A new metal binding domain involved in cadmium, cobalt and zinc transport

The P(1B)-ATPases, which couple cation transport across membranes to ATP hydrolysis, are central to metal homeostasis in all organisms. An important feature of P(1B)-ATPases is the presence of soluble metal binding domains that regulate transport activity. Only one type of MBD has been characterized extensively, but bioinformatics analyses indicate that a diversity of MBDs may exist in nature. Here we report the biochemical, structural, and functional characterization of a new MBD from the Cupriavidus metallidurans P(1B-4)-ATPase CzcP (CzcP MBD). The CzcP MBD binds two Cd(2+), Co(2+), or Zn(2+) ions in distinct and unique sites, and adopts an unexpected fold consisting of two fused ferredoxin-like domains. Both in vitro and in vivo activity assays using full length CzcP, truncated CzcP, and several variants indicate a regulatory role for the MBD and distinct functions for the two metal binding sites. Taken together, these findings elucidate a previously unknown MBD and suggest new regulatory mechanisms for metal transport by P(1B)-ATPases.