The Calcium Binding Sites Involved in the Regulation of the Purified Adenosine Triphosphatase of the Sarcoplasmic Reticulum

The role of the interaction of Ca2+ with the purified Ca2+-dependent ATPase in the regulation of enzyme activity has been investigated. It appears that the sensitivity to Ca2+ of the sarcoplasmic reticulum, resulting in activation and inhibition of ATPase activity, is intrinsic to the ATPase moiety of the membrane. Three types of Ca2+ binding sites have been found in equilibrium dialysis studies. In the absence of ATP there is approximately one of each per 105 daltons; the binding constants are 4 x 106 m-1 (α site), 4 x 104 (β site), and 1 x 103 (γ site). Addition of 1.5 mm ATP slightly increases the affinity of all sites and reduces the apparent capacity of the α and β sites. Study of ATP hydrolysis in parallel with calcium binding has shown that Ca2+ binding at the α site activates it and binding to the γ site inhibits it, while binding of Ca2+ to the β site appears not to be involved in the enzymatic regulation.