Nucleotide sequence and expression of gene nahH of plasmid NAH7 and homology with gene xylE of TOL pWWO

Abstract

The enzyme catechol 2,3-dioxygenase (C23O) encoded by the nahH gene of plasmid NAH7 converts catechol to alpha-hydroxymuconic epsilon-semialdehyde in Pseudomonas putida. We have cloned this structural gene into vectors pUC18 and pKT240, determined the nucleotide sequence and deduced the amino acid sequence. In comparison to the gene xylE of the TOL plasmid pWWO which encodes a similar C230 enzyme [Nakai et al. J. Biol. Chem. (1983b), 2923-2928], the respective G + C contents were 55% and 57%, the nucleotide sequences 81% homologous, and the amino acid homology 85%. The flanking sequences of the two C23O-coding genes also show homology. Clones of the nahH gene in Escherichia coli overproduce the protein product at least ten fold and the gene product was identified by polyacrylamide gel electrophoresis.