A hybrid between Na+,K+-ATPase and H+,K+-ATPase is sensitive to palytoxin, ouabain, and SCH 28080

Abstract

Na(+),K(+)-ATPase is inhibited by cardiac glycosides such as ouabain, and palytoxin, which do not inhibit gastric H(+),K(+)-ATPase. Gastric H(+),K(+)-ATPase is inhibited by SCH28080, which has no effect on Na(+),K(+)-ATPase. The goal of the current study was to identify amino acid sequences of the gastric proton-potassium pump that are involved in recognition of the pump-specific inhibitor SCH 28080. A chimeric polypeptide consisting of the rat sodium pump alpha3 subunit with the peptide Gln(905)-Val(930) of the gastric proton pump alpha subunit substituted in place of the original Asn(886)-Ala(911) sequence was expressed together with the gastric beta subunit in the yeast Saccharomyces cerevisiae. Yeast cells that express this subunit combination are sensitive to palytoxin, which interacts specifically with the sodium pump, and lose intracellular K(+) ions. The palytoxin-induced K(+) efflux is inhibited by the sodium pump-specific inhibitor ouabain and also by the gastric proton pump-specific inhibitor SCH 28080. The IC(50) for SCH 28080 inhibition of palytoxin-induced K(+) efflux is 14.3 +/- 2.4 microm, which is similar to the K(i) for SCH 28080 inhibition of ATP hydrolysis by the gastric H(+),K(+)-ATPase. In contrast, palytoxin-induced K(+) efflux from cells expressing either the native alpha3 and beta1 subunits of the sodium pump or the alpha3 subunit of the sodium pump together with the beta subunit of the gastric proton pump is inhibited by ouabain but not by SCH 28080. The acquisition of SCH 28080 sensitivity by the chimera indicates that the Gln(905)-Val(930) peptide of the gastric proton pump is likely to be involved in the interactions of the gastric proton-potassium pump with SCH 28080.