FEMS Microbiology Ecology 85 (1991) 293-300 © 1991 Federation of European Microbiological Societies 0168-6496/91/$03.50 Published by Elsevier ADONIS 016864969100082F 293 FEMSEC 00337 Evidence for inhibitory substrate interactions during cometabolism of 3,4-dichlorobenzoate by Acinetobacter sp. strain 4-CB1 P. Adriaens * and D.D. Focht Department of Soil and Environmental Sciences, University of California, Riverside, CA, U.S.A. Received 18 November 1990 Revision received 18 February 1991 Accepted 28 February 1991 Key words: Degradation pathway; Incompatibility; Inhibition kinetics; Pseudosubstrate; Resting cell 1. SUMMARY Acinetobacter sp. strain 4-CB1 cometabolized 3,4-dichlorobenzoate (3,4-DCB), via 3-chloro-4- hydroxybenzoate (3-C-4-OHB) and 4-carboxy- 1,2-benzoquinone, in the presence of 4-chloroben- zoate (4-CB) as a growth substrate. In resting cell incubations, 3,4-DCB acted competitively as an inhibitor with 4-CB metabolism, and as a sub- strate inhibitor of its own metabolism. The inhibi- tor constant K i and the affinity constant K,a were 800 and 181 #M, respectively with a maximal rate of 3,4-DCB disappearance of 18.8 nmol 3,4- DCB min -1 (mg protein) -1, in resting cells in- cubated solely with 3,4-DCB. Resting cells were less tolerant to 3,4-DCB than growing cells, as noted from the inhibition constants (Ki). More- Correspondence to: D.D. Foeht, Department of Soil and En- vironmental Sciences, University of California, Riverside, CA 92521, U.S.A. * Present address: Department of Civil Engineering, Stanford University, Stanford CA 94305, U.S.A. over, 3-C-4-OHB competitively inhibited 4-hy- droxybenzoate monooxygenase by acting as a pseudosubstrate (g i = 7.3 /tM). The next sequen- tial intermediate, 4-carboxy-l,2-benzoquinone, un- competitively inhibited 4-CB metabolism in rest- ing cell incubations. Thus, 3,4-DCB inhibited its own cometabolism as well as metabolism of 4-CB in Acinetobacter sp. strain 4-CB1. 2. INTRODUCTION Many chlorinated aromatic compounds have been demonstrated to be growth substrates for soil and water microorganisms, or are cometabolized to an array of chlorinated intermediates [1]. The biochemical basis of cometabolism has been ex- plained by the failure of the compound to induce the appropriate enzymes [2-4], by the inability of the compound to diffuse accross the cell mem- brane [1,3], or by the accumulation of toxic inter- mediates and inhibition of the enzymes [5-8]. Gibson et al. [9] suggested that the catechol di- Downloaded from https://academic.oup.com/femsle/article-abstract/85/4/293/487729 by guest on 17 June 2020