PMID- 10464296
OWN - NLM
STAT- MEDLINE
DCOM- 19991007
LR  - 20190508
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 274
IP  - 36
DP  - 1999 Sep 3
TI  - Structure and function of HNK-1 sulfotransferase. Identification of donor and
      acceptor binding sites by site-directed mutagenesis.
PG  - 25608-12
AB  - HNK-1 glycan, sulfo-->3GlcAbeta1-->3Galbeta1-->4GlcNAc-->R, is uniquely enriched 
      in neural cells and natural killer cells and is thought to play important roles
      in cell-cell interaction. HNK-1 glycan synthesis is dependent on HNK-1
      sulfotransferase (HNK-1ST), and cDNAs encoding human and rat HNK-1ST have been
      recently cloned. HNK-1ST belongs to the sulfotransferase gene family, which
      shares two homologous sequences in their catalytic domains. In the present study,
      we have individually mutated amino acid residues in these conserved sequences and
      determined how such mutations affect the binding to the donor substrate,
      adenosine 3'-phosphate 5'-phosphosulfate, and an acceptor. Mutations of Lys(128),
      Arg(189), Asp(190), Pro(191), and Ser(197) to Ala all abolished the enzymatic
      activity. When Lys(128) and Asp(190) were conservatively mutated to Arg and Glu, 
      respectively, however, the mutated enzymes still maintained residual activity,
      and both mutant enzymes still bound to adenosine 3',5'-diphosphate-agarose. K128R
      and D190E mutant enzymes, on the other hand, exhibited reduced affinity to the
      acceptor as demonstrated by kinetic studies. These findings, together with those 
      on the crystal structure of estrogen sulfotransferase and heparan sulfate
      N-deacetylase/sulfotransferase, suggest that Lys(128) may be close to the
      3-hydroxyl group of beta-glucuronic acid in a HNK-1 acceptor. In contrast, the
      effect by mutation at Asp(190) may be due to conformational change because this
      amino acid and Pro(191) reside in a transition of the secondary structure of the 
      enzyme. These results indicate that conserved amino acid residues in HNK-1ST play
      roles in maintaining a functional conformation and are directly involved in
      binding to donor and acceptor substrates.
FAU - Ong, E
AU  - Ong E
AD  - Glycobiology Program, Cancer Research Center, The Burnham Institute, La Jolla,
      California 92037, USA.
FAU - Yeh, J C
AU  - Yeh JC
FAU - Ding, Y
AU  - Ding Y
FAU - Hindsgaul, O
AU  - Hindsgaul O
FAU - Pedersen, L C
AU  - Pedersen LC
FAU - Negishi, M
AU  - Negishi M
FAU - Fukuda, M
AU  - Fukuda M
LA  - eng
GR  - P01 CA71932/CA/NCI NIH HHS/United States
GR  - R01 CA33895/CA/NCI NIH HHS/United States
PT  - Journal Article
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - EC 2.8.2.- (CHST10 protein, human)
RN  - EC 2.8.2.- (Chst10 protein, rat)
RN  - EC 2.8.2.- (HNK-1 sulfotransferase)
RN  - EC 2.8.2.- (Sulfotransferases)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Binding Sites/genetics
MH  - Enzyme Activation/genetics
MH  - Humans
MH  - Molecular Sequence Data
MH  - Mutagenesis, Site-Directed
MH  - Rats
MH  - Sequence Alignment
MH  - Structure-Activity Relationship
MH  - Substrate Specificity
MH  - Sulfotransferases/*chemistry/genetics/*metabolism
EDAT- 1999/08/28 00:00
MHDA- 1999/08/28 00:01
CRDT- 1999/08/28 00:00
PHST- 1999/08/28 00:00 [pubmed]
PHST- 1999/08/28 00:01 [medline]
PHST- 1999/08/28 00:00 [entrez]
AID - 10.1074/jbc.274.36.25608 [doi]
PST - ppublish
SO  - J Biol Chem. 1999 Sep 3;274(36):25608-12. doi: 10.1074/jbc.274.36.25608.