An essential role of N-terminal domain of copper chaperone in the enzymatic activation of Cu/Zn-superoxide dismutase

Abstract

Cu/Zn-superoxide dismutase (SOD1) is an enzyme that disproportionates superoxide anion into hydrogen peroxide and molecular oxygen. The enzymatic activity of SOD1 requires the binding of copper and zinc ions and also the formation of a conserved intramolecular disulfide bond. In a eukaryotic cell, a copper chaperone for SOD1 (CCS) has been known to supply a copper ion and also introduce the disulfide bond into SOD1; however, a mechanism controlling the CCS-dependent activation of SOD1 remains obscure. Here, we characterized CCS isolated from a human liver fluke, Clonorchis sinensis, and found that an N-terminal domain of CCS was essential in supplying a copper ion in SOD1. Regardless of the presence and absence of the N-terminal domain, CCS was able to bind a cuprous ion at the CxC motif of its C-terminal domain with quite high affinity (Kd~10−17). The copper-bound form of full-length CCS successfully activated C. sinensis SOD1, but that of CCS lacking the N-terminal domain did not. Nonetheless, the N-terminally truncated CCS with the bound copper ion was found to correctly introduce the disulfide bond into SOD1. Based upon these results, we propose that the N-terminal domain of CCS has roles in the release of the copper ion bound at the C-terminal domain of CCS to SOD1.