Two strains of Alcaligenes denitrificans, designated BRI 3010 and BRI 6011, were isolated from polychlorinated biphenyl (PCB) contaminated soil using 2,5-dichlorobenzoic acid (2,5-DCBA) and 2,4-DCBA, respectively, as sole carbon and energy sources. Both strains degraded 2-chlorobenzoic acid (2-CBA), 2,3-DCBA, and 2,5-DCBA. BRI 6011 alone degraded 2,4-DCBA. Metabolism of the chlorinated substrates resulted in the stoichiometric release of chloride, and degradation proceeded by intradiol cleavage of the aromatic ring. Growth of both strains on dichlorobenzoic acids induced pyrocatechase activities having catechol (catechol 1,2-dioxygenase) and chlorocatechols (chlorocatechol 1,2-dioxygenase) as substrates. Growth on 2-CBA and benzoic acid induced a pyrocatechase activity (catechol 1,2-dioxygenase) directed against catechol only. / The chlorocatechol 1,2-dioxygenase from BRI 6011 was purified, characterized, and compared with the chlorocatechol 1,2-dioxygenase from Pseudomonas sp. B13 and P. putida, organisms limited with respect to their CBA degradative versatility. These enzymes appear to be very similar based on biochemical and genetic data and possess sufficient broad substrate specificity to accommodate a wide range of chlorinated catechols, hence the increased versatility for chlorobenzoic acid degradation of A. denitrificans cannot be attributed to a more specialized chlorocatechol 1,2-dioxygenase. / Uptake of benzoic acid by BRI 3010 and BRI 6011 was inducible, exhibited saturation kinetics and the substrate was accumulated intracellularly against a concentration gradient by a factor of 8 and 10, respectively, indicative of active transport. Uptake of 2,4-DCBA by BRI 6011 was constitutive and saturation kinetics were not observed, suggesting passive diffusion of 2,4-DCBA and other CBAs into the cell down a concentration gradient. / Based on oxygen uptake experiments with whole cells, benzoic acid dioxygenase and chlorobenzoic acid dioxygenase activity was induced by benzoic acid and ortho-substituted chlorobenzoic acids, respectively. Since 2,4-DCBA diffuses across the membrane and the expected catecholic intermediates of 2,4-DCBA metabolism are metabolizable by BRI 3010, this suggests that the major difference between BRI 3010 and BRI 6011 might be the inability of the chlorobenzoic acid dioxygenase in BRI 3010 to recognize 2,4-DCBA as a substrate.

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