Multiple Protein Domains Contribute to the Action of the Copper Chaperone for Superoxide Dismutase

The Cu- and Zn-containing superoxide dismutase 1 (SOD1) largely obtains Cu in vivo by means of the action of the Cu chaperone CCS. Yet, in the case of mammalian SOD1, a secondary pathway of activation is apparent. Specifically, when human SOD1 is expressed in either yeast or mammalian cells that are null for CCS, the SOD1 enzyme retains a certain degree of activity. This CCS-independent activity is evident with both wild-type and mutant variants of SOD1 that have been associated with familial amyotrophic lateral sclerosis. We demonstrate here that the CCS-independent activation of mammalian SOD1 involves glutathione, particularly the reduced form, or GSH. A role for glutathione in CCS-independent activation was seen with human SOD1 molecules that were expressed in either yeast cells or immortalized fibroblasts. Compared with mammalian SOD1, the Saccharomyces cerevisiae enzyme cannot obtain Cu without CCS in vivo, and this total dependence on CCS involves the presence of dual prolines near the C terminus of the SOD1 polypeptide. Indeed, the insertion of such prolines into human SOD1 rendered this molecule refractory to CCS-independent activation. The possible implications of multiple pathways for SOD1 activation are discussed in the context of SOD1 evolutionary biology and familial amyotrophic lateral sclerosis

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“…The complete dependence of ySOD1 on CCS for activity (also shown in Fig. 5B) is consistent with several studies (4,13,31,32).…”

Section: Ccs-independent Activation Of Human Wild-type and Fals Mutantsupporting

confidence: 92%

Mechanisms for activating Cu- and Zn-containing superoxide dismutase in the absence of the CCS Cu chaperone

Carroll

Girouard

Ulloa

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

166

201

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“…The human CCS N-terminal domain 1 (D1) is homologous and shows the same fold of the Atx1-like metallochaperones and harbors the MXCXXC copper-binding motif typical of this family of proteins. Human D1 is essential for hSOD1 copper acquisition in vivo (32), whereas yeast D1 appears to be necessary only under copper-limiting conditions (33). CCS domain 2 (D2) is structurally similar to SOD1 and has been postulated to be critical for CCS-SOD1 protein recognition (33)(34)(35).…”

mentioning

confidence: 99%

Human superoxide dismutase 1 (hSOD1) maturation through interaction with human copper chaperone for SOD1 (hCCS)

Banci

Bertini

Cantini

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

128

149

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Copper chaperone for superoxide dismutase 1 (SOD1), CCS, is the physiological partner for the complex mechanism of SOD1 maturation. We report an in vitro model for human CCS-dependent SOD1 maturation based on the study of the interactions of human SOD1 (hSOD1) with full-length WT human CCS (hCCS), as well as with hCCS mutants and various truncated constructs comprising one or two of the protein’s three domains. The synergy between electrospray ionization mass spectrometry (ESI-MS) and NMR is fully exploited. This is an in vitro study of this process at the molecular level. Domain 1 of hCCS is necessary to load hSOD1 with Cu(I), requiring the heterodimeric complex formation with hSOD1 fostered by the interaction with domain 2. Domain 3 is responsible for the catalytic formation of the hSOD1 Cys-57–Cys-146 disulfide bond, which involves both hCCS Cys-244 and Cys-246 via disulfide transfer.

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“…Biochemical analysis demonstrates that yeast CCS is sufficient to incorporate Cu into SOD1 through direct transfer under restricted concentrations of intracellular free Cu. Moreover, structural and functional studies of yeast CCS indicate that this is accomplished by a complex intermolecular reaction involving a unique domain structure of the CCS molecule (7,8).…”

mentioning

confidence: 99%

Copper chaperone for superoxide dismutase is essential to activate mammalian Cu/Zn superoxide dismutase

Wong

Waggoner

Subramaniam

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

293

230

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Recent studies in Saccharomyces cerevisiae suggest that the delivery of copper to Cu͞Zn superoxide dismutase (SOD1) is mediated by a cytosolic protein termed the copper chaperone for superoxide dismutase (CCS). To determine the role of CCS in mammalian copper homeostasis, we generated mice with targeted disruption of CCS alleles (CCS ؊/؊ mice). Although CCS ؊/؊ mice are viable and possess normal levels of SOD1 protein, they reveal marked reductions in SOD1 activity when compared with control littermates. Metabolic labeling with 64 Cu demonstrated that the reduction of SOD1 activity in CCS ؊/؊ mice is the direct result of impaired Cu incorporation into SOD1 and that this effect was specific because no abnormalities were observed in Cu uptake, distribution, or incorporation into other cuproenzymes. Consistent with this loss of SOD1 activity, CCS ؊/؊ mice showed increased sensitivity to paraquat and reduced female fertility, phenotypes that are characteristic of SOD1-deficient mice. These results demonstrate the essential role of any mammalian copper chaperone and have important implications for the development of novel therapeutic strategies in familial amyotrophic lateral sclerosis.

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Multiple Protein Domains Contribute to the Action of the Copper Chaperone for Superoxide Dismutase

Cited by 174 publications

References 30 publications

Mechanisms for activating Cu- and Zn-containing superoxide dismutase in the absence of the CCS Cu chaperone

Mechanisms for activating Cu- and Zn-containing superoxide dismutase in the absence of the CCS Cu chaperone

Human superoxide dismutase 1 (hSOD1) maturation through interaction with human copper chaperone for SOD1 (hCCS)

Copper chaperone for superoxide dismutase is essential to activate mammalian Cu/Zn superoxide dismutase

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