Purification and characterization of platelet-activating factor acetylhydrolase from peritoneal fluid obtained from guinea pigs after endotoxin shock.
Platelet-activating factor (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine; PAF) acetyl-hydrolase is an enzyme that hydrolyzes the acetyl ester of PAF. We purified this enzyme, which accumulated in the peritoneal cavity during endotoxin shock, by ammonium sulfate precipitation, and sequential use of butyl-Toyopearl, heparin-Sepharose, Con A-Sepharose, chelating-Sepharose, and MonoQ column chromatographies. We identified a monomeric polypeptide with a molecular weight of approximately 63 kDa on SDS-PAGE. This molecular weight differs from those of previously described acetylhydrolases. The purified enzyme did not degrade phospholipids with a long chain fatty acyl group at the sn-2 position. In addition, the enzyme activity was not inhibited by either pBPB or quinacrine. Accordingly, this enzyme is distinct from phospholipase A2. In addition, this enzyme also hydrolyzed some oxidatively fragmented phospholipids with PAF-like biological activities such as 1-O-hexadecyl-2-glutaroyl-sn-glycero-3-phosphocholine and 1-O-hexadecyl-2-succinoyl-sn-glycero-3-phosphocholine.
Karasawa
K
K
Faculty of Pharmaceutical Sciences, Teikyo University, Kanagawa.
Yato
M
M
Setaka
M
M
Nojima
S
S
D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase; D000818:Animals; D001202:Ascitic Fluid / Q000201:enzymology*; D002845:Chromatography / Q000379:methods; D006168:Guinea Pigs; D006867:Hydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism; D006868:Hydrolysis; D008297:Male; D010755:Organophosphates / Q000494:pharmacology; D010084:Oxidation-Reduction; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism*; D054467:Phospholipases A2; D010743:Phospholipids / Q000378:metabolism; D012772:Shock, Septic / Q000201:enzymology*; D013379:Substrate Specificity
D010755:Organophosphates; D010743:Phospholipids; C091491:diisopropylphenylphosphate; D010664:Phenylmethylsulfonyl Fluoride; D006867:Hydrolases; D010741:Phospholipases A; D054467:Phospholipases A2; D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase