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Vinyl ketone reduction by three distinct Gluconobacter oxydans 621H enzymes

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Abstract

Three cytosolic NADPH-dependent flavin-associated proteins (Gox2107, Gox0502, and Gox2684) from Gluconobacter oxydans 621H were overproduced in Escherichia coli, and the recombinant enzymes were purified and characterized. Apparent native molecular masses of 65.2, 78.2, and 78.4 kDa were observed for Gox2107, Gox0502, and Gox2684, corresponding to a trimeric structure for Gox2107 and dimers for Gox0502 and Gox2684. Analysis of flavin content revealed Gox2107 was flavin adenine dinucleotide dependent, whereas Gox0502 and Gox2684 contained flavin mononucleotide. The enzymes were able to reduce vinyl ketones and quinones, reducing the olefinic bond of vinyl ketones as shown by 1H nuclear magnetic resonance. Additionally, Gox0502 and Gox2684 stereospecifically reduced 5S-(+)-carvone to 2R,5S-dihydrocarvone. All enzymes displayed highest activities with 3-butene-2-one and 1,4-naphthoquinone. Gox0502 and Gox2684 displayed a broader substrate spectrum also reducing short-chain α-diketones, whereas Gox2107 was most catalytically efficient.

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Acknowledgments

This project was supported by funds from Bundesministerium für Bildung und Forschung (BMBF) and University of Wisconsin—Milwaukee.

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Authors and Affiliations

  1. Department of Biological Sciences, University of Wisconsin—Milwaukee, 3209 N. Maryland Ave, Milwaukee, WI, 53211, USA

    Paul Schweiger & Shane Wesener

  2. Institut für Pharmazeutische Biologie, Nußallee 6, 53115, Bonn, Germany

    Harald Gross

  3. Institute für Mikrobiologie und Biotechnologie, 168 Meckenheimer Allee, 53115, Bonn, Germany

    Uwe Deppenmeier

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  1. Paul Schweiger
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Correspondence to Uwe Deppenmeier.

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Schweiger, P., Gross, H., Wesener, S. et al. Vinyl ketone reduction by three distinct Gluconobacter oxydans 621H enzymes. Appl Microbiol Biotechnol 80, 995–1006 (2008). https://doi.org/10.1007/s00253-008-1600-5

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  • DOI: https://doi.org/10.1007/s00253-008-1600-5

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