title: Purification and characterization of a phospholipase A2 from human osteoarthritic synovial fluid.
abstract: Phospholipase A2 (PLA2) from human osteoarthritic synovial fluid was purified to homogeneity in three steps. The NH2-terminal amino acid sequence and biochemical characteristics of the enzyme were identical to the Peak A PLA2 activity of rheumatoid synovial fluid (1). The enzyme exhibited an apparent mass of 14,000, an absolute Ca+(+)-dependence, an alkaline pH optimum, and was inhibited by sodium deoxycholate (DOC), NaCl and 0.5 M Tris-HCl. The enzyme strongly preferred phosphatidylethanolamine (PE) as substrate over phosphatidylcholine (PC) or phosphatidylinositol (PI), and hydrolyzed PE containing arachidonic acid or linoleic acid in the sn-2 position at similar rates. Heparin bound to the enzyme but did not inhibit catalytic activity. In addition, the human enzyme was not inhibited by the acidic 'chaperone' subunit of crotoxin despite considerable sequence similarity with the basic PLA2 subunit of the neurotoxin. The enzyme was capable of hydrolyzing E. coli membrane phospholipids in the presence of the neutrophil bactericidal/permeability increasing protein (BPI). This finding, coupled to the reported pro-inflammatory activity and presence of the enzyme in inflammatory cells, supports the hypothesis that it may be a component of the host defense mechanism which can, under certain conditions, contribute to the pathogenesis of inflammatory disease.
authors: Department of Biochemistry, Boehringer Ingelheim Pharmaceuticals, Inc., Ridgefield, CT 06877.
authors: T P
authors: TP
authors: Parks
authors: S
authors: S
authors: Lukas
authors: A F
authors: AF
authors: Hoffman
mesh_terms: D000595:Amino Acid Sequence; D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D002414:Cations, Monovalent; D003439:Crotoxin / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism; D006025:Glycosaminoglycans / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008565:Membrane Proteins; D008823:Micelles; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000378:metabolism; D010003:Osteoarthritis / Q000201:enzymology*; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010741:Phospholipases A / Q000302:isolation & purification* / Q000378:metabolism; D054467:Phospholipases A2; D010743:Phospholipids / Q000378:metabolism; D013379:Substrate Specificity; D013501:Surface-Active Agents / Q000494:pharmacology; D013582:Synovial Fluid / Q000201:enzymology*
chemical_list: D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins; D002414:Cations, Monovalent; D006025:Glycosaminoglycans; D008565:Membrane Proteins; D008823:Micelles; D010714:Phosphatidylethanolamines; D010743:Phospholipids; D013501:Surface-Active Agents; C021623:bactericidal permeability increasing protein; D003439:Crotoxin; D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium