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Isolation and Characterization of a Novel Phospholipase A1 Secreted from Human Platelets
Author(s) -
Bolen Alyssa Lynn,
Niessen Sherry,
Cravatt Benjamin,
BeranovaGiorgianni Sarka,
Tigyi Gabor
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.791.13
Subject(s) - lysophosphatidic acid , chemistry , biochemistry , phospholipase a2 , arachidonic acid , platelet , enzyme , thrombin , biology , receptor , immunology
Lysophosphatidic acid (LPA) is a phospholipid mediator that plays a role in human diseases including thrombosis, arteriosclerosis, angiogenesis, cancer metastasis, and inflammation. LPA is present in blood at low nanomolar concentrations, whereas in serum it reaches several micromolar in concentration. The mechanisms responsible for LPA production during blood coagulation remain unknown. Because in mammalian phospholipids linoleoleic acid and arachidonic acid are esterified to the sn‐2 position of the glycerol backbone and 83% of LPA in serum carries these fatty acids, we hypothesized that a phospholipase A 1 (PLA 1 ) is involved in its production. We have purified a novel PLA 1 from the microparticle‐free supernatant of thrombin‐activated human platelets that is distinct from known PLA 1 s and PLA 2 s. We have performed sequential chromatographic steps using Q‐Sepharose, Sephadex PD‐10, Butyl‐Sepharose, and MonoS columns and obtained a 150‐fold purified enzyme. Phospholipases are typically serine hydrolases and rhodamine‐ or biotin‐labeled flurorophosphonate probes abolished the PLA 1 activity. These affinity labels will be used for mass spectrometric identification of the labeled proteins. Because PLA 1 plays an important role in LPA production, further investigation of this enzyme may prove it to be an excellent therapeutic target for the inhibition of LPA production in disease.