Adenosine triphosphate-dependent copper transport in human liver.
OBJECTIVE
The recent cloning and sequencing of the Wilson disease gene indicates that hepatic copper (Cu) transport is mediated by a P-type ATPase. The location of this Cu-transporting protein within the hepatocyte is not known; in view of its proposed function and current concepts of hepatic Cu transport, it may reside in intracellular membranes (endoplasmic reticulum (ER), lysosomes) and/or in the bile canalicular membrane. The objective of this study was to establish characteristics and localization of ATP-dependent Cu transport in human liver.


METHODS
We have investigated Cu transport in vesicles of human liver plasma membranes showing a gradual increase in enrichment of canalicular domain markers: i.e. basolateral liver plasma membranes (blLPM), a mixed population of basolateral and canalicular (XLPM) and canalicular liver plasma membranes (cLPM).


RESULTS
In the presence of ATP (4 mM) and an ATP-regenerating system, uptake of radiolabeled Cu (64Cu, 10 microM) into cLPM vesicles and, to a lesser extent, into blLPM and XLPM was clearly stimulated when compared to control AMP values. Initial uptake rates of ATP-dependent Cu transport were 5.6, 7.8 and 13.7 nmol.min-1.mg-1 protein for blLPM, XLPM and cLPM, respectively, and showed no relationship with marker enzyme activity of ER and lysosomes (glucose-6-phosphatase and acid-phosphatase, respectively). Leucine aminopeptidase activity, as a marker for the cLPM, significantly correlated with ATP-dependent uptake rates measured in different membrane preparations: r = 0.70 (n = 9, p < 0.05). Estimated K(m) and Vmax values of ATP-dependent Cu uptake were 49.5 microM and 36.9 nmol.min-1.mg-1 protein, respectively.


CONCLUSIONS
This study provides biochemical evidence for the presence of an ATP-dependent Cu transport system in human liver (cCOP), mainly localized at the canalicular domain of the hepatocytic plasma membrane.
Dijkstra
M
M
Department of Pediatrics, Groningen Institute for Drug Studies, University Hospital Groningen, The Netherlands.
van den Berg
G J
GJ
Wolters
H
H
In't Veld
G
G
Slooff
M J
MJ
Heymans
H S
HS
Kuipers
F
F
Vonk
R J
RJ
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D001648:Bile Canaliculi / Q000378:metabolism; D001692:Biological Transport; D003300:Copper / Q000378:metabolism*; D006801:Humans; D008099:Liver / Q000378:metabolism*; D009994:Osmolar Concentration
D003300:Copper; D000255:Adenosine Triphosphate