Functional properties of an H,K-ATPase/Na,K-ATPase chimera.

Cultured pig kidney epithelial cells were transfected with a chimeric P-type ATPase catalytic subunit composed of the NH2-terminal half of the rat gastric H,K-ATPase and the COOH-terminal half of the rat Na,K-ATPase (alpha 1 isoform). Low concentrations of ouabain (< or = 0.2 mM) were used to inhibit completely the endogenous pig Na,K-ATPase and high concentrations (5 nM) to test the sensitivity of the chimeric rodent pump. In the presence of a low concentration of ouabain, a small but significant inhibition of residual Rb+(K+) influx by 5 mM ouabain was observed in only the transfected cells. Conditions were found in which a similar component of Rb+ influx was inhibited by the gastric H,K-ATPase inhibitor SCH28080, consistent with SCH28080 binding to the extracellular H1-H2 loop of this enzyme. These experiments demonstrate that this chimera behaves as a functional ion pump and indicate that the protein domains involved in cardiac glycoside binding are not confined to the amino-terminal half of the Na,K-ATPase.