Cloning and sequence analysis of two catechol-degrading gene clusters from a phenol-utilizing bacterium Pseudomonas putida SM25

Abstract

A 6.1 kb Sph I fragment from the genomic DNA of Pseudomonas putida SM 25 was cloned into the vector pUC19. The open reading frame of catB was found to consist of 1,122 nucleotides. The sequence alignment of the catB gene products from different kinds of bacteria revealed an overall identity ranging from 40 to 98%. The catC gene contained an open reading frame of 96 codons, from which a protein with a molecular mass of about 10.6 kDa was predicted. The amino acids in the proposed active-site region of CatC were found to be almost conserved, including the charged residues. Since the catBC genes in P putida SM25 were tightly linked, they could be regulated under coordinate transcription, and transcribed from a single promoter located upstream of the catB gene, as in P putida RB1.