Sodium‐sodium exchange through the sodium pump: the roles of ATP and ADP.

Abstract

A procedure was developed for preparing resealed red cell ghosts that contain ADP but very little ATP. The procedure involves lysis of the cells in a very large volume of lysing solution, resuspension of the ghosts in a small volume, the incorporation into the ghosts, before they resealed, of the adenylate kinase inhibitor P1,P5-di(adenosine-5''-)pentaphosphate (AP5A) and of hexokinase, and (i.v.) the removal of traces of ATP, formed by residual adenylate kinase activity, by the addition of glucose. Measurements of Na efflux from ghosts prepared in this way show that Na-Na exchange through the Na pump does not occur in the absence of ATP even if ADP is present. The .beta.:.gamma. imido analogue of ATP, which is incapable of phosphorylating Na, K-ATPase, cannot replace ATP in supporting Na-Na exchange. The outward movement of Na ions through the Na pump is associated with the transfer of a phosphoryl group from ATP to the enzyme, and the inward movement of Na ions through the pump is associated with the return of a phosphoryl group from the phosphoenzyme to ADP.