Ligand-Dependent Effects on the Conformational Equilibrium of the Na+,K+-ATPase As Monitored by Voltage Clamp Fluorometry

AUTOR(ES)

Geys, Stefan A.

FONTE

The Biophysical Society

RESUMO

Voltage clamp fluorometry was used to monitor conformational changes associated with electrogenic partial reactions of the Na+,K+-ATPase after changes in the concentration of internal sodium (Na+i) or external potassium (K+o). To probe the effects of the Na+i concentration on the Na+ branch of the Na+,K+-ATPase, oocytes were depleted of Na+i and then loaded with external sodium (Na+o) using the amiloride-sensitive epithelial sodium channel. The K+ branch of the Na+,K+-ATPase was studied by exposing the oocytes to different K+o concentrations in the presence and absence of Na+o to obtain additional information on the apparent affinity for K+o. Our results demonstrate that lowering the concentration of Na+i or increasing the amount of K+o in the external solution shifts the equilibrium toward E1/E1P. Furthermore, the K+o-induced relocation toward E1 occurs at a much lower K+o concentration when Na+o is absent, indicating a higher apparent affinity. Finally, voltage-dependent steps associated with the K+ branch or the Na+ branch of the Na+,K+-ATPase are affected by the K+o concentration or the Na+i concentration, respectively.

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