The process of hepatobiliary
copper (Cu) secretion is still poorly understood: Cu secretion as a complex with
glutathione and transport via a lysosomal pathway have been proposed. The recent cloning and sequencing of the gene for
Wilson disease indicates that Cu transport in liver cells may be mediated by a Cu transporting
P-type ATPase. Biochemical evidence for
ATP-dependent Cu transport in mammalian systems, however, has not been reported so far. We have investigated Cu transport in rat liver plasma membrane vesicles enriched in canalicular or basolateral membranes in the presence and absence of
ATP (4 mM) and an
ATP-regenerating system. The presence of
ATP clearly stimulated uptake of radiolabeled Cu (64Cu, 10 microM) into canalicular plasma membrane vesicles and, to a lesser extent, also into basolateral plasma membrane vesicles.
ATP-dependent Cu transport was dose-dependently inhibited by the
P-type ATPase inhibitor
vanadate, and showed saturation kinetics with an estimated Km of 8.6 microM and a Vmax of 6.9 nmol/min/mg
protein.
ATP-stimulated Cu uptake was similar in canalicular membrane vesicles of normal Wistar rats and those of mutant GY rats, expressing a
congenital defect in the activity of the
ATP-dependent canalicular
glutathione-conjugate transporter (
cMOAT). These studies demonstrate the presence of an
ATP-dependent Cu transporting system in isolated plasma membrane fractions of rat liver distinct from
cMOAT.