Coupling of the pathway of sulphur oxidation to dioxygen reduction: characterization of a novel membrane-bound thiosulphate:quinone oxidoreductase

Authors

  • Fabian H. Müller,

    1. Institute of Microbiology and Genetics, Darmstadt University of Technology, Schnittspahnstraße 10, D-64287 Darmstadt, Germany.
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  • Tiago M. Bandeiras,

    1. Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Apt 127, Av. da República (EAN), 2781-901 Oeiras, Portugal.
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  • Tim Urich,

    1. Institute of Microbiology and Genetics, Darmstadt University of Technology, Schnittspahnstraße 10, D-64287 Darmstadt, Germany.
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  • Miguel Teixeira,

    1. Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Apt 127, Av. da República (EAN), 2781-901 Oeiras, Portugal.
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  • Cláudio M. Gomes,

    1. Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Apt 127, Av. da República (EAN), 2781-901 Oeiras, Portugal.
    2. Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, 2825–114 Caparica, Portugal.
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  • Arnulf Kletzin

    Corresponding author
    1. Institute of Microbiology and Genetics, Darmstadt University of Technology, Schnittspahnstraße 10, D-64287 Darmstadt, Germany.
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E-mail Kletzin@bio.tu-darmstadt.de; Tel. (+49) 6151 16-5242; Fax (+49) 6151 16-2956.

Summary

Thiosulphate is one of the products of the initial step of the elemental sulphur oxidation pathway in the thermoacidophilic archaeon Acidianus ambivalens. A novel thiosulphate:quinone oxidoreductase (TQO) activity was found in the membrane extracts of aerobically grown cells of this organism. The enzyme was purified 21-fold from the solubilized membrane fraction. The TQO oxidized thiosulphate with tetrathionate as product and ferricyanide or decyl ubiquinone (DQ) as electron acceptors. The maximum specific activity with ferricyanide was 73.4 U (mg protein)−1 at 92°C and pH 6, with DQ it was 397 mU (mg protein)−1 at 80°C. The Km values were 2.6 mM for thiosulphate (kcat = 167 s−1),  3.4 mM  for  ferricyanide  and  5.87 µM for DQ. The enzymic activity was inhibited by sulphite (Ki = 5 µM), metabisulphite, dithionite and TritonX-100, but not by sulphate or tetrathionate. A mixture of caldariella quinone, sulfolobus quinone and menaquinone was non-covalently bound to the protein. No other cofactors were detected. Oxygen consumption was measured in membrane fractions upon thiosulphate addition, thus linking thiosulphate oxidation to dioxygen reduction, in what constitutes a novel activity among Archaea. The holoenzyme was composed of two subunits of apparent molecular masses of 28 and 16 kDa. The larger subunit appeared to be glycosylated and was identical to DoxA, and the smaller was identical to DoxD. Both subunits had been described previously as a part of the terminal quinol:oxygen oxidoreductase complex (cytochrome aa3).

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