Biphasic Ca2+ response of adenylate cyclase. The role of calmodulin in its activation by Ca2+ ions.
The Ca2+-dependent regulation of human platelet membrane adenylate cyclase has been studied. This enzyme exhibited a biphasic response to Ca2+ within a narrow range of Ca2+ concentrations (0.1-1.0 microM). At low Ca2+ (0.08-0.3 microM) adenylate cyclase was stimulated (Ka = 0.10 microM), whereas at higher Ca2+ (greater than 0.3 microM) the enzyme was inhibited to 70-80% control (Ki = 0.8 microM). Membrane fractions, prepared by washing in the presence of LaCl3 to remove endogenous calmodulin (approximately equal to 70-80% depletion), exhibited no stimulation of adenylate cyclase by Ca2+ but did show the inhibitory phase (Ki = 0.4 microM). The activation phase could be restored to La3+-washed membranes by addition of calmodulin (Ka = 3.0 nM). Under these conditions it was apparent that calmodulin reduced the sensitivity of adenylate cyclase to Ca2+ (Ki = 0.8 microM). Prostaglandin E1 (PGE1) did not alter Ki or Ka values for Ca2+. Calmodulin did not alter the EC50 for PGE1 stimulation of adenylate cyclase but increased the Vmax (1.5-fold). The calmodulin antagonist trifluoperazine potently inhibited adenylate cyclase in native membranes (80%) and to a much lesser extent in La3+-washed membranes (15%). This inhibition was due to interaction of trifluoperazine with endogenous calmodulin since trifluoperazine competitively antagonized the stimulatory effect of calmodulin on adenylate cyclase in La3+-washed membranes. We propose that biphasic Ca2+ regulation of platelet adenylate cyclase functions to both dampen (low Ca2+) and facilitate (high Ca2+) the haemostatic function of platelets.
D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000097:blood*; D001792:Blood Platelets / Q000201:enzymology; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000502:physiology*; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007811:Lanthanum; D014268:Trifluoperazine / Q000494:pharmacology
D000067956:Adenylyl Cyclase Inhibitors; D002147:Calmodulin; D014268:Trifluoperazine; D007811:Lanthanum; D000262:Adenylyl Cyclases; D002118:Calcium