Effects of alkali metal cations on phospho-enzyme levels and [³H]ouabain binding to (Na⁺ + K⁺)-ATPase

The effects of several alkali metal cations on the relationship between steady state phospho-enzyme levels and initial velocity and equilibrium levels of [³H]-ouabain binding to (Na⁺ + K⁺)-ATPase (ATP phosphohydrolase EC 3.6.1.3.) were examined. Only Na⁺ increased both phospho-enzyme and [³H] ouabain binding livels above those observed in the presence of Mg²⁺ alone. While Na⁺ stimulated phosphorylation with an apparent K_m of about 1 mM, its stimulation of [³H] ouabain binding was biphasic, the lower K_m for stimulation corresponding to the K_m for formation of phospho-enzyme. Among the other alkali metal cations, potassium, rubidium and lithium were at least eight times more effective in reducing phospho-enzyme levels than in reducing [³H] ouabain binding. This discrepancy is not due to the stability of the enzyme-ouabain complex, nor to any action on the rates of formation or dissociation of the enzyme-ouabain complex. The data thus suggest that [³H] ouabain interacts with the K⁺, Rb⁺ or Li⁺-enzyme complexes. For Li⁺, this hypothesis is further supported by the observation that Li⁺ can directly increase the equilibrium level of [³H]-ouabain binding to this enzyme under certain conditions.