Purification and characterization of a calcium-independent acidic phospholipase A2 from rat lung.
Several phospholipase A2 (PLA2) activities have been identified in rat lung homogenate and shown to be important in metabolism of lung phospholipids. One PLA2 activity is Ca(2+)-independent, active in vitro at pH 4, and inhibited by a substrate analogue, 1-hexadecyl-3-trifluoroethylglycero-sn-2-phosphomethanol (MJ33). Purification of this rat lung PLA2 by approx. 550-fold was carried out by sequential column chromatographies using DE-52, Sephacryl-100, heparin-Sepharose, and phenyl-Sepharose columns. The purified activity had an acidic pH optimum, was Ca(2+)-independent, was inhibited by MJ33 in a dose-dependent manner (50% inhibition at 3 mol%), was unaffected by treatment with p-bromophenacyl bromide or mercaptoethanol, and had a unique N-terminal amino acid sequence. The apparent molecular mass was 15 kDa on gel electrophoresis and activity was recovered in part by renaturation of protein from the gel. The properties of this enzyme suggest a new class of PLA2.
Wang
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R
Institute for Environmental Medicine, University of Pennsylvania Medical Center, Philadelphia 19104-6068.
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Jain
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D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D005994:Glycerophosphates / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008168:Lung / Q000201:enzymology*; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D054467:Phospholipases A2; D051381:Rats; D017207:Rats, Sprague-Dawley
D005994:Glycerophosphates; C078587:1-hexadecyl-3-trifluoroethylglycero-sn-2-phosphomethanol; D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium