Purification and characterization of 2,6-dichloro-p-hydroquinone chlorohydrolase from Flavobacterium sp. strain ATCC 39723.
AUTOR(ES)
Lee, J Y
RESUMO
The biochemistry of pentachlorophenol (PCP) degradation by Flavobacterium sp. strain ATCC 39723 has been studied, and two enzymes responsible for the conversion of PCP to 2,6-dichloro-p-hydroquinone (2,6-DiCH) have previously been purified and characterized. In this study, enzymatic activities consuming 2,6-DiCH were identified from the cell extracts of strain ATCC 39723. The enzyme was purified to apparent homogeneity by a purification scheme consisting of seven steps. Gel filtration chromatography showed a native molecular weight of about 40,000, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single protein of 42,500 Da. The purified enzyme converted 2,6-DiCH to 6-chlorohydroxyquinol (6-chloro-1,2,4-trihydroxybenzene), which was easily oxidized by molecular oxygen and hard to detect. The end product, 6-chlorohydroxyquinol, was detected only in the presence of a reductase and NADH in the reaction mixture. The enzyme dechlorinated 2,6-DiCH but not 2,5-DiCH. The enzyme required Fe2+ for activity and was severely inhibited by metal chelating agents. The optimal conditions for activity were pH 7.0 and 40 degrees C. The Kcat for 2,6-DiCH was 35 microM, and the kcat was 0.011 s-1.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=178861Documentos Relacionados
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