PMID- 10333503
OWN - NLM
STAT- MEDLINE
DCOM- 19990729
LR  - 20191210
IS  - 0264-6021 (Print)
IS  - 0264-6021 (Linking)
VI  - 340 ( Pt 2)
DP  - 1999 Jun 1
TI  - Identification of peroxisomal proteins by using M13 phage protein VI phage
      display: molecular evidence that mammalian peroxisomes contain a 2,4-dienoyl-CoA 
      reductase.
PG  - 561-8
AB  - To elucidate unknown mammalian peroxisomal enzymes and functions, we subjected
      M13 phage expressing fusions between the gene encoding protein VI and a rat liver
      cDNA library to an immunoaffinity selection process in vitro (biopanning) with
      the use of antibodies raised against peroxisomal subfractions. In an initial
      series of biopanning experiments, four different cDNA clones were obtained. These
      cDNA species encoded two previously identified peroxisomal enzymes, catalase and 
      urate oxidase, and two novel proteins that contained a C-terminal peroxisomal
      targeting signal (PTS1). A primary structure analysis of these novel proteins
      revealed that one, ending in the tripeptide AKL, is homologous to the yeast
      peroxisomal 2,4-dienoyl-CoA reductase (EC 1.3.1.34; DCR), an enzyme required for 
      the degradation of unsaturated fatty acids, and that the other, ending in the
      tripeptide SRL, is a putative member of the short-chain dehydrogenase/reductase
      (SDR) family, with three isoforms. Green fluorescent protein (GFP) fusions
      encoding GFP-DCR-AKL, GFP-DCR, GFP-SDR-SRL and GFP-SDR were expressed in
      mammalian cells. The analysis of the subcellular location of the recombinant
      fusion proteins confirmed the peroxisomal localization of GFP-DCR-AKL and
      GFP-SDR-SRL, as well as the functionality of the PTS1. That the AKL protein is
      indeed an NADPH-dependent DCR was demonstrated by showing DCR activity of the
      bacterially expressed protein. These results demonstrate at the molecular level
      that mammalian peroxisomes do indeed contain a DCR. In addition, the results
      presented here indicate that the protein VI display system is suitable for the
      isolation of rare cDNA clones from cDNA libraries and that this technology
      facilitates the identification of novel peroxisomal proteins.
FAU - Fransen, M
AU  - Fransen M
AD  - Department of Biology, University of California at San Diego, 9500 Gilman Drive, 
      La Jolla, CA 92093-0322, USA.
FAU - Van Veldhoven, P P
AU  - Van Veldhoven PP
FAU - Subramani, S
AU  - Subramani S
LA  - eng
SI  - GENBANK/AF044574
GR  - DK41737/DK/NIDDK NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - England
TA  - Biochem J
JT  - The Biochemical journal
JID - 2984726R
RN  - 0 (DNA Primers)
RN  - 0 (DNA, Complementary)
RN  - 0 (Luminescent Proteins)
RN  - 0 (Nuclear Proteins)
RN  - 0 (Recombinant Fusion Proteins)
RN  - 0 (Viral Proteins)
RN  - 147336-22-9 (Green Fluorescent Proteins)
RN  - EC 1.1.- (Alcohol Oxidoreductases)
RN  - EC 1.1.1.1 (Alcohol Dehydrogenase)
RN  - EC 1.1.1.184 (Carbonyl Reductase (NADPH))
RN  - EC 1.1.1.184 (DHRS2 protein, human)
RN  - EC 1.14.19.- (Fatty Acid Desaturases)
RN  - EC 1.3.- (Oxidoreductases Acting on CH-CH Group Donors)
RN  - EC 1.3.1.34 (2,4-dienoyl-CoA reductase)
SB  - IM
MH  - Alcohol Dehydrogenase/chemistry/genetics/metabolism
MH  - Alcohol Oxidoreductases
MH  - Amino Acid Sequence
MH  - Animals
MH  - Bacteriophage M13/*genetics
MH  - Base Sequence
MH  - Carbonyl Reductase (NADPH)
MH  - Cloning, Molecular
MH  - DNA Primers
MH  - DNA, Complementary
MH  - Fatty Acid Desaturases/chemistry/*genetics/metabolism
MH  - Green Fluorescent Proteins
MH  - Humans
MH  - Luminescent Proteins/genetics
MH  - Microbodies/enzymology/*metabolism
MH  - Molecular Sequence Data
MH  - Nuclear Proteins/chemistry/genetics/metabolism
MH  - *Oxidoreductases Acting on CH-CH Group Donors
MH  - Rats
MH  - Rats, Wistar
MH  - Recombinant Fusion Proteins/genetics
MH  - Sequence Homology, Amino Acid
MH  - Viral Proteins/*genetics
PMC - PMC1220285
EDAT- 1999/05/20 00:00
MHDA- 1999/05/20 00:01
CRDT- 1999/05/20 00:00
PHST- 1999/05/20 00:00 [pubmed]
PHST- 1999/05/20 00:01 [medline]
PHST- 1999/05/20 00:00 [entrez]
PST - ppublish
SO  - Biochem J. 1999 Jun 1;340 ( Pt 2):561-8.