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Applied and Environmental Microbiology, July 2000, p. 2928-2933, Vol. 66, No. 7
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Influence of Chlorine Substituents on Rates of Oxidation of Chlorinated Biphenyls by the Biphenyl Dioxygenase of Burkholderia sp. Strain LB400

Clint M. Arnett,1,dagger Juan V. Parales,2 and John D. Haddock1,*

Department of Microbiology, Southern Illinois University, Carbondale, Illinois,1 and Department of Microbiology and the Center for Biocatalysis and Bioprocessing, The University of Iowa, Iowa City, Iowa2

Received 20 January 2000/Accepted 5 May 2000

Biphenyl dioxygenase from Burkholderia (Pseudomonas) sp. strain LB400 catalyzes the first reaction of a pathway for the degradation of biphenyl and a broad range of chlorinated biphenyls (CBs). The effect of chlorine substituents on catalysis was determined by measuring the specific activity of the enzyme with biphenyl and 18 congeners. The catalytic oxygenase component was purified and incubated with individual CBs in the presence of electron transport proteins and cofactors that were required for enzyme activity. The rate of depletion of biphenyl from the assay mixture and the rate of formation of cis-biphenyl 2,3-dihydrodiol, the oxidation product, were almost equal, indicating that the assay accurately measured enzyme-specific activity. Four classes of CBs were defined based on their oxidation rates. Class I contained 3-CB and 2,5-CB, which gave rates that were approximately twice that of biphenyl. Class II contained 2,5,3',4'-CB, 2,3,2',5'-CB, 2,3,4,5-CB, 2,3,2',3'-CB, 2,4,5,2',5'-CB, 2,5,3'-CB, 2,5,4'-CB, 2-CB, and 3,4,5-CB, which gave rates that ranged from 97 to 35% of the biphenyl rate. Class III contained only 2,3,4,2',5'-CB, which gave a rate that was 4% of the biphenyl rate. Class IV contained 2,4,4'-CB, 2,4,2',4'-CB, 3,4,5,2'-CB, 3,4,5,3'-CB, 3,5,3',5'-CB, and 3,4,5,2',5'-CB, which showed no detectable depletion. Rates were not significantly correlated with the aqueous solubilities of the CBs or the number of chlorine substituents on the rings. Oxidation products were detected for all class I, II, and III congeners and were identified as chlorinated cis-dihydrodiols for classes I and II. The specificity of biphenyl dioxygenase for the CBs examined in this study was determined by the relative positions of the chlorine substituents on the aromatic rings rather than the number of chlorine substituents on the rings.


* Corresponding author. Mailing address: Department of Microbiology, 131 LSII, Southern Illinois University, Carbondale, IL 62901-6508. Phone: (618) 453-3818. Fax: (618) 453-8036. E-mail: haddock{at}micro.siu.edu.

dagger Present address: U.S. Army Construction Engineering Research Laboratory, Champaign, IL 61826-9005.


Applied and Environmental Microbiology, July 2000, p. 2928-2933, Vol. 66, No. 7
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.



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