Abstract
BackgroundThe strictly anaerobic, sulfate-reducing bacterium Desulfococcus biacutus can utilize acetone as sole carbon and energy source for growth. Whereas in aerobic and nitrate-reducing bacteria acetone is activated by carboxylation with CO2 to acetoacetate, D. biacutus involves CO as a cosubstrate for acetone activation through a different, so far unknown pathway. Proteomic studies indicated that, among others, a predicted medium-chain dehydrogenase/reductase (MDR) superfamily, zinc-dependent alcohol dehydrogenase (locus tag DebiaDRAFT_04514) is specifically and highly produced during growth with acetone.ResultsThe MDR gene DebiaDRAFT_04514 was cloned and overexpressed in E. coli. The purified recombinant protein required zinc as cofactor, and accepted NADH/NAD+ but not NADPH/NADP+ as electron donor/acceptor. The pH optimum was at pH 8, and the temperature optimum at 45 °C. Highest specific activities were observed for reduction of C3 - C5-aldehydes with NADH, such as propanal to propanol (380 ± 15 mU mg−1 protein), butanal to butanol (300 ± 24 mU mg−1), and 3-hydroxybutanal to 1,3-butanediol (248 ± 60 mU mg−1), however, the enzyme also oxidized 3-hydroxybutanal with NAD+ to acetoacetaldehyde (83 ± 18 mU mg−1).ConclusionThe enzyme might play a key role in acetone degradation by D. biacutus, for example as a bifunctional 3-hydroxybutanal dehydrogenase/reductase. Its recombinant production may represent an important step in the elucidation of the complete degradation pathway.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-016-0899-9) contains supplementary material, which is available to authorized users.
Highlights
The strictly anaerobic, sulfate-reducing bacterium Desulfococcus biacutus can utilize acetone as sole carbon and energy source for growth
One of the most prominent acetone-inducible proteins observed is encoded by gene (IMG locus tag) DebiaDRAFT_04514, and is annotated as medium-chain dehydrogenase/reductase (MDR) superfamily alcohol dehydrogenase (COG1063 in the Clusters of Orthologous Groups classification system)
Biochemicals (NADH, NADPH, NAD+ and NADP+) were purchased from Sigma-Aldrich (Germany). 3-Hydroxybutanal was synthesized by Dr Thomas Huhn and Fabian Schneider, Chemistry Department of University of Konstanz
Summary
The strictly anaerobic, sulfate-reducing bacterium Desulfococcus biacutus can utilize acetone as sole carbon and energy source for growth. Whereas in aerobic and nitrate-reducing bacteria acetone is activated by carboxylation with CO2 to acetoacetate, D. biacutus involves CO as a cosubstrate for acetone activation through a different, so far unknown pathway. ATP and CO as cosubstrates [8] This reaction was not observed in cell-free extract of butyrate-grown cells, the proposed acetone-activating enzyme, and most likely the entire acetone utilization pathway, is inducibly expressed in D. biacutus. One of the most prominent acetone-inducible proteins observed is encoded by gene (IMG locus tag) DebiaDRAFT_04514, and is annotated as medium-chain dehydrogenase/reductase (MDR) superfamily alcohol dehydrogenase (COG1063 in the Clusters of Orthologous Groups classification system). Other strongly induced proteins are a predicted thiamine diphosphate (TDP)-requiring enzyme (COG0028), and a cobalamin (B12)-binding subunit (COG2185) of a methylmalonyl-CoA mutase-like complex [9]
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