COPPER AND SILVER TRANSPORT BY COPB-ATPASE IN MEMBRANE-VESICLES OF ENTEROCOCCUS-HIRAE

The P-type ATPase, CopB, of Enterococcus hirae is required for the copper resistance displayed by this organism and thus was postulated to be a copper pump. Using Cu-64(+) and Ag-110m(+), we here show ATP-driven copper and silver accumulation catalyzed by CopB in native inside-out membrane vesicles of E. hirae. CopB ATPase exhibited an apparent K-m for Cu+ and Ag+ of 1 mu M and for ATP of 10 mu M. Transport was maximal at pH 6 and had an apparent V-max of 0.07 nmol . min(-1). mg(-1) for both copper and silver transport. Vanadate displayed a biphasic effect on transport: maximal inhibition was observed at 40 mu M vanadate for copper transport and 60 mu M for silver transport, respectively. At higher vanadate concentrations, these inhibitions were reversed. The CopB ATPase off. hirae is thus a pump for the extrusion of monovalent copper and silver ions, with copper probably being the natural substrate.