Interaction of Ca2+ with (Na+ + K+)-ATPase: properties of the Ca2+-stimulated phosphatase activity.

Ca2+ inhibited the Mg2+-dependent and K+-stimulated p-nitrophenylphosphatase activity of a highly purified preparation of dog kidney (Na+ + K+)-ATPase. In the absence of K+, however, a Mg2+-dependent and Ca2+-stimulated phosphatase was observed, the maximal velocity of which, at pH 7.2, was about 20% of that of the K+-stimulated phosphatase. The Ca2+-stimulated phosphatase, like the K+-stimulated activity, was inhibited by either ouabain or Na+ or ATP. Ouabain sensitivity was decreased with increase in Ca2+, but the K0.5 values of the inhibitory effects of Na+ and ATP were independent of Ca2+ concentration. Optimal pH was 7.0 for Ca2+-stimulated activity, and 7.8-8.2 for the K+-stimulated activity. The ratio of the two activities was the same in several enzyme preparations in different states of purity. The data indicate that (a) Ca2+-stimulated phosphatase is catalyzed by (Na+ + K+)-ATPase; (b) there is a site of Ca2+ action different from the site at which Ca2+ inhibits in competition with Mg2+; and (c) Ca2+ stimulation can not be explained easily by the action of Ca2+ at either the Na+ site or the K+ site.