Functional characterization of an E2P-like structure of the Na,K-ATPase that crystallized with K+

MÓNICA R. MONTES; MARIELA S. FERREIRA-GOMES; ROLANDO C. ROSSI

Tucuman

Congreso; XLI Reunión Anual de la Sociedad Argentina de Biofísica; 2012

Sociedad Argentina de Biofísica

The Na,K-ATPase generates electrochemical gradients exchanging three sodium ions for two potassium ions across the plasma membrane during each cycle of ATP hydrolysis. The crystal structure of the sodium pump with occluded K+ (or Rb+) could only be obtained in the presence of a magnesium fluoride complex (1), which was modeled as the tetrahedral (MgF4)2-. Complexes of magnesium with fluoride inhibit ATPase activity by stabilizing an E2P-like structure (2). Although so far three crystal structures of the K+-occluded state of the Na, K-ATPase in complex with (MgF4)2- have been obtained, there are no studies analyzing its kinetic behavior. In this work we investigated the rates at which Rb+ enters and leaves the enzyme-magnesium fluoride complex. We found that magnesium fluoride induces a transition of the Na,K-ATPase to the E2 conformation which occludes Rb+ at a very low rate. This occluded complex is unable to return to E1 and release the cation to the intracellular side as it occurs in the physiological cycle. 1-Morth J., Pedersen B., Toustrup-Jensen J., Sorensen T., Petersen., Andersen J., Bilsen V., and Nissen P., Nature, 450, 1043 (2007). 2-Clausen J., McIntosh D., Woolley D., Andersen J., J. Biol. Chem. 286, 11792 (2011) With grants of CONICET and UBACYT