Genetic and Biochemical Characterization of the Broad Spectrum Chlorobenzene Dioxygenase from Burkholderia Sp. Strain PS12 — Dechlorination of 1,2,4,5‐Tetrachlorobenzene

Abstract

The bacterium, Burkholderia (previously Pseudomonus) sp. strain PS12, reported earlier to degrade 1,2,4-trichlorobenzene is shown here to utilize also 1,2,4,S-tetrachlorobenzene (CL-benzene) as a growth substrate. To investigate the possibility that this organism attacks C1,-benzene with a chlorobenzene dioxygenase which concomitantly causes dehalogenation, and to analyze the substrate range of the initial enzyme, a 5503-bp DNA fragment from PS12, exhibiting high similarity to genes coding for class IIB dioxygenases, was cloned and expressed in Escherichia coli. The sequence includes the tec genes coding for the cr-subunit and /I-subunit of a terminal dioxygenase, a ferredoxin and a reductase. E. coli cells producing these proteins were able to dioxygenolytically attack a range of aromatic compounds including chlorinated benzenes and toluene, and also dinuclear aromatics such as biphenyl and dibenzo-p-dioxin. The enzyme was shown by "O0, incorporation experiments to dioxygenolytically attack a chlorosubstituted carbon atom of C1,-benzene, thereby forming an unstable diol intermediate which spontaneously rearomatizes with concomitant chloride elimination to the corresponding 3,4,6-trichlorocatechol (CLcatechol).

Degradation of chloroaromatics by Pseudomonas putida GJ31: assembled route for chlorobenzene degradation encoded by clusters on plasmid pKW1 and the chromosome

“…PS12 (Beil et al, 1997) and Acidovorax sp. P51 (van der Meer et al, 1991), Plasmid encodes 3-chlorocatechol meta-cleavage pathway and nearly identical in sequence (5478/5494 nt) to the todC1C2BADE cluster of P. putida strain F1 growing with toluene via a meta-cleavage pathway (Mosqueda et al, 1999;Zylstra & Gibson, 1989), and the high level of transcription in response to chlorobenzene validated its involvement in chlorobenzene degradation by strain GJ31.…”

Emergence of Multifunctional Oxygenase Activities by Random Priming Recombination

“…It was also reported that chlorobenzene dioxygenase of Burkholderia sp. strain PS12, which utilizes 1,2,4,5-tetrachlorobenzene, attacks dibenzo-p-dioxin preferentially by lateral dioxygenation (32). The same enzyme converts dibenzofuran into cis-1,2-dihydroxy-1,2-dihydrobenzofuran as a single metabolite.…”

“…strain NCIB9816-4, and 1,2,4,5-tetrachlorobenzen-utilizing Burkhorderia sp. strain PS12 also have broad substrate ranges (29,30,32,33,34).…”

Biodegradation of 2,4-dichlorophenol using Mycoplana dimorpha extracts and evaluation of kinetic parameters

“…J. Biotechnol. indicates that immobilized crude extract degradation is much faster than microbial degradation (Axcell and Geary 1973;Grund et al, 1975;Hartmann et al, 1979;Chang et al, 1992;Spiess and Gorisch 1996;Beil et al, 1997;Mars et al, 1997;Beil et al, 1998;Pollmann et al, 2001;Alfreider et al, 2003;Rapp and Gabriel-Jurgens 2003;Manikandan et al, 2007a;Manikandan et al, 2007b). The similar trend of decomposition was observed with the initial concentrations of 60 and 80 ppm with the percentage degradation at the end of 30 min is 72% for 60 ppm and 75% for 80 ppm (Manikandan et al, 2007d).…”