[Cloning and sequence analysis of 1,2,4-trichlorobenzene dioxygenase and dehydrogenase genes]

Pseudomonas nitroreducens J5-1 is able to use monochlorobenzene, 1,2-dichlorobenzene, 1,3-dichlorobenzene and 1,2,4-trichlorobenzene as sole carbon and energy sources, and it differs from those 1,2,4-trichlorobenzene degrading bacteria reported in substrate utilizing characters. PCR technique was used to amplify the genes of chlorobenzene dioxygenase and dehydrogenase of J5-1, and they were named as tcbA and tcbB, respectively. Homology analysis indicated that these genes and gene products were most closely related to those of Burkholderia sp. PS12. By alignment of the amino acid sequences of the a subunits of TcbAa (from J5-1) and TecA1 (from PS12), four amino acid residues from site 307 to site 310 were found to be different (I307L, M308T, I309V, Q310E), which probably retarded the preference for the substrate 1,2,4,5-tetrachlorobenzene. Furthermore, the phylogenetic analysis of the dioxygenase alpha subunits showed that TcbAa was belong to the toluene/diphenyl subfamily, and was most closely related to the poly-chlorinated benzene dioxygenase alpha subunit.