Characterization of the reaction kinetics of cations with the Na+/K+-ATPase measuring their effect on Rb+ occlusion.

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

San Carlos de Bariloche, Río Negro, Argentina

Congreso; Reunión internacional conjunta: XXXII reunión anual de la Sociedad Argentina de Biofísica (SAB), XXXIX reunión anual de la Sociedad Argentina de Investigaciones en Bioquímica y Biología Molecular (SAIB) y Bariloche Protein Symposium; 2003

Sociedad Argentina de Biofísica, y Sociedad Argentina de Investigaciones en Bioquímica y Biología Molecular

The active transport of Na+ and K+ across cell membranes is mediated by the Na+/K+ pump, which is an intrinsic membrane transport system, identical with the Na+/K+-ATPase. There is evidence that during the pump cycle 2 K+ become bound to the Na+/K+-ATPase in such a way that their release is slow. This type of binding is called occlusion because it is believed that their access to the bulk of the solvent is restricted by the protein structure. We here show that indirect information on the reactions between the enzyme and ligands like Na+, K+, Rb+, NH4+, Li+, Tl+ or Mg2+ can be obtained from their effects on the kinetics of Rb+ occlusion. To measure the time course of 86Rb+ occlusion each cation was either equilibrated with the enzyme before adding 20 µM 86Rb+ (condition A) or added to the enzyme together with 86Rb+ (condition B). When the cation was K+, Rb+, NH4+ or Tl+ the rate of 86Rb+ occlusion was much slower for condition A than for condition B. This result was expected since K+, Rb+ and Tl+ are known to become occluded themselves. When Na+, Mg2+ or Li+ were tested, no significant difference between the time courses for both conditions were observed. Compared to those obtained for K+ and its congeners results for Na+, Mg2+ and Li+, strongly suggest that neither cation forms occluded states under these conditions.