Purification and properties of benzyl alcohol dehydrogenase from a denitrifying Thauera sp.
Toluene and related aromatic compounds are anaerobically degraded by the denitrifying bacterium Thauera sp. strain K172 via oxidation to benzoyl-CoA. The postulated initial step is methylhydroxylation of toluene to benzyl alcohol, which is either a free or enzyme-bound intermediate. Cells grown with toluene or benzyl alcohol contained benzyl alcohol dehydrogenase, which is possibly the second enzyme in the proposed pathway. The enzyme was purified from benzyl-alcohol-grown cells and characterized. It has many properties in common with benzyl alcohol dehydrogenase from Acinetobacter and Pseudomonas species. The enzyme was active as a homotetramer of 160 kDa, with subunits of 40 kDa. It was NAD(+)-specific, had an alkaline pH optimum, and was inhibited by thiol-blocking agents. No evidence for a bound cofactor was obtained. Various benzyl alcohol analogues served as substrates, whereas non-aromatic alcohols were not oxidized. The N-terminal amino acid sequence indicates that the enzyme belongs to the class of long-chain Zn(2+)-dependent alcohol dehydrogenases, although it appears not to contain a metal ion that can be removed by complexing agents.
Biegert
T
T
Universität Ulm, Germany.
Altenschmidt
U
U
Eckerskorn
C
C
Fuchs
G
G
D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D000595:Amino Acid Sequence; D019905:Benzyl Alcohol; D001592:Benzyl Alcohols / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D002729:Chloromercuribenzoates / Q000494:pharmacology; D006089:Gram-Negative Anaerobic Bacteria / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009579:Nitrobenzoates / Q000494:pharmacology; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D001592:Benzyl Alcohols; D002614:Chelating Agents; D002729:Chloromercuribenzoates; D009579:Nitrobenzoates; D000429:Alcohol Oxidoreductases; C027016:benzyl alcohol dehydrogenase; D019905:Benzyl Alcohol