Institute of Biotechnology, Shanxi University, China
Received Date: October 18, 2012; Accepted Date: October 20, 2012; Published Date: October 22, 2012
Citation: Yang X (2012) Enzymes Related to Polychlorinated Biphenyl and Enzyme Kinetics. Biochem Anal Biochem 1:e124. doi: 10.4172/2161-1009.1000e124
Copyright: © 2012 Yang X. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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Polychlorinated biphenyls (PCBs) are now one of the most serious recalcitrant pollutants, the use of microorganisms is expected to be an effective tool for remediation of PCB-polluted environments. In the most PCB degrader, the conversion of biphenyl to a benzoic and other 2-hydroxypenta-2,4-dienoate is catalyzed by a series of enzymes: biphenyl dioxygease (BphA), dihydrodia dehydroxygenase (BphB), 2,3-dihydroxybiphenyl dioxygenase (BphC) and 2-hydroxyl- 6-oxo-6-phenylhexa-2,4-dienoic acid hydrolase (BphD). Biphenyl 2,3-dioxygenase catalyzes the oxygenation of two vicinal ortho-meta carbons of the biphenyl ring to yield a 2,3-dihydrodihydroxybiphenyl (DDBP). The dihydrodiol intermediate is then substrate for BphB, an NAD+ dependent dehydrogenase that produces 2,3-dihydroxybiphenyl (DHBP) for BphC, a ring fission dioxygenase. Following ring fission, hydrolytic cleavage by BphD produces benzoate and 2-hydroxypenta- 2,4-dienoate.
In Rhodococcus sp. strain R04, genes (bphBCA1A2A3A4D) encoding for biphenyl 2,3-dioxygenase (BphA1A2A3A4), cis-2,3-dihydro-2,3- dihydroxybiphenyl dehydrogenase (BphB), 2,3-dihydroxybiphenyl 1,2-dioxygenase (BphC) and 2-hydroxy-6-oxo-6-phenylhexa-2,4- dienoate hydrolase (HOPDA hydrolase, BphD), played an important role in degradation of biphenyl, and organized in an operon. The bph genes in strain R04 are preceded by bphB, and followed by bphC, bphA1A2A3A4 and bphD. The gene encoding the α-subunit of biphenyl dioxygenase (BphA1) separated from bphC by an orf2 of unknown function. Furthermore, a 330 bp and a 450 bp fragments are localized between bphA2 and bphA3, bphA3 and bphA4, respectively. A more detailed analysis revealed that bph genes organization in strain R04 varied from other PCB degrader.
We have characterized two enzymes BphC and BphD in the bph pathway, and proved them to be thermostable. Pre-steady-state analysis was carried out on wild-type BphD and the data simulated by curve-fitting. The results shows that Ser-110 plays a dominant role in the course of C–C cleavage; His-265 is responsible for ketonisation of the substrate and participates C–C cleavage along with Ser-110 and Trp-266; in the course of C–C cleavage catalyzed by the BphD, Trp- 266 also plays a very important role in this enzyme catalyzed reaction expect for catalytic triad (Ser-110, Asp-237, His-265).
Our objective is to provide the reader with some view of the 2,3-dihydroxybiphenyl 1,2-dioxygenase and biphenyl hydrolase, including (a) the regulation of PCB metabolism to several 2,3-dihydroxybiphenyl 1,2-dioxygenase, (b) the directed evolution of biphenyl hydrolase, (c) pre-steady-stable kinetics of biphenyl hydrolase and its mutants. We hope that the result of this endeavour is a repository of information that will be useful not only for the specialist to delve the wishes into the complexities of a specific area but also for those more casual readers just wanting to dip their toes in the water.