Characterization of the Menkes protein copper-binding domains and their role in copper-induced protein relocalization

Menkes disease is a fatal X-linked disorder of copper metabolism. The gene defective in Menkes disease (ATP7A) encodes a copper transporting P-type AT Pase (MNK or ATP7A) with six copper-binding domains at its N-terminus, MNK is normally localized to the trans Golgi network in cultured cells, but rel ocates to the plasma membrane in the presence of elevated extracellular cop per. In this study, the role of the six copper-binding domains on copper-in duced redistribution is investigated. In a recombinant clone, when all the wildtype copper-binding motifs are mutated from GMXCXXC to GMXSXXS and the cells grown in medium containing elevated copper, relocalization of the rec ombinant protein to the plasma membrane was not observed, Using the same as say with any one of the six copper-binding domains intact, MNK moves to the plasma membrane in a way indistinguishable from the wild-type protein. The refore, the copper-binding domains are vital for MNK trafficking and only a single domain is sufficient for this redistribution to occur.