The Cu chaperone CopZ is required for Cu homeostasis in <i>Rhodobacter capsulatus</i> and influences cytochrome <i>cbb</i>₃ oxidase assembly

Abstract

Cu homeostasis depends on a tightly regulated network of proteins that transport or sequester Cu, preventing the accumulation of this toxic metal while sustaining Cu supply for cuproproteins. In Rhodobacter capsulatus, Cu-detoxification and Cu delivery for cytochrome c oxidase (cbb₃ -Cox) assembly depend on two distinct Cu-exporting P_1B -type ATPases. The low-affinity CopA is suggested to export excess Cu and the high-affinity CcoI feeds Cu into a periplasmic Cu relay system required for cbb₃ -Cox biogenesis. In most organisms, CopA-like ATPases receive Cu for export from small Cu chaperones like CopZ. However, whether these chaperones are also involved in Cu export via CcoI-like ATPases is unknown. Here we identified a CopZ-like chaperone in R. capsulatus, determined its cellular concentration and its Cu binding activity. Our data demonstrate that CopZ has a strong propensity to form redox-sensitive dimers via two conserved cysteine residues. A ΔcopZ strain, like a ΔcopA strain, is Cu-sensitive and accumulates intracellular Cu. In the absence of CopZ, cbb₃ -Cox activity is reduced, suggesting that CopZ not only supplies Cu to P_1B -type ATPases for detoxification but also for cuproprotein assembly via CcoI. This finding was further supported by the identification of a ~150 kDa CcoI-CopZ protein complex in native R. capsulatus membranes.