Are the states that occlude rubidium obligatory intermediates of the Na+/K+-ATPase reaction?

S. B. KAUFMAN; R. M. GONZÁLEZ-LEBRERO; P. J. SCHWARZBAUM; J. G. NØRBY; P. J. GARRAHAN; R. C. ROSSI

JOURNAL OF BIOLOGICAL CHEMISTRY

American Society for Biochemistry and Molecular Biology

Año: 1999 vol. 274 p. 20779 - 20790

0021-9258

In the Albers-Post model, occlusion of K1 in the E2 conformer of the enzyme (E) is an obligatory step of Na+/K+-ATPase reaction. If this were so the ratio (Na+/K+-ATPase activity)/(concentration of occluded species) should be equal to the rate constant for deocclusion. We tested this prediction in a partially purified Na+/K+-ATPase from pig kidney by means of rapid filtration to measure the occlusion using the K+ congener Rb+. Assuming that always two Rb+ are occluded per enzyme, the steady-state levels of occluded forms and the kineticsof deocclusion were adequately described by the Albers-Post model over a very wide range of [ATP] and [Rb+]. The same happened with the kinetics of ATP hydrolysis. However, the value of the parameters that gave best fit differed from those for occlusion in such a way that the ratio (Na+/K+-ATPase activity)/(concentration of occluded species) became much larger than the rate constant for deocclusion when [Rb+] <10 mM. This points to the presence of an extra ATP hydrolysis that is not Na+-ATPase activity and that does not involve occlusion. A possible way of explaining this is to posit that the binding of a single Rb+ increases ATP hydrolysis without occlusion.