Activation of 3',5'-cyclic adenosine monophosphate phosphodiesterase by calcium ion and a protein activator.
3',5'-CAMP phosphodiesterase was partially purified from bovine cerebral cortex. A heat-stable activating factor was separated from the enzyme by chromatography on DEAE-cellulose. The enzyme in crude ammonium sulfate fractions was stimulated by 5 mM CaCl2. This stimulation was reversed by the calcium chelator EGTA. The main phosphodiesterase peak obtained by DEAE-cellulose chromatography was not stimulated by Ca2+. Upon addition of column effluent containing a heat stable factor, Ca2+ activation was restored. Protein activator was inactive when endogenous contaminating Ca2+ was complexed with EGTA. It was concluded that activation of phosphodiesterase requires the presence of both activator and Ca1+. From an analysis of activation of cGMP hydrolysis a kinetic model for the interaction of Ca2+ and protein activator with the phosphodiesterase was developed. Heterotropic cooperativity between the binding of Ca2+ and protein activator to the phosphodiesterase was observed, i.e., Ca1+ decreased the apparent dissociation constant for protein activator and protein activator decreased the apparent dissociation constant for Ca2+.
D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002417:Cattle; D002540:Cerebral Cortex / Q000201:enzymology*; D002614:Chelating Agents / Q000494:pharmacology; D002848:Chromatography, DEAE-Cellulose; D000242:Cyclic AMP; D006152:Cyclic GMP; D004789:Enzyme Activation / Q000187:drug effects; D007700:Kinetics; D008433:Mathematics; D010727:Phosphoric Diester Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D011506:Proteins / Q000494:pharmacology*
D002614:Chelating Agents; D011506:Proteins; D000242:Cyclic AMP; D010727:Phosphoric Diester Hydrolases; D006152:Cyclic GMP; D002118:Calcium