title: Mitochondrial Ccs1 contains a structural disulfide bond crucial for the import of this unconventional substrate by the disulfide relay system.
abstract: The copper chaperone for superoxide dismutase 1 (Ccs1) provides an important cellular function against oxidative stress. Ccs1 is present in the cytosol and in the intermembrane space (IMS) of mitochondria. Its import into the IMS depends on the Mia40/Erv1 disulfide relay system, although Ccs1 is, in contrast to typical substrates, a multidomain protein and lacks twin Cx(n)C motifs. We report on the molecular mechanism of the mitochondrial import of Saccharomyces cerevisiae Ccs1 as the first member of a novel class of unconventional substrates of the disulfide relay system. We show that the mitochondrial form of Ccs1 contains a stable disulfide bond between cysteine residues C27 and C64. In the absence of these cysteines, the levels of Ccs1 and Sod1 in mitochondria are strongly reduced. Furthermore, C64 of Ccs1 is required for formation of a Ccs1 disulfide intermediate with Mia40. We conclude that the Mia40/Erv1 disulfide relay system introduces a structural disulfide bond in Ccs1 between the cysteine residues C27 and C64, thereby promoting mitochondrial import of this unconventional substrate. Thus the disulfide relay system is able to form, in addition to double disulfide bonds in twin Cx(n)C motifs, single structural disulfide bonds in complex protein domains.
authors.affiliation: Adolf-Butenandt-Institut, Lehrstuhl für Physiologische Chemie, Ludwig-Maximilians-Universität München, 81377 München, Germany.
authors.forename: Dominik P
authors.initials: DP
authors.lastname: Gross
authors.forename: Caroline A
authors.initials: CA
authors.lastname: Burgard
authors.forename: Silvia
authors.initials: S
authors.lastname: Reddehase
authors.forename: Jeffry M
authors.initials: JM
authors.lastname: Leitch
authors.forename: Valeria C
authors.initials: VC
authors.lastname: Culotta
authors.forename: Kai
authors.initials: K
authors.lastname: Hell
mesh_terms: D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000378:metabolism*; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010084:Oxidation-Reduction; D050862:Oxidoreductases Acting on Sulfur Group Donors / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D017510:Protein Folding; D054730:Protein Interaction Domains and Motifs; D021381:Protein Transport / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics*; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism; D000072105:Superoxide Dismutase-1; D014161:Transduction, Genetic
chemical_list: C491960:CCS1 protein, S cerevisiae; D004220:Disulfides; C495320:MIA40 protein, S cerevisiae; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins; D018832:Molecular Chaperones; D029701:Saccharomyces cerevisiae Proteins; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D050862:Oxidoreductases Acting on Sulfur Group Donors; C073790:ERV1 protein, S cerevisiae; D003545:Cysteine