Cu-specific CopB transporter: Revising P-type ATPase classification.

The copper-transporting P-ATPases, which play a key role in cellular copper homeostasis, have been divided traditionally into two subfamilies, the P-ATPases or CopAs and the P-ATPases or CopBs. CopAs selectively export Cu whereas previous studies and bioinformatic analyses have suggested that CopBs are specific for Cu export. Biochemical and spectroscopic characterization of CopB (CopB) show that, while it does bind Cu, the binding site is not the prototypical P-ATPase transmembrane site and does not involve sulfur coordination as proposed previously. Most important, CopB exhibits metal-stimulated ATPase activity in response to Cu, but not Cu, indicating that it is actually a Cu transporter. X-ray absorption spectroscopic studies indicate that Cu is coordinated by four sulfur ligands, likely derived from conserved cysteine and methionine residues. The histidine-rich N-terminal region of CopB is required for maximal activity, but is inhibitory in the presence of divalent metal ions. Finally, reconsideration of the P-ATPase classification scheme suggests that the P- and PATPase subfamilies both comprise Cu transporters. These results are completely consistent with the known presence of only Cu within the reducing environment of the cytoplasm, which should eliminate the need for a Cu P-ATPase.