM103640/GM/NIGMS NIH HHS/United States
P41 GM103393/GM/NIGMS NIH HHS/United States
P41GM103473/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2015 Jul 10;290(28):17576-86. doi: 10.1074/jbc.M115.659680. Epub 2015 May 26.
Author Address: From the Macromolecular Crystallography Laboratory, Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
the Department of Chemistry and Biochemistry, University of Oklahoma, Norman, Oklahoma 73019.
the Energy Sciences Directorate/Photon Science Division, Brookhaven National Laboratory, Upton, New York 11973.
the Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Menlo Park, California 94025.
the Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, and.
the Department of Cell Biology, University of Oklahoma Health Sciences Center, Stanton L. Young Biomedical Research Center, Oklahoma City, Oklahoma 73104 marie-hanigan@ouhsc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1238
Author: Thaysen-Andersen, M., Mysling, S. and Højrup, P.
Year: 2009
Title: Site-specific glycoprofiling of N-linked glycopeptides using MALDI-TOF MS: strong correlation between signal strength and glycoform quantities
Journal: Anal Chem
Volume: 81
Issue: 10
Pages: 3933-43
Epub Date: 2009/04/11
Date: May 15
Short Title: Site-specific glycoprofiling of N-linked glycopeptides using MALDI-TOF MS: strong correlation between signal strength and glycoform quantities
Alternate Journal: Analytical chemistry
ISSN: 0003-2700
DOI: 10.1021/ac900231w
Accession Number: 19358553
Keywords: Chromatography, High Pressure Liquid/methods
Fluorescent Dyes/chemistry
Glycopeptides/*analysis/chemistry
Polysaccharides/analysis/chemistry
Sialic Acids/chemistry
Sialoglycoproteins/analysis/chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/*methods
Abstract: Site-specific glycoprofiling of N-linked glycopeptides using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is an emerging technique, but its quantitative accuracy lacks documentation. Thus, a systematic study of widely different glycopeptides was performed to determine the relationship between the relative abundances of the individual glycoforms and the MALDI-TOF MS signal strength. Glycopeptides derived from glycoproteins containing neutral glycans (ribonuclease B, IgG, and ovalbumin) were initially profiled and yielded excellent and reproducible quantitation (correlation coefficient r = 0.9958, n = 5) when evaluated against a normal phase HPLC 2-AB glycan profile. Similarly, precise quantitation was observed for various forms of N-glycans (free, permethylated, and fluorescence-labeled) using MS. In addition, three different sialoglycopeptides from fetuin were site-specifically profiled, and good correlation between peak intensities and relative abundances was found with only a minor loss of sialic acids (r = 0.9664, n = 5). For glycopeptide purification, a range of hydrophilic and graphite materials packed in microcolumn format proved capable of performing desalting without loss of quantitative information, but highlighted the column capacity as a critical parameter. In conclusion, MALDI-TOF MS signal strength of glycopeptides has been found to accurately reflect the relative quantities of glycoforms, providing that certain technical issues are considered, i.e., nonbiased sample handling, matrix choice, and instrumental settings. This enables rapid and sensitive site-specific glycoprofiling of N-glycan populations to promote biomarker discovery and elucidation of glycan structure/function relationships.
Notes: 1520-6882
Thaysen-Andersen, Morten
Mysling, Simon
Højrup, Peter
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Chem. 2009 May 15;81(10):3933-43. doi: 10.1021/ac900231w.
Author Address: Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, Odense M, DK-5230, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 794
Author: Thielens, N. M., Cseh, S., Thiel, S., Vorup-Jensen, T., Rossi, V., Jensenius, J. C. and Arlaud, G. J.
Year: 2001
Title: Interaction properties of human mannan-binding lectin (MBL)-associated serine proteases-1 and -2, MBL-associated protein 19, and MBL
Journal: J Immunol
Volume: 166
Issue: 8
Pages: 5068-77
Epub Date: 2001/04/06
Date: Apr 15
Short Title: Interaction properties of human mannan-binding lectin (MBL)-associated serine proteases-1 and -2, MBL-associated protein 19, and MBL
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
DOI: 10.4049/jimmunol.166.8.5068
Accession Number: 11290788
Keywords: Amino Acid Motifs/genetics
Animals
Calcium/metabolism
Carrier Proteins/*metabolism
Cations, Divalent/metabolism
Collectins
Complement C1s/genetics
Dimerization
Epidermal Growth Factor/genetics
Extracellular Matrix Proteins/genetics
Humans
Lectins/*metabolism
Mannans/metabolism
Mannose-Binding Protein-Associated Serine Proteases
Peptide Fragments/biosynthesis/genetics/metabolism
Receptors, Complement 3b/genetics
Recombinant Proteins/biosynthesis/chemistry/metabolism
Serine Endopeptidases/genetics/*metabolism
Spodoptera/genetics
Surface Plasmon Resonance
Abstract: The mannan-binding lectin (MBL) activation pathway of complement plays an important role in the innate immune defense against pathogenic microorganisms. In human serum, two MBL-associated serine proteases (MASP-1, MASP-2) and MBL-associated protein 19 (MAp19) were found to be associated with MBL. With a view to investigate the interaction properties of these proteins, human MASP-1, MASP-2, MAp19, as well as the N-terminal complement subcomponents C1r/C1s, Uegf, and bone morphogenetic protein-1-epidermal growth factor (CUB-EGF) segments of MASP-1 and MASP-2, were expressed in insect or human kidney cells, and MBL was isolated from human serum. Sedimentation velocity analysis indicated that the MASP-1 and MASP-2 CUB-EGF segments and the homologous protein MAp19 all behaved as homodimers (2.8-3.2 S) in the presence of Ca(2+). Although the latter two dimers were not dissociated by EDTA, their physical properties were affected. In contrast, the MASP-1 CUB-EGF homodimer was not sensitive to EDTA. The three proteins and full-length MASP-1 and MASP-2 showed no interaction with each other as judged by gel filtration and surface plasmon resonance spectroscopy. Using the latter technique, MASP-1, MASP-2, their CUB-EGF segments, and MAp19 were each shown to bind to immobilized MBL, with K:(D) values of 0.8 nM (MASP-2), 1.4 nM (MASP-1), 13.0 nM (MAp19 and MASP-2 CUB-EGF), and 25.7 nM (MASP-1 CUB-EGF). The binding was Ca(2+)-dependent and fully sensitive to EDTA in all cases. These data indicate that MASP-1, MASP-2, and MAp19 each associate as homodimers, and individually form Ca(2+)-dependent complexes with MBL through the CUB-EGF pair of each protein. This suggests that distinct MBL/MASP complexes may be involved in the activation or regulation of the MBL pathway.
Notes: Thielens, N M
Cseh, S
Thiel, S
Vorup-Jensen, T
Rossi, V
Jensenius, J C
Arlaud, G J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 2001 Apr 15;166(8):5068-77. doi: 10.4049/jimmunol.166.8.5068.
Author Address: Laboratoire d'Enzymologie Moléculaire, Institut de Biologie Structurale Jean-Pierre Ebel (Commissariat à l'Energie Atomique-Centre National de la Recherche Scientifique), Grenoble, France. thielens@ibs.fr
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 320
Author: Thielens, N. M., Van Dorsselaer, A., Gagnon, J. and Arlaud, G. J.
Year: 1990
Title: Chemical and functional characterization of a fragment of C1-s containing the epidermal growth factor homology region
Journal: Biochemistry
Volume: 29
Issue: 14
Pages: 3570-8
Epub Date: 1990/04/10
Date: Apr 10
Short Title: Chemical and functional characterization of a fragment of C1-s containing the epidermal growth factor homology region
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00466a021
Accession Number: 2141278
Keywords: Amino Acid Sequence
Calcium/metabolism
Complement C1 Inactivator Proteins
Complement C1r/metabolism
Complement C1s/*genetics/metabolism
Epidermal Growth Factor/*genetics
Fibrinolysin
Humans
Mass Spectrometry
Molecular Sequence Data
Pancreatic Elastase
Peptide Fragments/isolation & purification
Peptide Mapping
Sequence Homology, Nucleic Acid
Abstract: C1-s, one of the three subcomponents of C1-, the first component of complement, is a serine protease comprising two disulfide-linked chains, the B chain, containing the catalytic site, and the A chain, involved in Ca2+ binding and Ca2(+)-dependent interaction(s) with the other C1- subcomponents. In an attempt to identify the regions responsible for the latter functions, C1-s was submitted to limited proteolysis with plasmin, a treatment that split the A chain into three major fragments, alpha 1, alpha 2, and gamma. Fragment alpha 2, which comprised the epidermal growth factor-like (EGF-like) region of C1-s, was heterogeneous, starting at serine 97 or phenylalanine 105 and ending at lysine 195. This fragment was reduced and alkylated and then digested with elastase, and three peptides covering positions 131-135, 131-139, and 131-140 were characterized by amino acid analysis, Edman degradation, and mass spectrometry, showing that position 134 of C1-s is occupied partly by an asparagine (47%) and partly by an erythro-beta-hydroxyasparagine, in contrast with the homologous position (150) of C1-r which only contains erythro-beta-hydroxyasparagine. As measured by equilibrium dialysis, native alpha 2, like the other plasmin-cleavage fragments, did not retain the ability of intact C1-s to bind Ca2+. In the same way, plasmin cleavage abolished the ability of C1-s to dimerize or to associate with C1-r in the presence of Ca2+. In contrast, both alpha 2 and the N-terminal alpha 1 fragment, starting at serine 24 of the A chain, were able to compete significantly with intact C1s for the formation of the Ca2(+)-dependent C1-s-C1r-C1-r-C1-s tetramer.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Thielens, N M
Van Dorsselaer, A
Gagnon, J
Arlaud, G J
Comparative Study
Journal Article
United States
Biochemistry. 1990 Apr 10;29(14):3570-8. doi: 10.1021/bi00466a021.
Author Address: Département de Recherches Fondamentales, Centre d'Etudes Nucléaires de Grenoble, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 384
Author: Thim, L., Bjoern, S., Christensen, M., Nicolaisen, E. M., Lund-Hansen, T., Pedersen, A. H. and Hedner, U.
Year: 1988
Title: Amino acid sequence and posttranslational modifications of human factor VIIa from plasma and transfected baby hamster kidney cells
Journal: Biochemistry
Volume: 27
Issue: 20
Pages: 7785-93
Epub Date: 1988/10/04
Date: Oct 4
Short Title: Amino acid sequence and posttranslational modifications of human factor VIIa from plasma and transfected baby hamster kidney cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00420a030
Accession Number: 3264725
Keywords: Amino Acid Sequence
Animals
Cells, Cultured
Cricetinae
Factor VII/genetics/*metabolism
Factor VIIa
Glycosylation
Humans
Hydroxylation
Molecular Sequence Data
*Protein Processing, Post-Translational
Transfection
Abstract: Blood coagulation factor VII is a vitamin K dependent glycoprotein which in its activated form, factor VIIa, participates in the coagulation process by activating factor X and/or factor IX in the presence of Ca2+ and tissue factor. Three types of potential posttranslational modifications exist in the human factor VIIa molecule, namely, 10 gamma-carboxylated, N-terminally located glutamic acid residues, 1 beta-hydroxylated aspartic acid residue, and 2 N-glycosylated asparagine residues. In the present study, the amino acid sequence and posttranslational modifications of recombinant factor VIIa as purified from the culture medium of a transfected baby hamster kidney cell line have been compared to human plasma factor VIIa. By use of HPLC, amino acid analysis, peptide mapping, and automated Edman degradations, the protein backbone of recombinant factor VIIa was found to be identical with human factor VIIa. Neither recombinant factor VIIa nor human plasma factor VIIa was found to contain beta-hydroxyaspartic acid. In human plasma factor VIIa, the 10 N-terminally located glutamic acid residues were found to be fully gamma-carboxylated whereas 9 full and 1 partial gamma-carboxylated residues were found in the corresponding positions of the recombinant factor VIIa molecule. Asparagine residues 145 and 322 were found to be fully N-glycosylated in human plasma factor VIIa. In the recombinant factor VIIa, asparagine residue 322 was fully glycosylated whereas asparagine residue 145 was only partially (approximately 66%) glycosylated. Besides minor differences in the sialic acid and fucose contents, the overall carbohydrate compositions were nearly identical in recombinant factor VIIa and human plasma factor VIIa.(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Thim, L
Bjoern, S
Christensen, M
Nicolaisen, E M
Lund-Hansen, T
Pedersen, A H
Hedner, U
Journal Article
United States
Biochemistry. 1988 Oct 4;27(20):7785-93. doi: 10.1021/bi00420a030.
Author Address: Novo Research Institute, Bagsvaerd, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 901
Author: Thoma, R., Löffler, B., Stihle, M., Huber, W., Ruf, A. and Hennig, M.
Year: 2003
Title: Structural basis of proline-specific exopeptidase activity as observed in human dipeptidyl peptidase-IV
Journal: Structure
Volume: 11
Issue: 8
Pages: 947-59
Epub Date: 2003/08/09
Date: Aug
Short Title: Structural basis of proline-specific exopeptidase activity as observed in human dipeptidyl peptidase-IV
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/s0969-2126(03)00160-6
Accession Number: 12906826
Keywords: Adenosine Deaminase/metabolism
Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Dimerization
Dipeptidyl Peptidase 4/*chemistry/isolation & purification/metabolism
Enzyme Stability
Exopeptidases/*metabolism
Glycosylation
Humans
Ligands
Models, Molecular
Molecular Sequence Data
Oligopeptides/antagonists & inhibitors
Pichia/enzymology
*Proline
Protein Structure, Secondary
Protein Structure, Tertiary
Substrate Specificity
Water/chemistry
Abstract: Inhibition of dipeptidyl peptidase IV (DPP-IV), the main glucagon-like peptide 1 (GLP1)-degrading enzyme, has been proposed for the treatment of type II diabetes. We expressed and purified the ectodomain of human DPP-IV in Pichia pastoris and determined the X-ray structure at 2.1 A resolution. The enzyme consists of two domains, the catalytic domain, with an alpha/beta hydrolase fold, and a beta propeller domain with an 8-fold repeat of a four-strand beta sheet motif. The beta propeller domain contributes two important functions to the molecule that have not been reported for such structures, an extra beta sheet motif that forms part of the dimerization interface and an additional short helix with a double Glu sequence motif. The Glu motif provides recognition and a binding site for the N terminus of the substrates, as revealed by the complex structure with diprotin A, a substrate with low turnover that is trapped in the tetrahedral intermediate of the reaction in the crystal.
Notes: Thoma, Ralf
Löffler, Bernd
Stihle, Martine
Huber, Walter
Ruf, Armin
Hennig, Michael
Journal Article
United States
Structure. 2003 Aug;11(8):947-59. doi: 10.1016/s0969-2126(03)00160-6.
Author Address: F. Hoffmann-La Roche AG, Pharma Research Discovery, 4070 Basel, Switzerland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1420
Author: Thomas, C., Moraga, I., Levin, D., Krutzik, P. O., Podoplelova, Y., Trejo, A., Lee, C., Yarden, G., Vleck, S. E., Glenn, J. S., Nolan, G. P., Piehler, J., Schreiber, G. and Garcia, K. C.
Year: 2011
Title: Structural linkage between ligand discrimination and receptor activation by type I interferons
Journal: Cell
Volume: 146
Issue: 4
Pages: 621-32
Epub Date: 2011/08/23
Date: Aug 19
Short Title: Structural linkage between ligand discrimination and receptor activation by type I interferons
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2011.06.048
PMCID: PMC3166218
NIHMSID: NIHMS320698
Accession Number: 21854986
Keywords: Amino Acid Sequence
Cell Line, Tumor
Crystallography, X-Ray
Humans
Interferon Type I/*chemistry/metabolism
Interferon-alpha/*chemistry/metabolism
Ligands
Models, Molecular
Molecular Sequence Data
Receptors, Interferon/*metabolism
Sequence Alignment
Abstract: Type I Interferons (IFNs) are important cytokines for innate immunity against viruses and cancer. Sixteen human type I IFN variants signal through the same cell-surface receptors, IFNAR1 and IFNAR2, yet they can evoke markedly different physiological effects. The crystal structures of two human type I IFN ternary signaling complexes containing IFNα2 and IFNω reveal recognition modes and heterotrimeric architectures that are unique among the cytokine receptor superfamily but conserved between different type I IFNs. Receptor-ligand cross-reactivity is enabled by conserved receptor-ligand "anchor points" interspersed among ligand-specific interactions that "tune" the relative IFN-binding affinities, in an apparent extracellular "ligand proofreading" mechanism that modulates biological activity. Functional differences between IFNs are linked to their respective receptor recognition chemistries, in concert with a ligand-induced conformational change in IFNAR1, that collectively control signal initiation and complex stability, ultimately regulating differential STAT phosphorylation profiles, receptor internalization rates, and downstream gene expression patterns.
Notes: 1097-4172
Thomas, Christoph
Moraga, Ignacio
Levin, Doron
Krutzik, Peter O
Podoplelova, Yulia
Trejo, Angelica
Lee, Choongho
Yarden, Ganit
Vleck, Susan E
Glenn, Jeffrey S
Nolan, Garry P
Piehler, Jacob
Schreiber, Gideon
Garcia, K Christopher
R37 AI051321/AI/NIAID NIH HHS/United States
R01 AI087917/AI/NIAID NIH HHS/United States
R01 AI051321-10/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
R01-AI087917/AI/NIAID NIH HHS/United States
R01 AI051321/AI/NIAID NIH HHS/United States
R01-AI51321/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell. 2011 Aug 19;146(4):621-32. doi: 10.1016/j.cell.2011.06.048.
Author Address: Howard Hughes Medical Institute, Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 494
Author: Thomas, K. A., Baglan, N. C. and Bradshaw, R. A.
Year: 1981
Title: The amino acid sequence of the gamma-subunit of mouse submaxillary gland 7 S nerve growth factor
Journal: J Biol Chem
Volume: 256
Issue: 17
Pages: 9156-66
Epub Date: 1981/09/10
Date: Sep 10
Short Title: The amino acid sequence of the gamma-subunit of mouse submaxillary gland 7 S nerve growth factor
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7263706
Keywords: Amino Acid Sequence
Animals
Carboxypeptidase B
Carboxypeptidases
Cyanogen Bromide
Disulfides/analysis
Macromolecular Substances
Mice
*Nerve Growth Factors
Peptide Fragments
Protein Conformation
Submandibular Gland/*analysis
Abstract: The complete amino acid sequence of the gamma-subunit of mouse submaxillary gland 7 S nerve growth factor has been determined from analyses of the peptides generated by cyanogen bromide, trypsin, and chymotrypsin from the naturally occurring fragments. All peptides were sequenced automatically in a spinning-cup sequenator using Polybrene to minimize extractive losses by the solvents employed thoughout the degradation cycles. The gamma-subunit, a serine protease with arginine specificity, contains 233 amino acid residues and shares sequence homology with other proteases of this family. The five disulfide bonds of the gamma-subunit are a subset of the six disulfides present in bovine trypsin, as judged by the location of the half-cystine residues in the primary structure. An N-linked carbohydrate side chain is attached to Asn-78 in at least a majority of tee gamma-molecules.
Notes: Thomas, K A
Baglan, N C
Bradshaw, R A
AM 13362/AM/NIADDK NIH HHS/United States
NS 10229/NS/NINDS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1981 Sep 10;256(17):9156-66.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1012
Author: Thomassen, E. A., van Veen, H. A., van Berkel, P. H., Nuijens, J. H. and Abrahams, J. P.
Year: 2005
Title: The protein structure of recombinant human lactoferrin produced in the milk of transgenic cows closely matches the structure of human milk-derived lactoferrin
Journal: Transgenic Res
Volume: 14
Issue: 4
Pages: 397-405
Epub Date: 2005/10/06
Date: Aug
Short Title: The protein structure of recombinant human lactoferrin produced in the milk of transgenic cows closely matches the structure of human milk-derived lactoferrin
Alternate Journal: Transgenic research
ISSN: 0962-8819 (Print)
0962-8819
DOI: 10.1007/s11248-005-3233-0
Accession Number: 16201406
Keywords: Animals
Animals, Genetically Modified
Biopharmaceutics
Cattle/genetics
Crystallography, X-Ray
Glycosylation
Humans
Lactoferrin/biosynthesis/*chemistry
Milk/*metabolism
Protein Conformation
Recombinant Proteins/biosynthesis/*chemistry
Abstract: Human lactoferrin (hLF) is an iron-binding glycoprotein involved in the host defence against infection and excessive inflammation. As the availability of (human milk-derived) natural hLF is limited, alternative means of production of this biopharmaceutical are extensively researched. Here we report the crystal structure of recombinant hLF (rhLF) expressed in the milk of transgenic cows at a resolution of 2.4 A. To our knowledge, the first reported structure of a recombinant protein produced in milk of transgenic livestock. Even though rhLF contains oligomannose- and hybrid-type N-linked glycans next to complex-type glycans, which are the only glycans found on natural hLF, the structures are identical within the experimental error (r.m.s. deviation of only 0.28 A for the main-chain atoms). Of the differences in polymorphic amino acids between the natural and rhLF variant used, only the side-chain of Asp561 could be modeled into the rhLF electron density map. Taken together, the results confirm the structural integrity of the rhLF variant used in this study. It also confirms the validity of the transgenic cow mammary gland as a vehicle to produce recombinant human proteins.
Notes: Thomassen, Ellen A J
van Veen, Harrie A
van Berkel, Patrick H C
Nuijens, Jan H
Abrahams, Jan Pieter
Journal Article
Netherlands
Transgenic Res. 2005 Aug;14(4):397-405. doi: 10.1007/s11248-005-3233-0.
Author Address: Biophysical Structural Chemistry, Leiden Institute of Chemistry, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1495
Author: Thorn, A., Steinfeld, R., Ziegenbein, M., Grapp, M., Hsiao, H. H., Urlaub, H., Sheldrick, G. M., Gärtner, J. and Krätzner, R.
Year: 2012
Title: Structure and activity of the only human RNase T2
Journal: Nucleic Acids Res
Volume: 40
Issue: 17
Pages: 8733-42
Epub Date: 2012/06/28
Date: Sep 1
Short Title: Structure and activity of the only human RNase T2
Alternate Journal: Nucleic acids research
ISSN: 0305-1048 (Print)
0305-1048
DOI: 10.1093/nar/gks614
PMCID: PMC3458558
Accession Number: 22735700
Keywords: Amino Acid Sequence
Binding Sites
Copper/pharmacology
Crystallography, X-Ray
Endoribonucleases/*chemistry/genetics/metabolism
Glycosylation
Humans
Models, Molecular
Molecular Sequence Data
Mutation
Protein Folding
Structural Homology, Protein
Zinc/chemistry/pharmacology
Abstract: Mutations in the gene of human RNase T2 are associated with white matter disease of the human brain. Although brain abnormalities (bilateral temporal lobe cysts and multifocal white matter lesions) and clinical symptoms (psychomotor impairments, spasticity and epilepsy) are well characterized, the pathomechanism of RNase T2 deficiency remains unclear. RNase T2 is the only member of the Rh/T2/S family of acidic hydrolases in humans. In recent years, new functions such as tumor suppressing properties of RNase T2 have been reported that are independent of its catalytic activity. We determined the X-ray structure of human RNase T2 at 1.6 Å resolution. The α+β core fold shows high similarity to those of known T2 RNase structures from plants, while, in contrast, the external loop regions show distinct structural differences. The catalytic features of RNase T2 in presence of bivalent cations were analyzed and the structural consequences of known clinical mutations were investigated. Our data provide further insight into the function of human RNase T2 and may prove useful in understanding its mode of action independent of its enzymatic activity.
Notes: 1362-4962
Thorn, Andrea
Steinfeld, Robert
Ziegenbein, Marc
Grapp, Marcel
Hsiao, He-Hsuan
Urlaub, Henning
Sheldrick, George M
Gärtner, Jutta
Krätzner, Ralph
Journal Article
Research Support, Non-U.S. Gov't
Nucleic Acids Res. 2012 Sep 1;40(17):8733-42. doi: 10.1093/nar/gks614. Epub 2012 Jun 26.
Author Address: Department of Structural Chemistry, University of Göttingen, 37075 Göttingen, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1774
Author: Tian, J., Geng, Q., Ding, Y., Liao, J., Dong, M. Q., Xu, X. and Li, J.
Year: 2016
Title: O-GlcNAcylation Antagonizes Phosphorylation of CDH1 (CDC20 Homologue 1)
Journal: J Biol Chem
Volume: 291
Issue: 23
Pages: 12136-44
Epub Date: 2016/04/16
Date: Jun 3
Short Title: O-GlcNAcylation Antagonizes Phosphorylation of CDH1 (CDC20 Homologue 1)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.717850
PMCID: PMC4933264
Accession Number: 27080259
Keywords: Acetylglucosamine/*metabolism
Acylation
Anaphase-Promoting Complex-Cyclosome/genetics/*metabolism
Animals
Binding Sites/genetics
Cdh1 Proteins/genetics/*metabolism
Glycosylation
HeLa Cells
Humans
Immunoblotting
Mice
Mutation
N-Acetylglucosaminyltransferases/genetics/metabolism
Phosphorylation
Protein Binding
*Protein Processing, Post-Translational
Tandem Mass Spectrometry
Cdh1
O-linked N-acetylglucosamine (O-GlcNAc)
O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
anaphase promoting complex/cyclosome (APC/C)
cell cycle
mitosis
Abstract: The anaphase promoting complex/cyclosome (APC/C) orchestrates various aspects of the eukaryotic cell cycle. One of its co-activators, Cdh1, is subject to myriad post-translational modifications, such as phosphorylation and ubiquitination. Herein we identify the O-linked N-acetylglucosamine (O-GlcNAc) modification that occurs on Cdh1. Cdh1 is O-GlcNAcylated in cultured cells and mouse brain extracts. Mass spectrometry identifies an O-GlcNAcylated peptide that neighbors a known phosphorylation site. Cell synchronization and mutation studies reveal that O-GlcNAcylation of Cdh1 may antagonize its phosphorylation. Our results thus reveal a pivotal role of O-GlcNAcylation in regulating APC/C activity.
Notes: 1083-351x
Tian, Jie
Geng, Qizhi
Ding, Yuehe
Liao, Ji
Dong, Meng-Qiu
Xu, Xingzhi
Li, Jing
Orcid: 0000-0002-3977-1641
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2016 Jun 3;291(23):12136-44. doi: 10.1074/jbc.M116.717850. Epub 2016 Apr 14.
Author Address: From the Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China and.
National Institute of Biological Sciences, Beijing 102206, China.
From the Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China and xingzhi_xu@cnu.edu.cn.
From the Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China and jing_li@cnu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2011
Author: Tian, J. L. and Qin, H.
Year: 2019
Title: O-GlcNAcylation Regulates Primary Ciliary Length by Promoting Microtubule Disassembly
Journal: iScience
Volume: 12
Pages: 379-391
Epub Date: 2019/02/24
Date: Feb 22
Short Title: O-GlcNAcylation Regulates Primary Ciliary Length by Promoting Microtubule Disassembly
Alternate Journal: iScience
ISSN: 2589-0042
DOI: 10.1016/j.isci.2019.01.031
PMCID: PMC6374784
Accession Number: 30796923
Keywords: Biological Sciences
Cell Biology
Functional Aspects of Cell Biology
Molecular Biology
Abstract: The sensory organelle cilium is involved in sensing and transducing important signaling cascades in almost all cells of our body. These ciliary-mediated pathways affect cellular homeostasis and metabolisms profoundly. However, it is almost completely unknown whether the cellular metabolic state affects the assembly of cilia. This study is to investigate how O-linked β-N-acetylglucosamine (O-GlcNAc), a sensor of cellular nutrients, regulates the cilia length. Pharmacologic or genetic inhibition of O-GlcNAcylation led to longer cilia, and vice versa. Further biochemical assays revealed that both α-tubulin and HDAC6 (histone deacetylase 6) were O-GlcNAcylated in vivo. In vitro enzymatic assays showed that O-GlcNAcylation of either tubulin or HDAC6 promoted microtubule disassembly, which likely in turn caused ciliary shortening. Taken together, these results uncovered a negative regulatory role of O-GlcNAc in modulating the ciliary microtubule assembly. The cross talk between O-GlcNAc and cilium is likely critical for fine-tuning the cellular response to nutrients.
Notes: 2589-0042
Tian, Jie L
Qin, Hongmin
Journal Article
iScience. 2019 Feb 22;12:379-391. doi: 10.1016/j.isci.2019.01.031. Epub 2019 Jan 31.
Author Address: Molecular & Environmental Plant Sciences, Texas A&M University, College Station, TX, USA.
Molecular & Environmental Plant Sciences, Texas A&M University, College Station, TX, USA; Department of Biology, Texas A&M University, College Station, TX 77843, USA. Electronic address: hqin@bio.tamu.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1927
Author: Tidhar, R., Zelnik, I. D., Volpert, G., Ben-Dor, S., Kelly, S., Merrill, A. H., Jr. and Futerman, A. H.
Year: 2018
Title: Eleven residues determine the acyl chain specificity of ceramide synthases
Journal: J Biol Chem
Volume: 293
Issue: 25
Pages: 9912-9921
Epub Date: 2018/04/11
Date: Jun 22
Short Title: Eleven residues determine the acyl chain specificity of ceramide synthases
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA118.001936
PMCID: PMC6016465
Accession Number: 29632068
Keywords: Acyl Coenzyme A/*metabolism
Amino Acid Sequence
Base Sequence
CRISPR-Cas Systems
Ceramides/*metabolism
Humans
Oxidoreductases/antagonists & inhibitors/*classification/*metabolism
Sequence Homology
Sphingolipids/*metabolism
Substrate Specificity
*ceramide
*ceramide synthase
*lipid
*membrane
*sphingolipid
this article
Abstract: Lipids display large structural complexity, with ∼40,000 different lipids identified to date, ∼4000 of which are sphingolipids. A critical factor determining the biological activities of the sphingolipid, ceramide, and of more complex sphingolipids is their N-acyl chain length, which in mammals is determined by a family of six ceramide synthases (CerS). Little information is available about the CerS regions that determine specificity toward different acyl-CoA substrates. We previously demonstrated that substrate specificity resides in a region of ∼150 residues in the Tram-Lag-CLN8 domain. Using site-directed mutagenesis and biochemical analyses, we now narrow specificity down to an 11-residue sequence in a loop located between the last two putative transmembrane domains (TMDs) of the CerS. The specificity of a chimeric protein, CerS5((299-309→CerS2)), based on the backbone of CerS5 (which generates C16-ceramide), but containing 11 residues from CerS2 (which generates C22-C24-ceramides), was altered such that it generated C22-C24 and other ceramides. Moreover, a chimeric protein, CerS4((291-301→CerS2)), based on CerS4 (which normally generates C18-C22 ceramides) displayed significant activity toward C24:1-CoA. Additional data supported the notion that substitutions of these 11 residues alter the specificities of the CerS toward their cognate acyl-CoAs. Our findings may suggest that this short loop may restrict adjacent TMDs, leading to a more open conformation in the membrane, and that the CerS acting on shorter acyl-CoAs may have a longer, more flexible loop, permitting TMD flexibility. In summary, we have identified an 11-residue region that determines the acyl-CoA specificity of CerS.
Notes: 1083-351x
Tidhar, Rotem
Zelnik, Iris D
Volpert, Giora
Ben-Dor, Shifra
Orcid: 0000-0001-9604-1939
Kelly, Samuel
Merrill, Alfred H Jr
Futerman, Anthony H
R01 GM076217/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2018 Jun 22;293(25):9912-9921. doi: 10.1074/jbc.RA118.001936. Epub 2018 Apr 9.
Author Address: From the Department of Biomolecular Sciences and.
the Life Sciences Core Facilities, Weizmann Institute of Science, Rehovot 76100, Israel and.
the School of Biology and Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, Georgia 30332-0230.
From the Department of Biomolecular Sciences and tony.futerman@weizmann.ac.il.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 964
Author: Tiede, S., Cantz, M., Raas-Rothschild, A., Muschol, N., Bürger, F., Ullrich, K. and Braulke, T.
Year: 2004
Title: A novel mutation in UDP-N-acetylglucosamine-1-phosphotransferase gamma subunit (GNPTAG) in two siblings with mucolipidosis type III alters a used glycosylation site
Journal: Hum Mutat
Volume: 24
Issue: 6
Pages: 535
Epub Date: 2004/11/09
Date: Dec
Short Title: A novel mutation in UDP-N-acetylglucosamine-1-phosphotransferase gamma subunit (GNPTAG) in two siblings with mucolipidosis type III alters a used glycosylation site
Alternate Journal: Human mutation
ISSN: 1059-7794
DOI: 10.1002/humu.9293
Accession Number: 15532026
Keywords: Binding Sites
Child
DNA Mutational Analysis
Female
Glycosylation
Humans
Lysosomes/enzymology
Male
Mucolipidoses/enzymology/*genetics
Sequence Analysis, DNA
Transferases (Other Substituted Phosphate Groups)/*genetics
Abstract: The N-acetylglucosaminyl-1-phosphotransferase (termed phosphotransferase) catalyzes the initial step in the formation of mannose 6-phosphate (M6P) residues required for the efficient transport of soluble lysosomal enzymes. The phosphotransferase is a multisubunit enzyme composed of three subunits (alpha2beta2gamma2) that are products of two genes. The gene encoding the gamma-subunit (GNPTAG) appears to be defective in patients with mucolipidosis type III (ML III). We have analyzed the GNPTAG gene in two siblings with ML III showing elevated activities of several lysosomal enzymes in cultured fibroblasts serum and diminished activities in cultured fibroblasts. Immunoprecipitation of metabolically labeled cathepsin D (CtsD) from fibroblasts revealed that the sorting/transport of this lysosomal protease was affected. Addition of ammonium chloride inhibiting pH-dependent processes, such as the CtsD-M6P receptor interaction, indicated that 15 to 20% of the newly synthesized CtsD is transported in ML III fibroblasts in an M6P-dependent manner. By direct sequencing a novel homozygous mutation, c.347_349delACA (p.Asn116del), was identified affecting a potential N-linked glycosylation site. Western blot analysis of extracts from control fibroblasts detect a 97 kDa glycosylated dimer whereas ML III cells contain a GNPTAG dimer of reduced molecular mass. These data suggest that the loss of the used glycosylation site in the gamma subunit may affect the intracellular localization of GNPTAG and the overall efficiency of M6P formation.
Notes: 1098-1004
Tiede, Stephan
Cantz, Michael
Raas-Rothschild, Annick
Muschol, Nicole
Bürger, Friederike
Ullrich, Kurt
Braulke, Thomas
Case Reports
Journal Article
Research Support, Non-U.S. Gov't
United States
Hum Mutat. 2004 Dec;24(6):535. doi: 10.1002/humu.9293.
Author Address: Department of Biochemistry, Children's Hospital, Hamburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 406
Author: Titani, K., Kumar, S., Takio, K., Ericsson, L. H., Wade, R. D., Ashida, K., Walsh, K. A., Chopek, M. W., Sadler, J. E. and Fujikawa, K.
Year: 1986
Title: Amino acid sequence of human von Willebrand factor
Journal: Biochemistry
Volume: 25
Issue: 11
Pages: 3171-84
Epub Date: 1986/06/03
Date: Jun 3
Short Title: Amino acid sequence of human von Willebrand factor
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00359a015
Accession Number: 3524673
Keywords: Amino Acid Sequence
Chromatography, High Pressure Liquid
Cyanogen Bromide
Humans
Peptide Fragments/analysis
Peptide Hydrolases
*von Willebrand Factor
Abstract: The complete amino acid sequence of human von Willebrand factor (vWF) is presented. Most of the sequence was determined by analysis of the S-carboxymethylated protein. Some overlaps not provided by the protein sequence analysis were obtained from the sequence predicted by the nucleotide sequence of a cDNA clone [Sadler, J.E., Shelton-Inloes, B.B., Sorace, J., Harlan, M., Titani, K., & Davie, E.W. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 6391-6398]. The protein is composed of 2050 amino acid residues containing 12 Asn-linked and 10 Thr/Ser-linked oligosaccharide chains. One of the carbohydrate chains is linked to an Asn residue in the sequence Asn-Ser-Cys rather than the usual Asn-X-Ser/Thr sequence. The sequence of von Willebrand factor includes several regions bearing evidence of internal gene duplication of ancestral sequences. The protein also contains the tetrapeptide sequence Arg-Gly-Asp-Ser (at residues 1744-1747), which may be a cell attachment site, as in fibronectin. The amino- and carboxyl-terminal regions of the molecule contain clusters of half-cystinyl residues. The sequence is unique except for some homology to human complement factor B.
Notes: Titani, K
Kumar, S
Takio, K
Ericsson, L H
Wade, R D
Ashida, K
Walsh, K A
Chopek, M W
Sadler, J E
Fujikawa, K
HL16919/HL/NHLBI NIH HHS/United States
HL29595/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1986 Jun 3;25(11):3171-84. doi: 10.1021/bi00359a015.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 403
Author: Titani, K., Takio, K., Handa, M. and Ruggeri, Z. M.
Year: 1987
Title: Amino acid sequence of the von Willebrand factor-binding domain of platelet membrane glycoprotein Ib
Journal: Proc Natl Acad Sci U S A
Volume: 84
Issue: 16
Pages: 5610-4
Epub Date: 1987/08/01
Date: Aug
Short Title: Amino acid sequence of the von Willebrand factor-binding domain of platelet membrane glycoprotein Ib
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.84.16.5610
PMCID: PMC298912
Accession Number: 3497398
Keywords: Amino Acid Sequence
Binding Sites
Humans
Models, Molecular
Peptide Mapping
Platelet Membrane Glycoproteins/*analysis/metabolism
Protein Conformation
von Willebrand Factor/*analysis/metabolism
Abstract: We report the amino acid sequence of a 299-residue segment from the alpha chain of the human platelet membrane glycoprotein Ib. This includes the complete sequence of the amino-terminal tryptic fragment of 290 residues comprising the von Willebrand factor-binding domain. Two primary sets of overlapping fragments were obtained by cleavage of the S-carboxymethylated protein at methionyl and lysyl bonds following treatment with cyanogen bromide and Achromobacter protease I, respectively. Additional fragments were obtained by treatment of native glycocalicin with trypsin, Staphylococcus aureus V8 protease, and Serratia marcescens protease. Analysis of all these fragments provided data that allowed determination of the continuous sequence corresponding to approximately half of the alpha-chain polypeptide. This region of glycoprotein Ib is largely hydrophobic and contains only two N-linked and one O-linked carbohydrate chains. A hydrophilic region exists between residues 215 and 299, which contains a cluster of 10 negatively charged residues at 269-287. This area is likely to attract positively charged molecules. The hydrophilic, highly glycosylated (at serine and threonine residues) region corresponding to the previously described "macroglycopeptide" and representing the carboxyl-terminal half of the alpha chain is likely to begin at residue 292. The determined sequence of the alpha chain of glycoprotein Ib contains a region (residues 29-193) with seven repeats, which is indicative of gene duplication and is highly homologous to human leucine-rich alpha 2-glycoprotein. This protein sequence agrees completely with that deduced from the cDNA sequence reported by Lopez et al. [Lopez, J.A., Chung, D.W., Fujikawa, K., Hagen, F.S., Papayannopoulou, T. & Roth, G.J. (1987) Proc. Natl. Acad. Sci. USA 84, 5615-5619].
Notes: 1091-6490
Titani, K
Takio, K
Handa, M
Ruggeri, Z M
GM 15731/GM/NIGMS NIH HHS/United States
HL 29595/HL/NHLBI NIH HHS/United States
HL 31950/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1987 Aug;84(16):5610-4. doi: 10.1073/pnas.84.16.5610.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 360
Author: Tollersrud, O. K. and Aronson, N. N., Jr.
Year: 1989
Title: Purification and characterization of rat liver glycosylasparaginase
Journal: Biochem J
Volume: 260
Issue: 1
Pages: 101-8
Epub Date: 1989/05/15
Date: May 15
Short Title: Purification and characterization of rat liver glycosylasparaginase
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2600101
PMCID: PMC1138631
Accession Number: 2775174
Keywords: Amidohydrolases/*isolation & purification
Amino Acid Sequence
Animals
Aspartylglucosylaminase/*isolation & purification
Electrophoresis, Polyacrylamide Gel
Enzyme Stability
Hot Temperature
Hydrogen-Ion Concentration
Liver/*enzymology
Molecular Conformation
Molecular Sequence Data
Oligosaccharides/isolation & purification
Rats
Abstract: 1. Rat liver glycosylasparaginase [N4-(beta-N-acetylglucosaminyl)-L-asparaginase, EC 3.5.1.26] was purified to homogeneity by using salt fractionation, CM-cellulose and DEAE-cellulose chromatography, gel filtration on Ultrogel AcA-54, concanavalin A-Sepharose affinity chromatography, heat treatment at 70 degrees C and preparative SDS/polyacrylamide-gel electrophoresis. The purified enzyme had a specific activity of 3.8 mumol of N-acetylglucosamine/min per mg with N4-(beta-N-acetylglucosaminyl)-L-asparagine as substrate. 2. The native enzyme had a molecular mass of 49 kDa and was composed of two non-identical subunits joined by strong non-covalent forces and having molecular masses of 24 and 20 kDa as determined by SDS/polyacrylamide-gel electrophoresis. 3. The 20 kDa subunit contained one high-mannose-type oligosaccharide chain, and the 24 kDa subunit had one high-mannose-type and one complex-type oligosaccharide chain. 4. N-Terminal sequence analysis of each subunit revealed a frayed N-terminus of the 24 kDa subunit and an apparent N-glycosylation of Asn-15 in the same subunit. 5. The enzyme exhibited a broad pH maximum above 7. Two major isoelectric forms were found at pH 6.4 and 6.6. 6. Glycosylasparaginase was stable at 75 degrees C and in 5% (w/v) SDS at pH 7.0.
Notes: 1470-8728
Tollersrud, O K
Aronson, N N Jr
DK-33314/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Biochem J. 1989 May 15;260(1):101-8. doi: 10.1042/bj2600101.
Author Address: Department of Molecular and Cell Biology, Pennsylvania State University, University Park 16802.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 226
Author: Tomita, M. and Marchesi, V. T.
Year: 1975
Title: Amino-acid sequence and oligosaccharide attachment sites of human erythrocyte glycophorin
Journal: Proc Natl Acad Sci U S A
Volume: 72
Issue: 8
Pages: 2964-8
Epub Date: 1975/08/01
Date: Aug
Short Title: Amino-acid sequence and oligosaccharide attachment sites of human erythrocyte glycophorin
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.72.8.2964
PMCID: PMC432899
Accession Number: 1059087
Keywords: Amino Acids/analysis
Binding Sites
Blood Proteins/*analysis
Erythrocytes/*analysis
Glycoproteins/*analysis
Hexoses/analysis
Humans
Macromolecular Substances
Molecular Weight
Oligosaccharides/analysis
Protein Binding
Sialic Acids/analysis
Abstract: Glycophorin, the major sialoglycoprotein of the human erythrocyte membrane, is composed of 131 amino acids and an average of 16 oligosaccharide chains. Fifteen oligosaccharides are linked to threonine/serine residues via O-glycosidic bonds, and one more complex unit is attached to asparagine. The location of each of these oligosaccharides and the complete amino-acid sequence of this molecular have been determined by Edman degradation techniques. Glycophorin appears to be organized into three distinct "domains" on the basis of the locations of glycosylated amino acids and the clustering of residues of similar type. These include (i) a glycosylated segment composed of approximately 64 residues from the NH2-terminus, (ii) a "hydrophobic" segment of approximately 32 nonpolar residues, and (iii) a COOH-terminal segment, composed of approximately 35 residues, which has an unusual concentration of hydrophilic amino acids. This unique structure is consistent with the earlier suggestions that glycophorin is one of the major "intrinsic" membrane proteins which has a transmembrane orientation.
Notes: 1091-6490
Tomita, M
Marchesi, V T
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 1975 Aug;72(8):2964-8. doi: 10.1073/pnas.72.8.2964.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 591
Author: Tomiya, N., Awaya, J., Kurono, M., Hanzawa, H., Shimada, I., Arata, Y., Yoshida, T. and Takahashi, N.
Year: 1993
Title: Structural elucidation of a variety of GalNAc-containing N-linked oligosaccharides from human urinary kallidinogenase
Journal: J Biol Chem
Volume: 268
Issue: 1
Pages: 113-26
Epub Date: 1993/01/05
Date: Jan 5
Short Title: Structural elucidation of a variety of GalNAc-containing N-linked oligosaccharides from human urinary kallidinogenase
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 8416919
Keywords: Acetylgalactosamine/*analysis
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography
Gas Chromatography-Mass Spectrometry
Humans
Indicators and Reagents
Kallikreins/chemistry/*urine
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Abstract: Fifteen different structures of terminal GalNAc-containing N-linked oligosaccharides from human urinary kallidinogenase have been identified. These N-linked oligosaccharides were mostly neutral, because sialic acid content was lower than 0.13 mol of sialic acid/mol of sugar chain, and sulfate was not detected. The oligosaccharides were released from pepsin-digested protein by glycoamidase A (from almond) digestion. The reducing ends of the oligosaccharide chains were aminated with a fluorescent reagent, 2-aminopyridine. The resulting mixture of pyridylamino derivatives of the oligosaccharides were separated by high performance liquid chromatography on an ODS-silica column, and 15 oligosaccharides were isolated. The structure of each oligosaccharide fraction was analyzed by two-dimensional sugar mapping, component sugar analysis, high resolution proton nuclear magnetic resonance and methylation analysis. It was found that each N-linked oligosaccharide associated with human urinary kallidinogenase contains unsubstituted GalNAc residues at the nonreducing terminal. These 15 oligosaccharides include 5 biantennary, 7 triantennary, and 3 tetraantennary oligosaccharides.
Notes: Tomiya, N
Awaya, J
Kurono, M
Hanzawa, H
Shimada, I
Arata, Y
Yoshida, T
Takahashi, N
Journal Article
United States
J Biol Chem. 1993 Jan 5;268(1):113-26.
Author Address: Mie Research Laboratory, Sanwa Kagaku Kenkyusho Co. Ltd., Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 416
Author: Tomiya, N., Kurono, M., Ishihara, H., Tejima, S., Endo, S., Arata, Y. and Takahashi, N.
Year: 1987
Title: Structural analysis of N-linked oligosaccharides by a combination of glycopeptidase, exoglycosidases, and high-performance liquid chromatography
Journal: Anal Biochem
Volume: 163
Issue: 2
Pages: 489-99
Epub Date: 1987/06/01
Date: Jun
Short Title: Structural analysis of N-linked oligosaccharides by a combination of glycopeptidase, exoglycosidases, and high-performance liquid chromatography
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/0003-2697(87)90253-3
Accession Number: 3661998
Keywords: Amidohydrolases
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Glycoproteins
Glycoside Hydrolases
Myeloma Proteins
*Oligosaccharides
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Reference Standards
Abstract: A simple, sensitive, and rapid method for the analysis of structures of N-linked carbohydrates is reported. The method involves four steps: preparation of carbohydrate chains from glycopeptides by N-oligosaccharide glycopeptidase digestion; derivatization of the reducing ends of carbohydrate chains with a fluorescent reagent, 2-aminopyridine, by using sodium cyanoborohydride; separation of oligosaccharide derivatives by reverse-phase high-performance liquid chromatography; and structural analysis of oligosaccharides by sequential exoglycosidase digestion. The elution positions of 50 standard oligosaccharide derivatives were determined by HPLC. The structure of an unknown oligosaccharide can be characterized by comparison of its elution position with those of the standard compounds. The method was applied to elucidate the structures of oligosaccharides in the myeloma IgG protein, Yot.
Notes: Tomiya, N
Kurono, M
Ishihara, H
Tejima, S
Endo, S
Arata, Y
Takahashi, N
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Biochem. 1987 Jun;163(2):489-99. doi: 10.1016/0003-2697(87)90253-3.
Author Address: Mie Research Laboratory, Sanwa Kagaku Kenkyusho Co. Ltd., Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1677
Author: Torkko, J. M., Primo, M. E., Dirkx, R., Friedrich, A., Viehrig, A., Vergari, E., Borgonovo, B., Sönmez, A., Wegbrod, C., Lachnit, M., Münster, C., Sica, M. P., Ermácora, M. R. and Solimena, M.
Year: 2015
Title: Stability of proICA512/IA-2 and its targeting to insulin secretory granules require β4-sheet-mediated dimerization of its ectodomain in the endoplasmic reticulum
Journal: Mol Cell Biol
Volume: 35
Issue: 6
Pages: 914-27
Epub Date: 2015/01/07
Date: Mar
Short Title: Stability of proICA512/IA-2 and its targeting to insulin secretory granules require β4-sheet-mediated dimerization of its ectodomain in the endoplasmic reticulum
Alternate Journal: Molecular and cellular biology
ISSN: 0270-7306 (Print)
0270-7306
DOI: 10.1128/mcb.00994-14
PMCID: PMC4333099
Accession Number: 25561468
Keywords: Amino Acid Sequence
Animals
Cells, Cultured
Cytoplasmic Granules/*metabolism
Cytosol/metabolism
Dimerization
Endoplasmic Reticulum/*metabolism
Glycosylation
Insulin/*metabolism
Islets of Langerhans/metabolism
Molecular Sequence Data
Protein Structure, Tertiary
Rats
Receptor-Like Protein Tyrosine Phosphatases, Class 8/*metabolism
Secretory Vesicles/*metabolism
Abstract: The type 1 diabetes autoantigen ICA512/IA-2/RPTPN is a receptor protein tyrosine phosphatase of the insulin secretory granules (SGs) which regulates the size of granule stores, possibly via cleavage/signaling of its cytosolic tail. The role of its extracellular region remains unknown. Structural studies indicated that β2- or β4-strands in the mature ectodomain (ME ICA512) form dimers in vitro. Here we show that ME ICA512 prompts proICA512 dimerization in the endoplasmic reticulum. Perturbation of ME ICA512 β2-strand N-glycosylation upon S508A replacement allows for proICA512 dimerization, O-glycosylation, targeting to granules, and conversion, which are instead precluded upon G553D replacement in the ME ICA512 β4-strand. S508A/G553D and N506A/G553D double mutants dimerize but remain in the endoplasmic reticulum. Removal of the N-terminal fragment (ICA512-NTF) preceding ME ICA512 allows an ICA512-ΔNTF G553D mutant to exit the endoplasmic reticulum, and ICA512-ΔNTF is constitutively delivered to the cell surface. The signal for SG sorting is located within the NTF RESP18 homology domain (RESP18-HD), whereas soluble NTF is retained in the endoplasmic reticulum. Hence, we propose that the ME ICA512 β2-strand fosters proICA512 dimerization until NTF prevents N506 glycosylation. Removal of this constraint allows for proICA512 β4-strand-induced dimerization, exit from the endoplasmic reticulum, O-glycosylation, and RESP18-HD-mediated targeting to granules.
Notes: 1098-5549
Torkko, Juha M
Primo, M Evangelina
Dirkx, Ronald
Friedrich, Anne
Viehrig, Antje
Vergari, Elisa
Borgonovo, Barbara
Sönmez, Anke
Wegbrod, Carolin
Lachnit, Martina
Münster, Carla
Sica, Mauricio P
Ermácora, Mario R
Solimena, Michele
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Biol. 2015 Mar;35(6):914-27. doi: 10.1128/MCB.00994-14. Epub 2015 Jan 5.
Author Address: Paul Langerhans Institute Dresden, Uniklinikum Carl Gustav Carus, TU Dresden, Germany German Center for Diabetes Research (DZD e.V.), Neuherberg, Germany.
University of Buenos Aires, Buenos Aires, Argentina Instituto Multidisciplinario de Biología Celular, Consejo Nacional de Investigaciones Científicas y Técnicas, Buenos Aires, Argentina.
Paul Langerhans Institute Dresden, Uniklinikum Carl Gustav Carus, TU Dresden, Germany Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.
Instituto Multidisciplinario de Biología Celular, Consejo Nacional de Investigaciones Científicas y Técnicas, Buenos Aires, Argentina Departamento de Ciencia y Tecnología, Universidad Nacional de Quilmes, Bernal, Buenos Aires, Argentina.
Paul Langerhans Institute Dresden, Uniklinikum Carl Gustav Carus, TU Dresden, Germany German Center for Diabetes Research (DZD e.V.), Neuherberg, Germany Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany michele.solimena@tu-dresden.de.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 823
Author: Toth, J., Cutforth, T., Gelinas, A. D., Bethoney, K. A., Bard, J. and Harrison, C. J.
Year: 2001
Title: Crystal structure of an ephrin ectodomain
Journal: Dev Cell
Volume: 1
Issue: 1
Pages: 83-92
Epub Date: 2001/11/13
Date: Jul
Short Title: Crystal structure of an ephrin ectodomain
Alternate Journal: Developmental cell
ISSN: 1534-5807 (Print)
1534-5807
DOI: 10.1016/s1534-5807(01)00002-8
Accession Number: 11703926
Keywords: Amino Acid Sequence
Animals
Binding Sites/genetics
Crystallography
Ephrin-B2
Ligands
Membrane Proteins/*chemistry/*genetics/metabolism
Mice
Molecular Sequence Data
Mutation, Missense
Plant Proteins/chemistry
Protein Structure, Quaternary
Protein Structure, Tertiary
Abstract: Eph receptor tyrosine kinases and their membrane-associated ligands, the ephrins, are essential regulators of axon guidance, cell migration, segmentation, and angiogenesis. There are two classes of vertebrate ephrin ligands which have distinct binding specificities for their cognate receptors. Multimerization of the ligands is required for receptor activation, and ephrin ligands themselves signal intracellularly upon binding Eph receptors. We have determined the structure of the extracellular domain of mouse ephrin-B2. The ephrin ectodomain is an eight-stranded beta barrel with topological similarity to plant nodulins and phytocyanins. Based on the structure, we have identified potential surface determinants of Eph/ephrin binding specificity and a ligand dimerization region. The high sequence similarity among ephrin ectodomains indicates that all ephrins may be modeled upon the ephrin-B2 structure presented here.
Notes: Toth, J
Cutforth, T
Gelinas, A D
Bethoney, K A
Bard, J
Harrison, C J
P01 CA23767/CA/NCI NIH HHS/United States
R01 DC04209/DC/NIDCD NIH HHS/United States
R01 GM58256/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Dev Cell. 2001 Jul;1(1):83-92. doi: 10.1016/s1534-5807(01)00002-8.
Author Address: Boston Biomedical Research Institute, Watertown, Massachusetts 02472, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 907
Author: Tournamille, C., Filipe, A., Wasniowska, K., Gane, P., Lisowska, E., Cartron, J. P., Colin, Y. and Le Van Kim, C.
Year: 2003
Title: Structure-function analysis of the extracellular domains of the Duffy antigen/receptor for chemokines: characterization of antibody and chemokine binding sites
Journal: Br J Haematol
Volume: 122
Issue: 6
Pages: 1014-23
Epub Date: 2003/09/06
Date: Sep
Short Title: Structure-function analysis of the extracellular domains of the Duffy antigen/receptor for chemokines: characterization of antibody and chemokine binding sites
Alternate Journal: British journal of haematology
ISSN: 0007-1048 (Print)
0007-1048
DOI: 10.1046/j.1365-2141.2003.04533.x
Accession Number: 12956774
Keywords: Amino Acid Sequence
Antibodies, Monoclonal/metabolism
Binding Sites
Chemokines/metabolism
Chemokines, CXC/metabolism
Duffy Blood-Group System/genetics/*immunology/metabolism
Epitope Mapping
Glycosylation
Humans
K562 Cells
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Binding
Receptors, Cell Surface/genetics/*immunology/metabolism
Structure-Activity Relationship
Transfection
Abstract: The Duffy antigen/receptor for chemokines (DARC), a seven-transmembrane glycoprotein carrying the Duffy (Fy) blood group, acts as a widely expressed promiscuous chemokine receptor. In a structure-function study, we analysed the binding of chemokines and anti-Fy monoclonal antibodies (mAbs) to K562 cells expressing 39 mutant forms of DARC with alanine substitutions spread out on the four extracellular domains (ECDs). Using synthetic peptides, we defined previously the Fy6 epitope (22-FEDVW-26), and we characterized the Fya epitope as the linear sequence 41-YGANLE-46. In agreement with these results, mutations of F22-E23, V25 and Y41, G42, N44, L45 on ECD1 abolished the binding of anti-Fy6 and anti-Fya mAbs to K562 cells respectively, Anti-Fy3 binding was abolished by D58-D59 (ECD1), R124 (ECD2), D263 and D283 (ECD4) substitutions. Mutations of C51 (ECD1), C129 (ECD2), C195 (ECD3) and C276 (ECD4 severely reduced anti-Fy3 and CXC-chemokine ligand 8 (CXCL-8) binding. CXCL-8 binding was also abrogated by mutations of F22-E23, P50 (ECD1) and D263, R267, D283 (ECD4). These results defined the Fya epitope and suggested that (1) two disulphide bridges are involved in the creation of an active chemokine binding pocket; (2) a limited number of amino acids in ECDs 1-4 participate in CXCL-8 binding; and (3) Fy3 is a conformation-dependent epitope involving all ECDs. We also showed that N-glycosylation of DARC occurred on N16SS and did not influence antibody and chemokine binding.
Notes: Tournamille, Christophe
Filipe, Anne
Wasniowska, Kazimiera
Gane, Pierre
Lisowska, Elwira
Cartron, Jean-Pierre
Colin, Yves
Le Van Kim, Caroline
Journal Article
Research Support, Non-U.S. Gov't
England
Br J Haematol. 2003 Sep;122(6):1014-23. doi: 10.1046/j.1365-2141.2003.04533.x.
Author Address: INSERM U76, Institut National de la Transfusion Sanguine, Paris, France.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 922
Author: Towler, P., Staker, B., Prasad, S. G., Menon, S., Tang, J., Parsons, T., Ryan, D., Fisher, M., Williams, D., Dales, N. A., Patane, M. A. and Pantoliano, M. W.
Year: 2004
Title: ACE2 X-ray structures reveal a large hinge-bending motion important for inhibitor binding and catalysis
Journal: J Biol Chem
Volume: 279
Issue: 17
Pages: 17996-8007
Epub Date: 2004/02/03
Date: Apr 23
Short Title: ACE2 X-ray structures reveal a large hinge-bending motion important for inhibitor binding and catalysis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M311191200
PMCID: PMC7980034
Accession Number: 14754895
Keywords: Amino Acid Sequence
Amino Acids/chemistry
Angiotensin-Converting Enzyme 2
Binding Sites
Carboxypeptidases/*chemistry
Catalysis
Crystallography, X-Ray
Enzyme Inhibitors/pharmacology
Humans
Imidazoles/pharmacology
Leucine/analogs & derivatives/pharmacology
Models, Chemical
Models, Molecular
Molecular Sequence Data
Peptidyl-Dipeptidase A
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Receptors, Coronavirus
Receptors, Virus/chemistry
Sequence Homology, Amino Acid
Substrate Specificity
Zinc/chemistry
Abstract: The angiotensin-converting enzyme (ACE)-related carboxypeptidase, ACE2, is a type I integral membrane protein of 805 amino acids that contains one HEXXH + E zinc-binding consensus sequence. ACE2 has been implicated in the regulation of heart function and also as a functional receptor for the coronavirus that causes the severe acute respiratory syndrome (SARS). To gain further insights into this enzyme, the first crystal structures of the native and inhibitor-bound forms of the ACE2 extracellular domains were solved to 2.2- and 3.0-A resolution, respectively. Comparison of these structures revealed a large inhibitor-dependent hinge-bending movement of one catalytic subdomain relative to the other ( approximately 16 degrees ) that brings important residues into position for catalysis. The potent inhibitor MLN-4760 ((S,S)-2-[1-carboxy-2-[3-(3,5-dichlorobenzyl)-3H-imidazol4-yl]-ethylamino]-4-methylpentanoic acid) makes key binding interactions within the active site and offers insights regarding the action of residues involved in catalysis and substrate specificity. A few active site residue substitutions in ACE2 relative to ACE appear to eliminate the S(2)' substrate-binding subsite and account for the observed reactivity change from the peptidyl dipeptidase activity of ACE to the carboxypeptidase activity of ACE2.
Notes: 1083-351x
Towler, Paul
Staker, Bart
Prasad, Sridhar G
Menon, Saurabh
Tang, Jin
Parsons, Thomas
Ryan, Dominic
Fisher, Martin
Williams, David
Dales, Natalie A
Patane, Michael A
Pantoliano, Michael W
Journal Article
J Biol Chem. 2004 Apr 23;279(17):17996-8007. doi: 10.1074/jbc.M311191200. Epub 2004 Jan 30.
Author Address: Drug Discovery and Protein Sciences, Millennium Pharmaceuticals, Incorporated, Cambridge, Massachusetts 02139, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 489
Author: Townsend, R. R., Hilliker, E., Li, Y. T., Laine, R. A., Bell, W. R. and Lee, Y. C.
Year: 1982
Title: Carbohydrate structure of human fibrinogen. Use of 300-MHz 1H-NMR to characterize glycosidase-treated glycopeptides
Journal: J Biol Chem
Volume: 257
Issue: 16
Pages: 9704-10
Epub Date: 1982/08/25
Date: Aug 25
Short Title: Carbohydrate structure of human fibrinogen. Use of 300-MHz 1H-NMR to characterize glycosidase-treated glycopeptides
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7107587
Keywords: Carbohydrate Sequence
Carbohydrates/*analysis
Chemical Phenomena
Chemistry
Fibrinogen/*analysis
Glycopeptides/*analysis
Glycoside Hydrolases
Humans
Magnetic Resonance Spectroscopy
Methylation
Abstract: The carbohydrate composition of fibrinogen and constituent S-carboxymethylated chains was determined. All the carbohydrate on the parent molecule could be accounted for on the B beta and gamma chains. The A alpha chain was found to be devoid of carbohydrate. Glycopeptides were prepared from fibrinogen, B beta, and gamma chains by pronase digestion and subsequent chromatography on Sephadex G-50. The 300-MHz 1H-NMR spectra of glycopeptides from all three sources were consistent with biantennary type oligosaccharide chains. The glycopeptides resulting from exoglycosidase digestions were examined with 300-MHz 1H-NMR spectroscopy, and the changes in the signals caused by selective removal of sugar residues were studied. This technique enabled us to assign each anomeric proton to the corresponding monosaccharide unit independent of previous work. Our results indicate that fibrinogen glycopeptides are of the biantennary type, and are in complete agreement with the previously reported peak assignments (Vliegenthart, J. F. G., van Halbeek, H., and Dorland, L. (1981) Pure Appl. Chemn. 53, 45-77). Affinity chromatography of the glycopeptides on concanavalin A-Sepharose also showed that the glycopeptides from fibrinogen are greater than 95% biantennary oligosaccharide chains. Based on carbohydrate composition, 1H-NMR spectroscopy, sequential exoglycosidase digestion, and gas chromatography-mass spectrometry of derived partially methylated alditol acetates, we propose that fibrinogen contains four oligosaccharide chains of the structure shown below. (formula, see text)
Notes: Townsend, R R
Hilliker, E
Li, Y T
Laine, R A
Bell, W R
Lee, Y C
GM 27512/GM/NIGMS NIH HHS/United States
HL 06188/HL/NHLBI NIH HHS/United States
HL 07143/HL/NHLBI NIH HHS/United States
etc.
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1982 Aug 25;257(16):9704-10.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1494
Author: Townson, S. A., Martinez-Hackert, E., Greppi, C., Lowden, P., Sako, D., Liu, J., Ucran, J. A., Liharska, K., Underwood, K. W., Seehra, J., Kumar, R. and Grinberg, A. V.
Year: 2012
Title: Specificity and structure of a high affinity activin receptor-like kinase 1 (ALK1) signaling complex
Journal: J Biol Chem
Volume: 287
Issue: 33
Pages: 27313-25
Epub Date: 2012/06/22
Date: Aug 10
Short Title: Specificity and structure of a high affinity activin receptor-like kinase 1 (ALK1) signaling complex
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.377960
PMCID: PMC3431715
Accession Number: 22718755
Keywords: Activin Receptors, Type II/*chemistry/genetics/metabolism
Angiogenesis Inhibitors/chemistry/therapeutic use
Bone Morphogenetic Proteins/*chemistry/genetics/metabolism
Crystallography, X-Ray
Growth Differentiation Factor 2
Growth Differentiation Factors/*chemistry/genetics/metabolism
HEK293 Cells
Humans
Neovascularization, Pathologic/drug therapy/genetics/metabolism
Protein Structure, Quaternary
Protein Structure, Tertiary
Signal Transduction/drug effects/genetics
Abstract: Activin receptor-like kinase 1 (ALK1), an endothelial cell-specific type I receptor of the TGF-β superfamily, is an important regulator of normal blood vessel development as well as pathological tumor angiogenesis. As such, ALK1 is an important therapeutic target. Thus, several ALK1-directed agents are currently in clinical trials as anti-angiogenic cancer therapeutics. Given the biological and clinical importance of the ALK1 signaling pathway, we sought to elucidate the biophysical and structural basis underlying ALK1 signaling. The TGF-β family ligands BMP9 and BMP10 as well as the three type II TGF-β family receptors ActRIIA, ActRIIB, and BMPRII have been implicated in ALK1 signaling. Here, we provide a kinetic and thermodynamic analysis of BMP9 and BMP10 interactions with ALK1 and type II receptors. Our data show that BMP9 displays a significant discrimination in type II receptor binding, whereas BMP10 does not. We also report the crystal structure of a fully assembled ternary complex of BMP9 with the extracellular domains of ALK1 and ActRIIB. The structure reveals that the high specificity of ALK1 for BMP9/10 is determined by a novel orientation of ALK1 with respect to BMP9, which leads to a unique set of receptor-ligand interactions. In addition, the structure explains how BMP9 discriminates between low and high affinity type II receptors. Taken together, our findings provide structural and mechanistic insights into ALK1 signaling that could serve as a basis for novel anti-angiogenic therapies.
Notes: 1083-351x
Townson, Sharon A
Martinez-Hackert, Erik
Greppi, Chloe
Lowden, Patricia
Sako, Dianne
Liu, June
Ucran, Jeffrey A
Liharska, Katia
Underwood, Kathryn W
Seehra, Jasbir
Kumar, Ravindra
Grinberg, Asya V
Journal Article
J Biol Chem. 2012 Aug 10;287(33):27313-25. doi: 10.1074/jbc.M112.377960. Epub 2012 Jun 20.
Author Address: Acceleron Pharma, Cambridge, Massachusetts 02139, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1241
Author: Toyoda, Y., Sakurai, A., Mitani, Y., Nakashima, M., Yoshiura, K., Nakagawa, H., Sakai, Y., Ota, I., Lezhava, A., Hayashizaki, Y., Niikawa, N. and Ishikawa, T.
Year: 2009
Title: Earwax, osmidrosis, and breast cancer: why does one SNP (538G>A) in the human ABC transporter ABCC11 gene determine earwax type?
Journal: Faseb j
Volume: 23
Issue: 6
Pages: 2001-13
Epub Date: 2009/04/23
Date: Jun
Short Title: Earwax, osmidrosis, and breast cancer: why does one SNP (538G>A) in the human ABC transporter ABCC11 gene determine earwax type?
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638
DOI: 10.1096/fj.09-129098
Accession Number: 19383836
Keywords: ATP-Binding Cassette Transporters/*genetics/metabolism
Amino Acid Sequence
Animals
Antibodies/metabolism
Apocrine Glands/cytology/metabolism
Axilla/anatomy & histology
Base Sequence
Breast Neoplasms/*genetics/metabolism
Cell Line
Cerumen/*chemistry/metabolism
Ethnic Groups/genetics
Female
Genotype
Glycosylation
Humans
Molecular Sequence Data
Phenotype
*Polymorphism, Single Nucleotide
Proteasome Endopeptidase Complex/metabolism
Reproducibility of Results
Sequence Alignment
Sweat Gland Diseases/*genetics
Abstract: One single-nucleotide polymorphism (SNP), 538G>A (Gly180Arg), in the ABCC11 gene determines the type of earwax. The G/G and G/A genotypes correspond to the wet type of earwax, whereas A/A corresponds to the dry type. Wide ethnic differences exist in the frequencies of those alleles, reflecting global migratory waves of the ancestors of humankind. We herein provide the evidence that this genetic polymorphism has an effect on the N-linked glycosylation of ABCC11, intracellular sorting, and proteasomal degradation of the variant protein. Immunohistochemical studies with cerumen gland-containing tissue specimens revealed that the ABCC11 WT protein was localized in intracellular granules and large vacuoles, as well as at the luminal membrane of secretory cells in the cerumen gland, whereas granular or vacuolar localization was not detected for the SNP (Arg180) variant. This SNP variant lacking N-linked glycosylation is recognized as a misfolded protein in the endoplasmic reticulum and readily undergoes ubiquitination and proteasomal degradation, which determines the dry type of earwax as a mendelian trait with a recessive phenotype. For rapid genetic diagnosis of axillary osmidrosis and potential risk of breast cancer, we developed specific primers for the SmartAmp method that enabled us to clinically genotype the ABCC11 gene within 30 min.
Notes: 1530-6860
Toyoda, Yu
Sakurai, Aki
Mitani, Yasumasa
Nakashima, Masahiro
Yoshiura, Koh-ichiro
Nakagawa, Hiroshi
Sakai, Yasuo
Ota, Ikuko
Lezhava, Alexander
Hayashizaki, Yoshihide
Niikawa, Norio
Ishikawa, Toshihisa
Journal Article
Research Support, Non-U.S. Gov't
United States
FASEB J. 2009 Jun;23(6):2001-13. doi: 10.1096/fj.09-129098. Epub 2009 Apr 21.
Author Address: Department of Biomolecular Engineering, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259-B-60 Nagatsuta, Midori-ku, Yokohama 226-8501, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1832
Author: Traini, M., Kumaran, R., Thaysen-Andersen, M., Kockx, M., Jessup, W. and Kritharides, L.
Year: 2017
Title: N-glycosylation of human sphingomyelin phosphodiesterase acid-like 3A (SMPDL3A) is essential for stability, secretion and activity
Journal: Biochem J
Volume: 474
Issue: 7
Pages: 1071-1092
Epub Date: 2017/01/21
Date: Mar 8
Short Title: N-glycosylation of human sphingomyelin phosphodiesterase acid-like 3A (SMPDL3A) is essential for stability, secretion and activity
Alternate Journal: The Biochemical journal
ISSN: 0264-6021
DOI: 10.1042/bcj20160735
Accession Number: 28104755
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cell Line
Cricetulus
Glycosylation/drug effects
Humans
Indolizines/pharmacology
Leupeptins/pharmacology
Monocytes/cytology/drug effects/*enzymology
Mutagenesis, Site-Directed
Mutation
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/chemistry
Protease Inhibitors/pharmacology
Proteasome Endopeptidase Complex/drug effects/metabolism
Protein Sorting Signals
Protein Stability/drug effects
Recombinant Fusion Proteins/*chemistry/genetics/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Sphingomyelin Phosphodiesterase/*chemistry/genetics/metabolism
Swainsonine/pharmacology
Tunicamycin/pharmacology
*Smpdl3a
*glycoproteins
*glycosylation
*phosphodiesterase
*sphingomyelinase
Abstract: Sphingomyelin phosphodiesterase acid-like 3A (SMPDL3A) is a recently identified phosphodiesterase, which is a secreted N-linked glycoprotein. SMPDL3A is highly homologous to acid sphingomyelinase (aSMase), but unlike aSMase cannot cleave sphingomyelin. Rather, SMPDL3A hydrolyzes nucleotide tri- and diphosphates and their derivatives. While recent structural studies have shed light on these unexpected substrate preferences, many other aspects of SMPDL3A biology, which may give insight into its function in vivo, remain obscure. Here, we investigate the roles of N-glycosylation in the expression, secretion and activity of human SMPDL3A, using inhibitors of N-glycosylation and site-directed mutagenesis, with either THP-1 macrophages or CHO cells expressing human SMPDL3A. Tunicamycin (TM) treatment resulted in expression of non-glycosylated SMPDL3A that was not secreted, and was largely degraded by the proteasome. Proteasomal inhibition restored levels of SMPDL3A in TM-treated cells, although this non-glycosylated protein lacked phosphodiesterase activity. Enzymatic deglycosylation of purified recombinant SMPDL3A also resulted in significant loss of phosphodiesterase activity. Site-directed mutagenesis of individual N-glycosylation sites in SMPDL3A identified glycosylation of Asn69 and Asn222 as affecting maturation of its N-glycans and secretion. Glycosylation of Asn356 in SMPDL3A, an N-linked site conserved throughout the aSMase-like family, was critical for protection against proteasomal degradation and preservation of enzymatic activity. We provide the first experimental evidence for a predicted 22 residue N-terminal signal peptide in SMPDL3A, which is essential for facilitating glycosylation and is removed from the mature protein secreted from CHO cells. In conclusion, site-specific N-glycosylation is essential for the intracellular stability, secretion and activity of human SMPDL3A.
Notes: 1470-8728
Traini, Mathew
Kumaran, Raani
Thaysen-Andersen, Morten
Kockx, Maaike
Jessup, Wendy
Kritharides, Leonard
Journal Article
England
Biochem J. 2017 Mar 8;474(7):1071-1092. doi: 10.1042/BCJ20160735.
Author Address: Atherosclerosis Laboratory, ANZAC Research Institute, University of Sydney, NSW, Australia mathew.traini@sydney.edu.au.
Atherosclerosis Laboratory, ANZAC Research Institute, University of Sydney, NSW, Australia.
Department of Chemistry and Biomolecular Sciences, Macquarie University, NSW, Australia.
Department of Cardiology, Concord Repatriation General Hospital, Concord, NSW, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 260
Author: Treuheit, M. J., Costello, C. E. and Halsall, H. B.
Year: 1992
Title: Analysis of the five glycosylation sites of human alpha 1-acid glycoprotein
Journal: Biochem J
Volume: 283 ( Pt 1)
Issue: Pt 1
Pages: 105-12
Epub Date: 1992/04/01
Date: Apr 1
Short Title: Analysis of the five glycosylation sites of human alpha 1-acid glycoprotein
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2830105
PMCID: PMC1131000
Accession Number: 1567356
Keywords: Amino Acid Sequence
Binding Sites
Chromatography, High Pressure Liquid
Concanavalin A
Glycosylation
Hexoses/isolation & purification
Humans
Molecular Sequence Data
Orosomucoid/metabolism/*ultrastructure
Peptides/isolation & purification
Polysaccharides
Serine Endopeptidases/metabolism
Abstract: Orosomucoid (OMD) contains complex bi-, tri- and tetra-antennary glycan chains. Subfractionation of OMD into three molecular variants using concanavalin A lectin chromatography is based on variations in these complex structures. Standard h.p.l.c. profiles have been developed to analyse the percentage and distribution of the glycoforms present at each glycosylation site in OMD and its molecular variants. The ability to quantify the glycoforms present at each site allows us to extend the earlier results of others and resolve the remaining questions concerning the glycan structures of these variants. Most significantly, the proportions of bi-, tri- and tetra-antennary chains differ at each site for the three molecular variants. The most strongly retained variant from concanavalin A is uniquely capable of possessing biantennary chains at all five sites, whereas the unretained variant is completely devoid of biantennary chains. Only glycosylation site II of the five present is 100% biantennary in the retained and weakly retained variants. In addition, the two gene products of OMD were differentially glycosylated. Molecular masses of the glycoforms were verified by matrix-assisted u.v. laser desorption mass spectrometry. On the basis of the site distribution of oligosaccharides in the variants, efforts were made to understand the factors that control the processing of the carbohydrate chains in OMD. The results indicate that the 'site-directed' model of processing offers the most consistent explanation for the structures seen at the individual glycosylation sites of OMD.
Notes: 1470-8728
Treuheit, M J
Costello, C E
Halsall, H B
Journal Article
Biochem J. 1992 Apr 1;283 ( Pt 1)(Pt 1):105-12. doi: 10.1042/bj2830105.
Author Address: Department of Chemistry, University of Cincinnati, OH 45221-0172.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 916
Author: Trimble, R. B., Lubowski, C., Hauer, C. R., 3rd, Stack, R., McNaughton, L., Gemmill, T. R. and Kumar, S. A.
Year: 2004
Title: Characterization of N- and O-linked glycosylation of recombinant human bile salt-stimulated lipase secreted by Pichia pastoris
Journal: Glycobiology
Volume: 14
Issue: 3
Pages: 265-74
Epub Date: 2003/12/25
Date: Mar
Short Title: Characterization of N- and O-linked glycosylation of recombinant human bile salt-stimulated lipase secreted by Pichia pastoris
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwh036
Accession Number: 14693913
Keywords: Carbohydrate Sequence
Glycosylation
Humans
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Phosphorylation
Pichia/*metabolism
Polysaccharides/*analysis/*chemistry/metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Sterol Esterase/*chemistry/genetics/*metabolism
Sugar Alcohols/chemistry
Abstract: Recombinant human bile salt-stimulated lipase (hBSSL) was expressed in and secreted by Pichia pastoris, an organism exploited for the large-scale production of recombinant (glyco)proteins by bioprocessing technology. The 76.3-kDa glycoprotein was associated with 75-80 Man and a small amount of GlcNAc. hBSSL has one N-glycosylation site at Asn187, which was 38-40% occupied with a Man(10)GlcNAc(2) structure defined previously in Pichia as the oligosaccharide-lipid form of Man(9)GlcNAc(2) trimmed of the middle-arm terminal alpha 1,2-Man and elongated with Man alpha 1,2Man alpha 1,6-disaccharide attached to the lower-arm core alpha 1,3-Man (Trimble et al. [1991], J. Biol. Chem., 266, 22807-22817). The C-terminal 192 residues of hBSSL contain 16 Pro-rich 11-amino-acid repeats, which include 32 Ser/Thr residues as potential O-glycosylation sites. Using hBSSL as a platform to study Pichia's O-glycosylation capabilities, we found that nearly all of these sites were occupied by mannose-containing O-glycans, whose structures, after beta-elimination and purification, were assigned by (1)H NMR and, in some cases, by linkage-specific exoglycosidases and methylation analysis. The most abundant O-glycan was alpha 1,2-mannobiitol (55%), followed by alpha 1,2-mannotriitol (16%) and mannitol (10%) and a lesser amount was alpha 1,2-mannotetraitol. Unexpectedly, Man(5) and Man(6) O-glycans were present, which had the structure Man beta 1,2Man beta 1,2Man alpha 1,2(Man alpha 1,2)(1,2)mannitol. Also a small amount of a phosphorylated Man(6) O-glycan was characterized by MALDI-TOF MS postsource decay analysis as having the reducing-end mannitol disubstituted with a glycosidically linked phosphorylated Man and an unbranched Man(4) polymer elongated from a different mannitol carbon. This is the first report of the synthesis of beta-Man- and phosphate-containing O-linked constituents on glycoproteins synthesized by P. pastoris.
Notes: Trimble, Robert B
Lubowski, Catherine
Hauer, Charles R 3rd
Stack, Robert
McNaughton, Lynn
Gemmill, Trent R
Kumar, S Anand
GM23900/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 2004 Mar;14(3):265-74. doi: 10.1093/glycob/cwh036. Epub 2003 Dec 23.
Author Address: Wadsworth Center C-547, New York State Department of Health, PO Box 509, Albany, NY 12201-0509, USA. trimble@wadsworth.org
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1882
Author: Trinca, G. M., Goodman, M. L., Papachristou, E. K., D'Santos, C. S., Chalise, P., Madan, R., Slawson, C. and Hagan, C. R.
Year: 2018
Title: O-GlcNAc-Dependent Regulation of Progesterone Receptor Function in Breast Cancer
Journal: Horm Cancer
Volume: 9
Issue: 1
Pages: 12-21
Epub Date: 2017/09/21
Date: Feb
Short Title: O-GlcNAc-Dependent Regulation of Progesterone Receptor Function in Breast Cancer
Alternate Journal: Hormones & cancer
ISSN: 1868-8497 (Print)
1868-8497
DOI: 10.1007/s12672-017-0310-9
PMCID: PMC5775912
NIHMSID: NIHMS907397
Accession Number: 28929346
Keywords: Acetylglucosamine/genetics
Acylation/genetics
Breast Neoplasms/drug therapy/*genetics/pathology
Cell Line, Tumor
Cell Proliferation/genetics
Enzyme Inhibitors/therapeutic use
Female
Gene Regulatory Networks/genetics
Humans
MCF-7 Cells
N-Acetylglucosaminyltransferases/*genetics
Protein Processing, Post-Translational/*genetics
Receptors, Progesterone/*antagonists & inhibitors/genetics
Signal Transduction/drug effects
Abstract: Emerging clinical trial data implicate progestins in the development of breast cancer. While the role for the progesterone receptor (PR) in this process remains controversial, it is clear that PR, a steroid-activated nuclear receptor, alters the transcriptional landscape of breast cancer. PR interacts with many different types of proteins, including transcriptional co-activators and co-repressors, transcription factors, nuclear receptors, and proteins that post-translationally modify PR (i.e., kinases and phosphatases). Herein, we identify a novel interaction between PR and O-GlcNAc transferase (OGT), the enzyme that catalyzes the addition of a single N-acetylglucosamine sugar, referred to as O-GlcNAc, to acceptor serines and threonines in target proteins. This interaction between PR and OGT leads to the post-translational modification of PR by O-GlcNAc. Moreover, we show that O-GlcNAcylated PR is more transcriptionally active on PR-target genes, despite the observation that PR messenger RNA and protein levels are decreased when O-GlcNAc levels are high. O-GlcNAcylation in breast cancer is clinically relevant, as we show that O-GlcNAc levels are higher in breast cancer as compared to matched normal tissues, and PR-positive breast cancers have higher levels of OGT. These data predict that under conditions where O-GlcNAc levels are high (breast cancer), PR, through an interaction with the modifying enzyme OGT, will exhibit increased O-GlcNAcylation and potentiated transcriptional activity. Therapeutic strategies aimed at altering cellular O-GlcNAc levels may have profound effects on PR transcriptional activity in breast cancer.
Notes: 1868-8500
Trinca, Gloria M
Goodman, Merit L
Papachristou, Evangelia K
D'Santos, Clive S
Chalise, Prabhakar
Madan, Rashna
Slawson, Chad
Hagan, Christy R
R01DK100595/DK/NIDDK NIH HHS/United States
R00CA166643/CA/NCI NIH HHS/United States
V2015-025/V Foundation for Cancer Research/International
CCR16376147/KOMEN/Susan G. Komen/United States
W81XWH-16-1-0320/Congressionally Directed Medical Research Programs/International
R00 CA166643/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Horm Cancer. 2018 Feb;9(1):12-21. doi: 10.1007/s12672-017-0310-9. Epub 2017 Sep 19.
Author Address: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, MS3030, 3901 Rainbow Blvd, Kansas City, KS, 66160, USA.
Cancer Research UK Cambridge Institute, University of Cambridge, Cambridge, UK.
Department of Biostatistics, University of Kansas Medical Center, Kansas City, KS, 66160, USA.
Division of Hematology/Oncology, Department of Pathology, University of Kansas Medical Center, Kansas City, KS, 66160, USA.
Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, MS3030, 3901 Rainbow Blvd, Kansas City, KS, 66160, USA. chagan@kumc.edu.
Department of Cancer Biology, and University of Kansas Cancer Center, University of Kansas Medical Center, Kansas City, KS, 66160, USA. chagan@kumc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1411
Author: Tsachaki, M., Serlidaki, D., Fetani, A., Zarkou, V., Rozani, I., Ghiso, J. and Efthimiopoulos, S.
Year: 2011
Title: Glycosylation of BRI2 on asparagine 170 is involved in its trafficking to the cell surface but not in its processing by furin or ADAM10
Journal: Glycobiology
Volume: 21
Issue: 10
Pages: 1382-8
Epub Date: 2011/07/15
Date: Oct
Short Title: Glycosylation of BRI2 on asparagine 170 is involved in its trafficking to the cell surface but not in its processing by furin or ADAM10
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwr097
PMCID: PMC3167477
Accession Number: 21752865
Keywords: ADAM Proteins/genetics/*metabolism
ADAM10 Protein
Adaptor Proteins, Signal Transducing
Amyloid Precursor Protein Secretases/genetics/*metabolism
Asparagine/*genetics/metabolism
Cell Membrane/metabolism
Furin/*metabolism
Glycosylation
HEK293 Cells
Humans
Membrane Glycoproteins
Membrane Proteins/genetics/*metabolism
Protein Processing, Post-Translational
Transfection
Abstract: Two different mutated forms of BRI2 protein are linked with familial British and Danish dementias, which present neuropathological similarities with Alzheimer's disease. BRI2 is a type II transmembrane protein that is trafficked through the secretory pathway to the cell surface and is processed by furin and ADAM10 (a disintegrin and metalloproteinase domain 10) to release secreted fragments of unknown function. Its apparent molecular mass (42-44 kDa) is significantly higher than that predicted by the number and composition of amino acids (30 kDa) suggesting that BRI2 is glycosylated. In support, bioinformatics analysis indicated that BRI2 bears the consensus sequence Asn-Thr-Ser (residues 170-173) and could be N-glycosylated at Asn170. Given that N-glycosylation is considered essential for protein folding, processing and trafficking, we examined whether BRI2 is N-glycosylated. Treatment of HEK293 (human embryonic kidney) cells expressing BRI2 with the N-glycosylation inhibitor tunicamycin or mutation of Asn170 to alanine reduced its molecular mass by ~2 kDa. These data indicate that BRI2 is N-glycosylated at Asn170. To examine the effect of N-glycosylation on BRI2 trafficking at the cell surface, we performed biotinylation and (35)S methionine pulse-chase experiments. These experiments showed that mutation of Asn170 to alanine reduced BRI2 trafficking at the cell surface and its steady state levels at the plasma membrane. Furthermore, we obtained data indicating that this mutation did not affect cleavage of BRI2 by furin or ADAM10. Our results confirm the theoretical predictions that BRI2 is N-glycosylated at Asn170 and show that this post-translational modification is essential for its expression at the cell surface but not for its proteolytic processing.
Notes: 1460-2423
Tsachaki, Maria
Serlidaki, Despina
Fetani, Andriana
Zarkou, Vasiliki
Rozani, Ismini
Ghiso, Jorge
Efthimiopoulos, Spiros
AG030539/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Glycobiology. 2011 Oct;21(10):1382-8. doi: 10.1093/glycob/cwr097. Epub 2011 Jul 13.
Author Address: Division of Animal and Human Physiology, Department of Biology, National and Kapodistrian University of Athens, 157 84 Panepistimiopolis, Ilisia, Athens, Greece.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 378
Author: Tsuda, E., Goto, M., Murakami, A., Akai, K., Ueda, M., Kawanishi, G., Takahashi, N., Sasaki, R., Chiba, H., Ishihara, H. and et al.
Year: 1988
Title: Comparative structural study of N-linked oligosaccharides of urinary and recombinant erythropoietins
Journal: Biochemistry
Volume: 27
Issue: 15
Pages: 5646-54
Epub Date: 1988/07/26
Date: Jul 26
Short Title: Comparative structural study of N-linked oligosaccharides of urinary and recombinant erythropoietins
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00415a038
Accession Number: 3179269
Keywords: Carbohydrate Sequence
Chromatography, High Pressure Liquid
Erythropoietin/*analysis/urine
Glycoproteins/*analysis
Magnetic Resonance Spectroscopy
Mass Spectrometry
Molecular Sequence Data
Oligosaccharides/*analysis
Recombinant Proteins/analysis
Abstract: The structures of the N-linked oligosaccharides of the urinary erythropoietin (u-EPO) purified from urine of aplastic anemic patients were analyzed and compared with those for recombinant erythropoietin (r-EPO) prepared with baby hamster kidney (BHK) cells. Asparagine-linked neutral oligosaccharides were released from each EPO protein by N-oligosaccharide glycopeptidase (almond) digestion. The reducing ends of the oligosaccharide chains thus obtained were aminated with a fluorescent reagent, 2-aminopyridine, and the mixture of pyridylamino derivatives of the oligosaccharides was separated by high-performance liquid chromatography (HPLC) on an ODS silica column. More than 8 and 13 kinds of oligosaccharide fractions for u-EPO and r-EPO (BHK), respectively, were completely separated by the one-step HPLC procedure. The structure of each oligosaccharide thus isolated was analyzed by a combination of sequential exoglycosidase digestion and another kind of HPLC with an amide-silica column. Furthermore, high-resolution proton nuclear magnetic resonance (1H NMR) spectroscopy and methylation analyses were carried out in the case of r-EPO (BHK).(ABSTRACT TRUNCATED AT 250 WORDS)
Notes: Tsuda, E
Goto, M
Murakami, A
Akai, K
Ueda, M
Kawanishi, G
Takahashi, N
Sasaki, R
Chiba, H
Ishihara, H
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1988 Jul 26;27(15):5646-54. doi: 10.1021/bi00415a038.
Author Address: Research Institute of Life Science, Snow Brand Milk Products Company Ltd., Tochigi, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 412
Author: Tsuji, T. and Osawa, T.
Year: 1987
Title: The carbohydrate moiety of human platelet glycocalicin: the structures of the major Asn-linked sugar chains
Journal: J Biochem
Volume: 101
Issue: 1
Pages: 241-9
Epub Date: 1987/01/01
Date: Jan
Short Title: The carbohydrate moiety of human platelet glycocalicin: the structures of the major Asn-linked sugar chains
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a121897
Accession Number: 3571205
Keywords: Amino Acids/analysis
Asparagine/analysis
Blood Platelets/*analysis
Carbohydrates/*analysis
Chromatography, Affinity
Chromatography, Liquid
Electrophoresis, Polyacrylamide Gel
Humans
Oligosaccharides/analysis
Platelet Aggregation
*Platelet Glycoprotein GPIb-IX Complex
Platelet Membrane Glycoproteins/*blood/isolation & purification
Protein Denaturation
Abstract: Glycocalicin, a predominant glycoprotein on the human platelet surface, has been purified from a platelet suspension by means of sonication, ammonium sulfate precipitation and acid treatment followed by chromatography on columns of wheat germ agglutinin-Sepharose and Mono Q. Asparagine-linked (N-linked) oligosaccharides were released by hydrazinolysis, and then N-acetylated and reduced with NaBH4 or NaB3H4. The released carbohydrate chains were found to be of the complex-type from their interaction with immobilized lectin columns. The structures of the two major oligosaccharide-alditols separated by ion-exchange chromatography on a Mono Q column were investigated by means of methylation analysis, glycosidase digestion, and Smith periodate degradation, and they were assigned as typical di- and trisialylated complex-type oligosaccharide-alditols with two and three peripheral chains consisting of Gal-GlcNAc sequences, respectively.
Notes: Tsuji, T
Osawa, T
Journal Article
England
J Biochem. 1987 Jan;101(1):241-9. doi: 10.1093/oxfordjournals.jbchem.a121897.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 463
Author: Tsuji, T., Tsunehisa, S., Watanabe, Y., Yamamoto, K., Tohyama, H. and Osawa, T.
Year: 1983
Title: The carbohydrate moiety of human platelet glycocalicin
Journal: J Biol Chem
Volume: 258
Issue: 10
Pages: 6335-9
Epub Date: 1983/05/25
Date: May 25
Short Title: The carbohydrate moiety of human platelet glycocalicin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 6406478
Keywords: Blood Platelets/*analysis
Carbohydrate Conformation
Carbohydrates/analysis
Chromatography
Glycoproteins/*blood
Glycoside Hydrolases
Humans
Membrane Proteins/*blood
Methylation
Molecular Weight
Oligosaccharides/isolation & purification
*Platelet Glycoprotein GPIb-IX Complex
Serine
Threonine
Abstract: Glycocalicin, a predominant glycoprotein on the human platelet surface, has been purified from a platelet suspension by sonication, ammonium sulfate precipitation and acid treatment followed by chromatography on columns of wheat germ agglutinin-Sepharose and Sephacryl S-300. Ser/Thr-linked (O-linked) oligosaccharides were released by alkaline borohydride treatment, and fractionated by high performance liquid chromatography with an anion-exchange resin. The structure of a major oligosaccharide alditol separated by high performance liquid chromatography was investigated by a combination of compositional analyses, methylation and glycosidase treatments, and proposed to be a hexasaccharide alditol, NeuAc alpha 2-3Gal beta 1-4GlcNAc beta 1-6(NeuAc alpha 2-3Gal beta 1-3) N-acetylgalactosaminitol. We also found some sugar units which appeared to be intermediates in the biosynthetic pathway of the major hexasaccharide.
Notes: Tsuji, T
Tsunehisa, S
Watanabe, Y
Yamamoto, K
Tohyama, H
Osawa, T
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1983 May 25;258(10):6335-9.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1675
Author: Tsutsumi, N., Kimura, T., Arita, K., Ariyoshi, M., Ohnishi, H., Yamamoto, T., Zuo, X., Maenaka, K., Park, E. Y., Kondo, N., Shirakawa, M., Tochio, H. and Kato, Z.
Year: 2014
Title: The structural basis for receptor recognition of human interleukin-18
Journal: Nat Commun
Volume: 5
Pages: 5340
Epub Date: 2014/12/17
Date: Dec 15
Short Title: The structural basis for receptor recognition of human interleukin-18
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms6340
PMCID: PMC4275594
Accession Number: 25500532
Keywords: Amino Acid Sequence
Animals
Baculoviridae/genetics
Binding Sites
Crystallography, X-Ray
Gene Expression
Humans
Interleukin-1 Receptor Accessory Protein/chemistry/genetics
Interleukin-18/*chemistry/genetics
Interleukin-1beta/*chemistry/genetics
Molecular Sequence Data
Mutation
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Protein Subunits/*chemistry/genetics
Receptors, Interleukin/chemistry/genetics
Receptors, Interleukin-18/*chemistry/genetics
Recombinant Proteins/chemistry/genetics
Sequence Homology, Amino Acid
Sf9 Cells
Spodoptera
Abstract: Interleukin (IL)-18 is a proinflammatory cytokine that belongs to the IL-1 family and plays an important role in inflammation. The uncontrolled release of this cytokine is associated with severe chronic inflammatory disease. IL-18 forms a signalling complex with the IL-18 receptor α (Rα) and β (Rβ) chains at the plasma membrane, which induces multiple inflammatory cytokines. Here, we present a crystal structure of human IL-18 bound to the two receptor extracellular domains. Generally, the receptors' recognition mode for IL-18 is similar to IL-1β; however, certain notable differences were observed. The architecture of the IL-18 receptor second domain (D2) is unique among the other IL-1R family members, which presumably distinguishes them from the IL-1 receptors that exhibit a more promiscuous ligand recognition mode. The structures and associated biochemical and cellular data should aid in developing novel drugs to neutralize IL-18 activity.
Notes: 2041-1723
Tsutsumi, Naotaka
Kimura, Takeshi
Arita, Kyohei
Ariyoshi, Mariko
Ohnishi, Hidenori
Yamamoto, Takahiro
Zuo, Xiaobing
Maenaka, Katsumi
Park, Enoch Y
Kondo, Naomi
Shirakawa, Masahiro
Tochio, Hidehito
Kato, Zenichiro
P41 GM103311/GM/NIGMS NIH HHS/United States
NIGMS P41-GM103311/PHS HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Commun. 2014 Dec 15;5:5340. doi: 10.1038/ncomms6340.
Author Address: Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan.
Department of Pediatrics, Graduate School of Medicine, Gifu University, Yanagido 1-1, Gifu 501-1194, Japan.
Graduate School of Nanobioscience, Yokohama City University, 1-7-29 Suehiro-cho, Tsurumi-ku, Yokohama Kanagawa 230-0045, Japan.
1] Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan [2] Institute for Integrated Cell-Material Sciences, Kyoto University, Kyoto 606-8501, Japan.
X-Ray Science Division, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, Illinois 60439, USA.
Laboratory of Biomolecular Science and Center for Research and Education on Drug Discovery, Faculty of Pharmaceutical Sciences, Hokkaido University, , Kita-12, Nishi-6, Kita-ki, Sapporo 060-0812, Japan.
Research Institute of Green Science and Technology, Department of Bioscience, Graduate school of Science and Technology, Shizuoka University, 836 Ohya Suruga-ku, Shizuoka 422-8529, Japan.
1] Department of Pediatrics, Graduate School of Medicine, Gifu University, Yanagido 1-1, Gifu 501-1194, Japan [2] Heisei College of Health Sciences, 180 Kurono, Gifu 501-1131, Japan.
1] Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan [2] Core Research of Evolution Science (CREST), Japan Sciences and Technology Agency, Tokyo 102-0076, Japan.
Department of Biophysics, Graduate School of Science, Kyoto University, Kitashirakawa-oiwake, Sakyo-ku, Kyoto 606-8502, Japan.
1] Department of Pediatrics, Graduate School of Medicine, Gifu University, Yanagido 1-1, Gifu 501-1194, Japan [2] Biomedical Informatics, Medical Information Sciences Division, The United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, Gifu 501-1194, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 826
Author: Turk, D., Janjić, V., Stern, I., Podobnik, M., Lamba, D., Dahl, S. W., Lauritzen, C., Pedersen, J., Turk, V. and Turk, B.
Year: 2001
Title: Structure of human dipeptidyl peptidase I (cathepsin C): exclusion domain added to an endopeptidase framework creates the machine for activation of granular serine proteases
Journal: Embo j
Volume: 20
Issue: 23
Pages: 6570-82
Epub Date: 2001/12/01
Date: Dec 3
Short Title: Structure of human dipeptidyl peptidase I (cathepsin C): exclusion domain added to an endopeptidase framework creates the machine for activation of granular serine proteases
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/20.23.6570
PMCID: PMC125750
Accession Number: 11726493
Keywords: Amino Acid Sequence
Animals
Baculoviridae/metabolism
Binding Sites
Cathepsin C/*chemistry/*genetics
Cell Line
Dimerization
Endopeptidases/*chemistry
Humans
Insecta
Models, Molecular
Molecular Sequence Data
Mutation, Missense
Papillon-Lefevre Disease/genetics/metabolism
Protein Conformation
Protein Folding
Protein Structure, Tertiary
Recombinant Proteins/chemistry
Sequence Homology, Amino Acid
Serine Endopeptidases/*chemistry
Substrate Specificity
Syndrome
Abstract: Dipeptidyl peptidase I (DPPI) or cathepsin C is the physiological activator of groups of serine proteases from immune and inflammatory cells vital for defense of an organism. The structure presented shows how an additional domain transforms the framework of a papain-like endopeptidase into a robust oligomeric protease-processing enzyme. The tetrahedral arrangement of the active sites exposed to solvent allows approach of proteins in their native state; the massive body of the exclusion domain fastened within the tetrahedral framework excludes approach of a polypeptide chain apart from its termini; and the carboxylic group of Asp1 positions the N-terminal amino group of the substrate. Based on a structural comparison and interactions within the active site cleft, it is suggested that the exclusion domain originates from a metallo-protease inhibitor. The location of missense mutations, characterized in people suffering from Haim-Munk and Papillon-Lefevre syndromes, suggests how they disrupt the fold and function of the enzyme.
Notes: 1460-2075
Turk, D
Janjić, V
Stern, I
Podobnik, M
Lamba, D
Dahl, S W
Lauritzen, C
Pedersen, J
Turk, V
Turk, B
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2001 Dec 3;20(23):6570-82. doi: 10.1093/emboj/20.23.6570.
Author Address: Department of Biochemistry and Molecular Biology, Jozef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia. Dusan.Turk@ijs.si
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 600
Author: Turk, E., Kerner, C. J., Lostao, M. P. and Wright, E. M.
Year: 1996
Title: Membrane topology of the human Na+/glucose cotransporter SGLT1
Journal: J Biol Chem
Volume: 271
Issue: 4
Pages: 1925-34
Epub Date: 1996/01/26
Date: Jan 26
Short Title: Membrane topology of the human Na+/glucose cotransporter SGLT1
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.4.1925
Accession Number: 8567640
Keywords: Amino Acid Sequence
Animals
Biological Transport
Glycosylation
Humans
Membrane Glycoproteins/*chemistry
Molecular Sequence Data
Monosaccharide Transport Proteins/*chemistry
Mutagenesis, Site-Directed
Oocytes
Protein Structure, Secondary
Sodium-Glucose Transporter 1
Structure-Activity Relationship
Xenopus laevis
Abstract: The membrane topology of the human Na+/glucose cotransporter SGLT1 has been probed using N-glycosylation scanning mutants and nested truncations. Functional analysis proved essential for establishment of signal-anchor topology. The resultant model diverges significantly from previously held suppositions of structure based primarily on hydropathy analysis. SGLT1 incorporates 14 membrane spans. The N terminus resides extracellularly, and two hydrophobic regions form newly recognized membrane spans 4 and 12; the large charged domain near the C terminus is cytoplasmic. This model was evaluated further using two advanced empirically-based algorithms predictive of transmembrane helices. Helix ends were predicted using thermo-dynamically-based algorithms known to predict x-ray crystallographically determined transmembrane helix ends. Several considerations suggest the hydrophobic C terminus forms a 14th transmembrane helix, differentiating the eukaryotic members of the SGLT1 family from bacterial homologues. Our data inferentially indicate that these bacterial homologues incorporate 13 spans, with an extracellular N terminus. The model of SGLT1 secondary structure and the predicted helix ends signify information prerequisite for the rational design of further experiments on structure/function relationships.
Notes: Turk, E
Kerner, C J
Lostao, M P
Wright, E M
DK-44582/DK/NIDDK NIH HHS/United States
DK-44602/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1996 Jan 26;271(4):1925-34. doi: 10.1074/jbc.271.4.1925.
Author Address: Department of Physiology, UCLA School of Medicine 90095-1751, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 653
Author: Tvrdik, P., Asadi, A., Kozak, L. P., Nedergaard, J., Cannon, B. and Jacobsson, A.
Year: 1997
Title: Cig30, a mouse member of a novel membrane protein gene family, is involved in the recruitment of brown adipose tissue
Journal: J Biol Chem
Volume: 272
Issue: 50
Pages: 31738-46
Epub Date: 1998/02/12
Date: Dec 12
Short Title: Cig30, a mouse member of a novel membrane protein gene family, is involved in the recruitment of brown adipose tissue
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.50.31738
Accession Number: 9395518
Keywords: Acetyltransferases
Adipose Tissue, Brown/*cytology/metabolism
Amino Acid Sequence
Animals
Base Sequence
Body Temperature Regulation/physiology
Cell Division
Cloning, Molecular
Dexamethasone/pharmacology
Fatty Acid Elongases
Glucose Transporter Type 4
Glycoproteins/analysis
Male
Membrane Proteins/chemistry/genetics/*physiology
Mice
Models, Chemical
Molecular Sequence Data
Monosaccharide Transport Proteins/metabolism
*Muscle Proteins
Norepinephrine/pharmacology
Protein Structure, Secondary
RNA, Messenger/metabolism
Transfection
Abstract: We have identified a previously uncharacterized gene that is implicated in the thermogenic function of brown adipose tissue of mice. This gene, termed Cig30, is the first mammalian member of a novel gene family comprising several nematode and yeast genes, such as SUR4 and FEN1, mutation of which is associated with highly pleiotropic phenotypes. It codes for a 30-kDa plasma membrane glycoprotein with five putative transmembrane domains. The Cig30 mRNA was readily detected only in brown fat and liver. When animals were exposed to a 3-day cold stress, the Cig30 expression was selectively elevated in brown fat more than 200-fold. Similar increases were brought about in two other conditions of brown fat recruitment, namely during perinatal development and after cafeteria diet. The magnitude of Cig30 mRNA induction in the cold could be mimicked by chronic norepinephrine treatment in vivo. However, in primary cultures of brown adipocytes, a synergistic action of norepinephrine and dexamethasone was required for full expression of the gene, indicating that both catecholamines and glucocorticoids are required for the induction of Cig30. We propose that the CIG30 protein is involved in a pathway connected with brown fat hyperplasia.
Notes: Tvrdik, P
Asadi, A
Kozak, L P
Nedergaard, J
Cannon, B
Jacobsson, A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1997 Dec 12;272(50):31738-46. doi: 10.1074/jbc.272.50.31738.
Author Address: The Wenner-Gren Institute, The Arrhenius Laboratories F3, Stockholm University, S-106 91 Stockholm, Sweden. petr.tvrdik@zoofys.su.se
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1688
Author: Tykvart, J., Bařinka, C., Svoboda, M., Navrátil, V., Souček, R., Hubálek, M., Hradilek, M., Šácha, P., Lubkowski, J. and Konvalinka, J.
Year: 2015
Title: Structural and biochemical characterization of a novel aminopeptidase from human intestine
Journal: J Biol Chem
Volume: 290
Issue: 18
Pages: 11321-36
Epub Date: 2015/03/11
Date: May 1
Short Title: Structural and biochemical characterization of a novel aminopeptidase from human intestine
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M114.628149
PMCID: PMC4416838
Accession Number: 25752612
Keywords: Amino Acid Sequence
Animals
Crystallography, X-Ray
Dipeptidyl Peptidase 4/metabolism
Endopeptidases/metabolism
Gene Expression Regulation, Enzymologic
Glutamate Carboxypeptidase II/*chemistry/genetics/*metabolism
Humans
Intestines/*enzymology
Models, Molecular
Molecular Sequence Data
Protein Structure, Tertiary
Rats
Aminopeptidase
DPP IV Activity
Human Ileal Aminopeptidase
Intestinal Metabolism
Metalloprotease
Molecular Evolution
Pics
Protein Degradation
X-ray Crystallography
Abstract: N-acetylated α-linked acidic dipeptidase-like protein (NAALADase L), encoded by the NAALADL1 gene, is a close homolog of glutamate carboxypeptidase II, a metallopeptidase that has been intensively studied as a target for imaging and therapy of solid malignancies and neuropathologies. However, neither the physiological functions nor structural features of NAALADase L are known at present. Here, we report a thorough characterization of the protein product of the human NAALADL1 gene, including heterologous overexpression and purification, structural and biochemical characterization, and analysis of its expression profile. By solving the NAALADase L x-ray structure, we provide the first experimental evidence that it is a zinc-dependent metallopeptidase with a catalytic mechanism similar to that of glutamate carboxypeptidase II yet distinct substrate specificity. A proteome-based assay revealed that the NAALADL1 gene product possesses previously unrecognized aminopeptidase activity but no carboxy- or endopeptidase activity. These findings were corroborated by site-directed mutagenesis and identification of bestatin as a potent inhibitor of the enzyme. Analysis of NAALADL1 gene expression at both the mRNA and protein levels revealed the small intestine as the major site of protein expression and points toward extensive alternative splicing of the NAALADL1 gene transcript. Taken together, our data imply that the NAALADL1 gene product's primary physiological function is associated with the final stages of protein/peptide digestion and absorption in the human digestive system. Based on these results, we suggest a new name for this enzyme: human ileal aminopeptidase (HILAP).
Notes: 1083-351x
Tykvart, Jan
Bařinka, Cyril
Svoboda, Michal
Navrátil, Václav
Souček, Radko
Hubálek, Martin
Hradilek, Martin
Šácha, Pavel
Lubkowski, Jacek
Konvalinka, Jan
Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2015 May 1;290(18):11321-36. doi: 10.1074/jbc.M114.628149. Epub 2015 Mar 9.
Author Address: From the Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo n. 2, Prague 6, Czech Republic, the Departments of Biochemistry and.
the Institute of Biotechnology, Academy of Sciences of the Czech Republic, Vídeňská 1083, Prague 4, Czech Republic, and.
From the Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo n. 2, Prague 6, Czech Republic, Physical and Macromolecular Chemistry, Faculty of Natural Science, Charles University, Albertov 6, Prague 2, Czech Republic.
From the Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo n. 2, Prague 6, Czech Republic.
the Center for Cancer Research, Macromolecular Crystallography Laboratory, NCI, National Institutes of Health, Frederick, Maryland 21702-1201.
From the Gilead Sciences and IOCB Research Centre, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo n. 2, Prague 6, Czech Republic, the Departments of Biochemistry and jan.konvalinka@uochb.cas.cz.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1912
Author: Udagawa, T., Harita, Y., Miura, K., Mitsui, J., Ode, K. L., Morishita, S., Urae, S., Kanda, S., Kajiho, Y., Tsurumi, H., Ueda, H. R., Tsuji, S., Saito, A. and Oka, A.
Year: 2018
Title: Amnionless-mediated glycosylation is crucial for cell surface targeting of cubilin in renal and intestinal cells
Journal: Sci Rep
Volume: 8
Issue: 1
Pages: 2351
Epub Date: 2018/02/07
Date: Feb 5
Short Title: Amnionless-mediated glycosylation is crucial for cell surface targeting of cubilin in renal and intestinal cells
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/s41598-018-20731-4
PMCID: PMC5799345
Accession Number: 29402915
Keywords: Cell Membrane/metabolism
Cells, Cultured
Endoplasmic Reticulum/metabolism
Glycosylation
Humans
Intestinal Mucosa/metabolism
Kidney/metabolism
Membrane Proteins
Mutation, Missense
*Protein Transport
Proteins/genetics/*metabolism
Receptors, Cell Surface/genetics/*metabolism
Abstract: Mutations in either cubilin (CUBN) or amnionless (AMN) genes cause Imerslund-Gräsbeck syndrome (IGS), a hereditary disease characterised by anaemia attributed to selective intestinal malabsorption of cobalamin and low-molecular weight proteinuria. Although cubilin protein does not have a transmembrane segment, it functions as a multi-ligand receptor by binding to the transmembrane protein, amnionless. We established a system to quantitatively analyse membrane targeting of the protein complex in cultured renal and intestinal cells and analysed the pathogenic mechanisms of mutations found in IGS patients. A novel CUBN mutation, several previously reported CUBN missense mutations and all previously reported AMN missense mutations resulted in endoplasmic reticulum (ER) retention and completely inhibited amnionless-dependent plasma membrane expression of cubilin. The ER retention of cubilin and amnionless was confirmed in renal proximal tubular cells of a patient with IGS. Notably, the interaction between cubilin and amnionless was not sufficient, but amnionless-mediated glycosylation of cubilin was necessary for their surface expression. Quantitative mass spectrometry and mutagenesis demonstrated that N-linked glycosylation of at least 4 residues of cubilin protein was required for its surface targeting. These results delineated the molecular mechanisms of membrane trafficking of cubilin in renal and intestinal cells.
Notes: 2045-2322
Udagawa, Tomohiro
Harita, Yutaka
Miura, Kenichiro
Mitsui, Jun
Ode, Koji L
Morishita, Shinichi
Urae, Seiya
Kanda, Shoichiro
Kajiho, Yuko
Tsurumi, Haruko
Ueda, Hiroki R
Tsuji, Shoji
Orcid: 0000-0001-5602-5686
Saito, Akihiko
Oka, Akira
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2018 Feb 5;8(1):2351. doi: 10.1038/s41598-018-20731-4.
Author Address: Department of Pediatrics, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8655, Japan.
Department of Pediatrics and Developmental Biology, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8519, Japan.
Department of Pediatrics, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8655, Japan. haritay-ped@h.u-tokyo.ac.jp.
Department of Neurology, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8655, Japan.
Department of Systems Pharmacology, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8655, Japan.
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, 5-1-5, Kashiwanoha, Kashiwa-shi, Chiba, 277-8561, Japan.
Department of Applied Molecular Medicine, Niigata University Graduate School of Medical and Dental Sciences, 1-757, Asahimachidori, Chuo-ku, Niigata-shi, Niigata, 951-8510, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2044
Author: Ueda, Y., Moriwaki, K., Takeuchi, T., Higuchi, K. and Asahi, M.
Year: 2020
Title: O-GlcNAcylation-mediated degradation of FBXL2 stabilizes FOXM1 to induce cancer progression
Journal: Biochem Biophys Res Commun
Volume: 521
Issue: 3
Pages: 632-638
Epub Date: 2019/11/05
Date: Jan 15
Short Title: O-GlcNAcylation-mediated degradation of FBXL2 stabilizes FOXM1 to induce cancer progression
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2019.10.164
Accession Number: 31679690
Keywords: Acetylglucosamine/*metabolism
Acylation
Cell Line, Tumor
Cell Proliferation
Disease Progression
F-Box Proteins/*metabolism
Forkhead Box Protein M1/*metabolism
Humans
Protein Stability
Proteolysis
Stomach Neoplasms/*metabolism/pathology
Ubiquitination
*Cancer progression
*Fbxl2
*Foxm1
*O-GlcNAcylation
*Ubiquitination
Abstract: O-GlcNAcylation is a dynamic and reversible post-translational modification of cytonuclear molecules that regulates cellular signaling. Elevated O-GlcNAcylation is a general property of cancer and plays a critical role in cancer progression. We previously showed that the expression of FOXM1, a critical oncogenic transcription factor widely overexpressed in solid tumors, was elevated in MKN45 cells, a human gastric cancer cell line, by the O-GlcNAcase inhibitor Thiamet G (TMG), which induces augmented O-GlcNAcylation. Here, we identified FBXL2 E3 ubiquitin ligase as a new target of O-GlcNAcylation. Consistent with the results in MKN45 cells, FOXM1 expression was increased, accompanied by its decreased ubiquitination and degradation by TMG in the other gastric cancer cell lines, including NUGC-3 cells. We found that FBXL2 ubiquitinated FOXM1, and the interaction with FBXL2 and ubiquitination of FOXM1 were reduced by TMG in NUGC-3 cells. Interestingly, FBXL2 was also ubiquitinated, which was promoted by TMG in the cells. Moreover, FOXM1 expression and cell proliferation were reduced in FBXL2-induced NUGC-3 cells, and the reductions were attenuated by TMG, indicating that FOXM1 was stabilized by O-GlcNAcylation-mediated degradation of FBXL2 to induce cancer progression. These data suggest that elevated O-GlcNAcylation contributes to cancer progression by suppressing FBXL2-mediated degradation of FOXM1.
Notes: 1090-2104
Ueda, Yasuhiro
Moriwaki, Kazumasa
Takeuchi, Toshihisa
Higuchi, Kazuhide
Asahi, Michio
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2020 Jan 15;521(3):632-638. doi: 10.1016/j.bbrc.2019.10.164. Epub 2019 Oct 31.
Author Address: Second Department of Internal Medicine, Faculty of Medicine, Osaka Medical College, Osaka, 569-8686, Japan.
Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka, 569-8686, Japan.
Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka, 569-8686, Japan. Electronic address: masahi@osaka-med.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 495
Author: Uehara, H., Coligan, J. E. and Nathenson, S. G.
Year: 1981
Title: Amino acid sequence of the carboxyl-terminal hydrophilic region of the H-2Kb MHC alloantigen. Completion of the entire primary structure of the H-2Kb molecule
Journal: Biochemistry
Volume: 20
Issue: 21
Pages: 5940-5
Epub Date: 1981/10/13
Date: Oct 13
Short Title: Amino acid sequence of the carboxyl-terminal hydrophilic region of the H-2Kb MHC alloantigen. Completion of the entire primary structure of the H-2Kb molecule
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00524a003
Accession Number: 7306483
Keywords: Amino Acid Sequence
Animals
Cyanogen Bromide
H-2 Antigens/*genetics
*Isoantigens
*Major Histocompatibility Complex
Mice
Peptide Fragments/analysis
Protein Conformation
Trypsin
Abstract: The amino acid sequence of the COOH-terminal hydrophilic region of the H-2Kb histocompatibility antigen was determined. The sequence was completed by analyses of four CNBr fragments obtained from the intact molecule as well as tryptic peptides. This region was composed of 39 amino acid residues with a cluster of basic residues at the NH2 terminus and localized positions 308-346 of the H-2Kb molecule. These sequence data, together with those reported for the NH2-terminal 284 residues [Martinko, J. M., Uehara, H., Ewenstein, B. M., Kindt, T. J., Coligan, J. E., & Nathenson, S. G. (1980) Biochemistry 19, 6188-6193] and for the intramembranous segment [Uehara, H., Coligan, J. E., & Nathenson, S. G. (1981) Biochemistry (preceding paper in this issue)], provided the complete primary structure of the H-2Kb molecule. This is the first histocompatibility antigen for which the entire primary structure is determined.
Notes: Uehara, H
Coligan, J E
Nathenson, S G
AI-07289/AI/NIAID NIH HHS/United States
AI-10702/AI/NIAID NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1981 Oct 13;20(21):5940-5. doi: 10.1021/bi00524a003.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1010
Author: Uematsu, R., Furukawa, J., Nakagawa, H., Shinohara, Y., Deguchi, K., Monde, K. and Nishimura, S.
Year: 2005
Title: High throughput quantitative glycomics and glycoform-focused proteomics of murine dermis and epidermis
Journal: Mol Cell Proteomics
Volume: 4
Issue: 12
Pages: 1977-89
Epub Date: 2005/09/20
Date: Dec
Short Title: High throughput quantitative glycomics and glycoform-focused proteomics of murine dermis and epidermis
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M500203-MCP200
Accession Number: 16170054
Keywords: Amino Acid Sequence
Animals
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Databases, Protein
Epidermis/*chemistry
Glycopeptides/*chemistry/*isolation & purification
Glycoproteins/*chemistry
Glycosylation
Mice
Mice, Hairless
Molecular Sequence Data
Polysaccharides/chemistry/isolation & purification
Proteome/*chemistry
Proteomics/*methods
Skin/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Abstract: Despite recent advances in our understanding of the significance of the protein glycosylation, the throughput of protein glycosylation analysis is still too low to be applied to the exhaustive glycoproteomic analysis. Aiming to elucidate the N-glycosylation of murine epidermis and dermis glycoproteins, here we used a novel approach for focused proteomics. A gross N-glycan profiling (glycomics) of epidermis and dermis was first elucidated both qualitatively and quantitatively upon N-glycan derivatization with novel, stable isotope-coded derivatization reagents followed by MALDI-TOF(/TOF) analysis. This analysis revealed distinct features of the N-glycosylation profile of epidermis and dermis for the first time. A high abundance of high mannose type oligosaccharides was found to be characteristic of murine epidermis glycoproteins. Based on this observation, we performed high mannose type glycoform-focused proteomics by direct tryptic digestion of protein mixtures and affinity enrichment. We identified 15 glycoproteins with 19 N-glycosylation sites that carry high mannose type glycans by off-line LC-MALDI-TOF/TOF mass spectrometry. Moreover the relative quantity of microheterogeneity of different glycoforms present at each N-glycan binding site was determined. Glycoproteins identified were often contained in lysosomes (e.g. cathepsin L and gamma-glutamyl hydrolase), lamellar granules (e.g. glucosylceramidase and cathepsin D), and desmosomes (e.g. desmocollin 1, desmocollin 3, and desmoglein). Lamellar granules are organelles found in the terminally differentiating cells of keratinizing epithelia, and desmosomes are intercellular junctions in vertebrate epithelial cells, thus indicating that N-glycosylation of tissue-specific glycoproteins may contribute to increase the relative proportion of high mannose glycans. The striking roles of lysosomal enzymes in epidermis during lipid remodeling and desquamation may also reflect the observed high abundance of high mannose glycans.
Notes: Uematsu, Rie
Furukawa, Jun-ichi
Nakagawa, Hiroaki
Shinohara, Yasuro
Deguchi, Kisaburo
Monde, Kenji
Nishimura, Shin-Ichiro
Journal Article
Research Support, Non-U.S. Gov't
United States
Mol Cell Proteomics. 2005 Dec;4(12):1977-89. doi: 10.1074/mcp.M500203-MCP200. Epub 2005 Sep 16.
Author Address: Division of Biological Sciences, Graduate School of Science, Frontier Research Center for Post-Genomic Science and Technology, Hokkaido University, Sapporo 001-0021, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 369
Author: Ullrich, A., Bell, J. R., Chen, E. Y., Herrera, R., Petruzzelli, L. M., Dull, T. J., Gray, A., Coussens, L., Liao, Y. C., Tsubokawa, M. and et al.
Year: 1985
Title: Human insulin receptor and its relationship to the tyrosine kinase family of oncogenes
Journal: Nature
Volume: 313
Issue: 6005
Pages: 756-61
Epub Date: 1985/02/06
Date: Feb 28-Mar 6
Short Title: Human insulin receptor and its relationship to the tyrosine kinase family of oncogenes
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/313756a0
Accession Number: 2983222
Keywords: Amino Acid Sequence
Base Sequence
DNA/genetics
ErbB Receptors
Genes
Humans
Membrane Proteins/genetics
Molecular Weight
*Oncogenes
Protein Conformation
Protein Kinases/*genetics
Protein-Tyrosine Kinases
RNA, Messenger/genetics
*Receptor, Insulin/genetics
Receptors, Cell Surface/genetics
Abstract: We have deduced the entire 1,370-amino-acid sequence of the human insulin receptor precursor from a single complementary DNA clone. The precursor starts with a 27-amino-acid signal sequence, followed by the receptor alpha-subunit, a precursor processing enzyme cleavage site, then the beta-subunit containing a single 23-amino-acid transmembrane sequence. There are sequence homologies to human epidermal growth factor receptor and the members of the src family of oncogene products.
Notes: Ullrich, A
Bell, J R
Chen, E Y
Herrera, R
Petruzzelli, L M
Dull, T J
Gray, A
Coussens, L
Liao, Y C
Tsubokawa, M
AM-30075/AM/NIADDK NIH HHS/United States
AM35158/AM/NIADDK NIH HHS/United States
CA-16417/CA/NCI NIH HHS/United States
etc.
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
England
Nature. 1985 Feb 28-Mar 6;313(6005):756-61. doi: 10.1038/313756a0.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1308
Author: Ullrich, S., Münch, A., Neumann, S., Kremmer, E., Tatzelt, J. and Lichtenthaler, S. F.
Year: 2010
Title: The novel membrane protein TMEM59 modulates complex glycosylation, cell surface expression, and secretion of the amyloid precursor protein
Journal: J Biol Chem
Volume: 285
Issue: 27
Pages: 20664-74
Epub Date: 2010/04/30
Date: Jul 2
Short Title: The novel membrane protein TMEM59 modulates complex glycosylation, cell surface expression, and secretion of the amyloid precursor protein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.055608
PMCID: PMC2898310
Accession Number: 20427278
Keywords: Amyloid beta-Protein Precursor/genetics/*metabolism
Animals
Blotting, Northern
Brain/*metabolism
CHO Cells
COS Cells
Cell Line
Chlorocebus aethiops
Cricetinae
Cricetulus
Gene Knockdown Techniques
Genes, Reporter
Humans
Kidney
Membrane Proteins/deficiency/*genetics/metabolism
Nerve Tissue Proteins/deficiency/*genetics/metabolism
Polylysine
Protease Nexins
RNA, Small Interfering/genetics
Receptors, Cell Surface/metabolism
Abstract: Ectodomain shedding of the amyloid precursor protein (APP) by the two proteases alpha- and beta-secretase is a key regulatory event in the generation of the Alzheimer disease amyloid beta peptide (Abeta). At present, little is known about the cellular mechanisms that control APP shedding and Abeta generation. Here, we identified a novel protein, transmembrane protein 59 (TMEM59), as a new modulator of APP shedding. TMEM59 was found to be a ubiquitously expressed, Golgi-localized protein. TMEM59 transfection inhibited complex N- and O-glycosylation of APP in cultured cells. Additionally, TMEM59 induced APP retention in the Golgi and inhibited Abeta generation as well as APP cleavage by alpha- and beta-secretase cleavage, which occur at the plasma membrane and in the endosomes, respectively. Moreover, TMEM59 inhibited the complex N-glycosylation of the prion protein, suggesting a more general modulation of Golgi glycosylation reactions. Importantly, TMEM59 did not affect the secretion of soluble proteins or the alpha-secretase like shedding of tumor necrosis factor alpha, demonstrating that TMEM59 did not disturb the general Golgi function. The phenotype of TMEM59 transfection on APP glycosylation and shedding was similar to the one observed in cells lacking conserved oligomeric Golgi (COG) proteins COG1 and COG2. Both proteins are required for normal localization and activity of Golgi glycosylation enzymes. In summary, this study shows that TMEM59 expression modulates complex N- and O-glycosylation and suggests that TMEM59 affects APP shedding by reducing access of APP to the cellular compartments, where it is normally cleaved by alpha- and beta-secretase.
Notes: 1083-351x
Ullrich, Sylvia
Münch, Anna
Neumann, Stephanie
Kremmer, Elisabeth
Tatzelt, Jörg
Lichtenthaler, Stefan F
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Jul 2;285(27):20664-74. doi: 10.1074/jbc.M109.055608. Epub 2010 Apr 28.
Author Address: German Center for Neurodegenerative Diseases Munich (DZNE) and Adolf Butenandt-Institute, Biochemistry, Ludwig-Maximilians University Munich, 80336 Munich, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 355
Author: Urade, Y., Nagata, A., Suzuki, Y., Fujii, Y. and Hayaishi, O.
Year: 1989
Title: Primary structure of rat brain prostaglandin D synthetase deduced from cDNA sequence
Journal: J Biol Chem
Volume: 264
Issue: 2
Pages: 1041-5
Epub Date: 1989/01/15
Date: Jan 15
Short Title: Primary structure of rat brain prostaglandin D synthetase deduced from cDNA sequence
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2642896
Keywords: Amino Acid Sequence
Animals
Base Sequence
Brain/*enzymology
Cloning, Molecular
DNA/*genetics
Escherichia coli/enzymology/genetics
Genes
*Intramolecular Oxidoreductases
Isomerases/*genetics/metabolism
Lipocalins
Molecular Sequence Data
Molecular Weight
Protein Conformation
Rats
Recombinant Proteins/metabolism
Abstract: The amino acid sequence of rat brain prostaglandin D synthetase (Urade, Y., Fujimoto, N., and Hayaishi, O. (1985) J. Biol. Chem. 260, 12410-12415) was determined by a combination of cDNA and protein sequencing. cDNA clones specific for this enzyme were isolated from a lambda gt11 rat brain cDNA expression library. Nucleotide sequence analyses of cloned cDNA inserts revealed that this enzyme consisted of a 564- or 549-base pair open reading frame coding for a 188- or 183-amino acid polypeptide with a Mr of 21,232 or 20,749 starting at the first or second ATG. About 60% of the deduced amino acid sequence was confirmed by partial amino acid sequencing of tryptic peptides of the purified enzyme. The recognition sequence for N-glycosylation was seen at two positions of amino acid residues 51-53 (-Asn-Ser-Ser-) and 78-80 (-Asn-Leu-Thr-) counted from the first Met. Both sites were considered to be glycosylated with carbohydrate chains of Mr 3,000, since two smaller proteins with Mr 23,000 and 20,000 were found during deglycosylation of the purified enzyme (Mr 26,000) with N-glycanase. The prostaglandin D synthetase activity was detected in fusion proteins obtained from lysogens with recombinants coding from 34 and 19 nucleotides upstream and 47 and 77 downstream from the first ATG, indicating that the glycosyl chain and about 20 amino acid residues of N terminus were not essential for the enzyme activity. The amino acid composition of the purified enzyme indicated that about 20 residues of hydrophobic amino acids of the N terminus are post-translationally deleted, probably as a signal peptide. These results, together with the immunocytochemical localization of this enzyme to rough-surfaced endoplasmic reticulum and other nuclear membrane of oligodendrocytes (Urade, Y., Fujimoto, N., Kaneko, T., Konishi, A., Mizuno, N., and Hayaishi, O. (1987) J. Biol. Chem. 262, 15132-15136) suggest that this enzyme is a membrane-associated protein.
Notes: Urade, Y
Nagata, A
Suzuki, Y
Fujii, Y
Hayaishi, O
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1989 Jan 15;264(2):1041-5.
Author Address: Department of Enzymes and Metabolism, Osaka Bioscience Institute, Furuedai, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 219
Author: Vaith, P., Assmann, G. and Uhlenbruck, G.
Year: 1978
Title: Characterization of the oligosaccharide side chain of apolipoprotein C-III from human plasma very low density lipoproteins
Journal: Biochim Biophys Acta
Volume: 541
Issue: 2
Pages: 234-40
Epub Date: 1978/06/15
Date: Jun 15
Short Title: Characterization of the oligosaccharide side chain of apolipoprotein C-III from human plasma very low density lipoproteins
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/0304-4165(78)90396-3
Accession Number: 208636
Keywords: Alkalies
Apolipoproteins/*blood
Chemical Phenomena
Chemistry
Disaccharides/analysis
Humans
Lipoproteins, VLDL/*blood
Monosaccharides/analysis
Oligosaccharides/*blood
Oxidation-Reduction
Abstract: Apolipoprotein C-III1 and apolipoprotein C-III2 each contain one oligosaccharide side chain, bound O-glycosidically to threonine in position 74 of the amino acid sequence. The studies reported in this paper characterize these alkali labile oligosaccharides, thereby demonstrating the complete structure of apolipoprotein C-III. Monosaccharide analysis revealed the following sugar composition: D-galactose/N-acetyl-D-galactosamine/sialic acid 1 : 1 : 1 and 1 : 1 : 2 for apolipoprotein C-III1 and apolipoprotein C-III2, respectively. Treatment of desialylated apolipoproteins with alkaline borohydride released the reduced disaccharide beta-D-galactosyl-(1 leads to 3)-N-acetyl-D-galactosaminitol, which was detected by gas-liquid chromatography. Further studies employing periodate oxidation and Smith degradation indicated that the structure of the trisaccharide from apolipoprotein C-III1 was alpha-N-acetylneuraminyl-(2 leads to 3)-beta-D-galactosyl-(1 leads to 3)-N-acetyl-D-galactosaminitol. The tetrasaccharide structure from apolipoprotein C-III2 is made up of this trisaccharide plus one sialic acid residue linked to C6 of N-acetyl-D-galactosaminitol, as was shown by the assessment of chromogens formed upon alkaline degradation.
Notes: Vaith, P
Assmann, G
Uhlenbruck, G
Journal Article
Netherlands
Biochim Biophys Acta. 1978 Jun 15;541(2):234-40. doi: 10.1016/0304-4165(78)90396-3.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1546
Author: Vakhrushev, S. Y., Steentoft, C., Vester-Christensen, M. B., Bennett, E. P., Clausen, H. and Levery, S. B.
Year: 2013
Title: Enhanced mass spectrometric mapping of the human GalNAc-type O-glycoproteome with SimpleCells
Journal: Mol Cell Proteomics
Volume: 12
Issue: 4
Pages: 932-44
Epub Date: 2013/02/13
Date: Apr
Short Title: Enhanced mass spectrometric mapping of the human GalNAc-type O-glycoproteome with SimpleCells
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.O112.021972
PMCID: PMC3617340
Accession Number: 23399548
Keywords: Acetylgalactosamine/chemistry/*metabolism
Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line, Tumor
Chymotrypsin/chemistry
Glycoproteins/chemistry/isolation & purification/*metabolism
Glycosylation
Humans
Isoelectric Focusing
Molecular Sequence Data
Peptide Fragments/chemistry/isolation & purification
Peptide Mapping
*Protein Processing, Post-Translational
Proteolysis
Proteome/chemistry/isolation & purification/*metabolism
Tandem Mass Spectrometry/*methods
Trypsin/chemistry
Tyrosine/metabolism
Abstract: Characterizing protein GalNAc-type O-glycosylation has long been a major challenge, and as a result, our understanding of this glycoproteome is particularly poor. Recently, we presented a novel strategy for high throughput identification of O-GalNAc glycosites using zinc finger nuclease gene-engineered "SimpleCell" lines producing homogeneous truncated O-glycosylation. Total lysates of cells were trypsinized and subjected to lectin affinity chromatography enrichment, followed by identification of GalNAc O-glycopeptides by nLC-MS/MS, with electron transfer dissociation employed to specify sites of O-glycosylation. Here, we demonstrate a substantial improvement in the SimpleCell strategy by including an additional stage of lectin affinity chromatography on secreted glycoproteins from culture media (secretome) and by incorporating pre-fractionation of affinity-enriched glycopeptides via IEF before nLC-MS/MS. We applied these improvements to three human SimpleCells studied previously, and each yielded a substantial increase in the number of O-glycoproteins and O-glycosites identified. We found that analysis of the secretome was an important independent factor for increasing identifications, suggesting that further substantial improvements can also be sought through analysis of subcellular organelle fractions. In addition, we uncovered a substantial nonoverlapping set of O-glycoproteins and O-glycosites using an alternative protease digestion (chymotrypsin). In total, the improvements led to identification of 259 glycoproteins, of which 152 (59%) were novel compared with our previous strategy using the same three cell lines. With respect to individual glycosites, we identified a total of 856 sites, of which 508 (59%) were novel compared with our previous strategy; this includes four new identifications of O-GalNAc attached to tyrosine. Furthermore, we uncovered ≈ 220 O-glycosites wherein the peptides were clearly identified, but the glycosites could not be unambiguously assigned to specific positions. The improved strategy should greatly facilitate high throughput characterization of the human GalNAc-type O-glycoproteome as well as be applicable to analysis of other O-glycoproteomes.
Notes: 1535-9484
Vakhrushev, Sergey Y
Steentoft, Catharina
Vester-Christensen, Malene B
Bennett, Eric P
Clausen, Henrik
Levery, Steven B
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2013 Apr;12(4):932-44. doi: 10.1074/mcp.O112.021972. Epub 2013 Feb 11.
Author Address: Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark. seva@sund.ku.dk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1174
Author: Vakonakis, I., Langenhan, T., Prömel, S., Russ, A. and Campbell, I. D.
Year: 2008
Title: Solution structure and sugar-binding mechanism of mouse latrophilin-1 RBL: a 7TM receptor-attached lectin-like domain
Journal: Structure
Volume: 16
Issue: 6
Pages: 944-53
Epub Date: 2008/06/13
Date: Jun
Short Title: Solution structure and sugar-binding mechanism of mouse latrophilin-1 RBL: a 7TM receptor-attached lectin-like domain
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2008.02.020
PMCID: PMC2430599
Accession Number: 18547526
Keywords: Amino Acid Sequence
Amino Acid Substitution
Animals
Carbohydrates/chemistry
Lectins/chemistry
Mice
Models, Molecular
Molecular Sequence Data
Nuclear Magnetic Resonance, Biomolecular
Protein Binding
Protein Structure, Tertiary
Receptors, G-Protein-Coupled
Receptors, Peptide/*chemistry/genetics
Rhamnose/*chemistry
Sequence Homology, Amino Acid
Solutions
Abstract: Latrophilin-1 (Lat-1), a target receptor for alpha-Latrotoxin, is a putative G protein-coupled receptor implicated in synaptic function. The extracellular portion of Lat-1 contains a rhamnose binding lectin (RBL)-like domain of unknown structure. RBL domains, first isolated from the eggs of marine species, are also found in the ectodomains of other metazoan transmembrane proteins, including a recently discovered coreceptor of the neuronal axon guidance molecule SLT-1/Slit. Here, we describe a structure of this domain from the mouse Lat-1. RBL adopts a unique alpha/beta fold with long structured loops important for monosaccharide recognition, as shown in the structure of a complex with L-rhamnose. Sequence alignments and mutagenesis show that residues important for carbohydrate binding are often absent in other receptor-attached examples of RBL, including the SLT-1/Slit coreceptor. We postulate that this domain class facilitates direct protein-protein interactions in many transmembrane receptors.
Notes: 1878-4186
Vakonakis, Ioannis
Langenhan, Tobias
Prömel, Simone
Russ, Andreas
Campbell, Iain D
U54 GM062116/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Structure. 2008 Jun;16(6):944-53. doi: 10.1016/j.str.2008.02.020.
Author Address: Department of Biochemistry, University of Oxford, South Parks Road, Oxford, United Kingdom. ioannis.vakonakis@bioch.ox.ac.uk
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1028
Author: Valnickova, Z., Christensen, T., Skottrup, P., Thøgersen, I. B., Højrup, P. and Enghild, J. J.
Year: 2006
Title: Post-translational modifications of human thrombin-activatable fibrinolysis inhibitor (TAFI): evidence for a large shift in the isoelectric point and reduced solubility upon activation
Journal: Biochemistry
Volume: 45
Issue: 5
Pages: 1525-35
Epub Date: 2006/02/01
Date: Feb 7
Short Title: Post-translational modifications of human thrombin-activatable fibrinolysis inhibitor (TAFI): evidence for a large shift in the isoelectric point and reduced solubility upon activation
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi051956v
Accession Number: 16445295
Keywords: Carboxypeptidase B2/blood/*chemistry/isolation & purification
Chromatography, Liquid
Disulfides/chemistry
Glycosylation
Humans
Isoelectric Point
Mass Spectrometry
Peptides/chemistry/isolation & purification
Polysaccharides/chemistry
*Protein Processing, Post-Translational
Sensitivity and Specificity
Solubility
Abstract: Thrombin-activable fibrinolysis inhibitor (TAFI) is distinct from pancreatic procarboxypeptidase B in several ways. The enzymatic activity of TAFIa is unstable and decays with a half-life of a few minutes. During this study, we observed that (i) the isoelectric point (pI) of TAFI shifts dramatically from pH 5 toward pH 8 upon activation and (ii) TAFIa is significantly less soluble than TAFI. The structural bases for these observations were investigated by characterizing all post-translational modifications, including attached glycans and disulfide connectivity. The analyses revealed that all five potential N-glycosylation sites were utilized including Asn22, Asn51, Asn63, Asn86 (located in the activation peptide), and Asn219 (located in the catalytic domain). Asn219 was also found in an unglycosylated variant. Four of the glycans, Asn51, Asn63, Asn86, and Asn219 displayed microheterogeneity, while the glycan attached to Asn22 appeared to be homogeneous. In addition, bisecting GlcNAc attached to the trimannose core was detected, suggesting an origin other than the liver. Monosaccharide composition and LC-MS/MS analyses did not produce evidence for O glycosylation. TAFI contains eight cysteine residues, of which two, Cys69 and Cys383, are not involved in disulfides and contain free sulfhydryl groups. The remaining six cystines form disulfides, including Cys156-Cys169, Cys228-Cys252, and Cys243-Cys257. This pattern is homologous to pancreatic procarboxypeptidase B, and it is therefore unlikely that permutations in the cysteine connectivity are responsible for the enzymatic instability. LC-MS/MS analyses covering more than 90% of the TAFI amino acid sequence revealed no additional modifications. When these results are taken together, they suggest that the inherent instability of TAFIa is not caused by post-translational modifications. However, after activation, TAFIa loses 80% of the attached glycans, generating a large shift in pI and a propensity to precipitate. These changes are likely to significantly affect the properties of TAFIa as compared to TAFI.
Notes: Valnickova, Zuzana
Christensen, Trine
Skottrup, Peter
Thøgersen, Ida B
Højrup, Peter
Enghild, Jan J
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2006 Feb 7;45(5):1525-35. doi: 10.1021/bi051956v.
Author Address: Center for Insoluble Protein Structure (inSPIN) at the Department of Molecular Biology, Science Park, University of Aarhus, Gustav Wied's Vej 10C, 8000 Aarhus C, Denmark.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1301
Author: Van Antwerpen, P., Slomianny, M. C., Boudjeltia, K. Z., Delporte, C., Faid, V., Calay, D., Rousseau, A., Moguilevsky, N., Raes, M., Vanhamme, L., Furtmüller, P. G., Obinger, C., Vanhaeverbeek, M., Nève, J. and Michalski, J. C.
Year: 2010
Title: Glycosylation pattern of mature dimeric leukocyte and recombinant monomeric myeloperoxidase: glycosylation is required for optimal enzymatic activity
Journal: J Biol Chem
Volume: 285
Issue: 21
Pages: 16351-9
Epub Date: 2010/03/25
Date: May 21
Short Title: Glycosylation pattern of mature dimeric leukocyte and recombinant monomeric myeloperoxidase: glycosylation is required for optimal enzymatic activity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.089748
PMCID: PMC2871502
Accession Number: 20332087
Keywords: Animals
CHO Cells
Cricetinae
Cricetulus
Glycosylation
Humans
Neutrophils/*enzymology
Peroxidase/*chemistry/genetics/metabolism
Polysaccharides/*chemistry/genetics/metabolism
*Protein Multimerization
Protein Structure, Quaternary
Recombinant Proteins
Abstract: The involvement of myeloperoxidase (MPO) in various inflammatory conditions has been the scope of many recent studies. Besides its well studied catalytic activity, the role of its overall structure and glycosylation pattern in biological function is barely known. Here, the N-glycan composition of native dimeric human MPO purified from neutrophils and of monomeric MPO recombinantly expressed in Chinese hamster ovary cells has been investigated. Analyses showed the presence of five N-glycans at positions 323, 355, 391, 483, 729 in both proteins. Site by site analysis demonstrated a well conserved micro- and macro-heterogeneity and more complex-type N-glycans for the recombinant form. Comparison of biological functionality of glycosylated and deglycosylated recombinant MPO suggests that glycosylation is required for optimal enzymatic activity. Data are discussed with regard to biosynthesis and the three-dimensional structure of MPO.
Notes: 1083-351x
Van Antwerpen, Pierre
Slomianny, Marie-Christine
Boudjeltia, Karim Zouaoui
Delporte, Cedric
Faid, Valegh
Calay, Damien
Rousseau, Alexandre
Moguilevsky, Nicole
Raes, Martine
Vanhamme, Luc
Furtmüller, Paul G
Obinger, Christian
Vanhaeverbeek, Michel
Nève, Jean
Michalski, Jean-Claude
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 May 21;285(21):16351-9. doi: 10.1074/jbc.M109.089748. Epub 2010 Mar 23.
Author Address: Laboratory of Pharmaceutical Chemistry and the Analytical Platform of Institute of Pharmacy, UniversitéLibre de Bruxelles, Brussels, Belgium. pvantwer@ulb.ac.be
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 760
Author: van den Akker, F., Zhang, X., Miyagi, M., Huo, X., Misono, K. S. and Yee, V. C.
Year: 2000
Title: Structure of the dimerized hormone-binding domain of a guanylyl-cyclase-coupled receptor
Journal: Nature
Volume: 406
Issue: 6791
Pages: 101-4
Epub Date: 2000/07/14
Date: Jul 6
Short Title: Structure of the dimerized hormone-binding domain of a guanylyl-cyclase-coupled receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/35017602
Accession Number: 10894551
Keywords: Animals
Atrial Natriuretic Factor/metabolism
Binding Sites
COS Cells
Crystallography, X-Ray
Dimerization
Glycosylation
Models, Molecular
Protein Conformation
Protein Structure, Secondary
Receptors, Atrial Natriuretic Factor/*chemistry
Recombinant Proteins/chemistry
Abstract: The atrial natriuretic peptide (ANP) hormone is secreted by the heart in response to an increase in blood pressure. ANP exhibits several potent anti-hypertensive actions in the kidney, adrenal gland and vascular system. These actions are induced by hormone binding extracellularly to the ANP receptor, thereby activating its intracellular guanylyl cyclase domain for the production of cyclic GMP. Here we present the crystal structure of the glycosylated dimerized hormone-binding domain of the ANP receptor at 2.0-A resolution. The monomer comprises two interconnected subdomains, each encompassing a central beta-sheet flanked by alpha-helices, and exhibits the type I periplasmic binding protein fold. Dimerization is mediated by the juxtaposition of four parallel helices, arranged two by two, which brings the two protruding carboxy termini into close relative proximity. From affinity labelling and mutagenesis studies, the ANP-binding site maps to the side of the dimer crevice and extends to near the dimer interface. A conserved chloride-binding site is located in the membrane distal domain, and we found that hormone binding is chloride dependent. These studies suggest mechanisms for hormone activation and the allostery of the ANP receptor.
Notes: van den Akker, F
Zhang, X
Miyagi, M
Huo, X
Misono, K S
Yee, V C
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
England
Nature. 2000 Jul 6;406(6791):101-4. doi: 10.1038/35017602.
Author Address: Department of Molecular Biology, Lerner Research Institute, Cleveland Clinic Foundation, Ohio 44195, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 492
Author: van Halbeek, H., Dorland, L., Vliegenthart, J. F., Montreuil, J., Fournet, B. and Schmid, K.
Year: 1981
Title: Characterization of the microheterogeneity in glycoproteins by 500-MHz 1H-NMR spectroscopy of glycopeptide preparations. Application to a monofucosylated tetra-antennary glycopeptide fraction from human plasma alpha 1-acid glycoprotein
Journal: J Biol Chem
Volume: 256
Issue: 11
Pages: 5588-90
Epub Date: 1981/06/10
Date: Jun 10
Short Title: Characterization of the microheterogeneity in glycoproteins by 500-MHz 1H-NMR spectroscopy of glycopeptide preparations. Application to a monofucosylated tetra-antennary glycopeptide fraction from human plasma alpha 1-acid glycoprotein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7240157
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
*Glycopeptides
*Glycoproteins
Humans
Magnetic Resonance Spectroscopy
*Orosomucoid
Abstract: Five hundred-MHz 1H-NMR spectroscopy was employed to study a monofucosylated tetra-antennary glycopeptide fraction which was derived from human plasma alpha 1-acid glycoprotein. This fraction was earlier judged to be homogeneous by 360-MHz 1H-NMR spectroscopic analysis (Fournet, B., Montreuil, J., Strecker, G., Dorland, L., Haverkamp, J., Vliegenthart, J. F. G., Binette, J. P., and Schmid, K. (1978) Biochemistry 17, 5206-5214). The combination of the improved resolving power and the enhanced sensitivity of the 500-MHz 1H-NMR spectrometer afforded the elucidation of a new type of microheterogeneity with regard to the position of attachment of Fuc. Three isomeric compounds were identified. The major form contains Fuc alpha-(1 leads to 3) linked to GlcNac 7 of the tetra-antennary structure, as shown earlier. The two minor compounds, representing new structures, possess Fuc attached in alpha-(1 leads to 3) linkage to GlcNAc 7' or 5'. It is thus noteworthy that this spectral technique allows elucidation of structures of very closely related carbohydrate chains in a glycopeptide mixture.
Notes: van Halbeek, H
Dorland, L
Vliegenthart, J F
Montreuil, J
Fournet, B
Schmid, K
GM-10374/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1981 Jun 10;256(11):5588-90.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 497
Author: van Halbeek, H., Dorland, L., Vliegenthart, J. F., Schmid, K., Montreuil, J., Fournet, B. and Hull, W. E.
Year: 1980
Title: The application of 500-MHz H-NMR spectroscopy for the structure elucidation of N-acetyllactosamine type asparagine-bound carbohydrate chains of glycoproteins
Journal: FEBS Lett
Volume: 114
Issue: 1
Pages: 11-6
Epub Date: 1980/05/19
Date: May 19
Short Title: The application of 500-MHz H-NMR spectroscopy for the structure elucidation of N-acetyllactosamine type asparagine-bound carbohydrate chains of glycoproteins
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(80)80850-7
Accession Number: 7380009
Keywords: Amino Acid Sequence
Carbohydrate Sequence
Glycopeptides/blood
Humans
Magnetic Resonance Spectroscopy
*Orosomucoid/blood
Notes: van Halbeek, H
Dorland, L
Vliegenthart, J F
Schmid, K
Montreuil, J
Fournet, B
Hull, W E
Journal Article
England
FEBS Lett. 1980 May 19;114(1):11-6. doi: 10.1016/0014-5793(80)80850-7.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 944
Author: van Pesch, V., Lanaya, H., Renauld, J. C. and Michiels, T.
Year: 2004
Title: Characterization of the murine alpha interferon gene family
Journal: J Virol
Volume: 78
Issue: 15
Pages: 8219-28
Epub Date: 2004/07/16
Date: Aug
Short Title: Characterization of the murine alpha interferon gene family
Alternate Journal: Journal of virology
ISSN: 0022-538X (Print)
0022-538x
DOI: 10.1128/jvi.78.15.8219-8228.2004
PMCID: PMC446145
Accession Number: 15254193
Keywords: Alleles
Amino Acid Sequence
Animals
COS Cells
Gene Expression Profiling
Gene Expression Regulation
Genome
Glycosylation
Interferon-alpha/chemistry/classification/*genetics
Mice
Mice, Inbred C57BL
Molecular Sequence Data
Multigene Family
Promoter Regions, Genetic
Abstract: Mouse and human genomes carry more than a dozen genes coding for closely related alpha interferon (IFN-alpha) subtypes. IFN-alpha, as well as IFN-beta, IFN-kappa, IFN-epsilon, and limitin, are thought to bind the same receptor, raising the question of whether different IFN subtypes possess specific functions. As some confusion existed in the identity and characteristics of mouse IFN-alpha subtypes, the availability of data from the mouse genome sequence prompted us to characterize the murine IFN-alpha family. A total of 14 IFN-alpha genes were detected in the mouse genome, in addition to three IFN-alpha pseudogenes. Four IFN-alpha genes (IFN-alpha1, IFN-alpha7/10, IFN-alpha8/6, and IFN-alpha11) exhibited surprising allelic divergence between 129/Sv and C57BL/6 mice. All IFN-alpha subtypes were found to be stable at pH 2 and to exhibit antiviral activity. Interestingly, some IFN subtypes (IFN-alpha4, IFN-alpha11, IFN-alpha12, IFN-beta, and limitin) showed higher biological activity levels than others, whereas IFN-alpha7/10 exhibited lower activity. Most murine IFN-alpha turned out to be N-glycosylated. However, no correlation was found between N-glycosylation and activity. The various IFN-alpha subtypes displayed a good correlation between their antiviral and antiproliferative potencies, suggesting that IFN-alpha subtypes did not diverge primarily to acquire specific biological activities but probably evolved to acquire specific expression patterns. In L929 cells, IFN genes activated in response to poly(I*C) transfection or to viral infection were, however, similar.
Notes: 1098-5514
van Pesch, Vincent
Lanaya, Hanane
Renauld, Jean-Christophe
Michiels, Thomas
Journal Article
Research Support, Non-U.S. Gov't
J Virol. 2004 Aug;78(15):8219-28. doi: 10.1128/JVI.78.15.8219-8228.2004.
Author Address: University of Louvain, Christian de Duve Institute of Cellular Pathology, MIPA-VIRO 74-49, B-1200 Brussels, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 687
Author: van Rooijen, J. J., Jeschke, U., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1998
Title: Expression of N-linked sialyl Le(x) determinants and O-glycans in the carbohydrate moiety of human amniotic fluid transferrin during pregnancy
Journal: Glycobiology
Volume: 8
Issue: 11
Pages: 1053-64
Epub Date: 1998/09/30
Date: Nov
Short Title: Expression of N-linked sialyl Le(x) determinants and O-glycans in the carbohydrate moiety of human amniotic fluid transferrin during pregnancy
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/8.11.1053
Accession Number: 9751792
Keywords: Amino Acids/analysis
Amniotic Fluid/*chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Gel
Chromatography, Ion Exchange
Female
Humans
Lewis Blood Group Antigens/analysis/chemistry
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/*analysis/chemistry
Polyhydramnios/blood/metabolism
Polysaccharides/*analysis/chemistry
Pregnancy
Sialyl Lewis X Antigen
Transferrin/*chemistry
Abstract: Transferrin, a glycoprotein involved in iron transport in body fluids, was isolated from amniotic fluid of a hydramniospatient by sequential anion-exchange chromatography and gel filtration. The N-glycans of human amniotic fluid transferrin (hAFT) were enzymatically liberated by PNGase-F digestion, isolated by gel filtration and fractionated by (high-pH) anion-exchange chromatography. After alkaline borohydride treatment of native hAFT, the released O-glycans were isolated by gel filtration and fractionated by anion-exchange chroma-tography. Structure elucidation of 14 N- and 2 O-glycans was performed by 500 or 600 MHz1H-NMR spectroscopy. Besides conventional N-glycans established earlier for human serum transferrin (hST), new (alpha1-3)-fucosylated N-glycans were found, representing sialyl Le(x) elements. Furthermore, as compared to hST, a higher degree of (alpha1-6)-fucosylation and an increase in branching from di- to triantennary compounds has been detected. The presence of O-glycans is demonstrated for the first time in transferrin.
Notes: van Rooijen, J J
Jeschke, U
Kamerling, J P
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1998 Nov;8(11):1053-64. doi: 10.1093/glycob/8.11.1053.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, P.O. Box 80075, NL-3508 TB Utrecht, The Netherlands, Universität Rostock, Frauenklinik, P.O. Box 18055, Rostock, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 689
Author: van Rooijen, J. J., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1998
Title: Sulfated di-, tri- and tetraantennary N-glycans in human Tamm-Horsfall glycoprotein
Journal: Eur J Biochem
Volume: 256
Issue: 2
Pages: 471-87
Epub Date: 1998/10/06
Date: Sep 1
Short Title: Sulfated di-, tri- and tetraantennary N-glycans in human Tamm-Horsfall glycoprotein
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1046/j.1432-1327.1998.2560471.x
Accession Number: 9760189
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Chromatography/methods
Humans
Magnetic Resonance Spectroscopy
Male
Molecular Sequence Data
Mucoproteins/*urine
Oligosaccharides/analysis/*chemistry
Polysaccharides/*chemistry
Sequence Analysis
Sulfates/chemistry
Uromodulin
Abstract: The primary structures of 32 sulfated di-, tri- and tetraantennary N-glycans of human Tamm-Horsfall glycoprotein (THP) have been determined. THP was isolated from the urine of one healthy male donor. The intact carbohydrate chains were released by PNGase-F and fractionated via FPLC on Resource Q, HPLC on LiChrosorb-NH2, and high-pH anion-exchange chromatography on CarboPac PA-1. Characterizations were performed using 500-MHz and 600-MHz 1H-NMR spectroscopy, in combination with sialidase treatments. The type of characterized N-glycans ranged from monosulfated to trisulfated N-glycans, whereby the sulfate groups were present as 3-O-sulfated Gal (Gal3S) and 4-O-sulfated GalNAc (GalNAc4S). A compilation of the established structures is shown below. [structure in text]
Notes: van Rooijen, J J
Kamerling, J P
Vliegenthart, J F
Journal Article
England
Eur J Biochem. 1998 Sep 1;256(2):471-87. doi: 10.1046/j.1432-1327.1998.2560471.x.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 698
Author: van Rooijen, J. J., Voskamp, A. F., Kamerling, J. P. and Vliegenthart, J. F.
Year: 1999
Title: Glycosylation sites and site-specific glycosylation in human Tamm-Horsfall glycoprotein
Journal: Glycobiology
Volume: 9
Issue: 1
Pages: 21-30
Epub Date: 1999/01/13
Date: Jan
Short Title: Glycosylation sites and site-specific glycosylation in human Tamm-Horsfall glycoprotein
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/9.1.21
Accession Number: 9884403
Keywords: Amino Acid Sequence
Asparagine/metabolism
Binding Sites
Carbohydrate Conformation
Chromatography, Affinity
Chromatography, High Pressure Liquid
Glycosylation
Humans
Magnetic Resonance Spectroscopy
Male
Molecular Sequence Data
Mucoproteins/*chemistry/*urine
Peptide Fragments/chemistry/metabolism
Sequence Analysis
Serine Endopeptidases/metabolism
Spectrometry, Mass, Fast Atom Bombardment
Uromodulin
Abstract: The N-glycosylation sites of human Tamm-Horsfall glycoprotein from one healthy male donor have been characterized, based on an approach using endoproteinase Glu-C (V-8 protease, Staphylococcus aureus ) digestion and a combination of chromatographic techniques, automated Edman sequencing, and fast atom bombardment mass spectrometry. Seven out of the eight potential N-glycosylation sites, namely, Asn52, Asn56, Asn208, Asn251, Asn298, Asn372, and Asn489, turned out to be glycosylated, and the potential glycosylation site at Asn14, being close to the N-terminus, is not used. The carbohydrate microheterogeneity on three of the glycosylation sites was studied in more detail by high-pH anion-exchange chromatographic profiling and 500 MHz1H-NMR spectroscopy. Glycosylation site Asn489 contains mainly di- and tri-charged oligosaccharides which comprise, among others, the GalNAc4 S (beta1-4)GlcNAc terminal sequence. Only glycosylation site Asn251 bears oligomannose-type carbohydrate chains ranging from Man5GlcNAc2to Man8GlcNAc2, in addition to a small amount of complex-type structures. Profiling of the carbohydrate moieties of Asn208 indicates a large heterogeneity, similar to that established for native human Tamm-Horsfall glycoprotein, namely, multiply charged complex-type carbohydrate structures, terminated by sulfate groups, sialic acid residues, and/or the Sda-determinant.
Notes: van Rooijen, J J
Voskamp, A F
Kamerling, J P
Vliegenthart, J F
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1999 Jan;9(1):21-30. doi: 10.1093/glycob/9.1.21.
Author Address: Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, P.O. Box 80075, NL-3508 TB Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 667
Author: Vannier, B., Zhu, X., Brown, D. and Birnbaumer, L.
Year: 1998
Title: The membrane topology of human transient receptor potential 3 as inferred from glycosylation-scanning mutagenesis and epitope immunocytochemistry
Journal: J Biol Chem
Volume: 273
Issue: 15
Pages: 8675-9
Epub Date: 1998/05/16
Date: Apr 10
Short Title: The membrane topology of human transient receptor potential 3 as inferred from glycosylation-scanning mutagenesis and epitope immunocytochemistry
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.273.15.8675
Accession Number: 9535843
Keywords: Animals
Base Sequence
COS Cells
Calcium Channels/chemistry
Cell Membrane/metabolism/ultrastructure
DNA Primers
Epitopes/analysis/chemistry
Glycosylation
Humans
Immunohistochemistry
Ion Channels/*biosynthesis/*chemistry
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
*Protein Structure, Secondary
Recombinant Proteins/biosynthesis/chemistry
TRPC Cation Channels
Transfection
Abstract: Transient receptor potential (Trp) proteins form ion channels implicated in the calcium entry observed after stimulation of the phospholipase C pathway. Kyte-Doolittle analysis of the amino acid sequence of Trp proteins identifies seven hydrophobic regions (H1-H7) with potential of forming transmembrane segments. A limited sequence similarity to voltage-gated calcium channel alpha1 subunits lead to the prediction of six transmembrane (TM) segments flanked by intracellular N and C termini and a putative pore region between TM5 and TM6. However, experimental evidence supporting this model is missing. Using human Trp 3 to test Trp topology, we now confirm the intracellular nature of the termini by immunocytochemistry. We also demonstrate presence of a unique glycosylation site in position 418, which defines one extracellular loop between H2 and H3. After removal of this site and insertion of ten separate glycosylation sites, we defined two additional extracellular loops between H4 and H5, and H6 and H7. This demonstrated the existence of six transmembrane segments formed of H2-H7. Thus, the first hydrophobic region of Trp rather than being a transmembrane segment is intracellular and available for protein-protein interactions. A site placed in the center of the putative pore region was glycosylated, suggesting that this region may have been luminal and was reinserted into the membrane at a late stage of channel assembly.
Notes: Vannier, B
Zhu, X
Brown, D
Birnbaumer, L
GM-54235/GM/NIGMS NIH HHS/United States
HL-45198/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1998 Apr 10;273(15):8675-9. doi: 10.1074/jbc.273.15.8675.
Author Address: Department of Anesthesiology, University of California, Los Angeles, California, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 432
Author: Vauhkonen, M., Viitala, J., Parkkinen, J. and Rauvala, H.
Year: 1985
Title: High-mannose structure of apolipoprotein-B from low-density lipoproteins of human plasma
Journal: Eur J Biochem
Volume: 152
Issue: 1
Pages: 43-50
Epub Date: 1985/10/01
Date: Oct 1
Short Title: High-mannose structure of apolipoprotein-B from low-density lipoproteins of human plasma
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1985.tb09161.x
Accession Number: 4043085
Keywords: *Apolipoproteins B
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Concanavalin A
Glycopeptides/analysis
Humans
*Lipoproteins, LDL
Mannose
Ovalbumin/analysis
Transferrin/analysis
Abstract: Human plasma low-density lipoproteins were purified by flotation followed by gel filtration. The protein moiety of the lipoproteins, apolipoprotein-B, which was detected by polyacrylamide gel electrophoresis as the only protein component, contained 4.4% (by weight) carbohydrate. Glycopeptides liberated from apolipoprotein-B by pronase were fractionated by affinity chromatography on concanavalin-A--Sepharose. The results indicated that high-mannose glycopeptides interacting strongly with the lectin comprise about 37% of the total monosaccharides of apolipoprotein-B. Thus, as compared to the total serum glycoproteins having about 5% of their monosaccharides in high-mannose glycopeptides, low-density lipoproteins are relatively enriched in these structures amounting up to about 10% of the total high-mannose oligosaccharides in serum. The rest of the carbohydrates in low-density lipoproteins are suggested to be mainly biantennary acidic oligosaccharides interacting weakly with concanavalin A. The oligomannosidic chains from native low-density lipoproteins and isolated glycopeptides were released by digestion with endo-beta-N-acetylglucosaminidase H. Thin-layer chromatography of the released oligosaccharides indicated that apolipoprotein-B contains five different oligomannosidic structures varying in the number of the mannose residues from Man5GlcNAc to Man9GlcNAc. Separation of the per-O-benzoylated high-mannose oligosaccharides by high-pressure liquid chromatography revealed the same polymeric structures in a molar ratio (from Man5 to Man9) of 10:2:3:2:3. Apolipoprotein-B in low-density lipoproteins was calculated to contain five high-mannose chains in total. The different high-mannose oligosaccharides liberated by endo-beta-N-acetylglucosaminidase H were isolated with high-pressure liquid chromatography after reduction with NaBH4, and subjected to methylation analysis with gas-liquid chromatography--mass spectrometry. The data of these studies and the results of exoglycosidase treatment suggest the following structure for the main high-mannose oligosaccharide: (formula: see text) The higher polymeric structures are composed of chains in which the Man5GlcNAc structure is continued by one to four Man(alpha 1-2) residues.
Notes: Vauhkonen, M
Viitala, J
Parkkinen, J
Rauvala, H
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1985 Oct 1;152(1):43-50. doi: 10.1111/j.1432-1033.1985.tb09161.x.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1388
Author: Vecchio, A. J. and Malkowski, M. G.
Year: 2011
Title: The structural basis of endocannabinoid oxygenation by cyclooxygenase-2
Journal: J Biol Chem
Volume: 286
Issue: 23
Pages: 20736-45
Epub Date: 2011/04/15
Date: Jun 10
Short Title: The structural basis of endocannabinoid oxygenation by cyclooxygenase-2
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.230367
PMCID: PMC3121521
Accession Number: 21489986
Keywords: Amino Acid Substitution
Animals
Arachidonic Acids/*chemistry/metabolism
Cannabinoid Receptor Modulators/*chemistry/metabolism
Crystallography, X-Ray
Cyclooxygenase 2/*chemistry/genetics/metabolism
*Endocannabinoids
Glycerides/*chemistry/metabolism
Mice
Mutation, Missense
Oxidation-Reduction
Protein Binding
Protein Structure, Tertiary
Substrate Specificity
Abstract: The cyclooxygenases (COX-1 and COX-2) oxygenate arachidonic acid (AA) in the committed step of prostaglandin biogenesis. Substitutions of I434V, H513R, and I523V constitute the only differences in residues lining the cyclooxygenase channel between COX-1 and COX-2. These changes create a hydrophobic pocket in COX-2, with Arg-513 located at the base of the pocket, which has been exploited in the design of COX-2-selective inhibitors. Previous studies have shown that COX-2, but not COX-1, can oxygenate endocannabinoid substrates, including 2-arachidonoyl glycerol (2-AG). To investigate the isoform-specific structural basis of endocannabinoid binding to COX-2, we determined the crystal structure of the 2-AG isomer 1-arachidonoyl glycerol (1-AG) in complex with wild type and R513H murine (mu) COX-2 to 2.2 and 2.35 Å, respectively, and R513H muCOX-2 in complex with AA to 2.45 Å resolution. The 2,3-dihydroxypropyl moiety of 1-AG binds near the opening of the cyclooxygenase channel in the space vacated by the movement of the Leu-531 side chain, validating our previous hypothesis implicating the flexibility of the Leu-531 side chain as a determinant for the ability of COX-2 to oxygenate endocannabinoid substrates. Functional analyses carried out to compliment our structural findings indicated that Y355F and R513H muCOX-2 constructs had no effect on the oxygenation of 1-AG and 2-AG, whereas substitutions that resulted in a shortened side chain for Leu-531 had only modest effects. Both AA and 1-AG bind to R513H muCOX-2 in conformations similar to those observed in the co-crystal structures of these substrates with wild type enzyme.
Notes: 1083-351x
Vecchio, Alex J
Malkowski, Michael G
DMR0225180/GM/NIGMS NIH HHS/United States
P41 RR001646/RR/NCRR NIH HHS/United States
R01 GM077176/GM/NIGMS NIH HHS/United States
RR-01646/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2011 Jun 10;286(23):20736-45. doi: 10.1074/jbc.M111.230367. Epub 2011 Apr 13.
Author Address: Department of Structural Biology, Hauptman-Woodward Medical Research Institute,The State University of New York, Buffalo, New York 14203, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1312
Author: Vecchio, A. J., Simmons, D. M. and Malkowski, M. G.
Year: 2010
Title: Structural basis of fatty acid substrate binding to cyclooxygenase-2
Journal: J Biol Chem
Volume: 285
Issue: 29
Pages: 22152-63
Epub Date: 2010/05/14
Date: Jul 16
Short Title: Structural basis of fatty acid substrate binding to cyclooxygenase-2
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.119867
PMCID: PMC2903402
Accession Number: 20463020
Keywords: Amino Acid Substitution/genetics
Animals
Arachidonic Acid/chemistry/metabolism
Crystallography, X-Ray
Cyclooxygenase 2/*chemistry/*metabolism
Docosahexaenoic Acids/chemistry/metabolism
Eicosapentaenoic Acid/chemistry/metabolism
Fatty Acids/*chemistry/*metabolism
Leucine/metabolism
Mice
Models, Molecular
Molecular Conformation
Mutant Proteins/chemistry/metabolism
Oxygen/metabolism
Structure-Activity Relationship
Substrate Specificity
Abstract: The cyclooxygenases (COX-1 and COX-2) are membrane-associated heme-containing homodimers that generate prostaglandin H(2) from arachidonic acid (AA). Although AA is the preferred substrate, other fatty acids are oxygenated by these enzymes with varying efficiencies. We determined the crystal structures of AA, eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) bound to Co(3+)-protoporphyrin IX-reconstituted murine COX-2 to 2.1, 2.4, and 2.65 A, respectively. AA, EPA, and docosahexaenoic acid bind in different conformations in each monomer constituting the homodimer in their respective structures such that one monomer exhibits nonproductive binding and the other productive binding of the substrate in the cyclooxygenase channel. The interactions identified between protein and substrate when bound to COX-1 are conserved in our COX-2 structures, with the only notable difference being the lack of interaction of the carboxylate of AA and EPA with the side chain of Arg-120. Leu-531 exhibits a different side chain conformation when the nonproductive and productive binding modes of AA are compared. Unlike COX-1, mutating this residue to Ala, Phe, Pro, or Thr did not result in a significant loss of activity or substrate binding affinity. Determination of the L531F:AA crystal structure resulted in AA binding in the same global conformation in each monomer. We speculate that the mobility of the Leu-531 side chain increases the volume available at the opening of the cyclooxygenase channel and contributes to the observed ability of COX-2 to oxygenate a broad spectrum of fatty acid and fatty ester substrates.
Notes: 1083-351x
Vecchio, Alex J
Simmons, Danielle M
Malkowski, Michael G
R01 GM077176/GM/NIGMS NIH HHS/United States
P41 RR001646/RR/NCRR NIH HHS/United States
RR-01646/RR/NCRR NIH HHS/United States
DMR0225180/GM/NIGMS NIH HHS/United States
R01 GM115386/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2010 Jul 16;285(29):22152-63. doi: 10.1074/jbc.M110.119867. Epub 2010 May 12.
Author Address: Hauptman-Woodward Medical Research Institute, Buffalo, New York 14203, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1240
Author: Velho, A. M. and Jarvis, S. M.
Year: 2009
Title: Topological studies of hSVCT1, the human sodium-dependent vitamin C transporter and the influence of N-glycosylation on its intracellular targeting
Journal: Exp Cell Res
Volume: 315
Issue: 13
Pages: 2312-21
Epub Date: 2009/04/22
Date: Aug 1
Short Title: Topological studies of hSVCT1, the human sodium-dependent vitamin C transporter and the influence of N-glycosylation on its intracellular targeting
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2009.04.007
Accession Number: 19379732
Keywords: Animals
Antioxidants/metabolism
Ascorbic Acid/metabolism
COS Cells
Chlorocebus aethiops
Glycosylation
Humans
Mutagenesis, Site-Directed
Organic Anion Transporters, Sodium-Dependent/*chemistry/genetics/*metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/genetics/metabolism
Peptides/genetics/metabolism
*Protein Structure, Secondary
*Protein Transport
Sodium-Coupled Vitamin C Transporters
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Symporters/*chemistry/genetics/*metabolism
Abstract: The Na(+)-dependent transporters, hSVCT1 and hSVCT2, were assessed in COS-1 cells for their membrane topology. Antibodies to N- and C-termini of hSVCT1 and C-terminus of hSVCT2 identified positive immunofluorescence only after permeabilisation, suggesting these regions are intracellular. PNGase F treatment confirmed that WT hSVCT1 (approximately 70-100 kDa) is glycosylated and site-directed mutagenesis of the three putative N-glycosylation sites, Asn138, Asn144, Asn230, demonstrated that mutants N138Q and N144Q were glycosylated (approximately 68-90 kDa) with only 31-65% of WT l-ascorbic acid (AA) uptake while the glycosylation profile of N230Q remained unaltered (approximately 98% of WT activity). However, the N138Q/N144Q double mutant displayed barely detectable membrane expression at approximately 65 kDa, no apparent glycosylation and minimal AA uptake (<10%) with no discernible improvement in expression or activity when cultured at 28 degrees C or 37 degrees C. Marker protein immunocytochemistry with N138Q/N144Q identified intracellular aggregates with hSVCT1 localised at the nuclear membrane but absent at the plasma membrane thus implicating its role as a possible intracellular transporter and suggesting N-glycosylation is required for hSVCT1 membrane targeting. Also, Lys242 on the same putative hydrophilic loop as Asn230 after biotinylation was inaccessible from the extracellular side when analysed by MALDI-TOF MS. A new hSVCT1 secondary structure model supporting these findings is proposed.
Notes: 1090-2422
Velho, Albertina M
Jarvis, Simon M
Biotechnology and Biological Sciences Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Exp Cell Res. 2009 Aug 1;315(13):2312-21. doi: 10.1016/j.yexcr.2009.04.007. Epub 2009 Apr 18.
Author Address: Department of Biosciences University of Kent, CT2 7NJ, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1778
Author: Vendelboe, T. V., Harris, P., Zhao, Y., Walter, T. S., Harlos, K., El Omari, K. and Christensen, H. E.
Year: 2016
Title: The crystal structure of human dopamine β-hydroxylase at 2.9 Å resolution
Journal: Sci Adv
Volume: 2
Issue: 4
Pages: e1500980
Epub Date: 2016/05/07
Date: Apr
Short Title: The crystal structure of human dopamine β-hydroxylase at 2.9 Å resolution
Alternate Journal: Science advances
ISSN: 2375-2548
DOI: 10.1126/sciadv.1500980
PMCID: PMC4846438
Accession Number: 27152332
Keywords: Binding Sites
Catalytic Domain
Copper/chemistry
Crystallography, X-Ray
Dopamine/*metabolism
Dopamine beta-Hydroxylase/*chemistry/metabolism
Humans
Norepinephrine/metabolism
*Protein Conformation
Life sciences
biochemistry
biomolecules
crystal structure
dopamine
dopamine β-hydroxylase
neurochemistry
norepinephrine
structural biology
Abstract: The norepinephrine pathway is believed to modulate behavioral and physiological processes, such as mood, overall arousal, and attention. Furthermore, abnormalities in the pathway have been linked to numerous diseases, for example hypertension, depression, anxiety, Parkinson's disease, schizophrenia, Alzheimer's disease, attention deficit hyperactivity disorder, and cocaine dependence. We report the crystal structure of human dopamine β-hydroxylase, which is the enzyme converting dopamine to norepinephrine. The structure of the DOMON (dopamine β-monooxygenase N-terminal) domain, also found in >1600 other proteins, reveals a possible metal-binding site and a ligand-binding pocket. The catalytic core structure shows two different conformations: an open active site, as also seen in another member of this enzyme family [the peptidylglycine α-hydroxylating (and α-amidating) monooxygenase], and a closed active site structure, in which the two copper-binding sites are only 4 to 5 Å apart, in what might be a coupled binuclear copper site. The dimerization domain adopts a conformation that bears no resemblance to any other known protein structure. The structure provides new molecular insights into the numerous devastating disorders of both physiological and neurological origins associated with the dopamine system.
Notes: 2375-2548
Vendelboe, Trine V
Harris, Pernille
Orcid: 0000-0002-6806-4903
Zhao, Yuguang
Orcid: 0000-0001-8916-8552
Walter, Thomas S
Orcid: 0000-0002-0030-7235
Harlos, Karl
Orcid: 0000-0002-7266-4354
El Omari, Kamel
Orcid: 0000-0003-3506-6045
Christensen, Hans E M
Orcid: 0000-0002-9540-8679
Wellcome Trust/United Kingdom
G1100525/Medical Research Council/United Kingdom
075491/Z/04/Wellcome Trust/United Kingdom
MR/N00065X/1/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Sci Adv. 2016 Apr 8;2(4):e1500980. doi: 10.1126/sciadv.1500980. eCollection 2016 Apr.
Author Address: Department of Chemistry, Kemitorvet 207, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark.
Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford OX3 7BN, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1340
Author: Verdino, P., Witherden, D. A., Havran, W. L. and Wilson, I. A.
Year: 2010
Title: The molecular interaction of CAR and JAML recruits the central cell signal transducer PI3K
Journal: Science
Volume: 329
Issue: 5996
Pages: 1210-4
Epub Date: 2010/09/04
Date: Sep 3
Short Title: The molecular interaction of CAR and JAML recruits the central cell signal transducer PI3K
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1187996
PMCID: PMC2951132
NIHMSID: NIHMS235139
Accession Number: 20813955
Keywords: Animals
Binding Sites
CD28 Antigens/metabolism
CHO Cells
Cell Adhesion Molecules/*chemistry/*metabolism
Chemical Phenomena
Coxsackie and Adenovirus Receptor-Like Membrane Protein
Cricetinae
Cricetulus
Crystallization
Crystallography, X-Ray
Epithelium/immunology
Glycosylation
Hydrogen Bonding
Hydrophobic and Hydrophilic Interactions
Ligands
Mice
Phosphatidylinositol 3-Kinases/*metabolism
Protein Interaction Domains and Motifs
Protein Multimerization
Protein Structure, Tertiary
Receptors, Antigen, T-Cell, gamma-delta/immunology/metabolism
Receptors, Virus/*chemistry/*metabolism
*Signal Transduction
T-Lymphocyte Subsets/immunology/metabolism
Abstract: Coxsackie and adenovirus receptor (CAR) is the primary cellular receptor for group B coxsackieviruses and most adenovirus serotypes and plays a crucial role in adenoviral gene therapy. Recent discovery of the interaction between junctional adhesion molecule-like protein (JAML) and CAR uncovered important functional roles in immunity, inflammation, and tissue homeostasis. Crystal structures of JAML ectodomain (2.2 angstroms) and its complex with CAR (2.8 angstroms) reveal an unusual immunoglobulin-domain assembly for JAML and a charged interface that confers high specificity. Biochemical and mutagenesis studies illustrate how CAR-mediated clustering of JAML recruits phosphoinositide 3-kinase (P13K) to a JAML intracellular sequence motif as delineated for the alphabeta T cell costimulatory receptor CD28. Thus, CAR and JAML are cell signaling receptors of the immune system with implications for asthma, cancer, and chronic nonhealing wounds.
Notes: 1095-9203
Verdino, Petra
Witherden, Deborah A
Havran, Wendy L
Wilson, Ian A
R01 AI064811-01A1/AI/NIAID NIH HHS/United States
R01 AI052257/AI/NIAID NIH HHS/United States
R01 GM080301/GM/NIGMS NIH HHS/United States
AI42266/AI/NIAID NIH HHS/United States
AI52257/AI/NIAID NIH HHS/United States
R01 CA058896-16A1/CA/NCI NIH HHS/United States
R01 CA058896/CA/NCI NIH HHS/United States
AI064811/AI/NIAID NIH HHS/United States
R01 AI036964/AI/NIAID NIH HHS/United States
R37 AI042266/AI/NIAID NIH HHS/United States
R01 AI052257-05/AI/NIAID NIH HHS/United States
CA58896/CA/NCI NIH HHS/United States
R01 AI064811/AI/NIAID NIH HHS/United States
R37 AI042266-13/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2010 Sep 3;329(5996):1210-4. doi: 10.1126/science.1187996.
Author Address: Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1632
Author: Verstraete, K., van Schie, L., Vyncke, L., Bloch, Y., Tavernier, J., Pauwels, E., Peelman, F. and Savvides, S. N.
Year: 2014
Title: Structural basis of the proinflammatory signaling complex mediated by TSLP
Journal: Nat Struct Mol Biol
Volume: 21
Issue: 4
Pages: 375-82
Epub Date: 2014/03/19
Date: Apr
Short Title: Structural basis of the proinflammatory signaling complex mediated by TSLP
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.2794
Accession Number: 24632570
Keywords: Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray
Cytokines/*chemistry/immunology/metabolism
Humans
Interleukin-2/metabolism/physiology
Mice
Models, Immunological
Molecular Sequence Data
Protein Structure, Tertiary
Receptors, Interleukin-7/metabolism/physiology
Signal Transduction/*immunology
Abstract: Thymic stromal lymphopoietin (TSLP), a cytokine produced by epithelial cells at barrier surfaces, is pivotal for the development of widespread chronic inflammatory disorders such as asthma and atopic dermatitis. The structure of the mouse TSLP-mediated signaling complex reveals how TSLP establishes extensive interfaces with its cognate receptor (TSLPR) and the shared interleukin 7 receptor α-chain (IL-7Rα) to evoke membrane-proximal receptor-receptor contacts poised for intracellular signaling. Binding of TSLP to TSLPR is a mechanistic prerequisite for recruitment of IL-7Rα to the high-affinity ternary complex, which we propose is coupled to a structural switch in TSLP at the crossroads of the cytokine-receptor interfaces. Functional interrogation of TSLP-receptor interfaces points to putative interaction hotspots that could be exploited for antagonist design. Finally, we derive the structural rationale for the functional duality of IL-7Rα and establish a consensus for the geometry of ternary complexes mediated by interleukin 2 (IL-2)-family cytokines.
Notes: 1545-9985
Verstraete, Kenneth
van Schie, Loes
Vyncke, Laurens
Bloch, Yehudi
Tavernier, Jan
Pauwels, Ewald
Peelman, Frank
Savvides, Savvas N
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2014 Apr;21(4):375-82. doi: 10.1038/nsmb.2794. Epub 2014 Mar 16.
Author Address: Unit for Structural Biology, Laboratory for Protein Biochemistry and Biomolecular Engineering, Department of Biochemistry & Microbiology, Ghent University, Ghent, Belgium.
Department of Medical Protein Research, Vlaams Interuniversitair Instituut voor Biotechnologie and Ghent University, Ghent, Belgium.
Center for Molecular Modeling, Ghent University, Ghent, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1383
Author: Verstraete, K., Vandriessche, G., Januar, M., Elegheert, J., Shkumatov, A. V., Desfosses, A., Van Craenenbroeck, K., Svergun, D. I., Gutsche, I., Vergauwen, B. and Savvides, S. N.
Year: 2011
Title: Structural insights into the extracellular assembly of the hematopoietic Flt3 signaling complex
Journal: Blood
Volume: 118
Issue: 1
Pages: 60-8
Epub Date: 2011/03/11
Date: Jul 7
Short Title: Structural insights into the extracellular assembly of the hematopoietic Flt3 signaling complex
Alternate Journal: Blood
ISSN: 0006-4971
DOI: 10.1182/blood-2011-01-329532
Accession Number: 21389326
Keywords: Amino Acid Sequence
Crystallography, X-Ray
Cytokines/*chemistry/*metabolism
Extracellular Space/chemistry/metabolism
Hematopoiesis/*physiology
Hematopoietic Stem Cells/physiology
Humans
Ligands
Molecular Sequence Data
Protein Binding/physiology
Protein Structure, Tertiary/physiology
Signal Transduction/*physiology
Structure-Activity Relationship
Thermodynamics
*fms-Like Tyrosine Kinase 3/chemistry/genetics/metabolism
Abstract: The class III receptor tyrosine kinase (RTKIII) Fms-like tyrosine kinase receptor 3 (Flt3) and its cytokine ligand (FL) play central roles in hematopoiesis and the immune system, by establishing signaling cascades crucial for the development and homeostasis of hematopoietic progenitors and antigen-presenting dendritic cells. However, Flt3 is also one of the most frequently mutated receptors in hematologic malignancies and is currently a major prognostic factor and clinical target for acute myeloid leukemia. Here, we report the structural basis for the Flt3 ligand-receptor complex and unveil an unanticipated extracellular assembly unlike any other RTKIII/V complex characterized to date. FL induces dimerization of Flt3 via a remarkably compact binding epitope localized at the tip of extracellular domain 3 of Flt3, and it invokes a ternary complex devoid of homotypic receptor interactions. Comparisons of Flt3 with homologous receptors and available mutagenesis data for FL have allowed us to rationalize the unique features of the Flt3 extracellular assembly. Furthermore, thermodynamic dissection of complex formation points to a pronounced enthalpically driven binding event coupled to an entropic penalty. Together, our data suggest that the high-affinity Flt3:FL complex is driven in part by a single preformed binding epitope on FL reminiscent of a "lock-and-key" binding mode, thereby setting the stage for antagonist design.
Notes: 1528-0020
Verstraete, Kenneth
Vandriessche, Gonzalez
Januar, Mariska
Elegheert, Jonathan
Shkumatov, Alexander V
Desfosses, Ambroise
Van Craenenbroeck, Kathleen
Svergun, Dmitri I
Gutsche, Irina
Vergauwen, Bjorn
Savvides, Savvas N
Journal Article
Research Support, Non-U.S. Gov't
United States
Blood. 2011 Jul 7;118(1):60-8. doi: 10.1182/blood-2011-01-329532. Epub 2011 Mar 9.
Author Address: Unit for Structural Biology, Laboratory for Protein Biochemistry and Biomolecular Engineering (L-ProBE), Ghent University, Ghent, Belgium.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 474
Author: Viau, M., Constans, J., Debray, H. and Montreuil, J.
Year: 1983
Title: Isolation and characterization of the O-glycan chain of the human vitamin-D binding protein
Journal: Biochem Biophys Res Commun
Volume: 117
Issue: 1
Pages: 324-31
Epub Date: 1983/11/30
Date: Nov 30
Short Title: Isolation and characterization of the O-glycan chain of the human vitamin-D binding protein
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/0006-291x(83)91579-6
Accession Number: 6689265
Keywords: Carrier Proteins/*isolation & purification
Electrophoresis, Polyacrylamide Gel
Gas Chromatography-Mass Spectrometry
Glycopeptides/analysis
Humans
Isoelectric Focusing
Polysaccharides/isolation & purification
Vitamin D-Binding Protein
Abstract: On a highly purified preparation, the structure of the carbohydrate chain of the human vitamin D-binding protein was investigated and two genetic forms of this protein were considered (Gc 2 and Gc 1 proteins). It was found that only the Gc 1 protein (Gc1a isoform) was glycosylated, the glycan moiety representing about 1% of the protein. The structure of this O-glycosidically linked glycan was determined to be: Neu Ac alpha (2 leads to 3) Gal beta (1 leads to 3) GaINAc alpha (1 leads to 0) Ser (or Thr). A tetrasaccharidic O-glycan with two N-acetylneuraminic residues was also characterized. The vitamin D-binding protein is a rare example of a serum protein O-glycosylated only on some genetic forms.
Notes: Viau, M
Constans, J
Debray, H
Montreuil, J
Journal Article
United States
Biochem Biophys Res Commun. 1983 Nov 30;117(1):324-31. doi: 10.1016/0006-291x(83)91579-6.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1505
Author: Vigetti, D., Deleonibus, S., Moretto, P., Karousou, E., Viola, M., Bartolini, B., Hascall, V. C., Tammi, M., De Luca, G. and Passi, A.
Year: 2012
Title: Role of UDP-N-acetylglucosamine (GlcNAc) and O-GlcNAcylation of hyaluronan synthase 2 in the control of chondroitin sulfate and hyaluronan synthesis
Journal: J Biol Chem
Volume: 287
Issue: 42
Pages: 35544-35555
Epub Date: 2012/08/14
Date: Oct 12
Short Title: Role of UDP-N-acetylglucosamine (GlcNAc) and O-GlcNAcylation of hyaluronan synthase 2 in the control of chondroitin sulfate and hyaluronan synthesis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.402347
PMCID: PMC3471761
Accession Number: 22887999
Keywords: Amino Acid Substitution
Aorta/*metabolism/pathology
Cell Line
Chondroitin Sulfates/*biosynthesis/genetics
Diabetic Angiopathies/genetics/*metabolism/pathology
Glucuronosyltransferase/genetics/*metabolism
Glycosylation
Humans
Hyaluronan Synthases
Hyaluronic Acid/*biosynthesis/genetics
Muscle Proteins/genetics/*metabolism
Muscle, Smooth, Vascular/*metabolism/pathology
Mutation, Missense
Uridine Diphosphate N-Acetylglucosamine/genetics/*metabolism
Abstract: Hyaluronan (HA) is a glycosaminoglycan present in most tissue microenvironments that can modulate many cell behaviors, including proliferation, migration, and adhesive proprieties. In contrast with other glycosaminoglycans, which are synthesized in the Golgi, HA is synthesized at the plasma membrane by one or more of the three HA synthases (HAS1-3), which use cytoplasmic UDP-glucuronic acid and UDP-N-acetylglucosamine as substrates. Previous studies revealed the importance of UDP-sugars for regulating HA synthesis. Therefore, we analyzed the effect of UDP-GlcNAc availability and protein glycosylation with O-linked N-acetylglucosamine (O-GlcNAcylation) on HA and chondroitin sulfate synthesis in primary human aortic smooth muscle cells. Glucosamine treatment, which increases UDP-GlcNAc availability and protein O-GlcNAcylation, increased synthesis of both HA and chondroitin sulfate. However, increasing O-GlcNAcylation by stimulation with O-(2-acetamido-2-deoxy-d-glucopyranosylidene)amino-N-phenylcarbamate without a concomitant increase of UDP-GlcNAc increased only HA synthesis. We found that HAS2, the main synthase in aortic smooth muscle cells, can be O-GlcNAcylated on serine 221, which strongly increased its activity and its stability (t(&frac12;) >5 h versus ∼17 min without O-GlcNAcylation). S221A mutation prevented HAS2 O-GlcNAcylation, which maintained the rapid turnover rate even in the presence of GlcN and increased UDP-GlcNAc. These findings could explain the elevated matrix HA observed in diabetic vessels that, in turn, could mediate cell dedifferentiation processes critical in vascular pathologies.
Notes: 1083-351x
Vigetti, Davide
Deleonibus, Sara
Moretto, Paola
Karousou, Eugenia
Viola, Manuela
Bartolini, Barbara
Hascall, Vincent C
Tammi, Markku
De Luca, Giancarlo
Passi, Alberto
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Oct 12;287(42):35544-35555. doi: 10.1074/jbc.M112.402347. Epub 2012 Aug 10.
Author Address: Dipartimento di Scienze Chirurgiche e Morfologiche, Università degli Studi dell'Insubria, via J. H. Dunant 5, 21100 Varese, Italy.
Biomedical Engineering ND20, The Cleveland Clinic, Cleveland, Ohio 44195.
Institute of Biomedicine, School of Medicine, Faculty of Health Sciences, University of Eastern Finland, 70211 Kuopio, Finland.
Dipartimento di Scienze Chirurgiche e Morfologiche, Università degli Studi dell'Insubria, via J. H. Dunant 5, 21100 Varese, Italy. Electronic address: alberto.passi@uninsubria.it.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1347
Author: Vitorino, R., Alves, R., Barros, A., Caseiro, A., Ferreira, R., Lobo, M. C., Bastos, A., Duarte, J., Carvalho, D., Santos, L. L. and Amado, F. L.
Year: 2010
Title: Finding new posttranslational modifications in salivary proline-rich proteins
Journal: Proteomics
Volume: 10
Issue: 20
Pages: 3732-42
Epub Date: 2010/09/30
Date: Oct
Short Title: Finding new posttranslational modifications in salivary proline-rich proteins
Alternate Journal: Proteomics
ISSN: 1615-9853
DOI: 10.1002/pmic.201000261
Accession Number: 20879038
Keywords: Amino Acid Sequence
Chromatography, Liquid/*methods
Humans
Molecular Sequence Data
Protein Isoforms/*chemistry/genetics/metabolism
*Protein Processing, Post-Translational
Saliva/*chemistry
Salivary Proline-Rich Proteins/*chemistry/genetics/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/*methods
Abstract: Proline-rich proteins (PRPs) are the most complex family of salivary peptides with distinct isoforms and PTMs. Up to date, only the serine phosphorylation at positions 8, 17, and 22 have been experimentally observed on acidic PRP (aPRPs), and at position 8 on basic PRP1 and 2. The presence of a glucoronyl group at Ser17 was also noticed on aPRP. The main goal of this study was to identify new PTMs and distinct isoforms of salivary PRPs using LC-MALDI-TOF/TOF. Through the salivary peptidome characterization of 20 different subjects from Control, Diabetic, and Head and Neck Cancer groups, it was possible to identify the following species: (i) N-glycosylation sites: two in basic proline-rich protein 2 (bPRP2), one in bPRP3 and one in bPRP4; (ii) O-glycosylation sites: two in bPRP2 and one in aPRP; (iii) other terminal monosaccharide sites: six in bPRP1, two in bPRP2 and two in bPRP3; (iv) other modifications such as N-terminal pyro-Glu (two in bPRP1, six in bPRP2, eight in bPRP3 and nine in bPRP4); (v) phosphorylation in serine, three in bPRP1, one in bPRP2, one in bPRP3 and one in aPRP1; (vi) bPRP1 (allele S, allele M and variant CP5) and bPRP4 (allele M). In summary, salivary peptidome data analysis allowed the identification of 45 new PRP-modified residues, mainly due to glycosylation, phosphorylation and conversion of Gln to pyro-Glu. Moreover, comparing all subject groups, it was noticed a predominance of N-acetyl hexosamine modification on bPRPs in the Head and Neck Cancer patients.
Notes: 1615-9861
Vitorino, Rui
Alves, Renato
Barros, António
Caseiro, Armando
Ferreira, Rita
Lobo, Maria Calheiros
Bastos, Ana
Duarte, José
Carvalho, Davide
Santos, Lúcio Lara
Amado, Francisco L
Journal Article
Research Support, Non-U.S. Gov't
Germany
Proteomics. 2010 Oct;10(20):3732-42. doi: 10.1002/pmic.201000261.
Author Address: Department of Chemistry, University of Aveiro, QOPNA, Mass Spectrometry Center, Aveiro, Portugal.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1436
Author: Vivian, J. P., Duncan, R. C., Berry, R., O'Connor, G. M., Reid, H. H., Beddoe, T., Gras, S., Saunders, P. M., Olshina, M. A., Widjaja, J. M., Harpur, C. M., Lin, J., Maloveste, S. M., Price, D. A., Lafont, B. A., McVicar, D. W., Clements, C. S., Brooks, A. G. and Rossjohn, J.
Year: 2011
Title: Killer cell immunoglobulin-like receptor 3DL1-mediated recognition of human leukocyte antigen B
Journal: Nature
Volume: 479
Issue: 7373
Pages: 401-5
Epub Date: 2011/10/25
Date: Oct 23
Short Title: Killer cell immunoglobulin-like receptor 3DL1-mediated recognition of human leukocyte antigen B
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature10517
PMCID: PMC3723390
NIHMSID: NIHMS490428
Accession Number: 22020283
Keywords: Amino Acid Sequence
Binding Sites/genetics
HLA-B Antigens/*chemistry/genetics/*immunology
Humans
Models, Molecular
Mutant Proteins/chemistry/genetics/immunology
Polymorphism, Genetic/genetics
Protein Structure, Tertiary
Receptors, KIR3DL1/*chemistry/genetics/*immunology
Structure-Activity Relationship
beta 2-Microglobulin/chemistry/immunology
Abstract: Members of the killer cell immunoglobulin-like receptor (KIR) family, a large group of polymorphic receptors expressed on natural killer (NK) cells, recognize particular peptide-laden human leukocyte antigen (pHLA) class I molecules and have a pivotal role in innate immune responses. Allelic variation and extensive polymorphism within the three-domain KIR family (KIR3D, domains D0-D1-D2) affects pHLA binding specificity and is linked to the control of viral replication and the treatment outcome of certain haematological malignancies. Here we describe the structure of a human KIR3DL1 receptor bound to HLA-B*5701 complexed with a self-peptide. KIR3DL1 clamped around the carboxy-terminal end of the HLA-B*5701 antigen-binding cleft, resulting in two discontinuous footprints on the pHLA. First, the D0 domain, a distinguishing feature of the KIR3D family, extended towards β2-microglobulin and abutted a region of the HLA molecule with limited polymorphism, thereby acting as an 'innate HLA sensor' domain. Second, whereas the D2-HLA-B*5701 interface exhibited a high degree of complementarity, the D1-pHLA-B*5701 contacts were suboptimal and accommodated a degree of sequence variation both within the peptide and the polymorphic region of the HLA molecule. Although the two-domain KIR (KIR2D) and KIR3DL1 docked similarly onto HLA-C and HLA-B respectively, the corresponding D1-mediated interactions differed markedly, thereby providing insight into the specificity of KIR3DL1 for discrete HLA-A and HLA-B allotypes. Collectively, in association with extensive mutagenesis studies at the KIR3DL1-pHLA-B*5701 interface, we provide a framework for understanding the intricate interplay between peptide variability, KIR3D and HLA polymorphism in determining the specificity requirements of this essential innate interaction that is conserved across primate species.
Notes: 1476-4687
Vivian, Julian P
Duncan, Renee C
Berry, Richard
O'Connor, Geraldine M
Reid, Hugh H
Beddoe, Travis
Gras, Stephanie
Saunders, Philippa M
Olshina, Maya A
Widjaja, Jacqueline M L
Harpur, Christopher M
Lin, Jie
Maloveste, Sebastien M
Price, David A
Lafont, Bernard A P
McVicar, Daniel W
Clements, Craig S
Brooks, Andrew G
Rossjohn, Jamie
G0501963/Medical Research Council/United Kingdom
ZIA AI001026-04/Intramural NIH HHS/United States
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Nature. 2011 Oct 23;479(7373):401-5. doi: 10.1038/nature10517.
Author Address: Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 896
Author: Vocadlo, D. J., Hang, H. C., Kim, E. J., Hanover, J. A. and Bertozzi, C. R.
Year: 2003
Title: A chemical approach for identifying O-GlcNAc-modified proteins in cells
Journal: Proc Natl Acad Sci U S A
Volume: 100
Issue: 16
Pages: 9116-21
Epub Date: 2003/07/23
Date: Aug 5
Short Title: A chemical approach for identifying O-GlcNAc-modified proteins in cells
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1632821100
PMCID: PMC171382
Accession Number: 12874386
Keywords: Acetylglucosamine/*metabolism
Blotting, Western
Cell Nucleus/metabolism
Cloning, Molecular
Cytoplasm/metabolism
Dose-Response Relationship, Drug
Glycosylation
Hexosamines/chemistry
Humans
Jurkat Cells
Kinetics
Models, Chemical
Nuclear Pore/metabolism
Phosphotransferases (Phosphomutases)/genetics/metabolism
Precipitin Tests
Protein Processing, Post-Translational
Proteins/*metabolism
Recombinant Proteins/metabolism
Serine/metabolism
Substrate Specificity
Threonine/metabolism
Tumor Cells, Cultured
Abstract: The glycosylation of serine and threonine residues with a single GlcNAc moiety is a dynamic posttranslational modification of many nuclear and cytoplasmic proteins. We describe a chemical strategy directed toward identifying O-GlcNAc-modified proteins from living cells or proteins modified in vitro. We demonstrate, in vitro, that each enzyme in the hexosamine salvage pathway, and the enzymes that affect this dynamic modification (UDP-GlcNAc:polypeptidtyltransferase and O-GlcNAcase), tolerate analogues of their natural substrates in which the N-acyl side chain has been modified to bear a bio-orthogonal azide moiety. Accordingly, treatment of cells with N-azidoacetylglucosamine results in the metabolic incorporation of the azido sugar into nuclear and cytoplasmic proteins. These O-azidoacetylglucosamine-modified proteins can be covalently derivatized with various biochemical probes at the site of protein glycosylation by using the Staudinger ligation. The approach was validated by metabolic labeling of nuclear pore protein p62, which is known to be posttranslationally modified with O-GlcNAc. This strategy will prove useful for both the identification of O-GlcNAc-modified proteins and the elucidation of the specific residues that bear this saccharide.
Notes: 1091-6490
Vocadlo, David J
Hang, Howard C
Kim, Eun-Ju
Hanover, John A
Bertozzi, Carolyn R
R01 GM066047/GM/NIGMS NIH HHS/United States
GM066047/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2003 Aug 5;100(16):9116-21. doi: 10.1073/pnas.1632821100. Epub 2003 Jul 21.
Author Address: Center for New Directions in Organic Synthesis, Department of Chemistry, University of California, Berkeley, CA 94720, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 243
Author: Voigt, C. G., Maurer-Fogy, I. and Adolf, G. R.
Year: 1992
Title: Natural human tumor necrosis factor beta (lymphotoxin). Variable O-glycosylation at Thr7, proteolytic processing, and allelic variation
Journal: FEBS Lett
Volume: 314
Issue: 1
Pages: 85-8
Epub Date: 1992/12/07
Date: Dec 7
Short Title: Natural human tumor necrosis factor beta (lymphotoxin). Variable O-glycosylation at Thr7, proteolytic processing, and allelic variation
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(92)81467-z
Accession Number: 1451807
Keywords: Alleles
Amino Acid Sequence
Asparagine/metabolism
B-Lymphocytes/*chemistry
Genetic Variation
Glycosylation
Humans
Lymphotoxin-alpha/*genetics/isolation & purification/*metabolism
Molecular Sequence Data
Neuraminic Acids/analysis
*Protein Processing, Post-Translational
Threonine/metabolism
Abstract: Natural human tumor necrosis factor beta (TNF-beta) purified from supernatants of a human B-lymphoblastoid cell line was found to be heterogeneous in molecular mass, with seven components resolved by gel electrophoresis. All components are N-glycosylated at Asn62; N-glycosylation does not contribute to heterogeneity. In addition, part of the molecules are O-glycosylated at Thr7; O-glycosylation is heterogeneous due to variable decoration with neuraminic acid. The four lower molecular mass forms are derived from the full-length protein by trypsin-like proteolytic cleavage in the N-proximal region; these clipped molecules lack O-linked carbohydrates. Two allelic variants differing in amino acid position 26 (threonine/asparagine) were identified.
Notes: Voigt, C G
Maurer-Fogy, I
Adolf, G R
Journal Article
England
FEBS Lett. 1992 Dec 7;314(1):85-8. doi: 10.1016/0014-5793(92)81467-z.
Author Address: Ernst Boehringer-Institut für Arzneimittelforschung, Bender & Co. GmbH, Vienna, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1714
Author: Volkers, G., Worrall, L. J., Kwan, D. H., Yu, C. C., Baumann, L., Lameignere, E., Wasney, G. A., Scott, N. E., Wakarchuk, W., Foster, L. J., Withers, S. G. and Strynadka, N. C.
Year: 2015
Title: Structure of human ST8SiaIII sialyltransferase provides insight into cell-surface polysialylation
Journal: Nat Struct Mol Biol
Volume: 22
Issue: 8
Pages: 627-35
Epub Date: 2015/07/21
Date: Aug
Short Title: Structure of human ST8SiaIII sialyltransferase provides insight into cell-surface polysialylation
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.3060
Accession Number: 26192331
Keywords: Amino Acid Sequence
Animals
Binding Sites/genetics
Cells, Cultured
Chromatography, Thin Layer
Crystallography, X-Ray
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Kinetics
Mass Spectrometry/methods
Models, Molecular
Molecular Sequence Data
Mutation
Neural Cell Adhesion Molecules/chemistry/genetics/metabolism
Polysaccharides/chemistry/metabolism
Protein Binding
*Protein Structure, Tertiary
Recombinant Proteins/chemistry/metabolism
Sequence Homology, Amino Acid
Sialic Acids/chemistry/*metabolism
Sialyltransferases/*chemistry/genetics/*metabolism
Abstract: Sialyltransferases of the mammalian ST8Sia family catalyze oligo- and polysialylation of surface-localized glycoproteins and glycolipids through transfer of sialic acids from CMP-sialic acid to the nonreducing ends of sialic acid acceptors. The crystal structure of human ST8SiaIII at 1.85-Å resolution presented here is, to our knowledge, the first solved structure of a polysialyltransferase from any species, and it reveals a cluster of polysialyltransferase-specific structural motifs that collectively provide an extended electropositive surface groove for binding of oligo-polysialic acid chain products. The ternary complex of ST8SiaIII with a donor sugar analog and a sulfated glycan acceptor identified with a sialyltransferase glycan array provides insight into the residues involved in substrate binding, specificity and sialyl transfer.
Notes: 1545-9985
Volkers, Gesa
Worrall, Liam J
Kwan, David H
Yu, Ching-Ching
Baumann, Lars
Lameignere, Emilie
Wasney, Gregory A
Scott, Nichollas E
Wakarchuk, Warren
Foster, Leonard J
Withers, Stephen G
Strynadka, Natalie C J
GM098791/GM/NIGMS NIH HHS/United States
GM62116/GM/NIGMS NIH HHS/United States
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2015 Aug;22(8):627-35. doi: 10.1038/nsmb.3060. Epub 2015 Jul 20.
Author Address: 1] Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, Canada. [2] Centre for Blood Research, University of British Columbia, Vancouver, British Columbia, Canada.
1] Department of Chemistry, University of British Columbia, Vancouver, British Columbia, Canada. [2] Centre for High-Throughput Biology, University of British Columbia, Vancouver, British Columbia, Canada.
Department of Chemistry and Biology, Ryerson University, Toronto, Ontario, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1027
Author: Voss, D., Kern, A., Traggiai, E., Eickmann, M., Stadler, K., Lanzavecchia, A. and Becker, S.
Year: 2006
Title: Characterization of severe acute respiratory syndrome coronavirus membrane protein
Journal: FEBS Lett
Volume: 580
Issue: 3
Pages: 968-73
Epub Date: 2006/01/31
Date: Feb 6
Short Title: Characterization of severe acute respiratory syndrome coronavirus membrane protein
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2006.01.026
PMCID: PMC7094741
Accession Number: 16442106
Keywords: Animals
Cell Membrane/genetics/*metabolism
Chlorocebus aethiops
Coronavirus M Proteins
Glycosylation
Humans
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase/genetics/metabolism
Membrane Proteins/genetics/*metabolism
Protein Processing, Post-Translational/*physiology
Protein Transport/physiology
SARS Virus/genetics/*metabolism
Severe Acute Respiratory Syndrome/genetics/metabolism/virology
Vero Cells
Viral Matrix Proteins/genetics/*metabolism
Abstract: The coronavirus membrane protein (M) is the key player in the assembly of virions at intracellular membranes between endoplasmic-reticulum and Golgi-complex. Using a newly established human monoclonal anti-M antibody we detected glycosylated and nonglycosylated membrane-associated M in severe acute respiratory syndrome-associated coronavirus (SARS-CoV) infected cells and in purified virions. Further analyses revealed that M contained a single N-glycosylation site at asparagine 4. Recombinant M was transported to the plasma membrane and gained complex-type N-glycosylation. In SARS-CoV infected cells and in purified virions, however, N-glycosylation of M remained endoglycosidase H-sensitive suggesting that trimming of the N-linked sugar side chain is inhibited.
Notes: 1873-3468
Voss, Daniel
Kern, Anika
Traggiai, Elisabetta
Eickmann, Markus
Stadler, Konrad
Lanzavecchia, Antonio
Becker, Stephan
Journal Article
Research Support, Non-U.S. Gov't
FEBS Lett. 2006 Feb 6;580(3):968-73. doi: 10.1016/j.febslet.2006.01.026. Epub 2006 Jan 19.
Author Address: Institute for Virology, Philipps-University Marburg, Robert-Koch-Str. 17, 35037 Marburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 568
Author: Voss, T., Ergülen, E., Ahorn, H., Kubelka, V., Sugiyama, K., Maurer-Fogy, I. and Glössl, J.
Year: 1993
Title: Expression of human interferon omega 1 in Sf9 cells. No evidence for complex-type N-linked glycosylation or sialylation
Journal: Eur J Biochem
Volume: 217
Issue: 3
Pages: 913-9
Epub Date: 1993/11/01
Date: Nov 1
Short Title: Expression of human interferon omega 1 in Sf9 cells. No evidence for complex-type N-linked glycosylation or sialylation
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1993.tb18321.x
Accession Number: 8223648
Keywords: Animals
Baculoviridae/genetics
Base Sequence
Carbohydrate Sequence
Carbohydrates/analysis
Cell Line
Cells, Cultured
Chromatography, High Pressure Liquid
Circular Dichroism
Cloning, Molecular
DNA Primers
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Interferon Type I/*genetics/isolation & purification/metabolism
Mass Spectrometry
Molecular Sequence Data
Moths
N-Acetylneuraminic Acid
Protein Conformation
Sialic Acids/*metabolism
Spectrometry, Fluorescence
Abstract: Human interferon omega 1 (IFN-omega 1) was expressed in Spodoptera frugiperda Sf9 insect cells using the baculovirus expression system. Half of the protein purified by immunoaffinity chromatography was shown to be N-glycosylated at the same site as the natural IFN-omega 1. The degree of glycosylation was independent of the expression rate. While natural IFN-omega 1 was shown to carry complex-type oligosaccharides [Adolf, G. R., Maurer-Fogy, I., Kalsner, I. & Cantell, K. (1990) J. Biol. Chem. 265, 9290-9295], the insect cell produced protein which was demonstrated by lectin blot, mass spectroscopy and HPLC analysis to contain only the core oligosaccharide. Two different structures, (Man)2(GlcNAc)2[Fuc] and (Man)3(GlcNAc)2[Fuc] were identified. The fucosylation was identified to be (alpha 1-6)-linked to the core saccharide. Sialic acid residues were clearly absent. IFN-omega 1 expressed in S. frugiperda cells was shown to be partially truncated at the C-terminus by nine residues; its antiviral activity when glycosylated was significantly lower than the activity of IFN-omega 1 produced by Sendai-virus-stimulated leukocytes. Circular dichroism and fluorescence spectroscopy did not reveal any structural differences between glycosylated and nonglycosylated IFN-omega 1. This implies the importance of a complex-type glycosylation for the maximal biological activity of human IFN-omega 1.
Notes: Voss, T
Ergülen, E
Ahorn, H
Kubelka, V
Sugiyama, K
Maurer-Fogy, I
Glössl, J
Journal Article
England
Eur J Biochem. 1993 Nov 1;217(3):913-9. doi: 10.1111/j.1432-1033.1993.tb18321.x.
Author Address: Ernst-Boehringer Institut für Arzneimittelforschung, Department of Protein Chemistry, Bender & Co., Vienna, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1029
Author: Vosseller, K., Trinidad, J. C., Chalkley, R. J., Specht, C. G., Thalhammer, A., Lynn, A. J., Snedecor, J. O., Guan, S., Medzihradszky, K. F., Maltby, D. A., Schoepfer, R. and Burlingame, A. L.
Year: 2006
Title: O-linked N-acetylglucosamine proteomics of postsynaptic density preparations using lectin weak affinity chromatography and mass spectrometry
Journal: Mol Cell Proteomics
Volume: 5
Issue: 5
Pages: 923-34
Epub Date: 2006/02/03
Date: May
Short Title: O-linked N-acetylglucosamine proteomics of postsynaptic density preparations using lectin weak affinity chromatography and mass spectrometry
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.T500040-MCP200
Accession Number: 16452088
Keywords: Acetylglucosamine/*chemistry
Amino Acid Sequence
Animals
Cattle
Chromatography, Affinity/*methods
Lectins/*chemistry
Mass Spectrometry
Mice
Molecular Sequence Data
Peptides/chemistry
Proteomics/*methods
Synapses/*chemistry
Abstract: O-GlcNAc is a widespread dynamic carbohydrate modification of cytosolic and nuclear proteins with features analogous to phosphorylation. O-GlcNAc acts critically in many cellular processes, including signal transduction, protein degradation, and regulation of gene expression. However, the study of its specific regulatory functions has been limited by difficulties in mapping sites of O-GlcNAc modification. We report methods for direct enrichment and identification of in vivo O-GlcNAc-modified peptides through lectin weak affinity chromatography (LWAC) and mass spectrometry. The effectiveness of this strategy on complex peptide mixtures was demonstrated through enrichment of 145 unique O-GlcNAc-modified peptides from a postsynaptic density preparation. 65 of these O-GlcNAc-modified peptides were sequenced and belonged to proteins with diverse functions in synaptic transmission. Beta-elimination/Michael addition, MS(3) on O-GlcNAc neutral loss ions, and electron capture dissociation were shown to facilitate analysis of O-GlcNAc-modified peptides/sites from lectin weak affinity chromatography enriched postsynaptic density samples. Bassoon and Piccolo, proteins critical to synapse assembly and vesicle docking, were extensively modified by O-GlcNAc. In some cases, O-GlcNAc was mapped to peptides previously identified as phosphorylated, indicating potential interplay between these modifications. Shared substrate amino acid context was apparent in subsets of O-GlcNAc-modified peptides, including "PVST" and a novel "TTA" motif (two hydroxyl-containing amino acids adjacent to an alanine). The results suggest specific roles for O-GlcNAc modification in synaptic transmission, establish a basis for site-specific regulatory studies, and provide methods that will facilitate O-GlcNAc proteome analysis across a wide variety of cells and tissues.
Notes: Vosseller, Keith
Trinidad, Jonathan C
Chalkley, Robert J
Specht, Christian G
Thalhammer, Agnes
Lynn, Aenoch J
Snedecor, June O
Guan, Shenheng
Medzihradszky, Katalin F
Maltby, David A
Schoepfer, Ralf
Burlingame, Alma L
RR14606/RR/NCRR NIH HHS/United States
Wellcome Trust/United Kingdom
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Mol Cell Proteomics. 2006 May;5(5):923-34. doi: 10.1074/mcp.T500040-MCP200. Epub 2006 Feb 1.
Author Address: Mass Spectrometry Facility, Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, USA. kav27@drexel.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 372
Author: Waheed, A., Gottschalk, S., Hille, A., Krentler, C., Pohlmann, R., Braulke, T., Hauser, H., Geuze, H. and von Figura, K.
Year: 1988
Title: Human lysosomal acid phosphatase is transported as a transmembrane protein to lysosomes in transfected baby hamster kidney cells
Journal: Embo j
Volume: 7
Issue: 8
Pages: 2351-8
Epub Date: 1988/08/01
Date: Aug
Short Title: Human lysosomal acid phosphatase is transported as a transmembrane protein to lysosomes in transfected baby hamster kidney cells
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
PMCID: PMC457100
Accession Number: 3056714
Keywords: Acid Phosphatase/biosynthesis/genetics/*metabolism
Animals
Biological Transport
Cell Fractionation
Cell Line
Cell Membrane/*enzymology
Centrifugation, Density Gradient
DNA/genetics
Fluorescent Antibody Technique
Gene Expression Regulation
Humans
Immunohistochemistry
Lysosomes/*enzymology
Membrane Proteins/biosynthesis/genetics/*metabolism
Phosphorylation
Precipitin Tests
Protein Biosynthesis
Protein Precursors/biosynthesis/genetics/*metabolism
Transcription, Genetic
Transfection
Abstract: BHK cells transfected with human lysosomal acid phosphatase (LAP) cDNA (CT29) expressed 70-fold higher enzyme activities of acid phosphatase than non-transfected BHK cells. The CT29-LAP was synthesized in BHK cells as a heterogeneously glycosylated precursor that was tightly membrane associated. Transfer to the trans-Golgi was associated with a small increase in size (approximately 7 kd) and partial processing of the oligosaccharides to complex type structures. CT29-LAP was transferred into lysosomes as shown by subcellular fractionation, immunofluorescence and immunoelectron microscopy. Lack of mannose-6-phosphate residues suggested that transport does not involve mannose-6-phosphate receptors. Part of the membrane-associated CT29-LAP was processed to a soluble form. The mechanism that converts CT29-LAP into a soluble form was sensitive to NH4Cl, and reduced the size of the polypeptide by 7 kd. In vitro translation of CT29-derived cRNA in the presence of microsomal membranes yielded a CT29-LAP precursor that is protected from proteinase K except for a small peptide of approximately 2 kd. In combination with the sequence data available for LAP, these observations suggest that CT29-LAP is synthesized and transported to lysosomes as a transmembrane protein. In the lysosomes, CT29-LAP is released from the membrane by proteolytic cleavage, which removes a C-terminal peptide including the transmembrane domain and the cytosolic tail of 18 amino acids.
Notes: 1460-2075
Waheed, A
Gottschalk, S
Hille, A
Krentler, C
Pohlmann, R
Braulke, T
Hauser, H
Geuze, H
von Figura, K
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 1988 Aug;7(8):2351-8.
Author Address: Zentrum Biochemie, Universität Göttingen, FRG.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 701
Author: Wakabayashi, H., Natsuka, S., Mega, T., Otsuki, N., Isaji, M., Naotsuka, M., Koyama, S., Kanamori, T., Sakai, K. and Hase, S.
Year: 1999
Title: Novel proteoglycan linkage tetrasaccharides of human urinary soluble thrombomodulin, SO4-3GlcAbeta1-3Galbeta1-3(+/-Siaalpha2-6)Galbeta1-4Xyl
Journal: J Biol Chem
Volume: 274
Issue: 9
Pages: 5436-42
Epub Date: 1999/02/20
Date: Feb 26
Short Title: Novel proteoglycan linkage tetrasaccharides of human urinary soluble thrombomodulin, SO4-3GlcAbeta1-3Galbeta1-3(+/-Siaalpha2-6)Galbeta1-4Xyl
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.274.9.5436
Accession Number: 10026155
Keywords: Binding Sites, Antibody
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Gel
Chromatography, High Pressure Liquid
Humans
Hydrolysis
Molecular Sequence Data
Oligosaccharides/*chemistry/immunology/isolation & purification
Proteoglycans/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Thrombomodulin/*chemistry
Urine/chemistry
Abstract: O-linked sugar chains with xylose as a reducing end linked to human urinary soluble thrombomodulin were studied. Sugar chains were liberated by hydrazinolysis followed by N-acetylation and tagged with 2-aminopyridine. Two fractions containing pyridylaminated Xyl as a reducing end were collected. Their structures were determined by partial acid hydrolysis, two-dimensional sugar mapping combined with exoglycosidase digestions, methylation analysis, mass spectrometry, and NMR as SO4-3GlcAbeta1-3Galbeta1-3(+/-Siaalpha2-6)Galbeta1+ ++-4Xyl. These sugar chains could bind to an HNK-1 monoclonal antibody. This is believed to be the first example of a proteoglycan linkage tetrasaccharide with glucuronic acid 3-sulfate and sialic acid.
Notes: Wakabayashi, H
Natsuka, S
Mega, T
Otsuki, N
Isaji, M
Naotsuka, M
Koyama, S
Kanamori, T
Sakai, K
Hase, S
Journal Article
United States
J Biol Chem. 1999 Feb 26;274(9):5436-42. doi: 10.1074/jbc.274.9.5436.
Author Address: Department of Chemistry, Graduate School of Science, Osaka University, Toyonaka, Osaka 560-0043, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1529
Author: Walker, A. K., Soo, K. Y., Levina, V., Talbo, G. H. and Atkin, J. D.
Year: 2013
Title: N-linked glycosylation modulates dimerization of protein disulfide isomerase family A member 2 (PDIA2)
Journal: Febs j
Volume: 280
Issue: 1
Pages: 233-43
Epub Date: 2012/11/22
Date: Jan
Short Title: N-linked glycosylation modulates dimerization of protein disulfide isomerase family A member 2 (PDIA2)
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/febs.12063
Accession Number: 23167757
Keywords: Amino Acid Motifs
Asparagine/metabolism
Endoplasmic Reticulum/enzymology
Enzyme Stability
Glycosylation
HEK293 Cells
HeLa Cells
Histocompatibility Antigens Class I/metabolism
Humans
Immunoprecipitation
Mannose-Binding Lectins/metabolism
Membrane Proteins/metabolism
Procollagen-Proline Dioxygenase/metabolism
Protein Binding
Protein Disulfide-Isomerases/*metabolism
*Protein Multimerization
*Protein Processing, Post-Translational
Protein Transport
Abstract: Protein disulfide isomerase (PDI) family members are important enzymes for the correct folding and maturation of proteins that transit or reside in the endoplasmic reticulum (ER). The human PDI family comprises at least 19 members that differ in cell type expression, substrate specificity and post-translational modifications. PDI family A member 2 (PDIA2, previously known as PDIp) has a similar domain structure to prototypical PDI (also known as PDIA1), but the function and post-translational modifications of PDIA2 remain poorly understood. Unlike most PDI family members, PDIA2 contains three predicted N-linked glycosylation sites. By site-directed mutagenesis and enzymatic deglycosylation, we show here that all three Asn residues within the potential N-linked glycosylation sites of human PDIA2 (N127, N284 and N516) are glycosylated in human cells. Furthermore, mutation of N284 to glycosylation-null Gln increases formation of a highly stable disulfide-bonded PDIA2 dimer. Nevertheless, in HeLa cells, both wild-type and N127/284/516Q mutant PDIA2 proteins localize to the ER, but not the ER-Golgi intermediate compartment, suggesting that glycosylation is important for PDIA2 protein-protein interactions but not subcellular localization. Finally, we identified human major histocompatibility complex class 1 antigens (HLA-A,B,C) as potential binding partners of PDIA2, suggesting an involvement for PDIA2 in antigen presentation in addition to its previously described roles in autoimmunity and Parkinson's disease. These results further characterize this poorly defined member of the PDI family.
Notes: 1742-4658
Walker, Adam K
Soo, Kai Ying
Levina, Vita
Talbo, Gert H
Atkin, Julie D
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS J. 2013 Jan;280(1):233-43. doi: 10.1111/febs.12063. Epub 2012 Dec 14.
Author Address: Department of Biochemistry, La Trobe Institute for Molecular Science, La Trobe University, Bundoora, Victoria, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2067
Author: Walls, A. C., Park, Y. J., Tortorici, M. A., Wall, A., McGuire, A. T. and Veesler, D.
Year: 2020
Title: Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein
Journal: Cell
Volume: 181
Issue: 2
Pages: 281-292.e6
Epub Date: 2020/03/11
Date: Apr 16
Short Title: Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2020.02.058
PMCID: PMC7102599
NIHMSID: NIHMS1572097
Accession Number: 32155444
Keywords: Amino Acid Sequence
Angiotensin-Converting Enzyme 2
Antibodies, Neutralizing/metabolism/pharmacology
Antigens, Viral/chemistry/immunology/metabolism
Betacoronavirus/chemistry/*metabolism
Cell Line
Cryoelectron Microscopy
Humans
Models, Molecular
Peptidyl-Dipeptidase A/metabolism
Receptors, Virus/chemistry/metabolism
SARS Virus/metabolism
SARS-CoV-2
Spike Glycoprotein, Coronavirus/*chemistry/immunology/metabolism/*ultrastructure
Virus Internalization/drug effects
*SARS-CoV
*SARS-CoV-2
*antibodies
*coronavirus
*cryo-EM
*neutralizing antibodies
*spike glycoprotein
*viral receptor
Abstract: The emergence of SARS-CoV-2 has resulted in >90,000 infections and >3,000 deaths. Coronavirus spike (S) glycoproteins promote entry into cells and are the main target of antibodies. We show that SARS-CoV-2 S uses ACE2 to enter cells and that the receptor-binding domains of SARS-CoV-2 S and SARS-CoV S bind with similar affinities to human ACE2, correlating with the efficient spread of SARS-CoV-2 among humans. We found that the SARS-CoV-2 S glycoprotein harbors a furin cleavage site at the boundary between the S(1)/S(2) subunits, which is processed during biogenesis and sets this virus apart from SARS-CoV and SARS-related CoVs. We determined cryo-EM structures of the SARS-CoV-2 S ectodomain trimer, providing a blueprint for the design of vaccines and inhibitors of viral entry. Finally, we demonstrate that SARS-CoV S murine polyclonal antibodies potently inhibited SARS-CoV-2 S mediated entry into cells, indicating that cross-neutralizing antibodies targeting conserved S epitopes can be elicited upon vaccination.
Notes: 1097-4172
Walls, Alexandra C
Park, Young-Jun
Tortorici, M Alejandra
Wall, Abigail
McGuire, Andrew T
Veesler, David
HHSN272201700059C/AI/NIAID NIH HHS/United States
R01 GM120553/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Cell. 2020 Apr 16;181(2):281-292.e6. doi: 10.1016/j.cell.2020.02.058. Epub 2020 Mar 9.
Author Address: Department of Biochemistry, University of Washington, Seattle, WA 98195, USA.
Department of Biochemistry, University of Washington, Seattle, WA 98195, USA; Institute Pasteur & CNRS UMR 3569, Unité de Virologie Structurale, Paris 75015, France.
Vaccines and Infectious Diseases Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98195, USA.
Vaccines and Infectious Diseases Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98195, USA; Department of Global Health, University of Washington, Seattle, WA 98195, USA.
Department of Biochemistry, University of Washington, Seattle, WA 98195, USA. Electronic address: dveesler@uw.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 407
Author: Walsh, K. A., Titani, K., Takio, K., Kumar, S., Hayes, R. and Petra, P. H.
Year: 1986
Title: Amino acid sequence of the sex steroid binding protein of human blood plasma
Journal: Biochemistry
Volume: 25
Issue: 23
Pages: 7584-90
Epub Date: 1986/11/18
Date: Nov 18
Short Title: Amino acid sequence of the sex steroid binding protein of human blood plasma
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00371a048
Accession Number: 3542030
Keywords: Amino Acid Sequence
Cyanogen Bromide
Endopeptidases
Humans
Oligosaccharides/analysis
Peptide Fragments/analysis
*Serine Endopeptidases
*Sex Hormone-Binding Globulin
Trypsin
Abstract: The amino acid sequence of the sex steroid binding protein (SBP) from human plasma has been determined. The SBP subunit consists of a 373-residue polypeptide chain containing two disulfide bonds and three oligosaccharide chains. The sequence was solved primarily by analysis of peptides derived by cleavage at either lysyl or methionyl residues. In our preparations, approximately half of the protein molecules have the amino-terminal sequence Arg-Pro-Val-Leu-Pro; the other half lack Arg-Pro and begin with the valine. Preparations of Hammond et al. [Hammond, G. L., Robinson, P. A., Sugino, H., Ward, D. N., & Finne, J. (1986) J. Steroid Biochem. 24, 815] have an additional leucine at the amino terminus, making a total of 373 residues in the chain. Oligosaccharide chains are placed at Thr-7 and at Asn residues 351 and 367. The two disulfide bonds connect Cys-164 to Cys-188 and Cys-333 to Cys-361. The reported heterogeneity of preparations of the molecule may result in part from the amino-terminal microheterogeneity, in part from variations in the oligosaccharide moieties, and possibly in part from rearrangements involving cyclic imide formation in two Asn-Gly sequences. Certain hydrophobic segments are suggested as possible components of the steroid-binding sites. The protein shows no homology either with the cDNA-derived sequences of the estrogen and glucocorticoid receptors found by others to be homologous with each other or with any other protein sequence in the 1986 data base.
Notes: Walsh, K A
Titani, K
Takio, K
Kumar, S
Hayes, R
Petra, P H
GM-15731/GM/NIGMS NIH HHS/United States
HD-13956/HD/NICHD NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1986 Nov 18;25(23):7584-90. doi: 10.1021/bi00371a048.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1485
Author: Wan, C., Fu, J., Wang, Y., Miao, S., Song, W. and Wang, L.
Year: 2012
Title: Exosome-related multi-pass transmembrane protein TSAP6 is a target of rhomboid protease RHBDD1-induced proteolysis
Journal: PLoS One
Volume: 7
Issue: 5
Pages: e37452
Epub Date: 2012/05/25
Short Title: Exosome-related multi-pass transmembrane protein TSAP6 is a target of rhomboid protease RHBDD1-induced proteolysis
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0037452
PMCID: PMC3356283
Accession Number: 22624035
Keywords: Amino Acid Sequence
Cell Cycle Proteins
DNA Primers
DNA, Complementary/genetics
Exosomes/*metabolism
Flow Cytometry
Gene Knock-In Techniques
HEK293 Cells
Humans
Immunoblotting
Immunoprecipitation
Jurkat Cells
Mass Spectrometry
Molecular Sequence Data
Mutagenesis
Oncogene Proteins/genetics/*metabolism
Oxidoreductases
*Proteolysis
RNA Interference
Real-Time Polymerase Chain Reaction
Serine Endopeptidases/genetics/*metabolism
Abstract: We have previously reported that rhomboid domain containing 1 (RHBDD1), a mammalian rhomboid protease highly expressed in the testis, can cleave the Bcl-2 protein Bik. In this study, we identified a multi-pass transmembrane protein, tumor suppressor activated pathway-6 (TSAP6) as a potential substrate of RHBDD1. RHBDD1 was found to induce the proteolysis of TSAP6 in a dose- and activity-dependent manner. The cleavage of TSAP6 was not restricted to its glycosylated form and occurred in three different regions. In addition, mass spectrometry and mutagenesis analyses both indicated that the major cleavage site laid in the C-terminal of the third transmembrane domain of TSAP6. A somatic cell knock-in approach was used to genetically inactivate the endogenous RHBDD1 in HCT116 and RKO colon cancer cells. Exosome secretion was significantly elevated when RHBDD1 was inactivated in the two cells lines. The increased exosome secretion was verfied through the detection of certain exosomal components, including Tsg101, Tf-R, FasL and Trail. In addition, the elevation of exosome secretion by RHBDD1 inactivation was reduced when TSAP6 was knocked down, indicating that the role of RHBDD1 in regulating exosomal trafficking is very likely to be TSAP6-dependent. We found that the increase in FasL and Trail increased exosome-induced apoptosis in Jurkat cells. Taken together, our findings suggest that RHBDD1 is involved in the regulation of a nonclassical exosomal secretion pathway through the restriction of TSAP6.
Notes: 1932-6203
Wan, Chunhua
Fu, Jun
Wang, Yong
Miao, Shiying
Song, Wei
Wang, Linfang
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2012;7(5):e37452. doi: 10.1371/journal.pone.0037452. Epub 2012 May 18.
Author Address: National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, Peking Union Medical College, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 521
Author: Wang, C. S., Dashti, A., Jackson, K. W., Yeh, J. C., Cummings, R. D. and Tang, J.
Year: 1995
Title: Isolation and characterization of human milk bile salt-activated lipase C-tail fragment
Journal: Biochemistry
Volume: 34
Issue: 33
Pages: 10639-44
Epub Date: 1995/08/22
Date: Aug 22
Short Title: Isolation and characterization of human milk bile salt-activated lipase C-tail fragment
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00033a039
Accession Number: 7654718
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrates/analysis
Chromatography, Gel
Chymotrypsin/metabolism
Cyanogen Bromide
Enzyme Activation/drug effects
Glycosylation
Humans
Lectins/metabolism
Lipase/chemistry/*isolation & purification
Milk, Human/*enzymology
Molecular Sequence Data
Oligosaccharides/analysis/metabolism
Peptide Fragments/chemistry/*isolation & purification
*Sterol Esterase
Trypsin/metabolism
Abstract: Glycosylation positions and oligosaccharide characteristics in the proline-rich, mucin-like, C-terminal region (C-tail) of human milk bile salt-activated lipase (BAL) were studied in order to assess the possible physiological functions of this region. A large-scale purification method has been devised to purify the C-tail fragment from human milk BAL. Chymotryptic, tryptic, and cyanogen bromide cleavages of partially purified BAL and subsequent molecular sieve chromatography yielded 20-30 mg of C-tail fragment from 1 L of human milk. The N-terminal sequence and amino acid composition of the purified C-tail fragment establish that it is derived from residues 528-712 of the enzyme. The O-glycosylated carbohydrates of the C-tail fragment contain fucose, galactose, glucosamine, galactosamine, and neuraminic acid in a molar ratio of 1:3:2:1:0.3, respectively. beta-Elimination reaction revealed that nine threonine residues and less than one serine residue were glycosylated. Edman degradation of C-tail fragment and its pronase subfragment suggest a number of glycosylation sites which are flanked by a consensus motif of PVPP. We suggest that this motif may serve as a signal for O-glycosylation in the C-tail region of BAL. Immunochemical studies indicated that the oligosaccharide chains in the C-tail region of BAL contain Lewis x and Lewis a antigens and, less prominently, sialyl Lewis x and sialyl Lewis a antigens. C-tail fragment was also found to bind jacalin lectin. These observations suggest the possibility that the C-tail region may contribute to adhesive activity in the physiological function of BAL.
Notes: Wang, C S
Dashti, A
Jackson, K W
Yeh, J C
Cummings, R D
Tang, J
CA-37627/CA/NCI NIH HHS/United States
HD-23472/HD/NICHD NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1995 Aug 22;34(33):10639-44. doi: 10.1021/bi00033a039.
Author Address: Protein Studies Program, Oklahoma Medical Research Foundation, Oklahoma City 73104, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1506
Author: Wang, C. Y., Jenkitkasemwong, S., Duarte, S., Sparkman, B. K., Shawki, A., Mackenzie, B. and Knutson, M. D.
Year: 2012
Title: ZIP8 is an iron and zinc transporter whose cell-surface expression is up-regulated by cellular iron loading
Journal: J Biol Chem
Volume: 287
Issue: 41
Pages: 34032-43
Epub Date: 2012/08/18
Date: Oct 5
Short Title: ZIP8 is an iron and zinc transporter whose cell-surface expression is up-regulated by cellular iron loading
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.367284
PMCID: PMC3464513
Accession Number: 22898811
Keywords: Animals
Cation Transport Proteins/*biosynthesis/genetics
Cell Line, Tumor
Cell Membrane/genetics/*metabolism
HEK293 Cells
Humans
Ion Transport/physiology
Iron/*metabolism
Oocytes
Organ Specificity/physiology
Rats
Up-Regulation/*physiology
Xenopus laevis
Abstract: ZIP8 (SLC39A8) belongs to the ZIP family of metal-ion transporters. Among the ZIP proteins, ZIP8 is most closely related to ZIP14, which can transport iron, zinc, manganese, and cadmium. Here we investigated the iron transport ability of ZIP8, its subcellular localization, pH dependence, and regulation by iron. Transfection of HEK 293T cells with ZIP8 cDNA enhanced the uptake of (59)Fe and (65)Zn by 200 and 40%, respectively, compared with controls. Excess iron inhibited the uptake of zinc and vice versa. In RNA-injected Xenopus oocytes, ZIP8-mediated (55)Fe(2+) transport was saturable (K(0.5) of ∼0.7 μm) and inhibited by zinc. ZIP8 also mediated the uptake of (109)Cd(2+), (57)Co(2+), (65)Zn(2+) > (54)Mn(2+), but not (64)Cu (I or II). By using immunofluorescence analysis, we found that ZIP8 expressed in HEK 293T cells localized to the plasma membrane and partially in early endosomes. Iron loading increased total and cell-surface levels of ZIP8 in H4IIE rat hepatoma cells. We also determined by using site-directed mutagenesis that asparagine residues 40, 88, and 96 of rat ZIP8 are glycosylated and that N-glycosylation is not required for iron or zinc transport. Analysis of 20 different human tissues revealed abundant ZIP8 expression in lung and placenta and showed that its expression profile differs markedly from ZIP14, suggesting nonredundant functions. Suppression of endogenous ZIP8 expression in BeWo cells, a placental cell line, reduced iron uptake by ∼40%, suggesting that ZIP8 participates in placental iron transport. Collectively, these data identify ZIP8 as an iron transport protein that may function in iron metabolism.
Notes: 1083-351x
Wang, Chia-Yu
Jenkitkasemwong, Supak
Duarte, Stephanie
Sparkman, Brian K
Shawki, Ali
Mackenzie, Bryan
Knutson, Mitchell D
R01 DK080047/DK/NIDDK NIH HHS/United States
R01 DK080706/DK/NIDDK NIH HHS/United States
R24 CA086307/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Oct 5;287(41):34032-43. doi: 10.1074/jbc.M112.367284. Epub 2012 Aug 16.
Author Address: Food Science and Human Nutrition Department, University of Florida, Gainesville, Florida 32611, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1564
Author: Wang, D., Huang, B., Zhang, S., Yu, X., Wu, W. and Wang, X.
Year: 2013
Title: Structural basis for R-spondin recognition by LGR4/5/6 receptors
Journal: Genes Dev
Volume: 27
Issue: 12
Pages: 1339-44
Epub Date: 2013/06/13
Date: Jun 15
Short Title: Structural basis for R-spondin recognition by LGR4/5/6 receptors
Alternate Journal: Genes & development
ISSN: 0890-9369 (Print)
0890-9369
DOI: 10.1101/gad.219360.113
PMCID: PMC3701189
Accession Number: 23756652
Keywords: Amino Acid Sequence
Animals
Cell Line
Humans
*Models, Molecular
Molecular Sequence Data
Mutation
Protein Binding
Protein Structure, Tertiary
Receptors, G-Protein-Coupled/chemistry/genetics/metabolism
Sequence Alignment
Signal Transduction
Thrombospondins/*chemistry/genetics/*metabolism
Wnt signal
complex
ligand/receptor interaction
Abstract: The R-spondin (RSPO) family of secreted proteins (RSPO1-RSPO4) has pleiotropic functions in development and stem cell growth by strongly enhancing Wnt pathway activation. Recently, leucine-rich repeat-containing G-protein-coupled receptor 4 (LGR4), LGR5, and LGR6 have been identified as receptors for RSPOs. Here we report the complex structure of the LGR4 extracellular domain (ECD) with the RSPO1 N-terminal fragment (RSPO1-2F) containing two adjacent furin-like cysteine-rich domains (FU-CRDs). The LGR4-ECD adopts the anticipated TLR horseshoe structure and uses its concave surface close to the N termini to bind RSPO1-2F. Both the FU-CRD1 and FU-CRD2 domains of RSPO1 contribute to LGR4 interaction, and binding and cellular assays identified critical RSPO1 residues for its biological activities. Our results define the molecular mechanism by which the LGR4/5/6 receptors recognize RSPOs and also provide structural insights into the signaling difference between the LGR4/5/6 receptors and other members in the LGR family.
Notes: 1549-5477
Wang, Dongli
Huang, Binlu
Zhang, Senyan
Yu, Xiaojuan
Wu, Wei
Wang, Xinquan
Journal Article
Research Support, Non-U.S. Gov't
Genes Dev. 2013 Jun 15;27(12):1339-44. doi: 10.1101/gad.219360.113. Epub 2013 Jun 11.
Author Address: Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084, PR China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1336
Author: Wang, D., Zhang, S., Li, L., Liu, X., Mei, K. and Wang, X.
Year: 2010
Title: Structural insights into the assembly and activation of IL-1β with its receptors
Journal: Nat Immunol
Volume: 11
Issue: 10
Pages: 905-11
Epub Date: 2010/08/31
Date: Oct
Short Title: Structural insights into the assembly and activation of IL-1β with its receptors
Alternate Journal: Nature immunology
ISSN: 1529-2908
DOI: 10.1038/ni.1925
Accession Number: 20802483
Keywords: Amino Acid Sequence
Crystallography
Humans
Interleukin 1 Receptor Antagonist Protein/chemistry
Interleukin-1 Receptor Accessory Protein/*chemistry/metabolism
Interleukin-1beta/*chemistry/metabolism
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Tertiary
Receptors, Interleukin-1 Type II/*chemistry/metabolism
Sequence Alignment
Signal Transduction
Abstract: Interleukin 1β (IL-1β) is a key orchestrator of inflammation and host defense that exerts its effects through IL-1 receptor type I (IL-1RI) and IL-1 receptor accessory protein (IL-1RAcP). How IL-1RAcP is recruited by IL-1β-IL-1RI to form the signaling-competent complex remains elusive. Here we present the crystal structure of IL-1β bound to IL-1 receptor type II (IL-1RII) and IL-1RAcP. IL-1β-IL-1RII generated a composite binding surface to recruit IL-1RAcP. Biochemical analysis demonstrated that IL-1β-IL-1RI and IL-1β-IL-1RII interacted similarly with IL-1RAcP. It also showed the importance of two loops of IL-1 receptor antagonist (IL-1Ra) in determining its antagonism. Our results provide a structural basis for assembly and activation of the IL-1 receptor and offer a general cytokine-receptor architecture that governs the IL-1 family of cytokines.
Notes: 1529-2916
Wang, Dongli
Zhang, Senyan
Li, Liang
Liu, Xi
Mei, Kunrong
Wang, Xinquan
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Immunol. 2010 Oct;11(10):905-11. doi: 10.1038/ni.1925. Epub 2010 Aug 29.
Author Address: Center for Structural Biology, School of Life Sciences, Ministry of Education Key Laboratory of Bioinformatics, Tsinghua University, Beijing, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 875
Author: Wang, F., Nakouzi, A., Angeletti, R. H. and Casadevall, A.
Year: 2003
Title: Site-specific characterization of the N-linked oligosaccharides of a murine immunoglobulin M by high-performance liquid chromatography/electrospray mass spectrometry
Journal: Anal Biochem
Volume: 314
Issue: 2
Pages: 266-80
Epub Date: 2003/03/26
Date: Mar 15
Short Title: Site-specific characterization of the N-linked oligosaccharides of a murine immunoglobulin M by high-performance liquid chromatography/electrospray mass spectrometry
Alternate Journal: Analytical biochemistry
ISSN: 0003-2697 (Print)
0003-2697
DOI: 10.1016/s0003-2697(02)00693-0
Accession Number: 12654314
Keywords: Amino Acid Sequence
Animals
Antibodies, Monoclonal/analysis/chemistry/genetics
Binding Sites
Chromatography, High Pressure Liquid/*methods
Cryptococcus neoformans/immunology
Enzyme-Linked Immunosorbent Assay
Glycopeptides/analysis/chemistry/genetics
Glycosylation
Immunoglobulin Heavy Chains/chemistry/genetics
Immunoglobulin Light Chains/chemistry/genetics
Immunoglobulin M/analysis/*chemistry/genetics
Mice
Mice, Inbred BALB C
Molecular Sequence Data
Oligosaccharides/analysis/*chemistry
Peptide Mapping
Polysaccharides/immunology
Spectrometry, Mass, Electrospray Ionization/*methods
Structure-Activity Relationship
Time Factors
Abstract: Immunoglobulin M is an especially important product of the immune system because it plays a critical role in early protection against infections. In this report, the glycosylation pattern of the protective murine monoclonal IgM 12A1 to Cryptococcus neoformans polysaccharide was analyzed by high-performance liquid chromatography coupled with electrospray ionization mass spectrometry. Peptide mapping studies covering 88% of the deduced amino acid sequence indicated that of the six potential N-glycosylation sites in this antibody only five were utilized, as the tryptic peptide derived from monoclonal IgM 12A1 containing Asn-260 was recovered without carbohydrates. The oligosaccharide side chains of monoclonal IgM 12A1 were characterized at each of the N-glycosylation sites. Asn-166 possessed 20 monosialylated and nonsialylated, and fucosylated and nonfucosylated complex- and hybrid-type oligosaccharides and one high-mannose-type oligosaccharide. Thirteen oligosaccharides were attached to the site at Asn-401, including six complex-type, four hybrid-type, and three high-mannose-type oligosaccharides. Twelve hybrid-type oligosaccharides were attached to Asn-378, three of which had terminal sialic acids. Eleven hybrid-type oligosaccharides were attached to Asn-331, seven of which had terminal sialic acids. Only two high-mannose type oligosaccharides were attached to Asn-363. These results indicated great complexity in the structure and composition of oligosaccharides attached to individual IgM glycosylation sites.
Notes: Wang, Fang
Nakouzi, Antonio
Angeletti, Ruth Hogue
Casadevall, Arturo
AI 3342/AI/NIAID NIH HHS/United States
AI 33774/AI/NIAID NIH HHS/United States
HL 59842-01/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Anal Biochem. 2003 Mar 15;314(2):266-80. doi: 10.1016/s0003-2697(02)00693-0.
Author Address: Laboratory for Macromolecular Analysis and Proteomics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA. fawang@aecom.yu.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2071
Author: Wang, J., Wang, Z., Yuan, J., Wang, J. and Shen, X.
Year: 2020
Title: The positive feedback between ACSL4 expression and O-GlcNAcylation contributes to the growth and survival of hepatocellular carcinoma
Journal: Aging (Albany NY)
Volume: 12
Issue: 9
Pages: 7786-7800
Epub Date: 2020/05/02
Date: May 1
Short Title: The positive feedback between ACSL4 expression and O-GlcNAcylation contributes to the growth and survival of hepatocellular carcinoma
Alternate Journal: Aging
ISSN: 1945-4589
DOI: 10.18632/aging.103092
PMCID: PMC7244051
Accession Number: 32357142
Keywords: Apoptosis
Carcinoma, Hepatocellular/*genetics/metabolism/pathology
Cell Cycle
Cell Line, Tumor
Cell Proliferation
Coenzyme A Ligases/biosynthesis/*genetics
DNA, Neoplasm/*genetics
Down-Regulation
Female
*Gene Expression Regulation, Neoplastic
Humans
Liver Neoplasms/*genetics/metabolism/pathology
Male
Protein Processing, Post-Translational
Signal Transduction
*Acsl4
*Glut1
*O-GlcNAcylation
*hepatocellular carcinoma
Abstract: Acyl-CoA ligase 4 (ACSL4) has been reported to be overexpressed in hepatocellular carcinoma (HCC) and to enhance cell proliferation. However, the molecular mechanisms underlying the role of ACSL4 in HCC progression remain largely unclear. Here, we aimed to investigate whether and how O-GlcNAcylation and ACSL4 regulate each other and HCC progression. The clinical significance of ACSL4, O-GlcNAc and GLUT1 in HCC was determined by Pearson chi-squared test and Kaplan-Meier analysis. CCK-8, flow cytometry and in vivo tumour formation assays were performed to detect cell proliferation, apoptosis and tumorigenesis. IP technology was used to evaluate the relationship between ACSL4 and O-GlcNAc. ACSL4, GLUT1 and O-GlcNAc levels were elevated in HCC tissues and predicted poor prognosis in HCC patients. ACSL4 overexpression significantly promoted cell proliferation and tumorigenesis and inhibited cell apoptosis, whereas these effects were all obviously impaired when mTOR signalling was repressed or GLUT1 was downregulated. ACSL4 could be O-GlcNAcylated, and silencing of ACSL4 abolished the effects of O-GlcNAcylation on cell growth promotion and apoptosis inhibition. Collectively, this study demonstrates that ACSL4 contributes to the growth and survival of HCC by enhancing GLUT1-mediated O-GlcNAcylation. In turn, O-GlcNAcylation promotes HCC growth partially by increasing ACSL4 expression.
Notes: 1945-4589
Wang, Jiachen
Wang, Zhao
Yuan, Jiaxiang
Wang, Jiaxiang
Shen, Xinsheng
Journal Article
Aging (Albany NY). 2020 May 1;12(9):7786-7800. doi: 10.18632/aging.103092. Epub 2020 May 1.
Author Address: Department of Minimally Invasive Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, Henan, China.
Department of Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, Henan, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2021
Author: Wang, L., Chen, S., Zhang, J., Mao, S., Mao, W., Zhang, W., Guo, Y., Wu, Y., Wang, R., Yan, Y. and Yao, X.
Year: 2019
Title: Suppressed OGT expression inhibits cell proliferation and modulates EGFR expression in renal cell carcinoma
Journal: Cancer Manag Res
Volume: 11
Pages: 2215-2223
Epub Date: 2019/04/10
Short Title: Suppressed OGT expression inhibits cell proliferation and modulates EGFR expression in renal cell carcinoma
Alternate Journal: Cancer management and research
ISSN: 1179-1322 (Print)
1179-1322
DOI: 10.2147/cmar.s190642
PMCID: PMC6433112
Accession Number: 30962710
Keywords: Egfr
O-GlcNAcylation
Ogt
proliferation
renal cancer
Abstract: PURPOSE: O-linked N -acetylglucosamine (O-GlcNAc or O-GlcNAcylation) is a post-translational modification, which plays a vital role in the progression of various cancers. The purpose of the present study was to assess O-GlcNAcylation in human renal cell carcinoma (RCC). METHODS: O-GlcNAcylation levels and O-GlcNAc-transferase (OGT) expression in human RCC cell lines and 10 paired clinical tissues were detected by Western blot and Immunohistochemis-try. Then, the effects of O-GlcNAcylation on RCC cell proliferation in vitro were investigated by Cell Counting Kit-8 assay. A xenograft assay was performed to assess the in vivo effects of OGT knockdown in RCC cells. Cell apoptosis and cell cycle assays were performed by flow cytometry. Co-immunoprecipitation assays were used to assess epidermal growth factor receptor (EGFR) O-GlcNAcylation and the interaction between OGT and EGFR. RESULTS: O-GlcNAcylation levels and OGT expression were increased in RCC, and the high amounts correlated with poor patient prognosis. OGT knockdown significantly suppressed RCC cell proliferation in vitro and in vivo. Notably, EGFR was modulated by O-GlcNAcylation and directly interacted with OGT. CONCLUSION: These findings provide novel insights into the oncogenic roles of O-GlcNAcylation and OGT in the development of RCC, indicating that OGT might be used as a target for RCC therapy in the future.
Notes: 1179-1322
Wang, Longsheng
Chen, Shaojun
Zhang, Junfeng
Mao, Shiyu
Mao, Weipu
Zhang, Wentao
Guo, Yadong
Wu, Yuan
Wang, Ruiliang
Yan, Yang
Yao, Xudong
Journal Article
Cancer Manag Res. 2019 Mar 19;11:2215-2223. doi: 10.2147/CMAR.S190642. eCollection 2019.
Author Address: Department of Urology, Shanghai Tenth People's Hospital, Tongji University, School of Medicine, Shanghai 200072, China, yaoxudong1967@163.com.
Department of Urology, Shanghai General Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai 200080, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1100
Author: Wang, L. W., Dlugosz, M., Somerville, R. P., Raed, M., Haltiwanger, R. S. and Apte, S. S.
Year: 2007
Title: O-fucosylation of thrombospondin type 1 repeats in ADAMTS-like-1/punctin-1 regulates secretion: implications for the ADAMTS superfamily
Journal: J Biol Chem
Volume: 282
Issue: 23
Pages: 17024-31
Epub Date: 2007/03/31
Date: Jun 8
Short Title: O-fucosylation of thrombospondin type 1 repeats in ADAMTS-like-1/punctin-1 regulates secretion: implications for the ADAMTS superfamily
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M701065200
Accession Number: 17395588
Keywords: ADAMTS Proteins
Amino Acid Sequence
Blotting, Western
Electrophoresis, Polyacrylamide Gel
Extracellular Matrix Proteins/chemistry/genetics/*metabolism
Fucose/*metabolism
Glycosylation
Humans
Molecular Sequence Data
Mutagenesis, Site-Directed
Plasmids
Recombinant Proteins/chemistry/genetics/metabolism
Thrombospondin 1/*metabolism
Abstract: The ADAMTS superfamily contains several metalloproteases (ADAMTS proteases) as well as ADAMTS-like molecules that lack proteolytic activity. Their common feature is the presence of one or more thrombospondin type-1 repeats (TSRs) within a characteristic modular organization. ADAMTS like-1/punctin-1 has four TSRs. Previously, O-fucosylation on Ser or Thr mediated by the endoplasmic reticulum-localized enzyme protein-O-fucosyltransferase 2 (POFUT2) was described for TSRs of thrombospondin-1, properdin, and F-spondin within the sequence Cys-Xaa(1)-Xaa(2)-(Ser/Thr)-Cys-Xaa-Xaa-Gly (where the fucosylated residue is underlined). On mass spectrometric analysis of tryptic peptides from recombinant secreted human punctin-1, the appropriate peptides from TSR2, TSR3, and TSR4 were found to bear either a fucose monosaccharide (TSR3, TSR4) or a fucose-glucose disaccharide (TSR2, TSR3, TSR4). Although mass spectral analysis did not unambiguously identify the relevant peptide from TSR1, metabolic labeling of cells expressing TSR1 and the cysteine-rich module led to incorporation of [(3)H]fucose into this construct. Mutation of the putative modified Ser/Thr residues in TSR2, TSR3, and TSR4 led to significantly decreased levels of secreted punctin-1. Similarly, expression of punctin-1 in Lec-13 cells that are deficient in conversion of GDP-mannose to GDP-fucose substantially decreased the levels of secreted protein, which were restored upon culture in the presence of exogenous l-fucose. In addition, mutation of the single N-linked oligosaccharide in punctin-1 led to decreased levels of secreted punctin-1. Taken together, the data define a critical role for N-glycosylation and O-fucosylation in the biosynthesis of punctin-1. From a broad perspective, these data suggest that O-fucosylation may be a widespread post-translational modification in members of the ADAMTS superfamily with possible regulatory consequences.
Notes: Wang, Lauren W
Dlugosz, Malgosia
Somerville, Robert P T
Raed, Mona
Haltiwanger, Robert S
Apte, Suneel S
AR49930/AR/NIAMS NIH HHS/United States
GM61126/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
J Biol Chem. 2007 Jun 8;282(23):17024-31. doi: 10.1074/jbc.M701065200. Epub 2007 Mar 29.
Author Address: Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1260
Author: Wang, L. W., Leonhard-Melief, C., Haltiwanger, R. S. and Apte, S. S.
Year: 2009
Title: Post-translational modification of thrombospondin type-1 repeats in ADAMTS-like 1/punctin-1 by C-mannosylation of tryptophan
Journal: J Biol Chem
Volume: 284
Issue: 44
Pages: 30004-15
Epub Date: 2009/08/13
Date: Oct 30
Short Title: Post-translational modification of thrombospondin type-1 repeats in ADAMTS-like 1/punctin-1 by C-mannosylation of tryptophan
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.038059
PMCID: PMC2781554
Accession Number: 19671700
Keywords: ADAMTS Proteins
Amino Acid Substitution
Animals
Cell Line
Extracellular Matrix Proteins/genetics/*metabolism
Fucose
Glycosylation
Humans
Mannose/*metabolism
*Protein Processing, Post-Translational
Repetitive Sequences, Nucleic Acid
Thrombospondin 1/genetics/*metabolism
Tryptophan/*metabolism
Abstract: Protein C-mannosylation is the attachment of alpha-mannopyranose to tryptophan via a C-C linkage. This post-translational modification typically occurs within the sequence motif WXXW, which is frequently present in thrombospondin type-1 repeats (TSRs). TSRs are especially numerous in and a defining feature of the ADAMTS superfamily. We investigated the presence and functional significance of C-mannosylation of ADAMTS-like 1/punctin-1, which contains four TSRs (two with predicted C-mannosylation sites), using mass spectrometry, metabolic labeling, site-directed mutagenesis, and expression in C-mannosylation-defective Chinese hamster ovary cell variants. Analysis of tryptic fragments of recombinant human punctin-1 by mass spectrometry identified a peptide derived from TSR1 containing the (36)WDAWGPWSECSRTC(49) sequence of interest modified with two mannose residues and a Glc-Fuc disaccharide (O-fucosylation). Tandem mass spectrometry (MS/MS) and MS/MS/MS analysis demonstrated the characteristic cross-ring cleavage of C-mannose and identified the modified residues as Trp(39) and Trp(42). C-Mannosylation of TSR1 of the related protease ADAMTS5 was also identified. Metabolic labeling of CHO-K1 cells or Lec35.1 cells demonstrated incorporation of d-[2,6-(3)H]mannose in secreted punctin-1 from CHO-K1 cells but not Lec35.1 cells. Quantitation of punctin-1 secretion in Lec35.1 cells versus CHO-K1 cells suggested decreased secretion in Lec35.1 cells. Replacement of mannosylated Trp residues in TSR1 with either Ala or Phe affected punctin secretion efficiency. These data demonstrate that TSR1 from punctin-1 carries C-mannosylation in close proximity to O-linked fucose. Together, these modifications appear to provide a quality control mechanism for punctin-1 secretion.
Notes: 1083-351x
Wang, Lauren W
Leonhard-Melief, Christina
Haltiwanger, Robert S
Apte, Suneel S
R01 CA123071/CA/NCI NIH HHS/United States
AR53890/AR/NIAMS NIH HHS/United States
R56 AR049930/AR/NIAMS NIH HHS/United States
R01 AR049930/AR/NIAMS NIH HHS/United States
R01 CA123071-03/CA/NCI NIH HHS/United States
CA12307101/CA/NCI NIH HHS/United States
AR49930/AR/NIAMS NIH HHS/United States
R01 AR053890/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2009 Oct 30;284(44):30004-15. doi: 10.1074/jbc.M109.038059. Epub 2009 Aug 11.
Author Address: Department of Biomedical Engineering, Cleveland Clinic, Cleveland, Ohio 44195, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1570
Author: Wang, N., Shi, X., Jiang, L., Zhang, S., Wang, D., Tong, P., Guo, D., Fu, L., Cui, Y., Liu, X., Arledge, K. C., Chen, Y. H., Zhang, L. and Wang, X.
Year: 2013
Title: Structure of MERS-CoV spike receptor-binding domain complexed with human receptor DPP4
Journal: Cell Res
Volume: 23
Issue: 8
Pages: 986-93
Epub Date: 2013/07/10
Date: Aug
Short Title: Structure of MERS-CoV spike receptor-binding domain complexed with human receptor DPP4
Alternate Journal: Cell research
ISSN: 1001-0602 (Print)
1001-0602
DOI: 10.1038/cr.2013.92
PMCID: PMC3731569
Accession Number: 23835475
Keywords: Amino Acid Substitution
Animals
Binding Sites
Coronavirus/*metabolism
Dipeptidyl Peptidase 4/chemistry/genetics/*metabolism
Humans
Protein Binding
Protein Structure, Tertiary
SARS Virus/metabolism
Sf9 Cells
Spodoptera
Viral Proteins/chemistry/genetics/*metabolism
Virus Internalization
Abstract: The spike glycoprotein (S) of recently identified Middle East respiratory syndrome coronavirus (MERS-CoV) targets the cellular receptor, dipeptidyl peptidase 4 (DPP4). Sequence comparison and modeling analysis have revealed a putative receptor-binding domain (RBD) on the viral spike, which mediates this interaction. We report the 3.0 Å-resolution crystal structure of MERS-CoV RBD bound to the extracellular domain of human DPP4. Our results show that MERS-CoV RBD consists of a core and a receptor-binding subdomain. The receptor-binding subdomain interacts with DPP4 β-propeller but not its intrinsic hydrolase domain. MERS-CoV RBD and related SARS-CoV RBD share a high degree of structural similarity in their core subdomains, but are notably divergent in the receptor-binding subdomain. Mutagenesis studies have identified several key residues in the receptor-binding subdomain that are critical for viral binding to DPP4 and entry into the target cell. The atomic details at the interface between MERS-CoV RBD and DPP4 provide structural understanding of the virus and receptor interaction, which can guide development of therapeutics and vaccines against MERS-CoV infection.
Notes: 1748-7838
Wang, Nianshuang
Shi, Xuanling
Jiang, Liwei
Zhang, Senyan
Wang, Dongli
Tong, Pei
Guo, Dongxing
Fu, Lili
Cui, Ye
Liu, Xi
Arledge, Kelly C
Chen, Ying-Hua
Zhang, Linqi
Wang, Xinquan
Journal Article
Research Support, Non-U.S. Gov't
Cell Res. 2013 Aug;23(8):986-93. doi: 10.1038/cr.2013.92. Epub 2013 Jul 9.
Author Address: Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1158
Author: Wang, P., Hata, S., Xiao, Y., Murray, J. W. and Wolkoff, A. W.
Year: 2008
Title: Topological assessment of oatp1a1: a 12-transmembrane domain integral membrane protein with three N-linked carbohydrate chains
Journal: Am J Physiol Gastrointest Liver Physiol
Volume: 294
Issue: 4
Pages: G1052-9
Epub Date: 2008/03/01
Date: Apr
Short Title: Topological assessment of oatp1a1: a 12-transmembrane domain integral membrane protein with three N-linked carbohydrate chains
Alternate Journal: American journal of physiology. Gastrointestinal and liver physiology
ISSN: 0193-1857 (Print)
0193-1857
DOI: 10.1152/ajpgi.00584.2007
Accession Number: 18308854
Keywords: Amino Acid Sequence
Animals
Asparagine/metabolism
Cell Membrane/*metabolism
Fluorescent Antibody Technique
Glycosylation
HeLa Cells
Hepatocytes/*metabolism
Humans
Kinetics
Molecular Sequence Data
Mutagenesis, Site-Directed
Organic Anion Transporters, Sodium-Independent/chemistry/genetics/*metabolism
Protein Conformation
Protein Isoforms/metabolism
*Protein Processing, Post-Translational
Protein Structure, Tertiary
Protein Transport
Rats
Sulfobromophthalein/metabolism
Sulfur Radioisotopes/metabolism
Abstract: Organic anion transport protein 1a1 (oatp1a1), a prototypical member of the oatp family of highly homologous transport proteins, is expressed on the basolateral (sinusoidal) surface of rat hepatocytes. The organization of oatp1a1 within the plasma membrane has not been well defined, and computer-based models have predicted possible 12- as well as 10-transmembrane domain structures. Which of oatp1a1's four potential N-linked glycosylation sites are actually glycosylated and their influence on transport function have not been investigated in a mammalian system. In the present study, topology of oatp1a1 in the rat hepatocyte plasma membrane was examined by immunofluorescence analysis using an epitope-specific antibody designed to differentiate a 10- from a 12-transmembrane domain model. To map glycosylation sites, the asparagines at the each of the four N-linked glycosylation consensus sites were mutagenized to glutamines. Mutagenized oatp1a1 constructs were expressed in HeLa cells, and effects on protein expression and transport activity were assessed. These studies revealed that oatp1a1 is a 12-transmembrane-domain protein in which the second and fifth extracellular loops are glycosylated at asparagines 124, 135, and 492, whereas the potential glycosylation site at asparagine 62 is not utilized, consistent with its position in a transmembrane domain. Constructs in which more than one glycosylation site were eliminated had reduced transport activity but not necessarily reduced transporter expression. This was in accord with the finding that fully unglycosylated oatp1a1 was well expressed but located intracellularly with limited transport ability as a consequence of its reduced cell surface expression.
Notes: Wang, Pijun
Hata, Soichiro
Xiao, Yansen
Murray, John W
Wolkoff, Allan W
DK-23026/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
Am J Physiol Gastrointest Liver Physiol. 2008 Apr;294(4):G1052-9. doi: 10.1152/ajpgi.00584.2007. Epub 2008 Feb 28.
Author Address: Department of Medicine, Division of Hepatology, Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1736
Author: Wang, P., Peng, C., Liu, X., Liu, H., Chen, Y., Zheng, L., Han, B. and Pei, H.
Year: 2015
Title: OGT mediated histone H2B S112 GlcNAcylation regulates DNA damage response
Journal: J Genet Genomics
Volume: 42
Issue: 9
Pages: 467-75
Epub Date: 2015/09/27
Date: Sep 20
Short Title: OGT mediated histone H2B S112 GlcNAcylation regulates DNA damage response
Alternate Journal: Journal of genetics and genomics = Yi chuan xue bao
ISSN: 1673-8527 (Print)
1673-8527
DOI: 10.1016/j.jgg.2015.07.002
Accession Number: 26408091
Keywords: Acetylglucosamine/*metabolism
DNA Breaks, Double-Stranded
*DNA Damage
DNA End-Joining Repair
DNA Replication
Glycosylation
HCT116 Cells
HeLa Cells
Histones/*chemistry/*metabolism
Homologous Recombination
Humans
N-Acetylglucosaminyltransferases/*metabolism
Serine/*metabolism
DNA damage response
H2B S112 GlcNAcylation
Nbs1
Ogt
Abstract: O-GlcNAcylation is an important post-translational modification and has been implicated in many fundamental cellular processes. Recent studies showed that O-linked N-acetylglucosamine (GlcNAc) transferase (OGT) mediated O-GlcNAcylation of histone H2B Ser 112 (H2B S112 GlcNAcylation) plays an important role in gene transcription. However, the role of this histone modification in DNA damage response has not been studied yet. In this study, we found that OGT and OGT mediated H2B S112 GlcNAcylation are involved in DNA damage response for maintaining genomic stability and are required for resistance to many DNA-damaging and replication stress-inducing agents. OGT mediated H2B S112 GlcNAcylation increased locally upon the induction of double-strand breaks (DSBs), and depletion of OGT or overexpression of H2B S112A mutant impaired homologous recombination (HR) and nonhomologous end-joining (NHEJ). Mechanistically, H2B S112 GlcNAcylation could bind Nijmegen breakage syndrome 1 (NBS1) and regulate NBS1 foci formation. Taken together, our results demonstrate a new function of histone O-GlcNAcylation in DNA damage response (DDR).
Notes: Wang, Panfei
Peng, Changmin
Liu, Xia
Liu, Hailong
Chen, Yali
Zheng, Li
Han, Baolin
Pei, Huadong
Journal Article
Research Support, Non-U.S. Gov't
China
J Genet Genomics. 2015 Sep 20;42(9):467-75. doi: 10.1016/j.jgg.2015.07.002. Epub 2015 Jul 23.
Author Address: Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Lab, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, China; State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850, China; School of Nursing and Medical Technology, Jianghan University, Wuhan 430056, China.
Department of Radiotherapy, Tianjin Baodi Hospital, Baodi Clinical College of Tianjin Medical University, Tianjin 301800, China; School of Nursing and Medical Technology, Jianghan University, Wuhan 430056, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850, China.
School of Nursing and Medical Technology, Jianghan University, Wuhan 430056, China.
Department of Radiotherapy, Tianjin Baodi Hospital, Baodi Clinical College of Tianjin Medical University, Tianjin 301800, China. Electronic address: tjhanbaolin@163.com.
Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Lab, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, China; State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850, China. Electronic address: peihuadong@hotmail.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1486
Author: Wang, S., Huang, X., Sun, D., Xin, X., Pan, Q., Peng, S., Liang, Z., Luo, C., Yang, Y., Jiang, H., Huang, M., Chai, W., Ding, J. and Geng, M.
Year: 2012
Title: Extensive crosstalk between O-GlcNAcylation and phosphorylation regulates Akt signaling
Journal: PLoS One
Volume: 7
Issue: 5
Pages: e37427
Epub Date: 2012/05/26
Short Title: Extensive crosstalk between O-GlcNAcylation and phosphorylation regulates Akt signaling
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0037427
PMCID: PMC3358304
Accession Number: 22629392
Keywords: Cell Line
Cells, Cultured
Glycosylation
Humans
Phosphorylation
Protein Processing, Post-Translational/*physiology
Proto-Oncogene Proteins c-akt/*metabolism
Signal Transduction/*physiology
Abstract: O-linked N-acetylglucosamine glycosylations (O-GlcNAc) and O-linked phosphorylations (O-phosphate), as two important types of post-translational modifications, often occur on the same protein and bear a reciprocal relationship. In addition to the well documented phosphorylations that control Akt activity, Akt also undergoes O-GlcNAcylation, but the interplay between these two modifications and the biological significance remain unclear, largely due to the technique challenges. Here, we applied a two-step analytic approach composed of the O-GlcNAc immunoenrichment and subsequent O-phosphate immunodetection. Such an easy method enabled us to visualize endogenous glycosylated and phosphorylated Akt subpopulations in parallel and observed the inhibitory effect of Akt O-GlcNAcylations on its phosphorylation. Further studies utilizing mass spectrometry and mutagenesis approaches showed that O-GlcNAcylations at Thr 305 and Thr 312 inhibited Akt phosphorylation at Thr 308 via disrupting the interaction between Akt and PDK1. The impaired Akt activation in turn resulted in the compromised biological functions of Akt, as evidenced by suppressed cell proliferation and migration capabilities. Together, this study revealed an extensive crosstalk between O-GlcNAcylations and phosphorylations of Akt and demonstrated O-GlcNAcylation as a new regulatory modification for Akt signaling.
Notes: 1932-6203
Wang, Shuai
Huang, Xun
Sun, Danni
Xin, Xianliang
Pan, Qiuming
Peng, Shuying
Liang, Zhongjie
Luo, Cheng
Yang, Yiming
Jiang, Hualiang
Huang, Min
Chai, Wengang
Ding, Jian
Geng, Meiyu
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2012;7(5):e37427. doi: 10.1371/journal.pone.0037427. Epub 2012 May 22.
Author Address: Division of Antitumor Pharmacology, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1865
Author: Wang, S., Yang, F., Petyuk, V. A., Shukla, A. K., Monroe, M. E., Gritsenko, M. A., Rodland, K. D., Smith, R. D., Qian, W. J., Gong, C. X. and Liu, T.
Year: 2017
Title: Quantitative proteomics identifies altered O-GlcNAcylation of structural, synaptic and memory-associated proteins in Alzheimer's disease
Journal: J Pathol
Volume: 243
Issue: 1
Pages: 78-88
Epub Date: 2017/06/29
Date: Sep
Short Title: Quantitative proteomics identifies altered O-GlcNAcylation of structural, synaptic and memory-associated proteins in Alzheimer's disease
Alternate Journal: The Journal of pathology
ISSN: 0022-3417 (Print)
0022-3417
DOI: 10.1002/path.4929
PMCID: PMC5647145
NIHMSID: NIHMS889776
Accession Number: 28657654
Keywords: Alzheimer Disease/*metabolism/pathology/physiopathology/psychology
Autopsy
Biomarkers/metabolism
Brain/*metabolism/pathology/physiopathology
Case-Control Studies
Chromatography, Liquid
Glycosylation
Humans
*Memory
Nerve Tissue Proteins/*metabolism
*Protein Processing, Post-Translational
*Proteomics/methods
Reproducibility of Results
Synapses/*metabolism/pathology
*Synaptic Transmission
Tandem Mass Spectrometry
*Alzheimer's disease
*O-GlcNAcylation
*isobaric labelling
*mass spectrometry
Therapeutics. Other authors declare no competing financial interests.
Abstract: Protein modification by O-linked β-N-acetylglucosamine (O-GlcNAc) is emerging as an important factor in the pathogenesis of sporadic Alzheimer's disease (AD); however, detailed molecular characterization of this important protein post-translational modification at the proteome level has been highly challenging, owing to its low stoichiometry and labile nature. Herein, we report the most comprehensive, quantitative proteomics analysis for protein O-GlcNAcylation in postmortem human brain tissues with and without AD by the use of isobaric tandem mass tag labelling, chemoenzymatic photocleavage enrichment, and liquid chromatography coupled to mass spectrometry. A total of 1850 O-GlcNAc peptides covering 1094 O-GlcNAcylation sites were identified from 530 proteins in the human brain. One hundred and thirty-one O-GlcNAc peptides covering 81 proteins were altered in AD brains as compared with controls (q < 0.05). Moreover, alteration of O-GlcNAc peptide abundance could be attributed more to O-GlcNAcylation level than to protein level changes. The altered O-GlcNAcylated proteins belong to several structural and functional categories, including synaptic proteins, cytoskeleton proteins, and memory-associated proteins. These findings suggest that dysregulation of O-GlcNAcylation of multiple brain proteins may be involved in the development of sporadic AD. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Notes: 1096-9896
Wang, Sheng
Yang, Feng
Petyuk, Vladislav A
Shukla, Anil K
Monroe, Matthew E
Gritsenko, Marina A
Rodland, Karin D
Smith, Richard D
Qian, Wei-Jun
Gong, Cheng-Xin
Liu, Tao
Orcid: 0000-0001-9529-6550
P41 GM103493/GM/NIGMS NIH HHS/United States
U24 CA210955/CA/NCI NIH HHS/United States
U24 CA160019/CA/NCI NIH HHS/United States
U24 NS072026/NS/NINDS NIH HHS/United States
P30 AG019610/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Pathol. 2017 Sep;243(1):78-88. doi: 10.1002/path.4929. Epub 2017 Jul 28.
Author Address: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1864
Author: Wang, T., Yu, Q., Li, J., Hu, B., Zhao, Q., Ma, C., Huang, W., Zhuo, L., Fang, H., Liao, L., Eugene Chin, Y. and Jiang, Y.
Year: 2017
Title: O-GlcNAcylation of fumarase maintains tumour growth under glucose deficiency
Journal: Nat Cell Biol
Volume: 19
Issue: 7
Pages: 833-843
Epub Date: 2017/06/20
Date: Jul
Short Title: O-GlcNAcylation of fumarase maintains tumour growth under glucose deficiency
Alternate Journal: Nature cell biology
ISSN: 1465-7392
DOI: 10.1038/ncb3562
Accession Number: 28628081
Keywords: AMP-Activated Protein Kinases/metabolism
Activating Transcription Factor 2/genetics/metabolism
Animals
Cell Line, Tumor
*Cell Proliferation
DNA Methylation
F-Box Proteins/genetics/metabolism
Fumarate Hydratase/*metabolism
Gene Expression Regulation, Neoplastic
Glucose/*deficiency
Glycosylation
Humans
Jumonji Domain-Containing Histone Demethylases/genetics/metabolism
Male
Mice, Nude
Multiprotein Complexes
N-Acetylglucosaminyltransferases/genetics/metabolism
Pancreatic Neoplasms/*enzymology/genetics/pathology
Phosphorylation
Promoter Regions, Genetic
RNA Interference
Time Factors
Transcriptional Activation
Transfection
Tumor Burden
Abstract: Chromatin-associated fumarase (FH) affects histone methylation via its metabolic activity. However, whether this effect is involved in gene transcription remains to be clarified. In this study, we show that under glucose deprivation conditions, AMPK phosphorylates FH at Ser75, which in turn forms a complex with ATF2 and participates in promoter activation. FH-catalysed fumarate in promoter regions inhibits KDM2A demethylase activity, and thus maintains the H3K36me2 profile and facilitates gene expression for cell growth arrest. On the other hand, FH is found to be O-GlcNAcylated at the AMPK phosphorylation site; FH-ATF2-mediated downstream events are impeded by FH O-GlcNAcylation, especially in cancer cells that display robust O-GlcNAc transferase (OGT) activity. Consistently, the FH-Ser75 phosphorylation level inversely correlates with the OGT level and poor prognosis in pancreatic cancer patients. These findings uncover a previously uncharacterized mechanism underlying transcription regulation by FH and the linkage between dysregulated OGT activity and growth advantage of cancer cells under glucose deficiency.
Notes: 1476-4679
Wang, Ting
Orcid: 0000-0003-2257-0699
Yu, Qiujing
Li, Jingjie
Hu, Bin
Zhao, Qin
Ma, Chunmin
Huang, Wenhua
Zhuo, Lingang
Fang, Houqin
Liao, Lujian
Eugene Chin, Y
Jiang, Yuhui
Orcid: 0000-0001-6165-4524
Journal Article
England
Nat Cell Biol. 2017 Jul;19(7):833-843. doi: 10.1038/ncb3562. Epub 2017 Jun 19.
Author Address: The Institute of Cell Metabolism, Shanghai Key Laboratory of Pancreatic Disease, Shanghai General Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai 200080, China.
Department of Gastroenterology, Shanghai General Hospital, Shanghai Jiao Tong University, School of Medicine, Shanghai 200080, China.
Shanghai Key Laboratory of Regulatory Biology, School of Life Sciences, East China Normal University, Shanghai 200241, China.
Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai Jiao Tong University School of Medicine, 320 Yueyang Road, Shanghai 200031, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2079
Author: Wang, W., Kałuża, A., Nouta, J., Nicolardi, S., Ferens-Sieczkowska, M., Wuhrer, M., Lageveen-Kammeijer, G. S. M. and de Haan, N.
Year: 2021
Title: High-throughput glycopeptide profiling of prostate-specific antigen from seminal plasma by MALDI-MS
Journal: Talanta
Volume: 222
Pages: 121495
Epub Date: 2020/11/11
Date: Jan 15
Short Title: High-throughput glycopeptide profiling of prostate-specific antigen from seminal plasma by MALDI-MS
Alternate Journal: Talanta
ISSN: 0039-9140
DOI: 10.1016/j.talanta.2020.121495
Accession Number: 33167210
Keywords: *Glycopeptides
Glycosylation
Humans
Male
Polysaccharides
*Prostate-Specific Antigen
Semen
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Infertility
Mass spectrometry
Prostate-specific antigen
Seminal plasma
Sialic acids
Abstract: An altered total seminal plasma glycosylation has been associated with male infertility, and the highly abundant seminal plasma glycoprotein prostate-specific antigen (PSA) plays an important role in fertilization. However, the exact role of PSA glycosylation in male fertility is not clear. To understand the involvement of PSA glycosylation in the fertilization process, analytical methods are required to study the glycosylation of PSA from seminal plasma with a high glycoform resolution and in a protein-specific manner. In this study, we developed a novel, high-throughput PSA glycopeptide workflow, based on matrix-assisted laser desorption/ionization-mass spectrometry, allowing the discrimination of sialic acid linkage isomers via the derivatization of glycopeptides. The method was successfully applied on a cohort consisting of seminal plasma from infertile and fertile men (N = 102). Forty-four glycopeptides were quantified in all samples, showing mainly complex-type glycans with high levels of fucosylation and sialylation. In addition, N,N-diacetyllactosamine (LacdiNAc) motives were found as well as hybrid-type and high mannose-type structures. Our method showed a high intra- and interday repeatability and revealed no difference in PSA glycosylation between fertile and infertile men. Next to seminal plasma, the method is also expected to be of use for studying PSA glycopeptides derived from other biofluids and/or in other disease contexts.
Notes: 1873-3573
Wang, Wei
Kałuża, Anna
Nouta, Jan
Nicolardi, Simone
Ferens-Sieczkowska, Mirosława
Wuhrer, Manfred
Lageveen-Kammeijer, Guinevere S M
de Haan, Noortje
Journal Article
Netherlands
Talanta. 2021 Jan 15;222:121495. doi: 10.1016/j.talanta.2020.121495. Epub 2020 Aug 13.
Author Address: Leiden University Medical Center, Center for Proteomics and Metabolomics, 2300, RC, Leiden, the Netherlands.
Wrocław Medical University, Department of Chemistry and Immunochemistry, Curie-Skłodowska Str. 50, 50-369, Wrocław, Poland.
Leiden University Medical Center, Center for Proteomics and Metabolomics, 2300, RC, Leiden, the Netherlands. Electronic address: g.s.m.kammeijer@lumc.nl.
Leiden University Medical Center, Center for Proteomics and Metabolomics, 2300, RC, Leiden, the Netherlands. Electronic address: ndehaan@sund.ku.dk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1052
Author: Wang, X., Baloh, R. H., Milbrandt, J. and Garcia, K. C.
Year: 2006
Title: Structure of artemin complexed with its receptor GFRalpha3: convergent recognition of glial cell line-derived neurotrophic factors
Journal: Structure
Volume: 14
Issue: 6
Pages: 1083-92
Epub Date: 2006/06/13
Date: Jun
Short Title: Structure of artemin complexed with its receptor GFRalpha3: convergent recognition of glial cell line-derived neurotrophic factors
Alternate Journal: Structure (London, England : 1993)
ISSN: 0969-2126 (Print)
0969-2126
DOI: 10.1016/j.str.2006.05.010
Accession Number: 16765900
Keywords: Amino Acid Sequence
Cell Line
Crystallography
Dimerization
Glial Cell Line-Derived Neurotrophic Factor Receptors/agonists/*chemistry/metabolism
Humans
Hydrophobic and Hydrophilic Interactions
Ligands
Molecular Sequence Data
Nerve Growth Factors/*chemistry/metabolism
Nerve Tissue Proteins/*chemistry/metabolism
Neuroglia/metabolism
Protein Conformation
Protein Interaction Mapping
Abstract: Artemin (ARTN) is a member of the glial cell line-derived neurotrophic factor (GDNF) family ligands (GFLs) which regulate the development and maintenance of many neuronal populations in the mammalian nervous system. Here we report the 1.92 A crystal structure of the complex formed between ARTN and its receptor GFRalpha3, which is the initiating step in the formation of a ternary signaling complex containing the shared RET receptor. It represents a new receptor-ligand interaction mode for the TGF-beta superfamily that reveals both conserved and specificity-determining anchor points for all GFL-GFRalpha pairs. In tandem with the complex structure, cellular studies using receptor chimeras implicate dyad-symmetric composite interfaces for recruitment and dimerization of RET, leading to intracellular signaling. These studies should facilitate the functional dissection of the specific versus pleiotropic roles of this system in neurobiology, as well as its exploitation for therapeutic applications.
Notes: Wang, Xinquan
Baloh, Robert H
Milbrandt, Jeffrey
Garcia, K Christopher
AG13730/AG/NIA NIH HHS/United States
NS39358/NS/NINDS NIH HHS/United States
R01 HL077325/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Structure. 2006 Jun;14(6):1083-92. doi: 10.1016/j.str.2006.05.010.
Author Address: Howard Hughes Medical Institute, Stanford University School of Medicine, Department of Microbiology and Immunology, Stanford, California 94305-5124, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1016
Author: Wang, X., Rickert, M. and Garcia, K. C.
Year: 2005
Title: Structure of the quaternary complex of interleukin-2 with its alpha, beta, and gammac receptors
Journal: Science
Volume: 310
Issue: 5751
Pages: 1159-63
Epub Date: 2005/11/19
Date: Nov 18
Short Title: Structure of the quaternary complex of interleukin-2 with its alpha, beta, and gammac receptors
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.1117893
Accession Number: 16293754
Keywords: Crystallography, X-Ray
Humans
Interleukin Receptor Common gamma Subunit
Interleukin-2/*chemistry/metabolism/therapeutic use
Interleukin-2 Receptor alpha Subunit
Interleukin-2 Receptor beta Subunit
Models, Molecular
Mutation
Protein Binding
Protein Conformation
Receptors, Interleukin/*chemistry/metabolism
Receptors, Interleukin-2/*chemistry/genetics/metabolism
Recombinant Proteins/therapeutic use
Severe Combined Immunodeficiency/genetics
Abstract: Interleukin-2 (IL-2) is an immunoregulatory cytokine that acts through a quaternary receptor signaling complex containing alpha (IL-2Ralpha), beta (IL-2Rbeta), and common gamma chain (gc) receptors. In the structure of the quaternary ectodomain complex as visualized at a resolution of 2.3 angstroms, the binding of IL-2Ralpha to IL-2 stabilizes a secondary binding site for presentation to IL-2Rbeta. gammac is then recruited to the composite surface formed by the IL-2/IL-2Rbeta complex. Consistent with its role as a shared receptor for IL-4, IL-7, IL-9, IL-15, and IL-21, gammac forms degenerate contacts with IL-2. The structure of gammac provides a rationale for loss-of-function mutations found in patients with X-linked severe combined immunodeficiency diseases (X-SCID). This complex structure provides a framework for other gammac-dependent cytokine-receptor interactions and for the engineering of improved IL-2 therapeutics.
Notes: 1095-9203
Wang, Xinquan
Rickert, Mathias
Garcia, K Christopher
AI51321/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Science. 2005 Nov 18;310(5751):1159-63. doi: 10.1126/science.1117893.
Author Address: Howard Hughes Medical Institute, Department of Microbiology and Immunology, Stanford University School of Medicine, 299 Campus Drive, Fairchild D319, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1775
Author: Wang, X., Yuan, Z. F., Fan, J., Karch, K. R., Ball, L. E., Denu, J. M. and Garcia, B. A.
Year: 2016
Title: A Novel Quantitative Mass Spectrometry Platform for Determining Protein O-GlcNAcylation Dynamics
Journal: Mol Cell Proteomics
Volume: 15
Issue: 7
Pages: 2462-75
Epub Date: 2016/04/27
Date: Jul
Short Title: A Novel Quantitative Mass Spectrometry Platform for Determining Protein O-GlcNAcylation Dynamics
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.O115.049627
PMCID: PMC4937517
Accession Number: 27114449
Keywords: Boronic Acids/chemistry
Chromatography, Liquid/*methods
Glycosylation
HeLa Cells
Humans
Isotope Labeling
Protein Processing, Post-Translational
Proteome/*analysis/chemistry
Proteomics/*methods
Spectrometry, Mass, Electrospray Ionization/*methods
Abstract: Over the past decades, protein O-GlcNAcylation has been found to play a fundamental role in cell cycle control, metabolism, transcriptional regulation, and cellular signaling. Nevertheless, quantitative approaches to determine in vivo GlcNAc dynamics at a large-scale are still not readily available. Here, we have developed an approach to isotopically label O-GlcNAc modifications on proteins by producing (13)C-labeled UDP-GlcNAc from (13)C6-glucose via the hexosamine biosynthetic pathway. This metabolic labeling was combined with quantitative mass spectrometry-based proteomics to determine protein O-GlcNAcylation turnover rates. First, an efficient enrichment method for O-GlcNAc peptides was developed with the use of phenylboronic acid solid-phase extraction and anhydrous DMSO. The near stoichiometry reaction between the diol of GlcNAc and boronic acid dramatically improved the enrichment efficiency. Additionally, our kinetic model for turnover rates integrates both metabolomic and proteomic data, which increase the accuracy of the turnover rate estimation. Other advantages of this metabolic labeling method include in vivo application, direct labeling of the O-GlcNAc sites and higher confidence for site identification. Concentrating only on nuclear localized GlcNAc modified proteins, we are able to identify 105 O-GlcNAc peptides on 42 proteins and determine turnover rates of 20 O-GlcNAc peptides from 14 proteins extracted from HeLa nuclei. In general, we found O-GlcNAcylation turnover rates are slower than those published for phosphorylation or acetylation. Nevertheless, the rates widely varied depending on both the protein and the residue modified. We believe this methodology can be broadly applied to reveal turnovers/dynamics of protein O-GlcNAcylation from different biological states and will provide more information on the significance of O-GlcNAcylation, enabling us to study the temporal dynamics of this critical modification for the first time.
Notes: 1535-9484
Wang, Xiaoshi
Yuan, Zuo-Fei
Fan, Jing
Karch, Kelly R
Ball, Lauren E
Denu, John M
Garcia, Benjamin A
R01 DE020925/DE/NIDCR NIH HHS/United States
DP2 OD007447/OD/NIH HHS/United States
R01 GM110174/GM/NIGMS NIH HHS/United States
R37 GM059785/GM/NIGMS NIH HHS/United States
T32 GM071339/GM/NIGMS NIH HHS/United States
R01 GM059785/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Mol Cell Proteomics. 2016 Jul;15(7):2462-75. doi: 10.1074/mcp.O115.049627. Epub 2016 Apr 25.
Author Address: From the ‡Epigenetics Program, Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104;
§Department of Biomolecular Chemistry, University of Wisconsin, Madison, Wisconsin 53715;
¶Department of Cell and Molecular Pharmacology, Medical University of South Carolina, Charleston, South Carolina 29425.
From the ‡Epigenetics Program, Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104; bgarci@mail.med.upenn.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 636
Author: Wang, Y., Lee, G. F., Kelley, R. F. and Spellman, M. W.
Year: 1996
Title: Identification of a GDP-L-fucose:polypeptide fucosyltransferase and enzymatic addition of O-linked fucose to EGF domains
Journal: Glycobiology
Volume: 6
Issue: 8
Pages: 837-42
Epub Date: 1996/12/01
Date: Dec
Short Title: Identification of a GDP-L-fucose:polypeptide fucosyltransferase and enzymatic addition of O-linked fucose to EGF domains
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/6.8.837
Accession Number: 9023546
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cricetinae
Epidermal Growth Factor/chemistry/*metabolism
Fucose/*metabolism
Fucosyltransferases/*metabolism
Humans
Membrane Proteins/metabolism
Molecular Sequence Data
Protein Processing, Post-Translational
Rats
Abstract: An assay of GDP-fucose:polypeptide fucosyltransferase has been established. The enzyme catalyzes the reaction that attaches fucose through an O-glycosidic linkage to a conserved serine or threonine residue in EGF domains. The assay uses recombinant human factor VII EGF-1 domain as acceptor substrate and GDP-fucose as donor substrate. Synthetic peptides with sequences taken from five proteins previously shown to contain O-linked fucose (Harris and Spellman, 1993; Glycobiology, 3, 219-224) did not serve as efficient acceptor substrates. These synthetic peptides did not compromise complete EGF domains and did not contain all six cysteine residues that define the EGF structure. Therefore, the enzyme appears to require more than just a consensus primary sequence and likely requires that the EGF domain disulfide bonds be properly formed. The enzymatic reaction showed linear dependency of its activity on time, amount of enzyme, and substrates. Although the enzyme did not exhibit an absolute requirement for Mn2+, enzymatic activity did increase ten fold in the presence of 50 mM MnCl2. The in vitro glycosylation reaction resulted in complete conversion of the acceptor substrate to glycosylated product, and characterization of the purified product by electrospray mass spectrometry revealed that one fucose was added onto the polypeptide. Most of the enzymatic activity was found to be in the soluble fraction of CHO cell homogenates. However, when enzyme was prepared from rat liver in the presence of protease inhibitors, 37% of the activity was recovered by Triton X-100 extraction of the membrane particles after extensive aqueous washes. The result suggests that the enzyme is probably a membrane protein and, by analogy with other glycosyltransferases, probably has a 'stem' region that is very susceptible to proteolysis.
Notes: Wang, Y
Lee, G F
Kelley, R F
Spellman, M W
Journal Article
England
Glycobiology. 1996 Dec;6(8):837-42. doi: 10.1093/glycob/6.8.837.
Author Address: Department of Pharmacokinetics and Metabolism, Genentech, Inc., South San Francisco, CA 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1901
Author: Wang, Y., Liu, J., Jin, X., Zhang, D., Li, D., Hao, F., Feng, Y., Gu, S., Meng, F., Tian, M., Zheng, Y., Xin, L., Zhang, X., Han, X., Aravind, L. and Wei, M.
Year: 2017
Title: O-GlcNAcylation destabilizes the active tetrameric PKM2 to promote the Warburg effect
Journal: Proc Natl Acad Sci U S A
Volume: 114
Issue: 52
Pages: 13732-13737
Epub Date: 2017/12/13
Date: Dec 26
Short Title: O-GlcNAcylation destabilizes the active tetrameric PKM2 to promote the Warburg effect
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1704145115
PMCID: PMC5748163
Accession Number: 29229835
Keywords: Acylation
Cell Line
Female
Glucose/*metabolism
Humans
Lactic Acid/*metabolism
Male
Neoplasm Proteins/*metabolism
Neoplasms/*enzymology/pathology
*Protein Processing, Post-Translational
Pyruvate Kinase/*metabolism
*O-GlcNAcylation
*Pkm2
*Warburg effect
*cancer metabolism
Abstract: The Warburg effect, characterized by increased glucose uptake and lactate production, is a well-known universal across cancer cells and other proliferating cells. PKM2, a splice isoform of the pyruvate kinase (PK) specifically expressed in these cells, serves as a major regulator of this metabolic reprogramming with an adjustable activity subjected to numerous allosteric effectors and posttranslational modifications. Here, we have identified a posttranslational modification on PKM2, O-GlcNAcylation, which specifically targets Thr(405) and Ser(406), residues of the region encoded by the alternatively spliced exon 10 in cancer cells. We show that PKM2 O-GlcNAcylation is up-regulated in various types of human tumor cells and patient tumor tissues. The modification destabilized the active tetrameric PKM2, reduced PK activity, and led to nuclear translocation of PKM2. We also observed that the modification was associated with an increased glucose consumption and lactate production and enhanced level of lipid and DNA synthesis, indicating that O-GlcNAcylation promotes the Warburg effect. In vivo experiments showed that blocking PKM2 O-GlcNAcylation attenuated tumor growth. Thus, we demonstrate that O-GlcNAcylation is a regulatory mechanism for PKM2 in cancer cells and serves as a bridge between PKM2 and metabolic reprogramming typical of the Warburg effect.
Notes: 1091-6490
Wang, Yang
Liu, Jia
Jin, Xin
Zhang, Dapeng
Li, Dongxue
Hao, Fengqi
Feng, Yunpeng
Gu, Shan
Meng, Fanlin
Tian, Miaomiao
Zheng, Yi
Xin, Ling
Zhang, Xinbo
Han, Xue
Aravind, L
Wei, Min
Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2017 Dec 26;114(52):13732-13737. doi: 10.1073/pnas.1704145115. Epub 2017 Dec 11.
Author Address: Key Laboratory of Molecular Epigenetics of the Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, People's Republic of China.
Department of Biology, Saint Louis University, St. Louis, MO 63103.
Program of Bioinformatics and Computational Biology, Saint Louis University, St. Louis, MO 63103.
National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894.
Key Laboratory of Molecular Epigenetics of the Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, People's Republic of China; weim750@nenu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 833
Author: Wang, Y., Xu, A., Knight, C., Xu, L. Y. and Cooper, G. J.
Year: 2002
Title: Hydroxylation and glycosylation of the four conserved lysine residues in the collagenous domain of adiponectin. Potential role in the modulation of its insulin-sensitizing activity
Journal: J Biol Chem
Volume: 277
Issue: 22
Pages: 19521-9
Epub Date: 2002/03/26
Date: May 31
Short Title: Hydroxylation and glycosylation of the four conserved lysine residues in the collagenous domain of adiponectin. Potential role in the modulation of its insulin-sensitizing activity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M200601200
Accession Number: 11912203
Keywords: 3T3 Cells
Adipocytes/metabolism
Adiponectin
Amino Acid Sequence
Animals
COS Cells
Cell Differentiation
Chromatography, High Pressure Liquid
Dose-Response Relationship, Drug
Electrophoresis, Gel, Two-Dimensional
Glucose/metabolism
Glycosylation
Hepatocytes/metabolism
Humans
Insulin/*metabolism
*Intercellular Signaling Peptides and Proteins
Lysine/*chemistry
Male
Mice
Molecular Sequence Data
Peptides/chemistry
Protein Binding
Protein Isoforms
Protein Processing, Post-Translational
Protein Structure, Tertiary
Proteins/*chemistry/metabolism
Rats
Rats, Wistar
Recombinant Proteins/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Transfection
Trypsin/metabolism/pharmacology
Abstract: It has recently been shown that the fat-derived hormone adiponectin has the ability to decrease hyperglycemia and to reverse insulin resistance. However, bacterially produced full-length adiponectin is functionally inactive. Here, we show that endogenous adiponectin secreted by adipocytes is post-translationally modified into eight different isoforms, as shown by two-dimensional gel electrophoresis. Carbohydrate detection revealed that six of the adiponectin isoforms are glycosylated. The glycosylation sites were mapped to several lysines (residues 68, 71, 80, and 104) located in the collagenous domain of adiponectin, each having the surrounding motif of GXKGE(D). These four lysines were found to be hydroxylated and subsequently glycosylated. The glycosides attached to each of these four hydroxylated lysines are possibly glucosylgalactosyl groups. Functional analysis revealed that full-length adiponectin produced by mammalian cells is much more potent than bacterially generated adiponectin in enhancing the ability of subphysiological concentrations of insulin to inhibit gluconeogenesis in primary rat hepatocytes, whereas this insulin-sensitizing ability was significantly attenuated when the four glycosylated lysines were substituted with arginines. These results indicate that full-length adiponectin produced by mammalian cells is functionally active as an insulin sensitizer and that hydroxylation and glycosylation of the four lysines in the collagenous domain might contribute to this activity.
Notes: Wang, Yu
Xu, Aimin
Knight, Catriona
Xu, Lance Yi
Cooper, Garth J S
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2002 May 31;277(22):19521-9. doi: 10.1074/jbc.M200601200. Epub 2002 Mar 23.
Author Address: School of Biological Sciences, University of Auckland, Private Bag 92019, Auckland, 1001 New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 335
Author: Wang, Y. M., Hare, T. R., Won, B., Stowell, C. P., Scanlin, T. F., Glick, M. C., Hård, K., van Kuik, J. A. and Vliegenthart, J. F.
Year: 1990
Title: Additional fucosyl residues on membrane glycoproteins but not a secreted glycoprotein from cystic fibrosis fibroblasts
Journal: Clin Chim Acta
Volume: 188
Issue: 3
Pages: 193-210
Epub Date: 1990/05/01
Date: May
Short Title: Additional fucosyl residues on membrane glycoproteins but not a secreted glycoprotein from cystic fibrosis fibroblasts
Alternate Journal: Clinica chimica acta; international journal of clinical chemistry
ISSN: 0009-8981 (Print)
0009-8981
DOI: 10.1016/0009-8981(90)90201-3
Accession Number: 2387072
Keywords: Carbohydrate Sequence
Chromatography, Affinity
Cystic Fibrosis/*metabolism/pathology
Fibroblasts/*metabolism
Fibronectins/metabolism
Fucose/*metabolism
Glycopeptides/isolation & purification
Humans
Hydroxyapatites
Magnetic Resonance Spectroscopy/methods
Membrane Glycoproteins/isolation & purification/*metabolism
Molecular Sequence Data
Oligosaccharides/metabolism
Abstract: Glycopeptides derived from peripheral membrane glycoproteins of skin fibroblasts of seven patients with cystic fibrosis (CF) had an increase in fucosyl residues when compared with those of seven age, race and sex matched controls (Pediatr Res 1985;19:368-374). To further define these results, the membrane glycopeptides which bound to immobilized lentil lectin and thereby enriched in fucosyl residues linked alpha 1----6 to N-acetylglucosamine attached to asparagine, were Pronase digested, partially purified and examined by 500-MHz 1H-NMR spectroscopy. The CF derived glycopeptides had two different features when compared to those from Controls (1) an increased number of fucosyl residues linked alpha 1----6 to the N-acetylglucosamine attached to asparagine and (2) fucosyl residues linked alpha 1----3 to a branch N-acetylglucosamine. The glycopeptides from both sources were of the di and triantennary type containing sialic acid linked alpha 2----3 and alpha 2----6 to galactose in an approximate molar ratio of 3:2 and 2:1, from CF and Control, respectively. Glycopeptides derived from a glycoprotein, fibronectin, secreted from CF fibroblasts were also examined by 1H-NMR spectroscopy and showed no evidence of fucosyl residues linked alpha 1----3 to branch N-acetylglucosamine and a lesser percentage of core fucose than found in the peripheral membrane glycopeptides. These results define further the altered fucosylation of the CF peripheral membrane glycoproteins.
Notes: Wang, Y M
Hare, T R
Won, B
Stowell, C P
Scanlin, T F
Glick, M C
Hård, K
van Kuik, J A
Vliegenthart, J F
DK 16859/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Netherlands
Clin Chim Acta. 1990 May;188(3):193-210. doi: 10.1016/0009-8981(90)90201-3.
Author Address: Department of Pediatrics, School of Medicine, University of Pennsylvania, Philadelphia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1207
Author: Wang, Z., Park, K., Comer, F., Hsieh-Wilson, L. C., Saudek, C. D. and Hart, G. W.
Year: 2009
Title: Site-specific GlcNAcylation of human erythrocyte proteins: potential biomarker(s) for diabetes
Journal: Diabetes
Volume: 58
Issue: 2
Pages: 309-17
Epub Date: 2008/11/06
Date: Feb
Short Title: Site-specific GlcNAcylation of human erythrocyte proteins: potential biomarker(s) for diabetes
Alternate Journal: Diabetes
ISSN: 0012-1797 (Print)
0012-1797
DOI: 10.2337/db08-0994
PMCID: PMC2628603
Accession Number: 18984734
Keywords: Acetylglucosamine/*metabolism
Biomarkers/blood
Diabetes Mellitus/blood/*diagnosis/metabolism
Erythrocytes/*metabolism
Humans
Immunoblotting
Immunoprecipitation
Mass Spectrometry
Models, Biological
Proteins/*metabolism
Abstract: OBJECTIVE: O-linked N-acetylglucosamine (O-GlcNAc) is upregulated in diabetic tissues and plays a role in insulin resistance and glucose toxicity. Here, we investigated the extent of GlcNAcylation on human erythrocyte proteins and compared site-specific GlcNAcylation on erythrocyte proteins from diabetic and normal individuals. RESEARCH DESIGN AND METHODS: GlcNAcylated erythrocyte proteins or GlcNAcylated peptides were tagged and selectively enriched by a chemoenzymatic approach and identified by mass spectrometry. The enrichment approach was combined with solid-phase chemical derivatization and isotopic labeling to detect O-GlcNAc modification sites and to compare site-specific O-GlcNAc occupancy levels between normal and diabetic erythrocyte proteins. RESULTS: The enzymes that catalyze the cycling (addition and removal) of O-GlcNAc were detected in human erythrocytes. Twenty-five GlcNAcylated erythrocyte proteins were identified. Protein expression levels were compared between diabetic and normal erythrocytes. Thirty-five O-GlcNAc sites were reproducibly identified, and their site-specific O-GlcNAc occupancy ratios were calculated. CONCLUSIONS: GlcNAcylation is differentially regulated at individual sites on erythrocyte proteins in response to glycemic status. These data suggest not only that site-specific O-GlcNAc levels reflect the glycemic status of an individual but also that O-GlcNAc site occupancy on erythrocyte proteins may be eventually useful as a diagnostic tool for the early detection of diabetes.
Notes: 1939-327x
Wang, Zihao
Park, Kyoungsook
Comer, Frank
Hsieh-Wilson, Linda C
Saudek, Christopher D
Hart, Gerald W
R21 DK071280/DK/NIDDK NIH HHS/United States
N01-HV-28180/HV/NHLBI NIH HHS/United States
Howard Hughes Medical Institute/United States
R33 DK071280/DK/NIDDK NIH HHS/United States
N01HV28180/HL/NHLBI NIH HHS/United States
DK-71280/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Diabetes. 2009 Feb;58(2):309-17. doi: 10.2337/db08-0994. Epub 2008 Nov 4.
Author Address: Department of Biological Chemistry, School of Medicine, The Johns Hopkins University, Baltimore, Maryland, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1292
Author: Wang, Z., Udeshi, N. D., Slawson, C., Compton, P. D., Sakabe, K., Cheung, W. D., Shabanowitz, J., Hunt, D. F. and Hart, G. W.
Year: 2010
Title: Extensive crosstalk between O-GlcNAcylation and phosphorylation regulates cytokinesis
Journal: Sci Signal
Volume: 3
Issue: 104
Pages: ra2
Epub Date: 2010/01/14
Date: Jan 12
Short Title: Extensive crosstalk between O-GlcNAcylation and phosphorylation regulates cytokinesis
Alternate Journal: Science signaling
ISSN: 1945-0877
DOI: 10.1126/scisignal.2000526
PMCID: PMC2866299
NIHMSID: NIHMS189253
Accession Number: 20068230
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Antigens, Nuclear/metabolism
Binding Sites/genetics
Blotting, Western
CDC2 Protein Kinase/genetics/*metabolism
Cell Cycle Proteins
Centrosome/metabolism
Cytokinesis/genetics/*physiology
Glycosylation
HeLa Cells
Humans
Models, Biological
Molecular Sequence Data
N-Acetylglucosaminyltransferases/genetics/*metabolism
Neoplasm Proteins/metabolism
Nuclear Matrix-Associated Proteins/metabolism
Nuclear Pore Complex Proteins/metabolism
Nuclear Proteins/metabolism
Phosphorylation
Protein Binding
Proteomics
Repressor Proteins/metabolism
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
Spindle Apparatus/metabolism
Abstract: Like phosphorylation, the addition of O-linked beta-N-acetylglucosamine (O-GlcNAcylation) is a ubiquitous, reversible process that modifies serine and threonine residues on nuclear and cytoplasmic proteins. Overexpression of the enzyme that adds O-GlcNAc to target proteins, O-GlcNAc transferase (OGT), perturbs cytokinesis and promotes polyploidy, but the molecular targets of OGT that are important for its cell cycle functions are unknown. Here, we identify 141 previously unknown O-GlcNAc sites on proteins that function in spindle assembly and cytokinesis. Many of these O-GlcNAcylation sites are either identical to known phosphorylation sites or in close proximity to them. Furthermore, we found that O-GlcNAcylation altered the phosphorylation of key proteins associated with the mitotic spindle and midbody. Forced overexpression of OGT increased the inhibitory phosphorylation of cyclin-dependent kinase 1 (CDK1) and reduced the phosphorylation of CDK1 target proteins. The increased phosphorylation of CDK1 is explained by increased activation of its upstream kinase, MYT1, and by a concomitant reduction in the transcript for the CDK1 phosphatase, CDC25C. OGT overexpression also caused a reduction in both messenger RNA expression and protein abundance of Polo-like kinase 1, which is upstream of both MYT1 and CDC25C. The data not only illustrate the crosstalk between O-GlcNAcylation and phosphorylation of proteins that are regulators of crucial signaling pathways but also uncover a mechanism for the role of O-GlcNAcylation in regulation of cell division.
Notes: 1937-9145
Wang, Zihao
Udeshi, Namrata D
Slawson, Chad
Compton, Philip D
Sakabe, Kaoru
Cheung, Win D
Shabanowitz, Jeffrey
Hunt, Donald F
Hart, Gerald W
R01 GM037537-23/GM/NIGMS NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
S10 RR023025/RR/NCRR NIH HHS/United States
R01 GM037537/GM/NIGMS NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
Journal Article
Sci Signal. 2010 Jan 12;3(104):ra2. doi: 10.1126/scisignal.2000526.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 867
Author: Ward, J. L., Leung, G. P., Toan, S. V. and Tse, C. M.
Year: 2003
Title: Functional analysis of site-directed glycosylation mutants of the human equilibrative nucleoside transporter-2
Journal: Arch Biochem Biophys
Volume: 411
Issue: 1
Pages: 19-26
Epub Date: 2003/02/20
Date: Mar 1
Short Title: Functional analysis of site-directed glycosylation mutants of the human equilibrative nucleoside transporter-2
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1016/s0003-9861(02)00718-x
Accession Number: 12590919
Keywords: Amino Acid Substitution
Animals
Asparagine
Biological Transport
Cell Line
Cloning, Molecular
Equilibrative-Nucleoside Transporter 2/chemistry/*metabolism
Glycosylation
Humans
Kidney
Kinetics
Mutagenesis, Site-Directed
Recombinant Proteins/chemistry/metabolism
Swine
Transfection
Uridine/metabolism
Abstract: Protein glycosylation is important for nucleoside transport, and this has been demonstrated for the human equilibrative nucleoside transporter-1 (hENT1). It is not known whether glycosylation affects the functions of hENT2 or where hENT2 is glycosylated. We address these questions using N-glycosylation mutants (N48D, N57D, and N48/57D) and demonstrate that hENT2 is glycosylated at Asn(48) and Asn(57). Our results show that although the apparent affinities for [3H]uridine and [3H]cytidine of the mutants were indistinguishable from those of the wild-type protein, N-glycosylation was required for efficient targeting of hENT2 to the plasma membrane. All mutants had a two- to threefold increase in IC(50) for dipyridamole. N57D and N48/57D, but not N48D, also had a twofold increase in IC(50) for NBMPR. We conclude that the relative insensitivity of hENT2 to inhibitors is primarily due to its primary structure and not to glycosylation. Glycosylation modulates hENT1 function, but is not required for hENT2.
Notes: Ward, Jeffrey L
Leung, George P H
Toan, Shuy-Vang
Tse, Chung-Ming
K08DK02737/DK/NIDDK NIH HHS/United States
R01CA85428/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Arch Biochem Biophys. 2003 Mar 1;411(1):19-26. doi: 10.1016/s0003-9861(02)00718-x.
Author Address: Department of Medicine, Division of Gastroenterology, The Johns Hopkins University School of Medicine, Ross 925, 720 Rutland Avenune, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2073
Author: Watanabe, Y., Allen, J. D., Wrapp, D., McLellan, J. S. and Crispin, M.
Year: 2020
Title: Site-specific glycan analysis of the SARS-CoV-2 spike
Journal: Science
Volume: 369
Issue: 6501
Pages: 330-333
Epub Date: 2020/05/06
Date: Jul 17
Short Title: Site-specific glycan analysis of the SARS-CoV-2 spike
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.abb9983
PMCID: PMC7199903
Accession Number: 32366695
Keywords: Betacoronavirus/*chemistry
Binding Sites
Covid-19
Coronavirus Infections
Glycopeptides/chemistry/immunology
Glycosylation
Humans
Mass Spectrometry
Models, Molecular
Oligosaccharides/chemistry
Pandemics
Pneumonia, Viral
Polysaccharides/*chemistry
Protein Structure, Tertiary
Recombinant Proteins/chemistry/immunology
SARS-CoV-2
Spike Glycoprotein, Coronavirus/*chemistry/immunology
Abstract: The emergence of the betacoronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), represents a considerable threat to global human health. Vaccine development is focused on the principal target of the humoral immune response, the spike (S) glycoprotein, which mediates cell entry and membrane fusion. The SARS-CoV-2 S gene encodes 22 N-linked glycan sequons per protomer, which likely play a role in protein folding and immune evasion. Here, using a site-specific mass spectrometric approach, we reveal the glycan structures on a recombinant SARS-CoV-2 S immunogen. This analysis enables mapping of the glycan-processing states across the trimeric viral spike. We show how SARS-CoV-2 S glycans differ from typical host glycan processing, which may have implications in viral pathobiology and vaccine design.
Notes: 1095-9203
Watanabe, Yasunori
Orcid: 0000-0002-7952-1268
Allen, Joel D
Orcid: 0000-0003-2547-968x
Wrapp, Daniel
Orcid: 0000-0002-0538-9647
McLellan, Jason S
Orcid: 0000-0003-3991-542x
Crispin, Max
Orcid: 0000-0002-1072-2694
R01 AI127521/AI/NIAID NIH HHS/United States
UM1 AI144462/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2020 Jul 17;369(6501):330-333. doi: 10.1126/science.abb9983. Epub 2020 May 4.
Author Address: School of Biological Sciences, University of Southampton, Southampton SO17 1BJ, UK.
Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.
Division of Structural Biology, University of Oxford, Wellcome Centre for Human Genetics, Oxford OX3 7BN, UK.
Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX 78712, USA.
School of Biological Sciences, University of Southampton, Southampton SO17 1BJ, UK. max.crispin@soton.ac.uk.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 555
Author: Watson, E., Bhide, A. and van Halbeek, H.
Year: 1994
Title: Structure determination of the intact major sialylated oligosaccharide chains of recombinant human erythropoietin expressed in Chinese hamster ovary cells
Journal: Glycobiology
Volume: 4
Issue: 2
Pages: 227-37
Epub Date: 1994/04/01
Date: Apr
Short Title: Structure determination of the intact major sialylated oligosaccharide chains of recombinant human erythropoietin expressed in Chinese hamster ovary cells
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/4.2.227
Accession Number: 8054720
Keywords: Amidohydrolases
Animals
Asparagine
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Ion Exchange
Cricetinae
Erythropoietin/biosynthesis/*chemistry
Humans
Magnetic Resonance Spectroscopy/methods
Molecular Sequence Data
Oligosaccharides/*biosynthesis/*chemistry
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Recombinant Proteins/biosynthesis/*chemistry
Sialic Acids/*analysis
Transfection
Abstract: Recombinant human erythropoietin (rHuEPO) is used abundantly in the clinic to stimulate red blood cell growth in anaemic patients. The efficacy of the drug depends strongly on the extent of sialylation of its carbohydrate moiety. Prompted by conflicting literature reports on the issue, we reinvestigated the structures of the intact sialylated carbohydrate chains of rHuEPO expressed in Chinese hamster ovary (CHO) cells. The asparagine-linked oligosaccharides were released from rHuEPO with N-glycanase and fractionated by anion-exchange chromatography. The O-linked oligosaccharides were released under alkaline borohydride conditions. The primary structures of the major sialylated N- and O-type oligosaccharides were identified by 500-MHz 1H-NMR spectroscopy, supported by data from composition analysis, methylation analysis, low- and high-pH anion-exchange chromatography, and fast atom bombardment-mass spectrometry. The most abundant N-linked oligosaccharides in CHO cell-derived rHuEPO were found to be di-antennary, 2,4-branched tri-antennary, 2,6-branched tri-antennary and tetra-antennary chains (in the ratio of 7:6:5:82), with the latter containing between zero and three repeating N-acetyllactosamine units, in well-defined branches. The major (> 95%) di-, tri- and tetra-antennary structures are fully sialylated, i.e. they have two, three and four sialic acid residues, respectively, linked exclusively alpha(2-->3) to galactose residues. The majority (> 95%) of N-linked structures contain alpha(1-->6)-linked fucose at the proximal GlcNAc residue. The O-type mono- and disialyl oligosaccharides were characterized as a linear tri- and a branched tetra-saccharide, respectively.
Notes: Watson, E
Bhide, A
van Halbeek, H
P41-RR-05351/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1994 Apr;4(2):227-37. doi: 10.1093/glycob/4.2.227.
Author Address: Amgen Inc., Amgen Center, Thousand Oaks, CA 91320-1789.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1125
Author: Watts, J. C., Drisaldi, B., Ng, V., Yang, J., Strome, B., Horne, P., Sy, M. S., Yoong, L., Young, R., Mastrangelo, P., Bergeron, C., Fraser, P. E., Carlson, G. A., Mount, H. T., Schmitt-Ulms, G. and Westaway, D.
Year: 2007
Title: The CNS glycoprotein Shadoo has PrP(C)-like protective properties and displays reduced levels in prion infections
Journal: Embo j
Volume: 26
Issue: 17
Pages: 4038-50
Epub Date: 2007/08/19
Date: Sep 5
Short Title: The CNS glycoprotein Shadoo has PrP(C)-like protective properties and displays reduced levels in prion infections
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/sj.emboj.7601830
PMCID: PMC1950727
Accession Number: 17703189
Keywords: Amino Acid Sequence
Animals
Brain/*metabolism
Cell Line, Tumor
Cerebellum/metabolism
GPI-Linked Proteins
Glycoproteins/genetics/*physiology
Hippocampus/metabolism
Mice
Molecular Sequence Data
Nerve Tissue Proteins/genetics/*metabolism
Neurons/*metabolism
PrPC Proteins/*metabolism
Prion Diseases/*metabolism
Prions/*metabolism
Protein Binding
Abstract: The cellular prion protein, PrP(C), is neuroprotective in a number of settings and in particular prevents cerebellar degeneration mediated by CNS-expressed Doppel or internally deleted PrP ('DeltaPrP'). This paradigm has facilitated mapping of activity determinants in PrP(C) and implicated a cryptic PrP(C)-like protein, 'pi'. Shadoo (Sho) is a hypothetical GPI-anchored protein encoded by the Sprn gene, exhibiting homology and domain organization similar to the N-terminus of PrP. Here we demonstrate Sprn expression and Sho protein in the adult CNS. Sho expression overlaps PrP(C), but is low in cerebellar granular neurons (CGNs) containing PrP(C) and high in PrP(C)-deficient dendritic processes. In Prnp(0/0) CGNs, Sho transgenes were PrP(C)-like in their ability to counteract neurotoxic effects of either Doppel or DeltaPrP. Additionally, prion-infected mice exhibit a dramatic reduction in endogenous Sho protein. Sho is a candidate for pi, and since it engenders a PrP(C)-like neuroprotective activity, compromised neuroprotective activity resulting from reduced levels may exacerbate damage in prion infections. Sho may prove useful in deciphering several unresolved facets of prion biology.
Notes: 1460-2075
Watts, Joel C
Drisaldi, Bettina
Ng, Vivian
Yang, Jing
Strome, Bob
Horne, Patrick
Sy, Man-Sun
Yoong, Larry
Young, Rebecca
Mastrangelo, Peter
Bergeron, Catherine
Fraser, Paul E
Carlson, George A
Mount, Howard T J
Schmitt-Ulms, Gerold
Westaway, David
Journal Article
Research Support, Non-U.S. Gov't
EMBO J. 2007 Sep 5;26(17):4038-50. doi: 10.1038/sj.emboj.7601830. Epub 2007 Aug 16.
Author Address: Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 284
Author: Watzlawick, H., Walsh, M. T., Ehrhard, I., Slayter, H. S., Haupt, H., Schwick, H. G., Jourdian, G. W., Hase, S., Schmid, K. and Brossmer, R.
Year: 1991
Title: The effect of the carbohydrate moiety upon the size and conformation of human plasma galactoglycoprotein as judged by electron microscopy and circular dichroism. Structural studies of a glycoprotein after stepwise enzymic carbohydrate removal
Journal: Biochem J
Volume: 277 ( Pt 3)
Issue: Pt 3
Pages: 753-8
Epub Date: 1991/08/01
Date: Aug 1
Short Title: The effect of the carbohydrate moiety upon the size and conformation of human plasma galactoglycoprotein as judged by electron microscopy and circular dichroism. Structural studies of a glycoprotein after stepwise enzymic carbohydrate removal
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2770753
PMCID: PMC1151308
Accession Number: 1872812
Keywords: *Antigens, CD
Blood Proteins/*chemistry
Carbohydrate Sequence
Circular Dichroism
Glycoproteins/*blood/*chemistry
Glycoside Hydrolases/pharmacology
Humans
Leukosialin
Microscopy, Electron
Molecular Sequence Data
Protein Conformation
*Sialoglycoproteins
Abstract: Galactoglycoprotein is a unique human plasma protein [76% carbohydrate (23% N-acetylneuraminic acid, 20% galactose, 3% mannose, 1% fucose and 29% N-acetylgalactosamine plus N-acetylglucosamine) and 24% polypeptide, a single polypeptide chain of about 200 amino acid residues that is high in serine and threonine content] [Schmid, Mao, Kimura, Hayashi & Binette (1980) J. Biol. Chem. 255, 3221-3226]. Highly purified exoglycosidases with well-defined specificities were used to prepare five derivatives of galactoglycoprotein in which sequential residues of N-acetylneuraminic acid, galactose, N-acetylglucosamine, a second galactose and N-acetylgalactosamine were removed with 83% of the total carbohydrate cleaved. C.d. shows that native galactoglycoprotein and all derivatives in aqueous buffer are predominantly random coil, suggesting that removal of a large number of electrostatic net charges, as well as the major portion of the carbohydrate moiety, does not alter the secondary structure of the polypeptide chain. Examination of the size and conformation of tungsten-shadowed galactoglycoprotein and asialo and agalacto derivatives by electron microscopy shows the size and conformation of all three preparations to be similar, with only minor differences in particle length and width.
Notes: 1470-8728
Watzlawick, H
Walsh, M T
Ehrhard, I
Slayter, H S
Haupt, H
Schwick, H G
Jourdian, G W
Hase, S
Schmid, K
Brossmer, R
AR-10531/AR/NIAMS NIH HHS/United States
GM-14237/GM/NIGMS NIH HHS/United States
HL-26335/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Biochem J. 1991 Aug 1;277 ( Pt 3)(Pt 3):753-8. doi: 10.1042/bj2770753.
Author Address: Institut für Biochemie II, University of Heidelberg, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 245
Author: Watzlawick, H., Walsh, M. T., Yoshioka, Y., Schmid, K. and Brossmer, R.
Year: 1992
Title: Structure of the N- and O-glycans of the A-chain of human plasma alpha 2HS-glycoprotein as deduced from the chemical compositions of the derivatives prepared by stepwise degradation with exoglycosidases
Journal: Biochemistry
Volume: 31
Issue: 48
Pages: 12198-203
Epub Date: 1992/12/08
Date: Dec 8
Short Title: Structure of the N- and O-glycans of the A-chain of human plasma alpha 2HS-glycoprotein as deduced from the chemical compositions of the derivatives prepared by stepwise degradation with exoglycosidases
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00163a032
Accession Number: 1457416
Keywords: Blood Proteins/*analysis
Carbohydrate Sequence
Glycoside Hydrolases/metabolism
Humans
Hydrolysis
Molecular Sequence Data
Polysaccharides/*chemistry
alpha-2-HS-Glycoprotein
Abstract: The structure of the glycans of the A-chain of human plasma alpha 2HS-glycoprotein was established from the chemical compositions of its derivatives prepared by sequential enzymatic degradation of the carbohydrate moiety, from the determination of the kind and amount of the monosaccharides liberated after each step of the enzymatic digestion, and from the distinct specificity of the highly purified exoglycosidases. The exoglycosidases were three sialidases (Vibrio cholerae, fowl plague virus, and Arthrobacter ureafaciens), two beta-galactosidases (Streptococcus pneumoniae and bovine testis), one alpha-N-acetylgalactosaminidase, one beta-N-acetylglucosaminidase, and one alpha-mannosidase. Utilizing sialidases with different cleavage specificities, the number of alpha 2-3- and alpha 2-6-linked sialic acid residues could be separately determined. As to the beta-galactosidases, the enzyme isolated from S. pneumoniae cleaves only beta 1-4-linked galactose residues, whereas the bovine testes enzyme acts on both the beta 1-4- and beta 1-3-linked galactose residues. Jack bean beta-N-acetylglucosaminidase cleaves beta 1-2, beta 1-4, and beta 1-6 GlcNAc with higher activity for the beta 1-2. Jack bean alpha-mannosidase cleaves alpha 1-2, alpha 1-6, and alpha 1-3 Man with greater activity for alpha 1-2 and alpha 1-6. Bovine liver alpha-N-acetylgalactosaminidase cleaves O-linked GalNAc. On the basis of these results, the A-chain of alpha 2 HS-glycoprotein was found to possess two biantennary N-glycans and two O-linked trisaccharides.
Notes: Watzlawick, H
Walsh, M T
Yoshioka, Y
Schmid, K
Brossmer, R
GM-10374/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1992 Dec 8;31(48):12198-203. doi: 10.1021/bi00163a032.
Author Address: Institut für Biochemie II, Universität Heidelberg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 941
Author: Weber, A. N., Morse, M. A. and Gay, N. J.
Year: 2004
Title: Four N-linked glycosylation sites in human toll-like receptor 2 cooperate to direct efficient biosynthesis and secretion
Journal: J Biol Chem
Volume: 279
Issue: 33
Pages: 34589-94
Epub Date: 2004/06/03
Date: Aug 13
Short Title: Four N-linked glycosylation sites in human toll-like receptor 2 cooperate to direct efficient biosynthesis and secretion
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M403830200
Accession Number: 15173186
Keywords: Amino Acid Sequence
Animals
Binding Sites
Cell Line
Circular Dichroism
Computational Biology
Databases as Topic
Glycoside Hydrolases/metabolism
Glycosylation
Humans
Insecta
Leucine/chemistry
Membrane Glycoproteins/*biosynthesis/*chemistry/metabolism
Models, Molecular
Molecular Sequence Data
Mutagenesis
Mutagenesis, Site-Directed
Mutation
Platelet Glycoprotein GPIb-IX Complex/chemistry
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Receptors, Cell Surface/*biosynthesis/*chemistry/metabolism
Recombinant Proteins/chemistry
Sequence Homology, Amino Acid
Toll-Like Receptor 2
Toll-Like Receptor 4
Toll-Like Receptors
Transfection
Abstract: Most higher organisms have a system of innate immune defense that is mediated by a group of evolutionarily related, germ line-encoded receptors, so-called Toll-like receptors. In mammals Toll-like receptors signal in response to pathogen-associated microbial structures. For example, Toll-like receptor 2 appears to mediate responses to bacterial peptidoglycan and acylated lipoproteins and Toll-like receptor 4 to bacterial lipopolysaccharide. However, the structural principles that underlie recognition of these structures are poorly understood. Toll-like receptors have leucine-rich repeats in their extracellular domains and are thus believed to adopt solenoid structures, similar to that found in platelet glycoprotein Ib. Additionally, all Toll-like receptors contain N-linked glycosylation consensus sites, and Toll-like receptor 4 requires glycosylation for function. Toll-like receptor glycosylation is also likely to influence receptor surface representation, trafficking, and pattern recognition. Using circular dichroism spectroscopy, we show here that purified human Toll-like receptor 2 and 4 proteins have secondary structure contents similar to glycoprotein Ib. We have also analyzed where consensus glycosylation sites are located in the extracellular domains of other human Toll-like receptors. We found that there are significant differences in the location and degree of conservation between sites in different Toll-like receptors. Using site-directed mutagenesis, we have found that in Toll-like receptor 2 extracellular domain all four predicted glycosylation sites are substituted, although one site is inefficiently core-glycosylated and its removal drastically affects secretion. The remaining Toll-like receptor 2 glycosylation sites also contribute to efficient protein secretion, albeit to a lesser degree.
Notes: Weber, Alexander N R
Morse, Mary A
Gay, Nicholas J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2004 Aug 13;279(33):34589-94. doi: 10.1074/jbc.M403830200. Epub 2004 Jun 1.
Author Address: RA Pharmacology, RI CEDD, GlaxoSmithKline Medicines Research Centre, Stevenage SG1 2NY, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1852
Author: Wedepohl, S., Dernedde, J., Vahedi-Faridi, A., Tauber, R., Saenger, W. and Bulut, H.
Year: 2017
Title: Reducing Macro- and Microheterogeneity of N-Glycans Enables the Crystal Structure of the Lectin and EGF-Like Domains of Human L-Selectin To Be Solved at 1.9 Å Resolution
Journal: Chembiochem
Volume: 18
Issue: 13
Pages: 1338-1345
Epub Date: 2017/05/11
Date: Jul 4
Short Title: Reducing Macro- and Microheterogeneity of N-Glycans Enables the Crystal Structure of the Lectin and EGF-Like Domains of Human L-Selectin To Be Solved at 1.9 Å Resolution
Alternate Journal: Chembiochem : a European journal of chemical biology
ISSN: 1439-4227
DOI: 10.1002/cbic.201700220
Accession Number: 28489325
Keywords: Binding Sites
Calcium/*chemistry/metabolism
Cloning, Molecular
Crystallography, X-Ray
Epidermal Growth Factor/*chemistry/genetics/metabolism
Gene Expression
Glycosylation
HEK293 Cells
Humans
L-Selectin/*chemistry/genetics/metabolism
Lectins, C-Type/*chemistry/genetics/metabolism
Models, Molecular
Mutation
Polysaccharides/*chemistry/metabolism
Protein Binding
Protein Domains
Protein Interaction Domains and Motifs
Protein Structure, Secondary
Recombinant Proteins/chemistry/genetics/metabolism
*L-selectin
*crystal growth
*glycoproteins
*glycosylation
*mutagenesis
Abstract: L-Selectin, a cell-adhesion receptor on the surface of most leukocytes, contains seven N-glycosylation sites. In order to obtain the crystal structure of human L-selectin, we expressed a shortened version of L-selectin comprising the C-type lectin and EGF-like domains (termed LE) and systematically analysed mutations of the three glycosylation sites (Asn22, Asn66 and Asn139) in order to reduce macroheterogeneity. After we further removed microheterogeneity, we obtained crystals that diffracted X-rays up to 1.9 Å from a variant (LE010) with exchanges N22Q and N139Q and one GlcNAc(2) Man(5) N-glycan chain attached to Asn66. Crystal-structure analysis showed that the terminal mannose of GlcNAc(2) Man(5) of one LE010 molecule was coordinated to Ca(2+) in the binding site of a symmetry-related LE010. The orientation of the lectin and EGF-like domain was similar to the described "bent" conformation of E- and P-selectins. The Ca(2+) -binding site reflects the binding mode seen in E- and P-selectin structures co-crystallised with ligands.
Notes: 1439-7633
Wedepohl, Stefanie
Orcid: 0000-0001-5355-8137
Dernedde, Jens
Orcid: 0000-0001-5319-7677
Vahedi-Faridi, Ardeschir
Tauber, Rudolf
Saenger, Wolfram
Bulut, Haydar
Journal Article
Research Support, Non-U.S. Gov't
Germany
Chembiochem. 2017 Jul 4;18(13):1338-1345. doi: 10.1002/cbic.201700220. Epub 2017 Jun 6.
Author Address: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité Universitätsmedizin Berlin, CVK, Augustenburger Platz 1, 13353, Berlin, Germany.
Institut für Chemie und Biochemie/Kristallographie, Freie Universität Berlin, Takustrasse 6, 14195, Berlin, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1087
Author: Wehrman, T., He, X., Raab, B., Dukipatti, A., Blau, H. and Garcia, K. C.
Year: 2007
Title: Structural and mechanistic insights into nerve growth factor interactions with the TrkA and p75 receptors
Journal: Neuron
Volume: 53
Issue: 1
Pages: 25-38
Epub Date: 2007/01/02
Date: Jan 4
Short Title: Structural and mechanistic insights into nerve growth factor interactions with the TrkA and p75 receptors
Alternate Journal: Neuron
ISSN: 0896-6273 (Print)
0896-6273
DOI: 10.1016/j.neuron.2006.09.034
Accession Number: 17196528
Keywords: Animals
Binding Sites/physiology
Cell Line
Cell Membrane/*metabolism/ultrastructure
Dimerization
Extracellular Space/metabolism
Humans
Ligands
Macromolecular Substances/chemistry/metabolism
Mice
Models, Molecular
Nerve Growth Factor/*chemistry/metabolism
Neurons/*metabolism/ultrastructure
Protein Structure, Tertiary/physiology
Receptor, Nerve Growth Factor/*chemistry/metabolism
Receptor, trkA/*chemistry/metabolism
Signal Transduction/physiology
Abstract: Nerve growth factor engages two structurally distinct transmembrane receptors, TrkA and p75, which have been proposed to create a "high-affinity" NGF binding site through formation of a ternary TrkA/NGF/p75 complex. To define a structural basis for the high-affinity site, we have determined the three-dimensional structure of a complete extracellular domain of TrkA complexed with NGF. The complex reveals a crab-shaped homodimeric TrkA structure, but a mechanism for p75 coordination is not obvious. We investigated the heterodimerization of membrane-bound TrkA and p75, on intact mammalian cells, using a beta-gal protein-protein interaction system. We find that NGF dimerizes TrkA and that p75 exists on the cell surface as a preformed oligomer that is not dissociated by NGF. We find no evidence for a direct TrkA/p75 interaction. We propose that TrkA and p75 likely communicate through convergence of downstream signaling pathways and/or shared adaptor molecules, rather than through direct extracellular interactions.
Notes: Wehrman, Tom
He, Xiaolin
Raab, Bill
Dukipatti, Abhiram
Blau, Helen
Garcia, K Christopher
AG009521/AG/NIA NIH HHS/United States
AG020961/AG/NIA NIH HHS/United States
AG024987/AG/NIA NIH HHS/United States
EB005011/EB/NIBIB NIH HHS/United States
HD018179/HD/NICHD NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
United States
Neuron. 2007 Jan 4;53(1):25-38. doi: 10.1016/j.neuron.2006.09.034.
Author Address: Baxter Laboratory for Genetic Pharmacology, Department of Microbiology and Immunology, The Stem Cell Institute, Stanford University School of Medicine, Stanford, CA 94305, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1663
Author: Wei, H., Wang, D., Qian, Y., Liu, X., Fan, S., Yin, H. S. and Wang, X.
Year: 2014
Title: Structural basis for the specific recognition of IL-18 by its alpha receptor
Journal: FEBS Lett
Volume: 588
Issue: 21
Pages: 3838-43
Epub Date: 2014/09/28
Date: Nov 3
Short Title: Structural basis for the specific recognition of IL-18 by its alpha receptor
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1016/j.febslet.2014.09.019
Accession Number: 25261253
Keywords: Animals
Crystallography, X-Ray
Humans
Interleukin-1/metabolism
Interleukin-18/*chemistry/*metabolism
Interleukin-18 Receptor alpha Subunit/*chemistry/*metabolism
Ligands
Models, Molecular
Protein Binding
Protein Structure, Tertiary
Sf9 Cells
Spodoptera
Substrate Specificity
Surface Properties
Interleukin 18
Interleukin 18 receptor
Ligand-receptor recognition
X-ray structure
Abstract: Interleukin 18 (IL-18), a member of the IL-1 family of cytokines, is an important regulator of innate and acquired immune responses. It signals through its ligand-binding primary receptor IL-18Rα and accessory receptor IL-18Rβ. Here we report the crystal structure of IL-18 with the ectodomain of IL-18Rα, which reveals the structural basis for their specific recognition. It confirms that surface charge complementarity determines the ligand-binding specificity of primary receptors in the IL-1 receptor family. We suggest that IL-18 signaling complex adopts an architecture similar to other agonistic cytokines and propose a general ligand-receptor assembly and activation model for the IL-1 family.
Notes: 1873-3468
Wei, Hui
Wang, Dongli
Qian, Yun
Liu, Xi
Fan, Shilong
Yin, Hsien-Sheng
Wang, Xinquan
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2014 Nov 3;588(21):3838-43. doi: 10.1016/j.febslet.2014.09.019. Epub 2014 Sep 26.
Author Address: Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, Collaborative Innovation Center for Biotherapy, School of Life Sciences, Tsinghua University, Beijing 100084, China; Collaborative Innovation Center for Biotherapy, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, West China Medical School, Sichuan University, Chengdu, China.
Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, Collaborative Innovation Center for Biotherapy, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Institute of Bioinformatics and Structural Biology, National Tsing Hua University, Hsinchu, Taiwan.
Ministry of Education Key Laboratory of Protein Science, Center for Structural Biology, Collaborative Innovation Center for Biotherapy, School of Life Sciences, Tsinghua University, Beijing 100084, China; Collaborative Innovation Center for Biotherapy, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, West China Medical School, Sichuan University, Chengdu, China. Electronic address: xinquanwang@mail.tsinghua.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2005
Author: Wei, Y., Xiong, Z. J., Li, J., Zou, C., Cairo, C. W., Klassen, J. S. and Privé, G. G.
Year: 2019
Title: Crystal structures of human lysosomal EPDR1 reveal homology with the superfamily of bacterial lipoprotein transporters
Journal: Commun Biol
Volume: 2
Pages: 52
Epub Date: 2019/02/08
Short Title: Crystal structures of human lysosomal EPDR1 reveal homology with the superfamily of bacterial lipoprotein transporters
Alternate Journal: Communications biology
ISSN: 2399-3642
DOI: 10.1038/s42003-018-0262-9
PMCID: PMC6363788
Accession Number: 30729188
Keywords: Amino Acid Sequence
Animals
Baculoviridae/genetics/metabolism
Binding Sites
Cloning, Molecular
Crystallography, X-Ray
Escherichia coli/genetics/*metabolism
Escherichia coli Proteins/*chemistry/genetics/metabolism
G(M1) Ganglioside/*chemistry/metabolism
Gene Expression
Humans
Hydrophobic and Hydrophilic Interactions
Lysophospholipids/*chemistry/metabolism
Lysosomes/metabolism
Models, Molecular
Monoglycerides/*chemistry/metabolism
Neoplasm Proteins/*chemistry/genetics/metabolism
Periplasmic Binding Proteins/*chemistry/genetics/metabolism
Phylogeny
Plants/genetics/metabolism
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Folding
Protein Interaction Domains and Motifs
Protein Multimerization
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: EPDR1, a member of the ependymin-related protein family, is a relatively uncharacterized protein found in the lysosomes and secretomes of most vertebrates. Despite having roles in human disease and health, the molecular functions of EPDR1 remain unknown. Here, we present crystal structures of human EPDR1 and reveal that the protein adopts a fold previously seen only in bacterial proteins related to the LolA lipoprotein transporter. EPDR1 forms a homodimer with an overall shape resembling a half-shell with two non-overlapping hydrophobic grooves on the flat side of the hemisphere. EPDR1 can interact with membranes that contain negatively charged lipids, including BMP and GM1, and we suggest that EPDR1 may function as a lysosomal activator protein or a lipid transporter. A phylogenetic analysis reveals that the fold is more widely distributed than previously suspected, with representatives identified in all branches of cellular life.
Notes: 2399-3642
Wei, Yong
Orcid: 0000-0002-3314-118x
Xiong, Zi Jian
Orcid: 0000-0002-7170-0472
Li, Jun
Zou, Chunxia
Cairo, Christopher W
Orcid: 0000-0003-3363-8708
Klassen, John S
Privé, Gilbert G
Orcid: 0000-0002-0712-4319
CIHR/Canada
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Commun Biol. 2019 Feb 5;2:52. doi: 10.1038/s42003-018-0262-9. eCollection 2019.
Author Address: 1Princess Margaret Cancer Centre, Toronto, M5G 1L7 ON Canada.
2Department of Biochemistry, University of Toronto, Toronto, M5S 1A8 ON Canada.
3Alberta Glycomics Centre and Department of Chemistry, University of Alberta, Edmonton, T6G 2G2 AB Canada.
4Department of Medical Biophysics, University of Toronto, Toronto, M5G 1L7 ON Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1512
Author: Wei, Z., Lei, X., Seldin, M. M. and Wong, G. W.
Year: 2012
Title: Endopeptidase cleavage generates a functionally distinct isoform of C1q/tumor necrosis factor-related protein-12 (CTRP12) with an altered oligomeric state and signaling specificity
Journal: J Biol Chem
Volume: 287
Issue: 43
Pages: 35804-14
Epub Date: 2012/09/04
Date: Oct 19
Short Title: Endopeptidase cleavage generates a functionally distinct isoform of C1q/tumor necrosis factor-related protein-12 (CTRP12) with an altered oligomeric state and signaling specificity
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.365965
PMCID: PMC3476250
Accession Number: 22942287
Keywords: 3T3-L1 Cells
Adipocytes/cytology/*metabolism
Adipokines/genetics/*metabolism
Amino Acid Substitution
Animals
Chickens
Furin/genetics/*metabolism
Glucose/genetics/metabolism
Hepatocytes/cytology/metabolism
Humans
Insulin/genetics/*metabolism
MAP Kinase Signaling System/*physiology
Mice
Mitogen-Activated Protein Kinase Kinases/genetics/metabolism
Mutation, Missense
Protein Isoforms/genetics/metabolism
Protein Multimerization/physiology
Protein Processing, Post-Translational/*physiology
*Proteolysis
Xenopus
Zebrafish
Abstract: Adipose tissue-derived adipokines are an important class of secreted metabolic regulators that mediate tissue cross-talk to control systemic energy balance. We recently described C1q/TNF-related protein-12 (CTRP12), a novel insulin-sensitizing adipokine that regulates glucose metabolism in liver and adipose tissue. However, the biochemical properties of CTRP12 and its naturally occurring cleaved isoform have not been characterized. Here, we show that CTRP12 is a secreted hormone subjected to multiple functionally relevant posttranslational modifications at highly conserved residues. For example, Asn(39) is glycosylated, whereas Cys(85) mediates the assembly of higher order oligomeric structure. Endopeptidase cleavage at Lys(91) generates a cleaved globular gCTRP12 isoform, the expression of which is increased by insulin. PCSK3/furin was identified as the major proprotein convertase expressed by adipocytes that mediates the endogenous cleavage of CTRP12. Cleavage at Lys(91) is context-dependent: mutation of the charged Arg(93) to Ala on the P2' position enhanced cleavage, and triple mutations (K90A/K91A/R93A) abolished cleavage. Importantly, the two isoforms of CTRP12 differ in oligomeric structures and are functionally distinct. The full-length protein forms trimers and larger complexes, and the cleaved isoform consisted of predominantly dimers. Whereas full-length fCTRP12 strongly activated Akt signaling in H4IIE hepatocytes and 3T3-L1 adipocytes, gCTRP12 preferentially activated MAP kinase (ERK1/2 and p38 MAPK) signaling. Further, only fCTRP12 improved insulin-stimulated glucose uptake in adipocytes. These results reveal a novel mechanism controlling signaling specificity and function of a hormone via cleavage-dependent alteration in oligomeric state.
Notes: 1083-351x
Wei, Zhikui
Lei, Xia
Seldin, Marcus M
Wong, G William
R01 DK084171/DK/NIDDK NIH HHS/United States
DK084171/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Oct 19;287(43):35804-14. doi: 10.1074/jbc.M112.365965. Epub 2012 Aug 31.
Author Address: Department of Physiology and Center for Metabolism and Obesity Research, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1249
Author: Wei, Z., Yan, Y., Carrell, R. W. and Zhou, A.
Year: 2009
Title: Crystal structure of protein Z-dependent inhibitor complex shows how protein Z functions as a cofactor in the membrane inhibition of factor X
Journal: Blood
Volume: 114
Issue: 17
Pages: 3662-7
Epub Date: 2009/06/17
Date: Oct 22
Short Title: Crystal structure of protein Z-dependent inhibitor complex shows how protein Z functions as a cofactor in the membrane inhibition of factor X
Alternate Journal: Blood
ISSN: 0006-4971 (Print)
0006-4971
DOI: 10.1182/blood-2009-04-210021
PMCID: PMC2766681
Accession Number: 19528533
Keywords: Allosteric Site
Binding Sites
Blood Coagulation
Blood Proteins/*chemistry
Calcium/metabolism
Circular Dichroism
Crystallization
Crystallography, X-Ray
Factor X/*antagonists & inhibitors
Factor Xa/metabolism
Humans
Membranes/*metabolism
Phospholipids/metabolism
Protease Inhibitors/chemistry/pharmacology
Protein Binding
Protein Conformation
Recombinant Proteins/chemistry
Serpins/*chemistry
Abstract: Protein Z (PZ) binds to PZ-dependent inhibitor (ZPI) and accelerates the inhibition of the coagulation protease, activated factor X (FXa), in the presence of phospholipids and Ca2+. A 2.3A resolution crystal structure of PZ complexed with ZPI shows that ZPI is a typical serine protease inhibitor and that PZ has a serine protease fold with distorted oxyanion hole and S1 pocket. The 2 molecules bind with fully complementary surfaces spanning over 2400A(2) and involving extensive ionic and hydrophobic interactions. ZPI has an unusual shutter region with a negatively charged residue buried within the hydrophobic core of the molecule. This unique Asp(213) is critical in maintaining the balanced metastability required for optimal protease inhibition, especially when PZ is bound, with its replacement with Asn resulting in increased thermal stability, but decreased efficiency of protease inhibition. The structure of ZPI shows negatively and positively charged surfaces on top of the molecule, in keeping with mutagenesis studies in this work indicating exosite interactions with FXa when it docks on top of ZPI. As modeled in this study, the gamma-carboxy-glutamic acid-containing domains of PZ and FXa enable them to bind to the same phospholipid surfaces on platelet and other membranes, with optimal proximity for the inhibition of FXa by the complexed ZPI.
Notes: 1528-0020
Wei, Zhenquan
Yan, Yahui
Carrell, Robin W
Zhou, Aiwu
British Heart Foundation/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Blood. 2009 Oct 22;114(17):3662-7. doi: 10.1182/blood-2009-04-210021. Epub 2009 Jun 15.
Author Address: Department of Haematology, University of Cambridge, Cambridge Institute for Medical Research, Cambridge, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 282
Author: Weisshaar, G., Hiyama, J. and Renwick, A. G.
Year: 1991
Title: Site-specific N-glycosylation of human chorionic gonadotrophin--structural analysis of glycopeptides by one- and two-dimensional 1H NMR spectroscopy
Journal: Glycobiology
Volume: 1
Issue: 4
Pages: 393-404
Epub Date: 1991/09/01
Date: Sep
Short Title: Site-specific N-glycosylation of human chorionic gonadotrophin--structural analysis of glycopeptides by one- and two-dimensional 1H NMR spectroscopy
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/1.4.393
Accession Number: 1820200
Keywords: Acetylglucosamine/metabolism
Amino Acid Sequence
Asparagine/metabolism
Carbohydrate Sequence
Chorionic Gonadotropin/*chemistry/metabolism
Fucose/metabolism
Glycopeptides/*chemistry/isolation & purification
Glycosylation
Humans
Macromolecular Substances
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/*chemistry
Abstract: Glycopeptides representing individual N-glycosylation sites of the heterodimeric glycoprotein hormone human chorionic gonadotrophin (hCG) were obtained from subunits hCG alpha (N-glycosylated at Asn-52 and Asn-78) and hCG beta (N-glycosylated at Asn-13 and Asn-30) by digestion with trypsin and chymotrypsin, respectively. Following purification by reverse-phase HPLC and identification by amino acid sequencing, the glycopeptides were analysed by one- and two-dimensional 1H NMR spectroscopy. The results are summarized as follows: (i) oligosaccharides attached to Asn-52 of hCG alpha comprised monosialylated 'monoantenary' NeuAc alpha 2-3Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3[Man alpha 1-6]Man beta 1-4GlcNAc beta 1-4GlcNAc (N1-4'), disialylated diantennary NeuAc alpha 2-3Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3[NeuAc alpha 2-3-Gal beta 1-4GlcNAc beta 1-2Man alpha 1-6]Man beta 1-4GlcNAc beta 1-4GlcNAc (N2), and the monosialylated hybrid-type structures NeuAc alpha 2-3Gal beta 1-4GlcNAc beta 1-2Man alpha 1-3[Man alpha 1-3Man alpha 1-6]Man beta 1-4GlcNAc beta 1-4GlcNAc (N1-A) and NeuAc alpha 2-3Gal-beta 1-4GlcNAc beta 1-2Man alpha 1-3[Man alpha 1-3(Man alpha 1-6)Man alpha 1-6]Man beta 1-4GlcNAc beta 1-4GlcNAc (N1-AB) in a ratio approaching 5:2:2:1; (ii) Asn-78 of hCG alpha carried N2 and N1-4' almost exclusively (ratio approximately 3:2); (iii) both N-glycosylation sites of hCG beta contained predominantly component N2, partially (approximately 25%) and completely alpha 1-6-fucosylated at the N-acetylglucosamine linked to Asn-13 and Asn-30, respectively. The distinct site-specific distribution of the oligosaccharide structures among individual N-glycosylation sites of hCG appears to reflect primarily the influence of the surrounding protein structure on the substrate accessibility of the Golgi processing enzymes alpha-mannosidase II, GlcNAc transferase II and alpha 1,6-fucosyltransferase.
Notes: Weisshaar, G
Hiyama, J
Renwick, A G
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1991 Sep;1(4):393-404. doi: 10.1093/glycob/1.4.393.
Author Address: Department of Biochemistry, University of Auckland, New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 297
Author: Weisshaar, G., Hiyama, J., Renwick, A. G. and Nimtz, M.
Year: 1991
Title: NMR investigations of the N-linked oligosaccharides at individual glycosylation sites of human lutropin
Journal: Eur J Biochem
Volume: 195
Issue: 1
Pages: 257-68
Epub Date: 1991/01/01
Date: Jan 1
Short Title: NMR investigations of the N-linked oligosaccharides at individual glycosylation sites of human lutropin
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1991.tb15702.x
Accession Number: 1991473
Keywords: Amino Acid Sequence
Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, High Pressure Liquid
Glycosylation
Humans
Luteinizing Hormone/*chemistry
Magnetic Resonance Spectroscopy/methods
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Abstract: Human lutropin or luteinizing hormone (hLH) is a heterodimeric glycoprotein, composed of two subunits. hLH alpha (N-glycosylated at Asn52 and Asn78) and hLH beta (N-glycosylated at Asn30). The sugar chains were liberated by hydrazinolysis from intact hLH beta and from glycopeptides obtained after tryptic digestion of hLH alpha, subsequently reduced and fractionated as alditols by anion-exchange and ion-suppression amine-adsorption HPLC and identified mainly by one-dimensional (1D) and two-dimensional (2D) 1H-NMR spectroscopy. The results indicate predominantly diantennary. N-acetyllactosamine-type structures at all three glycosylation sites. The oligosaccharides attached to Asn52 (hLH alpha) and Asn30 (hLH beta) show a remarkably similar pattern, with mainly chain-terminating 4-sulphated 2-deoxy-2-N-acetylamino-D-galactose (GalNAc) and a sulphated/sialylated structure as the major single component. However, virtually all N-glycans on the beta subunit bear a fucose residue alpha 1-6-linked to the proximal GlcNAc, whereas those at Asn52 (and Asn78) of the alpha subunit are predominantly non-fucosylated. The oligosaccharides at Asn78 (hLH alpha) are sialylated rather than sulphated and contain the unique sequence NeuAc alpha 2-6 GalNAc beta 1-4GlcNAc beta 1-2 Man alpha 1-3 as part of the majority of mono- and disialylated compounds. The major single constituent at Asn78 has the following structure: [formula, see text]
Notes: Weisshaar, G
Hiyama, J
Renwick, A G
Nimtz, M
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1991 Jan 1;195(1):257-68. doi: 10.1111/j.1432-1033.1991.tb15702.x.
Author Address: Department of Biochemistry, University of Auckland, New Zealand.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 254
Author: Weitz, G. and Proia, R. L.
Year: 1992
Title: Analysis of the glycosylation and phosphorylation of the alpha-subunit of the lysosomal enzyme, beta-hexosaminidase A, by site-directed mutagenesis
Journal: J Biol Chem
Volume: 267
Issue: 14
Pages: 10039-44
Epub Date: 1992/05/15
Date: May 15
Short Title: Analysis of the glycosylation and phosphorylation of the alpha-subunit of the lysosomal enzyme, beta-hexosaminidase A, by site-directed mutagenesis
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1533633
Keywords: Amino Acid Sequence
Animals
Antibodies
Base Sequence
Cell Line
Codon
Glycosylation
Immunohistochemistry
Lysosomes/*enzymology
Macromolecular Substances
Molecular Sequence Data
*Mutagenesis, Site-Directed
Oligodeoxyribonucleotides
Oligopeptides/chemical synthesis/immunology
Phosphorylation
*Protein Processing, Post-Translational
Sequence Homology, Nucleic Acid
Transfection
beta-N-Acetylhexosaminidases/analysis/*genetics/metabolism
Abstract: The glycosylation and subsequent phosphorylation of mannose residues is a pivotal modification during the biosynthesis of lysosomal enzymes. We have identified the sites of N-linked glycosylation and oligosaccharide phosphorylation on the alpha-subunit of beta-hexosaminidase and have determined the influence of the oligosaccharides on the folding and transport of the enzyme. The potential glycosylation sequences, either singly or in combination, were eliminated through site-directed mutagenesis of the cDNA. By expression of the mutant cDNAs in COS-1 cells, each of the three glycosylation sites on the alpha-subunit was found to be modified by an oligosaccharide. One of the three oligosaccharides was the preferred site of phosphorylation. The absence of any individual oligosaccharide did not diminish the expression of the catalytic activity associated with the alpha-chain, implying proper folding and assembly of subunits. A profound effect was observed, however, when all three oligosaccharides were absent. The unglycosylated alpha-subunit, resulting from genetic alteration of all three glycosylation sites or synthesis of the wild-type protein in the presence of tunicamycin, was catalytically inactive. It was found to be improperly folded into an insoluble aggregate, linked through inappropriate disulfide bonds. The unglycosylated protein was trapped in the lumen of the endoplasmic reticulum and was found in a complex with the Ig heavy chain-binding protein, BiP. The properties of the nonglycosylated, misfolded alpha-subunit were similar to some mutant alpha-subunits in Tay-Sachs disease patients. The results indicate that the oligosaccharides are essential, although not in a site-specific manner, for proper folding and cellular transport of the alpha-subunit.
Notes: Weitz, G
Proia, R L
Comparative Study
Journal Article
United States
J Biol Chem. 1992 May 15;267(14):10039-44.
Author Address: Genetics and Biochemistry Branch, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1327
Author: Wells, L., Slawson, C. and Hart, G. W.
Year: 2011
Title: The E2F-1 associated retinoblastoma-susceptibility gene product is modified by O-GlcNAc
Journal: Amino Acids
Volume: 40
Issue: 3
Pages: 877-83
Epub Date: 2010/08/04
Date: Mar
Short Title: The E2F-1 associated retinoblastoma-susceptibility gene product is modified by O-GlcNAc
Alternate Journal: Amino acids
ISSN: 0939-4451 (Print)
0939-4451
DOI: 10.1007/s00726-010-0709-x
PMCID: PMC3030635
NIHMSID: NIHMS233111
Accession Number: 20680651
Keywords: 3T3 Cells
Acetylglucosamine/*metabolism
Animals
Cell Cycle
E2F1 Transcription Factor/genetics/*metabolism
Glycosylation
HeLa Cells
Humans
Mice
Phosphorylation
Protein Binding
Retinoblastoma Protein/genetics/*metabolism
Abstract: The retinoblastoma-susceptibility gene product (pRB) is a classical tumor suppressor. pRB regulates a number of cellular processes including proliferation, differentiation, and apoptosis. One of the essential mechanisms by which pRB, and the related p107 and p130 family members, act is through its interactions with the E2F class of transcription factors. E2F-1 transcription is necessary for entry into S-phase during the cell-cycle. pRB binds E2F-1 and represses transcription via recruitment of a histone deacetylase complex and by preventing co-activator complexes from binding E2F-1. Current dogma suggests that phosphorylation of pRB during mid- to late-G1 leads to release of E2F-1 and E2F-1 dependent transcriptional activation of essential S-phase genes. Here we show that pRB, and the related p107 protein, are modified by O-linked β-N-acetylglucosamine (O-GlcNAc) in an in vitro transcription/translation system. Furthermore, we show in vivo that pRB is more heavily glycosylated in G1 of the cell-cycle when pRB is known to be in an active, hypophosphorylated state. Finally, we demonstrate that E2F-1 associated pRB is modified by O-GlcNAc. These studies suggest that regulation of pRB function(s) may be controlled by dynamic O-GlcNAc modification, as well as phosphorylation.
Notes: 1438-2199
Wells, Lance
Slawson, Chad
Hart, Gerald W
CA83261/CA/NCI NIH HHS/United States
R01 CA042486-27/CA/NCI NIH HHS/United States
R01 DK061671-09/DK/NIDDK NIH HHS/United States
P30 DK079637/DK/NIDDK NIH HHS/United States
CA42486/CA/NCI NIH HHS/United States
R01 CA042486/CA/NCI NIH HHS/United States
DK075069/DK/NIDDK NIH HHS/United States
F32 CA083261/CA/NCI NIH HHS/United States
R01 DK061671/DK/NIDDK NIH HHS/United States
R01 DK075069/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Amino Acids. 2011 Mar;40(3):877-83. doi: 10.1007/s00726-010-0709-x. Epub 2010 Aug 1.
Author Address: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA 30602, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 858
Author: Wells, L., Vosseller, K., Cole, R. N., Cronshaw, J. M., Matunis, M. J. and Hart, G. W.
Year: 2002
Title: Mapping sites of O-GlcNAc modification using affinity tags for serine and threonine post-translational modifications
Journal: Mol Cell Proteomics
Volume: 1
Issue: 10
Pages: 791-804
Epub Date: 2002/11/20
Date: Oct
Short Title: Mapping sites of O-GlcNAc modification using affinity tags for serine and threonine post-translational modifications
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.m200048-mcp200
Accession Number: 12438562
Keywords: Acetylglucosamine/*chemistry/metabolism
Affinity Labels/*metabolism
Amino Acid Sequence
Animals
Cell Nucleus/metabolism
Chromatography, Affinity
Chromatography, Liquid
Cytoplasm/metabolism
Electrophoresis, Polyacrylamide Gel
Mass Spectrometry/methods
Nuclear Pore Complex Proteins/chemistry/metabolism
Peptides/chemistry
Phosphorylation
*Protein Processing, Post-Translational
Rats
Receptors, Cytoplasmic and Nuclear/metabolism
Reproducibility of Results
Serine/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Synapsins/isolation & purification/metabolism
Threonine/*metabolism
Abstract: Identifying sites of post-translational modifications on proteins is a major challenge in proteomics. O-Linked beta-N-acetylglucosamine (O-GlcNAc) is a dynamic nucleocytoplasmic modification more analogous to phosphorylation than to classical complex O-glycosylation. We describe a mass spectrometry-based method for the identification of sites modified by O-GlcNAc that relies on mild beta-elimination followed by Michael addition with dithiothreitol (BEMAD). Using synthetic peptides, we also show that biotin pentylamine can replace dithiothreitol as the nucleophile. The modified peptides can be efficiently enriched by affinity chromatography, and the sites can be mapped using tandem mass spectrometry. This same methodology can be applied to mapping sites of serine and threonine phosphorylation, and we provide a strategy that uses modification-specific antibodies and enzymes to discriminate between the two post-translational modifications. The BEMAD methodology was validated by mapping three previously identified O-GlcNAc sites, as well as three novel sites, on Synapsin I purified from rat brain. BEMAD was then used on a purified nuclear pore complex preparation to map novel sites of O-GlcNAc modification on the Lamin B receptor and the nucleoporin Nup155. This method is amenable for performing quantitative mass spectrometry and can also be adapted to quantify cysteine residues. In addition, our studies emphasize the importance of distinguishing between O-phosphate versus O-GlcNAc when mapping sites of serine and threonine post-translational modification using beta-elimination/Michael addition methods.
Notes: Wells, Lance
Vosseller, Keith
Cole, Robert N
Cronshaw, Janet M
Matunis, Michael J
Hart, Gerald W
1S10-RR14702/RR/NCRR NIH HHS/United States
CA43486/CA/NCI NIH HHS/United States
CA83261/CA/NCI NIH HHS/United States
DK61671/DK/NIDDK NIH HHS/United States
GM20528/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Mol Cell Proteomics. 2002 Oct;1(10):791-804. doi: 10.1074/mcp.m200048-mcp200.
Author Address: Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21218, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 359
Author: Wendorf, P., Geyer, R., Sziegoleit, A. and Linder, D.
Year: 1989
Title: Localization and characterization of the glycosylation site of human pancreatic elastase 1
Journal: FEBS Lett
Volume: 249
Issue: 2
Pages: 275-8
Epub Date: 1989/06/05
Date: Jun 5
Short Title: Localization and characterization of the glycosylation site of human pancreatic elastase 1
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/0014-5793(89)80640-4
Accession Number: 2737288
Keywords: Amino Acid Sequence
Amino Acids/analysis
Chromatography, Gel
Glycopeptides/isolation & purification
Glycosylation
Humans
Hydrolysis
Molecular Sequence Data
Pancreatic Elastase/*metabolism
Abstract: Crystalline elastase 1 from human pancreas was digested with trypsin. Two peptides, containing the potential N-glycosylation sites at Asn-86 and Asn-125, were isolated and analyzed by amino acid analysis, sequencing and carbohydrate component analysis. The results demonstrate that only Asn-86 is glycosylated.
Notes: Wendorf, P
Geyer, R
Sziegoleit, A
Linder, D
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 1989 Jun 5;249(2):275-8. doi: 10.1016/0014-5793(89)80640-4.
Author Address: Institut für Medizinische Mikrobiologie der Universität, Giessen, FRG.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 288
Author: Wendorf, P., Linder, D., Sziegoleit, A. and Geyer, R.
Year: 1991
Title: Carbohydrate structure of human pancreatic elastase 1
Journal: Biochem J
Volume: 278 ( Pt 2)
Issue: Pt 2
Pages: 505-14
Epub Date: 1991/09/01
Date: Sep 1
Short Title: Carbohydrate structure of human pancreatic elastase 1
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj2780505
PMCID: PMC1151374
Accession Number: 1898343
Keywords: Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Liquid
Electrophoresis, Polyacrylamide Gel
Glycopeptides/metabolism
Glycosylation
Humans
Hydrogen-Ion Concentration
Methylation
Molecular Sequence Data
Oligosaccharides/*chemistry/metabolism
Pancreas/*enzymology
Pancreatic Elastase/*chemistry/isolation & purification
Sugar Alcohols/chemistry
Trypsin/chemistry
Abstract: Human pancreatic elastase 1 (E1) is a glycoprotein containing two potential N-glycosylation sites, one of which carries a carbohydrate moiety [Wendorf, Geyer, Sziegoleit & Linder (1989) FEBS Lett. 249, 275-278]. In order to study its glycosylation, glycoprotein isolated from post-mortem pancreas tissue of 75 donors was digested with trypsin. Oligosaccharides were liberated from resulting glycopeptides by treatment with peptide-N4-(N-acetyl-beta-glycosaminyl)-asparagine amidase F, radiolabelled by reduction with KB3H4 and separated by h.p.l.c. and gel filtration. Major oligosaccharide alditol fractions, representing 67.8 mol% of total glycans, were characterized by methylation analysis and sequential degradation with exoglycosidases. The results revealed that about two-fifths of the partially truncated, mainly biantennary, complex-type glycans found comprised blood group A, B, Lea (or X), difucosyl A or difucosyl B determinants, which could be assigned to lactosamine antennae linked to Man(alpha 1-3)- residues of the sugar chains.
Notes: 1470-8728
Wendorf, P
Linder, D
Sziegoleit, A
Geyer, R
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 1991 Sep 1;278 ( Pt 2)(Pt 2):505-14. doi: 10.1042/bj2780505.
Author Address: Institut für Medizinische Mikrobiologie, Universität Giessen, Federal Republic of Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1543
Author: Weng, T. Y., Chiu, W. T., Liu, H. S., Cheng, H. C., Shen, M. R., Mount, D. B. and Chou, C. Y.
Year: 2013
Title: Glycosylation regulates the function and membrane localization of KCC4
Journal: Biochim Biophys Acta
Volume: 1833
Issue: 5
Pages: 1133-46
Epub Date: 2013/02/05
Date: May
Short Title: Glycosylation regulates the function and membrane localization of KCC4
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbamcr.2013.01.018
Accession Number: 23376777
Keywords: Animals
*Cell Membrane/genetics/metabolism
Cell Transformation, Neoplastic
Endoplasmic Reticulum/*metabolism
Gene Expression Regulation
*Glycosylation
HEK293 Cells
Humans
Mice
Mutagenesis, Site-Directed
Mutation
Polysaccharides/metabolism
Protein Folding
Protein Stability
Surface Properties
*Symporters/genetics
Tunicamycin/pharmacology
Abstract: Glycosylation plays a role in regulating many biological activities, including protein folding and cell surface expression of biomolecules. However, the importance of glycosylation for KCC4 function has not previously been demonstrated. Site-directed mutagenesis was performed on the four putative extracellular N-linked glycosylation sites of KCC4 to determine the role of these sites in KCC4 half-life, cell surface expression, and transporter activity, as well as in KCC4-dependent tumor formation. We showed that triple (N312/331/344/Q) and quadruple (N312/331/344/360/Q) mutations of N-linked glycosylation sites disrupt the N-linked glycosylation of KCC4, resulting in the accumulation of KCC4, predominantly in the endoplasmic reticulum (ER) and not at the cell surface. Further investigation indicated that mutations of the central two (N331/344/Q) N-linked glycosylation sites inhibit the membrane trafficking of KCC4. Our data suggest that the glycan moieties at the N331 and N344 sites were Endo H-resistant, complex-form structures, and that the N312 and N360 sites were Endo H-sensitive, high mannose-containing structures. Under hypotonic stress conditions, the ability to adapt to changes in intracellular chloride ion concentrations and RVD (regulatory volume decrease) activities were less efficient in cells containing the deglycosylated form of KCC4 that were not expressed at the cell surface. Deglycosylated forms of KCC4 also demonstrated decreased tumor formation and lung colonization in mouse xenografts. The difference in glycan complexity may account for the differential impact of each branch on the biological effects of KCC4. We propose that glycosylation is essential for the surface expression, stabilization, and bioactivity of KCC4.
Notes: Weng, Tzu-Yu
Chiu, Wen-Tai
Liu, Hsiao-Sheng
Cheng, Hung-Chi
Shen, Meng-Ru
Mount, David B
Chou, Cheng-Yang
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2013 May;1833(5):1133-46. doi: 10.1016/j.bbamcr.2013.01.018. Epub 2013 Jan 31.
Author Address: College of Medicine, National Cheng Kung University, Tainan, Taiwan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 342
Author: Wernette-Hammond, M. E., Lauer, S. J., Corsini, A., Walker, D., Taylor, J. M. and Rall, S. C., Jr.
Year: 1989
Title: Glycosylation of human apolipoprotein E. The carbohydrate attachment site is threonine 194
Journal: J Biol Chem
Volume: 264
Issue: 15
Pages: 9094-101
Epub Date: 1989/05/25
Date: May 25
Short Title: Glycosylation of human apolipoprotein E. The carbohydrate attachment site is threonine 194
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 2498325
Keywords: Animals
Apolipoproteins E/*biosynthesis/blood/genetics
Cell Line
Genetic Vectors
Glycosylation
HeLa Cells/metabolism
Humans
Models, Molecular
Plasmids
Protein Conformation
Restriction Mapping
*Threonine
Transfection
Abstract: The glycosylation of human apolipoprotein (apo) E was examined with purified plasma apoE and apoE produced by transfected cell lines. The carbohydrate attachment site of plasma apoE was localized to a single tryptic peptide (residues 192-206). Sequence analysis and amino sugar analysis of this peptide derived from asialo-, monosialo-, or disialo-apoE indicated that the carbohydrate moiety is attached only to Thr194 in monosialo- and disialo-apoE and that asialo-apoE is not glycosylated. Mammalian cells that normally do not express apoE were transfected with human apoE plasmid expression vectors to test the utilization of potential carbohydrate attachment sites and the role of apoE glycosylation in secretion. Site-specific mutants of apoE, designed to eliminate or alter glycosylation sites, were expressed in HeLa cells by acute transfection. Apolipoprotein E(Thr194----Ala) was secreted exclusively as the asialo isoform, confirming that Thr194 is the site of carbohydrate attachment in these cells and indicating that glycosylation of apoE is not essential for secretion. Apolipoprotein E(Thr194----Asn,Gly196----Ser), which introduces a potential site for N-glycosylation at position 194, was secreted with a higher apparent molecular weight than native, O-glycosylated apoE. Studies with tunicamycin indicated that this apoE was N-glycosylated at Asn194. Stably transfected cell lines expressing human apoE were prepared from wild-type Chinese hamster ovary (CHO) cells and from CHO ldlD cells, which are defective in glycosylation. The transfected wild-type cells secreted multiply sialylated apoE. The transfected ldlD cells also secreted high levels of apoE even in the absence of glycosylation, which confirms that glycosylation is not essential for secretion of apoE.
Notes: Wernette-Hammond, M E
Lauer, S J
Corsini, A
Walker, D
Taylor, J M
Rall, S C Jr
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1989 May 25;264(15):9094-101.
Author Address: Gladstone Foundation Laboratories for Cardiovascular Disease, San Francisco, California 94140-0608.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1438
Author: West, J. M., Zvonok, N., Whitten, K. M., Wood, J. T. and Makriyannis, A.
Year: 2012
Title: Mass spectrometric characterization of human N-acylethanolamine-hydrolyzing acid amidase
Journal: J Proteome Res
Volume: 11
Issue: 2
Pages: 972-81
Epub Date: 2011/11/02
Date: Feb 3
Short Title: Mass spectrometric characterization of human N-acylethanolamine-hydrolyzing acid amidase
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr200735a
PMCID: PMC3706083
NIHMSID: NIHMS347644
Accession Number: 22040171
Keywords: Amides
Amidohydrolases/*chemistry/isolation & purification/metabolism
Amino Acid Sequence
Chromatography, Gel
Endocannabinoids
Enzyme Precursors/chemistry/isolation & purification/metabolism
Ethanolamines
Glycosylation
HEK293 Cells
Humans
Kinetics
Molecular Sequence Data
Molecular Weight
Palmitic Acids
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism
Protein Subunits/chemistry/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/*methods
Abstract: N-Acylethanolamine-hydrolyzing acid amidase (NAAA) is a lysosomal enzyme that primarily degrades palmitoylethanolamine (PEA), a lipid amide that inhibits inflammatory responses. We developed a HEK293 cell line stably expressing the NAAA pro-enzyme (zymogen) and a single step chromatographic purification of the protein from the media. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry MALDI-TOF MS analysis of the zymogen (47.7 kDa) treated with peptide-N-glycosidase F (PNGase F) identified 4 glycosylation sites, and acid cleavage of the zymogen into α- and β-subunits (14.6 and 33.3 kDa) activated the enzyme. Size exclusion chromatography estimated the mass of the active enzyme as 45 ± 3 kDa, suggesting formation of an α/β heterodimer. MALDI-TOF MS fingerprinting covered more than 80% of the amino acid sequence, including the N-terminal peptides, and evidence for the lack of a disulfide bond between subunits. The significance of the cysteine residues was established by their selective alkylation resulting in almost complete loss of activity. The purified enzyme was kinetically characterized with PEA and a novel fluorogenic substrate, N-(4-methyl coumarin) palmitamide (PAMCA). The production of sufficient quantities of NAAA and a high throughput assay could be useful in discovering novel inhibitors and determining the structure and function of this enzyme.
Notes: 1535-3907
West, Jay M
Zvonok, Nikolai
Whitten, Kyle M
Wood, Jodianne T
Makriyannis, Alexandros
DA003801/DA/NIDA NIH HHS/United States
DA007312/DA/NIDA NIH HHS/United States
R37 DA003801/DA/NIDA NIH HHS/United States
P01 DA009158/DA/NIDA NIH HHS/United States
T32 DA007312/DA/NIDA NIH HHS/United States
R01 DA003801/DA/NIDA NIH HHS/United States
DA009158/DA/NIDA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
J Proteome Res. 2012 Feb 3;11(2):972-81. doi: 10.1021/pr200735a. Epub 2012 Jan 3.
Author Address: Center for Drug Discovery, Northeastern University , Boston, Massachusetts 02115, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1584
Author: West, M. B., Chen, Y., Wickham, S., Heroux, A., Cahill, K., Hanigan, M. H. and Mooers, B. H.
Year: 2013
Title: Novel insights into eukaryotic γ-glutamyltranspeptidase 1 from the crystal structure of the glutamate-bound human enzyme
Journal: J Biol Chem
Volume: 288
Issue: 44
Pages: 31902-13
Epub Date: 2013/09/21
Date: Nov 1
Short Title: Novel insights into eukaryotic γ-glutamyltranspeptidase 1 from the crystal structure of the glutamate-bound human enzyme
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.498139
PMCID: PMC3814782
Accession Number: 24047895
Keywords: Catalytic Domain
Crystallography, X-Ray
Glutamic Acid/*chemistry
Humans
Protein Structure, Secondary
Structure-Activity Relationship
gamma-Glutamyltransferase/*chemistry/genetics
Anomalous Diffraction
Enzyme Structure
Glycoprotein
Hydrolases
Oxidative Stress
Protease
Threonine Peptidase
gamma-Glutamylpeptidyltransferase
gamma-Glutamyltransferase
gamma-glutamyltranspeptidase
Abstract: The enzyme γ-glutamyltranspeptidase 1 (GGT1) is a conserved member of the N-terminal nucleophile hydrolase family that cleaves the γ-glutamyl bond of glutathione and other γ-glutamyl compounds. In animals, GGT1 is expressed on the surface of the cell and has critical roles in maintaining cysteine levels in the body and regulating intracellular redox status. Expression of GGT1 has been implicated as a potentiator of asthma, cardiovascular disease, and cancer. The rational design of effective inhibitors of human GGT1 (hGGT1) has been delayed by the lack of a reliable structural model. The available crystal structures of several bacterial GGTs have been of limited use due to differences in the catalytic behavior of bacterial and mammalian GGTs. We report the high resolution (1.67 Å) crystal structure of glutamate-bound hGGT1, the first of any eukaryotic GGT. Comparisons of the active site architecture of hGGT1 with those of its bacterial orthologs highlight key differences in the residues responsible for substrate binding, including a bimodal switch in the orientation of the catalytic nucleophile (Thr-381) that is unique to the human enzyme. Compared with several bacterial counterparts, the lid loop in the crystal structure of hGGT1 adopts an open conformation that allows greater access to the active site. The hGGT1 structure also revealed tightly bound chlorides near the catalytic residue that may contribute to catalytic activity. These are absent in the bacterial GGTs. These differences between bacterial and mammalian GGTs and the new structural data will accelerate the development of new therapies for GGT1-dependent diseases.
Notes: 1083-351x
West, Matthew B
Chen, Yunyu
Wickham, Stephanie
Heroux, Ann
Cahill, Kyle
Hanigan, Marie H
Mooers, Blaine H M
P41 RR012408/RR/NCRR NIH HHS/United States
R01 AI088011/AI/NIAID NIH HHS/United States
P41 GM103473/GM/NIGMS NIH HHS/United States
F32 CA128338/CA/NCI NIH HHS/United States
R56 CA57530/CA/NCI NIH HHS/United States
P20 GM103640/GM/NIGMS NIH HHS/United States
R56 CA057530/CA/NCI NIH HHS/United States
P20GM103640/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2013 Nov 1;288(44):31902-13. doi: 10.1074/jbc.M113.498139. Epub 2013 Sep 18.
Author Address: From the Departments of Cell Biology and.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1324
Author: West, M. B., Segu, Z. M., Feasley, C. L., Kang, P., Klouckova, I., Li, C., Novotny, M. V., West, C. M., Mechref, Y. and Hanigan, M. H.
Year: 2010
Title: Analysis of site-specific glycosylation of renal and hepatic γ-glutamyl transpeptidase from normal human tissue
Journal: J Biol Chem
Volume: 285
Issue: 38
Pages: 29511-24
Epub Date: 2010/07/14
Date: Sep 17
Short Title: Analysis of site-specific glycosylation of renal and hepatic γ-glutamyl transpeptidase from normal human tissue
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.145938
PMCID: PMC2937983
Accession Number: 20622017
Keywords: Aged
Amino Acid Sequence
Blotting, Western
Electrophoresis, Polyacrylamide Gel
Female
Glycosylation
Humans
In Vitro Techniques
Kidney/*enzymology
Liver/*enzymology
Middle Aged
Molecular Sequence Data
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tandem Mass Spectrometry
gamma-Glutamyltransferase/chemistry/*metabolism
Abstract: The cell surface glycoprotein γ-glutamyl transpeptidase (GGT) was isolated from healthy human kidney and liver to characterize its glycosylation in normal human tissue in vivo. GGT is expressed by a single cell type in the kidney. The spectrum of N-glycans released from kidney GGT constituted a subset of the N-glycans identified from renal membrane glycoproteins. Recent advances in mass spectrometry enabled us to identify the microheterogeneity and relative abundance of glycans on specific glycopeptides and revealed a broader spectrum of glycans than was observed among glycans enzymatically released from isolated GGT. A total of 36 glycan compositions, with 40 unique structures, were identified by site-specific glycan analysis. Up to 15 different glycans were observed at a single site, with site-specific variation in glycan composition. N-Glycans released from liver membrane glycoproteins included many glycans also identified in the kidney. However, analysis of hepatic GGT glycopeptides revealed 11 glycan compositions, with 12 unique structures, none of which were observed on kidney GGT. No variation in glycosylation was observed among multiple kidney and liver donors. Two glycosylation sites on renal GGT were modified exclusively by neutral glycans. In silico modeling of GGT predicts that these two glycans are located in clefts on the surface of the protein facing the cell membrane, and their synthesis may be subject to steric constraints. This is the first analysis at the level of individual glycopeptides of a human glycoprotein produced by two different tissues in vivo and provides novel insights into tissue-specific and site-specific glycosylation in normal human tissues.
Notes: 1083-351x
West, Matthew B
Segu, Zaneer M
Feasley, Christa L
Kang, Pilsoo
Klouckova, Iveta
Li, Chenglong
Novotny, Milos V
West, Christopher M
Mechref, Yehia
Hanigan, Marie H
F32CA128338/CA/NCI NIH HHS/United States
RR018942/RR/NCRR NIH HHS/United States
F32 CA128338/CA/NCI NIH HHS/United States
R01 CA057530/CA/NCI NIH HHS/United States
R56CA057530/CA/NCI NIH HHS/United States
R01CA57530/CA/NCI NIH HHS/United States
R56 CA057530/CA/NCI NIH HHS/United States
P41 RR018942/RR/NCRR NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Sep 17;285(38):29511-24. doi: 10.1074/jbc.M110.145938. Epub 2010 Jul 9.
Author Address: Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1803
Author: Whicher, J. R. and MacKinnon, R.
Year: 2016
Title: Structure of the voltage-gated K⁺ channel Eag1 reveals an alternative voltage sensing mechanism
Journal: Science
Volume: 353
Issue: 6300
Pages: 664-9
Epub Date: 2016/08/16
Date: Aug 12
Short Title: Structure of the voltage-gated K⁺ channel Eag1 reveals an alternative voltage sensing mechanism
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.aaf8070
PMCID: PMC5477842
NIHMSID: NIHMS867939
Accession Number: 27516594
Keywords: Animals
Binding Sites
Calmodulin/chemistry/pharmacology
Cryoelectron Microscopy
Ether-A-Go-Go Potassium Channels/antagonists & inhibitors/*chemistry/ultrastructure
*Ion Channel Gating
Potassium Channel Blockers/chemistry/pharmacology
Protein Binding
Protein Structure, Tertiary
Rats
Abstract: Voltage-gated potassium (K(v)) channels are gated by the movement of the transmembrane voltage sensor, which is coupled, through the helical S4-S5 linker, to the potassium pore. We determined the single-particle cryo-electron microscopy structure of mammalian K(v)10.1, or Eag1, bound to the channel inhibitor calmodulin, at 3.78 angstrom resolution. Unlike previous K(v) structures, the S4-S5 linker of Eag1 is a five-residue loop and the transmembrane segments are not domain swapped, which suggest an alternative mechanism of voltage-dependent gating. Additionally, the structure and position of the S4-S5 linker allow calmodulin to bind to the intracellular domains and to close the potassium pore, independent of voltage-sensor position. The structure reveals an alternative gating mechanism for K(v) channels and provides a template to further understand the gating properties of Eag1 and related channels.
Notes: 1095-9203
Whicher, Jonathan R
MacKinnon, Roderick
R01 GM043949/GM/NIGMS NIH HHS/United States
GM43949/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2016 Aug 12;353(6300):664-9. doi: 10.1126/science.aaf8070.
Author Address: Laboratory of Molecular Neurobiology and Biophysics, The Rockefeller University, Howard Hughes Medical Institute, 1230 York Avenue, New York, NY 10065, USA.
Laboratory of Molecular Neurobiology and Biophysics, The Rockefeller University, Howard Hughes Medical Institute, 1230 York Avenue, New York, NY 10065, USA. mackinn@rockefeller.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1470
Author: Whitaker, G. M., Lynn, F. C., McIntosh, C. H. and Accili, E. A.
Year: 2012
Title: Regulation of GIP and GLP1 receptor cell surface expression by N-glycosylation and receptor heteromerization
Journal: PLoS One
Volume: 7
Issue: 3
Pages: e32675
Epub Date: 2012/03/14
Short Title: Regulation of GIP and GLP1 receptor cell surface expression by N-glycosylation and receptor heteromerization
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0032675
PMCID: PMC3296735
Accession Number: 22412906
Keywords: Amino Acid Sequence
Animals
Cell Line
Chlorocebus aethiops
Consensus Sequence
Cyclic AMP/biosynthesis
Endoplasmic Reticulum/metabolism
Gene Expression Regulation
Glucagon-Like Peptide-1 Receptor
Glycosylation
Humans
Insulin/metabolism
Insulin Secretion
Ligands
Membrane Glycoproteins/chemistry/genetics/metabolism
Molecular Sequence Data
Mutation
Protein Isoforms
*Protein Multimerization
Receptors, Gastrointestinal Hormone/chemistry/genetics/*metabolism
Receptors, Glucagon/chemistry/genetics/*metabolism
Sequence Alignment
Abstract: In response to a meal, Glucose-dependent Insulinotropic Polypeptide (GIP) and Glucagon-like Peptide-1 (GLP-1) are released from gut endocrine cells into the circulation and interact with their cognate G-protein coupled receptors (GPCRs). Receptor activation results in tissue-selective pleiotropic responses that include augmentation of glucose-induced insulin secretion from pancreatic beta cells. N-glycosylation and receptor oligomerization are co-translational processes that are thought to regulate the exit of functional GPCRs from the ER and their maintenance at the plasma membrane. Despite the importance of these regulatory processes, their impact on functional expression of GIP and GLP-1 receptors has not been well studied. Like many family B GPCRs, both the GIP and GLP-1 receptors possess a large extracellular N-terminus with multiple consensus sites for Asn-linked (N)-glycosylation. Here, we show that each of these Asn residues is glycosylated when either human receptor is expressed in Chinese hamster ovary cells. N-glycosylation enhances cell surface expression and function in parallel but exerts stronger control over the GIP receptor than the GLP-1 receptor. N-glycosylation mainly lengthens receptor half-life by reducing degradation in the endoplasmic reticulum. N-glycosylation is also required for expression of the GIP receptor at the plasma membrane and efficient GIP potentiation of glucose-induced insulin secretion from the INS-1 pancreatic beta cell line. Functional expression of a GIP receptor mutant lacking N-glycosylation is rescued by co-expressed wild type GLP1 receptor, which, together with data obtained using Bioluminescence Resonance Energy Transfer, suggests formation of a GIP-GLP1 receptor heteromer.
Notes: 1932-6203
Whitaker, Gina M
Lynn, Francis C
McIntosh, Christopher H S
Accili, Eric A
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2012;7(3):e32675. doi: 10.1371/journal.pone.0032675. Epub 2012 Mar 7.
Author Address: Cardiovascular Research Group, University of British Columbia, Vancouver, British Columbia, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 281
Author: Whitehead, A. S., de Beer, M. C., Steel, D. M., Rits, M., Lelias, J. M., Lane, W. S. and de Beer, F. C.
Year: 1992
Title: Identification of novel members of the serum amyloid A protein superfamily as constitutive apolipoproteins of high density lipoprotein
Journal: J Biol Chem
Volume: 267
Issue: 6
Pages: 3862-7
Epub Date: 1992/02/25
Date: Feb 25
Short Title: Identification of novel members of the serum amyloid A protein superfamily as constitutive apolipoproteins of high density lipoprotein
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 1740433
Keywords: Amino Acid Sequence
Apolipoproteins/*blood/genetics
Base Sequence
DNA/genetics
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Lipoproteins, HDL/*blood/genetics
Liver/metabolism
Molecular Sequence Data
Serum Amyloid A Protein/*analysis/genetics
Abstract: A novel serum amyloid A protein (SAA) has been identified as a normal apolipoprotein component of non-acute phase high density lipoprotein. This novel SAA has been designated "constitutive" SAA (C-SAA) to distinguish it from "acute phase" SAA (A-SAA). C-SAA was partially sequenced, and immunochemical analyses indicated that it constitutes a distinct subclass of apolipoproteins within the SAA superfamily. A C-SAA cDNA clone was isolated from a human liver library and sequenced. The clone predicts a pre-C-SAA molecule of 130 residues from which an 18-residue leader peptide is cleaved. The 112-residue mature molecule is 8 residues longer than human A-SAA; the size difference is due to the presence of an octapeptide between positions 70 and 77 that is not found in the corresponding region of human A-SAA. Paradoxically, octapeptides of similar composition are found at similar positions in the A-SAAs of a number of other species. The C-SAA octapeptide specifies the first two residues of a NSS tripeptide, the only potential N-linked glycosylation site in the molecule. Studies indicate that approximately 50% of these sites are glycosylated, thereby giving rise to two size classes, 14 and 19 kDa, of C-SAA in vivo. Human acute phase liver contains little C-SAA mRNA relative to the levels of A-SAA mRNA, and the treatment of PLC/PRF/5 hepatoma cells with monocyte-conditioned medium does not induce C-SAA mRNA concentrations to detectable levels, in contrast to the massive induction of A-SAA mRNA observed. C-SAA is therefore not a major acute phase reactant.
Notes: Whitehead, A S
de Beer, M C
Steel, D M
Rits, M
Lelias, J M
Lane, W S
de Beer, F C
2T32A107306/PHS HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1992 Feb 25;267(6):3862-7.
Author Address: Department of Immunology, Children's Hospital, Boston, Massachusetts.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 914
Author: Whittington, D. A., Grubb, J. H., Waheed, A., Shah, G. N., Sly, W. S. and Christianson, D. W.
Year: 2004
Title: Expression, assay, and structure of the extracellular domain of murine carbonic anhydrase XIV: implications for selective inhibition of membrane-associated isozymes
Journal: J Biol Chem
Volume: 279
Issue: 8
Pages: 7223-8
Epub Date: 2003/12/09
Date: Feb 20
Short Title: Expression, assay, and structure of the extracellular domain of murine carbonic anhydrase XIV: implications for selective inhibition of membrane-associated isozymes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M310809200
Accession Number: 14660577
Keywords: Amino Acid Sequence
Animals
Binding Sites
COS Cells
Calibration
Carbonic Anhydrases/*chemistry/*genetics
Catalysis
Catalytic Domain
Cell Membrane/enzymology/metabolism
Crystallography, X-Ray
DNA, Complementary/metabolism
Humans
Ions
Mice
Mice, Inbred C57BL
Models, Molecular
Molecular Sequence Data
Protein Conformation
Protein Isoforms
Protein Structure, Tertiary
Sequence Homology, Amino Acid
Zinc/chemistry
Abstract: Carbonic anhydrase (CA) XIV is the most recently identified mammalian carbonic anhydrase isozyme, and its presence has been demonstrated in a number of tissues. Full-length CA XIV is a transmembrane protein composed of an extracellular catalytic domain, a single transmembrane helix, and a short intracellular polypeptide segment. The amino acid sequence identity of human CA XIV relative to the other membrane-associated isozymes (CA IV, CA IX, and CA XII) is 34-46%. We report here the expression and purification of both the full-length enzyme and a truncated, secretory form of murine CA XIV. Both forms of this isozyme are highly active, and both show an abrogation of activity in the presence of 0.2% SDS, in contrast to the behavior of murine CA IV. We also report the crystal structure of the extracellular domain of murine CA XIV at 2.8 A resolution and of an enzyme-acetazolamide complex at 2.9 A resolution. The structure shows a monomeric glycoprotein with a topology similar to that of other mammalian CA isozymes. Based on the x-ray crystallographic results, we compare and contrast known structures of membrane-associated CA isozymes to rationalize the structural elements responsible for the SDS resistance of CA IV and to discuss prospects for the design of selective inhibitors of membrane-associated CA isozymes.
Notes: Whittington, Douglas A
Grubb, Jeffrey H
Waheed, Abdul
Shah, Gul N
Sly, William S
Christianson, David W
DK40163/DK/NIDDK NIH HHS/United States
GM45614/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2004 Feb 20;279(8):7223-8. doi: 10.1074/jbc.M310809200. Epub 2003 Dec 3.
Author Address: Roy and Diana Vagelos Laboratories, Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1576
Author: Wibowo, A. S., Singh, M., Reeder, K. M., Carter, J. J., Kovach, A. R., Meng, W., Ratnam, M., Zhang, F. and Dann, C. E., 3rd
Year: 2013
Title: Structures of human folate receptors reveal biological trafficking states and diversity in folate and antifolate recognition
Journal: Proc Natl Acad Sci U S A
Volume: 110
Issue: 38
Pages: 15180-8
Epub Date: 2013/08/13
Date: Sep 17
Short Title: Structures of human folate receptors reveal biological trafficking states and diversity in folate and antifolate recognition
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1308827110
PMCID: PMC3780903
Accession Number: 23934049
Keywords: Animals
CHO Cells
Chromatography, Affinity
Cricetinae
Cricetulus
Crystallization
Folate Receptors, GPI-Anchored/*chemistry/genetics
Folic Acid/*metabolism
Folic Acid Antagonists/*metabolism
Humans
*Models, Molecular
Molecular Structure
Polymerase Chain Reaction
*Protein Conformation
Protein Transport/genetics
isothermal titration calorimetry
targeted drug delivery
Abstract: Antifolates, folate analogs that inhibit vitamin B9 (folic acid)-using cellular enzymes, have been used over several decades for the treatment of cancer and inflammatory diseases. Cellular uptake of the antifolates in clinical use occurs primarily via widely expressed facilitative membrane transporters. More recently, human folate receptors (FRs), high affinity receptors that transport folate via endocytosis, have been proposed as targets for the specific delivery of new classes of antifolates or folate conjugates to tumors or sites of inflammation. The development of specific, FR-targeted antifolates would be accelerated if additional biophysical data, particularly structural models of the receptors, were available. Here we describe six distinct crystallographic models that provide insight into biological trafficking of FRs and distinct binding modes of folate and antifolates to these receptors. From comparison of the structures, we delineate discrete structural conformations representative of key stages in the endocytic trafficking of FRs and propose models for pH-dependent conformational changes. Additionally, we describe the molecular details of human FR in complex with three clinically prevalent antifolates, pemetrexed (also Alimta), aminopterin, and methotrexate. On the whole, our data form the basis for rapid design and implementation of unique, FR-targeted, folate-based drugs for the treatment of cancer and inflammatory diseases.
Notes: 1091-6490
Wibowo, Ardian S
Singh, Mirage
Reeder, Kristen M
Carter, Joshua J
Kovach, Alexander R
Meng, Wuyi
Ratnam, Manohar
Zhang, Faming
Dann, Charles E 3rd
R01 CA166711/CA/NCI NIH HHS/United States
R01 GM094472/GM/NIGMS NIH HHS/United States
CA166711/CA/NCI NIH HHS/United States
GM094472/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2013 Sep 17;110(38):15180-8. doi: 10.1073/pnas.1308827110. Epub 2013 Aug 9.
Author Address: Department of Chemistry, Indiana University, Bloomington, IN 47405.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 966
Author: Wickham, L., Benjannet, S., Marcinkiewicz, E., Chretien, M. and Seidah, N. G.
Year: 2005
Title: Beta-amyloid protein converting enzyme 1 and brain-specific type II membrane protein BRI3: binding partners processed by furin
Journal: J Neurochem
Volume: 92
Issue: 1
Pages: 93-102
Epub Date: 2004/12/21
Date: Jan
Short Title: Beta-amyloid protein converting enzyme 1 and brain-specific type II membrane protein BRI3: binding partners processed by furin
Alternate Journal: Journal of neurochemistry
ISSN: 0022-3042 (Print)
0022-3042
DOI: 10.1111/j.1471-4159.2004.02840.x
Accession Number: 15606899
Keywords: Amino Acid Sequence
Amyloid Precursor Protein Secretases
Amyloid beta-Peptides/*metabolism
Animals
Aspartic Acid Endopeptidases/genetics/*metabolism
Brain/metabolism
Cell Line
Endopeptidases
Furin/*metabolism
Humans
Membrane Proteins/genetics/*metabolism
Mice
Mice, Transgenic
Molecular Sequence Data
Nerve Tissue Proteins
Protein Binding/physiology
Protein Interaction Mapping
*Protein Processing, Post-Translational
Transfection
Abstract: Using a yeast two-hybrid system, we screened a human brain cDNA library for possible interacting proteins with the C-terminal cytosolic tail of the beta-secretase beta-amyloid protein converting enzyme (BACE)1. This identified seven potential candidates, including the brain-specific type II membrane protein BRI3. Co-localization and co-immunoprecipitation experiments confirmed that BACE1 and BRI3 co-localize and interact with each other via the cytosolic tail of BACE1. Furthermore, pulse and pulse-chase analyses revealed that the pro-protein convertases furin, and to a lesser extent PC7 and PC5A, process BRI3 into a C-terminal secreted approximately 4-kDa product. Thus, furin efficiently processes both pro-BACE1 and its novel interacting protein pro-BRI3.
Notes: Wickham, Louise
Benjannet, Suzanne
Marcinkiewicz, Edwige
Chretien, Michel
Seidah, Nabil G
Journal Article
Research Support, Non-U.S. Gov't
England
J Neurochem. 2005 Jan;92(1):93-102. doi: 10.1111/j.1471-4159.2004.02840.x.
Author Address: Laboratory of Biochemical Neuroendocrinology, Clinical Research Institute of Montreal, Montreal, Quebec, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1078
Author: Wiesmann, C., Katschke, K. J., Yin, J., Helmy, K. Y., Steffek, M., Fairbrother, W. J., McCallum, S. A., Embuscado, L., DeForge, L., Hass, P. E. and van Lookeren Campagne, M.
Year: 2006
Title: Structure of C3b in complex with CRIg gives insights into regulation of complement activation
Journal: Nature
Volume: 444
Issue: 7116
Pages: 217-20
Epub Date: 2006/10/20
Date: Nov 9
Short Title: Structure of C3b in complex with CRIg gives insights into regulation of complement activation
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/nature05263
Accession Number: 17051150
Keywords: *Complement Activation
Complement C3-C5 Convertases/antagonists & inhibitors/metabolism
Complement C3b/*chemistry/*metabolism
Complement C3c/chemistry/metabolism
Complement C5/antagonists & inhibitors/metabolism
Crystallography, X-Ray
Humans
Models, Molecular
Mutation/genetics
Protein Binding
Protein Conformation
Receptors, Complement/*chemistry/genetics/*metabolism
Receptors, Complement 3b
Structure-Activity Relationship
Abstract: The complement system is a key part of the innate immune system, and is required for clearance of pathogens from the bloodstream. After exposure to pathogens, the third component of the complement system, C3, is cleaved to C3b which, after recruitment of factor B, initiates formation of the alternative pathway convertases. CRIg, a complement receptor expressed on macrophages, binds to C3b and iC3b mediating phagocytosis of the particles, but it is unknown how CRIg selectively recognizes proteolytic C3-fragments and whether binding of CRIg to C3b inhibits convertase activation. Here we present the crystal structure of C3b in complex with CRIg and, using CRIg mutants, provide evidence that CRIg acts as an inhibitor of the alternative pathway of complement. The structure shows that activation of C3 induces major structural rearrangements, including a dramatic movement (>80 A) of the thioester-bond-containing domain through which C3b attaches to pathogen surfaces. We show that CRIg is not only a phagocytic receptor, but also a potent inhibitor of the alternative pathway convertases. The structure provides insights into the complex macromolecular structural rearrangements that occur during complement activation and inhibition. Moreover, our structure-function studies relating the structural basis of complement activation and the means by which CRIg inhibits the convertases provide important clues to the development of therapeutics that target complement.
Notes: 1476-4687
Wiesmann, Christian
Katschke, Kenneth J
Yin, JianPing
Helmy, Karim Y
Steffek, Micah
Fairbrother, Wayne J
McCallum, Scott A
Embuscado, Lizette
DeForge, Laura
Hass, Philip E
van Lookeren Campagne, Menno
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
England
Nature. 2006 Nov 9;444(7116):217-20. doi: 10.1038/nature05263. Epub 2006 Oct 15.
Author Address: Department of Protein Engineering, 1 DNA Way, South San Francisco, California 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1500
Author: Wilke, S., Krausze, J. and Büssow, K.
Year: 2012
Title: Crystal structure of the conserved domain of the DC lysosomal associated membrane protein: implications for the lysosomal glycocalyx
Journal: BMC Biol
Volume: 10
Pages: 62
Epub Date: 2012/07/20
Date: Jul 19
Short Title: Crystal structure of the conserved domain of the DC lysosomal associated membrane protein: implications for the lysosomal glycocalyx
Alternate Journal: BMC biology
ISSN: 1741-7007
DOI: 10.1186/1741-7007-10-62
PMCID: PMC3409847
Accession Number: 22809326
Keywords: Amino Acid Sequence
*Conserved Sequence
Crystallography, X-Ray
Glycocalyx/*metabolism
Glycosylation
Humans
Hydrophobic and Hydrophilic Interactions
Intracellular Membranes/metabolism
Lysosomal-Associated Membrane Protein 3/*chemistry/*metabolism
Lysosomes/*metabolism
Models, Molecular
Molecular Sequence Data
Protein Structure, Secondary
Protein Structure, Tertiary
Abstract: BACKGROUND: The family of lysosome-associated membrane proteins (LAMP) comprises the multifunctional, ubiquitous LAMP-1 and LAMP-2, and the cell type-specific proteins DC-LAMP (LAMP-3), BAD-LAMP (UNC-46, C20orf103) and macrosialin (CD68). LAMPs have been implicated in a multitude of cellular processes, including phagocytosis, autophagy, lipid transport and aging. LAMP-2 isoform A acts as a receptor in chaperone-mediated autophagy. LAMP-2 deficiency causes the fatal Danon disease. The abundant proteins LAMP-1 and LAMP-2 are major constituents of the glycoconjugate coat present on the inside of the lysosomal membrane, the 'lysosomal glycocalyx'. The LAMP family is characterized by a conserved domain of 150 to 200 amino acids with two disulfide bonds. RESULTS: The crystal structure of the conserved domain of human DC-LAMP was solved. It is the first high-resolution structure of a heavily glycosylated lysosomal membrane protein. The structure represents a novel β-prism fold formed by two β-sheets bent by β-bulges and connected by a disulfide bond. Flexible loops and a hydrophobic pocket represent possible sites of molecular interaction. Computational models of the glycosylated luminal regions of LAMP-1 and LAMP-2 indicate that the proteins adopt a compact conformation in close proximity to the lysosomal membrane. The models correspond to the thickness of the lysosomal glycoprotein coat of only 5 to 12 nm, according to electron microscopy. CONCLUSION: The conserved luminal domain of lysosome-associated membrane proteins forms a previously unknown β-prism fold. Insights into the structure of the lysosomal glycoprotein coat were obtained by computational models of the LAMP-1 and LAMP-2 luminal regions.
Notes: 1741-7007
Wilke, Sonja
Krausze, Joern
Büssow, Konrad
Journal Article
Research Support, Non-U.S. Gov't
BMC Biol. 2012 Jul 19;10:62. doi: 10.1186/1741-7007-10-62.
Author Address: Department of Molecular Structural Biology, Helmholtz Centre for Infection Research, Inhoffenstr, 7, 38124 Braunschweig, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1831
Author: Wilkes, M., Madej, M. G., Kreuter, L., Rhinow, D., Heinz, V., De Sanctis, S., Ruppel, S., Richter, R. M., Joos, F., Grieben, M., Pike, A. C., Huiskonen, J. T., Carpenter, E. P., Kühlbrandt, W., Witzgall, R. and Ziegler, C.
Year: 2017
Title: Molecular insights into lipid-assisted Ca(2+) regulation of the TRP channel Polycystin-2
Journal: Nat Struct Mol Biol
Volume: 24
Issue: 2
Pages: 123-130
Epub Date: 2017/01/17
Date: Feb
Short Title: Molecular insights into lipid-assisted Ca(2+) regulation of the TRP channel Polycystin-2
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9985
DOI: 10.1038/nsmb.3357
Accession Number: 28092368
Keywords: Binding Sites
Calcium/chemistry
Calcium Signaling
Cryoelectron Microscopy
Glycosylation
HEK293 Cells
Humans
Models, Molecular
Phosphatidic Acids/chemistry
Phosphatidylcholines/chemistry
Protein Binding
Protein Conformation, alpha-Helical
Protein Domains
Protein Processing, Post-Translational
Protein Structure, Quaternary
TRPP Cation Channels/*chemistry
Abstract: Polycystin-2 (PC2), a calcium-activated cation TRP channel, is involved in diverse Ca(2+) signaling pathways. Malfunctioning Ca(2+) regulation in PC2 causes autosomal-dominant polycystic kidney disease. Here we report two cryo-EM structures of distinct channel states of full-length human PC2 in complex with lipids and cations. The structures reveal conformational differences in the selectivity filter and in the large exoplasmic domain (TOP domain), which displays differing N-glycosylation. The more open structure has one cation bound below the selectivity filter (single-ion mode, PC2(SI)), whereas multiple cations are bound along the translocation pathway in the second structure (multi-ion mode, PC2(MI)). Ca(2+) binding at the entrance of the selectivity filter suggests Ca(2+) blockage in PC2(MI), and we observed density for the Ca(2+)-sensing C-terminal EF hand in the unblocked PC2(SI) state. The states show altered interactions of lipids with the pore loop and TOP domain, thus reflecting the functional diversity of PC2 at different locations, owing to different membrane compositions.
Notes: 1545-9985
Wilkes, Martin
Madej, M Gregor
Orcid: 0000-0001-9181-6387
Kreuter, Lydia
Rhinow, Daniel
Heinz, Veronika
De Sanctis, Silvia
Ruppel, Sabine
Richter, Rebecca M
Joos, Friederike
Grieben, Marina
Pike, Ashley C W
Orcid: 0000-0001-9661-2607
Huiskonen, Juha T
Orcid: 0000-0002-0348-7323
Carpenter, Elisabeth P
Orcid: 0000-0001-9138-2937
Kühlbrandt, Werner
Witzgall, Ralph
Ziegler, Christine
092809/Z/10/Z/Wellcome Trust/United Kingdom
060208/Z/00/Z/Wellcome Trust/United Kingdom
093305/Z/10/Z/Wellcome Trust/United Kingdom
090532/Z/09/Z/Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Struct Mol Biol. 2017 Feb;24(2):123-130. doi: 10.1038/nsmb.3357. Epub 2017 Jan 16.
Author Address: Max Planck Institute of Biophysics, Department of Structural Biology, Frankfurt, Germany.
Biophysics II, Faculty of Biology and Preclinical Medicine, Regensburg, Germany.
Structural Genomics Consortium, University of Oxford, Oxford, UK.
Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.
Department of Anatomy, Faculty of Biology and Preclinical Medicine, Regensburg, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 614
Author: Wilkins, P. P., McEver, R. P. and Cummings, R. D.
Year: 1996
Title: Structures of the O-glycans on P-selectin glycoprotein ligand-1 from HL-60 cells
Journal: J Biol Chem
Volume: 271
Issue: 31
Pages: 18732-42
Epub Date: 1996/08/02
Date: Aug 2
Short Title: Structures of the O-glycans on P-selectin glycoprotein ligand-1 from HL-60 cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.271.31.18732
Accession Number: 8702529
Keywords: *Antigens, CD
Carbohydrate Sequence
Cell Line
Glycoside Hydrolases
Glycosylation
HL-60 Cells/*chemistry
Humans
Leukosialin
Ligands
Membrane Glycoproteins/*chemistry
Molecular Sequence Data
Molecular Structure
Oligosaccharides/chemistry
P-Selectin/isolation & purification/*metabolism
Polysaccharides/*chemistry
Sialoglycoproteins/chemistry/isolation & purification
Sialyl Lewis X Antigen
Abstract: P-selectin glycoprotein ligand-1 (PSGL-1) is a disulfide-bonded homodimeric mucin-like glycoprotein on leukocytes that interacts with both P- and E-selectin. In this report we describe the structures of the Ser/Thr-linked O-glycans of PSGL-1 synthesized by HL-60 cells metabolically radiolabeled with 3H-sugar precursors. In control studies, the O-glycans on CD43 (leukosialin), a mucin-like glycoprotein also expressed by HL-60 cells, were analyzed and compared to those of PSGL-1. O-Glycans were released from Ser/Thr residues by mild base/borohydride treatment of purified glycoproteins, and glycan structures were determined by a combination of techniques. In contrast to expectations, PSGL-1 is not heavily fucosylated; a majority of the O-glycans are disialylated or neutral forms of the core-2 tetrasaccharide Galbeta1-->4GlcNAcbeta1-->6(Galbeta1-->3)GalNAcOH++ +. A minority of the O-glycans are alpha-1,3-fucosylated that occur as two major species containing the sialyl Lewis x antigen; one species is a disialylated, monofucosylated glycan, and the other is a monosialylated, trifucosylated glycan having a polylactosamine backbone. CD43 lacks the fucosylated glycans found on PSGL-1 and is enriched for the nonfucosylated, disialylated core-2 hexasaccharide. These results demonstrate that PSGL-1 contains unique fucosylated O-glycans that are predicted to be critical for high affinity interactions between PSGL-1 and selectins.
Notes: Wilkins, P P
McEver, R P
Cummings, R D
P01 HL54804/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1996 Aug 2;271(31):18732-42. doi: 10.1074/jbc.271.31.18732.
Author Address: Department of Biochemistry, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73190, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1860
Author: Willems, A. P., Gundogdu, M., Kempers, M. J. E., Giltay, J. C., Pfundt, R., Elferink, M., Loza, B. F., Fuijkschot, J., Ferenbach, A. T., van Gassen, K. L. I., van Aalten, D. M. F. and Lefeber, D. J.
Year: 2017
Title: Mutations in N-acetylglucosamine (O-GlcNAc) transferase in patients with X-linked intellectual disability
Journal: J Biol Chem
Volume: 292
Issue: 30
Pages: 12621-12631
Epub Date: 2017/06/07
Date: Jul 28
Short Title: Mutations in N-acetylglucosamine (O-GlcNAc) transferase in patients with X-linked intellectual disability
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M117.790097
PMCID: PMC5535036
Accession Number: 28584052
Keywords: Cells, Cultured
Child
Child, Preschool
Cloning, Molecular
DNA/genetics/metabolism
Humans
Intellectual Disability/*genetics/metabolism
Male
*Mutation
N-Acetylglucosaminyltransferases/*genetics/metabolism
Recombinant Proteins/genetics/metabolism
*Congenital Disorders of Glycosylation
*Host Cell Factor 1 (HCF-1)
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
*X-linked Intellectual Disability
*glycobiology
*glycosyltransferase
*metabolic disease
this article
Abstract: N-Acetylglucosamine (O-GlcNAc) transferase (OGT) regulates protein O-GlcNAcylation, an essential and dynamic post-translational modification. The O-GlcNAc modification is present on numerous nuclear and cytosolic proteins and has been implicated in essential cellular functions such as signaling and gene expression. Accordingly, altered levels of protein O-GlcNAcylation have been associated with developmental defects and neurodegeneration. However, mutations in the OGT gene have not yet been functionally confirmed in humans. Here, we report on two hemizygous mutations in OGT in individuals with X-linked intellectual disability (XLID) and dysmorphic features: one missense mutation (p.Arg284Pro) and one mutation leading to a splicing defect (c.463-6T>G). Both mutations reside in the tetratricopeptide repeats of OGT that are essential for substrate recognition. We observed slightly reduced levels of OGT protein and reduced levels of its opposing enzyme O-GlcNAcase in both patient-derived fibroblasts, but global O-GlcNAc levels appeared to be unaffected. Our data suggest that mutant cells attempt to maintain global O-GlcNAcylation by down-regulating O-GlcNAcase expression. We also found that the c.463-6T>G mutation leads to aberrant mRNA splicing, but no stable truncated protein was detected in the corresponding patient-derived fibroblasts. Recombinant OGT bearing the p.Arg284Pro mutation was prone to unfolding and exhibited reduced glycosylation activity against a complex array of glycosylation substrates and proteolytic processing of the transcription factor host cell factor 1, which is also encoded by an XLID-associated gene. We conclude that defects in O-GlcNAc homeostasis and host cell factor 1 proteolysis may play roles in mediation of XLID in individuals with OGT mutations.
Notes: 1083-351x
Willems, Anke P
Gundogdu, Mehmet
Kempers, Marlies J E
Giltay, Jacques C
Pfundt, Rolph
Elferink, Martin
Loza, Bettina F
Fuijkschot, Joris
Ferenbach, Andrew T
van Gassen, Koen L I
van Aalten, Daan M F
Lefeber, Dirk J
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2017 Jul 28;292(30):12621-12631. doi: 10.1074/jbc.M117.790097. Epub 2017 Jun 5.
Author Address: Department of Neurology, Donders Institute for Brain, Cognition and Behaviour, Radboud University Medical Centre, 6500 HB Nijmegen, The Netherlands; Department of Laboratory Medicine, Translational Metabolic Laboratory, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, 6525 GA Nijmegen, The Netherlands.
Centre for Gene Regulation and Expression, School of Life Sciences, University of Dundee, DD1 5EH Dundee, Scotland, United Kingdom.
Department of Genetics, Radboud University Medical Centre, 6500 HB Nijmegen, The Netherlands.
Department of Genetics, University Medical Centre Utrecht, 3508 AB Utrecht, The Netherlands.
Department of Paediatrics, VieCuri Hospital, 5900 BX Venlo, The Netherlands.
Department of Paediatrics, Radboud University Medical Centre and Amalia Children's Hospital, 6500 HB Nijmegen, The Netherlands.
Centre for Gene Regulation and Expression, School of Life Sciences, University of Dundee, DD1 5EH Dundee, Scotland, United Kingdom. Electronic address: dmfvanaalten@dundee.ac.uk.
Department of Neurology, Donders Institute for Brain, Cognition and Behaviour, Radboud University Medical Centre, 6500 HB Nijmegen, The Netherlands; Department of Laboratory Medicine, Translational Metabolic Laboratory, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, 6525 GA Nijmegen, The Netherlands. Electronic address: Dirk.Lefeber@Radboudumc.nl.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 451
Author: Williams, A. F. and Gagnon, J.
Year: 1982
Title: Neuronal cell Thy-1 glycoprotein: homology with immunoglobulin
Journal: Science
Volume: 216
Issue: 4547
Pages: 696-703
Epub Date: 1982/05/14
Date: May 14
Short Title: Neuronal cell Thy-1 glycoprotein: homology with immunoglobulin
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.6177036
Accession Number: 6177036
Keywords: Amino Acid Sequence
Animals
Antigens, Surface/*immunology
Biological Evolution
Epitopes
Glycoproteins/*immunology
Immunoglobulin Constant Regions/immunology
Immunoglobulin Variable Region/immunology
Immunoglobulins/*immunology
Isoantibodies/biosynthesis
Protein Conformation
Thy-1 Antigens
Abstract: The amino acid sequences of mouse brain Thy-1 glycoproteins are shown to be homologous to those of variable-region immunoglobulin domains. There is also good homology with constant domains and beta 2-microglobulin; overall the results suggest that Thy-1 may be like the primordial immunoglobulin domain. Preliminary evidence for an invertebrate Thy-1 homolog supports this possibility.
Notes: Williams, A F
Gagnon, J
Comparative Study
Journal Article
Review
United States
Science. 1982 May 14;216(4547):696-703. doi: 10.1126/science.6177036.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 836
Author: Williams, R. T., Senior, P. V., Van Stekelenburg, L., Layton, J. E., Smith, P. J. and Dziadek, M. A.
Year: 2002
Title: Stromal interaction molecule 1 (STIM1), a transmembrane protein with growth suppressor activity, contains an extracellular SAM domain modified by N-linked glycosylation
Journal: Biochim Biophys Acta
Volume: 1596
Issue: 1
Pages: 131-7
Epub Date: 2002/05/02
Date: Apr 1
Short Title: Stromal interaction molecule 1 (STIM1), a transmembrane protein with growth suppressor activity, contains an extracellular SAM domain modified by N-linked glycosylation
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/s0167-4838(02)00211-x
Accession Number: 11983428
Keywords: Cloning, Molecular
Cytoplasm/chemistry
Extracellular Space/chemistry
Gene Expression Regulation
Glycosylation
Granulocyte Colony-Stimulating Factor/biosynthesis/chemistry
Humans
Immunoblotting
*Membrane Proteins
Mutagenesis, Site-Directed
Neoplasm Proteins/biosynthesis/*chemistry/genetics
Stromal Interaction Molecule 1
Transfection
Abstract: Stromal interaction molecule 1 (STIM1) is a cell surface transmembrane glycoprotein implicated in tumour growth control and stromal-haematopoietic cell interactions. A single sterile alpha motif (SAM) protein-protein interaction domain is modelled within its extracellular region, a subcellular localisation not previously described for other SAM domain-containing proteins. We have defined the transmembrane topology of STIM1 by determining the sites of N-linked glycosylation. We have confirmed that STIM1 is modified by N-linked glycosylation at two sites within the SAM domain itself, deduced as asparagine residues N131 and N171, demonstrating that STIM1 is translocated across the membrane of the endoplasmic reticulum such that the SAM domain resides within the endoplasmic reticulum (ER) lumen. Both N-linked oligosaccharides remain endoglycosidase H-sensitive, indicating absence of full processing within the ER and Golgi. This immature modification is nevertheless sufficient and critical for cell surface expression of STIM1. We show that STIM1-STIM1 homotypic interactions are mediated via the cytoplasmic rather than the extracellular region of STIM1, excluding an essential role for the SAM domain in these protein interactions. These studies provide the first evidence for an extracellular localisation of a SAM domain within any protein, and the first example of a SAM domain modified by N-linked glycosylation.
Notes: Williams, Richard T
Senior, Paul V
Van Stekelenburg, Leonie
Layton, Judith E
Smith, Peter J
Dziadek, Marie A
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2002 Apr 1;1596(1):131-7. doi: 10.1016/s0167-4838(02)00211-x.
Author Address: Department of Anatomy and Cell Biology, University of Melbourne, Melbourne, Victoria 3010, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1653
Author: Wilson, M. R., Hou, Z. and Matherly, L. H.
Year: 2014
Title: Substituted cysteine accessibility reveals a novel transmembrane 2-3 reentrant loop and functional role for transmembrane domain 2 in the human proton-coupled folate transporter
Journal: J Biol Chem
Volume: 289
Issue: 36
Pages: 25287-95
Epub Date: 2014/07/24
Date: Sep 5
Short Title: Substituted cysteine accessibility reveals a novel transmembrane 2-3 reentrant loop and functional role for transmembrane domain 2 in the human proton-coupled folate transporter
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M114.578252
PMCID: PMC4155690
Accession Number: 25053408
Keywords: Amino Acid Sequence
*Amino Acid Substitution
Binding Sites/genetics
Biological Transport/drug effects/genetics
Biotinylation
Blotting, Western
Cell Membrane/metabolism
Cysteine/chemistry/*genetics/metabolism
Folic Acid/metabolism
Folic Acid Antagonists/pharmacology
Glutamates/pharmacology
Guanine/analogs & derivatives/pharmacology
HeLa Cells
Humans
Kinetics
Methotrexate/metabolism
Molecular Sequence Data
*Mutation
Pemetrexed
Protein Structure, Secondary
Proton-Coupled Folate Transporter/chemistry/*genetics/metabolism
Tritium
Anticancer Drug
Cancer Therapy
Cysteine Accessibility
Folate
Folate Transport
Membrane Topology
Membrane Transport
Methanethiolsulfonate
Protein Structure
Reentrant Loop
Abstract: The proton-coupled folate transporter (PCFT) is a folate-proton symporter highly expressed in solid tumors that can selectively target cytotoxic antifolates to tumors under acidic microenvironment conditions. Predicted topology models for PCFT suggest that the loop domain between transmembrane domains (TMDs) 2 and 3 resides in the cytosol. Mutations involving Asp-109 or Arg-113 in the TMD2-3 loop result in loss of activity. By structural homology to other solute carriers, TMD2 may form part of the PCFT substrate binding domain. In this study we mutated the seven cysteine (Cys) residues of human PCFT to serine, creating Cys-less PCFT. Thirty-three single-Cys mutants spanning TMD2 and the TMD2-3 loop in a Cys-less PCFT background were transfected into PCFT-null HeLa cells. All 33 mutants were detected by Western blotting, and 28 were active for [(3)H]methotrexate uptake at pH 5.5. For the active residues, we performed pulldown assays with membrane-impermeable 2-aminoethyl methanethiosulfonate-biotin and streptavidin beads to determine their aqueous-accessibilities. Multiple residues in TMD2 and the TMD2-3 loop domain reacted with 2-aminoethyl methanethiosulfonate-biotin, establishing aqueous accessibilities. Pemetrexed pretreatment inhibited biotinylation of TMD2 mutants G93C and F94C, and biotinylation of these residues inhibited methotrexate transport activity. Our results suggest that the TMD 2-3 loop domain is aqueous-accessible and forms a novel reentrant loop structure. Residues in TMD2 form an aqueous transmembrane pathway for folate substrates, and Gly-93 and Phe-94 may contribute to a substrate binding domain. Characterization of PCFT structure is essential to understanding the transport mechanism including the critical determinants of substrate binding.
Notes: 1083-351x
Wilson, Mike R
Hou, Zhanjun
Matherly, Larry H
R01 CA053535/CA/NCI NIH HHS/United States
T32 CA009531/CA/NCI NIH HHS/United States
R01 CA53535/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2014 Sep 5;289(36):25287-95. doi: 10.1074/jbc.M114.578252. Epub 2014 Jul 22.
Author Address: From the Department of Oncology and.
From the Department of Oncology and the Molecular Therapeutics Program, Barbara Ann Karmanos Cancer Institute, Detroit, Michigan 48201.
From the Department of Oncology and the Molecular Therapeutics Program, Barbara Ann Karmanos Cancer Institute, Detroit, Michigan 48201 Department of Pharmacology, Wayne State University School of Medicine and matherly@karmanos.org.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 926
Author: Windpassinger, C., Auer-Grumbach, M., Irobi, J., Patel, H., Petek, E., Hörl, G., Malli, R., Reed, J. A., Dierick, I., Verpoorten, N., Warner, T. T., Proukakis, C., Van den Bergh, P., Verellen, C., Van Maldergem, L., Merlini, L., De Jonghe, P., Timmerman, V., Crosby, A. H. and Wagner, K.
Year: 2004
Title: Heterozygous missense mutations in BSCL2 are associated with distal hereditary motor neuropathy and Silver syndrome
Journal: Nat Genet
Volume: 36
Issue: 3
Pages: 271-6
Epub Date: 2004/02/26
Date: Mar
Short Title: Heterozygous missense mutations in BSCL2 are associated with distal hereditary motor neuropathy and Silver syndrome
Alternate Journal: Nature genetics
ISSN: 1061-4036 (Print)
1061-4036
DOI: 10.1038/ng1313
Accession Number: 14981520
Keywords: Bone and Bones/abnormalities
GTP-Binding Protein gamma Subunits/*genetics
Genetic Heterogeneity
Hereditary Sensory and Motor Neuropathy/*genetics
Humans
Motor Neurons/pathology
Mutation, Missense
Paraparesis/genetics
Syndrome
Abstract: Distal hereditary motor neuropathy (dHMN) or distal spinal muscular atrophy (OMIM #182960) is a heterogeneous group of disorders characterized by an almost exclusive degeneration of motor nerve fibers, predominantly in the distal part of the limbs. Silver syndrome (OMIM #270685) is a rare form of hereditary spastic paraparesis mapped to chromosome 11q12-q14 (SPG17) in which spasticity of the legs is accompanied by amyotrophy of the hands and occasionally also the lower limbs. Silver syndrome and most forms of dHMN are autosomal dominantly inherited with incomplete penetrance and a broad variability in clinical expression. A genome-wide scan in an Austrian family with dHMN-V (ref. 4) showed linkage to the locus SPG17, which was confirmed in 16 additional families with a phenotype characteristic of dHMN or Silver syndrome. After refining the critical region to 1 Mb, we sequenced the gene Berardinelli-Seip congenital lipodystrophy (BSCL2) and identified two heterozygous missense mutations resulting in the amino acid substitutions N88S and S90L. Null mutations in BSCL2, which encodes the protein seipin, were previously shown to be associated with autosomal recessive Berardinelli-Seip congenital lipodystrophy (OMIM #269700). We show that seipin is an integral membrane protein of the endoplasmic reticulum (ER). The amino acid substitutions N88S and S90L affect glycosylation of seipin and result in aggregate formation leading to neurodegeneration.
Notes: Windpassinger, Christian
Auer-Grumbach, Michaela
Irobi, Joy
Patel, Heema
Petek, Erwin
Hörl, Gerd
Malli, Roland
Reed, Johanna A
Dierick, Ines
Verpoorten, Nathalie
Warner, Thomas T
Proukakis, Christos
Van den Bergh, Peter
Verellen, Christine
Van Maldergem, Lionel
Merlini, Luciano
De Jonghe, Peter
Timmerman, Vincent
Crosby, Andrew H
Wagner, Klaus
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Genet. 2004 Mar;36(3):271-6. doi: 10.1038/ng1313. Epub 2004 Feb 22.
Author Address: Institute of Medical Biology and Human Genetics, Medical University Graz, Harrachgasse 21/8, A-8010 Graz, Austria.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1237
Author: Wollscheid, B., Bausch-Fluck, D., Henderson, C., O'Brien, R., Bibel, M., Schiess, R., Aebersold, R. and Watts, J. D.
Year: 2009
Title: Mass-spectrometric identification and relative quantification of N-linked cell surface glycoproteins
Journal: Nat Biotechnol
Volume: 27
Issue: 4
Pages: 378-86
Epub Date: 2009/04/08
Date: Apr
Short Title: Mass-spectrometric identification and relative quantification of N-linked cell surface glycoproteins
Alternate Journal: Nature biotechnology
ISSN: 1087-0156 (Print)
1087-0156
DOI: 10.1038/nbt.1532
PMCID: PMC2829300
NIHMSID: NIHMS169130
Accession Number: 19349973
Keywords: *Algorithms
Amino Acid Sequence
Cell Membrane/*metabolism
Glycoproteins/*analysis/*chemistry
Mass Spectrometry/*methods
Molecular Sequence Data
Abstract: Although the classification of cell types often relies on the identification of cell surface proteins as differentiation markers, flow cytometry requires suitable antibodies and currently permits detection of only up to a dozen differentiation markers in a single measurement. We use multiplexed mass-spectrometric identification of several hundred N-linked glycosylation sites specifically from cell surface-exposed glycoproteins to phenotype cells without antibodies in an unbiased fashion and without a priori knowledge. We apply our cell surface-capturing (CSC) technology, which covalently labels extracellular glycan moieties on live cells, to the detection and relative quantitative comparison of the cell surface N-glycoproteomes of T and B cells, as well as to monitor changes in the abundance of cell surface N-glycoprotein markers during T-cell activation and the controlled differentiation of embryonic stem cells into the neural lineage. A snapshot view of the cell surface N-glycoproteins will enable detection of panels of N-glycoproteins as potential differentiation markers that are currently not accessible by other means.
Notes: 1546-1696
Wollscheid, Bernd
Bausch-Fluck, Damaris
Henderson, Christine
O'Brien, Robert
Bibel, Miriam
Schiess, Ralph
Aebersold, Ruedi
Watts, Julian D
R01 AI051344-01/AI/NIAID NIH HHS/United States
N01-HV-28179/HV/NHLBI NIH HHS/United States
R01 AI051344/AI/NIAID NIH HHS/United States
N01HV28179/HL/NHLBI NIH HHS/United States
N01-HV-28179-22/HV/NHLBI NIH HHS/United States
R01-AI51344-01/AI/NIAID NIH HHS/United States
N01 HV028179/HV/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Biotechnol. 2009 Apr;27(4):378-86. doi: 10.1038/nbt.1532. Epub 2009 Apr 6.
Author Address: Institute for Systems Biology, Seattle, WA, USA. bernd.wollscheid@imsb.biol.ethz.ch
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1510
Author: Wong, A. H., Zhou, D. and Rini, J. M.
Year: 2012
Title: The X-ray crystal structure of human aminopeptidase N reveals a novel dimer and the basis for peptide processing
Journal: J Biol Chem
Volume: 287
Issue: 44
Pages: 36804-13
Epub Date: 2012/08/31
Date: Oct 26
Short Title: The X-ray crystal structure of human aminopeptidase N reveals a novel dimer and the basis for peptide processing
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.398842
PMCID: PMC3481283
Accession Number: 22932899
Keywords: Amino Acid Motifs
Angiotensins/chemistry
CD13 Antigens/*chemistry
Catalytic Domain
Coordination Complexes/chemistry
Crystallography, X-Ray
HEK293 Cells
Humans
Kinetics
Leucine/analogs & derivatives/chemistry
*Models, Molecular
Peptide Fragments/*chemistry
Peptides/chemistry
Protein Binding
Protein Interaction Domains and Motifs
Protein Structure, Quaternary
Proteolysis
Signal Transduction
Substrate Specificity
Zinc/chemistry
Abstract: Human aminopeptidase N (hAPN/hCD13) is a dimeric membrane protein and a member of the M1 family of zinc metallopeptidases. Within the rennin-angiotensin system, its enzymatic activity is responsible for processing peptide hormones angiotensin III and IV. In addition, hAPN is also involved in cell adhesion, endocytosis, and signal transduction and it is an important target for cancer therapy. Reported here are the high resolution x-ray crystal structures of the dimeric ectodomain of hAPN and its complexes with angiotensin IV and the peptidomimetic inhibitors, amastatin and bestatin. Each monomer of the dimer is found in what has been termed the closed form in other M1 enzymes and each monomer is characterized by an internal cavity surrounding the catalytic site as well as a unique substrate/inhibitor-dependent loop ordering, which in the case of the bestatin complex suggests a new route to inhibitor design. The hAPN structure provides the first example of a dimeric M1 family member and the observed structural features, in conjunction with a model for the open form, provide novel insights into the mechanism of peptide processing and signal transduction.
Notes: 1083-351x
Wong, Alan H M
Zhou, Dongxia
Rini, James M
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2012 Oct 26;287(44):36804-13. doi: 10.1074/jbc.M112.398842. Epub 2012 Aug 29.
Author Address: Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1198
Author: Wong, G. W., Krawczyk, S. A., Kitidis-Mitrokostas, C., Ge, G., Spooner, E., Hug, C., Gimeno, R. and Lodish, H. F.
Year: 2009
Title: Identification and characterization of CTRP9, a novel secreted glycoprotein, from adipose tissue that reduces serum glucose in mice and forms heterotrimers with adiponectin
Journal: Faseb j
Volume: 23
Issue: 1
Pages: 241-58
Epub Date: 2008/09/13
Date: Jan
Short Title: Identification and characterization of CTRP9, a novel secreted glycoprotein, from adipose tissue that reduces serum glucose in mice and forms heterotrimers with adiponectin
Alternate Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
ISSN: 0892-6638 (Print)
0892-6638
DOI: 10.1096/fj.08-114991
PMCID: PMC2626616
Accession Number: 18787108
Keywords: Adiponectin/chemistry/genetics/*metabolism
Adipose Tissue/*metabolism
Animals
*Blood Glucose
Cloning, Molecular
Gene Expression Regulation/physiology
Glycoproteins/genetics/*metabolism
Membrane Proteins/genetics/*metabolism
Mice
Abstract: Adiponectin is a major insulin-sensitizing, multimeric hormone derived from adipose tissue that acts on muscle and liver to regulate whole-body glucose and lipid metabolism. Here, we describe a novel and highly conserved paralog of adiponectin designated as C1q/TNF-related protein (CTRP) 9. Of all the CTRP paralogs, CTRP9 shows the highest degree of amino acid identity to adiponectin in its globular C1q domain. CTRP9 is expressed predominantly in adipose tissue and females expresses higher levels of the transcript than males. Moreover, its expression levels in ob/ob mice changed in an age-dependent manner, with significant up-regulation in younger mice. CTRP9 is a secreted glycoprotein with multiple post-translational modifications in its collagen domain that include hydroxylated prolines and hydroxylated and glycosylated lysines. It is secreted as multimers (predominantly trimers) from transfected cells and circulates in the mouse serum with levels varying according to sex and metabolic state of mice. Furthermore, CTRP9 and adiponectin can be secreted as heterooligomers when cotransfected into mammalian cells, and in vivo, adiponectin/CTRP9 complexes can be reciprocally coimmunoprecipitated from the serum of adiponectin and CTRP9 transgenic mice. Biochemical analysis demonstrates that adiponectin and CTRP9 associate via their globular C1q domain, and this interaction does not require their conserved N-terminal cysteines or their collagen domains. Furthermore, we show that adiponectin and CTRP9 form heterotrimers. In cultured myotubes, CTRP9 specifically activates AMPK, Akt, and p44/42 MAPK signaling pathways. Adenovirus-mediated overexpression of CTRP9 in obese (ob/ob) mice significantly lowered serum glucose levels. Collectively, these results suggest that CTRP9 is a novel adipokine, and further study of CTRP9 will yield novel mechanistic insights into its physiological and metabolic function.
Notes: 1530-6860
Wong, G William
Krawczyk, Sarah A
Kitidis-Mitrokostas, Claire
Ge, Guangtao
Spooner, Eric
Hug, Christopher
Gimeno, Ruth
Lodish, Harvey F
R37 DK047618/DK/NIDDK NIH HHS/United States
R37 DK47618-19/DK/NIDDK NIH HHS/United States
K08 HL077499/HL/NHLBI NIH HHS/United States
K08 HL077499-01/HL/NHLBI NIH HHS/United States
F32 DK067835/DK/NIDDK NIH HHS/United States
F32DK067835/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
FASEB J. 2009 Jan;23(1):241-58. doi: 10.1096/fj.08-114991. Epub 2008 Sep 11.
Author Address: Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 820
Author: Wong, G. W., Yasuda, S., Madhusudhan, M. S., Li, L., Yang, Y., Krilis, S. A., Sali, A. and Stevens, R. L.
Year: 2001
Title: Human tryptase epsilon (PRSS22), a new member of the chromosome 16p13.3 family of human serine proteases expressed in airway epithelial cells
Journal: J Biol Chem
Volume: 276
Issue: 52
Pages: 49169-82
Epub Date: 2001/10/17
Date: Dec 28
Short Title: Human tryptase epsilon (PRSS22), a new member of the chromosome 16p13.3 family of human serine proteases expressed in airway epithelial cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M108677200
Accession Number: 11602603
Keywords: Adult
Amino Acid Sequence
Animals
Base Sequence
Chromosomes, Human, Pair 16/genetics
Cloning, Molecular
Epithelial Cells/*enzymology/physiology
Gene Expression Regulation, Developmental/*physiology
Humans
Isoenzymes
Lung/anatomy & histology/embryology/physiology
Models, Molecular
Molecular Sequence Data
Phylogeny
Protein Structure, Tertiary
Recombinant Proteins/genetics/metabolism
Respiratory Mucosa/cytology/*enzymology/physiology
Sequence Alignment
Serine Endopeptidases/chemistry/classification/genetics/*metabolism
Substrate Specificity
Tissue Distribution
Tryptases
Abstract: Probing of the GenBank expressed sequence tag (EST) data base with varied human tryptase cDNAs identified two truncated ESTs that subsequently were found to encode overlapping portions of a novel human serine protease (designated tryptase epsilon or protease, serine S1 family member 22 (PRSS22)). The tryptase epsilon gene resides on chromosome 16p13.3 within a 2.5-Mb complex of serine protease genes. Although at least 7 of the 14 genes in this complex encode enzymatically active proteases, only one tryptase epsilon-like gene was identified. The trachea and esophagus were found to contain the highest steady-state levels of the tryptase epsilon transcript in adult humans. Although the tryptase epsilon transcript was scarce in adult human lung, it was present in abundance in fetal lung. Thus, the tryptase epsilon gene is expressed in the airways in a developmentally regulated manner that is different from that of other human tryptase genes. At the cellular level, tryptase epsilon is a major product of normal pulmonary epithelial cells, as well as varied transformed epithelial cell lines. Enzymatically active tryptase epsilon is also constitutively secreted from these cells. The amino acid sequence of human tryptase epsilon is 38-44% identical to those of human tryptase alpha, tryptase beta I, tryptase beta II, tryptase beta III, transmembrane tryptase/tryptase gamma, marapsin, and Esp-1/testisin. Nevertheless, comparative protein structure modeling and functional studies using recombinant material revealed that tryptase epsilon has a substrate preference distinct from that of its other family members. These data indicate that the products of the chromosome 16p13.3 complex of tryptase genes evolved to carry out varied functions in humans.
Notes: Wong, G W
Yasuda, S
Madhusudhan, M S
Li, L
Yang, Y
Krilis, S A
Sali, A
Stevens, R L
AI-23483/AI/NIAID NIH HHS/United States
GM-54762/GM/NIGMS NIH HHS/United States
HL-36110/HL/NHLBI NIH HHS/United States
HL-63284/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 2001 Dec 28;276(52):49169-82. doi: 10.1074/jbc.M108677200. Epub 2001 Oct 15.
Author Address: Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 690
Author: Wong, S. C., Zhang, L., Proefke, S. A. and Matherly, L. H.
Year: 1998
Title: Effects of the loss of capacity for N-glycosylation on the transport activity and cellular localization of the human reduced folate carrier
Journal: Biochim Biophys Acta
Volume: 1375
Issue: 1-2
Pages: 6-12
Epub Date: 1998/10/10
Date: Oct 15
Short Title: Effects of the loss of capacity for N-glycosylation on the transport activity and cellular localization of the human reduced folate carrier
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/s0005-2736(98)00118-7
Accession Number: 9767079
Keywords: Base Sequence
Biological Transport
Carrier Proteins/*metabolism
Cell Membrane/*metabolism
Fluorescent Antibody Technique
Folic Acid/metabolism
Glycosylation
Humans
K562 Cells
*Membrane Proteins
*Membrane Transport Proteins
Molecular Sequence Data
Reduced Folate Carrier Protein
Abstract: The role of N-glycosylation in reduced folate carrier (RFC) transport and membrane targeting was examined in transport-deficient K562 (K500E) cells transfected with human RFC cDNAs. Treatment of cells expressing wild-type RFC with tunicamycin (0-3 microg) resulted in a progressive shift of the approximately 85 kDa RFC on western blots to 65 kDa. At 3 microg/ml tunicamycin, the nearly complete loss of glycosylated RFC was accompanied by a approximately 25% decreased rate of methotrexate uptake. A deglycosylated RFC cDNA construct in which asparagine-58 was replaced by glutamine (Gln58-RFC) was expressed in K500E cells as a 65 kDa protein and restored transport capacity for methotrexate and (6S)5-formyl tetrahydrofolate. With both wild-type and Gln58-RFC constructs, expression of cDNA-encoded RFC protein far exceeded relative levels of RFC uptake. Wild-type and Gln58-RFCs containing a hemagglutinin (HA) epitope at the carboxyl terminus were similarly functional and, by immunofluorescence staining with rhodamine-conjugated anti-HA antibody, were localized to plasma membranes. Collectively, our results demonstrate that N-glycosylation of human RFC plays no significant role in either transport function or membrane targeting. The discrepancy between the stoichiometries of RFC expression and transport activity for both wild-type RFC and Gln58-RFC implies that identical regulatory controls and/or non-RFC transport components are necessary to completely restore transport function in the transfected cells.
Notes: Wong, S C
Zhang, L
Proefke, S A
Matherly, L H
CA53535/CA/NCI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
Netherlands
Biochim Biophys Acta. 1998 Oct 15;1375(1-2):6-12. doi: 10.1016/s0005-2736(98)00118-7.
Author Address: Experimental and Clinical Therapeutics Program, Barbara Ann Karmanos Cancer Institute and the Department of Pharmacology, School of Medicine, Wayne State University, 110 E. Warren Ave., Detroit, MI 48201, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1909
Author: Woo, C. M., Lund, P. J., Huang, A. C., Davis, M. M., Bertozzi, C. R. and Pitteri, S. J.
Year: 2018
Title: Mapping and Quantification of Over 2000 O-linked Glycopeptides in Activated Human T Cells with Isotope-Targeted Glycoproteomics (Isotag)
Journal: Mol Cell Proteomics
Volume: 17
Issue: 4
Pages: 764-775
Epub Date: 2018/01/21
Date: Apr
Short Title: Mapping and Quantification of Over 2000 O-linked Glycopeptides in Activated Human T Cells with Isotope-Targeted Glycoproteomics (Isotag)
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.RA117.000261
PMCID: PMC5880114
Accession Number: 29351928
Keywords: Acetylglucosamine/*metabolism
Cells, Cultured
Deuterium
Glycopeptides/*metabolism
Humans
Lymphocyte Activation
Protein Processing, Post-Translational
Proteomics
Proto-Oncogene Proteins c-jun/metabolism
T-Lymphocytes/*metabolism
Abstract: Post-translational modifications (PTMs) on proteins often function to regulate signaling cascades, with the activation of T cells during an adaptive immune response being a classic example. Mounting evidence indicates that the modification of proteins by O-linked N-acetylglucosamine (O-GlcNAc), the only mammalian glycan found on nuclear and cytoplasmic proteins, helps regulate T cell activation. Yet, a mechanistic understanding of how O-GlcNAc functions in T cell activation remains elusive, partly because of the difficulties in mapping and quantifying O-GlcNAc sites. Thus, to advance insight into the role of O-GlcNAc in T cell activation, we performed glycosite mapping studies via direct glycopeptide measurement on resting and activated primary human T cells with a technique termed Isotope Targeted Glycoproteomics. This approach led to the identification of 2219 intact O-linked glycopeptides across 1045 glycoproteins. A significant proportion (>45%) of the identified O-GlcNAc sites lie near or coincide with a known phosphorylation site, supporting the potential for PTM crosstalk. Consistent with other studies, we find that O-GlcNAc sites in T cells lack a strict consensus sequence. To validate our results, we employed gel shift assays based on conjugating mass tags to O-GlcNAc groups. Notably, we observed that the transcription factors c-JUN and JUNB show higher levels of O-GlcNAc glycosylation and higher levels of expression in activated T cells. Overall, our findings provide a quantitative characterization of O-GlcNAc glycoproteins and their corresponding modification sites in primary human T cells, which will facilitate mechanistic studies into the function of O-GlcNAc in T cell activation.
Notes: 1535-9484
Woo, Christina M
Lund, Peder J
Huang, Andrew C
Davis, Mark M
Bertozzi, Carolyn R
Pitteri, Sharon J
R01 CA200423/CA/NCI NIH HHS/United States
U01 CA207702/CA/NCI NIH HHS/United States
U19 AI057229/AI/NIAID NIH HHS/United States
HHMI/Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Mol Cell Proteomics. 2018 Apr;17(4):764-775. doi: 10.1074/mcp.RA117.000261. Epub 2018 Jan 19.
Author Address: From the ‡Departments of Chemistry, cwoo@chemistry.harvard.edu.
§Microbiology & Immunology, and.
‖Interdepartmental Program in Immunology.
§§KAYAK, Cambridge, Massachusetts 02142.
‡‡Howard Hughes Medical Institute; Stanford University, Stanford, California 94305.
From the ‡Departments of Chemistry.
¶Radiology.
**Canary Center for Cancer Early Detection.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 306
Author: Wormald, M. R., Wooten, E. W., Bazzo, R., Edge, C. J., Feinstein, A., Rademacher, T. W. and Dwek, R. A.
Year: 1991
Title: The conformational effects of N-glycosylation on the tailpiece from serum IgM
Journal: Eur J Biochem
Volume: 198
Issue: 1
Pages: 131-9
Epub Date: 1991/05/23
Date: May 23
Short Title: The conformational effects of N-glycosylation on the tailpiece from serum IgM
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1991.tb15995.x
Accession Number: 2040275
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Glycosylation
Humans
Immunoglobulin M/*metabolism
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/chemistry
Protein Conformation
Abstract: 1H-NMR spectroscopy has been used to study the conformation and dynamics of the isolated tailpiece from human serum immunoglobulin M, a 22-residue peptide containing a single asparagine glycosylation site. The peptide is isolated as a set of glycoforms, varying only in the sequence of the oligosaccharide attached at the glycosylation site. The oligosaccharides present have the general formula (Man)n(GlcNAc)2, with 45% having n = 6, 45% having n = 8 and 10% having n = 7 and/or 9. They have been identified and their NMR parameters compared to those found for the isolated oligosaccharides in free solution. The conformation and dynamics of the peptide component have also been studied, using NOE data and hydrogen-exchange experiments, and the results compared to those obtained from the aglycosyl peptide of the same sequence. The presence of the peptide is found to have no measurable effect on the conformation of the oligosaccharides. However, the presence of oligosaccharide causes a decrease in the conformational mobility of the backbone and sidechains of the peptide in the region of the glycosylation site. This is proposed to result from interactions between the oligosaccharide core and the amino acid side chains. Further, the conformation of the N-glycosidic linkage has been shown to be both rigid and planar. Thus, the conformational space available to an N-linked oligosaccharide in a glycoprotein relative to the protein may depend to a large extent upon the flexibility of the asparagine side chain. Various roles for the different glycoforms of the tail peptide are discussed.
Notes: Wormald, M R
Wooten, E W
Bazzo, R
Edge, C J
Feinstein, A
Rademacher, T W
Dwek, R A
Journal Article
Research Support, Non-U.S. Gov't
England
Eur J Biochem. 1991 May 23;198(1):131-9. doi: 10.1111/j.1432-1033.1991.tb15995.x.
Author Address: Department of Biochemistry, University of Oxford, England.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2041
Author: Worth, M., Hu, C. W., Li, H., Fan, D., Estevez, A., Zhu, D., Wang, A. and Jiang, J.
Year: 2019
Title: Targeted covalent inhibition of O-GlcNAc transferase in cells
Journal: Chem Commun (Camb)
Volume: 55
Issue: 88
Pages: 13291-13294
Epub Date: 2019/10/19
Date: Oct 31
Short Title: Targeted covalent inhibition of O-GlcNAc transferase in cells
Alternate Journal: Chemical communications (Cambridge, England)
ISSN: 1359-7345 (Print)
1359-7345
DOI: 10.1039/c9cc04560k
PMCID: PMC6823131
NIHMSID: NIHMS1056460
Accession Number: 31626249
Keywords: Enzyme Inhibitors/chemistry/*pharmacology
HEK293 Cells
Humans
MCF-7 Cells
Models, Molecular
N-Acetylglucosaminyltransferases/*antagonists & inhibitors/metabolism
Abstract: O-GlcNAc transferase (OGT) glycosylates numerous proteins and is implicated in many diseases. To date, most OGT inhibitors lack either sufficient potency or characterized specificity in cells. We report the first targeted covalent inhibitor that predominantly reacts with OGT but does not affect other functionally similar enzymes. This study provides a new strategy to interrogate cellular OGT functions and to investigate other glycosyltransferases.
Notes: 1364-548x
Worth, Matthew
Hu, Chia-Wei
Li, Hao
Fan, Dacheng
Estevez, Arielis
Orcid: 0000-0001-7588-2691
Zhu, Dongsheng
Wang, Ao
Jiang, Jiaoyang
Orcid: 0000-0001-8445-9924
R01 GM121718/GM/NIGMS NIH HHS/United States
Journal Article
Chem Commun (Camb). 2019 Oct 31;55(88):13291-13294. doi: 10.1039/c9cc04560k.
Author Address: Department of Chemistry, University of Wisconsin-Madison, Madison, Wisconsin 53705, USA.
Pharmaceutical Sciences Division, School of Pharmacy, University of Wisconsin-Madison, Madison, Wisconsin 53705, USA. jiaoyang.jiang@wisc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1037
Author: Wu, D., Zajonc, D. M., Fujio, M., Sullivan, B. A., Kinjo, Y., Kronenberg, M., Wilson, I. A. and Wong, C. H.
Year: 2006
Title: Design of natural killer T cell activators: structure and function of a microbial glycosphingolipid bound to mouse CD1d
Journal: Proc Natl Acad Sci U S A
Volume: 103
Issue: 11
Pages: 3972-7
Epub Date: 2006/03/16
Date: Mar 14
Short Title: Design of natural killer T cell activators: structure and function of a microbial glycosphingolipid bound to mouse CD1d
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0600285103
PMCID: PMC1449630
Accession Number: 16537470
Keywords: Animals
Antigens, CD1/chemistry/*metabolism
Antigens, CD1d
Binding Sites
Glycosphingolipids/*chemistry/*metabolism
Hydrogen Bonding
In Vitro Techniques
Killer Cells, Natural/*immunology/*metabolism
Lymphocyte Activation
Macromolecular Substances
Mice
Models, Molecular
Molecular Structure
Protein Structure, Quaternary
T-Lymphocytes/*immunology/*metabolism
Abstract: Natural killer T (NKT) cells provide an innate-type immune response upon T cell receptor interaction with CD1d-presented antigens. We demonstrate through equilibrium tetramer binding and antigen presentation assays with Valpha14i-positive NKT cell hybridomas that the Sphingomonas glycolipid alpha-galacturonosyl ceramide (GalA-GSL) is a NKT cell agonist that is significantly weaker than alpha-galactosylceramide (alpha-GalCer), the most potent known NKT agonist. For GalA-GSL, a shorter fatty acyl chain, an absence of the 4-OH on the sphingosine tail and a 6'-COOH group on the galactose moiety account for its observed antigenic potency. We further determined the crystal structure of mCD1d in complex with GalA-GSL at 1.8-A resolution. The overall binding mode of GalA-GSL to mCD1d is similar to that of the short-chain alpha-GalCer ligand PBS-25, but its sphinganine chain is more deeply inserted into the F' pocket due to alternate hydrogen-bonding interactions between the sphinganine 3-OH with Asp-80. Subsequently, a slight lateral shift (>1 A) of the galacturonosyl head group is noted at the CD1 surface compared with the galactose of alpha-GalCer. Because the relatively short C(14) fatty acid of GalA-GSL does not fully occupy the A' pocket, a spacer lipid is found that stabilizes this pocket. The lipid spacer was identified by GC/MS as a mixture of saturated and monounsaturated palmitic acid (C(16)). Comparison of available crystal structures of alpha-anomeric glycosphingolipids now sheds light on the structural basis of their differential antigenic potency and has led to the design and synthesis of NKT cell agonists with enhanced cell-based stimulatory activities compared with alpha-GalCer.
Notes: 1091-6490
Wu, Douglass
Zajonc, Dirk M
Fujio, Masakazu
Sullivan, Barbara A
Kinjo, Yuki
Kronenberg, Mitchell
Wilson, Ian A
Wong, Chi-Huey
R01 CA058896/CA/NCI NIH HHS/United States
AI45053/AI/NIAID NIH HHS/United States
F32 AI062015/AI/NIAID NIH HHS/United States
R01 AI045053/AI/NIAID NIH HHS/United States
AI062015/AI/NIAID NIH HHS/United States
CA52511/CA/NCI NIH HHS/United States
CA58896/CA/NCI NIH HHS/United States
R01 CA052511/CA/NCI NIH HHS/United States
GM62116/GM/NIGMS NIH HHS/United States
U54 GM062116/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Proc Natl Acad Sci U S A. 2006 Mar 14;103(11):3972-7. doi: 10.1073/pnas.0600285103. Epub 2006 Mar 6.
Author Address: Department of Chemistry and Molecular Biology and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1849
Author: Wu, D., Zhao, L., Feng, Z., Yu, C., Ding, J., Wang, L., Wang, F., Liu, D., Zhu, H., Xing, F., Conaway, J. W., Conaway, R. C., Cai, Y. and Jin, J.
Year: 2017
Title: O-Linked N-acetylglucosamine transferase 1 regulates global histone H4 acetylation via stabilization of the nonspecific lethal protein NSL3
Journal: J Biol Chem
Volume: 292
Issue: 24
Pages: 10014-10025
Epub Date: 2017/04/30
Date: Jun 16
Short Title: O-Linked N-acetylglucosamine transferase 1 regulates global histone H4 acetylation via stabilization of the nonspecific lethal protein NSL3
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M117.781401
PMCID: PMC5473209
Accession Number: 28450392
Keywords: Acetylation
Amino Acid Substitution
Animals
HEK293 Cells
HeLa Cells
Histone Acetyltransferases/antagonists & inhibitors/chemistry/genetics/*metabolism
Histones/*metabolism
Humans
Intracellular Signaling Peptides and Proteins
Isoenzymes/chemistry/genetics/metabolism
N-Acetylglucosaminyltransferases/antagonists &
inhibitors/chemistry/genetics/*metabolism
Nuclear Proteins/antagonists & inhibitors/chemistry/genetics/*metabolism
Point Mutation
Protein Isoforms/chemistry/genetics/metabolism
*Protein Processing, Post-Translational
Protein Stability
Protein Subunits/chemistry/genetics/metabolism
RNA Interference
Recombinant Fusion Proteins/chemistry/metabolism
Recombinant Proteins/chemistry/metabolism
Sf9 Cells
Spodoptera
Substrate Specificity
*NSL complex
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
*histone acetylase
*histone acetylation
*post-translational modification (PTM)
this article
Abstract: The human males absent on the first (MOF)-containing histone acetyltransferase nonspecific lethal (NSL) complex comprises nine subunits including the O-linked N-acetylglucosamine (O-GlcNAc) transferase, isoform 1 (OGT1). However, whether the O-GlcNAc transferase activity of OGT1 controls histone acetyltransferase activity of the NSL complex and whether OGT1 physically interacts with the other NSL complex subunits remain unclear. Here, we demonstrate that OGT1 regulates the activity of the NSL complex by mainly acetylating histone H4 Lys-16, Lys-5, and Lys-8 via O-GlcNAcylation and stabilization of the NSL complex subunit NSL3. Knocking down or overexpressing OGT1 in human cells remarkably affected the global acetylation of histone H4 residues Lys-16, Lys-5, and Lys-8. Because OGT1 is a subunit of the NSL complex, we also investigated the function of OGT1 in this complex. Co-transfection/co-immunoprecipitation experiments combined with in vitro O-GlcNAc transferase assays confirmed that OGT1 specifically binds to and O-GlcNAcylates NSL3. In addition, wheat germ agglutinin affinity purification verified the occurrence of O-GlcNAc modification on NSL3 in cells. Moreover, O-GlcNAcylation of NSL3 by wild-type OGT1 (OGT1-WT) stabilized NSL3. This stabilization was lost after co-transfection of NSL3 with an OGT1 mutant, OGT1(C964A), that lacks O-GlcNAc transferase activity. Furthermore, stabilization of NSL3 by OGT1-WT significantly increased the global acetylation levels of H4 Lys-5, Lys-8, and Lys-16 in cells. These results suggest that OGT1 regulates the activity of the NSL complex by stabilizing NSL3.
Notes: 1083-351x
Wu, Donglu
Zhao, Linhong
Feng, Zhitong
Yu, Chao
Ding, Jian
Wang, Lingyao
Wang, Fei
Liu, Da
Zhu, Huihui
Xing, Feiyang
Conaway, Joan W
Conaway, Ronald C
Cai, Yong
Jin, Jingji
Journal Article
J Biol Chem. 2017 Jun 16;292(24):10014-10025. doi: 10.1074/jbc.M117.781401. Epub 2017 Apr 27.
Author Address: From the School of Life Sciences.
School of Pharmacy, Changchun University of Chinese Medicine, Changchun 130117, China.
Stowers Institute for Medical Research, Kansas City, Missouri 64110, and.
Department of Biochemistry and Molecular Biology, Kansas University Medical Center, Lawrence, Kansas 66045.
From the School of Life Sciences, caiyong62@jlu.edu.cn.
National Engineering Laboratory for AIDS Vaccine, and.
Key Laboratory for Molecular Enzymology and Engineering, the Ministry of Education, Jilin University, Changchun 130012, China.
From the School of Life Sciences, jjjin@jlu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2045
Author: Wu, H., Song, S., Yan, A., Guo, X., Chang, L., Xu, L., Hu, L., Kuang, M., Liu, B., He, D., Zhao, R., Wang, L., Wu, X., Gu, J. and Ruan, Y.
Year: 2020
Title: RACK1 promotes the invasive activities and lymph node metastasis of cervical cancer via galectin-1
Journal: Cancer Lett
Volume: 469
Pages: 287-300
Epub Date: 2019/11/11
Date: Jan 28
Short Title: RACK1 promotes the invasive activities and lymph node metastasis of cervical cancer via galectin-1
Alternate Journal: Cancer letters
ISSN: 0304-3835
DOI: 10.1016/j.canlet.2019.11.002
Accession Number: 31705928
Keywords: Adult
Aged
Carcinoma, Squamous Cell/*genetics/pathology
Cell Line, Tumor
Cell Movement/genetics
Female
Galectin 1/*genetics
Humans
Lymph Nodes/metabolism/pathology
Lymphangiogenesis/genetics
Lymphatic Metastasis/*genetics/pathology
Middle Aged
Neoplasm Invasiveness/genetics/pathology
Neoplasm Proteins/*genetics
Prognosis
Receptors for Activated C Kinase/*genetics
Signal Transduction/genetics
Uterine Cervical Neoplasms/*genetics/pathology
*Human papillomavirus
*Integrin-β1
*Lymphangiogenesis
*O-GlcNAcylation
*miRNA-1275
Abstract: Cervical cancer remains the first leading cause of cancer-related mortality among female reproductive system malignancies worldwide, and invasion and lymph node metastasis of cervical cancer represent the major reason for its poor prognosis. In this study, we found that RACK1 facilitated tumor cell invasion and lymphatic tube formation in vitro, as well as promoted lymphangiogenesis and lymph node metastasis in vivo in a galectin-1-dependent manner. Mechanism studies revealed that RACK1 promoted the expression and secretion of galectin-1 by reducing miR-1275 levels. Additionally, RACK1 also augmented galectin-1-induced downstream MEK/ERK, FAK, and AKT signaling via integrin-β1 in cervical cancer cells. Tissue microarray confirmed that RACK1 was upregulated in squamous intraepithelial lesion and cancer, and RACK1 was positively correlated with invasion/metastasis phenotype, galectin-1 expression, and unfavorable prognosis in cervical cancer cases. Human papillomavirus E6 oncogene contributes to increased expression of RACK1 via the enhancement of its O-GlcNAcylation and protein stability. Together, our results demonstrate that RACK1 stimulates tumor invasion and lymph node metastasis of cervical cancer via galectin-1 and imply that targeting RACK1/galectin-1 axis provides promising means for cervical cancer treatment.
Notes: 1872-7980
Wu, Hao
Song, Shushu
Yan, Anqi
Guo, Xinying
Chang, Lei
Xu, Ling
Hu, Lan
Kuang, Mengzhen
Liu, Bo
He, Daochuan
Zhao, Ran
Wang, Lan
Wu, Xin
Gu, Jianxin
Ruan, Yuanyuan
Journal Article
Research Support, Non-U.S. Gov't
Ireland
Cancer Lett. 2020 Jan 28;469:287-300. doi: 10.1016/j.canlet.2019.11.002. Epub 2019 Nov 6.
Author Address: Key Laboratory of Glycoconjugate Research Ministry of Public Health, School of Basic Medical Sciences, Fudan University, Shanghai, PR China; Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai, PR China.
Key Laboratory of Female Reproductive Endocrine Related Diseases, Obstetrics and Gynecology Hospital, Fudan University, Shanghai, PR China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing, PR China.
Key Laboratory of Glycoconjugate Research Ministry of Public Health, School of Basic Medical Sciences, Fudan University, Shanghai, PR China; Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai, PR China. Electronic address: lanwang10@fudan.edu.cn.
Key Laboratory of Female Reproductive Endocrine Related Diseases, Obstetrics and Gynecology Hospital, Fudan University, Shanghai, PR China. Electronic address: sky_xin1980@aliyun.com.
Key Laboratory of Glycoconjugate Research Ministry of Public Health, School of Basic Medical Sciences, Fudan University, Shanghai, PR China; Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai, PR China. Electronic address: yuanyuanruan@fudan.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 957
Author: Wu, J., Hansen, G. H., Nilsson, A. and Duan, R. D.
Year: 2005
Title: Functional studies of human intestinal alkaline sphingomyelinase by deglycosylation and mutagenesis
Journal: Biochem J
Volume: 386
Issue: Pt 1
Pages: 153-60
Epub Date: 2004/10/02
Date: Feb 15
Short Title: Functional studies of human intestinal alkaline sphingomyelinase by deglycosylation and mutagenesis
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20041455
PMCID: PMC1134777
Accession Number: 15458386
Keywords: Animals
Binding Sites
COS Cells/enzymology
Cations, Divalent/*metabolism
Cell Membrane/enzymology
Cricetinae
Cricetulus
Culture Media, Conditioned
Endosomes/enzymology
Glycosylation/drug effects
Humans
Hydrogen-Ion Concentration
Immunohistochemistry
Intestines/*enzymology
Membrane Proteins/analysis
Mutagenesis, Site-Directed
Phosphodiesterase I/analysis
*Protein Processing, Post-Translational/drug effects
Protein Structure, Tertiary
Recombinant Fusion Proteins/metabolism
Sphingomyelin Phosphodiesterase/analysis/*chemistry/metabolism
Sphingomyelins/metabolism
Transfection
Tunicamycin/pharmacology
Abstract: Intestinal alk-SMase (alkaline sphingomyelinase) is an ectoenzyme related to the NPP (nucleotide phosphodiesterase) family. It has five potential N-glycosylation sites and predicated transmembrane domains at both the N- and C-termini. The amino acid residues forming the two metal-binding sites in NPP are conserved, and those of the active core are modified. We examined the functional changes of the enzyme induced by deglycosylation and mutagenesis. Treating alk-SMase cDNA-transfected COS-7 cells with tunicamycin rendered the expressed enzyme completely inactive. Mutations of the five potential N-glycosylation sites individually and in combination showed that these sites were all glycosylated and deficient glycosylation decreased the enzyme activity. Immunogold labelling showed that the wild-type enzyme was mainly located in the plasma membrane, whereas the C-terminal domain-truncated enzyme was released into the medium. Deglycosylation blocked the release of the enzyme that accumulated in endosome-like structures. The enzyme activity was also decreased by mutations of the residues forming the putative metal-binding sites and the active core. Substitution of the active core sequence with that of NPP or mutation of T75 in the core abolished the enzyme activity against sphingomyelin but failed to render the enzyme NPP active. Our results indicate that alk-SMase activity is severely affected by defective N-glycosylation and structural alterations of the putative metal-binding sites and the predicted active core.
Notes: 1470-8728
Wu, Jun
Hansen, Gert H
Nilsson, Ake
Duan, Rui-Dong
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2005 Feb 15;386(Pt 1):153-60. doi: 10.1042/BJ20041455.
Author Address: Gastroenterology Lab, Biomedical Center B11, Lund University, S-221 84 Lund, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1859
Author: Wu, L., Jiang, J., Jin, Y., Kallemeijn, W. W., Kuo, C. L., Artola, M., Dai, W., van Elk, C., van Eijk, M., van der Marel, G. A., Codée, J. D. C., Florea, B. I., Aerts, Jmfg, Overkleeft, H. S. and Davies, G. J.
Year: 2017
Title: Activity-based probes for functional interrogation of retaining β-glucuronidases
Journal: Nat Chem Biol
Volume: 13
Issue: 8
Pages: 867-873
Epub Date: 2017/06/06
Date: Aug
Short Title: Activity-based probes for functional interrogation of retaining β-glucuronidases
Alternate Journal: Nature chemical biology
ISSN: 1552-4450
DOI: 10.1038/nchembio.2395
Accession Number: 28581485
Keywords: Enzyme Inhibitors/chemical synthesis/chemistry/*pharmacology
Fluorescent Dyes/chemical synthesis/chemistry/*pharmacology
Glucuronidase/*metabolism
HEK293 Cells
Humans
Molecular Structure
Structure-Activity Relationship
Abstract: Humans express at least two distinct β-glucuronidase enzymes that are involved in disease: exo-acting β-glucuronidase (GUSB), whose deficiency gives rise to mucopolysaccharidosis type VII, and endo-acting heparanase (HPSE), whose overexpression is implicated in inflammation and cancers. The medical importance of these enzymes necessitates reliable methods to assay their activities in tissues. Herein, we present a set of β-glucuronidase-specific activity-based probes (ABPs) that allow rapid and quantitative visualization of GUSB and HPSE in biological samples, providing a powerful tool for dissecting their activities in normal and disease states. Unexpectedly, we find that the supposedly inactive HPSE proenzyme proHPSE is also labeled by our ABPs, leading to surprising insights regarding structural relationships between proHPSE, mature HPSE, and their bacterial homologs. Our results demonstrate the application of β-glucuronidase ABPs in tracking pathologically relevant enzymes and provide a case study of how ABP-driven approaches can lead to discovery of unanticipated structural and biochemical functionality.
Notes: 1552-4469
Wu, Liang
Orcid: 0000-0003-0294-7065
Jiang, Jianbing
Jin, Yi
Orcid: 0000-0002-6927-4371
Kallemeijn, Wouter W
Kuo, Chi-Lin
Orcid: 0000-0003-3748-5008
Artola, Marta
Orcid: 0000-0002-3051-3902
Dai, Wei
van Elk, Cas
Orcid: 0000-0001-9620-5534
van Eijk, Marco
van der Marel, Gijsbert A
Codée, Jeroen D C
Florea, Bogdan I
Aerts, Johannes M F G
Overkleeft, Herman S
Davies, Gideon J
Orcid: 0000-0002-7343-776x
Journal Article
United States
Nat Chem Biol. 2017 Aug;13(8):867-873. doi: 10.1038/nchembio.2395. Epub 2017 Jun 5.
Author Address: York Structural Biology Laboratory, Department of Chemistry, University of York, Heslington, York, UK.
Department of Bioorganic Synthesis, Leiden Institute of Chemistry, Leiden University, Leiden, the Netherlands.
Department of Medical Biochemistry, Leiden Institute of Chemistry, Leiden University, Leiden, the Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1986
Author: Wu, N., Jiang, M., Han, Y., Liu, H., Chu, Y., Liu, H., Cao, J., Hou, Q., Zhao, Y., Xu, B. and Xie, X.
Year: 2019
Title: O-GlcNAcylation promotes colorectal cancer progression by regulating protein stability and potential catcinogenic function of DDX5
Journal: J Cell Mol Med
Volume: 23
Issue: 2
Pages: 1354-1362
Epub Date: 2018/11/30
Date: Feb
Short Title: O-GlcNAcylation promotes colorectal cancer progression by regulating protein stability and potential catcinogenic function of DDX5
Alternate Journal: Journal of cellular and molecular medicine
ISSN: 1582-1838 (Print)
1582-1838
DOI: 10.1111/jcmm.14038
PMCID: PMC6349181
Accession Number: 30484950
Keywords: Animals
Apoptosis
Biomarkers, Tumor/genetics/*metabolism
Cell Movement
Cell Proliferation
Colorectal Neoplasms/genetics/metabolism/*pathology
DEAD-box RNA Helicases/*chemistry/genetics/*metabolism
Disease Progression
Gene Expression Regulation, Neoplastic
Glycosylation
Humans
Male
Mice
Mice, Inbred BALB C
Mice, Nude
N-Acetylglucosaminyltransferases/metabolism
*Protein Processing, Post-Translational
Protein Stability
Proto-Oncogene Proteins c-akt/genetics/*metabolism
TOR Serine-Threonine Kinases/genetics/*metabolism
Tumor Cells, Cultured
Xenograft Model Antitumor Assays
*Akt
*Colorectal cancer
*Ddx5
*O-GlcNAcylation
*mTOR
Abstract: The RNA helicase p68 (DDX5), a key player in RNA metabolism, belongs to the DEAD box family and is involved in the development of colorectal cancer. Here, we found both DDX5 and O-GlcNAcylation are up-regulated in colorectal cancer. In addition, DDX5 protein level is significantly positively correlated with the expression of O-GlcNAcylation. Although it was known DDX5 protein could be regulated by post-translational modification (PTM), how O-GlcNAcylation modification regulated of DDX5 remains unclear. Here we show that DDX5 interacts directly with OGT in the SW480 cell line, which is the only known enzyme that catalyses O-GlcNAcylation in humans. Meanwhile, O-GlcNAcylation could promote DDX5 protein stability. The OGT-DDX5 axis affects colorectal cancer progression mainly by regulating activation of the AKT/mTOR signalling pathway. Taken together, these results indicated that OGT-mediated O-GlcNAcylation stabilizes DDX5, promoting activation of the AKT/mTOR signalling pathway, thus accelerating colorectal cancer progression. This study not only reveals the novel functional of O-GlcNAcylation in regulating DDX5, but also reveals the carcinogenic effect of the OGT-DDX5 axis in colorectal cancer.
Notes: 1582-4934
Wu, Nan
Jiang, Mingzuo
Han, Yuying
Liu, Haiming
Chu, Yi
Liu, Hao
Cao, Jiayi
Hou, Qiuqiu
Zhao, Yu
Xu, Bing
Xie, Xin
Journal Article
Research Support, Non-U.S. Gov't
J Cell Mol Med. 2019 Feb;23(2):1354-1362. doi: 10.1111/jcmm.14038. Epub 2018 Nov 28.
Author Address: Laboratory of Tissue Engineering, Faculty of Life Science, Northwest University, Xi'an, Shaanxi, China.
State Key Laboratory of Cancer Biology, National Clinical Research Center for Digestive Diseases and Xijing Hospital of Digestive Diseases, Air Force Medical University (Fourth Military Medical University), Xi'an, Shaanxi, China.
College of Computer Science and Technology, Jilin University, Changchun, Jilin, China.
Department of Gastroenterology, Second Affiliated Hospital of Xi'an Jiao tong University, Xi'an, Shaanxi, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1005
Author: Wu, S. L., Kim, J., Hancock, W. S. and Karger, B.
Year: 2005
Title: Extended Range Proteomic Analysis (ERPA): a new and sensitive LC-MS platform for high sequence coverage of complex proteins with extensive post-translational modifications-comprehensive analysis of beta-casein and epidermal growth factor receptor (EGFR)
Journal: J Proteome Res
Volume: 4
Issue: 4
Pages: 1155-70
Epub Date: 2005/08/09
Date: Jul-Aug
Short Title: Extended Range Proteomic Analysis (ERPA): a new and sensitive LC-MS platform for high sequence coverage of complex proteins with extensive post-translational modifications-comprehensive analysis of beta-casein and epidermal growth factor receptor (EGFR)
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr050113n
Accession Number: 16083266
Keywords: Amino Acid Sequence
Animals
Caseins/*analysis/genetics
Cattle
Cell Line, Tumor
*Chromatography, Liquid/instrumentation/methods
ErbB Receptors/*analysis/genetics
Humans
*Mass Spectrometry/instrumentation/methods
Molecular Sequence Data
Peptides/analysis/genetics
*Protein Processing, Post-Translational
Proteome/*analysis
Proteomics/*methods
Abstract: We have developed a new and sensitive LC-MS platform, Extended Range Proteomic Analysis (ERPA), which is able to achieve very high sequence coverage and comprehensive characterization of post-translational modifications in complex proteins. This new platform provides advantages of both the top-down and bottom-up proteomic approaches by combining (i) digestion of the protein with an enzyme, such as Lys-C, which cuts less frequently than trypsin, leading to on average a higher molecular weight peptide size, (ii) high-performance LC separation of the resulting fragments, (iii) a new data acquisition strategy using the LTQ-FTMS, a hybrid mass spectrometer that couples a linear ion trap with a Fourier transform ion cyclotron resonance (FTICR) cell, for analysis of peptides in the range of 0.5 to 10 kDa, and (iv) new data analysis methods for assigning large peptide structures and determining the site of attachment of post-translational modifications as well as structural features from the accurate precursor mass together with MS(2) and MS(3) fragmentations. The LC retention of the Lys-C fragments is increased, relative to a tryptic digest, due to the generally greater hydrophobicity of the larger peptides, a result that is particularly important for peptides containing hydrophilic modifications such as glycosylation and phosphorylation. Furthermore, additional positively charged arginine and lysine residues in the Lys-C fragments enhance the sensitivity of the post-translationally modified phospho- and glycopeptides by at least 10-fold relative to tryptic fragments. In typical operation, the FTICR cell provides a survey scan with the high mass resolution (> 100 000) and accurate mass (<2 ppm) to characterize the higher charge-state precursor ions of the larger peptides. In parallel, the linear ion trap provides MS(2) and MS(3) fragmentation spectra, with a scan speed sufficiently fast for on-line LC-MS. Together, these data provide multiple means to determine or enhance the confidence of assignment of large or complicated peptide. Using ERPA, we demonstrate >95% sequence coverage in the analysis of two heavily phosphorylated and glycosylated proteins, beta-casein at the 50 fmole level and the epidermal growth factor receptor (EGFR) at the 1 pmole level. In summary, the combination of digestion strategy, high-performance separation, and the hybrid LTQ-FTMS instrument enables comprehensive characterization of large proteins, including posttranslational modifications.
Notes: Wu, Shiaw-Lin
Kim, Jeongkwon
Hancock, William S
Karger, Barry
GM 15847/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, P.H.S.
United States
J Proteome Res. 2005 Jul-Aug;4(4):1155-70. doi: 10.1021/pr050113n.
Author Address: Barnett Institute and Department of Chemistry, Northeastern University, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1969
Author: Wu, W., Zheng, X., Wang, J., Yang, T., Dai, W., Song, S., Fang, L., Wang, Y. and Gu, J.
Year: 2018
Title: O-GlcNAcylation on Rab3A attenuates its effects on mitochondrial oxidative phosphorylation and metastasis in hepatocellular carcinoma
Journal: Cell Death Dis
Volume: 9
Issue: 10
Pages: 970
Epub Date: 2018/09/22
Date: Sep 20
Short Title: O-GlcNAcylation on Rab3A attenuates its effects on mitochondrial oxidative phosphorylation and metastasis in hepatocellular carcinoma
Alternate Journal: Cell death & disease
DOI: 10.1038/s41419-018-0961-7
PMCID: PMC6148238
Accession Number: 30237463
Keywords: Acetylglucosamine/*metabolism
Adult
Animals
Blotting, Western
Carcinoma, Hepatocellular/genetics/*metabolism
Cell Line, Tumor
Cell Movement/physiology
Cell Proliferation/genetics/physiology
Disease Progression
Gene Expression Regulation, Neoplastic/genetics/physiology
Hep G2 Cells
Humans
Hydrogen Peroxide/metabolism
Immunohistochemistry
Immunoprecipitation
Liver Neoplasms/genetics/*metabolism
Male
Mice
Mice, Inbred BALB C
Mice, Nude
Middle Aged
Mitochondria/metabolism/pathology
Oxidative Phosphorylation
Oxygen Consumption/physiology
RNA, Messenger/metabolism
Reactive Oxygen Species/metabolism
Real-Time Polymerase Chain Reaction
Signal Transduction/physiology
Superoxides/metabolism
rab3A GTP-Binding Protein/genetics/*metabolism
Abstract: Rab3A is a small Ras-like GTPase critical for membrane traffic. Although the functions of Rab3A have been reported in several cancers, the roles of Rab3A in hepatocellular carcinoma (HCC) have never been determined. To investigate the potential roles of Rab3A in HCC progression, we first determined Rab3A levels in HCC tissues and observed upregulated mRNA and protein levels of Rab3A in most tumor tissues. However, in vitro data showed that decreasing Rab3A in most HCC cell lines conferred no significant effects and overexpressing Rab3A in PLC/PRF/5 cells even inhibited migration and invasion. Meanwhile, the upregulation of Rab3A in HCC patients did not correlate with metastasis or overall survival of HCC patients. These contradict data suggested that Rab3A might act as metastatic suppressor and its effects might be attenuated in most HCC cells. Further experiments revealed that O-GlcNAcylation on Rab3A was key for attenuating Rab3A-mediated effects by regulating its GTP-binding activity, and verified the effects of Rab3A and its aberrant O-GlcNAcylation on HCC metastasis in vitro and in vivo. We also found that Rab3A and its O-GlcNAcylation had opposite roles in mitochondria oxidative phosphorylation (mtOXPHOS), and their functions on HCC metastasis were partially depended on their effects on metabolic reprogramming.
Notes: 2041-4889
Wu, Weicheng
Orcid: 0000-0001-5605-3898
Zheng, Xixi
Wang, Jing
Yang, Tianxiao
Dai, Wenjuan
Song, Shushu
Fang, Lan
Wang, Yilin
Gu, Jianxin
Journal Article
Research Support, Non-U.S. Gov't
Cell Death Dis. 2018 Sep 20;9(10):970. doi: 10.1038/s41419-018-0961-7.
Author Address: Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai, China. Wuweicheng@fudan.edu.cn.
Key Laboratory of Glycoconjugate Research Ministry of Health, School of Basic Medical Sciences, Fudan University, Shanghai, China. Wuweicheng@fudan.edu.cn.
Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai, China.
Key Laboratory of Glycoconjugate Research Ministry of Health, School of Basic Medical Sciences, Fudan University, Shanghai, China.
Shanghai Tenth People's Hospital of Tongji University, School of Medicine and School of Life Science and Technology, Tongji University, Shanghai, China.
Department of Hepatic Surgery, Fudan University Shanghai Cancer Center, Shanghai Medical College, Fudan University, Shanghai, 200032, China. Linglingwangyi@126.com.
Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032, China. Linglingwangyi@126.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1146
Author: Wu, Y., Eigenbrot, C., Liang, W. C., Stawicki, S., Shia, S., Fan, B., Ganesan, R., Lipari, M. T. and Kirchhofer, D.
Year: 2007
Title: Structural insight into distinct mechanisms of protease inhibition by antibodies
Journal: Proc Natl Acad Sci U S A
Volume: 104
Issue: 50
Pages: 19784-9
Epub Date: 2007/12/14
Date: Dec 11
Short Title: Structural insight into distinct mechanisms of protease inhibition by antibodies
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0708251104
PMCID: PMC2148376
Accession Number: 18077410
Keywords: Animals
Antibodies/*chemistry/*metabolism/pharmacology
Binding Sites, Antibody
Binding, Competitive/immunology
Catalysis
Humans
Mice
Protease Inhibitors/*chemistry/*metabolism/pharmacology
Rabbits
Serine Endopeptidases/*immunology/metabolism
Abstract: To better understand how the relatively flat antigen-combining sites of antibodies interact with the concave shaped substrate-binding clefts of proteases, we determined the structures of two antibodies in complex with the trypsin-like hepatocyte growth-factor activator (HGFA). The two inhibitory antibodies, Ab58 and Ab75, were generated from a human Fab phage display library with synthetic diversity in the three complementarity determining regions (H1, H2, and H3) of the heavy chain, mimicking the natural diversity of the human Ig repertoire. Biochemical studies and the structures of the Fab58:HGFA (3.5-A resolution) and the Fab75:HGFA (2.2-A resolution) complexes revealed that Ab58 obstructed substrate access to the active site, whereas Ab75 allosterically inhibited substrate hydrolysis. In both cases, the antibodies interacted with the same protruding element (99-loop), which forms part of the substrate-binding cleft. Ab58 inserted its H1 and H2 loops in the cleft to occupy important substrate interaction sites (S3 and S2). In contrast, Ab75 bound at the backside of the cleft to a region corresponding to thrombin exosite II, which is known to interact with allosteric effector molecules. In agreement with the structural analysis, binding assays with active site inhibitors and enzymatic assays showed that Ab58 is a competitive inhibitor, and Ab75 is a partial competitive inhibitor. These results provide structural insight into antibody-mediated protease inhibition. They suggest that unlike canonical inhibitors, antibodies may preferentially target protruding loops at the rim of the substrate-binding cleft to interfere with the catalytic machinery of proteases without requiring long insertion loops.
Notes: 1091-6490
Wu, Yan
Eigenbrot, Charles
Liang, Wei-Ching
Stawicki, Scott
Shia, Steven
Fan, Bin
Ganesan, Rajkumar
Lipari, Michael T
Kirchhofer, Daniel
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Proc Natl Acad Sci U S A. 2007 Dec 11;104(50):19784-9. doi: 10.1073/pnas.0708251104. Epub 2007 Dec 5.
Author Address: Departments of Antibody Engineering and Protein Engineering, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1960
Author: Wu, Z. L., Tatge, T. J., Grill, A. E. and Zou, Y.
Year: 2018
Title: Detecting and Imaging O-GlcNAc Sites Using Glycosyltransferases: A Systematic Approach to Study O-GlcNAc
Journal: Cell Chem Biol
Volume: 25
Issue: 11
Pages: 1428-1435.e3
Epub Date: 2018/08/14
Date: Nov 15
Short Title: Detecting and Imaging O-GlcNAc Sites Using Glycosyltransferases: A Systematic Approach to Study O-GlcNAc
Alternate Journal: Cell chemical biology
ISSN: 2451-9448
DOI: 10.1016/j.chembiol.2018.07.007
Accession Number: 30100348
Keywords: Acetylglucosamine/*analysis/metabolism
Acylation
Animals
Binding Sites
CHO Cells
Cricetulus
HEK293 Cells
Humans
N-Acetylgalactosaminyltransferases/chemistry/*metabolism
N-Acetylglucosaminyltransferases/chemistry/*metabolism
Recombinant Proteins/chemistry/metabolism
*Akt1
*B3galnt2
*Ck2
*O-GlcNAc
*Oga
*Ogt
*Pfkfb3
*click chemistry
*glycan imaging
*glycosyltransferases
Abstract: O-GlcNAcylation is a reversible serine/threonine glycosylation for regulating protein activity and availability inside cells. In a given protein, O-GlcNAcylated and unoccupied O-linked β-N-acetylglucosamine (O-GlcNAc) sites are referred to as closed and open sites, respectively. The balance between open and closed sites is believed to be dynamically regulated. In this report, closed sites are detected using in vitro incorporation of GalNAz by B3GALNT2, and open sites are detected by in vitro incorporation of GlcNAz by O-GlcNAc transferase (OGT), via click chemistry. For assessing total O-GlcNAc sites, a sample is O-GlcNAcylated in vitro by OGT before detecting by B3GALNT2. The methods are demonstrated on purified recombinant proteins including CK2, AKT1, and PFKFB3, and cellular extracts of HEK cells. Through O-GlcNAc imaging, the modification degree of O-GlcNAc in nuclei of Chinese hamster ovary cells was estimated. The detection and imaging of both open and closed O-GlcNAc sites provide a systematic approach to study this important post-translational modification.
Notes: 2451-9448
Wu, Zhengliang L
Tatge, Timothy J
Grill, Alex E
Zou, Yonglong
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell Chem Biol. 2018 Nov 15;25(11):1428-1435.e3. doi: 10.1016/j.chembiol.2018.07.007. Epub 2018 Aug 9.
Author Address: Department of Enzyme, Bio-techne, R&D Systems, Inc., 614 McKinley Place N.E., Minneapolis, MN 55413, USA. Electronic address: leon.wu@bio-techne.com.
Department of Enzyme, Bio-techne, R&D Systems, Inc., 614 McKinley Place N.E., Minneapolis, MN 55413, USA.
Department of Product Support Process Improvement, Bio-techne, R&D Systems, Inc., 614 McKinley Place N.E., Minneapolis, MN 55413, USA.
Department of Antibody Application Quality Control, Bio-techne, R&D Systems, Inc., 614 McKinley Place N.E., Minneapolis, MN 55413, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 806
Author: Wuttke, M., Müller, S., Nitsche, D. P., Paulsson, M., Hanisch, F. G. and Maurer, P.
Year: 2001
Title: Structural characterization of human recombinant and bone-derived bone sialoprotein. Functional implications for cell attachment and hydroxyapatite binding
Journal: J Biol Chem
Volume: 276
Issue: 39
Pages: 36839-48
Epub Date: 2001/07/19
Date: Sep 28
Short Title: Structural characterization of human recombinant and bone-derived bone sialoprotein. Functional implications for cell attachment and hydroxyapatite binding
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M105689200
Accession Number: 11459848
Keywords: Amino Acid Sequence
Blotting, Western
Bone and Bones/metabolism
Carbohydrate Sequence
Carbohydrates/chemistry
Cell Adhesion
Cell Line
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Circular Dichroism
Dose-Response Relationship, Drug
Durapatite/*metabolism
Electrophoresis, Polyacrylamide Gel
Glycosylation
Humans
Integrin-Binding Sialoprotein
Mass Spectrometry
Methylation
Microscopy, Electron
Molecular Sequence Data
Oligosaccharides/metabolism
Polysaccharides/chemistry
Protein Binding
Protein Processing, Post-Translational
Protein Structure, Secondary
Recombinant Proteins/*chemistry/metabolism
Sialoglycoproteins/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Time Factors
ortho-Aminobenzoates/metabolism
Abstract: Human bone sialoprotein (BSP) comprises 15% of the total noncollagenous proteins in bone and is thought to be involved in bone mineralization and remodeling. Recent data suggest a role for BSP in breast cancer and the development of bone metastases. We have produced full-length recombinant BSP in a human cell line and purified the protein from human bone retaining the native structure with proper folding and post-translational modifications. Mass spectrometry of bone-derived BSP revealed an average mass of 49 kDa and for recombinant BSP 57 kDa. The post-translational modifications contribute 30-40%. Carbohydrate analysis revealed 10 different complex-type N-glycans on both proteins and eight different O-glycans on recombinant BSP, four of those were found on bone-derived BSP. We could identify eight threonines modified by O-glycans, leaving the C terminus of the protein free of glycans. The recombinant protein showed similar secondary structures as bone-derived BSP. BSP was visualized in electron microscopy as a globule linked to a thread-like structure. The affinity for hydroxyapatite was higher for bone-derived BSP than for recombinant BSP. Cell adhesion assays showed that the binding of BSP to cells can be reversibly diminished by denaturation.
Notes: Wuttke, M
Müller, S
Nitsche, D P
Paulsson, M
Hanisch, F G
Maurer, P
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 2001 Sep 28;276(39):36839-48. doi: 10.1074/jbc.M105689200. Epub 2001 Jul 17.
Author Address: Institute for Biochemistry II, Medical Faculty, University of Cologne, D-50931 Cologne, Germany.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1394
Author: Xiao, Y. P., Morice, A. H., Compton, S. J. and Sadofsky, L.
Year: 2011
Title: N-linked glycosylation regulates human proteinase-activated receptor-1 cell surface expression and disarming via neutrophil proteinases and thermolysin
Journal: J Biol Chem
Volume: 286
Issue: 26
Pages: 22991-3002
Epub Date: 2011/05/10
Date: Jul 1
Short Title: N-linked glycosylation regulates human proteinase-activated receptor-1 cell surface expression and disarming via neutrophil proteinases and thermolysin
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.204271
PMCID: PMC3123067
Accession Number: 21550978
Keywords: Animals
CHO Cells
Cathepsin G/genetics/metabolism
Cell Line, Transformed
Cricetinae
Cricetulus
Gene Expression Regulation/*physiology
Glycosylation
Humans
Myeloblastin/genetics/*metabolism
Pancreatic Elastase/genetics/*metabolism
Protein Structure, Tertiary
Rats
Receptor, PAR-1/*biosynthesis/genetics
Thermolysin/genetics/metabolism
Abstract: Proteinase-activated receptor 1 (PAR(1)) induces activation of platelet and vascular cells after proteolytic cleavage of its extracellular N terminus by thrombin. In pathological situations, other proteinases may be generated in the circulation and might modify the responses of PAR(1) by cleaving extracellular domains. In this study, epitope-tagged wild-type human PAR(1) (hPAR(1)) and a panel of N-linked glycosylation-deficient mutant receptors were permanently expressed in epithelial cells (Kirsten murine sarcoma virus-transformed rat kidney cells and CHO cells). We have analyzed the role of N-linked glycosylation in regulating proteinase activation/disarming and cell global expression of hPAR(1). We reported for the first time that glycosylation in the N terminus of hPAR(1) downstream of the tethered ligand (especially Asn(75)) governs receptor disarming to trypsin, thermolysin, and the neutrophil proteinases elastase and proteinase 3 but not cathepsin G. In addition, hPAR(1) is heavily N-linked glycosylated and sialylated in epithelial cell lines, and glycosylation occurs at all five consensus sites, namely, Asn(35), Asn(62), Asn(75), Asn(250), and Asn(259). Removing these N-linked glycosylation sequons affected hPAR(1) cell surface expression to varying degrees, and N-linked glycosylation at extracellular loop 2 (especially Asn(250)) of hPAR(1) is essential for optimal receptor cell surface expression and receptor stability.
Notes: 1083-351x
Xiao, Yu Pei
Morice, Alyn H
Compton, Steven J
Sadofsky, Laura
Journal Article
J Biol Chem. 2011 Jul 1;286(26):22991-3002. doi: 10.1074/jbc.M110.204271. Epub 2011 May 6.
Author Address: Division of Cardiovascular and Respiratory Studies, University of Hull, Hull York Medical School, East Yorkshire, HU16 5JQ, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1287
Author: Xie, C., Zhu, J., Chen, X., Mi, L., Nishida, N. and Springer, T. A.
Year: 2010
Title: Structure of an integrin with an alphaI domain, complement receptor type 4
Journal: Embo j
Volume: 29
Issue: 3
Pages: 666-79
Epub Date: 2009/12/25
Date: Feb 3
Short Title: Structure of an integrin with an alphaI domain, complement receptor type 4
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1038/emboj.2009.367
PMCID: PMC2830704
Accession Number: 20033057
Keywords: Animals
Antibodies/immunology/physiology
Antibody Specificity
CHO Cells
Cricetinae
Cricetulus
Disulfides/chemistry/metabolism
Humans
Integrin alphaXbeta2/*chemistry/immunology
Integrins/chemistry/immunology
Models, Biological
Models, Molecular
Protein Structure, Quaternary
Protein Structure, Secondary
Protein Structure, Tertiary
Abstract: We report the structure of an integrin with an alphaI domain, alpha(X)beta(2), the complement receptor type 4. It was earlier expected that a fixed orientation between the alphaI domain and the beta-propeller domain in which it is inserted would be required for allosteric signal transmission. However, the alphaI domain is highly flexible, enabling two betaI domain conformational states to couple to three alphaI domain states, and greater accessibility for ligand recognition. Although alpha(X)beta(2) is bent similarly to integrins that lack alphaI domains, the terminal domains of the alpha- and beta-legs, calf-2 and beta-tail, are oriented differently than in alphaI-less integrins. Linkers extending to the transmembrane domains are unstructured. Previous mutations in the beta(2)-tail domain support the importance of extension, rather than a deadbolt, in integrin activation. The locations of further activating mutations and antibody epitopes show the critical role of extension, and conversion from the closed to the open headpiece conformation, in integrin activation. Differences among 10 molecules in crystal lattices provide unprecedented information on interdomain flexibility important for modelling integrin extension and activation.
Notes: 1460-2075
Xie, Can
Zhu, Jianghai
Chen, Xing
Mi, Lizhi
Nishida, Noritaka
Springer, Timothy A
R01 AI072765/AI/NIAID NIH HHS/United States
AI 72765/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
EMBO J. 2010 Feb 3;29(3):666-79. doi: 10.1038/emboj.2009.367. Epub 2009 Dec 24.
Author Address: Department of Pathology, Harvard Medical School, Immune Disease Institute and Children's Hospital, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1756
Author: Xie, S., Jin, N., Gu, J., Shi, J., Sun, J., Chu, D., Zhang, L., Dai, C. L., Gu, J. H., Gong, C. X., Iqbal, K. and Liu, F.
Year: 2016
Title: O-GlcNAcylation of protein kinase A catalytic subunits enhances its activity: a mechanism linked to learning and memory deficits in Alzheimer's disease
Journal: Aging Cell
Volume: 15
Issue: 3
Pages: 455-64
Epub Date: 2016/02/04
Date: Jun
Short Title: O-GlcNAcylation of protein kinase A catalytic subunits enhances its activity: a mechanism linked to learning and memory deficits in Alzheimer's disease
Alternate Journal: Aging cell
ISSN: 1474-9718 (Print)
1474-9718
DOI: 10.1111/acel.12449
PMCID: PMC4854926
Accession Number: 26840030
Keywords: Acetylglucosamine/*metabolism
Alzheimer Disease/pathology
Animals
Brain/drug effects/enzymology
Cyclic AMP Response Element-Binding Protein/metabolism
Cyclic AMP-Dependent Protein Kinase Catalytic Subunits/*metabolism
Diazooxonorleucine/pharmacology
Down-Regulation/drug effects
Glycosylation/drug effects
HEK293 Cells
HeLa Cells
Humans
*Learning/drug effects
Male
Memory/drug effects
Memory Disorders/*pathology
Mice, Inbred C57BL
Phosphorylation/drug effects
Protein Transport/drug effects
Rats, Sprague-Dawley
Subcellular Fractions/drug effects/enzymology
tau Proteins/metabolism
*Alzheimer's disease
*O-GlcNAcylation
*learning and memory
*protein kinase A
Abstract: Alzheimer's disease (AD) is characterized clinically by memory loss and cognitive decline. Protein kinase A (PKA)-CREB signaling plays a critical role in learning and memory. It is known that glucose uptake and O-GlcNAcylation are reduced in AD brain. In this study, we found that PKA catalytic subunits (PKAcs) were posttranslationally modified by O-linked N-acetylglucosamine (O-GlcNAc). O-GlcNAcylation regulated the subcellular location of PKAcα and PKAcβ and enhanced their kinase activity. Upregulation of O-GlcNAcylation in metabolically active rat brain slices by O-(2-acetamido-2-deoxy-d-glucopyranosylidenamino) N-phenylcarbamate (PUGNAc), an inhibitor of N-acetylglucosaminidase, increased the phosphorylation of tau at the PKA site, Ser214, but not at the non-PKA site, Thr205. In contrast, in rat and mouse brains, downregulation of O-GlcNAcylation caused decreases in the phosphorylation of CREB at Ser133 and of tau at Ser214, but not at Thr205. Reduction in O-GlcNAcylation through intracerebroventricular injection of 6-diazo-5-oxo-l-norleucine (DON), the inhibitor of glutamine fructose-6-phosphate amidotransferase, suppressed PKA-CREB signaling and impaired learning and memory in mice. These results indicate that in addition to cAMP and phosphorylation, O-GlcNAcylation is a novel mechanism that regulates PKA-CREB signaling. Downregulation of O-GlcNAcylation suppresses PKA-CREB signaling and consequently causes learning and memory deficits in AD.
Notes: 1474-9726
Xie, Shutao
Jin, Nana
Gu, Jianlan
Shi, Jianhua
Sun, Jianming
Chu, Dandan
Zhang, Liang
Dai, Chun-Ling
Gu, Jin-Hua
Gong, Cheng-Xin
Iqbal, Khalid
Liu, Fei
Journal Article
Research Support, Non-U.S. Gov't
Aging Cell. 2016 Jun;15(3):455-64. doi: 10.1111/acel.12449. Epub 2016 Feb 3.
Author Address: Jiangsu Key Laboratory of Neuroregeneration, Co-innovation Center of Neuroregeneration, Nantong University, Nantong, Jiangsu, 226001, China.
Department of Neurochemistry, Inge Grundke-Iqbal Research Floor, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York, 10314, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1676
Author: Xing, J., Liu, H., Yang, H., Chen, R., Chen, Y. and Xu, J.
Year: 2014
Title: Upregulation of Unc-51-like kinase 1 by nitric oxide stabilizes SIRT1, independent of autophagy
Journal: PLoS One
Volume: 9
Issue: 12
Pages: e116165
Epub Date: 2014/12/30
Short Title: Upregulation of Unc-51-like kinase 1 by nitric oxide stabilizes SIRT1, independent of autophagy
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0116165
PMCID: PMC4277463
Accession Number: 25541949
Keywords: Animals
*Autophagy
Autophagy-Related Protein-1 Homolog
Cell Line
HEK293 Cells
Human Umbilical Vein Endothelial Cells
Humans
Intracellular Signaling Peptides and Proteins/*metabolism
Male
Mice
Mice, Inbred C57BL
Mice, Knockout
Nitric Oxide/*metabolism
Nitric Oxide Synthase Type III/genetics/metabolism
Proteasome Endopeptidase Complex/metabolism
Protein-Serine-Threonine Kinases/*metabolism
Proteolysis
Sirtuin 1/*metabolism
Up-Regulation
Abstract: SIRT1 is central to the lifespan and vascular health, but undergoes degradation that contributes to several medical conditions, including diabetes. How SIRT1 turnover is regulated remains unclear. However, emerging evidence suggests that endothelial nitric oxide synthase (eNOS) positively regulates SIRT1 protein expression. We recently identified NO as an endogenous inhibitor of 26S proteasome functionality with a cellular reporter system. Here we extended this finding to a novel pathway that regulates SIRT1 protein breakdown. In cycloheximide (CHX)-treated endothelial cells, NONOate, an NO donor, and A23187, an eNOS activator, significantly stabilized SIRT1 protein. Similarly, NO enhanced SIRT1 protein, but not mRNA expression, in CHX-free cells. NO also stabilized an autophagy-related protein unc-51 like kinase (ULK1), but did not restore SIRT1 protein levels in ULK1-siRNA-treated cells or in mouse embryonic fibroblasts (MEF) from Ulk1-/- mice. This suggests that ULK1 mediated the NO regulation of SIRT1. Furthermore, adenoviral overexpression of ULK1 increased SIRT1 protein expression, while ULK1 siRNA treatment decreased it. Rapamycin-induced autophagy did not mimic these effects, suggesting that the effects of ULK1 were autophagy-independent. Treatment with MG132, a proteasome inhibitor, or siRNA of β-TrCP1, an E3 ligase, prevented SIRT1 reduction induced by ULK1-siRNA. Mechanistically, ULK1 negatively regulated 26S proteasome functionality, which was at least partly mediated by O-linked-GlcNAc transferase (OGT), probably by increased O-GlcNAc modification of proteasomal subunit Rpt2. The NO-ULK1-SIRT1 axis was likely operative in the whole animal: both ULK1 and SIRT1 protein levels were significantly reduced in tissue homogenates in eNOS-knockout mice (lung) and in db/db mice where eNOS is downregulated (lung and heart). Taken together, the results show that NO stabilizes SIRT1 by regulating 26S proteasome functionality through ULK1 and OGT, but not autophagy, in endothelial cells.
Notes: 1932-6203
Xing, Junhui
Liu, Hongtao
Yang, Huabing
Chen, Rui
Chen, Yuguo
Xu, Jian
10SDG2600164/American Heart Association-American Stroke Association/United States
14BGIA20030027/American Heart Association-American Stroke Association/United States
P20 GM104934/GM/NIGMS NIH HHS/United States
5P20GM104934-08/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
PLoS One. 2014 Dec 26;9(12):e116165. doi: 10.1371/journal.pone.0116165. eCollection 2014.
Author Address: Department of Emergency, Qilu Hospital, Shandong University, Jinan, China; Department of Medicine and Harold Hamm Oklahoma Diabetes Center, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, United States of America.
Department of Medicine and Harold Hamm Oklahoma Diabetes Center, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, United States of America.
Department of Physiology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, United States of America.
Department of Emergency, Qilu Hospital, Shandong University, Jinan, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 814
Author: Xiong, J. P., Stehle, T., Diefenbach, B., Zhang, R., Dunker, R., Scott, D. L., Joachimiak, A., Goodman, S. L. and Arnaout, M. A.
Year: 2001
Title: Crystal structure of the extracellular segment of integrin alpha Vbeta3
Journal: Science
Volume: 294
Issue: 5541
Pages: 339-45
Epub Date: 2001/09/08
Date: Oct 12
Short Title: Crystal structure of the extracellular segment of integrin alpha Vbeta3
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1064535
PMCID: PMC2885948
NIHMSID: NIHMS143503
Accession Number: 11546839
Keywords: Amino Acid Motifs
Amino Acid Sequence
Binding Sites
Calcium/metabolism
Crystallization
Crystallography, X-Ray
Dimerization
Humans
Ligands
Metals/metabolism
Models, Molecular
Molecular Sequence Data
Mutation
Protein Conformation
Protein Folding
Protein Structure, Quaternary
Protein Structure, Secondary
Protein Structure, Tertiary
Protein Subunits
Receptors, Vitronectin/*chemistry/genetics/metabolism
Sequence Alignment
Abstract: Integrins are alphabeta heterodimeric receptors that mediate divalent cation-dependent cell-cell and cell-matrix adhesion through tightly regulated interactions with ligands. We have solved the crystal structure of the extracellular portion of integrin alphaVbeta3 at 3.1 A resolution. Its 12 domains assemble into an ovoid "head" and two "tails." In the crystal, alphaVbeta3 is severely bent at a defined region in its tails, reflecting an unusual flexibility that may be linked to integrin regulation. The main inter-subunit interface lies within the head, between a seven-bladed beta-propeller from alphaV and an A domain from beta3, and bears a striking resemblance to the Galpha/Gbeta interface in G proteins. A metal ion-dependent adhesion site (MIDAS) in the betaA domain is positioned to participate in a ligand-binding interface formed of loops from the propeller and betaA domains. MIDAS lies adjacent to a calcium-binding site with a potential regulatory function.
Notes: 1095-9203
Xiong, J P
Stehle, T
Diefenbach, B
Zhang, R
Dunker, R
Scott, D L
Joachimiak, A
Goodman, S L
Arnaout, M A
HL54227/HL/NHLBI NIH HHS/United States
P50 GM062414/GM/NIGMS NIH HHS/United States
DK48549/DK/NIDDK NIH HHS/United States
P50 GM062414-02/GM/NIGMS NIH HHS/United States
AI45716/AI/NIAID NIH HHS/United States
DK50305/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Science. 2001 Oct 12;294(5541):339-45. doi: 10.1126/science.1064535. Epub 2001 Sep 6.
Author Address: Renal Unit, Leukocyte Biology & Inflammation Program, Structural Biology Program, Massachusetts General Hospital and Harvard Medical School, 149 13th Street, Charlestown, MA 02129, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1791
Author: Xiong, Z. J., Huang, J., Poda, G., Pomès, R. and Privé, G. G.
Year: 2016
Title: Structure of Human Acid Sphingomyelinase Reveals the Role of the Saposin Domain in Activating Substrate Hydrolysis
Journal: J Mol Biol
Volume: 428
Issue: 15
Pages: 3026-42
Epub Date: 2016/06/29
Date: Jul 31
Short Title: Structure of Human Acid Sphingomyelinase Reveals the Role of the Saposin Domain in Activating Substrate Hydrolysis
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2016.06.012
Accession Number: 27349982
Keywords: Catalytic Domain/physiology
Humans
Hydrolysis
Lipids/physiology
Lysosomes/metabolism
Membranes/metabolism
Proline/metabolism
Saposins/*metabolism
Sphingomyelin Phosphodiesterase/*metabolism
Static Electricity
*Lipid hydrolase
*Niemann-Pick disease
*interfacial catalysis
*saposin
*sphingomyelin
Abstract: Acid sphingomyelinase (ASM) is a lysosomal phosphodiesterase that catalyzes the hydrolysis of sphingomyelin to produce ceramide and phosphocholine. While other lysosomal sphingolipid hydrolases require a saposin activator protein for full activity, the ASM polypeptide incorporates a built-in N-terminal saposin domain and does not require an external activator protein. Here, we report the crystal structure of human ASM and describe the organization of the three main regions of the enzyme: the N-terminal saposin domain, the proline-rich connector, and the catalytic domain. The saposin domain is tightly associated along an edge of the large, bowl-shaped catalytic domain and adopts an open form that exposes a hydrophobic concave surface approximately 30Å from the catalytic center. The calculated electrostatic potential of the enzyme is electropositive at the acidic pH of the lysosome, consistent with the strict requirement for the presence of acidic lipids in target membranes. Docking studies indicate that sphingomyelin binds with the ceramide-phosphate group positioned at the binuclear zinc center and molecular dynamic simulations indicate that the intrinsic flexibility of the saposin domain is important for monomer-dimer exchange and for membrane interactions. Overall, ASM uses a combination of electrostatic and hydrophobic interactions to cause local disruptions of target bilayers in order to bring the lipid headgroup to the catalytic center in a membrane-bound reaction.
Notes: 1089-8638
Xiong, Zi-Jian
Huang, Jingjing
Poda, Gennady
Pomès, Régis
Privé, Gilbert G
CIHR/Canada
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2016 Jul 31;428(15):3026-42. doi: 10.1016/j.jmb.2016.06.012. Epub 2016 Jun 24.
Author Address: Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada.
Drug Discovery, Ontario Institute for Cancer Research, Toronto, Ontario, Canada; Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada.
Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada; Molecular Structure and Function, Hospital for Sick Children, Toronto, Ontario, Canada.
Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada; Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada; Princess Margaret Cancer Centre, Toronto, Ontario, Canada. Electronic address: prive@uhnres.utoronto.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1967
Author: Xu, C., Liu, G. D., Feng, L., Zhang, C. H. and Wang, F.
Year: 2018
Title: Identification of O-GlcNAcylation Modification in Diabetic Retinopathy and Crosstalk with Phosphorylation of STAT3 in Retina Vascular Endothelium Cells
Journal: Cell Physiol Biochem
Volume: 49
Issue: 4
Pages: 1389-1402
Epub Date: 2018/09/12
Short Title: Identification of O-GlcNAcylation Modification in Diabetic Retinopathy and Crosstalk with Phosphorylation of STAT3 in Retina Vascular Endothelium Cells
Alternate Journal: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology
ISSN: 1015-8987
DOI: 10.1159/000493444
Accession Number: 30205382
Keywords: Animals
Caspase 3/metabolism
Cattle
Cell Hypoxia
Cells, Cultured
Diabetes Mellitus, Experimental/chemically induced/pathology
Diabetic Retinopathy/metabolism/*pathology
Endothelium, Vascular/cytology/metabolism
Female
Glucose/pharmacology
Glycosylation/drug effects
Humans
Janus Kinase 2/metabolism
N-Acetylglucosaminyltransferases/antagonists & inhibitors/metabolism
Phosphorylation/drug effects
Rats
Rats, Sprague-Dawley
Retina/cytology/metabolism
STAT3 Transcription Factor/antagonists & inhibitors/*metabolism
Triterpenes/pharmacology
Apoptosis
Diabetic retinopathy
O-GlcNAcylation
Phosphorylation
Stat3
Abstract: BACKGROUND/AIMS: Recently, we observed an increase in O-GlcNAc (O-linked-ß-N-acetylglucosamine) modification, and signal transducer and activator of transcription proteins 3 (STAT3) expression in primary retinal vascular endothelial cells (RVECs) under high glucose conditions and tissues altered by diabetic retinopathy (DR). In this study, we focused on the correlations between O-GlcNAcylation and STAT3 phosphorylation, and their potential effects with regards to DR. METHODS: Expression of O-GlcNAcylation and STAT3 were detected in DR-affected tissues and primary RVECs. The relationship between O-GlcNAcylation and STAT3 was further delineated by immunoprecipitation and Western blot analysis. Effects of O-GlcNAcylation on human RVEC apoptosis and involved protein expression were assayed with flow cytometry and Western blot. RESULTS: Global O-GlcNAcylation and pSTAT3 levels were significantly elevated in diabetic rat retina and primary RVECs under high glucose conditions. In vitro assays demonstrated that the Tyr705 site was sensitive to high glucose. While O-GlcNAcylation inhibited p727STAT3 expression, augmented O-GlcNAcylation could balance p705STAT3 expression within relatively high levels corresponding to vascular endothelial growth factor (VEGF) changes. Immunoprecipitation revealed that STAT3 was modified by O-GlcNAcylation and phosphorylation simultaneously. Next, we observed that overexpression of O-GlcNAcylation could relieve human RVEC apoptosis related to the JAK2-Tyr705STAT3-VEGF pathway. CONCLUSION: O-GlcNAcylation could relieve RVECs apoptosis through the STAT3 pathway in DR, and O-GlcNAcylation combined with STAT3 phosphorylation might open up new insights into the mechanisms of DR and other diabetic complications.
Notes: 1421-9778
Xu, Chong
Liu, Guo Dong
Feng, Le
Zhang, Cong Hui
Wang, Fang
Journal Article
Germany
Cell Physiol Biochem. 2018;49(4):1389-1402. doi: 10.1159/000493444. Epub 2018 Sep 11.
Author Address: Department of Ophthalmology, Shanghai Tenth People's Hospital Affiliated to Tongji University School of Medicine, Shanghai, China.
Nanshan Maternity and Child Healthcare Hospital of Shenzhen, Shenzhen, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1022
Author: Xu, H., Fu, Y., Tian, W. and Cohen, D. M.
Year: 2006
Title: Glycosylation of the osmoresponsive transient receptor potential channel TRPV4 on Asn-651 influences membrane trafficking
Journal: Am J Physiol Renal Physiol
Volume: 290
Issue: 5
Pages: F1103-9
Epub Date: 2005/12/22
Date: May
Short Title: Glycosylation of the osmoresponsive transient receptor potential channel TRPV4 on Asn-651 influences membrane trafficking
Alternate Journal: American journal of physiology. Renal physiology
ISSN: 1931-857X (Print)
1522-1466
DOI: 10.1152/ajprenal.00245.2005
Accession Number: 16368742
Keywords: Amino Acid Sequence
Animals
Biological Transport/*physiology
COS Cells
*Cell Size
Chickens
Chlorocebus aethiops
Electrophysiology
Glycosylation
Immunoblotting
Ion Channels/*physiology
Kidney/cytology
Membrane Potentials
Mice
Molecular Sequence Data
Phylogeny
Rats
Sequence Homology
Structure-Activity Relationship
TRPV Cation Channels/*genetics/*physiology
Transfection
Water-Electrolyte Balance
Abstract: We identified a consensus N-linked glycosylation motif within the pore-forming loop between the fifth and sixth transmembrane segments of the osmoresponsive transient receptor potential (TRP) channel TRPV4. Mutation of this residue from Asn to Gln (i.e., TRPV4(N651Q)) resulted in loss of a slower migrating band on anti-TRPV4 immunoblots and a marked reduction in lectin-precipitable TRPV4 immunoreactivity. HEK293 cells transiently transfected with the mutant TRPV4(N651Q) exhibited increased calcium entry in response to hypotonic stress relative to wild-type TRPV4 transfectants. This increase in hypotonicity responsiveness was associated with an increase in plasma membrane targeting of TRPV4(N651Q) relative to wild-type TRPV4 in both HEK293 and COS-7 cells but had no effect on overall channel abundance in whole cell lysates. Residue N651 of TRPV4 is immediately adjacent to the pore-forming loop. Although glycosylation in this vicinity has not been reported for a TRP channel, the structurally related hexahelical hyperpolarization-activated cyclic nucleotide-gated channel, HCN2, and the voltage-gated potassium channel, human ether-a-go-go-related (HERG), share a nearly identically situated and experimentally confirmed N-linked glycosylation site which promotes rather than limits channel insertion into the plasma membrane. These data point to a potentially conserved structural and functional feature influencing membrane trafficking across diverse members of the voltage-gated-like ion channel superfamily.
Notes: Xu, Hongshi
Fu, Yi
Tian, Wei
Cohen, David M
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
Am J Physiol Renal Physiol. 2006 May;290(5):F1103-9. doi: 10.1152/ajprenal.00245.2005. Epub 2005 Dec 20.
Author Address: Mailcode PP262, Oregon Health and Science Univ., 3314 S.W. US Veterans Hospital Rd., Portland, OR 97239, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1540
Author: Xu, L., Go, E. P., Finney, J., Moon, H., Lantz, M., Rebecchi, K., Desaire, H. and Mure, M.
Year: 2013
Title: Post-translational modifications of recombinant human lysyl oxidase-like 2 (rhLOXL2) secreted from Drosophila S2 cells
Journal: J Biol Chem
Volume: 288
Issue: 8
Pages: 5357-63
Epub Date: 2013/01/16
Date: Feb 22
Short Title: Post-translational modifications of recombinant human lysyl oxidase-like 2 (rhLOXL2) secreted from Drosophila S2 cells
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.C112.421768
PMCID: PMC3581389
Accession Number: 23319596
Keywords: Amino Acid Oxidoreductases/*metabolism
Animals
Breast Neoplasms/metabolism
Carbohydrates/chemistry
Catalytic Domain
Cell Line
Drosophila melanogaster
Glycosylation
Humans
Lysine/analogs & derivatives/chemistry
Mass Spectrometry/methods
Mutagenesis, Site-Directed
Neoplasm Invasiveness
Neoplasm Metastasis
Peptides/chemistry
Polysaccharides/chemistry
Protein Folding
*Protein Processing, Post-Translational
Quinones/chemistry
Recombinant Proteins/*chemistry
Solvents/chemistry
Abstract: Human lysyl oxidase-like 2 (hLOXL2) is highly up-regulated in metastatic breast cancer cells and tissues and induces epithelial-to-mesenchymal transition, the first step of metastasis/invasion. hloxl2 encodes four N-terminal scavenger receptor cysteine-rich domains and the highly conserved C-terminal lysyl oxidase (LOX) catalytic domain. Here, we assessed the extent of the post-translational modifications of hLOXL2 using truncated recombinant proteins produced in Drosophila S2 cells. The recombinant proteins are soluble, in contrast to LOX, which is consistently reported to require 2-6 m urea for solubilization. The recombinant proteins also show activity in tropoelastin oxidation. After phenylhydrazine derivatization and trypsin digestion, we used mass spectrometry to identify peptides containing the derivatized lysine tyrosylquinone cross-link at Lys-653 and Tyr-689, as well as N-linked glycans at Asn-455 and Asn-644. Disruption of N-glycosylation by site-directed mutagenesis or tunicamycin treatment completely inhibited secretion so that only small quantities of inclusion bodies were detected. The N-glycosylation site at Asn-644 in the LOX catalytic domain is not conserved in human LOX (hLOX), although the LOX catalytic domain of hLOX shares ∼50% identity and ∼70% homology with hLOXL2. The catalytic domain of hLOX was not secreted from S2 cells using the same expression system. These results suggest that the N-glycan at Asn-644 of hLOXL2 enhances the solubility and stability of the LOX catalytic domain.
Notes: 1083-351x
Xu, Li
Go, Eden P
Finney, Joel
Moon, HeeJung
Lantz, Mason
Rebecchi, Kathryn
Desaire, Heather
Mure, Minae
R01 GM079446/GM/NIGMS NIH HHS/United States
P20 RR015563/RR/NCRR NIH HHS/United States
R01RR026061/RR/NCRR NIH HHS/United States
R01 RR026061/RR/NCRR NIH HHS/United States
GM079446-02/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
J Biol Chem. 2013 Feb 22;288(8):5357-63. doi: 10.1074/jbc.C112.421768. Epub 2013 Jan 14.
Author Address: Department of Chemistry, The University of Kansas, Lawrence, Kansas 66045, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1634
Author: Xu, Q., Yang, C., Du, Y., Chen, Y., Liu, H., Deng, M., Zhang, H., Zhang, L., Liu, T., Liu, Q., Wang, L., Lou, Z. and Pei, H.
Year: 2014
Title: AMPK regulates histone H2B O-GlcNAcylation
Journal: Nucleic Acids Res
Volume: 42
Issue: 9
Pages: 5594-604
Epub Date: 2014/04/03
Date: May
Short Title: AMPK regulates histone H2B O-GlcNAcylation
Alternate Journal: Nucleic acids research
ISSN: 0305-1048 (Print)
0305-1048
DOI: 10.1093/nar/gku236
PMCID: PMC4027166
Accession Number: 24692660
Keywords: AMP-Activated Protein Kinases/*physiology
Acetylglucosamine/metabolism
Chromatin/metabolism
Energy Metabolism
Epigenesis, Genetic
Glycosylation
Hep G2 Cells
Histones/*metabolism
Homeostasis
Humans
N-Acetylglucosaminyltransferases/metabolism
Phosphorylation
*Protein Processing, Post-Translational
Transcription, Genetic
Abstract: Histone H2B O-GlcNAcylation is an important post-translational modification of chromatin during gene transcription. However, how this epigenetic modification is regulated remains unclear. Here we found that the energy-sensing adenosine-monophosphate-activated protein kinase (AMPK) could suppress histone H2B O-GlcNAcylation. AMPK directly phosphorylates O-linked β-N-acetylglucosamine (O-GlcNAc) transferase (OGT). Although this phosphorylation does not regulate the enzymatic activity of OGT, it inhibits OGT-chromatin association, histone O-GlcNAcylation and gene transcription. Conversely, OGT also O-GlcNAcylates AMPK and positively regulates AMPK activity, creating a feedback loop. Taken together, these results reveal a crosstalk between the LKB1-AMPK and the hexosamine biosynthesis (HBP)-OGT pathways, which coordinate together for the sensing of nutrient state and regulation of gene transcription.
Notes: 1362-4962
Xu, Qiuran
Yang, Caihong
Du, Yu
Chen, Yali
Liu, Hailong
Deng, Min
Zhang, Haoxing
Zhang, Lei
Liu, Tongzheng
Liu, Qingguang
Wang, Liewei
Lou, Zhenkun
Pei, Huadong
P50 CA108961/CA/NCI NIH HHS/United States
R01 CA129344/CA/NCI NIH HHS/United States
R01 CA130996/CA/NCI NIH HHS/United States
R01 CA148940/CA/NCI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Nucleic Acids Res. 2014 May;42(9):5594-604. doi: 10.1093/nar/gku236. Epub 2014 Apr 1.
Author Address: Department of Hepatobiliary Surgery, First Affiliated Hospital of Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi 710061, China.
Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850, China.
Division of Oncology Research, Mayo Clinic, Rochester, MN 55905, USA.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850, China Division of Oncology Research, Mayo Clinic, Rochester, MN 55905, USA.
Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic, Rochester, MN 55905, USA peihuadong@126.com.
Division of Oncology Research, Mayo Clinic, Rochester, MN 55905, USA peihuadong@126.com.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 100850, China peihuadong@126.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 993
Author: Xu, T., Logsdon, N. J. and Walter, M. R.
Year: 2005
Title: Structure of insect-cell-derived IL-22
Journal: Acta Crystallogr D Biol Crystallogr
Volume: 61
Issue: Pt 7
Pages: 942-50
Epub Date: 2005/06/29
Date: Jul
Short Title: Structure of insect-cell-derived IL-22
Alternate Journal: Acta crystallographica. Section D, Biological crystallography
ISSN: 0907-4449 (Print)
0907-4449
DOI: 10.1107/s0907444905009601
Accession Number: 15983417
Keywords: Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray
Drosophila melanogaster/chemistry
Glycosylation
Interleukins/*chemistry
Protein Conformation
Abstract: The crystal structure of interleukin-22 expressed in Drosophila melanogaster S2 cells (IL-22(Dm)) has been determined at 2.6 A resolution. IL-22(Dm) crystals contain six molecules in the asymmetric unit. Comparison of IL-22(Dm) and IL-22(Ec) (interleukin-22 produced in Escherichia coli) structures reveals that N-linked glycosylation causes only minor structural changes to the cytokine. However, 1-4 A main-chain differences are observed between the six IL-22(Dm) monomers at regions corresponding to the IL-22R1 and IL-10R2 binding sites. The structure of the carbohydrate and the conformational variation of IL22(Dm) provide new insights into IL-22 receptor recognition.
Notes: Xu, Ting
Logsdon, Naomi J
Walter, Mark R
AI47300/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
United States
Acta Crystallogr D Biol Crystallogr. 2005 Jul;61(Pt 7):942-50. doi: 10.1107/S0907444905009601. Epub 2005 Jun 24.
Author Address: Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1453
Author: Xu, X., Gårdsvoll, H., Yuan, C., Lin, L., Ploug, M. and Huang, M.
Year: 2012
Title: Crystal structure of the urokinase receptor in a ligand-free form
Journal: J Mol Biol
Volume: 416
Issue: 5
Pages: 629-41
Epub Date: 2012/01/31
Date: Mar 9
Short Title: Crystal structure of the urokinase receptor in a ligand-free form
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2011.12.058
Accession Number: 22285761
Keywords: Binding Sites
Cell Membrane/metabolism
Crystallization
Crystallography, X-Ray
Humans
Ligands
Mannose-Binding Lectins/*chemistry/genetics/metabolism
Membrane Glycoproteins/*chemistry/genetics/metabolism
Models, Molecular
Mutation
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Cell Surface/*chemistry/genetics/metabolism
Recombinant Proteins/chemistry/genetics/metabolism
Structure-Activity Relationship
Urokinase-Type Plasminogen Activator/metabolism
Vitronectin/metabolism
Abstract: The urokinase receptor urokinase-type plasminogen activator receptor (uPAR) is a surface receptor capable of not only focalizing urokinase-type plasminogen activator (uPA)-mediated fibrinolysis to the pericellular micro-environment but also promoting cell migration and chemotaxis. Consistent with this multifunctional role, uPAR binds several extracellular ligands, including uPA and vitronectin. Structural studies suggest that uPAR possesses structural flexibility. It is, however, not clear whether this flexibility is an inherent property of the uPAR structure per se or whether it is induced upon ligand binding. The crystal structure of human uPAR in its ligand-free state would clarify this issue, but such information remains unfortunately elusive. We now report the crystal structures of a stabilized, human uPAR (H47C/N259C) in its ligand-free form to 2.4 Å and in complex with amino-terminal fragment (ATF) to 3.2 Å. The structure of uPAR(H47C/N259C) in complex with ATF resembles the wild-type uPAR·ATF complex, demonstrating that these mutations do not perturb the uPA binding properties of uPAR. The present structure of uPAR(H47C/N259C) provides the first structural definition of uPAR in its ligand-free form, which represents one of the biologically active conformations of uPAR as defined by extensive biochemical studies. The domain boundary between uPAR DI-DII domains is more flexible than the DII-DIII domain boundary. Two important structural features are highlighted by the present uPAR structure. First, the DI-DIII domain boundary may face the cell membrane. Second, loop 130-140 of uPAR plays a dynamic role during ligand loading/unloading. Together, these studies provide new insights into uPAR structure-function relationships, emphasizing the importance of the inter-domain dynamics of this modular receptor.
Notes: 1089-8638
Xu, Xiang
Gårdsvoll, Henrik
Yuan, Cai
Lin, Lin
Ploug, Michael
Huang, Mingdong
R01 HL086584/HL/NHLBI NIH HHS/United States
HL086584/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2012 Mar 9;416(5):629-41. doi: 10.1016/j.jmb.2011.12.058. Epub 2012 Jan 21.
Author Address: Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1314
Author: Xu, Y., Kershaw, N. J., Luo, C. S., Soo, P., Pocock, M. J., Czabotar, P. E., Hilton, D. J., Nicola, N. A., Garrett, T. P. and Zhang, J. G.
Year: 2010
Title: Crystal structure of the entire ectodomain of gp130: insights into the molecular assembly of the tall cytokine receptor complexes
Journal: J Biol Chem
Volume: 285
Issue: 28
Pages: 21214-8
Epub Date: 2010/05/22
Date: Jul 9
Short Title: Crystal structure of the entire ectodomain of gp130: insights into the molecular assembly of the tall cytokine receptor complexes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.C110.129502
PMCID: PMC2898449
Accession Number: 20489211
Keywords: Contactins
Crystallography, X-Ray/methods
Cytokines/metabolism
Dimerization
Fibronectins/chemistry
Humans
Interleukin-6/*chemistry
Ligands
Molecular Conformation
Neural Cell Adhesion Molecules/*chemistry
Phosphorylation
Protein Binding
Protein Conformation
Protein Structure, Tertiary
Signal Transduction
Structure-Activity Relationship
Abstract: gp130 is the shared signal-transducing receptor subunit for the large and important family of interleukin 6-like cytokines. Previous x-ray structures of ligand-receptor complexes of this family lack the three membrane-proximal domains that are essential for signal transduction. Here we report the crystal structure of the entire extracellular portion of human gp130 (domains 1-6, D1-D6) at 3.6 A resolution, in an unliganded form, as well as a higher resolution structure of the membrane-proximal fibronectin type III domains (D4-D6) at 1.9 A. This represents the first atomic resolution structure of the complete ectodomain of any "tall" cytokine receptor. These structures show that other than a reorientation of the D1 domain, there is little structural change in gp130 upon ligand binding. They also reveal that the interface between the D4 and D5 domains forms an acute bend in the gp130 structure. Key residues at this interface are highly conserved across the entire tall receptor family, suggesting that this acute bend may be a common feature of these receptors. Importantly, this geometry positions the C termini of the membrane-proximal fibronectin type III domains of the tall cytokine receptors in close proximity within the transmembrane complex, favorable for receptor-associated Janus kinases to trans-phosphorylate and activate each other.
Notes: 1083-351x
Xu, Yibin
Kershaw, Nadia J
Luo, Cindy S
Soo, Priscilla
Pocock, Michael J
Czabotar, Peter E
Hilton, Douglas J
Nicola, Nicos A
Garrett, Thomas P J
Zhang, Jian-Guo
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Jul 9;285(28):21214-8. doi: 10.1074/jbc.C110.129502. Epub 2010 May 20.
Author Address: Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Parkville, Victoria 3052, Australia.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1691
Author: Xu, Y., Soo, P., Walker, F., Zhang, H. H., Redpath, N., Tan, C. W., Nicola, N. A., Adams, T. E., Garrett, T. P., Zhang, J. G. and Burgess, A. W.
Year: 2015
Title: LRIG1 extracellular domain: structure and function analysis
Journal: J Mol Biol
Volume: 427
Issue: 10
Pages: 1934-48
Epub Date: 2015/03/15
Date: May 22
Short Title: LRIG1 extracellular domain: structure and function analysis
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2015.03.001
Accession Number: 25765764
Keywords: Biosensing Techniques
Colonic Neoplasms/metabolism/pathology
Crystallography, X-Ray
ErbB Receptors/*chemistry/*metabolism
Extracellular Matrix/*metabolism
HEK293 Cells
Humans
Ligands
Membrane Glycoproteins/*chemistry/*metabolism
Microscopy, Fluorescence
Models, Molecular
Protein Conformation
Protein Structure, Tertiary
Proteins/chemistry/metabolism
Structure-Activity Relationship
Surface Plasmon Resonance
Tumor Cells, Cultured
EGFR inhibition
Lingo-1
leucine-rich repeat domain
stem cell marker
Abstract: We have expressed and purified three soluble fragments of the human LRIG1-ECD (extracellular domain): the LRIG1-LRR (leucine-rich repeat) domain, the LRIG1-3Ig (immunoglobulin-like) domain, and the LRIG1-LRR-1Ig fragment using baculovirus vectors in insect cells. The two LRIG1 domains crystallised so that we have been able to determine the three-dimensional structures at 2.3Å resolution. We developed a three-dimensional structure for the LRIG1-ECD using homology modelling based on the LINGO-1 structure. The LRIG1-LRR domain and the LRIG1-LRR-1Ig fragment are monomers in solution, whereas the LRIG1-3Ig domain appears to be dimeric. We could not detect any binding of the LRIG1 domains or the LRIG1-LRR-1Ig fragment to the EGF receptor (EGFR), either in solution using biosensor analysis or when the EGFR was expressed on the cell surface. The FLAG-tagged LRIG1-LRR-1Ig fragment binds weakly to colon cancer cells regardless of the presence of EGFRs. Similarly, neither the soluble LRIG1-LRR nor the LRIG1-3Ig domains nor the full-length LRIG1 co-expressed in HEK293 cells inhibited ligand-stimulated activation of cell-surface EGFR.
Notes: 1089-8638
Xu, Yibin
Soo, Priscilla
Walker, Francesca
Zhang, Hui Hua
Redpath, Nicholas
Tan, Chin Wee
Nicola, Nicos A
Adams, Timothy E
Garrett, Thomas P
Zhang, Jian-Guo
Burgess, Antony W
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2015 May 22;427(10):1934-48. doi: 10.1016/j.jmb.2015.03.001. Epub 2015 Mar 9.
Author Address: Structural Biology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia; Cancer and Haematology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia.
Cancer and Haematology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia.
Structural Biology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia; Department of Medical Biology, University of Melbourne, Parkville, Victoria 3010, Australia.
Structural Biology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia.
Cancer and Haematology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia; Department of Medical Biology, University of Melbourne, Parkville, Victoria 3010, Australia.
CSIRO Manufacturing Flagship, Parkville, Victoria 3052, Australia.
Cancer and Haematology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia; Department of Medical Biology, University of Melbourne, Parkville, Victoria 3010, Australia. Electronic address: zhang@wehi.edu.au.
Structural Biology Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3052, Australia; Department of Medical Biology, University of Melbourne, Parkville, Victoria 3010, Australia; Department of Surgery, RMH, University of Melbourne, Parkville, Victoria 3010, Australia. Electronic address: tburgess@wehi.edu.au.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1993
Author: Xu, Z., Isaji, T., Fukuda, T., Wang, Y. and Gu, J.
Year: 2019
Title: O-GlcNAcylation regulates integrin-mediated cell adhesion and migration via formation of focal adhesion complexes
Journal: J Biol Chem
Volume: 294
Issue: 9
Pages: 3117-3124
Epub Date: 2018/12/28
Date: Mar 1
Short Title: O-GlcNAcylation regulates integrin-mediated cell adhesion and migration via formation of focal adhesion complexes
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.RA118.005923
PMCID: PMC6398131
Accession Number: 30587575
Keywords: Acetylglucosamine/*metabolism
*Cell Adhesion
*Cell Movement
Focal Adhesion Kinase 1/metabolism
Focal Adhesions/*metabolism
Gene Knockdown Techniques
HEK293 Cells
HeLa Cells
Humans
Integrin beta1/*metabolism
N-Acetylglucosaminyltransferases/deficiency/genetics/*metabolism
Paxillin/metabolism
*Protein Processing, Post-Translational
Talin/metabolism
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT)
*cell migration
*focal adhesions
*glycosylation
*integrin
this article
Abstract: O-GlcNAcylation is a post-translational modification of a protein serine or threonine residue catalyzed by O-GlcNAc transferase (OGT) in the nucleus and cytoplasm. O-GlcNAcylation plays important roles in the cellular signaling that affect the different biological functions of cells, depending upon cell type. However, whether or not O-GlcNAcylation regulates cell adhesion and migration remains unclear. Here, we used the doxycycline-inducible short hairpin RNA (shRNA) system to establish an OGT knockdown (KD) HeLa cell line and found that O-GlcNAcylation is a key regulator for cell adhesion, migration, and focal adhesion (FA) complex formation. The expression levels of OGT and O-GlcNAcylation were remarkably suppressed 24 h after induction of doxycycline. Knockdown of OGT significantly promoted cell adhesion, but it suppressed the cell migration on fibronectin. The immunostaining with paxillin, a marker for FA plaque, clearly showed that the number of FAs was increased in the KD cells compared with that in the control cells. The O-GlcNAcylation levels of paxillin, talin, and focal adhesion kinase were down-regulated in KD cells. Interestingly, the complex formation between integrin β1, focal adhesion kinase, paxillin, and talin was greatly increased in KD cells. Consistently, levels of active integrin β1 were significantly enhanced in KD cells, whereas they were decreased in cells overexpressing OGT. The data suggest a novel regulatory mechanism for O-GlcNAcylation during FA complex formation, which thereby affects integrin activation and integrin-mediated functions such as cell adhesion and migration.
Notes: 1083-351x
Xu, Zhiwei
Isaji, Tomoya
Fukuda, Tomohiko
Wang, Yuqin
Gu, Jianguo
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2019 Mar 1;294(9):3117-3124. doi: 10.1074/jbc.RA118.005923. Epub 2018 Dec 26.
Author Address: From the Division of Regulatory Glycobiology, Institute of Molecular Biomembrane and Glycobiology, Tohoku Medical and Pharmaceutical University, 4-4-1 Komatsushima, Aoba-ku, Sendai Miyagi 981-8558, Japan and.
the Department of Pharmacology, Pharmacy College, Nantong University, Nantong, Jiangsu Province 226001, China.
From the Division of Regulatory Glycobiology, Institute of Molecular Biomembrane and Glycobiology, Tohoku Medical and Pharmaceutical University, 4-4-1 Komatsushima, Aoba-ku, Sendai Miyagi 981-8558, Japan and jgu@tohoku-mpu.ac.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1608
Author: Yagi, H., Nakagawa, N., Saito, T., Kiyonari, H., Abe, T., Toda, T., Wu, S. W., Khoo, K. H., Oka, S. and Kato, K.
Year: 2013
Title: AGO61-dependent GlcNAc modification primes the formation of functional glycans on α-dystroglycan
Journal: Sci Rep
Volume: 3
Pages: 3288
Epub Date: 2013/11/22
Date: Nov 21
Short Title: AGO61-dependent GlcNAc modification primes the formation of functional glycans on α-dystroglycan
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/srep03288
PMCID: PMC3836086
Accession Number: 24256719
Keywords: Animals
COS Cells
Cell Line
Chlorocebus aethiops
Dystroglycans/*chemistry/metabolism
Glucosamine/chemistry/*metabolism
Glycosyltransferases/deficiency/genetics/*metabolism
Laminin/chemistry/metabolism
Mice
Mice, Knockout
Mutation
N-Acetylglucosaminyltransferases/deficiency/genetics/*metabolism
Peptides/analysis
Phosphorylation
Polysaccharides/chemistry/*metabolism
Recombinant Proteins/biosynthesis/chemistry/genetics
Tandem Mass Spectrometry
Abstract: Dystroglycanopathy is a major class of congenital muscular dystrophy that is caused by a deficiency of functional glycans on α-dystroglycan (α-DG) with laminin-binding activity. A product of a recently identified causative gene for dystroglycanopathy, AGO61, acted in vitro as a protein O-mannose β-1, 4-N-acetylglucosaminyltransferase, although it was not functionally characterized. Here we show the phenotypes of AGO61-knockout mice and demonstrate that AGO61 is indispensable for the formation of laminin-binding glycans of α-DG. AGO61-knockout mouse brain exhibited abnormal basal lamina formation and a neuronal migration defect due to a lack of laminin-binding glycans. Furthermore, our results indicate that functional α-DG glycosylation was primed by AGO61-dependent GlcNAc modifications of specific threonine-linked mannosyl moieties of α-DG. These findings provide a key missing link for understanding how the physiologically critical glycan motif is displayed on α-DG and provides new insights on the pathological mechanisms of dystroglycanopathy.
Notes: 2045-2322
Yagi, Hirokazu
Nakagawa, Naoki
Saito, Takuya
Kiyonari, Hiroshi
Abe, Takaya
Toda, Tatsushi
Wu, Sz-Wei
Khoo, Kay-Hooi
Oka, Shogo
Kato, Koichi
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2013 Nov 21;3:3288. doi: 10.1038/srep03288.
Author Address: 1] Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabe-dori, Mizuho-ku, Nagoya 467-8603, Japan [2].
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 711
Author: Yamada, A., Kubo, K., Takeshita, T., Harashima, N., Kawano, K., Mine, T., Sagawa, K., Sugamura, K. and Itoh, K.
Year: 1999
Title: Molecular cloning of a glycosylphosphatidylinositol-anchored molecule CDw108
Journal: J Immunol
Volume: 162
Issue: 7
Pages: 4094-100
Epub Date: 1999/04/14
Date: Apr 1
Short Title: Molecular cloning of a glycosylphosphatidylinositol-anchored molecule CDw108
Alternate Journal: Journal of immunology (Baltimore, Md. : 1950)
ISSN: 0022-1767 (Print)
0022-1767
Accession Number: 10201933
Keywords: Amino Acid Sequence
Antigens, CD/chemistry/*genetics/metabolism
Base Sequence
Blood Group Antigens/*genetics/immunology
Cells, Cultured
Chromosome Mapping
Chromosomes, Human, Pair 15/immunology
Cloning, Molecular
GPI-Linked Proteins
Glycosylphosphatidylinositols/*metabolism
Humans
Membrane Glycoproteins/chemistry/*genetics/metabolism
Molecular Sequence Data
Organ Specificity/genetics/immunology
*Semaphorins
Transfection
Tumor Cells, Cultured
Abstract: CDw108, also known as the John-Milton-Hagen human blood group Ag, is an 80-kDa glycosylphosphatidylinositol (GPI)-anchored membrane glycoprotein that is preferentially expressed on activated lymphocytes and E. The molecular characteristics and biological function of the CDw108 were not clarified previously. In this manuscript, we identify the cDNA clone containing the entire coding sequence of the CDw108 gene and report its molecular characteristics. The 1998-base pairs of the open reading frame of the cloned cDNA encoded a protein of 666 amino acids (aa), including the 46 aa of the signal peptide and the 19 aa of the GPI-anchor motif. Thus, the membrane-anchoring form of CDw108 was the 602 aa, and the estimated molecular mass of the unglycosylated form was 68 kDa. The RGD (Arg-Gly-Asp) cell attachment sequence and the five potential N-linked glycosylation sites were located on the membrane-anchoring form. Flow cytometric and immunoprecipitation analyses of the CDw108 cDNA transfectants confirmed that the cloned cDNA encoded the native form of CDw108. The CDw108 mRNA was expressed in activated PBMCs as well as in the spleen, thymus, testis, placenta, and brain, but was not expressed in any other tissues tested. Radiation hybrid mapping indicated that the CDw108 gene was located in the middle of the long arm of chromosome 15 (15q23-24). This molecular information will be critical for understanding the biological function of the CDw108 Ag.
Notes: Yamada, A
Kubo, K
Takeshita, T
Harashima, N
Kawano, K
Mine, T
Sagawa, K
Sugamura, K
Itoh, K
Journal Article
Research Support, Non-U.S. Gov't
United States
J Immunol. 1999 Apr 1;162(7):4094-100.
Author Address: Cancer Vaccine Development Division, Kurume University Research Center for Innovative Cancer Therapy, Department of Immunology, Japan. akiymd@med.kurume-u.ac.jp
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1702
Author: Yamagata, A., Sato, Y., Goto-Ito, S., Uemura, T., Maeda, A., Shiroshima, T., Yoshida, T. and Fukai, S.
Year: 2015
Title: Structure of Slitrk2-PTPδ complex reveals mechanisms for splicing-dependent trans-synaptic adhesion
Journal: Sci Rep
Volume: 5
Pages: 9686
Epub Date: 2015/05/20
Date: May 19
Short Title: Structure of Slitrk2-PTPδ complex reveals mechanisms for splicing-dependent trans-synaptic adhesion
Alternate Journal: Scientific reports
ISSN: 2045-2322
DOI: 10.1038/srep09686
PMCID: PMC4437028
Accession Number: 25989451
Keywords: Animals
Binding Sites
Humans
Mice
*Models, Molecular
Protein Binding
*Protein Conformation
Protein Interaction Domains and Motifs
Receptor-Like Protein Tyrosine Phosphatases, Class 2/*chemistry/genetics/metabolism
Repressor Proteins/chemistry/metabolism
Structure-Activity Relationship
Synapses/*physiology
Abstract: Selective binding between pre- and postsynaptic adhesion molecules can induce synaptic differentiation. Here we report the crystal structure of a synaptogenic trans-synaptic adhesion complex between Slit and Trk-like family member 2 (Slitrk2) and receptor protein tyrosine phosphatase (RPTP) δ. The structure and site-directed mutational analysis revealed the structural basis of splicing-dependent adhesion between Slitrks and type IIa RPTPs for inducing synaptic differentiation.
Notes: 2045-2322
Yamagata, Atsushi
Sato, Yusuke
Goto-Ito, Sakurako
Uemura, Takeshi
Maeda, Asami
Shiroshima, Tomoko
Yoshida, Tomoyuki
Fukai, Shuya
Journal Article
Research Support, Non-U.S. Gov't
Sci Rep. 2015 May 19;5:9686. doi: 10.1038/srep09686.
Author Address: 1] Structural Biology Laboratory, Life Science Division, Synchrotron Radiation Research Organization and Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo 113-0032, Japan [2] Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Chiba 277-8501, Japan [3] CREST, JST, Saitama 332-0012, Japan.
1] Structural Biology Laboratory, Life Science Division, Synchrotron Radiation Research Organization and Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo 113-0032, Japan [2] CREST, JST, Saitama 332-0012, Japan.
1] CREST, JST, Saitama 332-0012, Japan [2] Department of Molecular and Cellular Physiology, Shinshu University School of Medicine, Nagano 390-8621, Japan [3] Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Nagano 390-8621, Japan.
1] Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan [2] PRESTO, JST, Saitama 332-0012, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1697
Author: Yamagata, A., Yoshida, T., Sato, Y., Goto-Ito, S., Uemura, T., Maeda, A., Shiroshima, T., Iwasawa-Okamoto, S., Mori, H., Mishina, M. and Fukai, S.
Year: 2015
Title: Mechanisms of splicing-dependent trans-synaptic adhesion by PTPδ-IL1RAPL1/IL-1RAcP for synaptic differentiation
Journal: Nat Commun
Volume: 6
Pages: 6926
Epub Date: 2015/04/25
Date: Apr 24
Short Title: Mechanisms of splicing-dependent trans-synaptic adhesion by PTPδ-IL1RAPL1/IL-1RAcP for synaptic differentiation
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms7926
PMCID: PMC4423211
Accession Number: 25908590
Keywords: Animals
Cell Adhesion
Coculture Techniques
HEK293 Cells
Humans
Immunoglobulins/genetics/metabolism
Interleukin-1 Receptor Accessory Protein/genetics/*metabolism
Mice
Models, Molecular
Mutation
Neurons/physiology
Protein Conformation
Protein Isoforms
RNA Splicing/*physiology
Receptor-Like Protein Tyrosine Phosphatases, Class 2/genetics/*metabolism
Synapses/*physiology
Abstract: Synapse formation is triggered through trans-synaptic interaction between pairs of pre- and postsynaptic adhesion molecules, the specificity of which depends on splice inserts known as 'splice-insert signaling codes'. Receptor protein tyrosine phosphatase δ (PTPδ) can bidirectionally induce pre- and postsynaptic differentiation of neurons by trans-synaptically binding to interleukin-1 receptor accessory protein (IL-1RAcP) and IL-1RAcP-like-1 (IL1RAPL1) in a splicing-dependent manner. Here, we report crystal structures of PTPδ in complex with IL1RAPL1 and IL-1RAcP. The first immunoglobulin-like (Ig) domain of IL1RAPL1 directly recognizes the first splice insert, which is critical for binding to IL1RAPL1. The second splice insert functions as an adjustable linker that positions the Ig2 and Ig3 domains of PTPδ for simultaneously interacting with the Ig1 domain of IL1RAPL1 or IL-1RAcP. We further identified the IL1RAPL1-specific interaction, which appears coupled to the first-splice-insert-mediated interaction. Our results thus reveal the decoding mechanism of splice-insert signaling codes for synaptic differentiation induced by trans-synaptic adhesion between PTPδ and IL1RAPL1/IL-1RAcP.
Notes: 2041-1723
Yamagata, Atsushi
Yoshida, Tomoyuki
Sato, Yusuke
Goto-Ito, Sakurako
Uemura, Takeshi
Maeda, Asami
Shiroshima, Tomoko
Iwasawa-Okamoto, Shiho
Mori, Hisashi
Mishina, Masayoshi
Fukai, Shuya
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2015 Apr 24;6:6926. doi: 10.1038/ncomms7926.
Author Address: 1] Structural Biology Laboratory, Life Science Division, Synchrotron Radiation Research Organization and Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo 113-0032, Japan [2] Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Chiba 277-8501, Japan [3] CREST, JST, Saitama 332-0012, Japan.
1] Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan [2] Department of Molecular Neurobiology and Pharmacology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033, Japan [3] PRESTO, JST, Saitama 332-0012, Japan.
Structural Biology Laboratory, Life Science Division, Synchrotron Radiation Research Organization and Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo 113-0032, Japan.
1] CREST, JST, Saitama 332-0012, Japan [2] Department of Molecular Neurobiology and Pharmacology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033, Japan [3] Department of Molecular and Cellular Physiology, Shinshu University School of Medicine, Nagano 390-8621, Japan [4] Institute for Biomedical Sciences, Interdisciplinary Cluster for Cutting Edge Research, Shinshu University, Nagano 390-8621, Japan.
1] Structural Biology Laboratory, Life Science Division, Synchrotron Radiation Research Organization and Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo 113-0032, Japan [2] CREST, JST, Saitama 332-0012, Japan.
1] Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan [2] PRESTO, JST, Saitama 332-0012, Japan.
Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan.
1] Department of Molecular Neurobiology and Pharmacology, Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033, Japan [2] Brain Science Laboratory, The Research Organization of Science and Technology, Ritsumeikan University, Shiga 525-8577, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 963
Author: Yamaguchi, S., Ishihara, H., Tamura, A., Yamada, T., Takahashi, R., Takei, D., Katagiri, H. and Oka, Y.
Year: 2004
Title: Endoplasmic reticulum stress and N-glycosylation modulate expression of WFS1 protein
Journal: Biochem Biophys Res Commun
Volume: 325
Issue: 1
Pages: 250-6
Epub Date: 2004/11/04
Date: Dec 3
Short Title: Endoplasmic reticulum stress and N-glycosylation modulate expression of WFS1 protein
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291X (Print)
0006-291x
DOI: 10.1016/j.bbrc.2004.10.017
Accession Number: 15522226
Keywords: Animals
Dithiothreitol/pharmacology
Endoplasmic Reticulum/*metabolism
Enzyme Inhibitors/pharmacology
*Gene Expression Regulation
Glycosylation
Islets of Langerhans/cytology/drug effects/metabolism
Membrane Proteins/chemistry/genetics/*metabolism
Mice
Mutagenesis, Site-Directed
Thapsigargin/pharmacology
Tunicamycin/pharmacology
Abstract: Mutations of the WFS1 gene are responsible for two hereditary diseases, Wolfram syndrome and low frequency sensorineural hearing loss. The WFS1 protein is a glycoprotein located in the endoplasmic reticulum (ER) membrane but its function is poorly understood. Herein we show WFS1 mRNA and protein levels in pancreatic islets to be increased with ER-stress inducers, thapsigargin and dithiothreitol. Another ER-stress inducer, the N-glycosylation inhibitor tunicamycin, also raised WFS1 mRNA but not protein levels. Site-directed mutagenesis showed both Asn-663 and Asn-748 to be N-glycosylated in mouse WFS1 protein. The glycosylation-defective WFS1 protein, in which Asn-663 and Asn-748 had been substituted with aspartate, exhibited an increased protein turnover rate. Consistent with this, the WFS1 protein was more rapidly degraded in the presence of tunicamycin. These data indicate that ER-stress and N-glycosylation play important roles in WFS1 expression and stability, and also suggest regulatory roles for this protein in ER-stress induced cell death.
Notes: Yamaguchi, Suguru
Ishihara, Hisamitsu
Tamura, Akira
Yamada, Takahiro
Takahashi, Rui
Takei, Daisuke
Katagiri, Hideki
Oka, Yoshitomo
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2004 Dec 3;325(1):250-6. doi: 10.1016/j.bbrc.2004.10.017.
Author Address: Division of Molecular Metabolism and Diabetes, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 453
Author: Yamashita, K., Hitoi, A., Tsuchida, Y., Nishi, S. and Kobata, A.
Year: 1983
Title: Sugar chain of alpha-fetoprotein produced in human yolk sac tumor
Journal: Cancer Res
Volume: 43
Issue: 10
Pages: 4691-5
Epub Date: 1983/10/01
Date: Oct
Short Title: Sugar chain of alpha-fetoprotein produced in human yolk sac tumor
Alternate Journal: Cancer research
ISSN: 0008-5472 (Print)
0008-5472
Accession Number: 6192908
Keywords: Carbohydrate Sequence
Electrophoresis, Paper
Female
Humans
Mesonephroma/*analysis
Oligosaccharides/analysis
Ovarian Neoplasms/*analysis
alpha-Fetoproteins/*analysis
Abstract: The carbohydrate moiety of alpha-fetoprotein purified from human yolk sac tumors grown in nude mice was quantitatively released from the polypeptide chain as oligosaccharides by hydrazinolysis. The oligosaccharides were separated into a neutral and two acidic oligosaccharides by paper electrophoresis. By sequential exoglycosidase digestion in combination with per-O-methylation study and periodate oxidation, their structures were determined to be: Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(GlcNAc beta 1 leads to 4) (Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAcOT, Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(GlcNAc beta 1 leads to 4) (Sia alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAcOT; and Sia alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(GlcNAc beta 1 leads to 4) (Sia alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4 GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAcOT, in which Gal is galactose, GlcNac is N-acetylglucosamine, Man is mannose, Fuc is fucose, Sia is sialic acid, and Subscript OT is the NaB3H4-reduced oligosaccharide.
Notes: Yamashita, K
Hitoi, A
Tsuchida, Y
Nishi, S
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
Cancer Res. 1983 Oct;43(10):4691-5.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 312
Author: Yamashita, K., Inui, K., Totani, K., Kochibe, N., Furukawa, M. and Okada, S.
Year: 1990
Title: Characteristics of asparagine-linked sugar chains of sphingolipid activator protein 1 purified from normal human liver and GM1 gangliosidosis (type 1) liver
Journal: Biochemistry
Volume: 29
Issue: 12
Pages: 3030-9
Epub Date: 1990/03/27
Date: Mar 27
Short Title: Characteristics of asparagine-linked sugar chains of sphingolipid activator protein 1 purified from normal human liver and GM1 gangliosidosis (type 1) liver
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00464a020
Accession Number: 2110822
Keywords: Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Affinity
G(M1) Ganglioside
Gangliosidoses/*metabolism
Glycoproteins/*isolation & purification
Glycoside Hydrolases
Humans
Liver/*analysis
Molecular Sequence Data
Neuraminidase
Oligosaccharides/*isolation & purification
Reference Values
Saposins
Sphingolipid Activator Proteins
Abstract: Asparagine-linked sugar chains of sphingolipid activator protein 1 (SAP-1) purified from normal human liver and GM1 gangliosidosis (type 1) liver were comparatively investigated. Oligosaccharides released from the two SAP-1 samples by hydrazinolysis were fractionated by paper electrophoresis and by Aleuria aurantia lectin-Sepharose and Bio-Gel P-4 (under 400 mesh) column chromatography. Structures of oligosaccharides in each fraction were estimated from data on their effective molecular sizes, behavior on immobilized lectin columns with different carbohydrate-binding specificities, results of sequential digestion by exoglycosidases with different aglycon specificities, and methylation analysis. Sugar chains of SAP-1 purified from normal human liver and from GM1 gangliosidosis (type 1) liver were different from each other, although both of them were derived from complex-type sugar chains. The sugar chains of the former were the following eight degradation products from complex-type sugar chains by exoglycosidases in lysosomes: Man alpha 1----6(Man alpha 1----3)Man beta 1----4GlcNAc beta 1----4GlcNAcOT, Man alpha 1----6(Man alpha 1----3)Man beta 1----4GlcNAc beta 1----4(Fuc alpha 1----6)GlcNAcOT, Man alpha 1----6Man beta 1----4GlcNAc beta 1----4GlcNAcOT, Man alpha 1----6Man beta 1----4GlcNAc beta 1----4(Fuc alpha 1----6)GlcNAcOT, Man beta 1----4GlcNAc beta 1----4GlcNAcOT, Man beta 1----4GlcNAc beta 1----4(Fuc alpha 1----6)GlcNAcOT, GlcNAc beta 1----4GlcNAcOT, and GlcNAcOT. In contrast to these, the sugar chains of the latter were sialylated and nonsialylated mono- to tetraantennary complex-type sugar chains that were not fully degraded due to a metabolic defect in acid beta-galactosidase activity.
Notes: Yamashita, K
Inui, K
Totani, K
Kochibe, N
Furukawa, M
Okada, S
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 1990 Mar 27;29(12):3030-9. doi: 10.1021/bi00464a020.
Author Address: Department of Biochemistry, Kobe University School of Medicine, Hyogo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 499
Author: Yamashita, K., Liang, C. J., Funakoshi, S. and Kobata, A.
Year: 1981
Title: Structural studies of asparagine-linked sugar chains of human ceruloplasmin. Structural characteristics of the triantennary complex type sugar chains of human plasma glycoproteins
Journal: J Biol Chem
Volume: 256
Issue: 3
Pages: 1283-9
Epub Date: 1981/02/10
Date: Feb 10
Short Title: Structural studies of asparagine-linked sugar chains of human ceruloplasmin. Structural characteristics of the triantennary complex type sugar chains of human plasma glycoproteins
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7451505
Keywords: Asparagine/*analysis
Carbohydrate Conformation
Carbohydrate Sequence
*Ceruloplasmin
*Glycoproteins/urine
Glycoside Hydrolases
Humans
Methylation
Oligosaccharides/analysis
Abstract: The structures of the oligosaccharides released from human ceruloplasmin by hydrazinolysis were studied by sequential exoglycosidase digestion in combination with methylation analysis and periodate oxidation. A molecule of ceruloplasmin was shown to have two biantennary oligosaccharides A-1 and one triantennary oligosaccharide A-2, the structures of which are shown below. (Formula see text) The abbreviations used are: NeuAc, N-acetylneuraminic acid; Gal, galactose; GlcNAc, N-acetylglucosamine; Man, mannose.
Notes: Yamashita, K
Liang, C J
Funakoshi, S
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1981 Feb 10;256(3):1283-9.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 527
Author: Yamashita, K., Taketa, K., Nishi, S., Fukushima, K. and Ohkura, T.
Year: 1993
Title: Sugar chains of human cord serum alpha-fetoprotein: characteristics of N-linked sugar chains of glycoproteins produced in human liver and hepatocellular carcinomas
Journal: Cancer Res
Volume: 53
Issue: 13
Pages: 2970-5
Epub Date: 1993/07/01
Date: Jul 1
Short Title: Sugar chains of human cord serum alpha-fetoprotein: characteristics of N-linked sugar chains of glycoproteins produced in human liver and hepatocellular carcinomas
Alternate Journal: Cancer research
ISSN: 0008-5472 (Print)
0008-5472
Accession Number: 7686446
Keywords: Biomarkers, Tumor/*analysis
Carbohydrate Sequence
Carcinoma, Hepatocellular/*blood/chemistry/metabolism
Chromatography, Affinity
Electrophoresis, Paper
Fetal Blood/*chemistry
Glycoproteins/biosynthesis/blood/*chemistry
Humans
Lectins
Liver/embryology/*metabolism
Liver Diseases/blood/metabolism
Liver Neoplasms/*blood/chemistry/metabolism
Molecular Sequence Data
Oligosaccharides/biosynthesis/blood/*chemistry
Phytohemagglutinins
Sensitivity and Specificity
alpha-Fetoproteins/*biosynthesis/metabolism
Abstract: Human serum alpha-fetoprotein (AFP) is elevated in not only hepatocellular carcinoma (HCC) but also benign liver diseases. AFP produced in HCC and benign liver diseases was separated into several isoforms corresponding to different sugar chain structures by several types of lectin affinity electrophoresis, and the HCC-specific AFP isoform was discriminated from those of benign liver diseases. Because a small amount of HCC-specific AFP isoform was detected in cord serum AFP, the whole sugar chain structures of human cord serum AFP were determined, as follows: Neu5Ac alpha 2-->Gal beta 1-->4GlcNAc beta 1-->2Man alpha 1-->6(Neu5Ac alpha 2 -->6Gal beta 1-->4GlcNAc beta 1-->2Man alpha 1-->3)Man beta 1-->4R1 and R2, Gal beta 1-->4GlcNAc beta 1-->2Man alpha 1-->6(Neu5Ac alpha 2-->6Gal beta 1 -->4GlcNAc beta 1-->2Man alpha 1-->3)Man beta 1-->4R1 and R2, and Neu5Ac alpha 2-->3Gal beta 1-->4GlcNAc beta 1-->2Man alpha 1-->6(Neu5Ac alpha 2 -->6Gal beta 1-->4GlcNAc beta 1-->2Man alpha 1-->3)Man beta 1-->4R1 and R2 in the ratio of 81.6:8.9:9.5. R1 and R2 denote GlcNAc beta 1-->4GlcNAcOT (subscript OT represents an NaB3H4-reduced oligosaccharide) and GlcNAc beta 1-->4(Fuc alpha 1-->6)GlcNAcOT, respectively, and the ratio between R1 and R2 in the respective fractions was approximately 19:1. In contrast, the sugar chain structure of HCC highly specific AFP isoform was found to comprise a monosialyl-biantennary sugar chain with additional fucosylation of the proximal N-acetylglucosamine. Fucosylation of AFP produced in fetal liver increased in inverse proportion to the gestation period, in weeks, indicating that fucosylation of AFP in HCC may be related to the dedifferentiation of human hepatocytes through malignant transformation.
Notes: Yamashita, K
Taketa, K
Nishi, S
Fukushima, K
Ohkura, T
Journal Article
Research Support, Non-U.S. Gov't
United States
Cancer Res. 1993 Jul 1;53(13):2970-5.
Author Address: Department of Biochemistry, Sasaki Institute, Tokyo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1476
Author: Yan, J. and Aldrich, R. W.
Year: 2012
Title: BK potassium channel modulation by leucine-rich repeat-containing proteins
Journal: Proc Natl Acad Sci U S A
Volume: 109
Issue: 20
Pages: 7917-22
Epub Date: 2012/05/02
Date: May 15
Short Title: BK potassium channel modulation by leucine-rich repeat-containing proteins
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1205435109
PMCID: PMC3356614
Accession Number: 22547800
Keywords: Brain/metabolism
DNA Primers/genetics
HEK293 Cells
Humans
Ion Channel Gating/physiology
Large-Conductance Calcium-Activated Potassium Channels/*metabolism
Male
Membrane Proteins/*metabolism
*Models, Molecular
Neoplasm Proteins/*chemistry/metabolism
Patch-Clamp Techniques
Real-Time Polymerase Chain Reaction
Testis/metabolism
Abstract: Molecular diversity of ion channel structure and function underlies variability in electrical signaling in nerve, muscle, and nonexcitable cells. Regulation by variable auxiliary subunits is a major mechanism to generate tissue- or cell-specific diversity of ion channel function. Mammalian large-conductance, voltage- and calcium-activated potassium channels (BK, K(Ca)1.1) are ubiquitously expressed with diverse functions in different tissues or cell types, consisting of the pore-forming, voltage- and Ca(2+)-sensing α-subunits (BKα), either alone or together with the tissue-specific auxiliary β-subunits (β1-β4). We recently identified a leucine-rich repeat (LRR)-containing membrane protein, LRRC26, as a BK channel auxiliary subunit, which causes an unprecedented large negative shift (∼140 mV) in voltage dependence of channel activation. Here we report a group of LRRC26 paralogous proteins, LRRC52, LRRC55, and LRRC38 that potentially function as LRRC26-type auxiliary subunits of BK channels. LRRC52, LRRC55, and LRRC38 produce a marked shift in the BK channel's voltage dependence of activation in the hyperpolarizing direction by ∼100 mV, 50 mV, and 20 mV, respectively, in the absence of calcium. They along with LRRC26 show distinct expression in different human tissues: LRRC26 and LRRC38 mainly in secretory glands, LRRC52 in testis, and LRRC55 in brain. LRRC26 and its paralogs are structurally and functionally distinct from the β-subunits and we designate them as a γ family of the BK channel auxiliary proteins, which potentially regulate the channel's gating properties over a spectrum of different tissues or cell types.
Notes: 1091-6490
Yan, Jiusheng
Aldrich, Richard W
R01 NS078152/NS/NINDS NIH HHS/United States
R21 NS075118/NS/NINDS NIH HHS/United States
NS075118/NS/NINDS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Proc Natl Acad Sci U S A. 2012 May 15;109(20):7917-22. doi: 10.1073/pnas.1205435109. Epub 2012 Apr 30.
Author Address: Section of Neurobiology, Center for Learning and Memory, University of Texas, Austin, TX 78712, USA. jshyan@mail.utexas.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2014
Author: Yan, R., Zhao, X., Lei, J. and Zhou, Q.
Year: 2019
Title: Structure of the human LAT1-4F2hc heteromeric amino acid transporter complex
Journal: Nature
Volume: 568
Issue: 7750
Pages: 127-130
Epub Date: 2019/03/15
Date: Apr
Short Title: Structure of the human LAT1-4F2hc heteromeric amino acid transporter complex
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/s41586-019-1011-z
Accession Number: 30867591
Keywords: Amino Acids/metabolism
Binding Sites
Biological Transport
Carboxylic Acids/chemistry/pharmacology
*Cryoelectron Microscopy
Disulfides/chemistry/metabolism
Fusion Regulatory Protein 1, Heavy Chain/antagonists &
inhibitors/*chemistry/metabolism/*ultrastructure
Humans
Large Neutral Amino Acid-Transporter
1/*chemistry/genetics/metabolism/*ultrastructure
Models, Molecular
Multiprotein Complexes/antagonists & inhibitors/chemistry/metabolism/ultrastructure
Norbornanes/chemistry/pharmacology
Protein Binding
Protein Conformation
Abstract: The L-type amino acid transporter 1 (LAT1; also known as SLC7A5) catalyses the cross-membrane flux of large neutral amino acids in a sodium- and pH-independent manner(1-3). LAT1, an antiporter of the amino acid-polyamine-organocation superfamily, also catalyses the permeation of thyroid hormones, pharmaceutical drugs, and hormone precursors such as L-3,4-dihydroxyphenylalanine across membranes(2-6). Overexpression of LAT1 has been observed in a wide range of tumour cells, and it is thus a potential target for anti-cancer drugs(7-11). LAT1 forms a heteromeric amino acid transporter complex with 4F2 cell-surface antigen heavy chain (4F2hc; also known as SLC3A2)-a type II membrane glycoprotein that is essential for the stability of LAT1 and for its localization to the plasma membrane(8,9). Despite extensive cell-based characterization of the LAT1-4F2hc complex and structural determination of its homologues in bacteria, the interactions between LAT1 and 4F2hc and the working mechanism of the complex remain largely unknown(12-19). Here we report the cryo-electron microscopy structures of human LAT1-4F2hc alone and in complex with the inhibitor 2-amino-2-norbornanecarboxylic acid at resolutions of 3.3 Å and 3.5 Å, respectively. LAT1 exhibits an inward open conformation. Besides a disulfide bond association, LAT1 also interacts extensively with 4F2hc on the extracellular side, within the membrane, and on the intracellular side. Biochemical analysis reveals that 4F2hc is essential for the transport activity of the complex. Together, our characterizations shed light on the architecture of the LAT1-4F2hc complex, and provide insights into its function and the mechanisms through which it might be associated with disease.
Notes: 1476-4687
Yan, Renhong
Zhao, Xin
Lei, Jianlin
Zhou, Qiang
Journal Article
England
Nature. 2019 Apr;568(7750):127-130. doi: 10.1038/s41586-019-1011-z. Epub 2019 Mar 13.
Author Address: Beijing Advanced Innovation Center for Structural Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China.
Technology Center for Protein Sciences, Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing, China.
Institute of Biology, Westlake Institute for Advanced Study, School of Life Sciences, Westlake University, Hangzhou, China. zhouqiang@westlake.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 560
Author: Yan, S. B., Chao, Y. B. and van Halbeek, H.
Year: 1993
Title: Novel Asn-linked oligosaccharides terminating in GalNAc beta (1-->4)[Fuc alpha (1-->3)]GlcNAc beta (1-->.) are present in recombinant human protein C expressed in human kidney 293 cells
Journal: Glycobiology
Volume: 3
Issue: 6
Pages: 597-608
Epub Date: 1993/12/01
Date: Dec
Short Title: Novel Asn-linked oligosaccharides terminating in GalNAc beta (1-->4)[Fuc alpha (1-->3)]GlcNAc beta (1-->.) are present in recombinant human protein C expressed in human kidney 293 cells
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/3.6.597
Accession Number: 8130392
Keywords: Asparagine/*chemistry
Blood Coagulation
Carbohydrate Conformation
Carbohydrate Sequence
Cells, Cultured
Chromatography, Ion Exchange
Gas Chromatography-Mass Spectrometry
Humans
Kidney/cytology/*metabolism
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Oligosaccharides/*analysis/biosynthesis
Protein C/biosynthesis/*chemistry/physiology
Recombinant Proteins/genetics/pharmacology
Abstract: Recombinant human Protein C (rHPC), expressed in human kidney 293 cells, has a higher anticoagulant activity than plasma HPC, while its in vivo circulatory half-life is essentially unaltered compared to that of the natural protein. In seeking to elucidate the molecular basis for the improved efficacy of the recombinant antithrombotic drug, we focused on the carbohydrate moiety of rHPC. Protein C is a heavily post-translationally modified serine protease with four N-glycosylation sites. Glycosyl composition analysis of rHPC revealed a 5-fold higher fucose content and a 2-fold lower sialic acid content compared to plasma HPC. In addition, we found that rHPC contains N-acetylgalactosamine (2.6 mol GalNAc/mol rHPC) in its Asn-linked oligosaccharides, while plasma HPC is devoid of GalNAc. The Asn-linked oligosaccharides of rHPC were released by N-glycanase and separated into 25 fractions by high-pH anion-exchange chromatography. The most abundant oligosaccharides were structurally characterized by glycosyl composition and linkage analysis, in conjunction with 1H-NMR spectroscopy at 600 MHz. The structure of the major neutral oligosaccharide in rHPC was determined to be: [formula: see text] Two representatives of the sialylated oligosaccharides in rHPC are: [formula: see text] and [formula: see text] Thus, many of the Asn-linked oligosaccharides in rHPC were found to terminate in GalNAc beta (1-->4)GlcNAc beta (1-->.), in NeuAc alpha (2-->6)GalNAc beta (1-->4)GlcNAc beta (1-->.), and/or in GalNAc beta (1-->4)[Fuc alpha (1-->3)]GlcNAc beta (1-->.). Since the latter trisaccharide was first [Yan, S.B., Chao, B.Y. and Van Halbeek,H. (1992) J. Cell. Biochem., 16D, 151] observed in the Asn-linked oligosaccharides of rHPC derived from human kidney 293 cells, we propose to label the GalNAc beta-(1-->4)[Fuc alpha (1-->3)]GlcNAc beta (1-->.) terminal trisaccharide the PC-293 determinant. The PC-293-containing oligosaccharides may contribute to the higher anticoagulant activity of rHPC as compared to plasma HPC.
Notes: Yan, S B
Chao, Y B
van Halbeek, H
P41-RR-05351/RR/NCRR NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
Glycobiology. 1993 Dec;3(6):597-608. doi: 10.1093/glycob/3.6.597.
Author Address: Eli Lilly and Company, Indianapolis, IN 46285.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1172
Author: Yan, Y., Scott, D. J., Wilkinson, T. N., Ji, J., Tregear, G. W. and Bathgate, R. A.
Year: 2008
Title: Identification of the N-linked glycosylation sites of the human relaxin receptor and effect of glycosylation on receptor function
Journal: Biochemistry
Volume: 47
Issue: 26
Pages: 6953-68
Epub Date: 2008/06/07
Date: Jul 1
Short Title: Identification of the N-linked glycosylation sites of the human relaxin receptor and effect of glycosylation on receptor function
Alternate Journal: Biochemistry
ISSN: 0006-2960
DOI: 10.1021/bi800535b
Accession Number: 18533687
Keywords: Amino Acid Sequence
Animals
Cell Line
Conserved Sequence
Cyclic AMP/metabolism
Glycosylation
Humans
Mice
Models, Molecular
Molecular Sequence Data
Mutation/genetics
Protein Structure, Tertiary
Rats
Receptors, G-Protein-Coupled/*chemistry/genetics/*metabolism
Receptors, Peptide/*chemistry/genetics/*metabolism
Sequence Alignment
Abstract: The relaxin receptor, RXFP1, is a member of the leucine-rich repeat-containing G-protein-coupled receptor (LGR) family. These receptors are characterized by a large extracellular ectodomain containing leucine-rich repeats which contain the primary ligand binding site. RXFP1 contains six putative Asn-linked glycosylation sites in the ectodomain at positions Asn-14, Asn-105, Asn-242, Asn-250, Asn-303, and Asn-346, which are highly conserved across species. N-Linked glycosylation is the most common post-translational modification of G-protein-coupled receptors, although its role in modulating receptor function differs. We herein investigate the actual N-linked glycosylation status of RXFP1 and the functional ramifications of these post-translational modifications. Site-directed mutagenesis was utilized to generate single- or multiple-glycosylation site mutants of FLAG-tagged human RXFP1 which were then transiently expressed in HEK-293T cells. Glycosylation status was analyzed by immunoprecipitation and Western blot and receptor function analyzed with an anti-FLAG ELISA, (33)P-H2 relaxin competition binding, and cAMP activity measurement. All of the potential N-glycosylation sites of RXFP1 were utilized in HEK-293T cells, and importantly, disruption of glycosylation at individual or combinations of double and triple sites had little effect on relaxin binding. However, combinations of glycosylation sites were required for cell surface expression and cAMP signaling. In particular, N-glycosylation at Asn-303 of RXFP1 was required for optimal intracellular cAMP signaling. Hence, as is the case for other LGR family members, N-glycosylation is essential for the transport of the receptor to the cell surface. Additionally, it is likely that glycosylation is also essential for the conformational changes required for G-protein coupling and subsequent cAMP signaling.
Notes: 1520-4995
Yan, Y
Scott, D J
Wilkinson, T N
Ji, J
Tregear, G W
Bathgate, R A D
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochemistry. 2008 Jul 1;47(26):6953-68. doi: 10.1021/bi800535b. Epub 2008 Jun 6.
Author Address: Department of Biochemistry and Molecular Biology, College of Life Sciences, The National Laboratory of Protein Engineering and Plant Genetic Engineering, Peking University, Beijing 100871, PR China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 348
Author: Yanagi, H., Yoshima, T., Ogawa, I. and Okamoto, M.
Year: 1989
Title: Recombinant human erythropoietin produced by Namalwa cells
Journal: Dna
Volume: 8
Issue: 6
Pages: 419-27
Epub Date: 1989/07/01
Date: Jul-Aug
Short Title: Recombinant human erythropoietin produced by Namalwa cells
Alternate Journal: DNA (Mary Ann Liebert, Inc.)
ISSN: 0198-0238 (Print)
0198-0238
DOI: 10.1089/dna.1.1989.8.419
Accession Number: 2550193
Keywords: Animals
Carbohydrate Sequence
Cell Line
Chromatography
Chromatography, Affinity
Chromatography, High Pressure Liquid
DNA/genetics
Durapatite
Erythropoietin/*genetics/metabolism/pharmacology
Humans
Hydroxyapatites
Iron/metabolism
Liver/metabolism
Mice
Molecular Sequence Data
Molecular Weight
Oligosaccharides/isolation & purification
Plasmids
Rats
Recombinant Proteins/isolation & purification/metabolism
Restriction Mapping
Abstract: To establish a practical exogenous gene expression system in human cells, a cDNA coding for human erythropoietin (EPO) was expressed in human B-lymphoblastoid Namalwa cells. The Namalwa-derived recombinant EPO was purified from the culture fluid by a simple three-step procedure. The Namalwa EPO showed an equivalent activity in vivo to that of human urinary EPO. Oligosaccharide structure analyses suggested that almost all N-linked oligosaccharide chains of Namalwa EPO are shared by urinary EPO. The two major N-linked oligosaccharides of Namalwa EPO were fucose-containing tetraantennary and fucose-containing triantennary structures.
Notes: Yanagi, H
Yoshima, T
Ogawa, I
Okamoto, M
Journal Article
United States
DNA. 1989 Jul-Aug;8(6):419-27. doi: 10.1089/dna.1.1989.8.419.
Author Address: Takarazuka Research Center, Sumitomo Chemical Co., Ltd., Hyogo, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 543
Author: Yang, C. Y., Gu, Z. W., Blanco-Vaca, F., Gaskell, S. J., Yang, M., Massey, J. B., Gotto, A. M., Jr. and Pownall, H. J.
Year: 1994
Title: Structure of human apolipoprotein D: locations of the intermolecular and intramolecular disulfide links
Journal: Biochemistry
Volume: 33
Issue: 41
Pages: 12451-5
Epub Date: 1994/10/18
Date: Oct 18
Short Title: Structure of human apolipoprotein D: locations of the intermolecular and intramolecular disulfide links
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi00207a011
Accession Number: 7918467
Keywords: Amino Acid Sequence
Apolipoprotein A-II/chemistry
Apolipoproteins/*chemistry/metabolism
Apolipoproteins D
Asparagine/metabolism
Chromatography, High Pressure Liquid
Chymotrypsin
Cyanogen Bromide
Cysteine/chemistry
Disulfides/*chemistry
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Peptide Fragments/chemistry
Protein Structure, Secondary
Trypsin
Abstract: We have determined the primary structure of human apolipoprotein D (apoD) by aligning peptides derived from digestions by cyanogen bromide, trypsin, and chymotrypsin. Our results confirm the primary structure derived from cDNA [Drayna et al. (1986) J. Biol. Chem. 261, 16535-16539]. ApoD consists of 169 amino acid residues, including 5 cysteines. Tryptic peptide analysis indicated that Cys41 and Cys16 are joined by a disulfide bridge. Using a combination of manual Edman degradations and mass spectrometric analysis on a purified cluster of chymotryptic fragments, we identified an intramolecular disulfide bridge between Cys8 and Cys114 and an intermolecular bridge between Cys116 of apoD and Cys6 of apoA-II. In addition, sites of N-glycosylation were found at Asn45 and Asn78. Because apoD contains two intramolecular disulfide linkages and has a high content of proline to disrupt alpha-helical structures, formation of the amphipathic helical regions that characterize the other soluble apolipoproteins is unlikely. We conclude that apoD binds to lipoprotein surfaces through structures other than alpha-helices, such as disulfide links.
Notes: Yang, C Y
Gu, Z W
Blanco-Vaca, F
Gaskell, S J
Yang, M
Massey, J B
Gotto, A M Jr
Pownall, H J
HL-27341/HL/NHLBI NIH HHS/United States
HL-30914/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Biochemistry. 1994 Oct 18;33(41):12451-5. doi: 10.1021/bi00207a011.
Author Address: Baylor College of Medicine, Houston, Texas 77030.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1994
Author: Yang, G., Zhou, R., Zhou, Q., Guo, X., Yan, C., Ke, M., Lei, J. and Shi, Y.
Year: 2019
Title: Structural basis of Notch recognition by human γ-secretase
Journal: Nature
Volume: 565
Issue: 7738
Pages: 192-197
Epub Date: 2019/01/02
Date: Jan
Short Title: Structural basis of Notch recognition by human γ-secretase
Alternate Journal: Nature
ISSN: 0028-0836
DOI: 10.1038/s41586-018-0813-8
Accession Number: 30598546
Keywords: Amino Acid Sequence
Amyloid Precursor Protein Secretases/chemistry/*metabolism/*ultrastructure
Amyloid beta-Protein Precursor/chemistry/metabolism
Animals
*Cryoelectron Microscopy
Humans
Mice
Models, Molecular
Protein Binding
Receptors, Notch/chemistry/*metabolism/*ultrastructure
Substrate Specificity
Abstract: Aberrant cleavage of Notch by γ-secretase leads to several types of cancer, but how γ-secretase recognizes its substrate remains unknown. Here we report the cryo-electron microscopy structure of human γ-secretase in complex with a Notch fragment at a resolution of 2.7 Å. The transmembrane helix of Notch is surrounded by three transmembrane domains of PS1, and the carboxyl-terminal β-strand of the Notch fragment forms a β-sheet with two substrate-induced β-strands of PS1 on the intracellular side. Formation of the hybrid β-sheet is essential for substrate cleavage, which occurs at the carboxyl-terminal end of the Notch transmembrane helix. PS1 undergoes pronounced conformational rearrangement upon substrate binding. These features reveal the structural basis of Notch recognition and have implications for the recruitment of the amyloid precursor protein by γ-secretase.
Notes: 1476-4687
Yang, Guanghui
Zhou, Rui
Zhou, Qiang
Guo, Xuefei
Yan, Chuangye
Ke, Meng
Lei, Jianlin
Shi, Yigong
Journal Article
Research Support, Non-U.S. Gov't
England
Nature. 2019 Jan;565(7738):192-197. doi: 10.1038/s41586-018-0813-8. Epub 2018 Dec 31.
Author Address: Beijing Advanced Innovation Center for Structural Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China.
Institute of Biology, Westlake Institute for Advanced Study, Hangzhou, China.
Technology Center for Protein Sciences, Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing, China.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China. shi-lab@tsinghua.edu.cn.
Institute of Biology, Westlake Institute for Advanced Study, Hangzhou, China. shi-lab@tsinghua.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 603
Author: Yang, S. X., Pollock, H. G. and Rawitch, A. B.
Year: 1996
Title: Glycosylation in human thyroglobulin: location of the N-linked oligosaccharide units and comparison with bovine thyroglobulin
Journal: Arch Biochem Biophys
Volume: 327
Issue: 1
Pages: 61-70
Epub Date: 1996/03/01
Date: Mar 1
Short Title: Glycosylation in human thyroglobulin: location of the N-linked oligosaccharide units and comparison with bovine thyroglobulin
Alternate Journal: Archives of biochemistry and biophysics
ISSN: 0003-9861 (Print)
0003-9861
DOI: 10.1006/abbi.1996.0093
Accession Number: 8615697
Keywords: Amino Acid Sequence
Animals
Carbohydrates/analysis
Cattle
Chromatography, Gel
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Glycopeptides/*chemistry/isolation & purification
Glycosylation
Humans
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Peptide Fragments/chemistry/isolation & purification
Peptide Mapping
*Protein Structure, Secondary
Sequence Homology, Amino Acid
Thyroglobulin/*chemistry/isolation & purification/metabolism
Thyroid Gland/chemistry/metabolism
Trypsin
Abstract: The amino acid sequence established for human thyroglobulin (hTG) from its cDNA sequence contains 20 putative N-linked glycosylation sites. We have characterized the glycopeptides contained in a tryptic digest of hTG in order to determine which sites are actually linked to carbohydrate. In addition, the distribution of oligosaccharide type(s) at these confirmed sites of N-linked glycosylation has been examined. Glycopeptides were purified using gel permeation chromatography followed by several steps of HPLC. The purified tryptic glycopeptides were characterized by gas phase sequencing and carbohydrate analysis and located within the amino acid sequence of thyroglobulin. Each of the recovered glycopeptides contained a consensus sequence for N-linked glycosylation. Of the 20 putative N-linked glycosylation sites in the human thyroglobulin polypeptide chain, 16 were shown to be actually glycosylated in the mature protein. Eight of these confirmed glycosylation sites (at positions 57, 465, 510, 729, 797, 1696, 1754, and 2230) appear to be linked to complex-type oligosaccharide units containing fucose and galactose in addition to mannose and glucosamine. Five sites (at positions 1200, 1329, 1993, 2275, and 2562) contain high mannose type units and two sites (at positions 179 and 1345) are linked to oligosaccharide units containing galactose in addition to mannose and glucosamine but no fucose and may be either hybrid or complex structures. In addition, position 928 was found to be degenerate in oligosaccharide structure and very different oligosaccharide composition types were found associated with peptides containing the same amino acid sequence. A high probability of a beta turn which would include the glycosylated asparagine residue was predicted for the amino acid sequence found at 13 of the 16 sites. The glycosylation pattern in hTG was also compared with the data recently reported for bovine thyroglobulin (bTG) (27) and as has been recently reported for bTG, no oligosaccharides of the high mannose type were found in the N-terminal portion of hTG. Only four of the 20 putative sites the sequence of hTG, at asparagine residues 91, 477, 1849, and 2102 were not represented in the purified glycopeptide population and are presumed to escape significant glycosylation.
Notes: Yang, S X
Pollock, H G
Rawitch, A B
NIDDK18896/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Arch Biochem Biophys. 1996 Mar 1;327(1):61-70. doi: 10.1006/abbi.1996.0093.
Author Address: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, 66160-7421, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2053
Author: Yang, S. Z., Xu, F., Yuan, K., Sun, Y., Zhou, T., Zhao, X., McDonald, J. M. and Chen, Y.
Year: 2020
Title: Regulation of pancreatic cancer TRAIL resistance by protein O-GlcNAcylation
Journal: Lab Invest
Volume: 100
Issue: 5
Pages: 777-785
Epub Date: 2020/01/04
Date: May
Short Title: Regulation of pancreatic cancer TRAIL resistance by protein O-GlcNAcylation
Alternate Journal: Laboratory investigation; a journal of technical methods and pathology
ISSN: 0023-6837 (Print)
0023-6837
DOI: 10.1038/s41374-019-0365-z
PMCID: PMC7183418
NIHMSID: NIHMS1546034
Accession Number: 31896813
Keywords: Acetylglucosamine/metabolism
Animals
Cell Line, Tumor
Drug Resistance, Neoplasm/*genetics
Humans
Male
Mice
Mice, Knockout
Mice, Nude
*N-Acetylglucosaminyltransferases/genetics/metabolism
*Pancreatic Neoplasms/genetics/metabolism
Signal Transduction/genetics
*TNF-Related Apoptosis-Inducing Ligand/genetics/metabolism
Abstract: TRAIL-activating therapy is promising in treating various cancers, including pancreatic cancer, a highly malignant neoplasm with poor prognosis. However, many pancreatic cancer cells are resistant to TRAIL-induced apoptosis despite their expression of intact death receptors (DRs). Protein O-GlcNAcylation is a versatile posttranslational modification that regulates various biological processes. Elevated protein O-GlcNAcylation has been recently linked to cancer cell growth and survival. In this study, we evaluated the role of protein O-GlcNAcylation in pancreatic cancer TRAIL resistance, and identified higher levels of O-GlcNAcylation in TRAIL-resistant pancreatic cancer cells. With gain- and loss-of-function of the O-GlcNAc-adding enzyme, O-GlcNActransferase (OGT), we determined that increasing O-GlcNAcylation rendered TRAIL-sensitive cells more resistant to TRA-8-induced apoptosis, while inhibiting O-GlcNAcylation promoted TRA-8-induced apoptosis in TRAIL-resistance cells. Furthermore, we demonstrated that OGT knockdown sensitized TRAIL-resistant cells to TRA-8 therapy in a mouse model in vivo. Mechanistic studies revealed direct O-GlcNAc modifications of DR5, which regulated TRA-8-induced DR5 oligomerization. We further defined that DR5 O-GlcNAcylation was independent of FADD, the adapter protein for the downstream death-inducing signaling. These studies have demonstrated an important role of protein O-GlcNAcylation in regulating TRAIL resistance of pancreatic cancer cells; and uncovered the contribution of O-GlcNAcylation to DR5 oligomerization and thus mediating DR-inducing signaling.
Notes: 1530-0307
Yang, Shan-Zhong
Xu, Fei
Yuan, Kaiyu
Sun, Yong
Zhou, Tong
Zhao, Xinyang
McDonald, Jay M
Chen, Yabing
Orcid: 0000-0001-9676-6657
I01 BX002296/BX/BLRD VA/United States
IK6 BX003617/BX/BLRD VA/United States
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Lab Invest. 2020 May;100(5):777-785. doi: 10.1038/s41374-019-0365-z. Epub 2020 Jan 2.
Author Address: Department of Pathology, University of Alabama at Birmingham, Birmingham, AL, USA.
Department of Gastrointestinal Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Department of Medicine, University of Alabama at Birmingham, Birmingham, AL, USA.
Department of Biochemistry, University of Alabama at Birmingham, Birmingham, AL, USA.
Research Department, Birmingham Veterans Affairs Medical Center, Birmingham, AL, 35294, USA.
Department of Pathology, University of Alabama at Birmingham, Birmingham, AL, USA. yabingchen@uabmc.edu.
Research Department, Birmingham Veterans Affairs Medical Center, Birmingham, AL, 35294, USA. yabingchen@uabmc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1984
Author: Yang, W., Ao, M., Hu, Y., Li, Q. K. and Zhang, H.
Year: 2018
Title: Mapping the O-glycoproteome using site-specific extraction of O-linked glycopeptides (EXoO)
Journal: Mol Syst Biol
Volume: 14
Issue: 11
Pages: e8486
Epub Date: 2018/11/22
Date: Nov 20
Short Title: Mapping the O-glycoproteome using site-specific extraction of O-linked glycopeptides (EXoO)
Alternate Journal: Molecular systems biology
ISSN: 1744-4292
DOI: 10.15252/msb.20188486
PMCID: PMC6243375
Accession Number: 30459171
Keywords: Cells, Cultured
Chromatography, Liquid
Glycoproteins/blood/*metabolism
Glycosylation
Humans
Kidney/*metabolism
Protein Processing, Post-Translational
Proteomics/*methods
Serum/*metabolism
Solid Phase Extraction
T-Lymphocytes/*metabolism
Tandem Mass Spectrometry
*O‐GalNAc
*O‐linked
*glycoproteomics
*glycosylation
*site‐specific
Abstract: Protein glycosylation is one of the most abundant post-translational modifications. However, detailed analysis of O-linked glycosylation, a major type of protein glycosylation, has been severely impeded by the scarcity of suitable methodologies. Here, a chemoenzymatic method is introduced for the site-specific extraction of O-linked glycopeptides (EXoO), which enabled the mapping of over 3,000 O-linked glycosylation sites and definition of their glycans on over 1,000 proteins in human kidney tissues, T cells, and serum. This large-scale localization of O-linked glycosylation sites demonstrated that EXoO is an effective method for defining the site-specific O-linked glycoproteome in different types of sample. Detailed structural analysis of the sites identified revealed conserved motifs and topological orientations facing extracellular space, the cell surface, the lumen of the Golgi, and the endoplasmic reticulum (ER). EXoO was also able to reveal significant differences in the O-linked glycoproteome of tumor and normal kidney tissues pointing to its broader use in clinical diagnostics and therapeutics.
Notes: 1744-4292
Yang, Weiming
Orcid: 0000-0002-5023-2155
Ao, Minghui
Hu, Yingwei
Li, Qing Kay
Zhang, Hui
U01 CA152813/CA/NCI NIH HHS/United States
U24 CA210985/CA/NCI NIH HHS/United States
R21 AI122382/AI/NIAID NIH HHS/United States
P01 HL107153/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Syst Biol. 2018 Nov 20;14(11):e8486. doi: 10.15252/msb.20188486.
Author Address: Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD, USA huizhang@jhu.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1071
Author: Yang, W. H., Kim, J. E., Nam, H. W., Ju, J. W., Kim, H. S., Kim, Y. S. and Cho, J. W.
Year: 2006
Title: Modification of p53 with O-linked N-acetylglucosamine regulates p53 activity and stability
Journal: Nat Cell Biol
Volume: 8
Issue: 10
Pages: 1074-83
Epub Date: 2006/09/12
Date: Oct
Short Title: Modification of p53 with O-linked N-acetylglucosamine regulates p53 activity and stability
Alternate Journal: Nature cell biology
ISSN: 1465-7392 (Print)
1465-7392
DOI: 10.1038/ncb1470
Accession Number: 16964247
Keywords: Acetylglucosamine/*metabolism
Breast Neoplasms/*metabolism
Cell Survival
Electrophoresis, Gel, Two-Dimensional
Humans
Immunoprecipitation
Lung Neoplasms/*metabolism
Phosphates/metabolism
Phosphorylation
Protein Processing, Post-Translational/drug effects
Proto-Oncogene Proteins c-mdm2/metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tumor Cells, Cultured
Tumor Suppressor Protein p53/*metabolism
Ubiquitin/*metabolism
Abstract: Post-translational addition of O-linked N-acetylglucosamine (O-GlcNAc) to p53 is known to occur, but the site of O-GlcNAcylation and its effects on p53 are not understood. Here, we show that Ser 149 of p53 is O-GlcNAcylated and that this modification is associated with decreased phosphorylation of p53 at Thr 155, which is a site that is targeted by the COP9 signalosome, resulting in decreased p53 ubiquitination. Accordingly, O-GlcNAcylation at Ser 149 stabilizes p53 by blocking ubiquitin-dependent proteolysis. Our results indicate that the dynamic interplay between O-GlcNAc and O-phosphate modifications coordinately regulate p53 stability and activity.
Notes: Yang, Won Ho
Kim, Ji Eun
Nam, Hyung Wook
Ju, Jung Won
Kim, Hoe Suk
Kim, Yu Sam
Cho, Jin Won
Journal Article
Research Support, Non-U.S. Gov't
England
Nat Cell Biol. 2006 Oct;8(10):1074-83. doi: 10.1038/ncb1470. Epub 2006 Sep 10.
Author Address: Department of Biology, Yonsei University, 134 Shinchon-dong, Seodaemun-gu, Seoul 120-749, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1281
Author: Yang, W. H., Park, S. Y., Ji, S., Kang, J. G., Kim, J. E., Song, H., Mook-Jung, I., Choe, K. M. and Cho, J. W.
Year: 2010
Title: O-GlcNAcylation regulates hyperglycemia-induced GPX1 activation
Journal: Biochem Biophys Res Commun
Volume: 391
Issue: 1
Pages: 756-61
Epub Date: 2009/12/01
Date: Jan 1
Short Title: O-GlcNAcylation regulates hyperglycemia-induced GPX1 activation
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2009.11.133
Accession Number: 19944066
Keywords: Acetylglucosamine/*metabolism
Acylation
Animals
Cell Line
Diabetes Mellitus/*enzymology
Enzyme Activation
Female
Glutathione Peroxidase/genetics/*metabolism
Humans
Hyperglycemia/*enzymology
Mice
Mice, Inbred C3H
Mice, Inbred C57BL
Rats
Abstract: Hyperglycemia induces activation of glutathione peroxidase 1 (GPX1), an anti-oxidant enzyme essential for cell survival during oxidative stress. However, the mechanism of GPX1 activation is unclear. Here, we report that hyperglycemia-induced protein glycosylation by O-linked N-acetylglucosamine (O-GlcNAc) is crucial for activation of GPX1 and for its binding to c-Abl and Arg kinases. GPX1 itself is modified with O-GlcNAc on its C-terminus. We also demonstrate that pharmacological injection of the O-GlcNAcase inhibitor NTZ induces GPX1 activation in the mouse liver. Our findings suggest a crucial role for GPX1 and its O-GlcNAc modification in hyperglycemia and diabetes mellitus.
Notes: 1090-2104
Yang, Won Ho
Park, Sang Yoon
Ji, Suena
Kang, Jeong Gu
Kim, Ji-Eun
Song, Hyundong
Mook-Jung, Inhee
Choe, Kwang-Min
Cho, Jin Won
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2010 Jan 1;391(1):756-61. doi: 10.1016/j.bbrc.2009.11.133. Epub 2009 Nov 26.
Author Address: Department of Biology, Yonsei University, Seoul 120-749, Republic of Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1208
Author: Yang, W. H., Park, S. Y., Nam, H. W., Kim, D. H., Kang, J. G., Kang, E. S., Kim, Y. S., Lee, H. C., Kim, K. S. and Cho, J. W.
Year: 2008
Title: NFkappaB activation is associated with its O-GlcNAcylation state under hyperglycemic conditions
Journal: Proc Natl Acad Sci U S A
Volume: 105
Issue: 45
Pages: 17345-50
Epub Date: 2008/11/08
Date: Nov 11
Short Title: NFkappaB activation is associated with its O-GlcNAcylation state under hyperglycemic conditions
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0806198105
PMCID: PMC2582288
Accession Number: 18988733
Keywords: Acetylation
Animals
Electrophoretic Mobility Shift Assay
Hyperglycemia/*metabolism
Immunoblotting
Immunoprecipitation
Luciferases
Mice
Mice, Knockout
Models, Biological
N-Acetylglucosaminyltransferases/*metabolism
NF-kappa B/genetics/*metabolism
Phosphorylation
Protein Processing, Post-Translational/*physiology
beta-N-Acetylhexosaminidases/*metabolism
Abstract: The transcription factor NFkappaB is activated by phosphorylation and acetylation and plays important roles in inflammatory and immune responses in the cell. Additionally, posttranslational modification of the NFkappaB p65 subunit by O-linked N-acetylglucosamine (O-GlcNAc) has been reported, but the modification site of O-GlcNAc on NFkappaB p65 and its exact function have not been elucidated. In this work, we show that O-GlcNAcylation of NFkappaB p65 decreases binding to IkappaB alpha and increases transcriptional activity under hyperglycemic conditions. Also, we demonstrate that both Thr-322 and Thr-352 of NFkappaB p65 can be modified with O-GlcNAc, but modification on Thr-352, not Thr-322, is important for transcriptional activation. Our findings suggest that site-specific O-GlcNAcylation may be a reason why NFkappaB activity increases continuously under hyperglycemic conditions.
Notes: 1091-6490
Yang, Won Ho
Park, Sang Yoon
Nam, Hyung Wook
Kim, Do Hyun
Kang, Jeong Gu
Kang, Eun Seok
Kim, Yu Sam
Lee, Hyun Chul
Kim, Kwan Soo
Cho, Jin Won
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2008 Nov 11;105(45):17345-50. doi: 10.1073/pnas.0806198105. Epub 2008 Nov 6.
Author Address: Department of Biology, Yonsei University, 134 Shinchon-dong, Seodaemun-gu, Seoul 120-749, Korea.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 801
Author: Yang, X., Su, K., Roos, M. D., Chang, Q., Paterson, A. J. and Kudlow, J. E.
Year: 2001
Title: O-linkage of N-acetylglucosamine to Sp1 activation domain inhibits its transcriptional capability
Journal: Proc Natl Acad Sci U S A
Volume: 98
Issue: 12
Pages: 6611-6
Epub Date: 2001/05/24
Date: Jun 5
Short Title: O-linkage of N-acetylglucosamine to Sp1 activation domain inhibits its transcriptional capability
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.111099998
PMCID: PMC34401
Accession Number: 11371615
Keywords: Acetylglucosamine/*metabolism
Binding Sites
DNA-Binding Proteins
Fungal Proteins/physiology
Glycosylation
HeLa Cells
Humans
*Saccharomyces cerevisiae Proteins
Sp1 Transcription Factor/chemistry/*metabolism
Streptozocin/pharmacology
Transcription Factors/physiology
*Transcription, Genetic
Abstract: The posttranslational modification of eukaryotic intracellular proteins by O-linked N-acetylglucosamine (O-GlcNAc) monosaccharides is essential for cell viability, yet its precise functional roles are largely unknown. O-GlcNAc transferase utilizes UDP-GlcNAc, the end product of hexosamine biosynthesis, to catalyze this modification. The availability of UDP-GlcNAc correlates with glycosylation levels of intracellular proteins as well as with transcriptional levels of some genes. Meanwhile, transcription factors and RNA polymerase II can be modified by O-GlcNAc. A linkage between transcription factor O-GlcNAcylation and transcriptional regulation therefore has been postulated. Here, we show that O-GlcNAcylation of a chimeric transcriptional activator containing the second activation domain of Sp1 decreases its transcriptional activity both in an in vitro transcription system and in living cells, which is in concert with our observation that O-GlcNAcylation of Sp1 activation domain blocks its in vitro and in vivo interactions with other Sp1 molecules and TATA-binding protein-associated factor II 110. Furthermore, overexpression of O-GlcNAc transferase specifically inhibits transcriptional activation by native Sp1 in cells. Thus, our studies provide direct evidence that O-GlcNAcylation of transcription factors is involved in transcriptional regulation.
Notes: 1091-6490
Yang, X
Su, K
Roos, M D
Chang, Q
Paterson, A J
Kudlow, J E
Journal Article
Research Support, U.S. Gov't, P.H.S.
Proc Natl Acad Sci U S A. 2001 Jun 5;98(12):6611-6. doi: 10.1073/pnas.111099998. Epub 2001 May 22.
Author Address: Departments of Cell Biology and Medicine, Division of Endocrinology and Metabolism, University of Alabama at Birmingham, Birmingham AL 35294, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 844
Author: Yang, X., Zhang, F. and Kudlow, J. E.
Year: 2002
Title: Recruitment of O-GlcNAc transferase to promoters by corepressor mSin3A: coupling protein O-GlcNAcylation to transcriptional repression
Journal: Cell
Volume: 110
Issue: 1
Pages: 69-80
Epub Date: 2002/08/02
Date: Jul 12
Short Title: Recruitment of O-GlcNAc transferase to promoters by corepressor mSin3A: coupling protein O-GlcNAcylation to transcriptional repression
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/s0092-8674(02)00810-3
Accession Number: 12150998
Keywords: Animal Population Groups
Animals
COS Cells
*Gene Expression Regulation
Gene Silencing
Genes, Reporter
Glycoproteins
Histone Deacetylases/metabolism/*physiology
Humans
N-Acetylglucosaminyltransferases/metabolism/*physiology
Promoter Regions, Genetic/physiology
Repressor Proteins/metabolism/*physiology
Sin3 Histone Deacetylase and Corepressor Complex
Transcription Factors/metabolism/*physiology
Transcription, Genetic
Tumor Cells, Cultured
Abstract: Transcription factors and RNA polymerase II can be modified by O-linked N-acetylglucosamine (O-GlcNAc) monosaccharides at serine or threonine residues, yet the precise functional roles of this modification are largely unknown. Here, we show that O-GlcNAc transferase (OGT), the enzyme that catalyzes this posttranslational modification, interacts with a histone deacetylase complex by binding to the corepressor mSin3A. Functionally, OGT and mSin3A cooperatively repress transcription in parallel with histone deacetylation. We propose that mSin3A targets OGT to promoters to inactivate transcription factors and RNA polymerase II by O-GlcNAc modification, which acts in concert with histone deacetylation to promote gene silencing in an efficient and specific manner.
Notes: Yang, Xiaoyong
Zhang, Fengxue
Kudlow, Jeffrey E
Journal Article
Research Support, Non-U.S. Gov't
United States
Cell. 2002 Jul 12;110(1):69-80. doi: 10.1016/s0092-8674(02)00810-3.
Author Address: Department of Medicine, Division of Endocrinology and Metabolism, Department of Cell Biology, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 935
Author: Yang, Y., Jun, C. D., Liu, J. H., Zhang, R., Joachimiak, A., Springer, T. A. and Wang, J. H.
Year: 2004
Title: Structural basis for dimerization of ICAM-1 on the cell surface
Journal: Mol Cell
Volume: 14
Issue: 2
Pages: 269-76
Epub Date: 2004/04/22
Date: Apr 23
Short Title: Structural basis for dimerization of ICAM-1 on the cell surface
Alternate Journal: Molecular cell
ISSN: 1097-2765 (Print)
1097-2765
DOI: 10.1016/s1097-2765(04)00204-7
Accession Number: 15099525
Keywords: Amino Acid Sequence
Animals
Binding Sites
CHO Cells
Cell Membrane/chemistry/*metabolism
Conserved Sequence
Cricetinae
Cricetulus
Crystallography, X-Ray
Dimerization
Disulfides
Glycosylation
Hydrophobic and Hydrophilic Interactions
Integrins/metabolism
Intercellular Adhesion Molecule-1/*chemistry
Models, Molecular
Molecular Sequence Data
Protein Structure, Secondary
Protein Structure, Tertiary
Sequence Homology, Amino Acid
Abstract: We have determined the 3.0 A crystal structure of the three C-terminal domains 3-5 (D3-D5) of ICAM-1. Combined with the previously known N-terminal two-domain structure (D1D2), a model of an entire ICAM-1 extracellular fragment has been constructed. This model should represent a general architecture of other ICAM family members, particularly ICAM-3 and ICAM-5. The observed intimate dimerization interaction at D4 and a stiff D4-D5 stem-like architecture provide a good structural explanation for the existence of preformed ICAM-1 cis dimers on the cell membrane. Together with another dimerization interface at D1, a band-like one-dimensional linear cluster of ICAM-1 on an antigen-presenting cell (APC) surface can be envisioned, which might explain the formation of an immunological synapse between an activated T cell and APC which is critical for T cell receptor signaling.
Notes: Yang, Yuting
Jun, Chang-Duk
Liu, Jin-Huan
Zhang, Rongguang
Joachimiak, Andrzej
Springer, Timothy A
Wang, Jia-Huai
HL48675/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
United States
Mol Cell. 2004 Apr 23;14(2):269-76. doi: 10.1016/s1097-2765(04)00204-7.
Author Address: Dana-Farber Cancer Institute, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1574
Author: Yang, Y., Liu, C., Lin, Y. L. and Li, F.
Year: 2013
Title: Structural insights into central hypertension regulation by human aminopeptidase A
Journal: J Biol Chem
Volume: 288
Issue: 35
Pages: 25638-25645
Epub Date: 2013/07/28
Date: Aug 30
Short Title: Structural insights into central hypertension regulation by human aminopeptidase A
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.494955
PMCID: PMC3757224
Accession Number: 23888046
Keywords: Antihypertensive Agents/chemistry/therapeutic use
Binding Sites
Calcium/*chemistry/metabolism
Catalytic Domain
Crystallography, X-Ray
Glutamyl Aminopeptidase/antagonists & inhibitors/*chemistry/metabolism
Humans
Hypertension/drug therapy/*enzymology
Peptides/chemistry
Protease Inhibitors/chemistry
Substrate Specificity
Aminopeptidase
Calcium-binding Proteins
Hypertension
Protein Structure
X-ray Crystallography
Abstract: Hypertension is regulated through both the central and systemic renin-angiotensin systems. In the central renin-angiotensin system, zinc-dependent aminopeptidase A (APA) up-regulates blood pressure by specifically cleaving the N-terminal aspartate, but not the adjacent arginine, from angiotensin II, a process facilitated by calcium. Here, we determined the crystal structures of human APA and its complexes with different ligands and identified a calcium-binding site in the S1 pocket of APA. Without calcium, the S1 pocket can bind both acidic and basic residues through formation of salt bridges with the charged side chains. In the presence of calcium, the binding of acidic residues is enhanced as they ligate the cation, whereas the binding of basic residues is no longer favorable due to charge repulsion. Of the peptidomimetic inhibitors of APA, amastatin has higher potency than bestatin by fitting better in the S1 pocket and interacting additionally with the S3' subsite. These results explain the calcium-modulated substrate specificity of APA in central hypertension regulation and can guide the design and development of brain-targeting antihypertensive APA inhibitors.
Notes: 1083-351x
Yang, Yang
Liu, Chang
Lin, Yi-Lun
Li, Fang
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2013 Aug 30;288(35):25638-25645. doi: 10.1074/jbc.M113.494955. Epub 2013 Jul 25.
Author Address: From the Department of Pharmacology, University of Minnesota Medical School, Minneapolis, Minnesota 55455.
From the Department of Pharmacology, University of Minnesota Medical School, Minneapolis, Minnesota 55455. Electronic address: lifang@umn.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1802
Author: Yao, B., Xu, Y., Wang, J., Qiao, Y., Zhang, Y., Zhang, X., Chen, Y., Wu, Q., Zhao, Y., Zhu, G., Sun, F., Li, Z. and Yuan, H.
Year: 2016
Title: Reciprocal regulation between O-GlcNAcylation and tribbles pseudokinase 2 (TRIB2) maintains transformative phenotypes in liver cancer cells
Journal: Cell Signal
Volume: 28
Issue: 11
Pages: 1703-12
Epub Date: 2016/08/16
Date: Nov
Short Title: Reciprocal regulation between O-GlcNAcylation and tribbles pseudokinase 2 (TRIB2) maintains transformative phenotypes in liver cancer cells
Alternate Journal: Cellular signalling
ISSN: 0898-6568
DOI: 10.1016/j.cellsig.2016.08.003
Accession Number: 27515988
Keywords: Acetylglucosamine/*metabolism
Animals
Biosynthetic Pathways/drug effects
Calcium-Calmodulin-Dependent Protein Kinases/*metabolism
Cell Line, Tumor
Cell Transformation, Neoplastic/drug effects/*metabolism/*pathology
Glucose/pharmacology
Glycosylation/drug effects
Hexosamines/biosynthesis
Humans
Intracellular Signaling Peptides and Proteins/*metabolism
Liver Neoplasms/*metabolism/*pathology
Mice, Nude
Models, Biological
Phenotype
Protein Binding/drug effects
Protein Stability/drug effects
Soluble Guanylyl Cyclase/metabolism
*Diabetes
*Gucy1a3
*Hexosamine biosynthesis pathway (HBP)
*High glucose
*Protein stability
Abstract: TRIB2 has been identified as an onco-protein, and O-GlcNAcylation of target proteins has been reported to stimulate transformative phenotypes in liver cancer cells. However, the relationships between TRIB2 and O-GlcNAcylation are still unknown. The aim of this study was to investigate whether and how O-GlcNAcylation and TRIB2 regulate each other. We found that stimulation of O-GlcNAcylation elevates TRIB2 by enhancing its protein stability. TRIB2 can be O-GlcNAcylated by the hexosamine biosynthesis pathway (HBP). Also, O-GlcNAcylation boosting of transformative phenotypes of liver cancer cells might occur in a TRIB2-dependent manner. Interestingly, TRIB2 stimulated the metabolism of HBP, demonstrating that TRIB2 has positive feedback on O-GlcNAcylation. Notably, TRIB2 was found to maintain the stability of guanylate cyclase 1 alpha 3 (GUCY1A3), a key component of HBP, by interacting GUCY1A3 and reducing its ubiquitination. Importantly, TRIB2-dependent regulation of metabolism, transformative phenotypes, and O-GlcNAcylation all rely on GUCY1A3. Mouse experiments demonstrate that O-GlcNAcylation of TRIB2 is much higher in the livers of diabetic mice compared to control mice, suggesting that O-GlcNAcylation of TRIB2 might be critical for diabetes-associated liver cancer. Collectively, we have uncovered a positive auto-regulatory feedback between O-GlcNAcylation and TRIB2, which might be regarded as a promising therapeutic target for liver cancer.
Notes: 1873-3913
Yao, Bingjie
Xu, Yanli
Wang, Jiayi
Qiao, Yongxia
Zhang, Yue
Zhang, Xiao
Chen, Yan
Wu, Qi
Zhao, Yinghui
Zhu, Guoqing
Sun, Fenyong
Li, Zhi
Yuan, Hong
Journal Article
England
Cell Signal. 2016 Nov;28(11):1703-12. doi: 10.1016/j.cellsig.2016.08.003. Epub 2016 Aug 8.
Author Address: Department of Clinical Laboratory, the First Hospital affiliated to Dalian Medical University, Dalian, Liaoning Province 116011, China.
Department of Pharmacy, Dongjing Town Community Health Service Center, Songjiang District, Shanghai 201619, China.
Department of Clinical Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China; Tongji University Advanced Institute of Translational Medicine, Shanghai 200092, China. Electronic address: karajan2@163.com.
School of Public Health, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
Department of Laboratory Centre, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
Department of Clinical Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
Department of Clinical Laboratory, Shanghai Yangpu Hospital of Tongji University, Shanghai 200090, China. Electronic address: lizhsh2001@163.com.
Department of Clinical Laboratory, the First Hospital affiliated to Dalian Medical University, Dalian, Liaoning Province 116011, China. Electronic address: yuanhonglab@163.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1786
Author: Yao, G., Zhang, S., Mahrhold, S., Lam, K. H., Stern, D., Bagramyan, K., Perry, K., Kalkum, M., Rummel, A., Dong, M. and Jin, R.
Year: 2016
Title: N-linked glycosylation of SV2 is required for binding and uptake of botulinum neurotoxin A
Journal: Nat Struct Mol Biol
Volume: 23
Issue: 7
Pages: 656-62
Epub Date: 2016/06/14
Date: Jul
Short Title: N-linked glycosylation of SV2 is required for binding and uptake of botulinum neurotoxin A
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb.3245
PMCID: PMC5033645
NIHMSID: NIHMS787259
Accession Number: 27294781
Keywords: Amino Acid Sequence
Antibodies, Monoclonal/*chemistry
Antidotes/*chemistry
Binding Sites
Biological Transport
Botulinum Toxins, Type A/*chemistry/metabolism
Cloning, Molecular
Clostridium botulinum/*chemistry/pathogenicity
Crystallography, X-Ray
Escherichia coli/genetics/metabolism
Gene Expression
Glycosylation
HEK293 Cells
Humans
Membrane Glycoproteins/antagonists & inhibitors/chemistry/genetics/*metabolism
Models, Molecular
Nerve Tissue Proteins/antagonists & inhibitors/chemistry/genetics/*metabolism
Protein Binding
Protein Domains
*Protein Processing, Post-Translational
Protein Structure, Secondary
Recombinant Fusion Proteins/chemistry/genetics/metabolism
Abstract: Botulinum neurotoxin serotype A1 (BoNT/A1), a licensed drug widely used for medical and cosmetic applications, exerts its action by invading motoneurons. Here we report a 2.0-Å-resolution crystal structure of the BoNT/A1 receptor-binding domain in complex with its neuronal receptor, glycosylated human SV2C. We found that the neuronal tropism of BoNT/A1 requires recognition of both the peptide moiety and an N-linked glycan on SV2. This N-glycan-which is conserved in all SV2 isoforms across vertebrates-is essential for BoNT/A1 binding to neurons and for its potent neurotoxicity. The glycan-binding interface on SV2 is targeted by a human BoNT/A1-neutralizing antibody currently licensed as an antibotulism drug. Our studies reveal a new paradigm of host-pathogen interactions, in which pathogens exploit conserved host post-translational modifications, thereby achieving highly specific receptor binding while also tolerating genetic changes across multiple isoforms of receptors.
Notes: 1545-9985
Yao, Guorui
Zhang, Sicai
Mahrhold, Stefan
Lam, Kwok-Ho
Stern, Daniel
Orcid: 0000-0001-9057-4283
Bagramyan, Karine
Perry, Kay
Orcid: 0000-0002-4046-1704
Kalkum, Markus
Rummel, Andreas
Dong, Min
Jin, Rongsheng
Orcid: 0000-0003-0348-7363
P41 GM103403/GM/NIGMS NIH HHS/United States
R01 NS080833/NS/NINDS NIH HHS/United States
P30 DK034854/DK/NIDDK NIH HHS/United States
R21 AI123920/AI/NIAID NIH HHS/United States
R01 AI096169/AI/NIAID NIH HHS/United States
R01 AI091823/AI/NIAID NIH HHS/United States
S10 RR029205/RR/NCRR NIH HHS/United States
Journal Article
Nat Struct Mol Biol. 2016 Jul;23(7):656-62. doi: 10.1038/nsmb.3245. Epub 2016 Jun 13.
Author Address: Department of Physiology and Biophysics, University of California, Irvine, Irvine, California, USA.
Department of Urology, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts, USA.
Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, USA.
Department of Surgery, Harvard Medical School, Boston, Massachusetts, USA.
Institut für Toxikologie, Medizinische Hochschule Hannover, Hannover, Germany.
Centre for Biological Threats and Special Pathogens-Biological Toxins (ZBS3), Robert Koch-Institut, Berlin, Germany.
Department of Molecular Immunology, Beckman Research Institute of City of Hope, Duarte, California, USA.
NE-CAT, Argonne National Laboratory, Department of Chemistry and Chemical Biology, Cornell University, Argonne, Illinois, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1401
Author: Yao, G., Zong, Y., Gu, S., Zhou, J., Xu, H., Mathews, II and Jin, R.
Year: 2011
Title: Crystal structure of the glutamate receptor GluA1 N-terminal domain
Journal: Biochem J
Volume: 438
Issue: 2
Pages: 255-63
Epub Date: 2011/06/07
Date: Sep 1
Short Title: Crystal structure of the glutamate receptor GluA1 N-terminal domain
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20110801
PMCID: PMC3296483
NIHMSID: NIHMS358187
Accession Number: 21639859
Keywords: Animals
Crystallography, X-Ray
Models, Molecular
Pliability
Protein Binding
Protein Multimerization
Protein Structure, Secondary
Protein Structure, Tertiary
Rats
Receptors, AMPA/*chemistry/isolation & purification
Abstract: The AMPA (α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid) subfamily of iGluRs (ionotropic glutamate receptors) is essential for fast excitatory neurotransmission in the central nervous system. The malfunction of AMPARs (AMPA receptors) has been implicated in many neurological diseases, including Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis. The active channels of AMPARs and other iGluR subfamilies are tetramers formed exclusively by assembly of subunits within the same subfamily. It has been proposed that the assembly process is controlled mainly by the extracellular ATD (N-terminal domain) of iGluR. In addition, ATD has also been implicated in synaptogenesis, iGluR trafficking and trans-synaptic signalling, through unknown mechanisms. We report in the present study a 2.5 Å (1 Å=0.1 nm) resolution crystal structure of the ATD of GluA1. Comparative analyses of the structure of GluA1-ATD and other subunits sheds light on our understanding of how ATD drives subfamily-specific assembly of AMPARs. In addition, analysis of the crystal lattice of GluA1-ATD suggests a novel mechanism by which the ATD might participate in inter-tetramer AMPAR clustering, as well as in trans-synaptic protein-protein interactions.
Notes: 1470-8728
Yao, Guorui
Zong, Yinong
Gu, Shenyan
Zhou, Jie
Xu, Huaxi
Mathews, Irimpan I
Jin, Rongsheng
R01GM090023/GM/NIGMS NIH HHS/United States
R01 AG038710/AG/NIA NIH HHS/United States
R01 AG021173/AG/NIA NIH HHS/United States
R01 GM090023/GM/NIGMS NIH HHS/United States
R21 AG033813/AG/NIA NIH HHS/United States
R21 AG033813-02/AG/NIA NIH HHS/United States
R01 GM090023-01/GM/NIGMS NIH HHS/United States
R21AG033813/AG/NIA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Biochem J. 2011 Sep 1;438(2):255-63. doi: 10.1042/BJ20110801.
Author Address: Fujian Provincial Key Laboratory of Neurodegenerative Disease and Aging Research, College of Medicine, Xiamen University, Fujian, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2077
Author: Yao, H., Song, Y., Chen, Y., Wu, N., Xu, J., Sun, C., Zhang, J., Weng, T., Zhang, Z., Wu, Z., Cheng, L., Shi, D., Lu, X., Lei, J., Crispin, M., Shi, Y., Li, L. and Li, S.
Year: 2020
Title: Molecular Architecture of the SARS-CoV-2 Virus
Journal: Cell
Volume: 183
Issue: 3
Pages: 730-738.e13
Epub Date: 2020/09/28
Date: Oct 29
Short Title: Molecular Architecture of the SARS-CoV-2 Virus
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2020.09.018
PMCID: PMC7474903
Accession Number: 32979942
Keywords: Animals
Betacoronavirus/*physiology/*ultrastructure
Chlorocebus aethiops
Cryoelectron Microscopy
Humans
Mass Spectrometry
Models, Molecular
Protein Conformation
SARS-CoV-2
Vero Cells
Viral Proteins/chemistry/ultrastructure
*Virus Assembly
Virus Cultivation
*SARS-CoV-2
*coronavirus
*cryo-EM
*cryo-electron tomography
*ribonucleoprotein
*spike glycoprotein
*subtomogram averaging
*virus structure
Abstract: SARS-CoV-2 is an enveloped virus responsible for the COVID-19 pandemic. Despite recent advances in the structural elucidation of SARS-CoV-2 proteins, the detailed architecture of the intact virus remains to be unveiled. Here we report the molecular assembly of the authentic SARS-CoV-2 virus using cryoelectron tomography (cryo-ET) and subtomogram averaging (STA). Native structures of the S proteins in pre- and postfusion conformations were determined to average resolutions of 8.7-11 Å. Compositions of the N-linked glycans from the native spikes were analyzed by mass spectrometry, which revealed overall processing states of the native glycans highly similar to that of the recombinant glycoprotein glycans. The native conformation of the ribonucleoproteins (RNPs) and their higher-order assemblies were revealed. Overall, these characterizations revealed the architecture of the SARS-CoV-2 virus in exceptional detail and shed light on how the virus packs its ∼30-kb-long single-segmented RNA in the ∼80-nm-diameter lumen.
Notes: 1097-4172
Yao, Hangping
Song, Yutong
Chen, Yong
Wu, Nanping
Xu, Jialu
Sun, Chujie
Zhang, Jiaxing
Weng, Tianhao
Zhang, Zheyuan
Wu, Zhigang
Cheng, Linfang
Shi, Danrong
Lu, Xiangyun
Lei, Jianlin
Crispin, Max
Shi, Yigong
Li, Lanjuan
Li, Sai
Journal Article
Research Support, Non-U.S. Gov't
Cell. 2020 Oct 29;183(3):730-738.e13. doi: 10.1016/j.cell.2020.09.018. Epub 2020 Sep 6.
Author Address: State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310003, Zhejiang Province, China; National Clinical Research Center for Infectious Diseases, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310003, Zhejiang Province, China.
School of Life Sciences, Tsinghua University, Beijing 100084, China; Beijing Advanced Innovation Center for Structural Biology & Frontier Research Center for Biological Structure, Beijing 100084, China.
School of Life Sciences, Tsinghua University, Beijing 100084, China; Beijing Advanced Innovation Center for Structural Biology & Frontier Research Center for Biological Structure, Beijing 100084, China; Tsinghua-Peking Center for Life Sciences, Beijing 100084, China.
School of Biological Sciences, University of Southampton, Southampton SO17 1BJ, UK.
School of Life Sciences, Tsinghua University, Beijing 100084, China; Beijing Advanced Innovation Center for Structural Biology & Frontier Research Center for Biological Structure, Beijing 100084, China; Tsinghua-Peking Center for Life Sciences, Beijing 100084, China; Key Laboratory of Structural Biology of Zhejiang Province, School of Life Sciences, Westlake University, 18 Shilongshan Road, Hangzhou 310024, Zhejiang Province, China; Institute of Biology, Westlake Institute for Advanced Study, 18 Shilongshan Road, Hangzhou 310024, Zhejiang Province, China.
State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310003, Zhejiang Province, China; National Clinical Research Center for Infectious Diseases, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310003, Zhejiang Province, China. Electronic address: ljli@zju.edu.cn.
School of Life Sciences, Tsinghua University, Beijing 100084, China; Beijing Advanced Innovation Center for Structural Biology & Frontier Research Center for Biological Structure, Beijing 100084, China; Tsinghua-Peking Center for Life Sciences, Beijing 100084, China. Electronic address: sai@tsinghua.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1536
Author: Yao, J., Hong, W., Huang, J., Zhan, K., Huang, H. and Hong, M.
Year: 2012
Title: N-Glycosylation dictates proper processing of organic anion transporting polypeptide 1B1
Journal: PLoS One
Volume: 7
Issue: 12
Pages: e52563
Epub Date: 2013/01/04
Short Title: N-Glycosylation dictates proper processing of organic anion transporting polypeptide 1B1
Alternate Journal: PloS one
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0052563
PMCID: PMC3527552
Accession Number: 23285087
Keywords: Amino Acid Sequence
Endoplasmic Reticulum/metabolism
Fluorescent Antibody Technique
Glycosylation
HEK293 Cells
Humans
Liver-Specific Organic Anion Transporter 1
Molecular Sequence Data
Mutant Proteins/metabolism
Mutation/genetics
Organic Anion Transporters/chemistry/*metabolism
*Protein Processing, Post-Translational
Protein Structure, Secondary
Abstract: Organic anion transporting polypeptides (OATPs) have been extensively recognized as key determinants of absorption, distribution, metabolism and excretion (ADME) of various drugs, xenobiotics and toxins. Putative N-glycosylation sites located in the extracellular loops 2 and 5 is considered a common feature of all OATPs and some members have been demonstrated to be glycosylated proteins. However, experimental evidence is still lacking on how such a post-translational modification affect the transport activity of OATPs and which of the putative glycosylation sites are utilized in these transporter proteins. In the present study, we substituted asparagine residues that are possibly involved in N-glycosylation with glutamine residues and identified three glycosylation sites (Asn134, Asn503 and Asn516) within the structure of OATP1B1, an OATP member that is mainly expressed in the human liver. Our results showed that Asn134 and Asn516 are used for glycosylation under normal conditions; however, when Asn134 was mutagenized, an additional asparagine at position 503 is involved in the glycosylation process. Simultaneously replacement of all three asparagines with glutamines led to significantly reduced protein level as well as loss of transport activity. Further studies revealed that glycosylation affected stability of the transporter protein and the unglycosylated mutant was retained within endoplasmic reticulum.
Notes: 1932-6203
Yao, Juan
Hong, Weifang
Huang, Jiujiu
Zhan, Kai
Huang, Hong
Hong, Mei
Journal Article
Research Support, Non-U.S. Gov't
PLoS One. 2012;7(12):e52563. doi: 10.1371/journal.pone.0052563. Epub 2012 Dec 20.
Author Address: College of Life Science, South China Agricultural University, Guangzhou, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 664
Author: Yao, P. J. and Coleman, P. D.
Year: 1998
Title: Reduction of O-linked N-acetylglucosamine-modified assembly protein-3 in Alzheimer's disease
Journal: J Neurosci
Volume: 18
Issue: 7
Pages: 2399-411
Epub Date: 1998/04/16
Date: Apr 1
Short Title: Reduction of O-linked N-acetylglucosamine-modified assembly protein-3 in Alzheimer's disease
Alternate Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience
ISSN: 0270-6474 (Print)
0270-6474
DOI: 10.1523/jneurosci.18-07-02399.1998
PMCID: PMC6793091
Accession Number: 9502801
Keywords: Acetylation
Acetylglucosamine/chemistry/*metabolism
Adaptor Proteins, Vesicular Transport
Aged
Aged, 80 and over
Alzheimer Disease/*metabolism/pathology
Cerebellum/chemistry/enzymology
Enzyme Inhibitors/*metabolism
Female
Frontal Lobe/chemistry/enzymology
Galactose
Galactosyltransferases/analysis
Humans
Male
Middle Aged
Molecular Weight
*Monomeric Clathrin Assembly Proteins
Nerve Tissue Proteins/*metabolism
Neurofibrillary Tangles/metabolism
Oligosaccharides/analysis/metabolism
Ovalbumin/analysis
Phosphoproteins/*metabolism
Phosphorylation
Tritium
Abstract: Abnormal protein processing and modification is associated with Alzheimer's disease (AD) pathology. The role of phosphorylation in AD has been studied extensively because the presumed abnormal phosphorylation of tau protein is believed to play a role in the formation of paired helical filaments. Glycosylation with O-linked N-acetylglucosamine (O-GlcNAc) to serine and threonine residues is a dynamic protein modification of intracellular proteins, and it shares similar features with protein phosphorylation. In this study, O-GlcNAc glycosylation of proteins from autopsied human brains with confirmed AD and non-AD age-matched controls was examined. O-GlcNAcylation was demonstrated by labeling protein extracts with [3H]galactose in the presence of galactosyltransferase and subsequent analyses of saccharide-protein linkage and saccharide structure. The number of O-GlcNAc-containing proteins and the overall O-GlcNAc level do not appear to be different between AD and control brain tissues. The only significant change observed is a marked reduction of O-GlcNAcylated clathrin assembly protein-3 (AP-3) in AD. The reduction is more evident in brain neocortical regions, and there appears to be a negative correlation between O-glycosylated AP-3 and the density of neurofibrillary tangles. These data suggest a possible association between the O-glycosylated AP-3 and AD pathology.
Notes: 1529-2401
Yao, P J
Coleman, P D
AG-08665/AG/NIA NIH HHS/United States
AG-09016/AG/NIA NIH HHS/United States
R01 AG-01121/AG/NIA NIH HHS/United States
etc.
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
J Neurosci. 1998 Apr 1;18(7):2399-411. doi: 10.1523/JNEUROSCI.18-07-02399.1998.
Author Address: Department of Neurobiology and Anatomy, University of Rochester Medical Center, Rochester, New York 14642, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 688
Author: Yao, P. J. and Coleman, P. D.
Year: 1998
Title: Reduced O-glycosylated clathrin assembly protein AP180: implication for synaptic vesicle recycling dysfunction in Alzheimer's disease
Journal: Neurosci Lett
Volume: 252
Issue: 1
Pages: 33-6
Epub Date: 1998/10/02
Date: Aug 7
Short Title: Reduced O-glycosylated clathrin assembly protein AP180: implication for synaptic vesicle recycling dysfunction in Alzheimer's disease
Alternate Journal: Neuroscience letters
ISSN: 0304-3940 (Print)
0304-3940
DOI: 10.1016/s0304-3940(98)00547-3
Accession Number: 9756352
Keywords: Acetylglucosamine/analysis/metabolism
Adaptor Proteins, Vesicular Transport
Alzheimer Disease/*metabolism/pathology
Clathrin/*metabolism
Enzyme Inhibitors/*metabolism
Galactose/pharmacokinetics
Glycosylation
Humans
*Monomeric Clathrin Assembly Proteins
Nerve Tissue Proteins/isolation & purification/*metabolism
Neurofibrillary Tangles/chemistry/metabolism
Neurofilament Proteins/analysis
Phosphoproteins/isolation & purification/*metabolism
Precipitin Tests
Synapses/chemistry/metabolism
Synaptic Vesicles/chemistry/*metabolism
Synaptophysin/analysis
Tritium
Abstract: Synapse loss is one of the neuropathologies in Alzheimer's disease (AD) that may play a crucial role in the mechanism of its distinct cognitive impairment and dementia. In a previous study [18], a significant reduction of O-glycosylated clathrin assembly protein AP180 was observed in neocortex of AD. The reduction correlated with the density of neurofibrillary tangles. In this study we further determine that the O-GlcNAc/AP180 ratio is not changed, but the level of AP180 protein decreases in AD. Furthermore, whereas the level of neurofilament (NF-M) remains relatively unchanged, another clathrin assembly protein, AP-2, is also reduced in AD along with a small loss of synaptophysin. Our findings suggest that synaptic vesicle recycling dysfunction may be involved in the pathology of synapse loss in AD.
Notes: Yao, P J
Coleman, P D
AG-08665/AG/NIA NIH HHS/United States
AG-09016/AG/NIA NIH HHS/United States
AG01121/AG/NIA NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Ireland
Neurosci Lett. 1998 Aug 7;252(1):33-6. doi: 10.1016/s0304-3940(98)00547-3.
Author Address: Department of Neurobiology and Anatomy, University of Rochester Medical Center, NY 14642, USA. pamela_yao@urmc.rochester.edu
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 330
Author: Yasuda, T., Awazu, S., Sato, W., Iida, R., Tanaka, Y. and Kishi, K.
Year: 1990
Title: Human genetically polymorphic deoxyribonuclease: purification, characterization, and multiplicity of urine deoxyribonuclease I
Journal: J Biochem
Volume: 108
Issue: 3
Pages: 393-8
Epub Date: 1990/09/01
Date: Sep
Short Title: Human genetically polymorphic deoxyribonuclease: purification, characterization, and multiplicity of urine deoxyribonuclease I
Alternate Journal: Journal of biochemistry
ISSN: 0021-924X (Print)
0021-924x
DOI: 10.1093/oxfordjournals.jbchem.a123212
Accession Number: 2277032
Keywords: Amino Acid Sequence
Antibodies/immunology
Carbohydrates/chemistry
Deoxyribonuclease I/chemistry/genetics/immunology/*urine
Electrophoresis, Polyacrylamide Gel
Humans
Isoenzymes/chemistry/genetics/immunology/*urine
Male
Middle Aged
Molecular Sequence Data
*Polymorphism, Genetic
Substrate Specificity
Abstract: A deoxyribonuclease I was purified from the urine of a 46-year-old male (a single individual) by using a series of column chromatographies to a homogeneous state as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme was found to be a glycoprotein, containing 1 fucose, 7 galactose, 10 mannose, 6 glucosamine, and 2 sialic acid residues per molecule. The N-terminal amino acid sequence up to the 27th residue of the enzyme was similar to that of pancreatic deoxyribonuclease I from bovine and other species. The catalytic properties of the enzyme derived from a single individual closely resembled those of deoxyribonuclease I purified from human urine collected from several volunteers [Ito, K. et al. (1984) J. Biochem. 95, 1399-1406]. The purified enzyme was found to consist of multiple forms with different pI values. These findings are compatible with the existence of genetic polymorphism of deoxyribonuclease I in human urine previously reported [Kishi, K. et al. (1989) Hum. Genet. 81, 295-297]. This multiplicity of the urine enzyme might be due to variations in the primary structure and/or differences in the content of sialic acid.
Notes: Yasuda, T
Awazu, S
Sato, W
Iida, R
Tanaka, Y
Kishi, K
Journal Article
Research Support, Non-U.S. Gov't
England
J Biochem. 1990 Sep;108(3):393-8. doi: 10.1093/oxfordjournals.jbchem.a123212.
Author Address: Department of Legal Medicine, Fukui Medical School.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1110
Author: Yasui, N., Nogi, T., Kitao, T., Nakano, Y., Hattori, M. and Takagi, J.
Year: 2007
Title: Structure of a receptor-binding fragment of reelin and mutational analysis reveal a recognition mechanism similar to endocytic receptors
Journal: Proc Natl Acad Sci U S A
Volume: 104
Issue: 24
Pages: 9988-93
Epub Date: 2007/06/06
Date: Jun 12
Short Title: Structure of a receptor-binding fragment of reelin and mutational analysis reveal a recognition mechanism similar to endocytic receptors
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.0700438104
PMCID: PMC1891246
Accession Number: 17548821
Keywords: Adaptor Proteins, Signal Transducing/metabolism
Amino Acid Sequence
Animals
Binding Sites
CHO Cells
Cell Adhesion Molecules, Neuronal/*chemistry
Cell Line
Cells, Cultured
Cerebral Cortex/cytology
Clone Cells
Conserved Sequence
Cricetinae
Cricetulus
Crystallography, X-Ray
DNA Mutational Analysis
Disulfides/chemistry
Electroporation
Extracellular Matrix Proteins/*chemistry
Glutamic Acid/chemistry
Histidine/chemistry
Humans
LDL-Receptor Related Proteins
Ligands
Mice
Models, Chemical
Models, Molecular
Molecular Sequence Data
Mutagenesis
Nerve Tissue Proteins/*chemistry/metabolism
Neurons/cytology/metabolism
Peptide Fragments/*chemistry/*genetics/isolation & purification/metabolism
Phosphorylation
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Lipoprotein/*metabolism
Recombinant Fusion Proteins/chemistry/metabolism
Serine Endopeptidases/*chemistry
Zinc/metabolism
Abstract: Reelin, a large secreted protein implicated in the cortical development of the mammalian brain, is composed of eight tandem concatenations of "reelin repeats" and binds to neuronal receptors belonging to the low-density lipoprotein receptor gene family. We found that both receptor-binding and subsequent Dab1 phosphorylation occur solely in the segment spanning the fifth and sixth reelin repeats (R5-6). Monomeric fragment exhibited a suboptimal level of signaling activity and artificial oligomerization resulted in a 10-fold increase in activity, indicating the critical importance of higher-order multimerization in physiological reelin. A 2.0-A crystal structure from the R5-6 fragment revealed not only a unique domain arrangement wherein two repeats were aligned side by side with the same orientation, but also the unexpected presence of bound Zn ions. Structure-guided alanine mutagenesis of R5-6 revealed that two Lys residues (Lys-2360 and Lys-2467) constitute a central binding site for the low-density lipoprotein receptor class A module in the receptor, indicating a strong similarity to the ligand recognition mode shared among the endocytic lipoprotein receptors.
Notes: 1091-6490
Yasui, Norihisa
Nogi, Terukazu
Kitao, Tomoe
Nakano, Yoshimi
Hattori, Mitsuharu
Takagi, Junichi
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2007 Jun 12;104(24):9988-93. doi: 10.1073/pnas.0700438104. Epub 2007 Jun 4.
Author Address: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 647
Author: Yeung, B., Porter, T. J. and Vath, J. E.
Year: 1997
Title: Direct isoform analysis of high-mannose-containing glycoproteins by on-line capillary electrophoresis electrospray mass spectrometry
Journal: Anal Chem
Volume: 69
Issue: 13
Pages: 2510-6
Epub Date: 1997/07/01
Date: Jul 1
Short Title: Direct isoform analysis of high-mannose-containing glycoproteins by on-line capillary electrophoresis electrospray mass spectrometry
Alternate Journal: Analytical chemistry
ISSN: 0003-2700 (Print)
0003-2700
DOI: 10.1021/ac9611172
Accession Number: 9265423
Keywords: Amino Acid Sequence
Bone Morphogenetic Protein 2
Bone Morphogenetic Proteins/analysis/chemistry
Chromatography, Ion Exchange
Electrophoresis, Capillary/*methods
Glycoproteins/*analysis/chemistry
Humans
Mannose/*analysis
Mass Spectrometry/*methods
Molecular Conformation
Molecular Sequence Data
Molecular Weight
Recombinant Proteins/analysis/chemistry
Ribonucleases/analysis/chemistry
Signal Processing, Computer-Assisted
*Transforming Growth Factor beta
Notes: Yeung, B
Porter, T J
Vath, J E
Journal Article
United States
Anal Chem. 1997 Jul 1;69(13):2510-6. doi: 10.1021/ac9611172.
Author Address: Biopharmaceutical Characterization and Analysis, Genetics Institute, Inc., Andover, Massacheusetts 01810.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1509
Author: Yi, W., Clark, P. M., Mason, D. E., Keenan, M. C., Hill, C., Goddard, W. A., 3rd, Peters, E. C., Driggers, E. M. and Hsieh-Wilson, L. C.
Year: 2012
Title: Phosphofructokinase 1 glycosylation regulates cell growth and metabolism
Journal: Science
Volume: 337
Issue: 6097
Pages: 975-80
Epub Date: 2012/08/28
Date: Aug 24
Short Title: Phosphofructokinase 1 glycosylation regulates cell growth and metabolism
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1222278
PMCID: PMC3534962
NIHMSID: NIHMS427904
Accession Number: 22923583
Keywords: Acetylglucosamine/metabolism
Acylation
Adenosine Triphosphate/metabolism
Animals
Cell Hypoxia
Cell Line
Cell Line, Tumor
*Cell Proliferation
Glucose/*metabolism
Glycolysis
Glycosylation
Humans
Lactic Acid/metabolism
Mice
Mice, Nude
N-Acetylglucosaminyltransferases/genetics/metabolism
NADP/metabolism
Neoplasms/*metabolism/*pathology
Pentose Phosphate Pathway
Phosphofructokinase-1, Liver Type/antagonists & inhibitors/chemistry/*metabolism
Abstract: Cancer cells must satisfy the metabolic demands of rapid cell growth within a continually changing microenvironment. We demonstrated that the dynamic posttranslational modification of proteins by O-linked β-N-acetylglucosamine (O-GlcNAcylation) is a key metabolic regulator of glucose metabolism. O-GlcNAcylation was induced at serine 529 of phosphofructokinase 1 (PFK1) in response to hypoxia. Glycosylation inhibited PFK1 activity and redirected glucose flux through the pentose phosphate pathway, thereby conferring a selective growth advantage on cancer cells. Blocking glycosylation of PFK1 at serine 529 reduced cancer cell proliferation in vitro and impaired tumor formation in vivo. These studies reveal a previously uncharacterized mechanism for the regulation of metabolic pathways in cancer and a possible target for therapeutic intervention.
Notes: 1095-9203
Yi, Wen
Clark, Peter M
Mason, Daniel E
Keenan, Marie C
Hill, Collin
Goddard, William A 3rd
Peters, Eric C
Driggers, Edward M
Hsieh-Wilson, Linda C
R01 GM084724/GM/NIGMS NIH HHS/United States
Howard Hughes Medical Institute/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Science. 2012 Aug 24;337(6097):975-80. doi: 10.1126/science.1222278.
Author Address: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1542
Author: Yin, X., Bern, M., Xing, Q., Ho, J., Viner, R. and Mayr, M.
Year: 2013
Title: Glycoproteomic analysis of the secretome of human endothelial cells
Journal: Mol Cell Proteomics
Volume: 12
Issue: 4
Pages: 956-78
Epub Date: 2013/01/25
Date: Apr
Short Title: Glycoproteomic analysis of the secretome of human endothelial cells
Alternate Journal: Molecular & cellular proteomics : MCP
ISSN: 1535-9476 (Print)
1535-9476
DOI: 10.1074/mcp.M112.024018
PMCID: PMC3617342
Accession Number: 23345538
Keywords: Amino Acid Sequence
Carbohydrate Conformation
Carbohydrate Sequence
Cells, Cultured
Culture Media, Conditioned
Glycoproteins/chemistry/*metabolism
Glycosylation
Human Umbilical Vein Endothelial Cells/*metabolism
Humans
Molecular Sequence Data
Peptide Fragments/chemistry
*Protein Processing, Post-Translational
Proteome/chemistry/*metabolism
Tetradecanoylphorbol Acetate/pharmacology
Abstract: Previous proteomics studies have partially unraveled the complexity of endothelial protein secretion but have not investigated glycosylation, a key modification of secreted and membrane proteins for cell communication. In this study, human umbilical vein endothelial cells were kept in serum-free medium before activation by phorbol-12-myristate-13 acetate, a commonly used secretagogue that induces exocytosis of endothelial vesicles. In addition to 123 secreted proteins, the secretome was particularly rich in membrane proteins. Glycopeptides were enriched by zwitterionic hydrophilic interaction liquid chromatography resins and were either treated with PNGase F and H2(18)O or directly analyzed using a recently developed workflow combining higher-energy C-trap dissociation (HCD) with electron-transfer dissociation (ETD) for a hybrid linear ion trap-orbitrap mass spectrometer. After deglycosylation with PNGase F in the presence of H2(18)O, 123 unique peptides displayed (18)O-deamidation of asparagine, corresponding to 86 proteins with a total of 121 glycosylation sites. Direct glycopeptide analysis via HCD-ETD identified 131 glycopeptides from 59 proteins and 118 glycosylation sites, of which 41 were known, 51 were predicted, and 26 were novel. Two methods were compared: alternating HCD-ETD and HCD-product-dependent ETD. The former detected predominantly high-intensity, multiply charged glycopeptides, whereas the latter preferentially selected precursors with complex/hybrid glycans for fragmentation. Validation was performed by means of glycoprotein enrichment and analysis of the input, the flow-through, and the bound fraction. This study represents the most comprehensive characterization of endothelial protein secretion to date and demonstrates the potential of new HCD-ETD workflows for determining the glycosylation status of complex biological samples.
Notes: 1535-9484
Yin, Xiaoke
Bern, Marshall
Xing, Qiuru
Ho, Jenny
Viner, Rosa
Mayr, Manuel
FS/13/2/29892/British Heart Foundation/United Kingdom
Department of Health/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
Mol Cell Proteomics. 2013 Apr;12(4):956-78. doi: 10.1074/mcp.M112.024018. Epub 2013 Jan 23.
Author Address: The King's British Heart Foundation Centre, King's College London, London SE5 9NU, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 761
Author: Yoon, C., Johnston, S. C., Tang, J., Stahl, M., Tobin, J. F. and Somers, W. S.
Year: 2000
Title: Charged residues dominate a unique interlocking topography in the heterodimeric cytokine interleukin-12
Journal: Embo j
Volume: 19
Issue: 14
Pages: 3530-41
Epub Date: 2000/07/19
Date: Jul 17
Short Title: Charged residues dominate a unique interlocking topography in the heterodimeric cytokine interleukin-12
Alternate Journal: The EMBO journal
ISSN: 0261-4189 (Print)
0261-4189
DOI: 10.1093/emboj/19.14.3530
PMCID: PMC313992
Accession Number: 10899108
Keywords: Arginine/genetics/metabolism
Binding Sites
Crystallography, X-Ray
Dimerization
Disulfides/chemistry/metabolism
Drug Design
Epitopes/chemistry/metabolism
Growth Hormone/chemistry/metabolism
Humans
Interleukin-12/antagonists & inhibitors/*chemistry/genetics/metabolism
Models, Molecular
Molecular Weight
Mutagenesis, Site-Directed
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, Cytokine/antagonists & inhibitors/chemistry/genetics/metabolism
Receptors, Somatotropin/chemistry/metabolism
Signal Transduction
Static Electricity
Structure-Activity Relationship
Abstract: Human interleukin-12 (IL-12, p70) is an early pro-inflammatory cytokine, comprising two disulfide-linked subunits, p35 and p40. We solved the crystal structures of monomeric human p40 at 2.5 A and the human p70 complex at 2.8 A resolution, which reveals that IL-12 is similar to class 1 cytokine-receptor complexes. They also include the first description of an N-terminal immunoglobulin-like domain, found on the p40 subunit. Several charged residues from p35 and p40 intercalate to form a unique interlocking topography, shown by mutagenesis to be critical for p70 formation. A central arginine residue from p35 projects into a deep pocket on p40, which may be an ideal target for a small molecule antagonist of IL-12 formation.
Notes: 1460-2075
Yoon, C
Johnston, S C
Tang, J
Stahl, M
Tobin, J F
Somers, W S
Journal Article
EMBO J. 2000 Jul 17;19(14):3530-41. doi: 10.1093/emboj/19.14.3530.
Author Address: Departments of Musculoskeletal Sciences and Biological Chemistry, Wyeth Research, 87 Cambridge Park Drive, Cambridge, MA 02140, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1291
Author: Yoshida-Moriguchi, T., Yu, L., Stalnaker, S. H., Davis, S., Kunz, S., Madson, M., Oldstone, M. B., Schachter, H., Wells, L. and Campbell, K. P.
Year: 2010
Title: O-mannosyl phosphorylation of alpha-dystroglycan is required for laminin binding
Journal: Science
Volume: 327
Issue: 5961
Pages: 88-92
Epub Date: 2010/01/02
Date: Jan 1
Short Title: O-mannosyl phosphorylation of alpha-dystroglycan is required for laminin binding
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1180512
PMCID: PMC2978000
NIHMSID: NIHMS248430
Accession Number: 20044576
Keywords: Animals
Carbohydrate Conformation
Cell Line
Dystroglycans/chemistry/*metabolism
Glycosylation
Humans
Laminin/*metabolism
Magnetic Resonance Spectroscopy
Mannose/*metabolism
Mass Spectrometry
Membrane Proteins/metabolism
Mice
Mice, Inbred C57BL
Muscle, Skeletal/metabolism
Muscular Dystrophies/metabolism
Muscular Dystrophy, Animal/metabolism
N-Acetylglucosaminyltransferases/genetics/metabolism
Phosphorylation
Protein Binding
Recombinant Proteins/chemistry/metabolism
Abstract: Alpha-dystroglycan (alpha-DG) is a cell-surface glycoprotein that acts as a receptor for both extracellular matrix proteins containing laminin-G domains and certain arenaviruses. Receptor binding is thought to be mediated by a posttranslational modification, and defective binding with laminin underlies a subclass of congenital muscular dystrophy. Using mass spectrometry- and nuclear magnetic resonance (NMR)-based structural analyses, we identified a phosphorylated O-mannosyl glycan on the mucin-like domain of recombinant alpha-DG, which was required for laminin binding. We demonstrated that patients with muscle-eye-brain disease and Fukuyama congenital muscular dystrophy, as well as mice with myodystrophy, commonly have defects in a postphosphoryl modification of this phosphorylated O-linked mannose, and that this modification is mediated by the like-acetylglucosaminyltransferase (LARGE) protein. These findings expand our understanding of the mechanisms that underlie congenital muscular dystrophy.
Notes: 1095-9203
Yoshida-Moriguchi, Takako
Yu, Liping
Stalnaker, Stephanie H
Davis, Sarah
Kunz, Stefan
Madson, Michael
Oldstone, Michael B A
Schachter, Harry
Wells, Lance
Campbell, Kevin P
R01 AI009484-40/AI/NIAID NIH HHS/United States
R21 AI055540/AI/NIAID NIH HHS/United States
R01 AI045927/AI/NIAID NIH HHS/United States
U54 NS053672/NS/NINDS NIH HHS/United States
AI55540/AI/NIAID NIH HHS/United States
R01 AI055540/AI/NIAID NIH HHS/United States
P41 RR018502/RR/NCRR NIH HHS/United States
R01 AI045927-09/AI/NIAID NIH HHS/United States
R01 AI045927-10/AI/NIAID NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 AI009484/AI/NIAID NIH HHS/United States
R01 AI045927-08/AI/NIAID NIH HHS/United States
P30 DK054759/DK/NIDDK NIH HHS/United States
1U54NS053672/NS/NINDS NIH HHS/United States
P30 DK 54759/DK/NIDDK NIH HHS/United States
R01 AI009484-41/AI/NIAID NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2010 Jan 1;327(5961):88-92. doi: 10.1126/science.1180512.
Author Address: Howard Hughes Medical Institute, University of Iowa Roy J. and Lucille A. Carver College of Medicine, 4283 Carver Biomedical Research Building, 285 Newton Road, Iowa City, IA 52242-1101, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1168
Author: Yoshikawa, F., Sato, Y., Tohyama, K., Akagi, T., Hashikawa, T., Nagakura-Takagi, Y., Sekine, Y., Morita, N., Baba, H., Suzuki, Y., Sugano, S., Sato, A. and Furuichi, T.
Year: 2008
Title: Opalin, a transmembrane sialylglycoprotein located in the central nervous system myelin paranodal loop membrane
Journal: J Biol Chem
Volume: 283
Issue: 30
Pages: 20830-40
Epub Date: 2008/05/21
Date: Jul 25
Short Title: Opalin, a transmembrane sialylglycoprotein located in the central nervous system myelin paranodal loop membrane
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M801314200
PMCID: PMC3258930
Accession Number: 18490449
Keywords: Animals
Axons/*metabolism
COS Cells
Central Nervous System/*metabolism
Chlorocebus aethiops
Dogs
HeLa Cells
Humans
Mice
Mice, Inbred C57BL
Mice, Inbred ICR
Myelin Proteins/*chemistry
Myelin Sheath/*chemistry
Rabbits
Rats
Sialoglycoproteins/*chemistry
Abstract: In contrast to compact myelin, the series of paranodal loops located in the outermost lateral region of myelin is non-compact; the intracellular space is filled by a continuous channel of cytoplasm, the extracellular surfaces between neighboring loops keep a definite distance, but the loop membranes have junctional specializations. Although the proteins that form compact myelin have been well studied, the protein components of paranodal loop membranes are not fully understood. This report describes the biochemical characterization and expression of Opalin as a novel membrane protein in paranodal loops. Mouse Opalin is composed of a short N-terminal extracellular domain (amino acid residues 1-30), a transmembrane domain (residues 31-53), and a long C-terminal intracellular domain (residues 54-143). Opalin is enriched in myelin of the central nervous system, but not that of the peripheral nervous system of mice. Enzymatic deglycosylation showed that myelin Opalin contained N- and O-glycans, and that the O-glycans, at least, had negatively charged sialic acids. We identified two N-glycan sites at Asn-6 and Asn-12 and an O-glycan site at Thr-14 in the extracellular domain. Site-directed mutations at the glycan sites impaired the cell surface localization of Opalin. In addition to the somata and processes of oligodendrocytes, Opalin immunoreactivity was observed in myelinated axons in a spiral fashion, and was concentrated in the paranodal loop region. Immunogold electron microscopy demonstrated that Opalin was localized at particular sites in the paranodal loop membrane. These results suggest a role for highly sialylglycosylated Opalin in an intermembranous function of the myelin paranodal loops in the central nervous system.
Notes: 1083-351x
Yoshikawa, Fumio
Sato, Yumi
Tohyama, Koujiro
Akagi, Takumi
Hashikawa, Tsutomu
Nagakura-Takagi, Yuko
Sekine, Yukiko
Morita, Noriyuki
Baba, Hiroko
Suzuki, Yutaka
Sugano, Sumio
Sato, Akira
Furuichi, Teiichi
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2008 Jul 25;283(30):20830-40. doi: 10.1074/jbc.M801314200. Epub 2008 May 19.
Author Address: Laboratory for Molecular Neurogenesis and Laboratory for Neural Architecture, RIKEN Brain Science Institute, Wako, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 498
Author: Yoshima, H., Furthmayr, H. and Kobata, A.
Year: 1980
Title: Structures of the asparagine-linked sugar chains of glycophorin A
Journal: J Biol Chem
Volume: 255
Issue: 20
Pages: 9713-8
Epub Date: 1980/10/25
Date: Oct 25
Short Title: Structures of the asparagine-linked sugar chains of glycophorin A
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7430095
Keywords: Asparagine/*analysis
Carbohydrate Conformation
Carbohydrate Sequence
*Glycophorins
Glycoside Hydrolases
Humans
Protein Binding
*Sialoglycoproteins
Sugar Alcohols/analysis
Abstract: Glycophorin A isolated from human erythrocytes contains one asparagine-linked sugar chain in one molecule. This sugar chain was successfully released as an oligosaccharide fraction by hydrazinolysis. The oligosaccharide fraction was separated into four components (N, A-1, A-2', and A-2) by paper electrophoresis. Complete structures of the major acidic oligosaccharides (A-1 and A-2) were elucidated by sequential exoglycosidase digestion in combination with methylation study and periodate oxidation as follows: NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(3) [Gal beta 1 leads to 4GlcNAc beta 1 leads to to 2Man alpha 1 leads to 3(6)] (GlcNAc beta 1 leads to 4)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAc and NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3) (GlcNAc beta 1 leads to 4)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6) GlcNAc. The abbreviations used are: NeuAc, N-acetylneuraminic acid; GlcNAc, N-acetylglucosamine; Fuc, fucose.
Notes: Yoshima, H
Furthmayr, H
Kobata, A
AM21714-01CO9S/AM/NIADDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1980 Oct 25;255(20):9713-8.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 493
Author: Yoshima, H., Matsumoto, A., Mizuochi, T., Kawasaki, T. and Kobata, A.
Year: 1981
Title: Comparative study of the carbohydrate moieties of rat and human plasma alpha 1-acid glycoproteins
Journal: J Biol Chem
Volume: 256
Issue: 16
Pages: 8476-84
Epub Date: 1981/08/25
Date: Aug 25
Short Title: Comparative study of the carbohydrate moieties of rat and human plasma alpha 1-acid glycoproteins
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
Accession Number: 7263664
Keywords: Animals
Asparagine
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Gel
Glycoside Hydrolases
Humans
*Orosomucoid
Rats
Species Specificity
Abstract: Rat and human alpha 1-acid glycoproteins contain 5 and 6 acidic asparagine-linked sugar chains in 1 molecule, respectively. The structures of these sugar chains were studied by sequential exoglycosidase digestion in combination with methylation analysis after being released from the polypeptide back bone by hydrazinolysis. All oligosaccharides obtained from both glycoproteins contain N-acetylneuraminic acid. Sialidase digestion of the acidic oligosaccharides from rat sample released three biantennary complex type sugar chains and three triantennary complex type sugar chains, the outer chains of which are composed of Gal beta 1 leads to 3GlcNAc and/or Gal beta 1 leads to 4GlcNAc. In the original acidic oligosaccharides, sialic acids occur as three type linkages: NeuAc alpha 2 leads to 3Gal, NeuAc alpha 2 leads to 6Gal, and Gal beta 1 leads to 3(NeuAc alpha 2 leads to 6)GlcNAc. In addition to the five sugar chains reported by Fournet, B., Montreuil, J., Strecker, G., Dorland, L., Haverkamp, J., Vilegenthart, J. F. G., Binette, J. P., and Schmid, K. ((1978) Biochemistry 17, 5206-5214), the structures of three new sugar chains of human alpha 1-acid glycoprotein were elucidated.
Notes: Yoshima, H
Matsumoto, A
Mizuochi, T
Kawasaki, T
Kobata, A
Journal Article
Research Support, Non-U.S. Gov't
United States
J Biol Chem. 1981 Aug 25;256(16):8476-84.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1069
Author: Young, A. R., Chan, E. Y., Hu, X. W., Köchl, R., Crawshaw, S. G., High, S., Hailey, D. W., Lippincott-Schwartz, J. and Tooze, S. A.
Year: 2006
Title: Starvation and ULK1-dependent cycling of mammalian Atg9 between the TGN and endosomes
Journal: J Cell Sci
Volume: 119
Issue: Pt 18
Pages: 3888-900
Epub Date: 2006/08/31
Date: Sep 15
Short Title: Starvation and ULK1-dependent cycling of mammalian Atg9 between the TGN and endosomes
Alternate Journal: Journal of cell science
ISSN: 0021-9533 (Print)
0021-9533
DOI: 10.1242/jcs.03172
Accession Number: 16940348
Keywords: Animals
Autophagy-Related Protein-1 Homolog
Autophagy-Related Proteins
Endosomes/*metabolism
Green Fluorescent Proteins/metabolism
Humans
Intracellular Signaling Peptides and Proteins/*metabolism
Membrane Proteins/chemistry/*metabolism/ultrastructure
Protein Transport
Protein-Serine-Threonine Kinases/*metabolism
Rats
Recombinant Fusion Proteins/metabolism
rab GTP-Binding Proteins/metabolism
trans-Golgi Network/*metabolism/ultrastructure
Abstract: Autophagy, fundamentally a lysosomal degradation pathway, functions in cells during normal growth and certain pathological conditions, including starvation, to maintain homeostasis. Autophagosomes are formed through a mechanism that is not well understood, despite the identification of many genes required for autophagy. We have studied the mammalian homologue of Atg9p, a multi-spanning transmembrane protein essential in yeast for autophagy, to gain a better understanding of the function of this ubiquitious protein. We show that both the N- and C-termini of mammalian Atg9 (mAtg9) are cytosolic, and predict that mAtg9 spans the membrane six times. We find that mAtg9 is located in the trans-Golgi network and late endosomes and colocalizes with TGN46, the cation-independent mannose-6-phosphate receptor, Rab7 and Rab9. Amino acid starvation or rapamycin treatment, which upregulates autophagy, causes a redistribution of mAtg9 from the TGN to peripheral, endosomal membranes, which are positive for the autophagosomal marker GFP-LC3. siRNA-mediated depletion of the putative mammalian homologue of Atg1p, ULK1, inhibits this starvation-induced redistribution. The redistribution of mAtg9 also requires PI 3-kinase activity, and is reversed after restoration of amino acids. We speculate that starvation-induced autophagy, which requires mAtg9, may rely on an alteration of the steady-state trafficking of mAtg9, in a Atg1-dependent manner.
Notes: Young, Andrew R J
Chan, Edmond Y W
Hu, Xiao Wen
Köchl, Robert
Crawshaw, Samuel G
High, Stephen
Hailey, Dale W
Lippincott-Schwartz, Jennifer
Tooze, Sharon A
Journal Article
England
J Cell Sci. 2006 Sep 15;119(Pt 18):3888-900. doi: 10.1242/jcs.03172. Epub 2006 Aug 29.
Author Address: Cancer Research UK London Research Institute, 44 Lincoln's Inn Fields, London, WC2A 3PX, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1361
Author: Yu, C., Sonnen, A. F., George, R., Dessailly, B. H., Stagg, L. J., Evans, E. J., Orengo, C. A., Stuart, D. I., Ladbury, J. E., Ikemizu, S., Gilbert, R. J. and Davis, S. J.
Year: 2011
Title: Rigid-body ligand recognition drives cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor triggering
Journal: J Biol Chem
Volume: 286
Issue: 8
Pages: 6685-96
Epub Date: 2010/12/16
Date: Feb 25
Short Title: Rigid-body ligand recognition drives cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor triggering
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M110.182394
PMCID: PMC3057841
Accession Number: 21156796
Keywords: Animals
Antigens, CD/*chemistry/genetics/immunology
B7-1 Antigen/*chemistry/genetics/immunology
B7-2 Antigen/*chemistry/genetics/immunology
Binding Sites
CHO Cells
CTLA-4 Antigen
Cricetinae
Cricetulus
Crystallography, X-Ray
Humans
Protein Structure, Tertiary
Receptors, Antigen, T-Cell/*chemistry/genetics/immunology
Thermodynamics
Abstract: The inhibitory T-cell surface-expressed receptor, cytotoxic T lymphocyte-associated antigen-4 (CTLA-4), which belongs to the class of cell surface proteins phosphorylated by extrinsic tyrosine kinases that also includes antigen receptors, binds the related ligands, B7-1 and B7-2, expressed on antigen-presenting cells. Conformational changes are commonly invoked to explain ligand-induced "triggering" of this class of receptors. Crystal structures of ligand-bound CTLA-4 have been reported, but not the apo form, precluding analysis of the structural changes accompanying ligand binding. The 1.8-Å resolution structure of an apo human CTLA-4 homodimer emphasizes the shared evolutionary history of the CTLA-4/CD28 subgroup of the immunoglobulin superfamily and the antigen receptors. The ligand-bound and unbound forms of both CTLA-4 and B7-1 are remarkably similar, in marked contrast to B7-2, whose binding to CTLA-4 has elements of induced fit. Isothermal titration calorimetry reveals that ligand binding by CTLA-4 is enthalpically driven and accompanied by unfavorable entropic changes. The similarity of the thermodynamic parameters determined for the interactions of CTLA-4 with B7-1 and B7-2 suggests that the binding is not highly specific, but the conformational changes observed for B7-2 binding suggest some level of selectivity. The new structure establishes that rigid-body ligand interactions are capable of triggering CTLA-4 phosphorylation by extrinsic kinase(s).
Notes: 1083-351x
Yu, Chao
Sonnen, Andreas F-P
George, Roger
Dessailly, Benoit H
Stagg, Loren J
Evans, Edward J
Orengo, Christine A
Stuart, David I
Ladbury, John E
Ikemizu, Shinji
Gilbert, Robert J C
Davis, Simon J
081894/Wellcome Trust/United Kingdom
G1000099/Medical Research Council/United Kingdom
G1100525/Medical Research Council/United Kingdom
G19/3/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 Feb 25;286(8):6685-96. doi: 10.1074/jbc.M110.182394. Epub 2010 Dec 14.
Author Address: Nuffield Department of Clinical Medicine and MRC Human Immunology Unit, The University of Oxford, John Radcliffe Hospital, Headington, Oxford OX3 9DU, United Kingdom.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2028
Author: Yu, M., Chu, S., Fei, B., Fang, X. and Liu, Z.
Year: 2019
Title: O-GlcNAcylation of ITGA5 facilitates the occurrence and development of colorectal cancer
Journal: Exp Cell Res
Volume: 382
Issue: 2
Pages: 111464
Epub Date: 2019/06/17
Date: Sep 15
Short Title: O-GlcNAcylation of ITGA5 facilitates the occurrence and development of colorectal cancer
Alternate Journal: Experimental cell research
ISSN: 0014-4827
DOI: 10.1016/j.yexcr.2019.06.009
Accession Number: 31202709
Keywords: Acetylglucosamine/*metabolism
Acylation
Apoptosis
Carcinogenesis/*metabolism/*pathology
Cell Line, Tumor
Cell Proliferation
Colorectal Neoplasms/*metabolism
Humans
Integrin alpha5/*metabolism
Up-Regulation
*Colorectal cancer
*Itga5
*O-GlcNAcylation
*Proliferation
*Tumorigenesis
Abstract: BACKGROUND/OBJECTIVE: Integrin α5 (ITGA5) as one member of integrins family, plays an important role in promoting cancer cell metastasis and invasion through inducing the communications among different cells or cells with extracellular matrix (ECM). However, the mechanisms underlying ITGA5 in colorectal cancer (CRC) progression need to be explored, especially for its O-GlcNAcylation. To this end, the current study was performed to explore the effects of O-GlcNAcylation on ITGA5 expression, as well as to probe the effects of ITGA5 O-GlcNAcylation on CRC progression. METHODS: The expression profiles of ITGA5, OGT and O-GlcNAc in CRC tissues and cells were detected by immunohistochemistry (IHC), RT-PCR and western blotting. CCK-8, flow cytometry and xenotransplantation assays were used to assess cell growth, apoptosis and tumorigenesis. Immunoprecipitation (IP), in vitro O-GlcNAcylation of ITGA5 and enzymatic labelling of O-GlcNAc assays were used to detect the O-GlcNAcylation of ITGA5 protein. RESULTS: The expression of ITGA5, OGT and O-GlcNAc were all elevated in CRC tissues and cells compared with the normal tissues and cells. Up-regulation of ITGA5 in CRC RKO cells enhanced cell growth and tumorigenesis while decreased cell apoptosis, while down-regulation of ITGA5 in CRC SW620 cells decreased cell growth and tumorigenesis and induced cell apoptosis. Besides, PUGNAc, GlcN or PUGNAc + GlcNAc treatment increased ITGA5 protein expression in RKO and SW620 cells, as well as increased its protein stability via enhancing its O-GlcNAcylation. CONCLUSION: Collectively, the present study makes clear that ITGA5 overexpression accelerates the progression of CRC, which is closely associated to its enhanced O-GlcNAcylation.
Notes: 1090-2422
Yu, Miao
Chu, Songtao
Fei, Bingyuan
Fang, Xuedong
Liu, Zhuo
Journal Article
United States
Exp Cell Res. 2019 Sep 15;382(2):111464. doi: 10.1016/j.yexcr.2019.06.009. Epub 2019 Jun 13.
Author Address: Department of Gastrointestinal Colorectal and Anal Surgery, CHINA-JAPAN Union Hospital of Jilin University, Changchun, Jilin Province, 130000, China.
Department of Forensic Medicine of Basic Medical College, Beihua University, Jilin, Jilin Province, 132013, China.
Department of Gastrointestinal Colorectal and Anal Surgery, CHINA-JAPAN Union Hospital of Jilin University, Changchun, Jilin Province, 130000, China. Electronic address: fangxuedong@medmail.com.cn.
Department of Gastrointestinal Colorectal and Anal Surgery, CHINA-JAPAN Union Hospital of Jilin University, Changchun, Jilin Province, 130000, China. Electronic address: doctorliu2019@163.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1922
Author: Yu, P., Hu, L., Xie, J., Chen, S., Huang, L., Xu, Z., Liu, X., Zhou, Q., Yuan, P., Yan, X., Jin, J., Shen, Y., Zhu, W., Fu, L., Chen, Q., Yu, J., Hu, J., Cao, Q., Wan, R. and Hong, K.
Year: 2018
Title: O-GlcNAcylation of cardiac Nav1.5 contributes to the development of arrhythmias in diabetic hearts
Journal: Int J Cardiol
Volume: 260
Pages: 74-81
Epub Date: 2018/03/14
Date: Jun 1
Short Title: O-GlcNAcylation of cardiac Nav1.5 contributes to the development of arrhythmias in diabetic hearts
Alternate Journal: International journal of cardiology
ISSN: 0167-5273
DOI: 10.1016/j.ijcard.2018.02.099
Accession Number: 29530619
Keywords: Acetylglucosamine/*metabolism
Acylation/physiology
Animals
Arrhythmias, Cardiac/etiology/*metabolism/physiopathology
Diabetes Mellitus, Experimental/complications/*metabolism/physiopathology
HEK293 Cells
Humans
Male
NAV1.5 Voltage-Gated Sodium Channel/*metabolism
Rats
Rats, Sprague-Dawley
*Arrhythmias
*Diabetic mellitus
*Nav1.5
*O-GlcNAc
Abstract: BACKGROUND: Cardiovascular complications are major causes of mortality and morbidity in diabetic patients. The mechanisms underlying the progression of diabetic heart (DH) to ventricular arrhythmias are unclear. O-linked GlcNAcylation (O-GlcNAc) is a reversible post-translational modification for the regulation of diverse cellular processes. The purpose of this study was to assess whether the cardiac voltage-gated sodium channel (Nav1.5) is subjected to O-linked GlcNAcylation (O-GlcNAc), which plays an essential role in DH-induced arrhythmias. METHODS AND RESULTS: In this study, Sprague-Dawley rats (male, 200-230 g) were treated with a single high-dose of streptozotocin (STZ, 80 mg/kg) to generate a rat model of diabetes. STZ-induced 3-month diabetic rats displayed increased susceptibility to ventricular arrhythmias. The elevated O-GlcNAc modification was correlated with decreases in both total and cytoplasmic Nav1.5 expression in vivo and in vitro. In addition, both co-immunoprecipitation and immunostaining assays demonstrated that hyperglycemia could increase the O-GlcNAc-modified Nav1.5 levels and decrease the interaction between Nav1.5 and Nav1.5-binding proteins Nedd4-2/SAP-97. Furthermore, patch-clamp measurements in HEK-293 T cells showed that Nav1.5 current densities decreased by 30% after high-glucose treatment, and the sodium currents increased via O-GlcNAc inhibition. CONCLUSION: Our data suggested that hyperglycemia increased the O-GlcNAc modification of Nav1.5 expression and decreased the interaction between Nav1.5 and Nedd4-2/SAP-97, which led to the abnormal expression and distribution of Nav1.5, loss of function of the sodium channel, and prolongation of the PR/QT interval. Excessive O-GlcNAc modification of Nav1.5 is a novel signaling event, which may be an underlying contributing factor for the development of the arrhythmogenesis in DH.
Notes: 1874-1754
Yu, Peng
Hu, Lili
Xie, Jinyan
Chen, Sisi
Huang, Lin
Xu, Zixuan
Liu, Xiao
Zhou, Qiongqiong
Yuan, Ping
Yan, Xia
Jin, Jiejin
Shen, Yang
Zhu, Wengen
Fu, Linghua
Chen, Qi
Yu, Jianhua
Hu, Jianxin
Cao, Qing
Wan, Rong
Hong, Kui
Journal Article
Netherlands
Int J Cardiol. 2018 Jun 1;260:74-81. doi: 10.1016/j.ijcard.2018.02.099. Epub 2018 Feb 27.
Author Address: Department of Cardiovascular Medicine, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China; Jiangxi Key Laboratory of Molecular Medicine, Nanchang, Jiangxi 330006, China.
Department of Cardiovascular Medicine, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China; Department of Nephrology, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, China.
Jiangxi Key Laboratory of Molecular Medicine, Nanchang, Jiangxi 330006, China.
Department of General Surgery, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, China.
Department of Cardiovascular Medicine, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China.
Department of Cardiovascular Medicine, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China; Jiangxi Key Laboratory of Molecular Medicine, Nanchang, Jiangxi 330006, China. Electronic address: rong87223@163.com.
Department of Cardiovascular Medicine, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China; Jiangxi Key Laboratory of Molecular Medicine, Nanchang, Jiangxi 330006, China. Electronic address: hongkui88@163.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1979
Author: Yu, T., Liang, L., Zhao, X. and Yin, Y.
Year: 2018
Title: Structural and biochemical studies of the extracellular domain of Myelin protein zero-like protein 1
Journal: Biochem Biophys Res Commun
Volume: 506
Issue: 4
Pages: 883-890
Epub Date: 2018/11/06
Date: Dec 2
Short Title: Structural and biochemical studies of the extracellular domain of Myelin protein zero-like protein 1
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2018.10.161
Accession Number: 30392906
Keywords: Amino Acid Sequence
Asparagine/chemistry
Calorimetry
Concanavalin A
Crystallography, X-Ray
Glycosylation
Humans
Intracellular Signaling Peptides and Proteins/*chemistry/*metabolism
Phosphoproteins/*chemistry/*metabolism
Protein Domains
Protein Multimerization
*Crystallography
*Hcc
*Mpzl1
*Metastasis
*N-linked glycosylation
Abstract: Myelin protein zero-like protein 1 (MPZL1) is a member of the immunoglobulin superfamily, and is also a receptor of concanavalin A (ConA). MPZL1 is upregulated in hepatocellular carcinoma (HCC) and accelerates migration of HCC cells. However, function of MPZL1 as a receptor of ConA and its role in HCC development are largely unknown. To elucidate the functional basis, we have determined the crystal structure of the extracellular domain of MPZL1 at 2.7 Å resolution. Overall, it folds like a typical immunoglobulin variable-like domain that is much like MPZ. Unexpectedly, we found Asn50 is a unique glycosylation site and the glycosylation mediates its interaction with ConA. Furthermore, we also found that MPZL1 exists as a homodimer in the crystal, in which hydrogen bonds between Ser86 and Val145 play an important role. Our results demonstrate that glycosylation of Asn50 is essential for its function as a receptor of ConA. We propose that dimerization of MPZL1 participates in control of its signal transmission in cell adhesion.
Notes: 1090-2104
Yu, Tianshu
Liang, Ling
Zhao, Xuyang
Yin, Yuxin
Journal Article
Research Support, Non-U.S. Gov't
United States
Biochem Biophys Res Commun. 2018 Dec 2;506(4):883-890. doi: 10.1016/j.bbrc.2018.10.161. Epub 2018 Nov 2.
Author Address: Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, 100871, China.
Department of Biophysics, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, 100191, China; Institute of Systems Biomedicine, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, 100191, China.
Institute of Systems Biomedicine, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, 100191, China.
Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, 100871, China; Institute of Systems Biomedicine, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, 100191, China; Department of Pathology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, 100191, China. Electronic address: yinyuxin@hsc.pku.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 635
Author: Yu, X. C., Sturrock, E. D., Wu, Z., Biemann, K., Ehlers, M. R. and Riordan, J. F.
Year: 1997
Title: Identification of N-linked glycosylation sites in human testis angiotensin-converting enzyme and expression of an active deglycosylated form
Journal: J Biol Chem
Volume: 272
Issue: 6
Pages: 3511-9
Epub Date: 1997/02/07
Date: Feb 7
Short Title: Identification of N-linked glycosylation sites in human testis angiotensin-converting enzyme and expression of an active deglycosylated form
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.272.6.3511
Accession Number: 9013598
Keywords: Animals
CHO Cells
Chromatography, High Pressure Liquid
Cricetinae
Cricetulus
Female
Glycosylation
Humans
Male
Ovary/metabolism
Peptidyl-Dipeptidase A/*metabolism
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Testis/*enzymology
Abstract: The sites of glycosylation of Chinese hamster ovary cell expressed testicular angiotensin-converting enzyme (tACE) have been determined by matrix-assisted laser desorption ionization/time of flight/mass spectrometry of peptides generated by proteolytic and cyanogen bromide digestion. Two of the seven potential N-linked glycosylation sites, Asn90 and Asn109, were found to be fully glycosylated by analysis of peptides before and after treatment with a series of glycosidases and with endoproteinase Asp-N. The mass spectra of the glycopeptides exhibit characteristic clusters of peaks which indicate the N-linked glycans in tACE to be mostly of the biantennary, fucosylated complex type. This structural information was used to demonstrate that three other sites, Asn155, Asn337, and Asn586, are partially glycosylated, whereas Asn72 appears to be fully glycosylated. The only potential site that was not modified is Asn620. Sequence analysis of tryptic peptides obtained from somatic ACE (human kidney) identified six glycosylated and one unglycosylated Asn. Only one of these glycosylation sites had a counterpart in tACE. Comparison of the two proteins reveals a pattern in which amino-terminal N-linked sites are preferred. The functional significance of glycosylation was examined with a tACE mutant lacking the O-glycan-rich first amino-terminal 36 residues and truncated at Ser625. When expressed in the presence of the alpha-glucosidase I inhibitor N-butyldeoxynojirimycin and treated with endoglycosidase H to remove all but the terminal N-acetylglucosamine residues, it retained full enzymatic activity, was electrophoretically homogeneous, and is a good candidate for crystallographic studies.
Notes: Yu, X C
Sturrock, E D
Wu, Z
Biemann, K
Ehlers, M R
Riordan, J F
GM05472/GM/NIGMS NIH HHS/United States
HL34704/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1997 Feb 7;272(6):3511-9. doi: 10.1074/jbc.272.6.3511.
Author Address: Center for Biochemical and Biophysical Sciences and Medicine, Harvard Medical School, Boston, Massachusetts 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1111
Author: Yu, Y., Hoffhines, A. J., Moore, K. L. and Leary, J. A.
Year: 2007
Title: Determination of the sites of tyrosine O-sulfation in peptides and proteins
Journal: Nat Methods
Volume: 4
Issue: 7
Pages: 583-8
Epub Date: 2007/06/15
Date: Jul
Short Title: Determination of the sites of tyrosine O-sulfation in peptides and proteins
Alternate Journal: Nature methods
ISSN: 1548-7091 (Print)
1548-7091
DOI: 10.1038/nmeth1056
Accession Number: 17558413
Keywords: Amino Acid Sequence
Humans
Molecular Sequence Data
Peptides/*chemistry
Protein Processing, Post-Translational
Proteins/*chemistry
Sulfotransferases/chemistry
Tandem Mass Spectrometry/*methods
Tyrosine/*analogs & derivatives/analysis/chemistry
Abstract: Tyrosine O-sulfation is a key post-translational modification that regulates protein-protein interactions in extracellular space. We describe a subtractive strategy to determine the sites of tyrosine O-sulfation in proteins. Hydroxyl groups on unsulfated tyrosines are blocked by stoichiometric acetylation in a one-step reaction using sulfosuccinimidyl acetate (S-NHSAc) in the presence of imidazole at pH 7.0. The presence of sulfotyrosine is indicated by the detection of free tyrosine after tandem mass spectrometry (MS/MS) analysis under conditions in which the sulfuryl group of sulfotyrosine is labile. Since phosphorylation and sulfation of tyrosine are isobaric, we used alkaline phosphatase treatment to distinguish these two modifications. Using this methodology we identified the sites and the order of sulfation of several peptides mediated by purified human tyrosylprotein sulfotransferases (TPSTs), and unambiguously determined the tyrosine sulfation sites in mouse lumican and human vitronectin.
Notes: Yu, Yonghao
Hoffhines, Adam J
Moore, Kevin L
Leary, Julie A
R01 HL074015/HL/NHLBI NIH HHS/United States
R01 HL074015-04/HL/NHLBI NIH HHS/United States
GM63581/GM/NIGMS NIH HHS/United States
HL 074015/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
Nat Methods. 2007 Jul;4(7):583-8. doi: 10.1038/nmeth1056. Epub 2007 Jun 10.
Author Address: Genome Center, Department of Chemistry, University of California, Davis, California 95616, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1537
Author: Yu, Y., Zhu, J., Huang, P. S., Wang, J. H., Pullen, N. and Springer, T. A.
Year: 2013
Title: Domain 1 of mucosal addressin cell adhesion molecule has an I1-set fold and a flexible integrin-binding loop
Journal: J Biol Chem
Volume: 288
Issue: 9
Pages: 6284-94
Epub Date: 2013/01/09
Date: Mar 1
Short Title: Domain 1 of mucosal addressin cell adhesion molecule has an I1-set fold and a flexible integrin-binding loop
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M112.413153
PMCID: PMC3585063
Accession Number: 23297416
Keywords: Animals
CHO Cells
Cell Adhesion/physiology
Cell Adhesion Molecules/chemistry/genetics/metabolism
Cricetinae
Cricetulus
Crystallography, X-Ray
Humans
Immunoglobulins/*chemistry/genetics/metabolism
Integrin alpha4/chemistry/genetics/metabolism
Integrin beta Chains/chemistry/genetics/metabolism
Mice
Mucoproteins/*chemistry/genetics/metabolism
Protein Structure, Secondary
Protein Structure, Tertiary
Abstract: Mucosal addressin cell adhesion molecule (MAdCAM) binds integrin α4β7. Their interaction directs lymphocyte homing to mucosa-associated lymphoid tissues. The interaction between the two immunoglobulin superfamily (IgSF) domains of MAdCAM and integrin α4β7 is unusual in its ability to mediate either rolling adhesion or firm adhesion of lymphocytes on vascular surfaces. We determined four crystal structures of the IgSF domains of MAdCAM to test for unusual structural features that might correlate with this functional diversity. Higher resolution 1.7- and 1.4-Å structures of the IgSF domains of MAdCAM in a previously described crystal lattice revealed two alternative conformations of the integrin-binding loop, which were deformed by large lattice contacts. New crystal forms in the presence of two different Fabs to MAdCAM demonstrate a shift in IgSF domain topology from the I2- to I1-set, with a switch of integrin-binding loop from CC' to CD. The I1-set fold and CD loop appear biologically relevant. The different conformations seen in crystal structures suggest that the integrin-binding loop of MAdCAM is inherently flexible. This contrasts with rigidity of the corresponding loops in vascular cell adhesion molecule, intercellular adhesion molecule (ICAM)-1, ICAM-2, ICAM-3, and ICAM-5 and may reflect a specialization of MAdCAM to mediate both rolling and firm adhesion by binding to different α4β7 integrin conformations.
Notes: 1083-351x
Yu, Yamei
Zhu, Jianghai
Huang, Po-Ssu
Wang, Jia-Huai
Pullen, Nick
Springer, Timothy A
P01 HL103526/HL/NHLBI NIH HHS/United States
HL103526/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2013 Mar 1;288(9):6284-94. doi: 10.1074/jbc.M112.413153. Epub 2013 Jan 7.
Author Address: Program in Cellular and Molecular Medicine, Children's Hospital Boston, and Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02215, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1625
Author: Yuasa, T., Amo, K., Ishikura, S., Nagaya, H., Uchiyama, K., Hashida, S. and Ebina, Y.
Year: 2014
Title: Development of in vitro model of insulin receptor cleavage induced by high glucose in HepG2 cells
Journal: Biochem Biophys Res Commun
Volume: 445
Issue: 1
Pages: 236-43
Epub Date: 2014/02/11
Date: Feb 28
Short Title: Development of in vitro model of insulin receptor cleavage induced by high glucose in HepG2 cells
Alternate Journal: Biochemical and biophysical research communications
ISSN: 0006-291x
DOI: 10.1016/j.bbrc.2014.01.187
Accession Number: 24508798
Keywords: Acetylglucosamine/metabolism
Acylation/drug effects
Blood Glucose/metabolism
Blotting, Western
Calcium/metabolism
Cell Line, Tumor
Diabetes Mellitus/blood
Dose-Response Relationship, Drug
Enzyme-Linked Immunosorbent Assay
Glucose/*pharmacology
HEK293 Cells
HeLa Cells
Hep G2 Cells
Humans
Models, Biological
N-Acetylglucosaminyltransferases/genetics/metabolism
Peptide Hydrolases/metabolism
Proteolysis/*drug effects
RNA Interference
Receptor, Insulin/blood/*metabolism
Time Factors
Calcium-dependent protease
HepG2 cells
O-GlcNAcylation
Shedding
Soluble insulin receptor
Abstract: Soluble insulin receptor (sIR), the ectodomain of IR, has been detected in human plasma, and its concentration parallels that of blood glucose in patients with diabetes. IR has a pivotal role in glucose homeostasis and diabetes development; therefore, cleavage of IR promoted by hyperglycemia is involved in insulin resistance and glucose toxicity. To elucidate the physiology of sIR, we developed an in vitro model mimicking the changes in sIR levels in plasma from patients with diabetes. Among four human cell lines that expressed IR, spontaneous cleavage of IR occurred only in HepG2 cells. The molecular characteristics of sIR derived from HepG2 cells were similar to those of sIR detected in human plasma. The concentration of sIR in the medium did not differ between basal and high-glucose conditions in the initial 24-h period, but increasing the duration of pre-stimulation (>48 h) led to a significant increase in sIR levels in cells exposed to high glucose. Additionally, glucose-dependent increment of sIR was reversible in this model. These results are consistent with the observation of plasma sIR in patients with diabetes. Using this model, O-linked N-acetylglucosamine modification was determined to be involved in high-glucose-induced IR cleavage. A calcium-dependent protease was shown to cleave IR extracellularly. These findings show that this in vitro model could be useful for determining the molecular mechanism underlying IR cleavage.
Notes: 1090-2104
Yuasa, Tomoyuki
Amo, Kikuko
Ishikura, Shuhei
Nagaya, Hisao
Uchiyama, Keiji
Hashida, Seiichi
Ebina, Yousuke
Journal Article
United States
Biochem Biophys Res Commun. 2014 Feb 28;445(1):236-43. doi: 10.1016/j.bbrc.2014.01.187. Epub 2014 Feb 6.
Author Address: Division of Molecular Genetics, Institute for Enzyme Research, Tokushima University, 3-18-15 Kuramotocho, Tokushima 770-8503, Japan.
Institute for Health Sciences, Tokushima Bunri University, Tokushima 770-8514, Japan.
Division of Molecular Genetics, Institute for Enzyme Research, Tokushima University, 3-18-15 Kuramotocho, Tokushima 770-8503, Japan. Electronic address: ebina@za3.so-net.ne.jp.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 322
Author: Yuen, C. T., Carr, S. A. and Feizi, T.
Year: 1990
Title: The spectrum of N-linked oligosaccharide structures detected by enzymic microsequencing on a recombinant soluble CD4 glycoprotein from Chinese hamster ovary cells
Journal: Eur J Biochem
Volume: 192
Issue: 2
Pages: 523-8
Epub Date: 1990/09/11
Date: Sep 11
Short Title: The spectrum of N-linked oligosaccharide structures detected by enzymic microsequencing on a recombinant soluble CD4 glycoprotein from Chinese hamster ovary cells
Alternate Journal: European journal of biochemistry
ISSN: 0014-2956 (Print)
0014-2956
DOI: 10.1111/j.1432-1033.1990.tb19256.x
Accession Number: 2209609
Keywords: Animals
*CD4 Antigens/genetics
Carbohydrate Conformation
Carbohydrate Sequence
Cell Line
Chromatography, Gel
Cricetinae
Cricetulus
Electrophoresis, Paper
Female
Humans
Molecular Sequence Data
Neuraminidase
Oligosaccharides/*isolation & purification
Ovary
Recombinant Proteins
Transfection
Abstract: Structures of the N-linked oligosaccharides of a recombinant soluble form of human CD4 glycoprotein (sCD4) have been investigated by enzymic microsequencing. The glycoprotein has two N-glycosylation sites, Asn271 and Asn300, at both of which evidence for the presence of complex type biantennary sialo-oligosaccharides has been obtained previously by mass spectrometric analyses [Carr, S.A., Hemling, M.E., Folena-Wasserman, G., Sweet, R.W., Anumula, K., Barr, J.R., Huddleston, M.J. & Taylor, P. (1989) J. Biol. Chem. 264, 21,286-21,295]. Among oligosaccharides released from sCD4 by hydrazinolysis and labelled with NaB3H4, neutral (12.8%) and acidic (87.2%) oligosaccharides were detected by paper electrophoresis. The latter were rendered neutral following sialidase treatment indicating that acidity was due exclusively to the presence of sialic acid residues. By enzymic microsequencing of the sialidase-treated oligosaccharides (fractionated on affinity columns of Ricinis communis agglutinin 120 and concanavalin A) in conjunction with methylation data from the earlier study, 14 sequences were identified. These accounted for over 80% of the sialidase-treated oligosaccharides of sCD4 as follows: [formula: see text] where +/- indicates residues present on only a proportion of chains. The spectrum of oligosaccharide structures released from each glycosylation site was assessed as being similar to that of total oligosaccharides on the basis of their chromatographic profiles on the lectin columns and on Bio-Gel P-4.
Notes: Yuen, C T
Carr, S A
Feizi, T
Journal Article
England
Eur J Biochem. 1990 Sep 11;192(2):523-8. doi: 10.1111/j.1432-1033.1990.tb19256.x.
Author Address: Glycoconjugates Section, Medical Research Council Clinical Research Centre, Harrow, England.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 967
Author: Yusa, A., Kitajima, K. and Habuchi, O.
Year: 2005
Title: N-linked oligosaccharides are required to produce and stabilize the active form of chondroitin 4-sulphotransferase-1
Journal: Biochem J
Volume: 388
Issue: Pt 1
Pages: 115-21
Epub Date: 2005/01/05
Date: May 15
Short Title: N-linked oligosaccharides are required to produce and stabilize the active form of chondroitin 4-sulphotransferase-1
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
DOI: 10.1042/bj20041573
PMCID: PMC1186699
Accession Number: 15628971
Keywords: Animals
CHO Cells
COS Cells
Chlorocebus aethiops
Cricetinae
Cricetulus
Enzyme Activation
Mutagenesis, Site-Directed
Mutation
Oligosaccharides/chemistry/*metabolism
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/chemistry/metabolism
Recombinant Proteins
Sulfotransferases/chemistry/genetics/*metabolism
Tunicamycin
Abstract: C4ST-1 (chondroitin 4-sulphotransferase-1) transfers sulphate to position 4 of N-acetylgalactosamine in chondroitin. We showed previously that purified C4ST-1 from the culture medium of rat chondrosarcoma cells was a glycoprotein containing approx. 35% N-linked oligosaccharides. In the present paper, we investigated the functional role of the N-linked oligosaccharides attached to C4ST-1. We found that (i) treatment of recombinant C4ST-1 with peptide N-glycosidase F caused a marked decrease in activity, (ii) production of the active form of C4ST-1 by COS-7 cells transfected with cDNA of C4ST-1 was inhibited by tunicamycin, (iii) deletion of the N-glycosylation site located at the C-terminal region of C4ST-1 abolished activity, (iv) attachment of a single N-glycan at the C-terminal region supported production of the active form of C4ST-1, but the resulting recombinant enzyme was much more unstable at 37 degrees C than the control recombinant protein, and (v) truncation of C-terminal region up to the N-glycosylation site at the C-terminal region resulted in total loss of activity. These observations strongly suggest that N-linked oligosaccharides attached to C4ST-1 contribute to the production and stability of the active form of C4ST-1. In addition, the N-linked oligosaccharide at the C-terminal region appears to affect the glycosylation pattern of recombinant C4ST; a broad protein band of the wildtype protein resulting from microheterogeneity of N-linked oligosaccharides disappeared and four discrete protein bands with different numbers of N-linked oligosaccharides appeared when the N-linked oligosaccharide at the C-terminal region was deleted.
Notes: 1470-8728
Yusa, Akiko
Kitajima, Ken
Habuchi, Osami
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2005 May 15;388(Pt 1):115-21. doi: 10.1042/BJ20041573.
Author Address: Department of Chemistry, Aichi University of Education, Kariya, Aichi 448-8542, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1123
Author: Yuzawa, S., Opatowsky, Y., Zhang, Z., Mandiyan, V., Lax, I. and Schlessinger, J.
Year: 2007
Title: Structural basis for activation of the receptor tyrosine kinase KIT by stem cell factor
Journal: Cell
Volume: 130
Issue: 2
Pages: 323-34
Epub Date: 2007/07/31
Date: Jul 27
Short Title: Structural basis for activation of the receptor tyrosine kinase KIT by stem cell factor
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2007.05.055
Accession Number: 17662946
Keywords: Amino Acid Sequence
Binding Sites
Crystallography, X-Ray
Dimerization
Disease
Enzyme Activation
Humans
Ligands
Models, Molecular
Molecular Sequence Data
Mutation/genetics
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Proto-Oncogene Proteins c-kit/*chemistry/*metabolism
Stem Cell Factor/*metabolism
Structure-Activity Relationship
Abstract: Stem Cell Factor (SCF) initiates its multiple cellular responses by binding to the ectodomain of KIT, resulting in tyrosine kinase activation. We describe the crystal structure of the entire ectodomain of KIT before and after SCF stimulation. The structures show that KIT dimerization is driven by SCF binding whose sole role is to bring two KIT molecules together. Receptor dimerization is followed by conformational changes that enable lateral interactions between membrane proximal Ig-like domains D4 and D5 of two KIT molecules. Experiments with cultured cells show that KIT activation is compromised by point mutations in amino acids critical for D4-D4 interaction. Moreover, a variety of oncogenic mutations are mapped to the D5-D5 interface. Since key hallmarks of KIT structures, ligand-induced receptor dimerization, and the critical residues in the D4-D4 interface, are conserved in other receptors, the mechanism of KIT stimulation unveiled in this report may apply for other receptor activation.
Notes: Yuzawa, Satoru
Opatowsky, Yarden
Zhang, Zhongtao
Mandiyan, Valsan
Lax, Irit
Schlessinger, Joseph
AR 051448/AR/NIAMS NIH HHS/United States
AR 051886/AR/NIAMS NIH HHS/United States
P50 AR 054086/AR/NIAMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
United States
Cell. 2007 Jul 27;130(2):323-34. doi: 10.1016/j.cell.2007.05.055.
Author Address: Department of Pharmacology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1623
Author: Yuzwa, S. A., Cheung, A. H., Okon, M., McIntosh, L. P. and Vocadlo, D. J.
Year: 2014
Title: O-GlcNAc modification of tau directly inhibits its aggregation without perturbing the conformational properties of tau monomers
Journal: J Mol Biol
Volume: 426
Issue: 8
Pages: 1736-52
Epub Date: 2014/01/22
Date: Apr 17
Short Title: O-GlcNAc modification of tau directly inhibits its aggregation without perturbing the conformational properties of tau monomers
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2014.01.004
Accession Number: 24444746
Keywords: Acetylglucosamine/chemistry/metabolism
Alzheimer Disease/metabolism
Animals
Fluorescence Resonance Energy Transfer
Humans
Mice
Microtubules/chemistry/metabolism
Mutagenesis, Site-Directed
Nuclear Magnetic Resonance, Biomolecular
Protein Conformation
Protein Folding
Protein Multimerization
Protein Processing, Post-Translational
Protein Structure, Secondary
Recombinant Proteins/chemistry/genetics/metabolism
Tubulin/chemistry/metabolism
tau Proteins/*chemistry/genetics/metabolism
Alzheimer's disease
NMR spectroscopy
glycosylation
paired helical filaments
tauopathy
Abstract: The aggregation of the microtubule-associated protein tau into paired helical filaments to form neurofibrillary tangles constitutes one of the pathological hallmarks of Alzheimer's disease. Tau is post-translationally modified by the addition of N-acetyl-D-glucosamine O-linked to several serine and threonine residues (O-GlcNAc). Previously, increased O-GlcNAcylation of tau has been shown to block the accumulation of tau aggregates within a tauopathy mouse model. Here we show that O-GlcNAc modification of full-length human tau impairs the rate and extent of its heparin-induced aggregation without perturbing its activity toward microtubule polymerization. O-GlcNAcylation, however, does not impact the "global-fold" of tau as measured by a Förster resonance energy transfer assay. Similarly, nuclear magnetic resonance studies demonstrated that O-GlcNAcylation only minimally perturbs the local structural and dynamic features of a tau fragment (residues 353-408) spanning the last microtubule binding repeat to the major GlcNAc-acceptor Ser400. These data indicate that the inhibitory effects of O-GlcNAc on tau aggregation may result from enhanced monomer solubility or the destabilization of fibrils or soluble aggregates, rather than by altering the conformational properties of the monomeric protein. This work further underscores the potential of targeting the O-GlcNAc pathway for potential Alzheimer's disease therapeutics.
Notes: 1089-8638
Yuzwa, Scott A
Cheung, Adrienne H
Okon, Mark
McIntosh, Lawrence P
Vocadlo, David J
MOP 275394/Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
England
J Mol Biol. 2014 Apr 17;426(8):1736-52. doi: 10.1016/j.jmb.2014.01.004. Epub 2014 Jan 18.
Author Address: Department of Molecular Biology and Biochemistry, Simon Fraser University, 8888 University Drive, Burnaby, BC, Canada V5A 1S6; Department of Chemistry, Simon Fraser University, 8888 University Drive, Burnaby, BC, Canada V5A 1S6.
Department of Biochemistry and Molecular Biology, Department of Chemistry, and the Michael Smith Laboratories, University of British Columbia, 2350 Health Sciences Mall, Vancouver, BC, Canada V6T 1Z3.
Department of Molecular Biology and Biochemistry, Simon Fraser University, 8888 University Drive, Burnaby, BC, Canada V5A 1S6; Department of Chemistry, Simon Fraser University, 8888 University Drive, Burnaby, BC, Canada V5A 1S6. Electronic address: dvocadlo@sfu.ca.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1463
Author: Yuzwa, S. A., Shan, X., Macauley, M. S., Clark, T., Skorobogatko, Y., Vosseller, K. and Vocadlo, D. J.
Year: 2012
Title: Increasing O-GlcNAc slows neurodegeneration and stabilizes tau against aggregation
Journal: Nat Chem Biol
Volume: 8
Issue: 4
Pages: 393-9
Epub Date: 2012/03/01
Date: Feb 26
Short Title: Increasing O-GlcNAc slows neurodegeneration and stabilizes tau against aggregation
Alternate Journal: Nature chemical biology
ISSN: 1552-4450
DOI: 10.1038/nchembio.797
Accession Number: 22366723
Keywords: Acetylglucosamine/chemistry/*metabolism
Adaptor Proteins, Signal Transducing/metabolism
Alzheimer Disease/metabolism/pathology
Animals
Carbohydrate Conformation
Disease Models, Animal
Enzyme Inhibitors/pharmacology
Female
Humans
Mice
Mice, Transgenic
N-Acetylglucosaminyltransferases/antagonists & inhibitors/metabolism
Neurodegenerative Diseases/drug therapy/metabolism/pathology
Neurons/drug effects/metabolism/pathology
Phosphorylation
Pyrans/pharmacology
Thiazoles/pharmacology
tau Proteins/genetics/*metabolism
Abstract: Oligomerization of tau is a key process contributing to the progressive death of neurons in Alzheimer's disease. Tau is modified by O-linked N-acetylglucosamine (O-GlcNAc), and O-GlcNAc can influence tau phosphorylation in certain cases. We therefore speculated that increasing tau O-GlcNAc could be a strategy to hinder pathological tau-induced neurodegeneration. Here we found that treatment of hemizygous JNPL3 tau transgenic mice with an O-GlcNAcase inhibitor increased tau O-GlcNAc, hindered formation of tau aggregates and decreased neuronal cell loss. Notably, increases in tau O-GlcNAc did not alter tau phosphorylation in vivo. Using in vitro biochemical aggregation studies, we found that O-GlcNAc modification, on its own, hinders tau oligomerization. O-GlcNAc also inhibits thermally induced aggregation of an unrelated protein, TAK-1 binding protein, suggesting that a basic biochemical function of O-GlcNAc may be to prevent protein aggregation. These results also suggest O-GlcNAcase as a potential therapeutic target that could hinder progression of Alzheimer's disease.
Notes: 1552-4469
Yuzwa, Scott A
Shan, Xiaoyang
Macauley, Matthew S
Clark, Thomas
Skorobogatko, Yuliya
Vosseller, Keith
Vocadlo, David J
Canadian Institutes of Health Research/Canada
Journal Article
Research Support, Non-U.S. Gov't
United States
Nat Chem Biol. 2012 Feb 26;8(4):393-9. doi: 10.1038/nchembio.797.
Author Address: Department of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, British Columbia, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1332
Author: Yuzwa, S. A., Yadav, A. K., Skorobogatko, Y., Clark, T., Vosseller, K. and Vocadlo, D. J.
Year: 2011
Title: Mapping O-GlcNAc modification sites on tau and generation of a site-specific O-GlcNAc tau antibody
Journal: Amino Acids
Volume: 40
Issue: 3
Pages: 857-68
Epub Date: 2010/08/14
Date: Mar
Short Title: Mapping O-GlcNAc modification sites on tau and generation of a site-specific O-GlcNAc tau antibody
Alternate Journal: Amino acids
ISSN: 0939-4451
DOI: 10.1007/s00726-010-0705-1
Accession Number: 20706749
Keywords: Acetylglucosamine/*metabolism
Amino Acid Sequence
Animals
Antibodies/*analysis
Brain/metabolism
Glycosylation
Humans
Mass Spectrometry
Molecular Sequence Data
Peptide Mapping/instrumentation/*methods
Rabbits
Rats
tau Proteins/*chemistry/genetics/immunology/*metabolism
Abstract: The microtubule-associated protein tau is known to be post-translationally modified by the addition of N-acetyl-D: -glucosamine monosaccharides to certain serine and threonine residues. These O-GlcNAc modification sites on tau have been challenging to identify due to the inherent complexity of tau from mammalian brains and the fact that the O-GlcNAc modification typically has substoichiometric occupancy. Here, we describe a method for the production of recombinant O-GlcNAc modified tau and, using this tau, we have mapped sites of O-GlcNAc on tau at Thr-123 and Ser-400 using mass spectrometry. We have also detected the presence of a third O-GlcNAc site on either Ser-409, Ser-412, or Ser-413. Using this information we have raised a rabbit polyclonal IgG antibody (3925) that detects tau O-GlcNAc modified at Ser-400. Further, using this antibody we have detected the Ser-400 tau O-GlcNAc modification in rat brain, which confirms the validity of this in vitro mapping approach. The identification of these O-GlcNAc sites on tau and this antibody will enable both in vivo and in vitro experiments designed to understand the possible functional roles of O-GlcNAc on tau.
Notes: 1438-2199
Yuzwa, Scott A
Yadav, Anuj K
Skorobogatko, Yuliya
Clark, Thomas
Vosseller, Keith
Vocadlo, David J
Canadian Institutes of Health Research/Canada
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't
Austria
Amino Acids. 2011 Mar;40(3):857-68. doi: 10.1007/s00726-010-0705-1. Epub 2010 Aug 13.
Author Address: Department of Molecular Biology and Biochemistry, Simon Fraser University, 8888 University Drive, Burnaby, BC, Canada.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 997
Author: Zajonc, D. M., Cantu, C., 3rd, Mattner, J., Zhou, D., Savage, P. B., Bendelac, A., Wilson, I. A. and Teyton, L.
Year: 2005
Title: Structure and function of a potent agonist for the semi-invariant natural killer T cell receptor
Journal: Nat Immunol
Volume: 6
Issue: 8
Pages: 810-8
Epub Date: 2005/07/12
Date: Aug
Short Title: Structure and function of a potent agonist for the semi-invariant natural killer T cell receptor
Alternate Journal: Nature immunology
ISSN: 1529-2908 (Print)
1529-2908
DOI: 10.1038/ni1224
PMCID: PMC2045075
NIHMSID: NIHMS4829
Accession Number: 16007091
Keywords: Amino Acid Motifs
Animals
Antigens, CD1/biosynthesis
Antigens, CD1d
Cell Wall/metabolism
Crystallography, X-Ray
Dose-Response Relationship, Drug
Flow Cytometry
Galactosylceramides/chemistry
Glycosphingolipids/chemistry
Hot Temperature
Humans
Isoelectric Focusing
Killer Cells, Natural/chemistry/*immunology
Ligands
Lipids/chemistry
Lymphocyte Activation
Mice
Models, Chemical
Models, Molecular
Mutation
Protein Conformation
Receptors, Antigen, T-Cell/chemistry/*immunology
Receptors, Antigen, T-Cell, alpha-beta/*chemistry/*physiology
Recombinant Proteins/chemistry
Structure-Activity Relationship
Surface Plasmon Resonance
Temperature
Th1 Cells/metabolism
Th2 Cells/metabolism
Time Factors
Tissue Distribution
Abstract: Natural killer T cells express a conserved, semi-invariant alphabeta T cell receptor that has specificity for self glycosphingolipids and microbial cell wall alpha-glycuronosylceramide antigens presented by CD1d molecules. Here we report the crystal structure of CD1d in complex with a short-chain synthetic variant of alpha-galactosylceramide at a resolution of 2.2 A. This structure elucidates the basis for the high specificity of these microbial ligands and explains the restriction of the alpha-linkage as a unique pathogen-specific pattern-recognition motif. Comparison of the binding of altered lipid ligands to CD1d and T cell receptors suggested that the differential T helper type 1-like and T helper type 2-like properties of natural killer T cells may originate largely from differences in their 'loading' in different cell types and hence in their tissue distribution in vivo.
Notes: 1529-2916
Zajonc, Dirk M
Cantu, Carlos 3rd
Mattner, Jochen
Zhou, Dapeng
Savage, Paul B
Bendelac, Albert
Wilson, Ian A
Teyton, Luc
AI053725/AI/NIAID NIH HHS/United States
R01 CA058896/CA/NCI NIH HHS/United States
P01 AI053725/AI/NIAID NIH HHS/United States
CA58896/CA/NCI NIH HHS/United States
P01 AI053725-010002/AI/NIAID NIH HHS/United States
GM62116/GM/NIGMS NIH HHS/United States
U54 GM062116/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Nat Immunol. 2005 Aug;6(8):810-8. doi: 10.1038/ni1224. Epub 2005 Jul 10.
Author Address: Department of Molecular Biology and The Scripps Research Institute, La Jolla, California 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 977
Author: Zajonc, D. M., Crispin, M. D., Bowden, T. A., Young, D. C., Cheng, T. Y., Hu, J., Costello, C. E., Rudd, P. M., Dwek, R. A., Miller, M. J., Brenner, M. B., Moody, D. B. and Wilson, I. A.
Year: 2005
Title: Molecular mechanism of lipopeptide presentation by CD1a
Journal: Immunity
Volume: 22
Issue: 2
Pages: 209-19
Epub Date: 2005/02/23
Date: Feb
Short Title: Molecular mechanism of lipopeptide presentation by CD1a
Alternate Journal: Immunity
ISSN: 1074-7613 (Print)
1074-7613
DOI: 10.1016/j.immuni.2004.12.009
Accession Number: 15723809
Keywords: Antigen Presentation/*immunology
Antigens, CD1/chemistry/*immunology/metabolism
Cells, Cultured
Crystallization
Crystallography, X-Ray
Humans
Hydrogen Bonding
Ligands
Lipoproteins/chemical synthesis/chemistry/*immunology
Lymphocyte Activation/drug effects/immunology
Models, Molecular
Oxazoles/chemical synthesis/chemistry/immunology
Peptides/chemical synthesis/chemistry/*immunology
Protein Binding
Protein Conformation
Receptors, Antigen, T-Cell/chemistry/immunology/metabolism
Substrate Specificity
Sulfoglycosphingolipids/chemistry
T-Lymphocytes/chemistry/drug effects/immunology/metabolism
Abstract: CD1a is expressed on Langerhans cells (LCs) and dendritic cells (DCs), where it mediates T cell recognition of glycolipid and lipopeptide antigens that contain either one or two alkyl chains. We demonstrate here that CD1a-restricted T cells can discriminate the peptide component of didehydroxymycobactin lipopeptides. Structure analysis of CD1a cocrystallized with a synthetic mycobactin lipopeptide at 2.8 A resolution further reveals that the single alkyl chain is inserted deep within the A' pocket of the groove, whereas its two peptidic branches protrude along the F' pocket to the outer, alpha-helical surface of CD1a for recognition by the TCR. Remarkably, the cyclized lysine branch of the peptide moiety lies in the shallow F' pocket in a conformation that closely mimics that of the alkyl chain in the CD1a-sulfatide structure. Thus, this structural study illustrates how a single chain lipid can be presented by CD1 and that the peptide moiety of the lipopeptide is recognized by the TCR.
Notes: Zajonc, Dirk M
Crispin, M D Max
Bowden, Thomas A
Young, David C
Cheng, Tan-Yun
Hu, Jingdan
Costello, Catherine E
Rudd, Pauline M
Dwek, Raymond A
Miller, Marvin J
Brenner, Michael B
Moody, D Branch
Wilson, Ian A
AI054193/AI/NIAID NIH HHS/United States
AI28973/AI/NIAID NIH HHS/United States
AI30988/AI/NIAID NIH HHS/United States
AI49313/AI/NIAID NIH HHS/United States
AR48632/AR/NIAMS NIH HHS/United States
CA58896/CA/NCI NIH HHS/United States
GM62116/GM/NIGMS NIH HHS/United States
RR10888/RR/NCRR NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Immunity. 2005 Feb;22(2):209-19. doi: 10.1016/j.immuni.2004.12.009.
Author Address: Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 890
Author: Zajonc, D. M., Elsliger, M. A., Teyton, L. and Wilson, I. A.
Year: 2003
Title: Crystal structure of CD1a in complex with a sulfatide self antigen at a resolution of 2.15 A
Journal: Nat Immunol
Volume: 4
Issue: 8
Pages: 808-15
Epub Date: 2003/07/02
Date: Aug
Short Title: Crystal structure of CD1a in complex with a sulfatide self antigen at a resolution of 2.15 A
Alternate Journal: Nature immunology
ISSN: 1529-2908 (Print)
1529-2908
DOI: 10.1038/ni948
Accession Number: 12833155
Keywords: Antigens, CD1/*chemistry/metabolism
Autoantigens/*chemistry/metabolism
Binding Sites
Crystallography
Humans
Sulfoglycosphingolipids/*chemistry/metabolism
Abstract: CD1 antigens bind a variety of self and foreign lipid and glycolipid antigens for presentation to CD1-restricted T cell receptors (TCRs). Here we report the crystal structure of human CD1a in complex with a sulfatide self antigen at a resolution of 2.15 A. The lipid adopts an S-shaped conformation, with the sphingosine chain completely buried in the A' pocket and the fatty acid chain emerging from the interface of the A' pocket into the more exposed F' pocket. The headgroup is anchored in the A'-F' junction and protrudes into the F' pocket for TCR recognition. Because the A' pocket is narrow with a fixed terminus, it can act as a molecular 'ruler' to select alkyl chains of a particular length.
Notes: Zajonc, Dirk M
Elsliger, Marc A
Teyton, Luc
Wilson, Ian A
AI53725/AI/NIAID NIH HHS/United States
CA58896/CA/NCI NIH HHS/United States
GM62116/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
Nat Immunol. 2003 Aug;4(8):808-15. doi: 10.1038/ni948. Epub 2003 Jun 29.
Author Address: Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 721
Author: Zamze, S., Harvey, D. J., Pesheva, P., Mattu, T. S., Schachner, M., Dwek, R. A. and Wing, D. R.
Year: 1999
Title: Glycosylation of a CNS-specific extracellular matrix glycoprotein, tenascin-R, is dominated by O-linked sialylated glycans and "brain-type" neutral N-glycans
Journal: Glycobiology
Volume: 9
Issue: 8
Pages: 823-31
Epub Date: 1999/07/16
Date: Aug
Short Title: Glycosylation of a CNS-specific extracellular matrix glycoprotein, tenascin-R, is dominated by O-linked sialylated glycans and "brain-type" neutral N-glycans
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/9.8.823
Accession Number: 10406848
Keywords: Animals
*Brain Chemistry
Carbohydrate Conformation
Carbohydrate Sequence
Chromatography, Gel
Chromatography, High Pressure Liquid
Extracellular Matrix Proteins/*chemistry/isolation & purification/metabolism
Glycoside Hydrolases
Glycosylation
Mice
Molecular Sequence Data
Oligosaccharides/*chemistry/isolation & purification
Polysaccharides/*chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Tenascin/*chemistry/isolation & purification/metabolism
Abstract: As a member of the tenascin family of extracellular matrix glycoproteins, tenascin-R is located exclusively in the CNS. It is believed to play a role in myelination and axonal stabilization and, through repulsive properties, may contribute to the lack of regeneration of CNS axons following damage. The contrary functions of the tenascins have been localized to the different structural domains of the protein. However, little is known concerning the influence of the carbohydrate conjugated to the many potential sites for N - and O -glycosylation (10-20% by weight). As a first analytical requirement, we show that >80% of the N -glycans in tenascin-R are neutral and dominated by complex biantennary structures. These display the "brain-type" characteristics of outer-arm- and core-fucosylation, a bisecting N -acetylglucosamine and, significantly, an abundance of antennae truncation. In some structures, truncation resulted in only a single mannose residue remaining on the 3-arm, a particularly unusual consequence of the N -glycan processing pathway. In contrast to brain tissue, hybrid and oligomannosidic N -glycans were either absent or in low abundance. A high relative abundance of O -linked sialylated glycans was found. This was associated with a significant potential for O -linked glycosylation sites and multivalent display of the sialic acid residues. These O -glycans were dominated by the disialylated structure, NeuAcalpha2-3Galbeta1-3(NeuAcalpha2-6)GalNAc. The possibility that these O -glycans enable tenascin-R to interact in the CNS either with the myelin associated glycoprotein or with sialoadhesin on activated microglia is discussed.
Notes: Zamze, S
Harvey, D J
Pesheva, P
Mattu, T S
Schachner, M
Dwek, R A
Wing, D R
Wellcome Trust/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
England
Glycobiology. 1999 Aug;9(8):823-31. doi: 10.1093/glycob/9.8.823.
Author Address: Glycobiology Institute, Department of Biochemistry, South Parks Road, Oxford OX1 3QU, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1393
Author: Zaro, B. W., Yang, Y. Y., Hang, H. C. and Pratt, M. R.
Year: 2011
Title: Chemical reporters for fluorescent detection and identification of O-GlcNAc-modified proteins reveal glycosylation of the ubiquitin ligase NEDD4-1
Journal: Proc Natl Acad Sci U S A
Volume: 108
Issue: 20
Pages: 8146-51
Epub Date: 2011/05/05
Date: May 17
Short Title: Chemical reporters for fluorescent detection and identification of O-GlcNAc-modified proteins reveal glycosylation of the ubiquitin ligase NEDD4-1
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1102458108
PMCID: PMC3100932
Accession Number: 21540332
Keywords: 3T3 Cells
Animals
Endosomal Sorting Complexes Required for Transport/*metabolism
Fluorescent Dyes/analysis
Glycosylation
Mice
Nedd4 Ubiquitin Protein Ligases
*Protein Processing, Post-Translational
Proteins/analysis/*metabolism
Proteomics/*methods
Ubiquitin-Protein Ligases/*metabolism
Abstract: The dynamic modification of nuclear and cytoplasmic proteins by the monosaccharide N-acetyl-glucosamine (GlcNAc) continues to emerge as an important regulator of many biological processes. Herein we describe the development of an alkynyl-modified GlcNAc analog (GlcNAlk) as a new chemical reporter of O-GlcNAc modification in living cells. This strategy is based on metabolic incorporation of reactive functionality into the GlcNAc biosynthetic pathway. When combined with the Cu(I)-catalyzed [3 + 2] azide-alkyne cycloaddition, this chemical reporter allowed for the robust in-gel fluorescent visualization of O-GlcNAc and affinity enrichment and identification of O-GlcNAc-modified proteins. Using in-gel fluorescence detection, we characterized the metabolic fates of GlcNAlk and the previously reported azido analog, GlcNAz. We confirmed previous results that GlcNAz can be metabolically interconverted to GalNAz, whereas GlcNAlk does not, thereby yielding a more specific metabolic reporter of O-GlcNAc modification. We also used GlcNAlk, in combination with a biotin affinity tag, to identify 374 proteins, 279 of which were not previously reported, and we subsequently confirmed the enrichment of three previously uncharacterized proteins. Finally we confirmed the O-GlcNAc modification of the ubiquitin ligase NEDD4-1, the first reported glycosylation of this protein.
Notes: 1091-6490
Zaro, Balyn W
Yang, Yu-Ying
Hang, Howard C
Pratt, Matthew R
R01 GM087544/GM/NIGMS NIH HHS/United States
1R01GM087544 O1A2/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2011 May 17;108(20):8146-51. doi: 10.1073/pnas.1102458108. Epub 2011 May 3.
Author Address: Department of Chemistry, University of Southern California, Los Angeles, CA 90089, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1524
Author: Zauner, G., Hoffmann, M., Rapp, E., Koeleman, C. A., Dragan, I., Deelder, A. M., Wuhrer, M. and Hensbergen, P. J.
Year: 2012
Title: Glycoproteomic analysis of human fibrinogen reveals novel regions of O-glycosylation
Journal: J Proteome Res
Volume: 11
Issue: 12
Pages: 5804-14
Epub Date: 2012/10/12
Date: Dec 7
Short Title: Glycoproteomic analysis of human fibrinogen reveals novel regions of O-glycosylation
Alternate Journal: Journal of proteome research
ISSN: 1535-3893
DOI: 10.1021/pr3005937
Accession Number: 23050552
Keywords: Amino Acid Sequence
Binding Sites
Chromatography, Liquid/methods
Electrophoresis, Polyacrylamide Gel
Endopeptidase K/chemistry
Fibrinogen/*analysis/chemistry
Glycopeptides/*analysis/chemistry
Glycosylation
Humans
Molecular Sequence Data
Nanotechnology
Oligosaccharides/analysis/chemistry
Polysaccharides/chemistry
Proteolysis
Proteome/*analysis/chemistry
Proteomics/methods
Sequence Analysis, Protein
Spectrometry, Mass, Electrospray Ionization/*methods
Trypsin/chemistry
Abstract: Human fibrinogen is a 340 kDa, soluble plasma glycoprotein composed of paired sets of three subunits (α, β, γ). The protein plays a crucial role in protecting the vascular network against the loss of blood after tissue injury. The beta and gamma subunits each contain one N-glycosylation site, each of which is occupied by a biantennary N-glycan. So far O-linked oligosaccharides have rarely been described. Here, we make use of tryptic- and proteinase K-generated fibrinogen glycopeptides for the detailed analysis of the protein's O-glycosylation by combining information obtained from both one- and two-dimensional nanoLC-ESI-ion trap (IT)-MS approaches. Glycopeptides were analyzed by ion trap-MS/MS which displayed fragmentations of glycosidic linkages and some peptide backbone cleavages. MS3 spectra of the generated O-glycopeptides showed cleavages of the peptide backbone and provided essential information on the peptide sequence. The previously reported N-glycan attachment sites of human fibrinogen could be confirmed. Moreover, we describe seven novel O-glycosylation regions in human fibrinogen, all occupied by a monosialylated T-antigen. Our findings may help to improve the general understanding of human fibrinogen in the blood clotting process.
Notes: 1535-3907
Zauner, Gerhild
Hoffmann, Marcus
Rapp, Erdmann
Koeleman, Carolien A M
Dragan, Irina
Deelder, André M
Wuhrer, Manfred
Hensbergen, Paul J
Journal Article
Research Support, Non-U.S. Gov't
United States
J Proteome Res. 2012 Dec 7;11(12):5804-14. doi: 10.1021/pr3005937. Epub 2012 Oct 29.
Author Address: Department of Parasitology, Biomolecular Mass Spectrometry Unit, Leiden University Medical Center, Leiden, The Netherlands. gzauner@lumc.nl
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1296
Author: Zavialov, A. V., Yu, X., Spillmann, D., Lauvau, G. and Zavialov, A. V.
Year: 2010
Title: Structural basis for the growth factor activity of human adenosine deaminase ADA2
Journal: J Biol Chem
Volume: 285
Issue: 16
Pages: 12367-77
Epub Date: 2010/02/12
Date: Apr 16
Short Title: Structural basis for the growth factor activity of human adenosine deaminase ADA2
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M109.083527
PMCID: PMC2852975
Accession Number: 20147294
Keywords: Adaptor Proteins, Signal Transducing/*chemistry/genetics/*physiology
Adenosine Deaminase/*chemistry/genetics/physiology
Amino Acid Sequence
Amino Acid Substitution
Catalytic Domain/genetics
Coformycin/pharmacology
Crystallography, X-Ray
DNA-Binding Proteins
Enzyme Inhibitors/pharmacology
Growth Substances/chemistry/genetics/physiology
Humans
In Vitro Techniques
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Multimerization
Recombinant Proteins/chemistry/genetics/metabolism
Sequence Deletion
Sequence Homology, Amino Acid
Signal Transduction
Static Electricity
Thermodynamics
Transcription Factors/*chemistry/genetics/*physiology
Abstract: Two distinct adenosine deaminases, ADA1 and ADA2, are found in humans. ADA1 has an important role in lymphocyte function and inherited mutations in ADA1 result in severe combined immunodeficiency. The recently isolated ADA2 belongs to the novel family of adenosine deaminase growth factors (ADGFs), which play an important role in tissue development. The crystal structures of ADA2 and ADA2 bound to a transition state analogue presented here reveal the structural basis of the catalytic/signaling activity of ADGF/ADA2 proteins. In addition to the catalytic domain, the structures discovered two ADGF/ADA2-specific domains of novel folds that mediate the protein dimerization and binding to the cell surface receptors. This complex architecture is in sharp contrast with that of monomeric single domain ADA1. An extensive glycosylation and the presence of a conserved disulfide bond and a signal peptide in ADA2 strongly suggest that ADA2, in contrast to ADA1, is specifically designed to act in the extracellular environment. The comparison of catalytic sites of ADA2 and ADA1 demonstrates large differences in the arrangement of the substrate-binding pockets. These structural differences explain the substrate and inhibitor specificity of adenosine deaminases and provide the basis for a rational design of ADA2-targeting drugs to modulate the immune system responses in pathophysiological conditions.
Notes: 1083-351x
Zavialov, Anton V
Yu, Xiaodi
Spillmann, Dorothe
Lauvau, Grégoire
Zavialov, Andrey V
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2010 Apr 16;285(16):12367-77. doi: 10.1074/jbc.M109.083527. Epub 2010 Feb 9.
Author Address: Department of Molecular Biology, Uppsala Biomedical Centre, Swedish University of Agricultural Sciences, Uppsala, Sweden.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1307
Author: Zeidan, Q., Wang, Z., De Maio, A. and Hart, G. W.
Year: 2010
Title: O-GlcNAc cycling enzymes associate with the translational machinery and modify core ribosomal proteins
Journal: Mol Biol Cell
Volume: 21
Issue: 12
Pages: 1922-36
Epub Date: 2010/04/23
Date: Jun 15
Short Title: O-GlcNAc cycling enzymes associate with the translational machinery and modify core ribosomal proteins
Alternate Journal: Molecular biology of the cell
ISSN: 1059-1524 (Print)
1059-1524
DOI: 10.1091/mbc.e09-11-0941
PMCID: PMC2883937
Accession Number: 20410138
Keywords: Acetylglucosamine/*metabolism
Adenoviridae/metabolism
Animals
Cell Line, Tumor
Cell Nucleolus/enzymology
Glycosylation
Humans
Mass Spectrometry
Mice
N-Acetylglucosaminyltransferases/*metabolism
Phosphorylation
*Protein Biosynthesis
*Protein Processing, Post-Translational
Rats
Ribosomal Proteins/*biosynthesis/chemistry
Ribosome Subunits, Large, Eukaryotic/metabolism
Ribosomes/enzymology
Abstract: Protein synthesis is globally regulated through posttranslational modifications of initiation and elongation factors. Recent high-throughput studies have identified translation factors and ribosomal proteins (RPs) as substrates for the O-GlcNAc modification. Here we determine the extent and abundance of O-GlcNAcylated proteins in translational preparations. O-GlcNAc is present on many proteins that form active polysomes. We identify twenty O-GlcNAcylated core RPs, of which eight are newly reported. We map sites of O-GlcNAc modification on four RPs (L6, L29, L32, and L36). RPS6, a component of the mammalian target of rapamycin (mTOR) signaling pathway, follows different dynamics of O-GlcNAcylation than nutrient-induced phosphorylation. We also show that both O-GlcNAc cycling enzymes OGT and OGAse strongly associate with cytosolic ribosomes. Immunofluorescence experiments demonstrate that OGAse is present uniformly throughout the nucleus, whereas OGT is excluded from the nucleolus. Moreover, nucleolar stress only alters OGAse nuclear staining, but not OGT staining. Lastly, adenovirus-mediated overexpression of OGT, but not of OGAse or GFP control, causes an accumulation of 60S subunits and 80S monosomes. Our results not only establish that O-GlcNAcylation extensively modifies RPs, but also suggest that O-GlcNAc play important roles in regulating translation and ribosome biogenesis.
Notes: 1939-4586
Zeidan, Quira
Wang, Zihao
De Maio, Antonio
Hart, Gerald W
P30 DK079637/DK/NIDDK NIH HHS/United States
Journal Article
Mol Biol Cell. 2010 Jun 15;21(12):1922-36. doi: 10.1091/mbc.e09-11-0941. Epub 2010 Apr 21.
Author Address: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, MD 21205-2185, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1248
Author: Zelcer, N., Hong, C., Boyadjian, R. and Tontonoz, P.
Year: 2009
Title: LXR regulates cholesterol uptake through Idol-dependent ubiquitination of the LDL receptor
Journal: Science
Volume: 325
Issue: 5936
Pages: 100-4
Epub Date: 2009/06/13
Date: Jul 3
Short Title: LXR regulates cholesterol uptake through Idol-dependent ubiquitination of the LDL receptor
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075 (Print)
0036-8075
DOI: 10.1126/science.1168974
PMCID: PMC2777523
NIHMSID: NIHMS151722
Accession Number: 19520913
Keywords: Animals
Cell Line, Tumor
Cholesterol/*metabolism
DNA-Binding Proteins/agonists/*metabolism
Homeostasis
Humans
Ligands
Lipoproteins, LDL/blood/metabolism
Liver/metabolism
Liver X Receptors
Mice
Mice, Inbred C57BL
Orphan Nuclear Receptors
Promoter Regions, Genetic
RNA, Messenger/genetics/metabolism
Receptors, Cytoplasmic and Nuclear/agonists/*metabolism
Receptors, LDL/genetics/*metabolism
Transcription, Genetic
Ubiquitin-Protein Ligases
Ubiquitination
Abstract: Cellular cholesterol levels reflect a balance between uptake, efflux, and endogenous synthesis. Here we show that the sterol-responsive nuclear liver X receptor (LXR) helps maintain cholesterol homeostasis, not only through promotion of cholesterol efflux but also through suppression of low-density lipoprotein (LDL) uptake. LXR inhibits the LDL receptor (LDLR) pathway through transcriptional induction of Idol (inducible degrader of the LDLR), an E3 ubiquitin ligase that triggers ubiquitination of the LDLR on its cytoplasmic domain, thereby targeting it for degradation. LXR ligand reduces, whereas LXR knockout increases, LDLR protein levels in vivo in a tissue-selective manner. Idol knockdown in hepatocytes increases LDLR protein levels and promotes LDL uptake. Conversely, adenovirus-mediated expression of Idol in mouse liver promotes LDLR degradation and elevates plasma LDL levels. The LXR-Idol-LDLR axis defines a complementary pathway to sterol response element-binding proteins for sterol regulation of cholesterol uptake.
Notes: 1095-9203
Zelcer, Noam
Hong, Cynthia
Boyadjian, Rima
Tontonoz, Peter
P01 HL090553-01A10003/HL/NHLBI NIH HHS/United States
HL090553/HL/NHLBI NIH HHS/United States
R01 HL066088-09/HL/NHLBI NIH HHS/United States
HL066088/HL/NHLBI NIH HHS/United States
HL030568/HL/NHLBI NIH HHS/United States
P01 HL090553/HL/NHLBI NIH HHS/United States
Howard Hughes Medical Institute/United States
R01 HL066088/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Science. 2009 Jul 3;325(5936):100-4. doi: 10.1126/science.1168974. Epub 2009 Jun 11.
Author Address: Howard Hughes Medical Institute and Department of Pathology and Laboratory Medicine, University of California, Los Angeles (UCLA), Los Angeles, CA 90095, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 722
Author: Zelenski, N. G., Rawson, R. B., Brown, M. S. and Goldstein, J. L.
Year: 1999
Title: Membrane topology of S2P, a protein required for intramembranous cleavage of sterol regulatory element-binding proteins
Journal: J Biol Chem
Volume: 274
Issue: 31
Pages: 21973-80
Epub Date: 1999/07/27
Date: Jul 30
Short Title: Membrane topology of S2P, a protein required for intramembranous cleavage of sterol regulatory element-binding proteins
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.274.31.21973
Accession Number: 10419520
Keywords: Amino Acid Sequence
Animals
*CCAAT-Enhancer-Binding Proteins
CHO Cells
Cell Line
Cell Membrane/metabolism/ultrastructure
Cricetinae
DNA-Binding Proteins/chemistry/genetics/*metabolism
Glycosylation
Humans
Kidney
Membrane Proteins/*chemistry/genetics/*metabolism
Metalloendopeptidases/*chemistry/*metabolism
Molecular Sequence Data
Mutagenesis, Insertional
Nuclear Proteins/chemistry/genetics/*metabolism
Protein Conformation
Recombinant Fusion Proteins/biosynthesis
Sterol Regulatory Element Binding Protein 1
Transcription Factors/metabolism
Transfection
Abstract: In sterol-depleted mammalian cells, a two-step proteolytic process releases the NH(2)-terminal domains of sterol regulatory element-binding proteins (SREBPs) from membranes of the endoplasmic reticulum (ER). These domains translocate into the nucleus, where they activate genes of cholesterol and fatty acid biosynthesis. The SREBPs are oriented in the membrane in a hairpin fashion, with the NH(2)- and COOH-terminal domains facing the cytosol and a single hydrophilic loop projecting into the lumen. The first cleavage occurs at Site-1 within the ER lumen to generate an intermediate that is subsequently released from the membrane by cleavage at Site-2, which lies within the first transmembrane domain. A membrane protein, designated S2P, a putative zinc metalloprotease, is required for this cleavage. Here, we use protease protection and glycosylation site mapping to define the topology of S2P in ER membranes. Both the NH(2) and COOH termini of S2P face the cytosol. Most of S2P is hydrophobic and appears to be buried in the membrane. All three of the long hydrophilic sequences of S2P can be glycosylated, indicating that they all project into the lumen. The HEIGH sequence of S2P, which contains two potential zinc-coordinating residues, is contained within a long hydrophobic segment. Aspartic acid 467, located approximately 300 residues away from the HEIGH sequence, appears to provide the third coordinating residue for the active site zinc. This residue, too, is located in a hydrophobic sequence. The hydrophobicity of these sequences suggests that the active site of S2P is located within the membrane in an ideal position to cleave its target, a Leu-Cys bond in the first transmembrane helix of SREBPs.
Notes: Zelenski, N G
Rawson, R B
Brown, M S
Goldstein, J L
HL20948/HL/NHLBI NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
United States
J Biol Chem. 1999 Jul 30;274(31):21973-80. doi: 10.1074/jbc.274.31.21973.
Author Address: Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, Texas 75235, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2020
Author: Zhang, C., Xie, F., Li, L., Zhang, C., Zhang, Y., Ying, W., Liu, L., Yan, X., Yin, F. and Zhang, L.
Year: 2019
Title: Hepatocyte nuclear factor 1 alpha (HNF1A) regulates transcription of O-GlcNAc transferase in a negative feedback mechanism
Journal: FEBS Lett
Volume: 593
Issue: 10
Pages: 1050-1060
Epub Date: 2019/04/07
Date: May
Short Title: Hepatocyte nuclear factor 1 alpha (HNF1A) regulates transcription of O-GlcNAc transferase in a negative feedback mechanism
Alternate Journal: FEBS letters
ISSN: 0014-5793
DOI: 10.1002/1873-3468.13381
Accession Number: 30953348
Keywords: Acylation
*Feedback, Physiological
*Gene Expression Regulation
HEK293 Cells
Hepatocyte Nuclear Factor 1-alpha/*metabolism
Humans
N-Acetylglucosaminyltransferases/*genetics/metabolism
*Protein Processing, Post-Translational
Substrate Specificity
Tandem Mass Spectrometry
Transcription, Genetic
* Ogt
*Hnf1a
*negative feedback mechanism
*transcriptional regulation
Abstract: O-GlcNAc transferase (OGT)-catalyzed protein O-GlcNAcylation is implicated in diverse cellular events. In the present study, we report the regulation of ogt transcription by the hepatocyte nuclear factor 1 homologue A (HNF1A) in HEK293T cells. We first identified a core ogt promoter (-150 to +200 bp) and confirmed its binding to the transcription factor HNF1A. We found that HNF1A regulates ogt transcription in a time-dependent manner and that O-GlcNAcylation of HNF1A represses ogt transcription. Electron-transfer dissociation based tandem mass spectrometry analysis revealed 14 O-GlcNAc sites on HNF1A, six of which are predominantly modified, including Ser(303/304) , Ser(471) , Ser(560) and Thr(563/564) . We further found that loss of O-GlcNAcylation at Ser(303/304) or Thr(563/564) significantly elevates ogt transcription. These findings highlight a negative feedback mechanism for ogt transcription, which partially explains the homeostasis of cellular O-GlcNAcylation.
Notes: 1873-3468
Zhang, Chuanhui
Xie, Fei
Li, Ling
Zhang, Cheng
Zhang, Yong
Ying, Wantao
Liu, Li
Yan, Xuli
Yin, Futao
Zhang, Lianwen
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2019 May;593(10):1050-1060. doi: 10.1002/1873-3468.13381. Epub 2019 May 14.
Author Address: College of Pharmacy and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin, China.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, China.
Key Lab of Transplant Engineering and Immunology, MOH, West China-Washington Mitochondria and Metabolism Research Center, West China Hospital, Sichuan University, Chengdu, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 952
Author: Zhang, E. Y., Phelps, M. A., Banerjee, A., Khantwal, C. M., Chang, C., Helsper, F. and Swaan, P. W.
Year: 2004
Title: Topology scanning and putative three-dimensional structure of the extracellular binding domains of the apical sodium-dependent bile acid transporter (SLC10A2)
Journal: Biochemistry
Volume: 43
Issue: 36
Pages: 11380-92
Epub Date: 2004/09/08
Date: Sep 14
Short Title: Topology scanning and putative three-dimensional structure of the extracellular binding domains of the apical sodium-dependent bile acid transporter (SLC10A2)
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi049270a
Accession Number: 15350125
Keywords: Amino Acid Sequence
Binding Sites/genetics
Carbohydrate Conformation
Computer Simulation
Consensus Sequence
Extracellular Space/*chemistry/genetics/*metabolism
Glutamic Acid/genetics
Glycosylation
Humans
Ligands
Models, Chemical
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Organic Anion Transporters, Sodium-Dependent/*chemistry/genetics/*metabolism
Peptide Fragments/chemistry/genetics/metabolism
Polymorphism, Genetic
Proline/genetics
Protein Binding/genetics
Protein Folding
Protein Structure, Tertiary/genetics
Sequence Homology, Amino Acid
Serine/genetics
Symporters/*chemistry/genetics/*metabolism
Abstract: The apical sodium-dependent bile acid transporter (ASBT, SLC10A2) facilitates the enterohepatic circulation of bile salts and plays a key role in cholesterol metabolism. The membrane topology of ASBT was initially scanned using a consensus topography analysis that predominantly predicts a seven transmembrane (TM) domain configuration adhering to the "positive inside" rule. Membrane topology was further evaluated and confirmed by N-glycosylation-scanning mutagenesis, as reporter sites inserted in the putative extracellular loops 1 and 3 were glycosylated. On the basis of a 7TM topology, we built a three-dimensional model of ASBT using an approach of homology-modeling and remote-threading techniques for the extramembranous domains using bacteriorhodopsin as a scaffold for membrane attachment points; the model was refined using energy minimizations and molecular dynamics simulations. Ramachandran scores and other geometric indicators show that the model is comparable in quality to the crystal structures of similar proteins. Simulated annealing and docking of cholic acid, a natural substrate, onto the protein surface revealed four distinct binding sites. Subsequent site-directed mutagenesis of the predicted binding domain further validated the model. This model agrees further with available data for a pathological mutation (P290S) because the mutant model after in silico mutagenesis loses the ability to bind bile acids.
Notes: Zhang, Eric Y
Phelps, Mitch A
Banerjee, Antara
Khantwal, Chandra M
Chang, Cheng
Helsper, Freek
Swaan, Peter W
DK061425/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Validation Study
United States
Biochemistry. 2004 Sep 14;43(36):11380-92. doi: 10.1021/bi049270a.
Author Address: Department of Pharmaceutical Sciences, University of Maryland, 20 Penn Street, Baltimore, Maryland 21201, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1183
Author: Zhang, H., Casasnovas, J. M., Jin, M., Liu, J. H., Gahmberg, C. G., Springer, T. A. and Wang, J. H.
Year: 2008
Title: An unusual allosteric mobility of the C-terminal helix of a high-affinity alphaL integrin I domain variant bound to ICAM-5
Journal: Mol Cell
Volume: 31
Issue: 3
Pages: 432-7
Epub Date: 2008/08/12
Date: Aug 8
Short Title: An unusual allosteric mobility of the C-terminal helix of a high-affinity alphaL integrin I domain variant bound to ICAM-5
Alternate Journal: Molecular cell
ISSN: 1097-2765 (Print)
1097-2765
DOI: 10.1016/j.molcel.2008.06.022
PMCID: PMC2603608
NIHMSID: NIHMS65487
Accession Number: 18691975
Keywords: Allosteric Regulation
Animals
CD11a Antigen/*chemistry/*metabolism
CHO Cells
Cell Adhesion Molecules/chemistry/*metabolism
Cricetinae
Cricetulus
Crystallography, X-Ray
Models, Molecular
Mutant Proteins/*chemistry/*metabolism
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Abstract: Integrins are cell surface receptors that transduce signals bidirectionally across the plasma membrane. The key event of integrin signaling is the allosteric regulation between its ligand-binding site and the C-terminal helix (alpha7) of integrin's inserted (I) domain. A significant axial movement of the alpha7 helix is associated with the open, active conformation of integrins. We describe the crystal structure of an engineered high-affinity I domain from the integrin alpha(L)beta(2) (LFA-1) alpha subunit in complex with the N-terminal two domains of ICAM-5, an adhesion molecule expressed in telencephalic neurons. The finding that the alpha7 helix swings out and inserts into a neighboring I domain in an upside-down orientation in the crystals implies an intrinsically unusual mobility of this helix. This remarkable feature allows the alpha7 helix to trigger integrin's large-scale conformational changes with little energy penalty. It serves as a mechanistic example of how a weakly bound adhesion molecule works in signaling.
Notes: 1097-4164
Zhang, Hongmin
Casasnovas, Jose M
Jin, Moonsoo
Liu, Jin-huan
Gahmberg, Carl G
Springer, Timothy A
Wang, Jia-huai
P01 HL048675/HL/NHLBI NIH HHS/United States
P01 HL048675-08A10003/HL/NHLBI NIH HHS/United States
R37 CA031798/CA/NCI NIH HHS/United States
R37 CA031798-29/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Mol Cell. 2008 Aug 8;31(3):432-7. doi: 10.1016/j.molcel.2008.06.022.
Author Address: Department of Medical Oncology and Department of Cancer Biology, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 882
Author: Zhang, H., Li, X. J., Martin, D. B. and Aebersold, R.
Year: 2003
Title: Identification and quantification of N-linked glycoproteins using hydrazide chemistry, stable isotope labeling and mass spectrometry
Journal: Nat Biotechnol
Volume: 21
Issue: 6
Pages: 660-6
Epub Date: 2003/05/20
Date: Jun
Short Title: Identification and quantification of N-linked glycoproteins using hydrazide chemistry, stable isotope labeling and mass spectrometry
Alternate Journal: Nature biotechnology
ISSN: 1087-0156 (Print)
1087-0156
DOI: 10.1038/nbt827
Accession Number: 12754519
Keywords: Amino Acid Sequence
Blood Proteins/analysis/chemistry/isolation & purification
Chromatography, High Pressure Liquid/*methods
Glycoproteins/*analysis/chemistry/*isolation & purification
Isotope Labeling/*methods
Mass Spectrometry/*methods
Molecular Sequence Data
Oxidation-Reduction
Peptides/analysis/chemistry/metabolism
Polysaccharides/analysis/chemistry/metabolism
Resins, Synthetic
alpha 1-Antichymotrypsin/analysis/chemistry/isolation & purification
alpha 1-Antitrypsin/analysis/chemistry/isolation & purification
alpha-2-HS-Glycoprotein
Abstract: Quantitative proteome profiling using stable isotope protein tagging and automated tandem mass spectrometry (MS/MS) is an emerging technology with great potential for the functional analysis of biological systems and for the detection of clinical diagnostic or prognostic marker proteins. Owing to the enormous complexity of proteomes, their comprehensive analysis is an as-yet-unresolved technical challenge. However, biologically or clinically important information can be obtained if specific, information-rich protein classes, or sub-proteomes, are isolated and analyzed. Glycosylation is the most common post-translational modification. Here we describe a method for the selective isolation, identification and quantification of peptides that contain N-linked carbohydrates. It is based on the conjugation of glycoproteins to a solid support using hydrazide chemistry, stable isotope labeling of glycopeptides and the specific release of formerly N-linked glycosylated peptides via peptide- N-glycosidase F (PNGase F). The recovered peptides are then identified and quantified by MS/MS. We applied the approach to the analysis of plasma membrane proteins and proteins contained in human blood serum.
Notes: Zhang, Hui
Li, Xiao-Jun
Martin, Daniel B
Aebersold, Ruedi
K08CA97282-01/CA/NCI NIH HHS/United States
R33 CA93302/CA/NCI NIH HHS/United States
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.
Validation Study
United States
Nat Biotechnol. 2003 Jun;21(6):660-6. doi: 10.1038/nbt827. Epub 2003 May 18.
Author Address: Institute for Systems Biology, 1441 N 34th Street, Seattle, Washington 98103-8904, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1854
Author: Zhang, H., Qiao, A., Yang, D., Yang, L., Dai, A., de Graaf, C., Reedtz-Runge, S., Dharmarajan, V., Zhang, H., Han, G. W., Grant, T. D., Sierra, R. G., Weierstall, U., Nelson, G., Liu, W., Wu, Y., Ma, L., Cai, X., Lin, G., Wu, X., Geng, Z., Dong, Y., Song, G., Griffin, P. R., Lau, J., Cherezov, V., Yang, H., Hanson, M. A., Stevens, R. C., Zhao, Q., Jiang, H., Wang, M. W. and Wu, B.
Year: 2017
Title: Structure of the full-length glucagon class B G-protein-coupled receptor
Journal: Nature
Volume: 546
Issue: 7657
Pages: 259-264
Epub Date: 2017/05/18
Date: Jun 8
Short Title: Structure of the full-length glucagon class B G-protein-coupled receptor
Alternate Journal: Nature
ISSN: 0028-0836 (Print)
0028-0836
DOI: 10.1038/nature22363
PMCID: PMC5492955
NIHMSID: NIHMS868095
Accession Number: 28514451
Keywords: Allosteric Site/drug effects
Benzamides/chemistry/metabolism/pharmacology
Cell Membrane/metabolism
Cross-Linking Reagents/chemistry
Crystallography, X-Ray
Deuterium Exchange Measurement
Disulfides/chemistry
Humans
Ligands
Models, Molecular
Molecular Dynamics Simulation
Phenylurea Compounds/chemistry/metabolism/pharmacology
Protein Domains
Protein Stability
Receptors, Glucagon/agonists/*chemistry/*classification/metabolism
Abstract: The human glucagon receptor, GCGR, belongs to the class B G-protein-coupled receptor family and plays a key role in glucose homeostasis and the pathophysiology of type 2 diabetes. Here we report the 3.0 Å crystal structure of full-length GCGR containing both the extracellular domain and transmembrane domain in an inactive conformation. The two domains are connected by a 12-residue segment termed the stalk, which adopts a β-strand conformation, instead of forming an α-helix as observed in the previously solved structure of the GCGR transmembrane domain. The first extracellular loop exhibits a β-hairpin conformation and interacts with the stalk to form a compact β-sheet structure. Hydrogen-deuterium exchange, disulfide crosslinking and molecular dynamics studies suggest that the stalk and the first extracellular loop have critical roles in modulating peptide ligand binding and receptor activation. These insights into the full-length GCGR structure deepen our understanding of the signalling mechanisms of class B G-protein-coupled receptors.
Notes: 1476-4687
Zhang, Haonan
Qiao, Anna
Yang, Dehua
Yang, Linlin
Dai, Antao
de Graaf, Chris
Reedtz-Runge, Steffen
Dharmarajan, Venkatasubramanian
Zhang, Hui
Han, Gye Won
Grant, Thomas D
Sierra, Raymond G
Weierstall, Uwe
Nelson, Garrett
Liu, Wei
Wu, Yanhong
Ma, Limin
Cai, Xiaoqing
Lin, Guangyao
Wu, Xiaoai
Geng, Zhi
Dong, Yuhui
Song, Gaojie
Griffin, Patrick R
Lau, Jesper
Cherezov, Vadim
Yang, Huaiyu
Hanson, Michael A
Stevens, Raymond C
Zhao, Qiang
Jiang, Hualiang
Wang, Ming-Wei
Wu, Beili
P41 GM103393/GM/NIGMS NIH HHS/United States
P41 RR001209/RR/NCRR NIH HHS/United States
R01 GM108635/GM/NIGMS NIH HHS/United States
R21 DA042298/DA/NIDA NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nature. 2017 Jun 8;546(7657):259-264. doi: 10.1038/nature22363. Epub 2017 May 17.
Author Address: CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zuchongzhi Road, Pudong, Shanghai 201203, China.
University of Chinese Academy of Sciences, No. 19A Yuquan Road, Beijing 100049, China.
The National Center for Drug Screening, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 189 Guo Shou Jing Road, Pudong, Shanghai 201203, China.
Department of Pharmacology, School of Basic Medical Sciences, Zhengzhou University, 100 Science Avenue, Zhengzhou 450001, China.
Division of Medicinal Chemistry, Faculty of Sciences, Amsterdam Institute for Molecules, Medicines and Systems (AIMMS), Vrije Universiteit Amsterdam, De Boelelaan 1108, Amsterdam 1081 HZ, The Netherlands.
Novo Nordisk A/S, Novo Nordisk Park, Måløv 2760, Denmark.
Department of Molecular Medicine, The Scripps Research Institute, 130 Scripps Way, Jupiter, Florida 33458, USA.
Department of Chemistry, Bridge Institute, University of Southern California, 3430 S. Vermont Avenue, Los Angeles, California 90089, USA.
Hauptman-Woodward Institute, SUNY at Buffalo, 700 Ellicott Street, Buffalo, New York 14203, USA.
Linac Coherent Light Source (LCLS), SLAC National Accelerator Laboratory, Menlo Park, California 94025, USA.
Department of Physics, Arizona State University, Tempe, Arizona 85287, USA.
Biodesign Center for Applied Structural Discovery, Biodesign Institute, School of Molecular Sciences, Arizona State University, Tempe, Arizona 85287, USA.
School of Life Science and Technology, ShanghaiTech University, 393 Hua Xia Zhong Road, Pudong, Shanghai 201210, China.
Novo Nordisk Research Centre China, No. 20 Life Science Park Road, Changping District, Beijing 102206, China.
Beijing Synchrotron Radiation Facility, Institute of High Energy Physics, Chinese Academy of Sciences, Beijing 100049, China.
iHuman Institute, ShanghaiTech University, 393 Hua Xia Zhong Road, Shanghai 201210, China.
Drug Discovery and Design Center, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zuchongzhi Road, Pudong, Shanghai 201203, China.
GPCR Consortium, San Marcos, California 92078, USA.
State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zuchongzhi Road, Pudong, Shanghai 201203, China.
CAS Center for Excellence in Biomacromolecules, Chinese Academy of Sciences, Beijing 100101, China.
School of Pharmacy, Fudan University, 826 Zhangheng Road, Shanghai 201203, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1869
Author: Zhang, J., Lei, H., Chen, Y., Ma, Y. T., Jiang, F., Tan, J., Zhang, Y. and Li, J. D.
Year: 2017
Title: Enzymatic O-GlcNAcylation of α-synuclein reduces aggregation and increases SDS-resistant soluble oligomers
Journal: Neurosci Lett
Volume: 655
Pages: 90-94
Epub Date: 2017/07/05
Date: Aug 10
Short Title: Enzymatic O-GlcNAcylation of α-synuclein reduces aggregation and increases SDS-resistant soluble oligomers
Alternate Journal: Neuroscience letters
ISSN: 0304-3940
DOI: 10.1016/j.neulet.2017.06.034
Accession Number: 28673834
Keywords: Benzothiazoles
Humans
N-Acetylglucosaminyltransferases/*chemistry
Protein Aggregates
Protein Multimerization
Protein Processing, Post-Translational
Sodium Dodecyl Sulfate/chemistry
Solubility
Thiazoles/chemistry
alpha-Synuclein/*chemistry
Aggregation
O-GlcNAcylation
SDS-resistant oligomers
α-Synuclein
Abstract: Neurodegenerative diseases including dementia with Lewy bodies, Lewy body variant of Alzheimer's disease, and Parkinson's disease are associated with the aberrant aggregation of α-synuclein, which is influenced by several post-translational modifications (PTMs). O-GlcNAcylation is one PTM that has an important role in many fundamental processes. The O-GlcNAcylation of endogenous α-synuclein at residues 53, 64, 72 and 87 has been reported in an unbiased mass spectrum analysis. The consequences of O-GlcNAcylation at residues 72 or 87 have been studied by using a synthetic α-synuclein bearing O-GlcNAcylation at threonine residue 72 or serine 87, respectively. O-GlcNAcylation at Thr72 or Ser87 suppresses the aggregation of α-synuclein. However, the effect of enzymatic O-GlcNAcylation of α-synuclein at multiple residues is not clear. Here, we successfully generated O-GlcNAcylated α-synuclein by co-expressing a shorter form of OGT (sOGT) with α-synuclein. The O-GlcNAcylation inhibited α-synuclein aggregation and promoted the formation of soluble SDS-resistant and Thioflavine T negative oligomers. Our data warrant further studies on the role of O-GlcNAcylation in the progression/treatment of Parkinson's disease in animal models.
Notes: 1872-7972
Zhang, Jiaming
Lei, Haozhi
Chen, Yubei
Ma, Yan-Tao
Jiang, Fang
Tan, Jieqiong
Zhang, Yi
Li, Jia-Da
Journal Article
Ireland
Neurosci Lett. 2017 Aug 10;655:90-94. doi: 10.1016/j.neulet.2017.06.034. Epub 2017 Jul 1.
Author Address: State Key Laboratory of Medical Genetics and School of Life Sciences, Central South University, Changsha, Hunan 410078, China.
Shanghai Institute of Appllied Physics, Chinese Academy of Sciences, Shanghai 201800, China.
Shanghai Institute of Appllied Physics, Chinese Academy of Sciences, Shanghai 201800, China. Electronic address: zhangyi@sinap.ac.cn.
State Key Laboratory of Medical Genetics and School of Life Sciences, Central South University, Changsha, Hunan 410078, China; Department of Pharmacology, Hubei University of Science and Technology, Xianning, Hubei 437100, China. Electronic address: lijiada@sklmg.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2023
Author: Zhang, N., Zhu, T., Yu, K., Shi, M., Wang, X., Wang, L., Huang, T., Li, W., Liu, Y. and Zhang, J.
Year: 2019
Title: Elevation of O-GlcNAc and GFAT expression by nicotine exposure promotes epithelial-mesenchymal transition and invasion in breast cancer cells
Journal: Cell Death Dis
Volume: 10
Issue: 5
Pages: 343
Epub Date: 2019/04/26
Date: Apr 24
Short Title: Elevation of O-GlcNAc and GFAT expression by nicotine exposure promotes epithelial-mesenchymal transition and invasion in breast cancer cells
Alternate Journal: Cell death & disease
DOI: 10.1038/s41419-019-1577-2
PMCID: PMC6482138
Accession Number: 31019204
Keywords: Acylation
Breast Neoplasms/metabolism/pathology
CCAAT-Enhancer-Binding Protein-beta/metabolism
Cell Line, Tumor
Cell Movement/drug effects
Epithelial-Mesenchymal Transition/*drug effects
Female
Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing)/antagonists &
inhibitors/genetics/*metabolism
Humans
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/*metabolism
Nicotine/*pharmacology
Promoter Regions, Genetic
RNA Interference
RNA, Small Interfering/metabolism
Transcription Factor CHOP/metabolism
Abstract: Cigarette smoking has been shown to be a carcinogenic factor in breast cancer. Nicotine (Nic), an active component of tobacco, has been found to induce epithelial-mesenchymal transition (EMT) in breast cancer cells. However, the alterations in protein O-GlcNAcylation in Nic-mediated tumorigenesis and malignization mechanisms are less well studied. Herein, we found that cellular O-GlcNAcylation dramatically increased in human breast cancer cells with EMT activation induced by Nic. Elevated O-GlcNAcylation subsequently promoted Nic-induced EMT activation and increased cell migratory abbility. In addition, we demonstrated that a differentiation factor for the mammary epithelium, CCAAT/enhancer-binding protein B (CEBPB), was involved in Nic-induced hyper-O-GlcNAcylation via transcriptional regulation of the expression of the key enzyme glutamine: fructose-6-phosphate amidotransferase (GFAT) and thus increased the flux through the hexosamine biosynthetic pathway (HBP). Finally, elevated O-GlcNAcylation of the transcriptional repressor C/EBP homologous protein (CHOP) suppressed its heterodimerization with CEBPB and facilitated the DNA-binding activity of CEBPB, further generating positive feedback that enhanced EMT upon Nic stimulation. In conclusion, our results have revealed a new regulatory mechanism involving CEBPB/GFAT-induced hyper-O-GlcNAcylation that plays a key role in EMT and smoking-mediated breast cancer progression.
Notes: 2041-4889
Zhang, Nana
Zhu, Tong
Yu, Kairan
Shi, Meiyun
Wang, Xue
Wang, Lingyan
Huang, Tianmiao
Li, Wenli
Liu, Yubo
Zhang, Jianing
Journal Article
Research Support, Non-U.S. Gov't
Cell Death Dis. 2019 Apr 24;10(5):343. doi: 10.1038/s41419-019-1577-2.
Author Address: School of Life Science & Medicine, Dalian University of Technology, Panjin, China.
School of Life Science & Biotechnology, Dalian University of Technology, Dalian, China.
School of Life Science & Medicine, Dalian University of Technology, Panjin, China. liuyubo@dlut.edu.cn.
School of Life Science & Medicine, Dalian University of Technology, Panjin, China. jnzhang@dlut.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1621
Author: Zhang, Q., Liu, X., Gao, W., Li, P., Hou, J., Li, J. and Wong, J.
Year: 2014
Title: Differential regulation of the ten-eleven translocation (TET) family of dioxygenases by O-linked β-N-acetylglucosamine transferase (OGT)
Journal: J Biol Chem
Volume: 289
Issue: 9
Pages: 5986-96
Epub Date: 2014/01/08
Date: Feb 28
Short Title: Differential regulation of the ten-eleven translocation (TET) family of dioxygenases by O-linked β-N-acetylglucosamine transferase (OGT)
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M113.524140
PMCID: PMC3937666
Accession Number: 24394411
Keywords: 5-Methylcytosine/analogs & derivatives
Active Transport, Cell Nucleus/physiology
Cell Nucleus/*enzymology/genetics
Cytosine/analogs & derivatives/metabolism
DNA Methylation/*physiology
Dioxygenases/genetics/*metabolism
Glucose/genetics/metabolism
HeLa Cells
Humans
N-Acetylglucosaminyltransferases/genetics/*metabolism
DNA Methylation
Glucose Metabolism
Histone Modification
O-GlcNAcylation
Protein Export
Tet3
Abstract: The ten-eleven translocation (TET) family of dioxygenases (TET1/2/3) converts 5-methylcytosine to 5-hydroxymethylcytosine and provides a vital mechanism for DNA demethylation. However, how TET proteins are regulated is largely unknown. Here we report that the O-linked β-GlcNAc (O-GlcNAc) transferase (OGT) is not only a major TET3-interacting protein but also regulates TET3 subcellular localization and enzymatic activity. OGT catalyzes the O-GlcNAcylation of TET3, promotes TET3 nuclear export, and, consequently, inhibits the formation of 5-hydroxymethylcytosine catalyzed by TET3. Although TET1 and TET2 also interact with and can be O-GlcNAcylated by OGT, neither their subcellular localization nor their enzymatic activity are affected by OGT. Furthermore, we show that the nuclear localization and O-GlcNAcylation of TET3 are regulated by glucose metabolism. Our study reveals the differential regulation of TET family proteins by OGT and a novel link between glucose metabolism and DNA epigenetic modification.
Notes: 1083-351x
Zhang, Qiao
Liu, Xiaoguang
Gao, Wenqi
Li, Pishun
Hou, Jingli
Li, Jiwen
Wong, Jiemin
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2014 Feb 28;289(9):5986-96. doi: 10.1074/jbc.M113.524140. Epub 2014 Jan 6.
Author Address: From the Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai 200241, China and.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1199
Author: Zhang, Q., Meng, Y., Zhang, L., Chen, J. and Zhu, D.
Year: 2009
Title: RNF13: a novel RING-type ubiquitin ligase over-expressed in pancreatic cancer
Journal: Cell Res
Volume: 19
Issue: 3
Pages: 348-57
Epub Date: 2008/09/17
Date: Mar
Short Title: RNF13: a novel RING-type ubiquitin ligase over-expressed in pancreatic cancer
Alternate Journal: Cell research
ISSN: 1001-0602
DOI: 10.1038/cr.2008.285
Accession Number: 18794910
Keywords: Adult
Aged
Amino Acid Sequence
Carcinoma in Situ/enzymology/pathology
Cell Line, Tumor
Cell Membrane/enzymology
Cell Proliferation
Endoplasmic Reticulum/enzymology
Glycoproteins/metabolism
Glycosylation
Golgi Apparatus/enzymology
Humans
MicroRNAs
Middle Aged
Molecular Sequence Data
Neoplasm Invasiveness
Pancreatic Neoplasms/*enzymology/pathology
Pancreatitis, Chronic/enzymology/pathology
*RING Finger Domains
Sequence Analysis, Protein
Tenascin/metabolism
Ubiquitin-Protein Ligases/chemistry/*metabolism
Abstract: Protein ubiquitination by E3 ubiquitin ligases plays an important role in cancer development. In this study, we provide experimental evidence that a RING-finger-containing protein RNF13 is an ER/Golgi membrane-associated E3 ubiquitin ligase and its RING finger domain is required for the ubiquitin ligase activity. Immunohistochemical analysis of pancreatic ductal adenocarcinoma (PDAC) and paracancerous normal tissues from 72 patients documented RNF13 over-expression in 30 tumor samples (41.7%, 30/72), and its expression was significantly associated with histological grading (P = 0.024). In addition, RNF13 was detected in precancerous lesions: tubular complexes in chronic pancreatitis (CP) and pancreatic intraepithelial neoplasia (PanIN) (79.3%, 23/29 and 62.8%, 22/35, respectively). Moreover, RNF13 staining was significantly correlated with Tenascin-C expression (P = 0.004) in PDAC samples, further supporting the role of RNF13 in cancer progression. Over-expression of wild type but not RING domain-mutant RNF13 in pancreatic MiaPaca-2 cancer cells increased invasive potential and gelatinolytic activity by matrix metalloproteinase-9. Taken together, these findings reveal that RNF13 is a novel E3 ubiquitin ligase involved in pancreatic carcinogenesis; ubiquitin-mediated modification of proteins by RNF13 may participate in pancreatic cancer development.
Notes: 1748-7838
Zhang, Qiang
Meng, Yunxiao
Zhang, Lei
Chen, Jie
Zhu, Dahai
Journal Article
Research Support, Non-U.S. Gov't
England
Cell Res. 2009 Mar;19(3):348-57. doi: 10.1038/cr.2008.285.
Author Address: National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, Tsinghua University, 5 Dong Dan San Tiao, Beijing, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1872
Author: Zhang, Q., Na, Q. and Song, W.
Year: 2017
Title: Moderate mammalian target of rapamycin inhibition induces autophagy in HTR8/SVneo cells via O-linked β-N-acetylglucosamine signaling
Journal: J Obstet Gynaecol Res
Volume: 43
Issue: 10
Pages: 1585-1596
Epub Date: 2017/07/12
Date: Oct
Short Title: Moderate mammalian target of rapamycin inhibition induces autophagy in HTR8/SVneo cells via O-linked β-N-acetylglucosamine signaling
Alternate Journal: The journal of obstetrics and gynaecology research
ISSN: 1341-8076
DOI: 10.1111/jog.13410
Accession Number: 28691313
Keywords: Apoptosis
Autophagy/*physiology
Beclin-1/*metabolism
Cell Line
Humans
N-Acetylglucosaminyltransferases/*metabolism
Qb-SNARE Proteins/*metabolism
Qc-SNARE Proteins/*metabolism
Signal Transduction/*physiology
TOR Serine-Threonine Kinases/antagonists & inhibitors/*metabolism
Trophoblasts/*metabolism
O-GlcNAc
autophagy
mTOR
phosphorylation
Abstract: AIM: Autophagy, a highly regulated process with a dual role (pro-survival or pro-death), has been implicated in adverse pregnancy outcomes. The aim of this study was to explore the mechanism whereby mammalian target of rapamycin (mTOR) signaling regulates autophagy by modulating protein O-GlcNAcylation in human trophoblasts. METHODS: HTR8/SVneo cells were incubated in serum-free medium for different time intervals or treated with varying doses of Torin1. Protein expression and cell apoptosis were detected by immunoblotting and flow cytometry, respectively. RESULTS: Short-term serum starvation or slight suppression of mTOR signaling promoted autophagy and decreased apoptosis in HTR8/SVneo cells. Conversely, prolonged serum starvation or excessive inhibition of mTOR reduced autophagy and enhanced cell apoptosis. Both serum starvation and mTOR signaling suppression reduced protein O-GlcNAcylation. Upregulation and downregulation of O-linked β-N-acetylglucosamine (O-GlcNAc) levels attenuated and augmented autophagy, respectively. Moderate mTOR inhibition-induced autophagy was blocked by upregulation of protein O-GlcNAcylation. Furthermore, immunoprecipitation studies revealed that Beclin1 and synaptosome associated protein 29 (SNAP29) could be O-GlcNAcylated, and that slight mTOR inhibition resulted in decreased O-GlcNAc modification of Beclin1 and SNAP29. Notably, we observed an inverse correlation between phosphorylation (Ser15) and O-GlcNAcylation of Beclin1. CONCLUSION: mTOR signaling inhibition played dual roles in regulating autophagy and apoptosis in HTR8/SVneo cells. Moderate mTOR suppression might induce autophagy via modulating O-GlcNAcylation of Beclin1 and SNAP29. Moreover, the negative interplay between Beclin1 O-GlcNAcylation and phosphorylation (Ser15) may be involved in autophagy regulation by mTOR signaling.
Notes: 1447-0756
Zhang, Qiuxia
Na, Quan
Song, Weiwei
Orcid: 0000-0002-8567-7680
Journal Article
Australia
J Obstet Gynaecol Res. 2017 Oct;43(10):1585-1596. doi: 10.1111/jog.13410. Epub 2017 Jul 10.
Author Address: Department of Obstetrics and Gynecology, Shengjing Hospital, China Medical University, Shenyang, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 732
Author: Zhang, Q. X., Pilquil, C. S., Dewald, J., Berthiaume, L. G. and Brindley, D. N.
Year: 2000
Title: Identification of structurally important domains of lipid phosphate phosphatase-1: implications for its sites of action
Journal: Biochem J
Volume: 345 Pt 2
Issue: Pt 2
Pages: 181-4
Epub Date: 2000/01/06
Date: Jan 15
Short Title: Identification of structurally important domains of lipid phosphate phosphatase-1: implications for its sites of action
Alternate Journal: The Biochemical journal
ISSN: 0264-6021 (Print)
0264-6021
PMCID: PMC1220744
Accession Number: 10620492
Keywords: Amino Acid Sequence
Animals
Catalytic Domain
Cell Membrane/enzymology
Cells, Cultured
Conserved Sequence
Fibroblasts/cytology
Glycosylation
Lysophospholipids/*metabolism
Membrane Proteins
Mice
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Phosphatidate Phosphatase/genetics/*metabolism
Precipitin Tests
Protein Processing, Post-Translational
Rats
Abstract: Lipid phosphate phosphatase-1 (LPP-1) dephosphorylates exogenous lysophosphatidate and thereby regulates the activation of lysophosphatidate receptors and cell division. Mutation of seven amino acids in three conserved domains of mouse LPP-1 abolished its activity. A glycosylation site was demonstrated between conserved Domains 1 and 2. LPP-1 is expressed in the plasma membrane, and the present results demonstrate the active site to be located on the outer surface.
Notes: 1470-8728
Zhang, Q X
Pilquil, C S
Dewald, J
Berthiaume, L G
Brindley, D N
Journal Article
Research Support, Non-U.S. Gov't
Biochem J. 2000 Jan 15;345 Pt 2(Pt 2):181-4.
Author Address: Department of Biochemistry, Lipid and Lipoprotein Research Group, University of Alberta, 357 Heritage Medical Research Centre, Edmonton, Alberta, Canada T6G 2S2.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1592
Author: Zhang, S., Jiang, K., Sun, C., Lu, H. and Liu, Y.
Year: 2013
Title: Quantitative analysis of site-specific N-glycans on sera haptoglobin β chain in liver diseases
Journal: Acta Biochim Biophys Sin (Shanghai)
Volume: 45
Issue: 12
Pages: 1021-9
Epub Date: 2013/10/10
Date: Dec
Short Title: Quantitative analysis of site-specific N-glycans on sera haptoglobin β chain in liver diseases
Alternate Journal: Acta biochimica et biophysica Sinica
ISSN: 1672-9145
DOI: 10.1093/abbs/gmt110
Accession Number: 24103369
Keywords: Adolescent
Adult
Amino Acid Sequence
Binding Sites
Carcinoma, Hepatocellular/blood/*metabolism
Chromatography, Liquid
Female
Glycopeptides/analysis
Glycosylation
Haptoglobins/chemistry/*metabolism
Hepatitis B/blood/metabolism
Humans
Liver Cirrhosis/blood/metabolism
Liver Diseases/blood/*metabolism
Liver Neoplasms/blood/*metabolism
Male
Middle Aged
Molecular Sequence Data
Polysaccharides/chemistry/*metabolism
Spectrometry, Mass, Electrospray Ionization
Young Adult
haptoglobin
liver diseases
mass spectrometry
site-specific N-glycan
Abstract: The site-specific characterization of N-glycans in glycoproteins with the potential of clinical application is important. In our previous report, the overall N-glycans of sera haptoglobin (Hp) β chain were found to be different in liver diseases. Hp β chain contains four potential sites of N-glycosylation. In this study, we investigated the potential change of N-glycans on Hp β chain in a site-specific fashion. Sera Hp β chain in healthy individuals as well as patients with hepatitis B virus (HBV), liver cirrhosis (LC) and hepatocellular carcinoma (HCC) were purified, digested and subjected to liquid chromatography-electrospray ionization-higher energy collision dissociation mass spectrometry, which allowed identification and structure determination of the glycopeptide, as well as the relative quantification of glycans present on each glycopeptide. The quantitative results revealed that the sialylation of NLFLN(207)HSEN(211)ATAK and the fucosylated structure at all glycopeptides increased significantly in LC and HCC patients compared with those in HBV patients and healthy individuals. A set of different N-glycan patterns of Hp β chain in various liver diseases has been determined. Thus, the sialylated and fucosylated glycoforms of Hp β chain might be related to early hepatocarcinogenesis and also might be useful as novel differential markers for LC and HCC patients.
Notes: 1745-7270
Zhang, Shu
Jiang, Kai
Sun, Chun
Lu, Haojie
Liu, Yinkun
Journal Article
Research Support, Non-U.S. Gov't
China
Acta Biochim Biophys Sin (Shanghai). 2013 Dec;45(12):1021-9. doi: 10.1093/abbs/gmt110. Epub 2013 Oct 8.
Author Address: Liver Cancer Institute, Zhongshan Hospital, Key Labolatory of Carcinogenesis and Cancer Invasion, Fudan University, Ministry of Education, Shanghai 200032, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1883
Author: Zhang, S., Li, N., Zeng, W., Gao, N. and Yang, M.
Year: 2017
Title: Cryo-EM structures of the mammalian endo-lysosomal TRPML1 channel elucidate the combined regulation mechanism
Journal: Protein Cell
Volume: 8
Issue: 11
Pages: 834-847
Epub Date: 2017/09/25
Date: Nov
Short Title: Cryo-EM structures of the mammalian endo-lysosomal TRPML1 channel elucidate the combined regulation mechanism
Alternate Journal: Protein & cell
ISSN: 1674-800X (Print)
1674-800x
DOI: 10.1007/s13238-017-0476-5
PMCID: PMC5676595
Accession Number: 28936784
Keywords: Animals
Calcium/*metabolism
Cryoelectron Microscopy
*Endocytosis
Endosomes/*metabolism
Gene Expression
HEK293 Cells
Humans
Hydrogen-Ion Concentration
Lysosomes/metabolism
Mice
Models, Biological
Mucolipidoses/genetics/*metabolism/pathology
Nanostructures/chemistry/ultrastructure
Phosphatidylinositols/*metabolism
Transgenes
Transient Receptor Potential Channels/*chemistry/genetics/metabolism
combined regulation mechanism
mTRPML1
mucolipidosis type IV
structual comparisons
Abstract: TRPML1 channel is a non-selective group-2 transient receptor potential (TRP) channel with Ca(2+) permeability. Located mainly in late endosome and lysosome of all mammalian cell types, TRPML1 is indispensable in the processes of endocytosis, membrane trafficking, and lysosome biogenesis. Mutations of TRPML1 cause a severe lysosomal storage disorder called mucolipidosis type IV (MLIV). In the present study, we determined the cryo-electron microscopy (cryo-EM) structures of Mus musculus TRPML1 (mTRPML1) in lipid nanodiscs and Amphipols. Two distinct states of mTRPML1 in Amphipols are added to the closed state, on which could represent two different confirmations upon activation and regulation. The polycystin-mucolipin domain (PMD) may sense the luminal/extracellular stimuli and undergo a "move upward" motion during endocytosis, thus triggering the overall conformational change in TRPML1. Based on the structural comparisons, we propose TRPML1 is regulated by pH, Ca(2+), and phosphoinositides in a combined manner so as to accommodate the dynamic endocytosis process.
Notes: 1674-8018
Zhang, Sensen
Li, Ningning
Zeng, Wenwen
Gao, Ning
Yang, Maojun
Journal Article
Protein Cell. 2017 Nov;8(11):834-847. doi: 10.1007/s13238-017-0476-5. Epub 2017 Sep 21.
Author Address: Ministry of Education Key Laboratory of Protein Science, Tsinghua-Peking Joint Center for Life Sciences, Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing, 100084, China.
State Key Laboratory of Membrane Biology, Peking-Tsinghua Center for Life Sciences, School of Life Science, Peking University, Beijing, 100871, China.
Institute for Immunology and School of Medicine, Tsinghua-Peking Joint Center for Life Sciences, Tsinghua University, Beijing, 100084, China.
State Key Laboratory of Membrane Biology, Peking-Tsinghua Center for Life Sciences, School of Life Science, Peking University, Beijing, 100871, China. gaon@pku.edu.cn.
Ministry of Education Key Laboratory of Protein Science, Tsinghua-Peking Joint Center for Life Sciences, Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing, 100084, China. maojunyang@tsinghua.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1424
Author: Zhang, S., Roche, K., Nasheuer, H. P. and Lowndes, N. F.
Year: 2011
Title: Modification of histones by sugar β-N-acetylglucosamine (GlcNAc) occurs on multiple residues, including histone H3 serine 10, and is cell cycle-regulated
Journal: J Biol Chem
Volume: 286
Issue: 43
Pages: 37483-95
Epub Date: 2011/09/08
Date: Oct 28
Short Title: Modification of histones by sugar β-N-acetylglucosamine (GlcNAc) occurs on multiple residues, including histone H3 serine 10, and is cell cycle-regulated
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M111.284885
PMCID: PMC3199494
Accession Number: 21896475
Keywords: Acetylglucosamine/genetics/*metabolism
Acylation
Amino Acid Substitution
Cell Cycle/*physiology
Glycosylation
HEK293 Cells
HeLa Cells
Histones/genetics/*metabolism
Humans
K562 Cells
Mutation, Missense
Phosphorylation
Protein Processing, Post-Translational/*physiology
Serine/genetics/metabolism
Transcription, Genetic/physiology
Uridine Diphosphate N-Acetylglucosamine/genetics/metabolism
Abstract: The monosaccharide, β-N-acetylglucosamine (GlcNAc), can be added to the hydroxyl group of either serines or threonines to generate an O-linked β-N-acetylglucosamine (O-GlcNAc) residue (Love, D. C., and Hanover, J. A. (2005) Sci. STKE 2005 312, 1-14; Hart, G. W., Housley, M. P., and Slawson, C. (2007) Nature 446, 1017-1022). This post-translational protein modification, termed O-GlcNAcylation, is reversible, analogous to phosphorylation, and has been implicated in many cellular processes. Here, we present evidence that in human cells all four core histones of the nucleosome are substrates for this glycosylation in the relative abundance H3, H4/H2B, and H2A. Increasing the intracellular level of UDP-GlcNAc, the nucleotide sugar donor substrate for O-GlcNAcylation enhanced histone O-GlcNAcylation and partially suppressed phosphorylation of histone H3 at serine 10 (H3S10ph). Expression of recombinant H3.3 harboring an S10A mutation abrogated histone H3 O-GlcNAcylation relative to its wild-type version, consistent with H3S10 being a site of histone O-GlcNAcylation (H3S10glc). Moreover, O-GlcNAcylated histones were lost from H3S10ph immunoprecipitates, whereas immunoprecipitation of either H3K4me3 or H3K9me3 (active or inactive histone marks, respectively) resulted in co-immunoprecipitation of O-GlcNAcylated histones. We also examined histone O-GlcNAcylation during cell cycle progression. Histone O-GlcNAcylation is high in G(1) cells, declines throughout the S phase, increases again during late S/early G(2), and persists through late G(2) and mitosis. Thus, O-GlcNAcylation is a novel histone post-translational modification regulating chromatin conformation during transcription and cell cycle progression.
Notes: 1083-351x
Zhang, Suisheng
Roche, Kevin
Nasheuer, Heinz-Peter
Lowndes, Noel Francis
Journal Article
Research Support, Non-U.S. Gov't
J Biol Chem. 2011 Oct 28;286(43):37483-95. doi: 10.1074/jbc.M111.284885. Epub 2011 Sep 6.
Author Address: Genome Stability Laboratory, Center for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, University Road, Galway, Ireland.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1853
Author: Zhang, W., Liu, T., Dong, H., Bai, H., Tian, F., Shi, Z., Chen, M., Wang, J., Qin, W. and Qian, X.
Year: 2017
Title: Synthesis of a Highly Azide-Reactive and Thermosensitive Biofunctional Reagent for Efficient Enrichment and Large-Scale Identification of O-GlcNAc Proteins by Mass Spectrometry
Journal: Anal Chem
Volume: 89
Issue: 11
Pages: 5810-5817
Epub Date: 2017/05/17
Date: Jun 6
Short Title: Synthesis of a Highly Azide-Reactive and Thermosensitive Biofunctional Reagent for Efficient Enrichment and Large-Scale Identification of O-GlcNAc Proteins by Mass Spectrometry
Alternate Journal: Analytical chemistry
ISSN: 0003-2700
DOI: 10.1021/acs.analchem.6b04960
Accession Number: 28510447
Abstract: O-linked β-N-acetylglucosamine (O-GlcNAc) is a ubiquitous post-translational modification of proteins in eukaryotic cells. Despite their low abundance, O-GlcNAc-modified proteins play many important roles in regulating gene expression, signal transduction, and cell cycle. Aberrant O-GlcNAc proteins are correlated with many major human diseases, such as Alzheimer's disease, diabetes, and cancer. Because of the extremely low stoichiometry of O-GlcNAc proteins, enrichment is required before mass spectrometry analysis for large-scale identification and in-depth understanding of their cellular function. In this work, we designed and synthesized a novel thermosensitive immobilized triarylphosphine reagent as a convenient tool for efficient enrichment of azide-labeled O-GlcNAc proteins from complex biological samples. Immobilization of triarylphosphine on highly water-soluble thermosensitive polymer largely increases its solubility and reactivity in aqueous solution. As a result, facilitated coupling is achieved between triarylphosphine and azide-labeled O-GlcNAc proteins via Staudinger ligation, due to the increased triarylphosphine concentration, reduced interfacial mass transfer resistance, and steric hindrance in homogeneous reaction. Furthermore, solubility of the polymer from complete dissolution to full precipitation can be easily controlled by simply adjusting the environmental temperature. Therefore, facile sample recovery can be achieved by increasing the temperature to precipitate the polymer-O-GlcNAc protein conjugates from solution. This novel immobilized triarylphosphine reagent enables efficient enrichment and sensitive detection of more than 1700 potential O-GlcNAc proteins from HeLa cell using mass spectrometry, demonstrating its potential as a general strategy for low-abundance target enrichment.
Notes: 1520-6882
Zhang, Wanjun
Liu, Tong
Dong, Hangyan
Bai, Haihong
Tian, Fang
Shi, Zhaomei
Chen, Mingli
Wang, Jianhua
Orcid: 0000-0003-2175-3610
Qin, Weijie
Orcid: 0000-0002-7633-9786
Qian, Xiaohong
Journal Article
Research Support, Non-U.S. Gov't
United States
Anal Chem. 2017 Jun 6;89(11):5810-5817. doi: 10.1021/acs.analchem.6b04960. Epub 2017 May 16.
Author Address: National Center for Protein Sciences Beijing, State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine , Beijing 102206, PR China.
Research Center for Analytical Sciences, College of Sciences, Northeastern University , Shenyang 110819, PR China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1194
Author: Zhang, W., Na, T. and Peng, J. B.
Year: 2008
Title: WNK3 positively regulates epithelial calcium channels TRPV5 and TRPV6 via a kinase-dependent pathway
Journal: Am J Physiol Renal Physiol
Volume: 295
Issue: 5
Pages: F1472-84
Epub Date: 2008/09/05
Date: Nov
Short Title: WNK3 positively regulates epithelial calcium channels TRPV5 and TRPV6 via a kinase-dependent pathway
Alternate Journal: American journal of physiology. Renal physiology
ISSN: 1931-857X (Print)
1522-1466
DOI: 10.1152/ajprenal.90229.2008
PMCID: PMC2584897
Accession Number: 18768590
Keywords: Amino Acid Substitution
Animals
Calcium/metabolism
Calcium Channels/genetics/*physiology
Cell Size
Colchicine/pharmacology
Cytochalasin D/pharmacology
Electrophysiology
Exocytosis/drug effects/physiology
Female
Glycosylation
Humans
Membrane Potentials/drug effects
Oocytes/cytology/metabolism/physiology
Phosphotransferases/genetics/*metabolism
Protein Transport/drug effects/physiology
Protein-Serine-Threonine Kinases/genetics/*metabolism
Sequence Deletion
Sodium/metabolism
Sodium Chloride Symporters/genetics/physiology
TRPV Cation Channels/genetics/*physiology
Xenopus laevis
Abstract: WNK3, a member of the With No Lysine (K) family of protein serine/threonine kinases, was shown to regulate members of the SLC12A family of cation-chloride cotransporters and the renal outer medullary K+ channel ROMK and Cl(-) channel SLC26A9. To evaluate the effect of WNK3 on TRPV5, a renal epithelial Ca2+ channel that serves as a gatekeeper for active Ca2+ reabsorption, WNK3 and TRPV5 were coexpressed in Xenopus laevis oocytes and the function and expression of TRPV5 were subsequently examined. An 82.7 +/- 7.1% increase in TRPV5-mediated Ca2+ uptake was observed when WNK3 was coexpressed. A similar increase in TRPV5-mediated Na+ current was observed with the voltage-clamp technique. WNK3 also enhanced Ca2+ influx and Na+ current mediated by TRPV6, which is the closest homolog of TRPV5 that mediates active intestinal Ca2+ absorption. The kinase domain of WNK3 alone was sufficient to increase TRPV5-mediated Ca2+ transport, and the positive regulatory effect was abolished by the kinase-inactive D294A mutation in WNK3, indicating a kinase-dependent mechanism. The complexly glycosylated TRPV5 that appears at the plasma membrane was increased by WNK3. The exocytosis of TRPV5 was increased by WNK3, and the effect of WNK3 on TRPV5 was abolished by the microtubule inhibitor colchicine. The increased plasma membrane expression of TRPV5 was likely due to the enhanced delivery of mature TRPV5 to the plasma membrane from its intracellular pool via the secretory pathway. These results indicate that WNK3 is a positive regulator of the transcellular Ca2+ transport pathway.
Notes: 1522-1466
Zhang, Wei
Na, Tao
Peng, Ji-Bin
R01 DK072154/DK/NIDDK NIH HHS/United States
R01-DK-072154/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Am J Physiol Renal Physiol. 2008 Nov;295(5):F1472-84. doi: 10.1152/ajprenal.90229.2008. Epub 2008 Sep 3.
Author Address: Nephrology Research and Training Center, Division of Nephrology, Department of Medicine, University of Alabama at Birmingham, ZRB 625, 1900 University Blvd., Birmingham, AL 35294-0006, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1851
Author: Zhang, X., Qiao, Y., Wu, Q., Chen, Y., Zou, S., Liu, X., Zhu, G., Zhao, Y., Chen, Y., Yu, Y., Pan, Q., Wang, J. and Sun, F.
Year: 2017
Title: The essential role of YAP O-GlcNAcylation in high-glucose-stimulated liver tumorigenesis
Journal: Nat Commun
Volume: 8
Pages: 15280
Epub Date: 2017/05/06
Date: May 5
Short Title: The essential role of YAP O-GlcNAcylation in high-glucose-stimulated liver tumorigenesis
Alternate Journal: Nature communications
ISSN: 2041-1723
DOI: 10.1038/ncomms15280
PMCID: PMC5424161
Accession Number: 28474680
Keywords: Acetylglucosamine/*metabolism
Adaptor Proteins, Signal Transducing/metabolism
Animals
Carcinogenesis/drug effects/*metabolism/pathology
Cell Line, Tumor
Gene Expression Regulation, Neoplastic/drug effects
Glucose/*toxicity
Glycosylation/drug effects
Humans
Liver Neoplasms/genetics/*metabolism/pathology
Male
Mice, Inbred BALB C
N-Acetylglucosaminyltransferases/metabolism
Phosphoproteins/metabolism
Protein Stability/drug effects
Threonine/metabolism
Transcription Factors
Transcription, Genetic/drug effects
beta-Transducin Repeat-Containing Proteins/metabolism
Abstract: O-GlcNAcylation has been implicated in the tumorigenesis of various tissue origins, but its function in liver tumorigenesis is not clear. Here, we demonstrate that O-GlcNAcylation can enhance the expression, stability and function of Yes-associated protein (YAP), the downstream transcriptional regulator of the Hippo pathway and a potent oncogenic factor in liver cancer. O-GlcNAcylation induces transformative phenotypes of liver cancer cells in a YAP-dependent manner. An O-GlcNAc site of YAP was identified at Thr241, and mutating this site decreased the O-GlcNAcylation, stability, and pro-tumorigenic capacities of YAP, while increasing YAP phosphorylation. Importantly, we found via in vitro cell-based and in vivo mouse model experiments that O-GlcNAcylation of YAP was required for high-glucose-induced liver tumorigenesis. Interestingly, a positive feedback between YAP and global cellular O-GlcNAcylation is also uncovered. We conclude that YAP O-GlcNAcylation is a potential therapeutic intervention point for treating liver cancer associated with high blood glucose levels and possibly diabetes.
Notes: 2041-1723
Zhang, Xiao
Qiao, Yongxia
Wu, Qi
Chen, Yan
Zou, Shaowu
Liu, Xiangfan
Zhu, Guoqing
Zhao, Yinghui
Chen, Yuxin
Yu, Yongchun
Pan, Qiuhui
Wang, Jiayi
Sun, Fenyong
Journal Article
Research Support, Non-U.S. Gov't
Nat Commun. 2017 May 5;8:15280. doi: 10.1038/ncomms15280.
Author Address: Department of Clinical Laboratory Medicine, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
School of Public Health, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
Department of Hepatopancreatobiliary, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
Faculty of Medical Laboratory Science, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China.
Shanghai Municipal Hospital of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai 200071, China.
Department of Central Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai 200072, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1926
Author: Zhang, X., Wang, Q., Wu, J., Wang, J., Shi, Y. and Liu, M.
Year: 2018
Title: Crystal structure of human lysyl oxidase-like 2 (hLOXL2) in a precursor state
Journal: Proc Natl Acad Sci U S A
Volume: 115
Issue: 15
Pages: 3828-3833
Epub Date: 2018/03/28
Date: Apr 10
Short Title: Crystal structure of human lysyl oxidase-like 2 (hLOXL2) in a precursor state
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1720859115
PMCID: PMC5899467
Accession Number: 29581294
Keywords: Amino Acid Motifs
Amino Acid Oxidoreductases/*chemistry/metabolism
Catalysis
Copper/chemistry/metabolism
Crystallography, X-Ray
Humans
Lysine/analogs & derivatives/chemistry/metabolism
Quinones/chemistry/metabolism
Zinc/chemistry/metabolism
*crystal structure
*human LOXL2
*lysyl oxidase
*precursor state
Abstract: Lysyl oxidases (LOXs), a type of copper- and lysyl tyrosylquinone (LTQ) -dependent amine oxidase, catalyze the oxidative deamination of lysine residues of extracellular matrix (ECM) proteins such as elastins and collagens and generate aldehyde groups. The oxidative deamination of lysine represents the foundational step for the cross-linking of elastin and collagen and thus is crucial for ECM modeling. Despite their physiological significance, the structure of this important family of enzymes remains elusive. Here we report the crystal structure of human lysyl oxidase-like 2 (hLOXL2) at 2.4-Å resolution. Unexpectedly, the copper-binding site of hLOXL2 is occupied by zinc, which blocks LTQ generation and the enzymatic activity of hLOXL2 in our in vitro assay. Biochemical analysis confirms that copper loading robustly activates hLOXL2 and supports LTQ formation. Furthermore, the LTQ precursor residues in the structure are distanced by 16.6 Å, corroborating the notion that the present structure may represent a precursor state and that pronounced conformational rearrangements would be required for protein activation. The structure presented here establishes an important foundation for understanding the structure-function relationship of LOX proteins and will facilitate LOX-targeting drug discovery.
Notes: 1091-6490
Zhang, Xi
Wang, Qifan
Wu, Jianping
Wang, Jiawei
Shi, Yigong
Liu, Minhao
Journal Article
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2018 Apr 10;115(15):3828-3833. doi: 10.1073/pnas.1720859115. Epub 2018 Mar 26.
Author Address: Beijing Advanced Innovation Center for Structural Biology, Tsinghua University, 100084 Beijing, China.
Tsinghua-Peking Joint Center for Life Sciences, Tsinghua University, 100084 Beijing, China.
School of Life Sciences, Tsinghua University, 100084 Beijing, China.
School of Medicine, Tsinghua University, 100084 Beijing, China.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua University, 100084 Beijing, China; shi-lab@tsinghua.edu.cn liuminhao@biomed.tsinghua.edu.cn.
Institute of Biology, Westlake Institute for Advanced Study, Westlake University, 310064 Hangzhou, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1783
Author: Zhang, Z., Costa, F. C., Tan, E. P., Bushue, N., DiTacchio, L., Costello, C. E., McComb, M. E., Whelan, S. A., Peterson, K. R. and Slawson, C.
Year: 2016
Title: O-Linked N-Acetylglucosamine (O-GlcNAc) Transferase and O-GlcNAcase Interact with Mi2β Protein at the Aγ-Globin Promoter
Journal: J Biol Chem
Volume: 291
Issue: 30
Pages: 15628-40
Epub Date: 2016/05/28
Date: Jul 22
Short Title: O-Linked N-Acetylglucosamine (O-GlcNAc) Transferase and O-GlcNAcase Interact with Mi2β Protein at the Aγ-Globin Promoter
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M116.721928
PMCID: PMC4957047
Accession Number: 27231347
Keywords: Animals
GATA1 Transcription Factor/genetics/metabolism
Gene Silencing/*physiology
Humans
K562 Cells
Mice
Mice, Transgenic
N-Acetylglucosaminyltransferases/genetics/*metabolism
Nuclear Proteins/genetics/metabolism
*Response Elements
Transcription Factors/genetics/metabolism
beta-N-Acetylhexosaminidases/genetics/*metabolism
gamma-Globins/*biosynthesis/genetics
*Chd4
*Fog
*GATA transcription factor
*Globin
*O-GlcNAc-transferase
*O-GlcNAcase
*O-GlcNAcylation
*O-linked N-acetylglucosamine (O-GlcNAc)
*post-translational modification (PTM)
*transcription
Abstract: One mode of γ-globin gene silencing involves a GATA-1·FOG-1·Mi2β repressor complex that binds to the -566 GATA site relative to the (A)γ-globin gene cap site. However, the mechanism of how this repressor complex is assembled at the -566 GATA site is unknown. In this study, we demonstrate that the O-linked N-acetylglucosamine (O-GlcNAc) processing enzymes, O-GlcNAc-transferase (OGT) and O-GlcNAcase (OGA), interact with the (A)γ-globin promoter at the -566 GATA repressor site; however, mutation of the GATA site to GAGA significantly reduces OGT and OGA promoter interactions in β-globin locus yeast artificial chromosome (β-YAC) bone marrow cells. When WT β-YAC bone marrow cells are treated with the OGA inhibitor Thiamet-G, the occupancy of OGT, OGA, and Mi2β at the (A)γ-globin promoter is increased. In addition, OGT and Mi2β recruitment is increased at the (A)γ-globin promoter when γ-globin becomes repressed in postconception day E18 human β-YAC transgenic mouse fetal liver. Furthermore, we show that Mi2β is modified with O-GlcNAc, and both OGT and OGA interact with Mi2β, GATA-1, and FOG-1. Taken together, our data suggest that O-GlcNAcylation is a novel mechanism of γ-globin gene regulation mediated by modulating the assembly of the GATA-1·FOG-1·Mi2β repressor complex at the -566 GATA motif within the promoter.
Notes: 1083-351x
Zhang, Zhen
Costa, Flávia C
Tan, Ee Phie
Bushue, Nathan
DiTacchio, Luciano
Costello, Catherine E
McComb, Mark E
Whelan, Stephen A
Peterson, Kenneth R
Slawson, Chad
R01 DK100595/DK/NIDDK NIH HHS/United States
HHSN268201000031C/HL/NHLBI NIH HHS/United States
P20 GM104936/GM/NIGMS NIH HHS/United States
R01 HL111264/HL/NHLBI NIH HHS/United States
P41 GM104603/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Biol Chem. 2016 Jul 22;291(30):15628-40. doi: 10.1074/jbc.M116.721928. Epub 2016 May 26.
Author Address: From the Department of Biochemistry and Molecular Biology.
IntelligeneDx Cancer Genomics, Olathe, Kansas 66061.
Pharmacology, Toxicology and Therapeutics.
Department of Biochemistry and Center for Biomedical Mass Spectrometry, Boston University School of Medicine, Boston, Massachusetts 02118, and.
From the Department of Biochemistry and Molecular Biology, Anatomy and Cell Biology, and Cancer Center, Institute for Reproductive Health and Regenerative Medicine, and kpeterson@kumc.edu.
From the Department of Biochemistry and Molecular Biology, Cancer Center, Institute for Reproductive Health and Regenerative Medicine, and Alzheimer's Disease Center, University of Kansas Medical Center, Kansas City, Kansas 66160, cslawson@kumc.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1356
Author: Zhao, J., Liu, Y. H., Reichert, P., Pflanz, S. and Pramanik, B.
Year: 2010
Title: Glycosylation analysis of interleukin-23 receptor: elucidation of glycosylation sites and characterization of attached glycan structures
Journal: J Mass Spectrom
Volume: 45
Issue: 12
Pages: 1416-25
Epub Date: 2010/11/06
Date: Dec
Short Title: Glycosylation analysis of interleukin-23 receptor: elucidation of glycosylation sites and characterization of attached glycan structures
Alternate Journal: Journal of mass spectrometry : JMS
ISSN: 1076-5174
DOI: 10.1002/jms.1858
Accession Number: 21053369
Keywords: Acetylglucosamine/chemistry/metabolism
Amino Acid Sequence
Carbohydrate Conformation
Chromatography, High Pressure Liquid
Glycopeptides/chemistry/metabolism
Glycosylation
Humans
Molecular Sequence Data
Polysaccharides/*chemistry/*metabolism
Receptors, Interleukin/*chemistry/*metabolism
Recombinant Proteins/chemistry/metabolism
Tandem Mass Spectrometry
Abstract: Interleukin-23 (IL-23) is a heterodimeric cytokine, a central factor in chronic/autoimmune inflammation. It signals through a heterodimeric receptor consisting of IL-23r, which is heavily glycosylated. The structural characterization of IL-23r has not been reported. In this work, glycosylation profiles of soluble recombinant human IL-23r (rhIL-23r) were established using mass spectrometry (MS), which included defining glycosylation sites, degree of glycosylation occupancy of each site and structure of attached oligosaccharides. Specifically, precursor ion scan of oxonium ion protonated N-acetylglucosamine (GlcNAc(+)) (m/z 204) was performed using a triple quadrupole MS instrument to locate the retention time of glycopeptides. Both the glycopeptides and their corresponding deglycosylated forms in each collected HPLC fraction were studied by liquid chromatography-tandem mass spectrometry (LC-MS/MS) (LTQ-Orbitrap) for glycosylation site profiling. The attached glycan structures were elucidated by collision-induced dissociation (CID) fragmentation of target glycopeptides in combination with accurate mass measurement. Eight glycosylation sites were identified on IL-23r (Asn24, Asn209, Asn239, Asn157, Asn118, Asn250, Asn58 and Asn6). Most of the glycosylation sites were > 95% occupied except Asn250 and Asn6. Those two sites were 88% and 45% occupied by estimation from trypsin digestion and were 55% and 42% occupied from LysC digestion. Multiple glycoforms were observed in IL-23r. Most of them were bi-, tri- or tetra-antennary complex type structures with fucose and sialic acid. High mannose and hybrid type glycans were only observed on Asn157. The structural characterization on IL-23r glycosylation provides useful information for better understanding of the biological function of IL-23r.
Notes: 1096-9888
Zhao, Jia
Liu, Yan-Hui
Reichert, Paul
Pflanz, Stefan
Pramanik, Birendra
Journal Article
England
J Mass Spectrom. 2010 Dec;45(12):1416-25. doi: 10.1002/jms.1858.
Author Address: Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, NJ 07033, USA. jia.zhao@merck.com
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2052
Author: Zhao, L., Li, M., Wei, T., Feng, C., Wu, T., Shah, J. A., Liu, H., Wang, F., Cai, Y. and Jin, J.
Year: 2019
Title: O-GlcNAc-Modification of NSL3 at Thr755 Site Maintains the Holoenzyme Activity of MOF/NSL Histone Acetyltransfease Complex
Journal: Int J Mol Sci
Volume: 21
Issue: 1
Epub Date: 2019/12/29
Date: Dec 25
Short Title: O-GlcNAc-Modification of NSL3 at Thr755 Site Maintains the Holoenzyme Activity of MOF/NSL Histone Acetyltransfease Complex
Alternate Journal: International journal of molecular sciences
ISSN: 1422-0067
DOI: 10.3390/ijms21010173
PMCID: PMC6981688
Accession Number: 31881804
Keywords: Acetylation
HEK293 Cells
Histone Acetyltransferases/chemistry/*metabolism
Holoenzymes/chemistry/metabolism
Humans
Intracellular Signaling Peptides and Proteins/chemistry/genetics/*metabolism
N-Acetylglucosaminyltransferases/antagonists & inhibitors/genetics/metabolism
Protein Binding
Protein Stability
Proteolysis
RNA Interference
RNA, Small Interfering/metabolism
Threonine/metabolism
Ubiquitin-Conjugating Enzymes/metabolism
O-GlcNAc-modification
histone acetyltransferase
nonspecific lethal protein NSL3
protein degradation
ubiquitin
Abstract: Both OGT1 (O-linked β-N-acetylglucosamine (O-GlcNAc) transferase isoform 1) and NSL3 (nonspecific lethal protein 3) are crucial components of the MOF (males absent on the first)/NSL histone acetyltransferase complex. We previously described how global histone H4 acetylation levels were modulated by OGT1/O-GlcNAcylation-mediated NSL3 stability. However, the specific modification site of NSL3 and its molecular mechanism of protein stability remain unknown. Here, we present evidence from biochemical experiments arguing that O-GlcNAcylation of NSL3 at Thr755 is tightly associated with holoenzyme activity of the MOF/NSL complex. Using in vitro O-GlcNAc-transferase assays combined with mass spectrometry, we suppose that the residue Thr755 on NSL3 C-terminus is the major site O-GlcNAc-modified by OGT1. Importantly, O-GlcNAcylation of this site is involved in the regulation of the ubiquitin-degradation of NSL3, because this site mutation (T755A) promotes the ubiquitin-mediated degradation of NSL3. Further in-depth research found that ubiquitin conjugating enzyme E2 S (UBE2S) accelerated the degradation of NSL3 via direct binding to it. Interestingly, OGT1 and UBE2S competitively bind to NSL3, suggesting the coordination of OGT1-UBE2S in regulating NSL3 stability. Furthermore, O-GlcNAcylation of NSL3 Thr755 site regulates the histone H4 acetylation levels at lysine 5, 8, and 16, suggesting that the O-GlcNAcylation of NSL3 at Thr755 is required for maintaining the integrity and holoenzyme activity of the MOF/NSL complex. In colony formation assays, we found that the integrity of the complex impacts the proliferation of the lung carcinoma type II epithelium-like A549 cells. Taken together, our results provide new insight into the elucidation of the molecular mechanism of the MOF/NSL complex.
Notes: 1422-0067
Zhao, Linhong
Li, Min
Wei, Tao
Feng, Chang
Wu, Tingting
Shah, Junaid Ali
Liu, Hongsen
Wang, Fei
Cai, Yong
Jin, Jingji
31571316 & 31771421/National Natural Science Foundation of China/
Journal Article
Int J Mol Sci. 2019 Dec 25;21(1):173. doi: 10.3390/ijms21010173.
Author Address: School of Life Sciences, Jilin University, Changchun City, Jilin 130012, China.
School of Pharmacy, Changchun University of Chinese Medicine, Changchun City, Jilin 130117, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1139
Author: Zhao, L. Y., Tsuboi, K., Okamoto, Y., Nagahata, S. and Ueda, N.
Year: 2007
Title: Proteolytic activation and glycosylation of N-acylethanolamine-hydrolyzing acid amidase, a lysosomal enzyme involved in the endocannabinoid metabolism
Journal: Biochim Biophys Acta
Volume: 1771
Issue: 11
Pages: 1397-405
Epub Date: 2007/11/06
Date: Nov
Short Title: Proteolytic activation and glycosylation of N-acylethanolamine-hydrolyzing acid amidase, a lysosomal enzyme involved in the endocannabinoid metabolism
Alternate Journal: Biochimica et biophysica acta
ISSN: 0006-3002 (Print)
0006-3002
DOI: 10.1016/j.bbalip.2007.10.002
Accession Number: 17980170
Keywords: Amidohydrolases/chemistry/genetics/*metabolism
Amino Acid Sequence
Amino Acid Substitution
Asparagine/chemistry
Cannabinoid Receptor Modulators/*metabolism
Cell Line
*Endocannabinoids
Enzyme Activation
Enzyme Stability
Glycosylation
Humans
Lysosomes/enzymology
Molecular Sequence Data
Molecular Weight
Mutagenesis, Site-Directed
Peptide Hydrolases/metabolism
Protein Processing, Post-Translational
Recombinant Proteins/chemistry/genetics/metabolism
Abstract: N-acylethanolamine-hydrolyzing acid amidase (NAAA) is a lysosomal enzyme hydrolyzing bioactive N-acylethanolamines, including anandamide and N-palmitoylethanolamine. Previously, we suggested that NAAA is glycosylated and proteolytically cleaved. Here, we investigated the mechanism and significance of the cleavage of human NAAA overexpressed in human embryonic kidney 293 cells. Western blotting with anti-NAAA antibody revealed that most of NAAA in the cell homogenate was the cleaved 30-kDa form. However, some of NAAA were released outside the cells and the extracellular enzyme was mostly the uncleaved 48-kDa form. When incubated at pH 4.5, the 48-kDa form was time-dependently converted to the 30-kDa form with concomitant increase in the N-palmitoylethanolamine-hydrolyzing activity. The purified 48-kDa form was also cleaved and activated. However, the cleavage did not proceed at pH 7.4 or in the presence of p-chloromercuribenzoic acid. The mutant C126S was resistant to the cleavage and remained inactive. These results suggested that this specific proteolysis is a self-catalyzed activation step. We next determined N-glycosylation sites of human NAAA by site-directed mutagenesis addressed to asparagine residues in six potential N-glycosylation sites. The results exhibited that Asn-37, Asn-107, Asn-309, and Asn-333 are actual N-glycosylation sites. The glycosylation appeared to play an important role in stabilizing the enzyme protein.
Notes: Zhao, Li-Ying
Tsuboi, Kazuhito
Okamoto, Yasuo
Nagahata, Shunichiro
Ueda, Natsuo
Journal Article
Research Support, Non-U.S. Gov't
Netherlands
Biochim Biophys Acta. 2007 Nov;1771(11):1397-405. doi: 10.1016/j.bbalip.2007.10.002. Epub 2007 Oct 13.
Author Address: Department of Biochemistry, Kagawa University School of Medicine, 1750-1 Ikenobe, Miki, Kagawa 761-0793, Japan.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 954
Author: Zhao, M., Boja, E. S., Hoodbhoy, T., Nawrocki, J., Kaufman, J. B., Kresge, N., Ghirlando, R., Shiloach, J., Pannell, L., Levine, R. L., Fales, H. M. and Dean, J.
Year: 2004
Title: Mass spectrometry analysis of recombinant human ZP3 expressed in glycosylation-deficient CHO cells
Journal: Biochemistry
Volume: 43
Issue: 38
Pages: 12090-104
Epub Date: 2004/09/24
Date: Sep 28
Short Title: Mass spectrometry analysis of recombinant human ZP3 expressed in glycosylation-deficient CHO cells
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi048958k
Accession Number: 15379548
Keywords: Amino Acid Sequence
Animals
CHO Cells
Cricetinae
Disulfides/chemistry/metabolism
Egg Proteins/*chemistry/genetics/isolation & purification/*metabolism
Gene Expression
Glycosylation
Humans
Mass Spectrometry
Membrane Glycoproteins/*chemistry/genetics/isolation & purification/*metabolism
Mice
Molecular Sequence Data
Receptors, Cell Surface/*chemistry/genetics/isolation & purification/*metabolism
Recombinant Proteins/*chemistry/genetics/isolation & purification/*metabolism
Sequence Alignment
Zona Pellucida Glycoproteins
Abstract: The zona pellucida is an extracellular matrix that mediates taxon-specific fertilization in which human sperm will not bind to mouse eggs. The mouse zona pellucida is composed of three glycoproteins (ZP1, ZP2, ZP3). The primary structure of each has been deduced from the cDNA nucleic acid sequence, and each has been analyzed by mass spectrometry. However, determination of the secondary structure and processing of the human zona proteins have been hampered by the paucity of biological material. To investigate if taxon-specific sperm-egg recognition was ascribable to structural differences in a zona protein required for matrix formation, recombinant human ZP3 was expressed in CHO-Lec3.2.8.1 cells and compared to mouse ZP3. With nearly complete coverage, LC-QTOF mass spectrometry was used to determine the cleavage of an N-terminal signal peptide (amino acids 1-22) and the release of secreted ZP3 from a C-terminal transmembrane domain (amino acids 379-424). The resultant N-terminal glutamine was cyclized to pyroglutamate (pyrGln(23)), and several C-terminal peptides were detected, including one ending at Asn(350). The disulfide bond linkages of eight cysteine residues in the conserved zona domain were ascertained (Cys(46)/Cys(140), Cys(78)/Cys(99), Cys(217)/Cys(282), Cys(239)/Cys(300)), but the precise linkage of two additional disulfide bonds was indeterminate due to clustering of the remaining four cysteine residues (Cys(319), Cys(321), Cys(322), Cys(327)). Three of the four potential N-linked oligosaccharide binding sites (Asn(125), Asn(147), Asn(272)) were occupied, and clusters of O-glycans were observed within two regions, amino acids 156-173 and 260-281. Taken together, these data indicate that human and mouse ZP3 proteins are quite similar, and alternative explanations of taxon-specific sperm binding warrant exploration.
Notes: Zhao, Ming
Boja, Emily S
Hoodbhoy, Tanya
Nawrocki, Joseph
Kaufman, Jeanne B
Kresge, Nicole
Ghirlando, Rodolfo
Shiloach, Joseph
Pannell, Lewis
Levine, Rodney L
Fales, Henry M
Dean, Jurrien
Journal Article
United States
Biochemistry. 2004 Sep 28;43(38):12090-104. doi: 10.1021/bi048958k.
Author Address: Laboratory of Cellular and Developmental Biology, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1649
Author: Zhao, N., Zhang, A. S., Worthen, C., Knutson, M. D. and Enns, C. A.
Year: 2014
Title: An iron-regulated and glycosylation-dependent proteasomal degradation pathway for the plasma membrane metal transporter ZIP14
Journal: Proc Natl Acad Sci U S A
Volume: 111
Issue: 25
Pages: 9175-80
Epub Date: 2014/06/14
Date: Jun 24
Short Title: An iron-regulated and glycosylation-dependent proteasomal degradation pathway for the plasma membrane metal transporter ZIP14
Alternate Journal: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424 (Print)
0027-8424
DOI: 10.1073/pnas.1405355111
PMCID: PMC4078863
Accession Number: 24927598
Keywords: Cation Transport Proteins/genetics/*metabolism
Cell Membrane/genetics/*metabolism
Endocytosis/physiology
Glycosylation
Hep G2 Cells
Humans
Iron/*metabolism
Proteasome Endopeptidase Complex/genetics/*metabolism
Protein Structure, Tertiary
*Proteolysis
Slc39a14
hereditary hemochromatosis
Abstract: Protein degradation is instrumental in regulating cellular function. Plasma membrane proteins targeted for degradation are internalized and sorted to multivesicular bodies, which fuse with lysosomes, where they are degraded. ZIP14 is a newly identified iron transporter with multitransmembrane domains. In an attempt to dissect the molecular mechanisms by which iron regulates ZIP14 levels, we found that ZIP14 is endocytosed, extracted from membranes, deglycosylated, and degraded by proteasomes. This pathway did not depend on the retrograde trafficking to the endoplasmic reticulum and thus did not involve the well-defined endoplasmic reticulum-associated protein degradation pathway. Iron inhibited membrane extraction of internalized ZIP14, resulting in higher steady-state levels of ZIP14. Asparagine-linked (N-linked) glycosylation of ZIP14, particularly the glycosylation at N102, was required for efficient membrane extraction of ZIP14 and therefore is necessary for its iron sensitivity. These findings highlight the importance of proteasomes in the degradation of endocytosed plasma membrane proteins.
Notes: 1091-6490
Zhao, Ningning
Zhang, An-Sheng
Worthen, Christal
Knutson, Mitchell D
Enns, Caroline A
DK054488/DK/NIDDK NIH HHS/United States
R01 DK072166/DK/NIDDK NIH HHS/United States
R37 DK054488/DK/NIDDK NIH HHS/United States
R01 DK054488/DK/NIDDK NIH HHS/United States
GM071338-08/GM/NIGMS NIH HHS/United States
R01 DK080706/DK/NIDDK NIH HHS/United States
DK072166/DK/NIDDK NIH HHS/United States
T32 GM071338/GM/NIGMS NIH HHS/United States
R01 DK080765/DK/NIDDK NIH HHS/United States
DK080706/DK/NIDDK NIH HHS/United States
DK080765/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Proc Natl Acad Sci U S A. 2014 Jun 24;111(25):9175-80. doi: 10.1073/pnas.1405355111. Epub 2014 Jun 9.
Author Address: Department of Cell and Developmental Biology, Oregon Health and Science University, Portland, OR 97239; and.
Food Science and Human Nutrition Department, University of Florida, Gainesville, FL 32611.
Department of Cell and Developmental Biology, Oregon Health and Science University, Portland, OR 97239; and ennsca@ohsu.edu.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1668
Author: Zhao, P., Schulz, T. C., Sherrer, E. S., Weatherly, D. B., Robins, A. J. and Wells, L.
Year: 2015
Title: The human embryonic stem cell proteome revealed by multidimensional fractionation followed by tandem mass spectrometry
Journal: Proteomics
Volume: 15
Issue: 2-3
Pages: 554-66
Epub Date: 2014/11/05
Date: Jan
Short Title: The human embryonic stem cell proteome revealed by multidimensional fractionation followed by tandem mass spectrometry
Alternate Journal: Proteomics
ISSN: 1615-9853 (Print)
1615-9853
DOI: 10.1002/pmic.201400132
PMCID: PMC4591931
NIHMSID: NIHMS723868
Accession Number: 25367160
Keywords: Acetylglucosamine/analysis/metabolism
Cell Fractionation
Cell Line
Embryonic Stem Cells/chemistry/*metabolism
Humans
Phosphorylation
Protein Processing, Post-Translational
Proteome/*analysis/metabolism
Proteomics
Tandem Mass Spectrometry
Cell biology
Embryonic stem cells
O-GlcNAc
Secretome
Shotgun proteomics
Abstract: Human embryonic stem cells (hESCs) have received considerable attention due to their therapeutic potential and usefulness in understanding early development and cell fate commitment. In order to appreciate the unique properties of these pluripotent, self-renewing cells, we have performed an in-depth multidimensional fractionation followed by LC-MS/MS analysis of the hESCs harvested from defined media to elucidate expressed, phosphorylated, O-linked β-N-acetylglucosamine (O-GlcNAc) modified, and secreted proteins. From the triplicate analysis, we were able to assign more than 3000 proteins with less than 1% false-discovery rate. This analysis also allowed us to identify nearly 500 phosphorylation sites and 68 sites of O-GlcNAc modification with the same high confidence. Investigation of the phosphorylation sites allowed us to deduce the set of kinases that are likely active in these cells. We also identified more than 100 secreted proteins of hESCs that likely play a role in extracellular matrix formation and remodeling, as well as autocrine signaling for self-renewal and maintenance of the undifferentiated state. Finally, by performing in-depth analysis in triplicate, spectral counts were obtained for these proteins and posttranslationally modified peptides, which will allow us to perform relative quantitative analysis between these cells and any derived cell type in the future.
Notes: 1615-9861
Zhao, Peng
Schulz, Thomas C
Sherrer, Eric S
Weatherly, D Brent
Robins, Allan J
Wells, Lance
P41 GM103490/GM/NIGMS NIH HHS/United States
P41GM103490/GM/NIGMS NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Proteomics. 2015 Jan;15(2-3):554-66. doi: 10.1002/pmic.201400132. Epub 2014 Dec 17.
Author Address: Complex Carbohydrate Research Center, University of Georgia, Athens, GA, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1410
Author: Zhao, P., Viner, R., Teo, C. F., Boons, G. J., Horn, D. and Wells, L.
Year: 2011
Title: Combining high-energy C-trap dissociation and electron transfer dissociation for protein O-GlcNAc modification site assignment
Journal: J Proteome Res
Volume: 10
Issue: 9
Pages: 4088-104
Epub Date: 2011/07/12
Date: Sep 2
Short Title: Combining high-energy C-trap dissociation and electron transfer dissociation for protein O-GlcNAc modification site assignment
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/pr2002726
PMCID: PMC3172619
NIHMSID: NIHMS314262
Accession Number: 21740066
Keywords: Acetylglucosamine/*chemistry/metabolism
Amino Acid Sequence
Antibodies, Monoclonal/chemistry
Glycosylation
HEK293 Cells
Humans
Molecular Sequence Data
Peptide Fragments/*chemistry/metabolism
Protein Processing, Post-Translational
Proteomics/*methods
Tandem Mass Spectrometry/*methods
Abstract: Mass spectrometry-based studies of proteins that are post-translationally modified by O-linked β-N-acetylglucosamine (O-GlcNAc) are challenged in effectively identifying the sites of modification while simultaneously sequencing the peptides. Here we tested the hypothesis that a combination of high-energy C-trap dissociation (HCD) and electron transfer dissociation (ETD) could specifically target the O-GlcNAc modified peptides and elucidate the amino acid sequence while preserving the attached GlcNAc residue for accurate site assignment. By taking advantage of the recently characterized O-GlcNAc-specific IgG monoclonal antibodies and the combination of HCD and ETD fragmentation techniques, O-GlcNAc modified proteins were enriched from HEK293T cells and subsequently characterized using the LTQ Orbitrap Velos ETD (Thermo Fisher Scientific) mass spectrometer. In our data set, 83 sites of O-GlcNAc modification are reported with high confidence confirming that the HCD/ETD combined approach is amenable to the detection and site assignment of O-GlcNAc modified peptides. Realizing HCD triggered ETD fragmentation on a linear ion trap/Orbitrap platform for more in-depth analysis and application of this technique to other post-translationally modified proteins are currently underway. Furthermore, this report illustrates that the O-GlcNAc transferase appears to demonstrate promiscuity with regards to the hydroxyl-containing amino acid modified in short stretches of primary sequence of the glycosylated polypeptides.
Notes: 1535-3907
Zhao, Peng
Viner, Rosa
Teo, Chin Fen
Boons, Geert-Jan
Horn, David
Wells, Lance
P41RR018502/RR/NCRR NIH HHS/United States
P41 RR018502-10/RR/NCRR NIH HHS/United States
P41 RR018502/RR/NCRR NIH HHS/United States
R01 DK075069-05/DK/NIDDK NIH HHS/United States
R01DK075069/DK/NIDDK NIH HHS/United States
R01 DK075069/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
J Proteome Res. 2011 Sep 2;10(9):4088-104. doi: 10.1021/pr2002726. Epub 2011 Jul 25.
Author Address: Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602, United States.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1757
Author: Zhao, Y., Ren, J., Harlos, K. and Stuart, D. I.
Year: 2016
Title: Structure of glycosylated NPC1 luminal domain C reveals insights into NPC2 and Ebola virus interactions
Journal: FEBS Lett
Volume: 590
Issue: 5
Pages: 605-12
Epub Date: 2016/02/06
Date: Mar
Short Title: Structure of glycosylated NPC1 luminal domain C reveals insights into NPC2 and Ebola virus interactions
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1002/1873-3468.12089
PMCID: PMC4819692
Accession Number: 26846330
Keywords: Amino Acid Sequence
Animals
Carrier Proteins/*chemistry/*metabolism
Ebolavirus/*metabolism
Glycoproteins/*metabolism
Glycosylation
HEK293 Cells
Humans
Intracellular Signaling Peptides and Proteins
Membrane Glycoproteins/*chemistry/*metabolism
Models, Molecular
Molecular Sequence Data
Protein Binding
Protein Structure, Tertiary
Vesicular Transport Proteins
Ebola virus receptor
Ebola virus susceptibility
Npc1
Npc2
Niemann-Pick disease type C
cholesterol transport
Abstract: Niemann-pick type C1 (NPC1) is an endo/lysosomal membrane protein involved in intracellular cholesterol trafficking, and its luminal domain C is an essential endosomal receptor for Ebola and Marburg viruses. We have determined the crystal structure of glycosylated NPC1 luminal domain C and find all seven possible sites are glycosylated. Mapping the disease mutations onto the glycosylated structure reveals a potential binding face for NPC2. Knowledge-based docking of NPC1 onto Ebola viral glycoprotein and sequence analysis of filovirus susceptible and refractory species reveals four critical residues, H418, Q421, F502 and F504, some or all of which are likely responsible for the species-specific susceptibility to the virus infection.
Notes: 1873-3468
Zhao, Yuguang
Ren, Jingshan
Harlos, Karl
Stuart, David I
MR/N00065X/1/MRC_/Medical Research Council/United Kingdom
WT_/Wellcome Trust/United Kingdom
G1100525/MRC_/Medical Research Council/United Kingdom
090532/Z/09/Z/WT_/Wellcome Trust/United Kingdom
G1000099/MRC_/Medical Research Council/United Kingdom
Journal Article
Research Support, Non-U.S. Gov't
FEBS Lett. 2016 Mar;590(5):605-12. doi: 10.1002/1873-3468.12089. Epub 2016 Feb 23.
Author Address: Division of Structural Biology, University of Oxford, Headington, Oxford, UK.
Diamond Light Source Ltd, Didcot, UK.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 880
Author: Zhen, Y., Caprioli, R. M. and Staros, J. V.
Year: 2003
Title: Characterization of glycosylation sites of the epidermal growth factor receptor
Journal: Biochemistry
Volume: 42
Issue: 18
Pages: 5478-92
Epub Date: 2003/05/07
Date: May 13
Short Title: Characterization of glycosylation sites of the epidermal growth factor receptor
Alternate Journal: Biochemistry
ISSN: 0006-2960 (Print)
0006-2960
DOI: 10.1021/bi027101p
PMCID: PMC2765783
NIHMSID: NIHMS118921
Accession Number: 12731890
Keywords: Amino Acid Sequence
Animals
Binding Sites/genetics
CHO Cells
Carbohydrate Conformation
Carbohydrate Sequence
Cells, Cultured/metabolism
Chromatography, High Pressure Liquid
Cricetinae
ErbB Receptors/*chemistry/*metabolism
Gas Chromatography-Mass Spectrometry
Glycosylation
Humans
Lung Neoplasms/metabolism
Molecular Sequence Data
Peptide Fragments/chemistry
Peptide Mapping
Protein Binding
Recombinant Proteins/chemistry/isolation & purification/metabolism
Trypsin/pharmacology
Abstract: The epidermal growth factor receptor is a transmembrane glycoprotein that mediates the cellular responses to epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha). In this study of the human EGF receptor naturally expressed in A431 cells, the glycosylation sites of the full-length, membrane-bound receptor and of a secreted form of the receptor were characterized by mass spectrometry. Our data show that the naturally expressed human EGF receptor is fully glycosylated on eight of the 11 canonical sites; two of the sites are not glycosylated, and one is partially glycosylated, a pattern of site-usage similar but not identical to those reported for the recombinant human EGF receptor heterologously expressed in Chinese hamster ovary cells. We also confirm the partial glycosylation of an atypical NNC site first identified in the receptor expressed in Chinese hamster ovary cells. We show that an additional canonical site in the secreted form of the receptor is fully glycosylated. While the pattern of glycosylation is the same for the sites shared by the full-length and the secreted forms of the receptor, the oligosaccharides of the full-length receptor are more extensively processed. Finally, we provide evidence that in addition to the known secreted form of the receptor, a proteolytic cleavage product of the receptor corresponding to the full extracytoplasmic, ligand-binding domain is present in the conditioned medium.
Notes: 1520-4995
Zhen, Yuejun
Caprioli, Richard M
Staros, James V
R01 GM58008/GM/NIGMS NIH HHS/United States
R01 GM058008/GM/NIGMS NIH HHS/United States
R01 GM058008-04/GM/NIGMS NIH HHS/United States
R01 GM058008-07/GM/NIGMS NIH HHS/United States
R01 GM058008-02/GM/NIGMS NIH HHS/United States
R01 GM058008-05/GM/NIGMS NIH HHS/United States
R01 GM058008-09/GM/NIGMS NIH HHS/United States
R01 GM55056/GM/NIGMS NIH HHS/United States
R01 GM058008-10/GM/NIGMS NIH HHS/United States
R01 GM055056/GM/NIGMS NIH HHS/United States
R01 GM058008-06/GM/NIGMS NIH HHS/United States
R01 GM058008-03/GM/NIGMS NIH HHS/United States
R01 DK25489/DK/NIDDK NIH HHS/United States
R01 GM058008-08/GM/NIGMS NIH HHS/United States
Comparative Study
Journal Article
Research Support, U.S. Gov't, P.H.S.
Biochemistry. 2003 May 13;42(18):5478-92. doi: 10.1021/bi027101p.
Author Address: Department of Biological Sciences, Vanderbilt University, Nashville, Tennessee 37235, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1724
Author: Zheng, Y., Li, X., Qian, X., Wang, Y., Lee, J. H., Xia, Y., Hawke, D. H., Zhang, G., Lyu, J. and Lu, Z.
Year: 2015
Title: Secreted and O-GlcNAcylated MIF binds to the human EGF receptor and inhibits its activation
Journal: Nat Cell Biol
Volume: 17
Issue: 10
Pages: 1348-55
Epub Date: 2015/08/19
Date: Oct
Short Title: Secreted and O-GlcNAcylated MIF binds to the human EGF receptor and inhibits its activation
Alternate Journal: Nature cell biology
ISSN: 1465-7392 (Print)
1465-7392
DOI: 10.1038/ncb3222
PMCID: PMC4785887
NIHMSID: NIHMS759610
Accession Number: 26280537
Keywords: Acetylglucosamine/*metabolism
Animals
Cell Line
Cell Line, Tumor
Cell Proliferation/drug effects
Enzyme Activation/drug effects
Epidermal Growth Factor/metabolism/pharmacology
ErbB Receptors/genetics/*metabolism
Extracellular Signal-Regulated MAP Kinases/metabolism
Glioma/genetics/metabolism/pathology
Humans
Immunoblotting
JNK Mitogen-Activated Protein Kinases/metabolism
Macrophage Migration-Inhibitory Factors/genetics/*metabolism
Matrix Metalloproteinase 13/metabolism
Mice, Nude
Microscopy, Fluorescence
Mutation
Phosphorylation/drug effects
Protein Binding
RNA Interference
Serine/metabolism
Survival Analysis
Threonine/genetics/metabolism
Transplantation, Heterologous
Abstract: Activation of epidermal growth factor receptor (EGFR), which occurs in many types of tumour, promotes tumour progression. However, no extracellular antagonist of human EGFR has been identified. We found that human macrophage migration inhibitory factor (MIF) is O-GlcNAcylated at Ser 112/Thr 113 at its carboxy terminus. The naturally secreted and O-GlcNAcylated MIF binds to EGFR, thereby inhibiting the binding of EGF to EGFR and EGF-induced EGFR activation, phosphorylation of ERK and c-Jun, cell invasion, proliferation and brain tumour formation. Activation of EGFR through mutation or its ligand binding enhances the secretion of MMP13, which degrades extracellular MIF, and results in abrogation of the negative regulation of MIF on EGFR. The finding that EGFR activation downregulates its antagonist in the tumour microenvironment represents an important feedforward mechanism for human tumour cells to enhance EGFR signalling and promote tumorigenesis.
Notes: 1476-4679
Zheng, Yanhua
Li, Xinjian
Qian, Xu
Wang, Yugang
Lee, Jong-Ho
Xia, Yan
Hawke, David H
Zhang, Gang
Lyu, Jianxin
Lu, Zhimin
P30 CA016672/CA/NCI NIH HHS/United States
1R0 CA169603/CA/NCI NIH HHS/United States
1R01 NS089754/NS/NINDS NIH HHS/United States
1S10 OD012304-01/OD/NIH HHS/United States
CA016672/CA/NCI NIH HHS/United States
2R01 CA109035/CA/NCI NIH HHS/United States
R01 CA169603/CA/NCI NIH HHS/United States
R01 CA109035/CA/NCI NIH HHS/United States
2P50 CA127001/CA/NCI NIH HHS/United States
S10 OD012304/OD/NIH HHS/United States
R01 NS089754/NS/NINDS NIH HHS/United States
P50 CA127001/CA/NCI NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Nat Cell Biol. 2015 Oct;17(10):1348-55. doi: 10.1038/ncb3222. Epub 2015 Aug 17.
Author Address: Department of Neuro-Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.
Department of Systems Biology, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.
Department of Surgical Oncology, Affiliated Hospital of Hebei University, Baoding, Hebei 071000, China.
Key Laboratory of Laboratory Medicine, Ministry of Education, Zhejiang Provincial Key Laboratory of Medical Genetics, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China.
Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.
Cancer Biology Program, The University of Texas Graduate School of Biomedical Sciences at Houston, Houston, Texas 77030, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2074
Author: Zhou, D., Tian, X., Qi, R., Peng, C. and Zhang, W.
Year: 2021
Title: Identification of 22 N-glycosites on spike glycoprotein of SARS-CoV-2 and accessible surface glycopeptide motifs: Implications for vaccination and antibody therapeutics
Journal: Glycobiology
Volume: 31
Issue: 1
Pages: 69-80
Epub Date: 2020/06/11
Date: Jan 9
Short Title: Identification of 22 N-glycosites on spike glycoprotein of SARS-CoV-2 and accessible surface glycopeptide motifs: Implications for vaccination and antibody therapeutics
Alternate Journal: Glycobiology
ISSN: 0959-6658 (Print)
0959-6658
DOI: 10.1093/glycob/cwaa052
PMCID: PMC7313968
Accession Number: 32518941
Keywords: Amino Acid Motifs
*Antibodies, Viral/immunology/therapeutic use
COVID-19/immunology/*therapy
*COVID-19 Vaccines/chemistry/immunology
*Glycopeptides/chemistry/immunology
Humans
Immunization, Passive
*SARS-CoV-2/chemistry/immunology
*Spike Glycoprotein, Coronavirus/chemistry/metabolism
*antibody
*cryogenic electron microscopy structure
*crystal structures
*epitope prediction
*glycopeptide/SARS-CoV-2
Abstract: Coronaviruses hijack human enzymes to assemble the sugar coat on their spike glycoproteins. The mechanisms by which human antibodies may recognize the antigenic viral peptide epitopes hidden by the sugar coat are unknown. Glycosylation by insect cells differs from the native form produced in human cells, but insect cell-derived influenza vaccines have been approved by the US Food and Drug Administration. In this study, we analyzed recombinant severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein secreted from BTI-Tn-5B1-4 insect cells, by trypsin and chymotrypsin digestion followed by mass spectrometry analysis. We acquired tandem mass spectrometry (MS/MS) spectrums for glycopeptides of all 22 predicted N-glycosylated sites. We further analyzed the surface accessibility of spike proteins according to cryogenic electron microscopy and homolog-modeled structures and available antibodies that bind to SARS-CoV-1. All 22 N-glycosylated sites of SARS-CoV-2 are modified by high-mannose N-glycans. MS/MS fragmentation clearly established the glycopeptide identities. Electron densities of glycans cover most of the spike receptor-binding domain of SARS-CoV-2, except YQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQ, similar to a region FSPDGKPCTPPALNCYWPLNDYGFYTTTGIGYQ in SARS-CoV-1. Other surface-exposed domains include those located on central helix, connecting region, heptad repeats and N-terminal domain. Because the majority of antibody paratopes bind to the peptide portion with or without sugar modification, we propose a snake-catching model for predicted paratopes: a minimal length of peptide is first clamped by a paratope and sugar modifications close to the peptide either strengthen or do not hinder the binding.
Notes: 1460-2423
Zhou, Dapeng
Tian, Xiaoxu
Qi, Ruibing
Peng, Chao
Zhang, Wen
Journal Article
Research Support, Non-U.S. Gov't
Glycobiology. 2021 Jan 9;31(1):69-80. doi: 10.1093/glycob/cwaa052.
Author Address: Tongji University School of Medicine, 1239 Siping Road, Shanghai 200092, China.
Shanghai Pudong New Area Mental Health Center affiliated with Tongji University School of Medicine, 165 Sanlin Road, Shanghai 200124, China.
National Facility for Protein Science in Shanghai, Zhangjiang Lab, Shanghai Advanced Research Institute, Chinese Academy of Science, 333 Haike Road, Shanghai 201210, China.
Innovation Team of Small Animal Infectious Disease, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Science, 518 Ziyue Road, Shanghai 200241, China.
Fudan University Pudong Medical Center, Institutes of Biomedical Sciences, 200433 Gongwei Road, Shanghai, China.
Department of Systems Biology for Medicine, Shanghai Medical College, Fudan University, 138 Yixueyuan Road, Shanghai 200032, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1765
Author: Zhou, F., Huo, J., Liu, Y., Liu, H., Liu, G., Chen, Y. and Chen, B.
Year: 2016
Title: Elevated glucose levels impair the WNT/β-catenin pathway via the activation of the hexosamine biosynthesis pathway in endometrial cancer
Journal: J Steroid Biochem Mol Biol
Volume: 159
Pages: 19-25
Epub Date: 2016/03/01
Date: May
Short Title: Elevated glucose levels impair the WNT/β-catenin pathway via the activation of the hexosamine biosynthesis pathway in endometrial cancer
Alternate Journal: The Journal of steroid biochemistry and molecular biology
ISSN: 0960-0760
DOI: 10.1016/j.jsbmb.2016.02.015
Accession Number: 26923859
Keywords: *Biosynthetic Pathways
Cell Line, Tumor
Endometrial Neoplasms/*metabolism
Female
Glucose/*physiology
Glycosylation
Hexosamines/*biosynthesis
Humans
Protein Processing, Post-Translational
*Wnt Signaling Pathway
beta Catenin/metabolism
Elevated glucose levels
Endometrial cancer
O-GlcNAc
WNT/β-catenin
Abstract: Endometrial cancer (EC) is one of the most common gynecological malignancies in the world. Associations between fasting glucose levels (greater than 5.6mmol/L) and the risk of cancer fatality have been reported. However, the underlying link between glucose metabolic disease and EC remains unclear. In the present study, we explored the influence of elevated glucose levels on the WNT/β-catenin pathway in EC. Previous studies have suggested that elevated concentrations of glucose can drive the hexosamine biosynthesis pathway (HBP) flux, thereby enhancing the O-GlcNAc modification of proteins. Here, we cultured EC cell lines, AN3CA and HEC-1-B, with various concentrations of glucose. Results showed that when treated with high levels of glucose, both lines showed increased expression of β-catenin and O-GlcNAcylation levels; however, these effects could be abolished by the HBP inhibitors, Azaserine and 6-Diazo-5-oxo-l-norleucine, and be restored by glucosamine. Moreover the AN3CA and HEC-1-B cells that were cultured with or without PUGNAc, an inhibitor of the O-GlcNAcase, showed that PUGNAc increased β-catenin levels. The results suggest that elevated glucose levels increase β-catenin expression via the activation of the HBP in EC cells. Subcellular fractionation experiments showed that AN3CA cells had a higher expression of intranuclear β-catenin in high glucose medium. Furthermore, TOP/FOP-Flash and RT-PCR results showed that glucose-induced increased expression of β-catenin triggered the transcription of target genes. In conclusion, elevated glucose levels, via HBP, increase the O-GlcNAcylation level, thereby inducing the over expression of β-catenin and subsequent transcription of the target genes in EC cells.
Notes: 1879-1220
Zhou, Fuxing
Huo, Junwei
Liu, Yu
Liu, Haixia
Liu, Gaowei
Chen, Ying
Chen, Biliang
Journal Article
England
J Steroid Biochem Mol Biol. 2016 May;159:19-25. doi: 10.1016/j.jsbmb.2016.02.015. Epub 2016 Feb 26.
Author Address: Department of Gynecology and Obstetrics, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi 710032, China.
Department of Gynecology and Obstetrics, The First Hospital of Yulin, Yulin, Shaanxi 718000, China.
Department of Natural Medicine, School of Pharmacy, The Fourth Military Medical University, Xi'an, Shaanxi 710032, China.
Department of Gynecology and Obstetrics, Xijing Hospital, The Fourth Military Medical University, Xi'an, Shaanxi 710032, China. Electronic address: chenbiliang1962@sina.com.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1169
Author: Zhou, F., Su, J., Fu, L., Yang, Y., Zhang, L., Wang, L., Zhao, H., Zhang, D., Li, Z. and Zha, X.
Year: 2008
Title: Unglycosylation at Asn-633 made extracellular domain of E-cadherin folded incorrectly and arrested in endoplasmic reticulum, then sequentially degraded by ERAD
Journal: Glycoconj J
Volume: 25
Issue: 8
Pages: 727-40
Epub Date: 2008/05/21
Date: Nov
Short Title: Unglycosylation at Asn-633 made extracellular domain of E-cadherin folded incorrectly and arrested in endoplasmic reticulum, then sequentially degraded by ERAD
Alternate Journal: Glycoconjugate journal
ISSN: 0282-0080 (Print)
0282-0080
DOI: 10.1007/s10719-008-9133-9
Accession Number: 18491227
Keywords: Asparagine/chemistry
Base Sequence
Binding Sites/genetics
Cadherins/*chemistry/genetics/*metabolism
Cell Line, Tumor
DNA Primers/genetics
Endoplasmic Reticulum/*metabolism
Female
Glycosylation
Golgi Apparatus/metabolism
Humans
Models, Molecular
Mutagenesis, Site-Directed
Proteasome Endopeptidase Complex/metabolism
Protein Folding
Protein Structure, Tertiary
Recombinant Proteins/chemistry/genetics/metabolism
Ubiquitin/metabolism
Abstract: The human E-cadherin is a single transmembrane domain protein involved in Ca(2+)-dependent cell-cell adhesion. In a previous study, we demonstrated that all of four potential N-glycosylation sites in E-cadherin are occupied by N-glycans in human breast carcinoma cells in vivo and the elimination of N-glycan at Asn-633 dramatically affected E-cadherin expression and made it degraded. In this study we investigated the molecular mechanism of E-cadherin, which lacks N-glycosylation at Asn-633 (M4), degradation and the role of the N-glycan at Asn-633 in E-cadherin folding. We treated cells stably expressed M4 E-cadherin with MG123, DMM, respectively. Either MG132 or DMM could efficiently block degradation of M4 E-cadherin. M4 E-cadherin was recognized as the substrate of ERAD and was retro-translocated from ER lumen to cytoplasm by p97. It was observed that the ration of M4 E-cadherin binding to calnexin was significantly increased compared with that of other variants, suggesting that it was a misfolded protein, though cytoplasmic domain of M4 E-cadherin could associate with beta-catenin. Furthermore, we found that N-glycans of M4 E-cadherin were modified in immature high mannose type, suggesting that it could not depart to Golgi apparatus. In conclusion, this study revealed that N-glycosylation at Asn-633 is essential for E-cadherin expression, folding and trafficking.
Notes: Zhou, Feng
Su, Jianmin
Fu, Le
Yang, Yong
Zhang, Lineng
Wang, Liying
Zhao, Hongbo
Zhang, Diancai
Li, Zengxia
Zha, Xiliang
Journal Article
Research Support, Non-U.S. Gov't
United States
Glycoconj J. 2008 Nov;25(8):727-40. doi: 10.1007/s10719-008-9133-9. Epub 2008 May 20.
Author Address: Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai, 200032, People's Republic of China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1447
Author: Zhou, K., Kanai, R., Lee, P., Wang, H. W. and Modis, Y.
Year: 2012
Title: Toll-like receptor 5 forms asymmetric dimers in the absence of flagellin
Journal: J Struct Biol
Volume: 177
Issue: 2
Pages: 402-9
Epub Date: 2011/12/17
Date: Feb
Short Title: Toll-like receptor 5 forms asymmetric dimers in the absence of flagellin
Alternate Journal: Journal of structural biology
ISSN: 1047-8477
DOI: 10.1016/j.jsb.2011.12.002
Accession Number: 22173220
Keywords: Flagellin/*chemistry
Glycosylation
Humans
Models, Molecular
Particle Size
Protein Binding
Protein Interaction Domains and Motifs
Protein Multimerization
Protein Processing, Post-Translational
Protein Structure, Quaternary
Structural Homology, Protein
Surface Properties
Toll-Like Receptor 5/*chemistry
Abstract: The structure of full-length human TLR5 determined by electron microscopy single-particle image reconstruction at 26Å resolution shows that TLR5 forms an asymmetric homodimer via ectodomain interactions. The structure shows that like TLR9, TLR5 dimerizes in the absence of ligand. The asymmetry of the dimer suggests that TLR5 may recognize two flagellin molecules cooperatively to establish an optimal flagellin response threshold. A TLR5 homology model was generated and fitted into the electron microscopy structure. All seven predicted N-linked glycosylation sites are exposed on the molecular surface, away from the dimer interface. Glycosylation at the first five sites was confirmed by tandem mass spectrometry. Two aspartate residues proposed to interact with flagellin (Asp294 and Asp366) are sterically occluded by a glycan at position 342. In contrast, the central region of the ectodomains near the dimer interface is unobstructed by glycans. Ligand binding in this region would be consistent with the ligand binding sites of other TLRs.
Notes: 1095-8657
Zhou, Kaifeng
Kanai, Ryuta
Lee, Phong
Wang, Hong-Wei
Modis, Yorgo
Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
United States
J Struct Biol. 2012 Feb;177(2):402-9. doi: 10.1016/j.jsb.2011.12.002. Epub 2011 Dec 8.
Author Address: Department of Molecular Biophysics & Biochemistry, Yale University, New Haven, CT 06520, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1995
Author: Zhou, R., Yang, G., Guo, X., Zhou, Q., Lei, J. and Shi, Y.
Year: 2019
Title: Recognition of the amyloid precursor protein by human γ-secretase
Journal: Science
Volume: 363
Issue: 6428
Epub Date: 2019/01/12
Date: Feb 15
Short Title: Recognition of the amyloid precursor protein by human γ-secretase
Alternate Journal: Science (New York, N.Y.)
ISSN: 0036-8075
DOI: 10.1126/science.aaw0930
Accession Number: 30630874
Keywords: Alzheimer Disease/metabolism
Amyloid Precursor Protein Secretases/*chemistry
Amyloid beta-Protein Precursor/*chemistry
*Catalytic Domain
Cryoelectron Microscopy
Humans
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
*Proteolysis
Receptors, Notch/chemistry
Abstract: Cleavage of amyloid precursor protein (APP) by the intramembrane protease γ-secretase is linked to Alzheimer's disease (AD). We report an atomic structure of human γ-secretase in complex with a transmembrane (TM) APP fragment at 2.6-angstrom resolution. The TM helix of APP closely interacts with five surrounding TMs of PS1 (the catalytic subunit of γ-secretase). A hybrid β sheet, which is formed by a β strand from APP and two β strands from PS1, guides γ-secretase to the scissile peptide bond of APP between its TM and β strand. Residues at the interface between PS1 and APP are heavily targeted by recurring mutations from AD patients. This structure, together with that of γ-secretase bound to Notch, reveal contrasting features of substrate binding, which may be applied toward the design of substrate-specific inhibitors.
Notes: 1095-9203
Zhou, Rui
Orcid: 0000-0002-0782-4761
Yang, Guanghui
Orcid: 0000-0002-6835-1611
Guo, Xuefei
Orcid: 0000-0001-7135-0954
Zhou, Qiang
Lei, Jianlin
Orcid: 0000-0002-9384-8742
Shi, Yigong
Orcid: 0000-0003-2030-168x
Journal Article
United States
Science. 2019 Feb 15;363(6428):eaaw0930. doi: 10.1126/science.aaw0930. Epub 2019 Jan 10.
Author Address: Beijing Advanced Innovation Center for Structural Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Institute of Biology, Westlake Institute for Advanced Study, Westlake University, 18 Shilongshan Road, Xihu District, Hangzhou 310024, Zhejiang Province, China.
School of Life Sciences, Westlake University, 18 Shilongshan Road, Xihu District, Hangzhou 310024, Zhejiang Province, China.
Technology Center for Protein Sciences, Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Beijing Advanced Innovation Center for Structural Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China. shi-lab@tsinghua.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1891
Author: Zhou, X., Li, M., Su, D., Jia, Q., Li, H., Li, X. and Yang, J.
Year: 2017
Title: Cryo-EM structures of the human endolysosomal TRPML3 channel in three distinct states
Journal: Nat Struct Mol Biol
Volume: 24
Issue: 12
Pages: 1146-1154
Epub Date: 2017/11/07
Date: Dec
Short Title: Cryo-EM structures of the human endolysosomal TRPML3 channel in three distinct states
Alternate Journal: Nature structural & molecular biology
ISSN: 1545-9993 (Print)
1545-9985
DOI: 10.1038/nsmb.3502
PMCID: PMC5747366
NIHMSID: NIHMS928247
Accession Number: 29106414
Keywords: Animals
Binding Sites/physiology
Cell Line
Cryoelectron Microscopy/*methods
Endosomes/*metabolism
HEK293 Cells
Humans
Ion Channel Gating/physiology
Lysosomes/metabolism
Models, Molecular
Protein Domains/*physiology
Protein Structure, Secondary
Sf9 Cells
Spodoptera
Transient Receptor Potential Channels/genetics/*metabolism
Abstract: TRPML3 channels are mainly localized to endolysosomes and play a critical role in the endocytic pathway. Their dysfunction causes deafness and pigmentation defects in mice. TRPML3 activity is inhibited by low endolysosomal pH. Here we present cryo-electron microscopy (cryo-EM) structures of human TRPML3 in the closed, agonist-activated, and low-pH-inhibited states, with resolutions of 4.06, 3.62, and 4.65 Å, respectively. The agonist ML-SA1 lodges between S5 and S6 and opens an S6 gate. A polycystin-mucolipin domain (PMD) forms a luminal cap. S1 extends into this cap, forming a 'gating rod' that connects directly to a luminal pore loop, which undergoes dramatic conformational changes in response to low pH. S2 extends intracellularly and interacts with several intracellular regions to form a 'gating knob'. These unique structural features, combined with the results of electrophysiological studies, indicate a new mechanism by which luminal pH and other physiological modulators such as PIP(2) regulate TRPML3 by changing S1 and S2 conformations.
Notes: 1545-9985
Zhou, Xiaoyuan
Li, Minghui
Su, Deyuan
Jia, Qi
Li, Huan
Li, Xueming
Yang, Jian
R01 GM085234/GM/NIGMS NIH HHS/United States
Journal Article
Nat Struct Mol Biol. 2017 Dec;24(12):1146-1154. doi: 10.1038/nsmb.3502. Epub 2017 Nov 6.
Author Address: Beijing Advanced Innovation Center for Structural Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China.
Department of Biological Sciences, Columbia University, New York, New York, USA.
Key Laboratory of Animal Models and Human Disease Mechanisms of Chinese Academy of Sciences, Key Laboratory of Bioactive Peptides of Yunnan Province, and Ion Channel Research and Drug Development Center, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China.
Department of Orthopedic Oncology, Shanghai Changzheng Hospital, The Second Military Medical University, Shanghai, China.
Kunming College of Life Science, University of Chinese Academy of Sciences, Kunming, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1875
Author: Zhou, X. E., He, Y., de Waal, P. W., Gao, X., Kang, Y., Van Eps, N., Yin, Y., Pal, K., Goswami, D., White, T. A., Barty, A., Latorraca, N. R., Chapman, H. N., Hubbell, W. L., Dror, R. O., Stevens, R. C., Cherezov, V., Gurevich, V. V., Griffin, P. R., Ernst, O. P., Melcher, K. and Xu, H. E.
Year: 2017
Title: Identification of Phosphorylation Codes for Arrestin Recruitment by G Protein-Coupled Receptors
Journal: Cell
Volume: 170
Issue: 3
Pages: 457-469.e13
Epub Date: 2017/07/29
Date: Jul 27
Short Title: Identification of Phosphorylation Codes for Arrestin Recruitment by G Protein-Coupled Receptors
Alternate Journal: Cell
ISSN: 0092-8674 (Print)
0092-8674
DOI: 10.1016/j.cell.2017.07.002
PMCID: PMC5567868
NIHMSID: NIHMS891443
Accession Number: 28753425
Keywords: Amino Acid Sequence
Animals
Arrestins/*chemistry/metabolism
Chromatography, Liquid
Humans
Mice
Models, Molecular
Phosphorylation
Rats
Rhodopsin/*chemistry/metabolism
Sequence Alignment
Tandem Mass Spectrometry
X-Rays
Gpcr
Grk
arrestin
biased signaling
drug discovery
membrane proteins
phosphorylation codes
rhodopsin
Abstract: G protein-coupled receptors (GPCRs) mediate diverse signaling in part through interaction with arrestins, whose binding promotes receptor internalization and signaling through G protein-independent pathways. High-affinity arrestin binding requires receptor phosphorylation, often at the receptor's C-terminal tail. Here, we report an X-ray free electron laser (XFEL) crystal structure of the rhodopsin-arrestin complex, in which the phosphorylated C terminus of rhodopsin forms an extended intermolecular β sheet with the N-terminal β strands of arrestin. Phosphorylation was detected at rhodopsin C-terminal tail residues T336 and S338. These two phospho-residues, together with E341, form an extensive network of electrostatic interactions with three positively charged pockets in arrestin in a mode that resembles binding of the phosphorylated vasopressin-2 receptor tail to β-arrestin-1. Based on these observations, we derived and validated a set of phosphorylation codes that serve as a common mechanism for phosphorylation-dependent recruitment of arrestins by GPCRs.
Notes: 1097-4172
Zhou, X Edward
He, Yuanzheng
de Waal, Parker W
Gao, Xiang
Kang, Yanyong
Van Eps, Ned
Yin, Yanting
Pal, Kuntal
Goswami, Devrishi
White, Thomas A
Barty, Anton
Latorraca, Naomi R
Chapman, Henry N
Hubbell, Wayne L
Dror, Ron O
Stevens, Raymond C
Cherezov, Vadim
Gurevich, Vsevolod V
Griffin, Patrick R
Ernst, Oliver P
Melcher, Karsten
Xu, H Eric
R01 DK066202/DK/NIDDK NIH HHS/United States
R01 GM109955/GM/NIGMS NIH HHS/United States
T32 GM008294/GM/NIGMS NIH HHS/United States
R01 EY005216/EY/NEI NIH HHS/United States
R01 DK071662/DK/NIDDK NIH HHS/United States
R01 GM108635/GM/NIGMS NIH HHS/United States
R01 EY011500/EY/NEI NIH HHS/United States
R01 GM087413/GM/NIGMS NIH HHS/United States
R35 GM122491/GM/NIGMS NIH HHS/United States
Journal Article
Cell. 2017 Jul 27;170(3):457-469.e13. doi: 10.1016/j.cell.2017.07.002.
Author Address: VARI-SIMM Center, Center for Structure and Function of Drug Targets, CAS-Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China; Laboratory of Structural Sciences, Center for Structural Biology and Drug Discovery, Van Andel Research Institute, Grand Rapids, MI 49503, USA.
Laboratory of Structural Sciences, Center for Structural Biology and Drug Discovery, Van Andel Research Institute, Grand Rapids, MI 49503, USA.
Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada.
Department of Molecular Medicine, The Scripps Research Institute, Scripps Florida, Jupiter, FL 33458, USA.
Center for Free Electron Laser Science, Deutsches Elektronen-Synchrotron DESY, 22607 Hamburg, Germany.
Department of Computer Science, Stanford University, Stanford, CA 94305, USA; Department of Molecular and Cellular Physiology, Stanford University, Stanford, CA 94305, USA; Department of Structural Biology, Stanford University, Stanford, CA 94305, USA; Institute for Computational and Mathematical Engineering, Stanford University, Stanford, CA 94305, USA; Biophysics Program, Stanford University, Stanford, CA 94305, USA.
Center for Free Electron Laser Science, Deutsches Elektronen-Synchrotron DESY, 22607 Hamburg, Germany; Centre for Ultrafast Imaging, 22761 Hamburg, Germany.
Jules Stein Eye Institute and Department of Chemistry and Biochemistry, University of California, Los Angeles, CA 90095, USA.
Department of Chemistry, Bridge Institute, University of Southern California, Los Angeles, CA 90089, USA; iHuman Institute, ShanghaiTech University, 2F Building 6, 99 Haike Road, Pudong New District, Shanghai 201210, China.
Department of Chemistry, Bridge Institute, University of Southern California, Los Angeles, CA 90089, USA.
Department of Pharmacology, Vanderbilt University, Nashville, TN 37232, USA.
Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada.
VARI-SIMM Center, Center for Structure and Function of Drug Targets, CAS-Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China; Laboratory of Structural Sciences, Center for Structural Biology and Drug Discovery, Van Andel Research Institute, Grand Rapids, MI 49503, USA. Electronic address: eric.xu@vai.org.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 961
Author: Zhou, Y. B., Liu, F., Zhu, Z. D., Zhu, H., Zhang, X., Wang, Z. Q., Liu, J. H. and Han, Z. G.
Year: 2004
Title: N-glycosylation is required for efficient secretion of a novel human secreted glycoprotein, hPAP21
Journal: FEBS Lett
Volume: 576
Issue: 3
Pages: 401-7
Epub Date: 2004/10/23
Date: Oct 22
Short Title: N-glycosylation is required for efficient secretion of a novel human secreted glycoprotein, hPAP21
Alternate Journal: FEBS letters
ISSN: 0014-5793 (Print)
0014-5793
DOI: 10.1016/j.febslet.2004.09.039
Accession Number: 15498570
Keywords: Amino Acid Sequence
Animals
Base Sequence
Chromosomes, Human, Pair 2/*genetics
Conserved Sequence
Cricetinae
DNA Primers
Glycoproteins/*genetics/metabolism
Glycosylation
Humans
Molecular Sequence Data
Molecular Weight
Open Reading Frames
Phylogeny
Reverse Transcriptase Polymerase Chain Reaction
Sequence Alignment
Sequence Homology, Amino Acid
Abstract: The present study reported the isolation and characterization of a novel human secreted protein, named as hPAP21 (human protease-associated domain-containing protein, 21 kDa), encoded by the hypothetical gene chromosome 2 open reading frame 7 (C2orf7) that contains signal peptide in its N-terminus, without transmembrane domain, except for PA domain in its middle. Western blotting assay indicated that the c-Myc tagged hPAP21 could be secreted into culture medium in the transfected Chinese hamster ovary cells. However, the molecular weights, whatever intracellular (28 kDa) or extracellular (30 kDa) forms, are larger than that of the prediction. To define whether the glycosylation was important process for its secretion, endoglycosidase H (Endo H) and PNGase F (PNG F) were employed to evaluate the effect of glycosylation types on secretion of hPAP21. Interestingly, the extracellular forms were primarily sensitive to PNG F, not Endo H, implying that complex N-glycosylation could be required for the secretion of hPAP21. Furthermore, N-glycosylation of Asn171 was confirmed as potential crucial process for the secretory protein via site-directed mutagenesis assay. All data will be contributed to the understanding of molecular functions of hPAP21.
Notes: Zhou, Yu-Bo
Liu, Feng
Zhu, Zhi-Dong
Zhu, Hong
Zhang, Xin
Wang, Zhi-Qin
Liu, Jian-Hua
Han, Ze-Guang
Journal Article
Research Support, Non-U.S. Gov't
England
FEBS Lett. 2004 Oct 22;576(3):401-7. doi: 10.1016/j.febslet.2004.09.039.
Author Address: College of Life Science and Biotechnology, Shanghai Jiaotong University, 1954 Huashan Road, Shanghai 200030, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2003
Author: Zhu, G., Qian, M., Lu, L., Chen, Y., Zhang, X., Wu, Q., Liu, Y., Bian, Z., Yang, Y., Guo, S., Wang, J., Pan, Q. and Sun, F.
Year: 2019
Title: O-GlcNAcylation of YY1 stimulates tumorigenesis in colorectal cancer cells by targeting SLC22A15 and AANAT
Journal: Carcinogenesis
Epub Date: 2019/02/05
Date: Jan 30
Short Title: O-GlcNAcylation of YY1 stimulates tumorigenesis in colorectal cancer cells by targeting SLC22A15 and AANAT
Alternate Journal: Carcinogenesis
ISSN: 0143-3334
DOI: 10.1093/carcin/bgz010
Accession Number: 30715269
Abstract: Emerging studies have revealed that O-GlcNAcylation plays pivotal roles in the tumorigenesis of colorectal cancers. However, the underlying mechanism still remains largely unknown. Here, we demonstrated that YY1 was O-GlcNAcylated by OGT and O-GlcNAcylation of YY1 could increase the protein expression by enhancing its stability. O-GlcNAcylation facilitated transformative phenotypes of CRC cell in a YY1-dependent manner. Also, O-GlcNAcylation stimulates YY1-dependent transcriptional activity. Besides, we also identified the oncoproteins, SLC22A15 and AANAT, which were regulated by YY1 directly, are responsible for the YY1 stimulated tumorigenesis. Furthermore, we identified the main putative O-GlcNAc site of YY1 at Thr236, and mutating of this site decreased the pro-tumorigenic capacities of YY1. We concluded that O-GlcNAcylation of YY1 stimulates tumorigenesis in CRC cells by targeting SLC22A15 and AANAT, suggesting that YY1 O-GlcNAcylation might be a potential effective therapeutic target for treating CRC.
Notes: 1460-2180
Zhu, Guoqing
Qian, Mingping
Lu, Liesheng
Chen, Yan
Zhang, Xiao
Wu, Qi
Liu, Ya
Bian, Zhixuan
Yang, Yueyue
Guo, Susu
Wang, Jiayi
Pan, Qiuhui
Sun, Fenyong
Journal Article
England
Carcinogenesis. 2019 Jan 30. doi: 10.1093/carcin/bgz010.
Author Address: Department of Clinical Laboratory, Shanghai Tenth People's Hospital of Tongji University, Shanghai, China.
Department of General Surgery, Shanghai Tenth People's Hospital of Tongji University, Shanghai, China.
Department of Clinical Medicine, Shanghai Children's Medical Center, Shanghai Jiaotong University School of Medicine, Shanghai, China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1773
Author: Zhu, G., Tao, T., Zhang, D., Liu, X., Qiu, H., Han, L., Xu, Z., Xiao, Y., Cheng, C. and Shen, A.
Year: 2016
Title: O-GlcNAcylation of histone deacetylases 1 in hepatocellular carcinoma promotes cancer progression
Journal: Glycobiology
Volume: 26
Issue: 8
Pages: 820-833
Epub Date: 2016/04/10
Date: Aug
Short Title: O-GlcNAcylation of histone deacetylases 1 in hepatocellular carcinoma promotes cancer progression
Alternate Journal: Glycobiology
ISSN: 0959-6658
DOI: 10.1093/glycob/cww025
Accession Number: 27060025
Keywords: Acylation
Antigens, CD
Cadherins/genetics/metabolism
Carcinoma, Hepatocellular/enzymology/*genetics/pathology
Cell Movement
Cell Proliferation
Cyclin-Dependent Kinase Inhibitor p21/genetics/metabolism
Disease Progression
*Gene Expression Regulation, Neoplastic
HEK293 Cells
Hep G2 Cells
Histone Deacetylase 1/*genetics/metabolism
Histones/genetics/metabolism
Humans
Liver Neoplasms/enzymology/*genetics/pathology
N-Acetylglucosaminyltransferases/*genetics/metabolism
*Protein Processing, Post-Translational
*O-GlcNAc transferase
*O-GlcNAcylation
*cell proliferation
*hepatocellular carcinoma
*histone deacetylase-1
Abstract: Hepatocellular carcinoma (HCC) is a malignant tumor originating in the liver. Previous studies have indicated that O-GlcNAc transferase (OGT) and histone deacetylase-1 (HDAC1) play important roles in the pathogenesis of HCC. In the present study, we investigated the physical link between OGT and HDAC1. The O-GlcNAcylation of HDAC1 is overexpressed in HCC. We found that HDAC1 has two major sites of O-GlcNAcylation in its histone deacetylase domain. HDAC1 O-GlcNAcylation increases the activated phosphorylation of HDAC1, which enhances its enzyme activity. HDAC1 O-GlcNAc mutants promote the p21 transcription regulation through affecting the acetylation levels of histones from chromosome, and then influence the proliferation of HCC cells. We also found that mutants of O-GlcNAcylation site of HDAC1 affect invasion and migration of HepG2 cells. E-cadherin level is highly up-regulated in HDAC1 O-GlcNAc mutant-treated liver cancer cells, which inhibit the occurrence and development of HCC. Our findings suggest that OGT promotes the O-GlcNAc modification of HDAC1in the development of HCC. Therefore, inhibiting O-GlcNAcylation of HDAC1 may repress the progression of HCC.
Notes: 1460-2423
Zhu, Guizhou
Tao, Tao
Zhang, Dongmei
Liu, Xiaojuan
Qiu, Huiyuan
Han, LiJian
Xu, Zhiwei
Xiao, Ying
Cheng, Chun
Shen, Aiguo
Journal Article
England
Glycobiology. 2016 Aug;26(8):820-833. doi: 10.1093/glycob/cww025. Epub 2016 Apr 8.
Author Address: The Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 19 Qixiu Road, Nantong, 226001 Jiangsu, People's Republic of China.
Department of Co-innovation Center of Neuroregeneration, Nantong University, 19 Qixiu Road, Nantong, 226001 Jiangsu, People's Republic of China.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 2064
Author: Zhu, J., Lin, Y. H., Dingess, K. A., Mank, M., Stahl, B. and Heck, A. J. R.
Year: 2020
Title: Quantitative Longitudinal Inventory of the N-Glycoproteome of Human Milk from a Single Donor Reveals the Highly Variable Repertoire and Dynamic Site-Specific Changes
Journal: J Proteome Res
Volume: 19
Issue: 5
Pages: 1941-1952
Epub Date: 2020/03/04
Date: May 1
Short Title: Quantitative Longitudinal Inventory of the N-Glycoproteome of Human Milk from a Single Donor Reveals the Highly Variable Repertoire and Dynamic Site-Specific Changes
Alternate Journal: Journal of proteome research
ISSN: 1535-3893 (Print)
1535-3893
DOI: 10.1021/acs.jproteome.9b00753
PMCID: PMC7252941
Accession Number: 32125861
Keywords: Glycopeptides
Glycoproteins
Humans
*Milk, Human
*Proteome
Proteomics
*HILIC-based platform
*N-glycopeptide enrichment
*N-glycosylation
*glycopeptides
*glycoprotein
*human milk
*lactation
*mass spectrometry
*milk proteins
*targeted N-glycopeptide quantification
employees of Danone Nutricia Research. J.Z and K.A.D were enrolled as PhD students
at Utrecht University during this study and received partial financial support from
Danone Nutricia Research. None of the authors have further conflicts of interest
with regard to the content of this manuscript.
Abstract: Protein N-glycosylation on human milk proteins assists in protecting an infant's health and functions among others as competitive inhibitors of pathogen binding and immunomodulators. Due to the individual uniqueness of each mother's milk and the overall complexity and temporal changes of protein N-glycosylation, analysis of the human milk N-glycoproteome requires longitudinal personalized approaches, providing protein- and N-site-specific quantitative information. Here, we describe an automated platform using hydrophilic-interaction chromatography (HILIC)-based cartridges enabling the proteome-wide monitoring of intact N-glycopeptides using just a digest of 150 μg of breast milk protein. We were able to map around 1700 glycopeptides from 110 glycoproteins covering 191 glycosites, of which 43 sites have not been previously reported with experimental evidence. We next quantified 287 of these glycopeptides originating from 50 glycoproteins using a targeted proteomics approach. Although each glycoprotein, N-glycosylation site, and attached glycan revealed distinct dynamic changes, we did observe a few general trends. For instance, fucosylation, especially terminal fucosylation, increased across the lactation period. Building on the improved glycoproteomics approach outlined above, future studies are warranted to reveal the potential impact of the observed glycosylation microheterogeneity on the healthy development of infants.
Notes: 1535-3907
Zhu, Jing
Lin, Yu-Hsien
Dingess, Kelly A
Mank, Marko
Stahl, Bernd
Heck, Albert J R
Orcid: 0000-0002-2405-4404
Journal Article
Research Support, Non-U.S. Gov't
J Proteome Res. 2020 May 1;19(5):1941-1952. doi: 10.1021/acs.jproteome.9b00753. Epub 2020 Mar 27.
Author Address: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Netherlands Proteomics Center, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Beijing Institute of Nutritional Resources, 100069 Beijing, China.
Danone Nutricia Research, Uppsalalaan 12, 3584 CT Utrecht, The Netherlands.
Chemical Biology & Drug Discovery, Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, 3584 CG Utrecht, The Netherlands.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1484
Author: Zhu, J., Yan, J. and Thornhill, W. B.
Year: 2012
Title: N-glycosylation promotes the cell surface expression of Kv1.3 potassium channels
Journal: Febs j
Volume: 279
Issue: 15
Pages: 2632-44
Epub Date: 2012/05/23
Date: Aug
Short Title: N-glycosylation promotes the cell surface expression of Kv1.3 potassium channels
Alternate Journal: The FEBS journal
ISSN: 1742-464x
DOI: 10.1111/j.1742-4658.2012.08642.x
Accession Number: 22613618
Keywords: Acetylglucosamine/metabolism
Amino Acid Sequence
Amino Acid Substitution
Animals
Binding Sites
CHO Cells
Cell Membrane/metabolism
Cricetinae
Cricetulus
Glycosylation
Kv1.3 Potassium Channel/*chemistry/genetics/*metabolism
Molecular Sequence Data
Mutagenesis, Site-Directed
Polysaccharides/chemistry/metabolism
Rats
Recombinant Proteins/chemistry/genetics/metabolism
Abstract: The voltage-gated potassium channel Kv1.3 plays an essential role in modulating membrane excitability in many cell types. Kv1.3 is a heavily glycosylated membrane protein. Two successive N-glycosylation consensus sites, N228NS and N229ST, are present on the S1-S2 linker of rat Kv1.3. Our data suggest that Kv1.3 contains only one N-glycan and it is predominantly attached to N229 in the S1-S2 extracellular linker. Preventing N-glycosylation of Kv1.3 significantly decreased its surface protein level and surface conductance density level, which were ∼ 49% and ∼ 46% respectively of the level of wild type. Supplementation of N-acetylglucosamine (GlcNAc), l-fucose or N-acetylneuraminic acid to the culture medium promoted Kv1.3 surface protein expression, whereas supplementation of d-glucose, d-mannose or d-galactose did not. Among the three effective monosaccharides/derivatives, adding GlcNAc appeared to reduce sialic acid content and increase the degree of branching in the N-glycan of Kv1.3, suggesting that the N-glycan structure and composition had changed. Furthermore, the cell surface half-life of the Kv1.3 surface protein was increased upon GlcNAc supplementation, indicating that it had decreased internalization. The GlcNAc effect appears to apply mainly to membrane proteins containing complex type N-glycans. Thus, N-glycosylation promotes Kv1.3 cell surface expression; supplementation of GlcNAc increased Kv1.3 surface protein level and decreased its internalization, presumably by a combined effect of decreased branch size and increased branching of the N-glycan.
Notes: 1742-4658
Zhu, Jing
Yan, Jenny
Thornhill, William B
Journal Article
England
FEBS J. 2012 Aug;279(15):2632-44. doi: 10.1111/j.1742-4658.2012.08642.x. Epub 2012 Jun 15.
Author Address: Department of Biological Sciences and Center for Cancer, Genetic Diseases and Gene Regulation, Fordham University, Bronx, NY, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1717
Author: Zhu, L., Tao, T., Zhang, D., Liu, X., Ke, K. and Shen, A.
Year: 2015
Title: NOS1AP O-GlcNAc Modification Involved in Neuron Apoptosis Induced by Excitotoxicity
Journal: Int J Mol Sci
Volume: 16
Issue: 7
Pages: 16560-75
Epub Date: 2015/07/22
Date: Jul 21
Short Title: NOS1AP O-GlcNAc Modification Involved in Neuron Apoptosis Induced by Excitotoxicity
Alternate Journal: International journal of molecular sciences
ISSN: 1422-0067
DOI: 10.3390/ijms160716560
PMCID: PMC4519966
Accession Number: 26197318
Keywords: Acetylglucosamine/*metabolism
Adaptor Proteins, Signal Transducing/genetics/*metabolism
Animals
*Apoptosis
Glutamic Acid/toxicity
Glycosylation
HEK293 Cells
Humans
Male
Neurons/drug effects/*metabolism
Nitric Oxide Synthase Type I/metabolism
PC12 Cells
*Protein Processing, Post-Translational
Rats
Rats, Sprague-Dawley
Nos1ap
O-GlcNAc modification
excitotoxicity
Abstract: O-Linked N-acetylglucosamine, or O-GlcNAc, is a dynamic post-translational modification that cycles on and off serine and threonine residues of nucleocytoplasmic and mitochondrial proteins. In addition to cancer and inflammation diseases, O-GlcNAc modification appears to play a critical role during cell apoptosis and stress response, although the precise mechanisms are still not very clear. Here we found that nitric oxide synthase adaptor (NOS1AP), which plays an important part in glutamate-induced neuronal apoptosis, carries the modification of O-GlcNAc. Mass spectrometry analysis identified Ser47, Ser183, Ser204, Ser269, Ser271 as O-GlcNAc sites. Higher O-GlcNAc of NOS1AP was detected during glutamate-induced neuronal apoptosis. Furthermore, with O-GlcNAc sites of NOS1AP mutated, the interaction of NOS1AP and neuronal nitric oxide syntheses (nNOS) decreases. Finally, during glutamate-induced neuronal apoptosis, decreasing the O-GlcNAc modification of NOS1AP results in more severe neuronal apoptosis. All these results suggest that O-GlcNAc modification of NOS1AP exerts protective effects during glutamate-induced neuronal apoptosis.
Notes: 1422-0067
Zhu, Liang
Tao, Tao
Zhang, Dongmei
Liu, Xiaojuan
Ke, Kaifu
Shen, Aiguo
Journal Article
Research Support, Non-U.S. Gov't
Int J Mol Sci. 2015 Jul 21;16(7):16560-75. doi: 10.3390/ijms160716560.
Author Address: Department of Neurology, Affiliated Hospital of Nantong University, the Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. zhuliang_nt@163.com.
The Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. zhuliang_nt@163.com.
Department of Neurology, Affiliated Hospital of Nantong University, the Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. nttaotao@ntu.edu.cn.
The Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. nttaotao@ntu.edu.cn.
Department of Neurology, Affiliated Hospital of Nantong University, the Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. zdm@ntu.edu.cn.
The Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. zdm@ntu.edu.cn.
Department of Neurology, Affiliated Hospital of Nantong University, the Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. lxj@ntu.edu.cn.
The Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. lxj@ntu.edu.cn.
Department of Neurology, Affiliated Hospital of Nantong University, the Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. kekaifu_nt@126.com.
Department of Neurology, Affiliated Hospital of Nantong University, the Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. shag@ntu.edu.cn.
The Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, Nantong 226000, China. shag@ntu.edu.cn.
Department of Co-innovation Center of Neuroregeneration, Nantong University, Nantong 226000, China. shag@ntu.edu.cn.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1678
Author: Zhu, Q., Kao, L., Azimov, R., Abuladze, N., Newman, D. and Kurtz, I.
Year: 2015
Title: Interplay between disulfide bonding and N-glycosylation defines SLC4 Na+-coupled transporter extracellular topography
Journal: J Biol Chem
Volume: 290
Issue: 9
Pages: 5391-404
Epub Date: 2015/01/09
Date: Feb 27
Short Title: Interplay between disulfide bonding and N-glycosylation defines SLC4 Na+-coupled transporter extracellular topography
Alternate Journal: The Journal of biological chemistry
ISSN: 0021-9258 (Print)
0021-9258
DOI: 10.1074/jbc.M114.619320
PMCID: PMC4342456
Accession Number: 25568315
Keywords: Amino Acid Sequence
Binding Sites/genetics
Cysteine/*chemistry/genetics/metabolism
Disulfides/*chemistry/metabolism
Glycosylation
HEK293 Cells
Humans
Immunoblotting
Ion Transport/genetics
Microscopy, Fluorescence
Molecular Sequence Data
Mutation
*Protein Folding
Protein Multimerization
Protein Structure, Secondary
Sequence Homology, Amino Acid
Sodium-Bicarbonate Symporters/*chemistry/genetics/metabolism
Disulfide
Membrane Protein
Protein Folding
Transporter
Abstract: The extracellular loop 3 (EL-3) of SLC4 Na(+)-coupled transporters contains 4 highly conserved cysteines and multiple N-glycosylation consensus sites. In the electrogenic Na(+)-HCO3(-) cotransporter NBCe1-A, EL-3 is the largest extracellular loop and is predicted to consist of 82 amino acids. To determine the structural-functional importance of the conserved cysteines and the N-glycosylation sites in NBCe1-A EL-3, we analyzed the potential interplay between EL-3 disulfide bonding and N-glycosylation and their roles in EL-3 topological folding. Our results demonstrate that the 4 highly conserved cysteines form two intramolecular disulfide bonds, Cys(583)-Cys(585) and Cys(617)-Cys(642), respectively, that constrain EL-3 in a folded conformation. The formation of the second disulfide bond is spontaneous and unaffected by the N-glycosylation state of EL-3 or the first disulfide bond, whereas formation of the first disulfide bond relies on the presence of the second disulfide bond and is affected by N-glycosylation. Importantly, EL-3 from each monomer is adjacently located at the NBCe1-A dimeric interface. When the two disulfide bonds are missing, EL-3 adopts an extended conformation highly accessible to protease digestion. This unique adjacent parallel location of two symmetrically folded EL-3 loops from each monomer resembles a domain-like structure that is potentially important for NBCe1-A function in vivo. Moreover, the formation of this unique structure is critically dependent on the finely tuned interplay between disulfide bonding and N-glycosylation in the membrane processed NBCe1-A dimer.
Notes: 1083-351x
Zhu, Quansheng
Kao, Liyo
Azimov, Rustam
Abuladze, Natalia
Newman, Debra
Kurtz, Ira
R01 DK077162/DK/NIDDK NIH HHS/United States
DK077162/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
J Biol Chem. 2015 Feb 27;290(9):5391-404. doi: 10.1074/jbc.M114.619320. Epub 2015 Jan 7.
Author Address: From the Department of Medicine and quzhu@mednet.ucla.edu.
From the Department of Medicine and.
From the Department of Medicine and Brain Research Institute, David Geffen School of Medicine, UCLA, Los Angeles, California 90095-1689.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 866
Author: Zhu, Y., Rudensky, A. Y., Corper, A. L., Teyton, L. and Wilson, I. A.
Year: 2003
Title: Crystal structure of MHC class II I-Ab in complex with a human CLIP peptide: prediction of an I-Ab peptide-binding motif
Journal: J Mol Biol
Volume: 326
Issue: 4
Pages: 1157-74
Epub Date: 2003/02/19
Date: Feb 28
Short Title: Crystal structure of MHC class II I-Ab in complex with a human CLIP peptide: prediction of an I-Ab peptide-binding motif
Alternate Journal: Journal of molecular biology
ISSN: 0022-2836 (Print)
0022-2836
DOI: 10.1016/s0022-2836(02)01437-7
Accession Number: 12589760
Keywords: Amino Acid Sequence
Animals
Antigens, Differentiation, B-Lymphocyte/chemistry/*metabolism
Binding Sites
Crystallography, X-Ray
*Genes, MHC Class II
Histocompatibility Antigens Class II/*chemistry/*metabolism
Humans
Hydrogen Bonding
Macromolecular Substances
Mice
Models, Molecular
Molecular Sequence Data
Protein Binding
*Protein Structure, Tertiary
Recombinant Fusion Proteins/genetics/metabolism
Sequence Alignment
Abstract: Association between the class II major histocompatibility complex (MHC) and the class II invariant chain-associated peptide (CLIP) occurs naturally as an intermediate step in the MHC class II processing pathway. Here, we report the crystal structure of the murine class II MHC molecule I-A(b) in complex with human CLIP at 2.15A resolution. The structure of I-A(b) accounts, via the peptide-binding groove's unique physicochemistry, for the distinct peptide repertoire bound by this allele. CLIP adopts a similar conformation to peptides bound by other I-A alleles, reinforcing the notion that CLIP is presented as a conventional peptide antigen. When compared to the related HLA-DR3/CLIP complex structure, the CLIP peptide displays a slightly different conformation and distinct interaction pattern with residues in I-A(b). In addition, after examining the published sequences of peptides presented by I-A(b), we discuss the possibility of predicting peptide alignment in the I-A(b) binding groove using a simple scoring matrix.
Notes: Zhu, Yuerong
Rudensky, Alexander Y
Corper, Adam L
Teyton, Luc
Wilson, Ian A
CA-58896/CA/NCI NIH HHS/United States
DK55037/DK/NIDDK NIH HHS/United States
Journal Article
Research Support, U.S. Gov't, P.H.S.
England
J Mol Biol. 2003 Feb 28;326(4):1157-74. doi: 10.1016/s0022-2836(02)01437-7.
Author Address: Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Database Provider: NLM
Language: eng

Reference Type:  Journal Article
Record Number: 1250
Author: Zong, X., Schieder, M., Cuny, H., Fenske, S., Gruner, C., Rötzer, K., Griesbeck, O., Harz, H., Biel, M. and Wahl-Schott, C.
Year: 2009
Title: The two-pore channel TPCN2 mediates NAADP-dependent Ca(2+)-release from lysosomal stores
Journal: Pflugers Arch
Volume: 458
Issue: 5
Pages: 891-9
Epub Date: 2009/06/27
Date: Sep
Short Title: The two-pore channel TPCN2 mediates NAADP-dependent Ca(2+)-release from lysosomal stores
Alternate Journal: Pflugers Archiv : European journal of physiology
ISSN: 0031-6768 (Print)
0031-6768
DOI: 10.1007/s00424-009-0690-y
PMCID: PMC2719734
Accession Number: 19557428
Keywords: Amino Acid Sequence
Animal Structures/metabolism
Animals
COS Cells
Calcium/*metabolism
Calcium Channels/*physiology
Calcium Signaling/drug effects/*physiology
Cell Line
Chlorocebus aethiops
Cloning, Molecular
Endoplasmic Reticulum/metabolism
Enzyme Inhibitors/pharmacology
Glycosylation
Humans
Ion Channel Gating/physiology
Lysosomal-Associated Membrane Protein 1/metabolism
Lysosomes/*metabolism
Mice
Mice, Inbred Strains
Molecular Sequence Data
NADP/*analogs & derivatives/metabolism/pharmacology
Sequence Homology, Amino Acid
Thapsigargin/pharmacology
Transfection
Vacuolar Proton-Translocating ATPases/antagonists & inhibitors
Abstract: Second messenger-induced Ca(2+)-release from intracellular stores plays a key role in a multitude of physiological processes. In addition to 1,4,5-inositol trisphosphate (IP(3)), Ca(2+), and cyclic ADP ribose (cADPR) that trigger Ca(2+)-release from the endoplasmatic reticulum (ER), nicotinic acid adenine dinucleotide phosphate (NAADP) has been identified as a cellular metabolite that mediates Ca(2+)-release from lysosomal stores. While NAADP-induced Ca(2+)-release has been found in many tissues and cell types, the molecular identity of the channel(s) conferring this release remained elusive so far. Here, we show that TPCN2, a novel member of the two-pore cation channel family, displays the basic properties of native NAADP-dependent Ca(2+)-release channels. TPCN2 transcripts are widely expressed in the body and encode a lysosomal protein forming homomers. TPCN2 mediates intracellular Ca(2+)-release after activation with low-nanomolar concentrations of NAADP while it is desensitized by micromolar concentrations of this second messenger and is insensitive to the NAADP analog nicotinamide adenine dinucleotide phosphate (NADP). Furthermore, TPCN2-mediated Ca(2+)-release is almost completely abolished when the capacity of lysosomes for storing Ca(2+) is pharmacologically blocked. By contrast, TPCN2-specific Ca(2+)-release is unaffected by emptying ER-based Ca(2+) stores. In conclusion, these findings indicate that TPCN2 is a major component of the long-sought lysosomal NAADP-dependent Ca(2+)-release channel.
Notes: 1432-2013
Zong, Xiangang
Schieder, Michael
Cuny, Hartmut
Fenske, Stefanie
Gruner, Christian
Rötzer, Katrin
Griesbeck, Oliver
Harz, Hartmann
Biel, Martin
Wahl-Schott, Christian
Journal Article
Research Support, Non-U.S. Gov't
Pflugers Arch. 2009 Sep;458(5):891-9. doi: 10.1007/s00424-009-0690-y. Epub 2009 Jun 26.
Author Address: Center for Integrated Protein Science CIPS-M and Zentrum für Pharmaforschung, Department Pharmazie, Ludwig-Maximilians-Universität München, Butenandtstr. 5-13, 81377, Munich, Germany.
Database Provider: NLM
Language: eng