The alpha-subunit of the Na,K-ATPase has catalytic activity independent of the beta-subunit.

All catalytic activities of the Na,K-ATPase have been ascribed to the alpha-subunit; however, normal activity requires the presence of the beta-subunit. Using recombinant baculoviruses to infect insect cells, we demonstrate that the alpha-subunit, without the beta-subunit, has catalytic activity. During the normal catalytic cycle of the Na,K-ATPase, the alpha-subunit is transiently phosphorylated by ATP at an aspartate residue. This phosphorylation requires Na+, in the presence of K+ the enzyme undergoes rapid dephosphorylation. In contrast, phosphorylation of the independent alpha-subunit by ATP occurs in the presence of Mg2+, does not require Na+ or K+, and is not affected by ouabain. The phosphorylation is, however, inhibited by EGTA and increasing ionic strength. Chemical properties of the alpha-subunit phosphointermediate are consistent with phosphorylation at the normal aspartyl residue. Membranes from cells infected with the recombinant alpha baculovirus exhibit an EGTA-sensitive Mg(2+)-ATPase activity that is not present in the uninfected cells. The Mg(2+)-ATPase of the alpha-infected cells is reduced under conditions of high ionic strength and completely inhibited by EGTA. Thus the phosphorylation of the unassociated alpha-subunit is representative of the ATPase activity of the enzyme. These results suggest that the alpha-subunit of the Na,K-ATPase can catalyze an activity not normally associated with the enzyme and demonstrate that the bea-subunit plays an important role in conferring normal activity to the enzyme complex.