title: NADPH-dependent reductive ortho dehalogenation of 2,4-dichlorobenzoic acid in Corynebacterium sepedonicum KZ-4 and Coryneform bacterium strainNTB-1 via 2,4-dichlorobenzoyl coenzyme A.
abstract: Corynebacterium sepedonicum KZ-4, described earlier as a strain capable of growth on 2,4-dichlorobenzoate (G.M. Zaitsev and Y.N. Karasevich, Mikrobiologiya 54:356-369, 1985), is known to metabolize this substrate via 4-hydroxybenzoate and protocatechuate, and evidence consistent with an initial reductive dechlorination step to form 4-chlorobenzoate was found in another coryneform bacterium, strain NTB-1 (W.J.J. van den Tweel, J.B. Kok, and J.A.M. de Bont, Appl. Environ. Microbiol. 53:810-815, 1987). 2-Chloro-4-fluorobenzoate was found to be converted stoichiometrically to 4-fluorobenzoate by resting cells of strain KZ-4, compatible with a reductive process. Experiments with cell extracts demonstrated that Mg - ATP and coenzyme A (CoA) were required to stimulate reductive dehalogenation, consistent with the intermediacy of 2-chloro-4-fluoro-benzoyl-CoA and 2,4-dichlorobenzoyl-CoA thioesters. 2,4-Dichlorobenzoyl-CoA was shown to be converted to 4-chlorobenzoyl-CoA in a novel NADPH-dependent reaction in extracts of both KZ-4 and NTB-1. In addition to the ligase and reductive dehalogenase activities, hydrolytic 4-chlorobenzoyl-CoA dehalogenase and thioesterase activities, 4-hydroxybenzoate 3-monooxygenase, and protocatechuate 3,4-dioxygenase activities were demonstrated to be present in the soluble fraction of KZ-4 extracts following ultracentrifugation. We propose that the pathway for 2,4-dichlorobenzoate catabolism in strains KZ-4 and NTB-1 involves formation of 2,4-dichlorobenzoyl-CoA, NADPH-dependent ortho dehalogenation yielding 4-chlorobenzoyl-CoA, hydrolytic removal of chlorine from the para position to generate 4-hydroxybenzoyl-CoA, hydrolysis to form 4-hydroxybenzoate, oxidation to yield protocatechuate, and oxidative ring cleavage.
authors.affiliation: Center for Microbial Ecology, Michigan State University, East Lansing 48824, USA.
authors.forename: V
authors.initials: V
authors.lastname: Romanov
authors.forename: R P
authors.initials: RP
authors.lastname: Hausinger
mesh_terms: D000421:Alcaligenes / Q000201:enzymology*; D001565:Benzoates / Q000378:metabolism; D001673:Biodegradation, Environmental; D002713:Chlorine / Q000378:metabolism; D002723:Chlorobenzoates / Q000378:metabolism*; D003065:Coenzyme A / Q000378:metabolism*; D003066:Coenzyme A Ligases / Q000378:metabolism; D003352:Corynebacterium / Q000201:enzymology*; D006867:Hydrolases / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010105:Oxygenases / Q000378:metabolism; D013869:Thiolester Hydrolases / Q000378:metabolism
chemical_list: D001565:Benzoates; D002723:Chlorobenzoates; C070735:2,4-dichlorobenzoic acid; D002713:Chlorine; D009249:NADP; D006899:Mixed Function Oxygenases; D010105:Oxygenases; D006867:Hydrolases; D013869:Thiolester Hydrolases; C084333:4-chlorobenzoyl coenzyme A dehalogenase; C077846:4-halobenzoate-CoA ligase; D003066:Coenzyme A Ligases; C040768:4-chlorobenzoic acid; D003065:Coenzyme A; C022893:4-fluorobenzoic acid