Isolation and characterization of an alcohol dehydrogenase gene from the octylphenol polyethoxylate degrader Pseudomonas putida S-5.
Octylphenol polyethoxylate (OPEO(n)) biodegradation by Pseudomonas putida S-5 under aerobic conditions is initiated by the oxidation of its terminal alcohol group by alcohol dehydrogenase. A DNA fragment, containing an alcohol dehydrogenase gene (adh1), was isolated using a combination of degenerate PCR and inverse PCR. The predicted translation product of adh1 showed significant sequence similarity to bacterial alcohol dehydrogenases. Furthermore, a flavin-binding motif and signature patterns conserved in type III FAD-dependent alcohol oxidases were detected. Two open reading frames (ORFs) were found upstream of adh1, encoding a putative acyl-CoA synthetase and a putative esterase. Downstream of adh1 and located on the opposite strand was an ORF encoding a putative aldehyde dehydrogenase. Transcription analysis using RT-PCR showed that adh1 is cotranscribed with the putative acyl-CoA synthetase and esterase genes during growth on OPEO(n). ADH1 overproduced in Escherichia coli exhibited activity not only toward various alcohols, including OPEO(n)s, but also toward primary aliphatic and aromatic aldehydes.
Tasaki
Yuji
Y
Department of Materials Engineering, Nagaoka National College of Technology, Niigata, Japan. ytasaki@nagaoka-ct.ac.jp
Yoshikawa
Hiromichi
H
Tamura
Hiroto
H
D000426:Alcohol Dehydrogenase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D017830:Octoxynol / Q000737:chemistry / Q000378:metabolism*; D016958:Pseudomonas putida / Q000201:enzymology* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity
D017830:Octoxynol; D000426:Alcohol Dehydrogenase