All catalytic activities of the Na,K-ATPase have been ascribed to the α- subunit; however, normal activity requires the presence of the β-subunit. Using recombinant baculoviruses to infect insect cells, we demonstrate that the α-subunit, without the β-subunit, has catalytic activity. During the normal catalytic cycle of the Na,K-ATPase, the α-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 α-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 α-subunit phosphointermediate are consistent with phosphorylation at the normal aspartyl residue. Membranes from cells infected with the recombinant α baculovirus exhibit an EGTA- sensitive Mg2+-ATPase activity that is not present in the uninfected cells. The Mg2+-ATPase of the α-infected cells is reduced under conditions of high ionic strength and completely inhibited by EGTA. Thus the phosphorylation of the unassociated α-subunit is representative of the ATPase activity of the enzyme. These results suggest that the α-subunit of the Na,K-ATPase can catalyze an activity not normally associated with the enzyme and demonstrate that the β-subunit plays an important role in conferring normal activity to the enzyme complex.
|Number of pages||6|
|Journal||Journal of Biological Chemistry|
|State||Published - Sep 23 1994|