Structure/Function Studies of the Gamma Subunit of the Na,K‐ATPase

: The Na,K-ATPase γ subunit is present primarily in kidney as two splice variants, γa and γb, which differ only at their extracellular N-termini. Two distinct effects of gamma are seen in biochemical Na,K-ATPase assays of mammalian (HeLa) cells transfected with γa or γb, namely, (i) a decrease in K′ATP probably secondary to a shift in steady-state E1 E2 poise in favor of E1 and (ii) an increase in cytoplasmic K+/Na+ antagonism seen as an increase in K′Na at high K+ concentration. Mutagenesis experiments involving alterations in extramembranous regions of γ indicate that different regions mediate the aforementioned distinct effects and that the effects appear to be long range. Studies of ouabain-sensitive fluxes with intact cells confirm the γ effects seen with membranes and also suggest an additional effect (increase) in apparent affinity for extracellular K+. Alteration in gamma function was also evidenced in the behavior of a G41 R mutation within the transmembrane domain of gamma. G41R is associated with autosomal dominant renal magnesium wasting. Our studies show that this mutation in the γb variant retards trafficking of γ, but not αβ pumps, to the cell surface and abolishes functional effects of γ, consistent with the conclusion that the Mg2+ transport defect is secondary to loss of γ modulation of Na,K-ATPase function.