Differences in binding properties of two proton pump inhibitors on the gastric H+,K+-ATPase in vivo

Abstract

Restoration of acid secretion after treatment with covalently-bound proton pump inhibitors may depend on protein turnover and on reversal of inhibition by reducing agents such as glutathione. Glutathione incubation of the H(+),K(+)-ATPase isolated from omeprazole or pantoprazole-treated rats reversed 88% of the omeprazole inhibition but none of the pantoprazole inhibition. The present study was designed to measure binding properties of omeprazole or pantoprazole in vivo. Rats were injected with (14)C-omeprazole or (14)C-pantoprazole after acid stimulation. The specific binding to the gastric H(+),K(+)-ATPase was measured at timed intervals as well as reversal of binding by glutathione reduction. The stoichiometry of omeprazole and pantoprazole binding to the catalytic subunit of the H(+),K(+)-ATPase was 2 moles of inhibitor per mole of the H(+),K(+)-ATPase phosphoenzyme. Omeprazole bound to one cysteine between transmembrane segments 5/6 and one between 7/8, pantoprazole only to the two cysteines in the TM5/6 domain. Loss of drug from the pump was biphasic, the fast component accounted for 84% of omeprazole binding and 51% of pantoprazole binding. Similarly, only 16% of omeprazole binding but 40% of pantoprazole binding was not reversed by glutathione. The residence time of omeprazole and pantoprazole on the ATPase in vivo depends on the reversibility of binding. Binding of pantoprazole at cysteine 822 is irreversible whereas that of omeprazole at cysteine 813 and 892 is reversible both in vivo and in vitro. This is consistent with the luminal exposure of cysteine 813 and 892 and the intra-membranal location of cysteine 822 in the 3D structure of the H(+),K(+)-ATPase.