ADENOSINE TRIPHOSPHATE-DEPENDENT COPPER TRANSPORT IN ISOLATED RAT-LIVER PLASMA-MEMBRANES

The process of hepatobiliary copper (Cu) secretion is still poorly und erstood: Cu secretion as a complex with glutathione and transport via a lysosomal pathway have been proposed. The recent cloning and sequenc ing of the gene for Wilson disease indicates that Cu transport in live r cells may be mediated by a Cu transporting P-type ATPase. Biochemica l evidence for ATP-dependent Cu transport in mammalian systems, howeve r, has not been reported so far. We have investigated Cu transport in rat liver plasma membrane vesicles enriched in canalicular or basolate ral membranes in the presence and absence of ATP (4 mM) and an ATP-reg enerating system. The presence of ATP clearly stimulated uptake of rad iolabeled Cu ((CU)-C-64, 10 mu M) into canalicular plasma membrane ves icles and, to a lesser extent, also into basolateral plasma membrane v esicles. ATP-dependent Cu transport was dose-dependently inhibited by the P-type ATPase inhibitor vanadate, and showed saturation kinetics w ith an estimated K-m of 8.6 mu M and a V-max of 6.9 nmol/min/mg protei n. ATP-stimulated Cu uptake was similar in canalicular membrane vesicl es of normal Wistar rats and those of mutant GY rats, expressing a con genital defect in the activity of the ATP-dependent canalicular glutat hione-conjugate transporter (cMOAT). These studies demonstrate the pre sence of an ATP-dependent Cu transporting system in isolated plasma me mbrane fractions of rat liver distinct from cMOAT.