title: Cloning, functional expression and characterization of a bifunctional 3-hydroxybutanal dehydrogenase /reductase involved in acetone metabolism by Desulfococcus biacutus.
abstract: 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 CO2to 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.
  
  
  
  The 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-1protein), 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).
  
  
  
  The 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.
authors.affiliation: Department of Biology, University of Konstanz, Postbox 649, D-78457, Konstanz, Germany.
authors.forename: Jasmin
authors.initials: J
authors.lastname: Frey
authors.affiliation: Department of Biology, University of Konstanz, Postbox 649, D-78457, Konstanz, Germany.
authors.forename: Hendrik
authors.initials: H
authors.lastname: Rusche
authors.affiliation: Department of Biology, University of Konstanz, Postbox 649, D-78457, Konstanz, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Schink
authors.affiliation: Department of Biology, University of Konstanz, Postbox 649, D-78457, Konstanz, Germany. david.schleheck@uni-konstanz.de.
authors.forename: David
authors.initials: D
authors.lastname: Schleheck
mesh_terms: D000079:Acetaldehyde / Q000031:analogs & derivatives / Q000378:metabolism; D000096:Acetone / Q000737:chemistry / Q000378:metabolism*; D000426:Alcohol Dehydrogenase / Q000378:metabolism; D000447:Aldehydes / Q000737:chemistry / Q000378:metabolism*; D001421:Bacteria, Anaerobic / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D000440:Butanols / Q000378:metabolism; D002072:Butylene Glycols / Q000737:chemistry; D002248:Carbon Monoxide / Q000378:metabolism; D019976:Cloning, Organism; D003067:Coenzymes / Q000378:metabolism; D020564:Deltaproteobacteria / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D004789:Enzyme Activation; D057075:Enzyme Assays; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D053858:Metabolic Networks and Pathways / Q000235:genetics; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010088:Oxidoreductases / Q000235:genetics* / Q000378:metabolism*; D020005:Propanols / Q000378:metabolism; D040901:Proteomics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013696:Temperature; D015032:Zinc / Q000378:metabolism
chemical_list: D000447:Aldehydes; D001426:Bacterial Proteins; D000440:Butanols; D002072:Butylene Glycols; D003067:Coenzymes; D020005:Propanols; D011994:Recombinant Proteins; D009243:NAD; D000096:Acetone; C028491:1,3-butylene glycol; D009249:NADP; C057552:acetoacetaldehyde; D002248:Carbon Monoxide; C116609:3-hydroxybutanal; C005556:propionaldehyde; D010088:Oxidoreductases; D000426:Alcohol Dehydrogenase; D000079:Acetaldehyde; C018475:butyraldehyde; D015032:Zinc
keywords: Acetone activation; Bifunctional MDR superfamily oxidoreductase; Carbonylation; Sulfate-reducing bacteria