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PMID:18614531

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Citation

Duncan, RE, Sarkadi-Nagy, E, Jaworski, K, Ahmadian, M and Sul, HS (2008) Identification and functional characterization of adipose-specific phospholipase A2 (AdPLA). J. Biol. Chem. 283:25428-36

Abstract

Phospholipases A(2) (PLA(2)s) catalyze hydrolysis of fatty acids from the sn-2 position of phospholipids. Here we report the identification and characterization of a membrane-associated intracellular calcium-dependent, adipose-specific PLA(2) that we named AdPLA (adipose-specific phospholipase A(2)). We found that AdPLA was highly expressed specifically in white adipose tissue and was induced during preadipocyte differentiation into adipocytes. Clearance of AdPLA by immunoprecipitation significantly decreased PLA activity in white adipose tissue lysates but had no effect on liver lysates, where expression was hardly detectable. In characterizing AdPLA, we employed radiochemical assays with TLC analysis of the enzyme activity of lysates from COS-7 cells overexpressing AdPLA. For kinetic studies, we produced purified recombinant AdPLA for use in a lipoxidase-coupled spectrophotometric assay. AdPLA generated free fatty acid and lysophospholipid from phosphatidylcholine with a preference for hydrolysis at the sn-2 position. Although we found low but detectable lysophospholipase activity, AdPLA showed no significant activity against a variety of other lipid substrates. Calcium was found to activate AdPLA but was not essential for activity. Studies with known phospholipase inhibitors, including bromoenolactone, methyl arachidonyl fluorophosphate, AACOCF(3), 7,7-dimethyl-5,8-eicosadienoic acid, and thioetheramide, supported that AdPLA is a phospholipase. Mutational studies showed that His-23 and Cys-113 are critical for activity of AdPLA and suggested that AdPLA is likely a His/Cys PLA(2). Overall, although AdPLA is similar to other histidine phospholipases in pH and calcium dependence, AdPLA showed different characteristics in many regards, including predicted catalytic mechanism. AdPLA may therefore represent the first member of a new group of PLA(2)s, group XVI.

Links

PubMed PMC2533091 Online version:10.1074/jbc.M804146200

Keywords

3T3 Cells; Adipocytes/metabolism; Adipose Tissue/enzymology; Amides/pharmacology; Animals; Arachidonic Acids/pharmacology; COS Cells; Cercopithecus aethiops; Fatty Acids, Unsaturated/pharmacology; Lysophospholipids/chemistry; Mice; Models, Biological; Phosphatidylcholines/chemistry; Phospholipases A2/chemistry; Phospholipases A2/metabolism; Sulfides/pharmacology

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

MOUSE:PLAT3

located_in

GO:0048471: perinuclear region of cytoplasm

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

MOUSE:PLAT3

located_in

GO:0005783: endoplasmic reticulum

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

MOUSE:PLAT3

involved_in

GO:0008654: phospholipid biosynthetic process

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete

MOUSE:PLAT3

NOT|enables

GO:0016746: acyltransferase activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

MOUSE:PLAT3

enables

GO:0004623: phospholipase A2 activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete


See also

References

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