Open AccessGenetic and Biochemical Characterization of the Broad Spectrum Chlorobenzene Dioxygenase from Burkholderia Sp. Strain PS12 — Dechlorination of 1,2,4,5‐Tetrachlorobenzene
Author(s) -
Beil Stefan,
Happe Birgitta,
Timmis Kenneth Nigel,
Pieper Dietmar Helmut
Publication year - 1997
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1997.00190.x
Subject(s) - chlorobenzene , strain (injury) , broad spectrum , dioxygenase , characterization (materials science) , microbiology and biotechnology , reductive dechlorination , burkholderia , chemistry , biology , biodegradation , materials science , nanotechnology , genetics , combinatorial chemistry , organic chemistry , bacteria , enzyme , catalysis , anatomy
The bacterium, Burkholderia (previously Pseudomonas ) sp. strain PS12, reported earlier to degrade 1,2,4‐trichlorobenzene is shown here to utilize also 1,2,4,5‐tetrachlorobenzene (Cl 4 ‐benzene) as a growth substrate. To investigate the possibility that this organism attacks Cl 4 ‐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 α‐subunit and β‐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 18 O 2 incorporation experiments to dioxygenolytically attack a chlorosubstituted carbon atom of Cl 4 ‐benzene, thereby forming an unstable diol intermediate which spontaneously rearomatizes with concomitant chloride elimination to the corresponding 3,4,6‐trichlorocatechol (Cl 3 ‐catechol).