Catechol 2,3-Dioxygenase from Pseudomonassp. Strain ND6: Gene Sequence and Enzyme Characterization

The catechol 2,3-dioxygenase (C23O) gene in naphthalene catabolic plasmid pND6-1 of Pseudomonassp. ND6 was cloned and sequenced. The C23O gene was consisted of 924 nucleotides and encoded a polypeptide of molecular weight 36 kDa containing 307 amino acid residues. The C23O of Pseudomonassp. ND6 exhibited 93% and 89% identities in amino acid sequence with C23Os encoded by naphthalene catabolic plasmid NAH7 from Pseudomonas putidaG7 and the chromosome of Pseudomonas stutzeriAN10 respectively. The Pseudomonassp. ND6 C23O gene was overexpressed in Escherichia coliDH 5α using the lacpromoter of pUC18, and its gene product was purified by DEAE-Sephacel and Phenyl-Sepharose CL-4B chromatography. The enzymology experiments indicated that the specific activity and thermostability of C23O from Pseudomonassp. ND6 were better than those of C23O from Pseudomonas putidaG7.