Human myocardial cytosolic Ca(2+)-independent phospholipase A2 is modulated by ATP. Concordant ATP-induced alterations in enzyme kinetics and mechanism-based inhibition.
Although phospholipases A2 (PLsA2) have been implicated as enzymic mediators of electrophysiological dysfunction during myocardial ischaemia and reperfusion in man, no information on the regulation of human myocardial PLsA2 is available. We now report that human myocardial cytosolic Ca(2+)-independent PLA2 is modulated by ATP through mechanisms which are independent of protein phosphorylation, since: (1) ATP and its non-hydrolysable analogues reversibly augment the initial rate of crude (but not purified) human myocardial cytosolic PLA2 activity 3-4 fold; (2) ATP and its non-hydrolysable analogues dramatically attenuate the rate of thermal denaturation of human myocardial cytosolic PLA2 activity; (3) ATP and its non-hydrolysable analogues alter the sensitivity of human myocardial cytosolic PLA2 to mechanism-based inhibition by (E)-6-(bromomethylene)tetrahydro-3-(1-naphthalenyl)-2H-pyran-2-one; (4) adenine nucleotide triphosphates are more potent than guanine nucleotide triphosphates in activating and stabilizing human myocardial cytosolic PLA2; (5) adenosine 5'-[beta gamma-methylene]triphosphate is more potent than its guanosine counterpart in protecting an essential thiol residue(s) in human myocardial cytosolic PLA2 from covalent modification by 5,5'-dithiobis-(2-nitrobenzoic acid); and (6) ATP-dependent activation and stabilization of human myocardial cytosolic PLA2 are mediated by a cytosolic protein(s) which can be functionally reconstituted with purified human myocardial cytosolic PLA2 catalytic polypeptide. Collectively these results demonstrate that multiple physical and kinetic properties of human myocardial cytosolic Ca(2+)-independent PLA2 are dramatically influenced by dynamic interactions with ATP.
Hazen
S L
SL
Department of Internal Medicine, Washington University School of Medicine, St. Louis, MO 63110.
Gross
R W
RW
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004795:Enzyme Stability; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives; D006801:Humans; D007700:Kinetics; D009206:Myocardium / Q000201:enzymology*; D010741:Phospholipases A / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D054467:Phospholipases A2; D011489:Protein Denaturation; D013438:Sulfhydryl Compounds / Q000378:metabolism
D013438:Sulfhydryl Compounds; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D010741:Phospholipases A; D054467:Phospholipases A2