The roles of Rhodobacter sphaeroides copper chaperones PCuAC and Sco (PrrC) in the assembly of the copper centers of the aa3-type and the cbb3-type cytochrome c oxidases

The α proteobacter Rhodobacter sphaeroides accumulates two cytochrome c oxidases (CcO) in its cytoplasmic membrane during aerobic growth: a mitochondrial-like aa3-type CcO containing a di-copper CuA center and mono-copper CuB, plus a cbb3-type CcO that contains CuB but lacks CuA. Three copper chaperones are located in the periplasm of R. sphaeroides, PCuAC, PrrC (Sco) and Cox11. Cox11 is required to assemble CuB of the aa3-type but not the cbb3-type CcO. PrrC is homologous to mitochondrial Sco1; Sco proteins are implicated in CuA assembly in mitochondria and bacteria, and with CuB assembly of the cbb3-type CcO. PCuAC is present in many bacteria, but not mitochondria. PCuAC of Thermus thermophilus metallates a CuA center in vitro, but its in vivo function has not been explored. Here, the extent of copper center assembly in the aa3- and cbb3-type CcOs of R. sphaeroides has been examined in strains lacking PCuAC, PrrC, or both. The absence of either chaperone strongly lowers the accumulation of both CcOs in the cells grown in low concentrations of Cu2 +. The absence of PrrC has a greater effect than the absence of PCuAC and PCuAC appears to function upstream of PrrC. Analysis of purified aa3-type CcO shows that PrrC has a greater effect on the assembly of its CuA than does PCuAC, and both chaperones have a lesser but significant effect on the assembly of its CuB even though Cox11 is present. Scenarios for the cellular roles of PCuAC and PrrC are considered. The results are most consistent with a role for PrrC in the capture and delivery of copper to CuA of the aa3-type CcO and to CuB of the cbb3-type CcO, while the predominant role of PCuAC may be to capture and deliver copper to PrrC and Cox11. This article is part of a Special Issue entitled: Biogenesis/Assembly of Respiratory Enzyme Complexes.

► The presence of Sco strongly enhances the assembly of CuA of aa3-type CcO.
► The presence of Sco strongly enhances the assembly of CuB of cbb3-type CcO.
► The Cu chaperone PCuAC enhances assembly but requires the presence of Sco.