title: Assignment of the CO-sensitive carboxyl group in mitochondrial forms of cytochrome c oxidase using yeast mutants.
abstract: Point mutations of E243D and I67N were introduced into subunit I of a 6histidine-tagged (6H-WT) form of yeast Saccharomyces cerevisiae mitochondrial cytochrome c oxidase. The two mutants (6H-E243D(I) and 6H-I67N(I)) were purified and showed ≈50 and 10% of the 6H-WT turnover number. Light-induced CO photolysis FTIR difference spectra of the 6H-WT showed a peak/trough at 1749/1740cm(-1), as seen in bovine CcO, which downshifted by 7cm(-1) in D(2)O. The bands shifted to 1736/1762cm(-1) in 6H-E243D(I), establishing that the carboxyl group affected by CO binding in mitochondrial CcOs is E243. In 6H-I67N(I), the trough at 1740cm(-1) was shifted to 1743cm(-1) and its accompanying peak intensity was greatly reduced. This confirms that the I67N mutation interferes with conformational alterations around E243. This article is part of a Special Issue entitled: 17th European Bioenergetics Conference (EBEC 2012).
authors.affiliation: Institute of Structural and Molecular Biology, University College, London, UK.
authors.forename: Amandine
authors.initials: A
authors.lastname: Maréchal
authors.forename: Brigitte
authors.initials: B
authors.lastname: Meunier
authors.forename: Peter R
authors.initials: PR
authors.lastname: Rich
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002248:Carbon Monoxide / Q000737:chemistry / Q000378:metabolism; D002417:Cattle; D003576:Electron Transport Complex IV / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D010782:Photolysis; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D024101:Mitochondrial Proteins; D029701:Saccharomyces cerevisiae Proteins; D002248:Carbon Monoxide; D003576:Electron Transport Complex IV