Regulation of brain cyclic nucleotide phosphodiesterase by calmodulin. A quantitative analysis.
Comparison of the parameters of Ca and Sr binding to bovine brain calmodulin with the activation of bovine brain phosphodiesterase by Ca2+ and Sr2+ at different calmodulin concentrations allows a quantitative description of the mechanism of activation of the enzyme. Equilibrium dialysis studies show that calmodulin possesses three high affinity (K'diss = 6 micoM) and one low affinity (K'diss = 200 microM) sites for Ca2+. All four sites display the same affinity for Sr2+ with K'diss = 180 microM. In the presence of calmodulin, soluble bovine brain phosphodiesterase is activated by Sr2+ to the same extent as by Ca2+. The activation of the enzyme shows the same Ca2+/Sr2+ selectivity ratio of 30 as the binding of the metal ions to calmodulin. Based on the findings that the Ca2+ or Sr2+ concentration at half-maximal activation of the enzyme depends on the concentration of calmodulin present, a quantitative analysis of activation was carried out as a function of the four calmodulin-metal complex species (CaM . Men). The data show that the activating species are CaM . Ca3, CaM . Ca4 or CaM . Sr3, CaM . Sr4. The interaction of these activating species with phosphodiesterase follows the Hill equation with a dissociation constant of 10(-9) M and a Hill coefficient of 2, irrespective of the binding characteristics of Ca2+ or Sr2+. The latter value agrees well with the fact that phosphodiesterase possesses two binding sites for calmodulin.
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002417:Cattle; D007700:Kinetics; D046911:Macromolecular Substances; D008274:Magnesium / Q000494:pharmacology; D011485:Protein Binding; D013324:Strontium / Q000494:pharmacology
D002135:Calcium-Binding Proteins; D002147:Calmodulin; D046911:Macromolecular Substances; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D008274:Magnesium; D002118:Calcium; D013324:Strontium