Evidence for a Proton Transfer Network and a Required Persulfide-Bond-Forming Cysteine Residue in Ni-Containing Carbon Monoxide Dehydrogenases

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OAK-B135 Carbon monoxide dehydrogenase from Moorella thermoacetica catalyzes the reversible oxidation of CO to CO2 at a nickel-iron-sulfur active-site called the C-cluster. Mutants of a proposed proton transfer pathway and of a cysteine residue recently found to form a persulfide bond with the C-cluster were characterized. Four semi-conserved histidine residues were individually mutated to alanine. His116 and His122 were essential to catalysis, while His113 and His119 attenuated catalysis but were not essential. Significant activity was ''rescued'' by a double mutant where His116 was replaced by Ala and His was also introduced at position 115. Activity was also rescued in double ... continued below

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Kim, Eun Jin; Feng, Jian; Bramlett, Matthew R. & Lindahl, Paul A. May 18, 2004.

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  • Texas A & M University
    Publisher Info: Texas A&M University, College Station, TX (United States)
    Place of Publication: College Station, Texas

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OAK-B135 Carbon monoxide dehydrogenase from Moorella thermoacetica catalyzes the reversible oxidation of CO to CO2 at a nickel-iron-sulfur active-site called the C-cluster. Mutants of a proposed proton transfer pathway and of a cysteine residue recently found to form a persulfide bond with the C-cluster were characterized. Four semi-conserved histidine residues were individually mutated to alanine. His116 and His122 were essential to catalysis, while His113 and His119 attenuated catalysis but were not essential. Significant activity was ''rescued'' by a double mutant where His116 was replaced by Ala and His was also introduced at position 115. Activity was also rescued in double mutants where His122 was replaced by Ala and His was simultaneously introduced at either position 121 or 123. Activity was also ''rescued'' by replacing His with Cys at position 116. Mutation of conserved Lys587 near the C-cluster attenuated activity but did not eliminate it. Activity was virtually abolished in a double mutant where Lys587 and His113 were both changed to Ala. Mutations of conserved Asn284 also attenuated activity. These effects suggest the presence of a network of amino acid residues responsible for proton transfer rather than a single linear pathway. The Ser mutant of the persulfide-forming Cys316 was essentially inactive and displayed no EPR signals originating from the C-cluster. Electronic absorption and metal analysis suggests that the C-cluster is absent in this mutant. The persulfide bond appears to be essential for either the assembly or stability of the C-cluster, and/or for eliciting the redox chemistry of the C-cluster required for catalytic activity.

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INIS; OSTI as DE00826210

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  • Journal Name: Biochemistry; Journal Volume: 43; Journal Issue: 19; Other Information: Published in Biochemistry, Volume 43, No.19; DOI 10.1021/bi036062u

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  • Report No.: DOE/ER/15177-5
  • Grant Number: FG03-01ER15177
  • DOI: 10.1021/bi036062u | External Link
  • Office of Scientific & Technical Information Report Number: 826210
  • Archival Resource Key: ark:/67531/metadc782601

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  • May 18, 2004

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  • Dec. 3, 2015, 9:30 a.m.

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  • May 6, 2016, 2:14 p.m.

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Kim, Eun Jin; Feng, Jian; Bramlett, Matthew R. & Lindahl, Paul A. Evidence for a Proton Transfer Network and a Required Persulfide-Bond-Forming Cysteine Residue in Ni-Containing Carbon Monoxide Dehydrogenases, article, May 18, 2004; College Station, Texas. (digital.library.unt.edu/ark:/67531/metadc782601/: accessed August 22, 2017), University of North Texas Libraries, Digital Library, digital.library.unt.edu; crediting UNT Libraries Government Documents Department.