Genetic and biochemical analyses of the tec operon suggest a route for evolution of chlorobenzene degradation genes

The TecA broad-spectrum chlorobenzene dioxygenase of Burkholderia sp. strai n PS12 catalyzes the first step in the mineralization of 1,2,4,5-tetrachlor obenzene. The catabolic genes were localized on a small plasmid that belong s to the IncP beta incompatibility group. PCR analysis of the genetic envir onment of the fec genes indicated high similarity to the transposon-organiz ed catabolic tcb chlorobenzene degradation genes of Pseudomonas sp. strain P51. Sequence analysis of the regions flanking the tecA genes revealed an u pstream open reading frame (ORF) with high similarity to the todF 2-hydroxy -6-oxo-2,4-heptadienoate hydrolase gene of Pseudomonas putida Fl and a disc ontinuous downstream ORF showing high similarity to the todE catechol 2,3-d ioxygenase gene of strain F1. Both homologues in strain P51 exist only as d eletion remnants. We suggest that different genetic events thus led to inac tivation of the perturbing meta-cleavage enzymes in strains P51 and PS12 du ring the evolution of efficient chlorobenzene degradation pathways. Biochem ical characterization of TodF-like protein TlpF and a genetically refunctio nalized TodE-like protein, TlpE, produced in Escherichia coli provided data consistent with the proposed relationships.