Tricyclohexyltin hydroxide effects on cationic and substrate activation kinetics of beta-adrenergic-stimulated cardiac Ca2+-ATPase.
Previous studies from this laboratory have indicated that tricyclohexyltin hydroxide (Plictran) is a potent inhibitor of both basal- and isoproterenol-stimulated cardiac sarcoplasmic reticulum (SR) Ca2+-ATPase, with an estimated IC-50 of 2.5 X 10(-8) M. The present studies were initiated to evaluate the mechanism of inhibition of Ca2+-ATPase by Plictran. Data on substrate and cationic activation kinetics of Ca2+-ATPase indicated alteration of Vmax and Km by Plictran (1 and 5 X 10(-8) M), suggesting a mixed type of inhibition. The beta-adrenergic agonist isoproterenol increased Vmax of both ATP- and Ca2+-dependent enzyme activities. However, the Km of enzyme was decreased only for Ca2+. Plictran inhibited isoproterenol-stimulated Ca2+-ATPase activity by altering both Vmax and Km of ATP as well as Ca2+-dependent enzyme activities, suggesting that after binding to a single independent site, Plictran inhibits enzyme catalysis by decreasing the affinity of enzyme for ATP as well as for Ca2+. Preincubation of enzyme with 15 microM cAMP or the addition of 2mM ATP to the reaction mixture resulted in slight activation of Plictran-inhibited enzyme. Pretreatment of SR with 5 X 10(-7) M propranolol and 5 X 10(-8) M Plictran resulted in inhibition of basal activity in addition to the loss of stimulated activity. Preincubation of heart SR preparation with 5 X 10(-5) M coenzyme A in combination with 5 X 10(-8) M Plictran partly restored the beta-adrenergic stimulation. These results suggest that some critical sites common to both basal- and beta-adrenergic-stimulated Ca2+-ATPase are sensitive to binding by Plictran, and the resultant conformational change may lead to inhibition of beta-adrenergic stimulation.
Sahib
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Department of Neurology, University of Mississippi Medical Center, Jackson 39216.
Desaiah
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D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000032:analysis / Q000037:antagonists & inhibitors*; D000242:Cyclic AMP / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D007545:Isoproterenol / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D009206:Myocardium / Q000201:enzymology*; D010710:Phosphates / Q000032:analysis; D011433:Propranolol / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D013379:Substrate Specificity; D014220:Trialkyltin Compounds / Q000494:pharmacology*
D010710:Phosphates; D014220:Trialkyltin Compounds; D000255:Adenosine Triphosphate; C001765:cyhexatin; D011433:Propranolol; D000242:Cyclic AMP; D000252:Calcium-Transporting ATPases; D007545:Isoproterenol; D002118:Calcium