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Please use this identifier to cite or link to this item: http://repository.iitr.ac.in/handle/123456789/26942
Title: Structural basis of the enhanced pollutant-degrading capabilities of an engineered biphenyl dioxygenase
Authors: Dhindwal S.
Gomez-Gil L.
Neau D.B.
Pham T.T.M.
Sylvestre M.
Eltis L.D.
Bolin J.T.
Kumar, Pravindra R.Manish
Published in: Journal of Bacteriology
Abstract: Biphenyl dioxygenase, the first enzyme of the biphenyl catabolic pathway, is a major determinant of which polychlorinated biphenyl (PCB) congeners are metabolized by a given bacterial strain. Ongoing efforts aim to engineer BphAE, the oxygenase component of the enzyme, to efficiently transform a wider range of congeners. BphAEII9, a variant of BphAELB400 in which a sevenresidue segment, 335TFNNIRI341, has been replaced by the corresponding segment of BphAEB356, 333GINTIRT339, transforms a broader range of PCB congeners than does either BphAELB400 or BphAEB356, including 2,6-dichlorobiphenyl, 3,3=-dichlorobiphenyl, 4,4=-dichlorobiphenyl, and 2,3,4=-trichlorobiphenyl. To understand the structural basis of the enhanced activity of BphAEII9, we have determined the three-dimensional structure of this variant in substrate-free and biphenyl-bound forms. Structural comparison with BphAELB400 reveals a flexible active-site mouth and a relaxed substrate binding pocket in BphAEII9 that allow it to bind different congeners and which could be responsible for the enzyme's altered specificity. Biochemical experiments revealed that BphAEII9 transformed 2,3,4=-trichlorobiphenyl and 2,2=,5,5=-tetrachlorobiphenyl more efficiently than did BphAELB400 and BphAEB356. BphAEII9 also transformed the insecticide dichlorodiphenyltrichloroethane (DDT) more efficiently than did either parental enzyme (apparent kcat/Km of 2.2±0.5mM-1 s-1, versus 0.9±0.5mM-1 s-1 for BphAEB356). Studies of docking of the enzymes with these three substrates provide insight into the structural basis of the different substrate selectivities and regiospecificities of the enzymes. © 2016, American Society for Microbiology.
Citation: Journal of Bacteriology, 198(10): 1499-1512
URI: https://doi.org/10.1128/JB.00952-15
http://repository.iitr.ac.in/handle/123456789/26942
Issue Date: 2016
Publisher: American Society for Microbiology
Keywords: 2,3,4' trichlorobiphenyl
2,6 dichlorobiphenyl
3,3' dichlorobiphenyl
4,4' dichlorobiphenyl
biphenyl
biphenyl dioxygenase
chlorphenotane
dioxygenase
oxygenase
unclassified drug
biphenyl
biphenyl derivative
protein binding
Article
comparative study
enzyme activity
enzyme binding
enzyme degradation
enzyme engineering
enzyme specificity
enzyme structure
enzyme substrate
molecular docking
multigene family
priority journal
chemistry
enzyme active site
genetics
metabolism
molecular model
procedures
protein conformation
protein engineering
X ray crystallography
Biphenyl Compounds
Catalytic Domain
Crystallography, X-Ray
Models, Molecular
Molecular Docking Simulation
Oxygenases
Protein Binding
Protein Conformation
Protein Engineering
Substrate Specificity
ISSN: 219193
Author Scopus IDs: 36082537700
18233632900
18042121400
54791518500
7005984229
7003863696
57197844041
55064809000
Author Affiliations: Dhindwal, S., Department of Biotechnology, Indian Institute of Technology Roorkee, Roorkee, Uttarakhand, India
Gomez-Gil, L., Department of Microbiology and Biochemistry, Life Sciences Institute, The University of British Columbia, Vancouver, BC, Canada
Neau, D.B., Department of Chemistry and Chemical Biology, Cornell University, Northeastern Collaborative Access Team, Argonne National Laboratory, Argonne, IL, United States
Pham, T.T.M., Institut National de Recherche Scientifique (INRS-Institut Armand-Frappier), Laval, QC, United States
Sylvestre, M., Institut National de Recherche Scientifique (INRS-Institut Armand-Frappier), Laval, QC, United States
Eltis, L.D., Department of Microbiology and Biochemistry, Life Sciences Institute, The University of British Columbia, Vancouver, BC, Canada
Bolin, J.T., Department of Biological Sciences, Center for Cancer Research, Purdue University, West Lafayette, IN, United States
Kumara, P., Department of Biotechnology, Indian Institute of Technology Roorkee, Roorkee, Uttarakhand, India
Funding Details: This work was supported by the DRDO, India. This research used resources of the Advanced Photon Source, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by the Argonne National Laboratory under contract no. DE-AC02-06CH11357. Use of BioCARS was also supported by the National Institute of General Medical Sciences (NIGMS) of the National Institutes of Health under grant number R24GM111072. D.B.N. is supported by grant number GM103403 from the NIGMS. This work, including the efforts of Lindsay D. Eltis, was funded by Gouvernement du Canada / Natural Sciences and Engineering Research Council of Canada (NSERC) (Discovery grant). This work, including the efforts of Pravindra Kumar, was funded by Defence Research and Development Organisation (DRDO) (ERIP/ER/1000391/M/01/1390). National Institutes of Health, NIH; U.S. Department of Energy, USDOE; National Institute of General Medical Sciences, NIGMS: P41GM103403, R24GM111072; Office of Science, SC; Argonne National Laboratory, ANL: DE-AC02-06CH11357; Government of Canada; Natural Sciences and Engineering Research Council of Canada, NSERC; Defence Research and Development Organisation, DRDO: ERIP/ER/1000391/M/01/1390
Corresponding Author: Kumara, P.; Department of Biotechnology, India; email: pravshai@gmail.com
Appears in Collections:Journal Publications [BT]

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