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Structural Basis of the Enhanced Pollutant-Degrading Capabilities of an Engineered Biphenyl Dioxygenase

Dhindwal, Sonali; Gomez-Gil, Leticia; Neau, David; Pham, Thi Thanh My; Sylvestre, Michel; Eltis, Lindsay; Bolin, Jeffrey; Kumar, Pravindra (2016). Structural Basis of the Enhanced Pollutant-Degrading Capabilities of an Engineered Biphenyl Dioxygenase Journal of Bacteriology , vol. 198 , nº 10. p. 1499-1512. DOI: 10.1128/jb.00952-15.

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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 BphAEII9, a variant of BphAELB400 in which a seven-residue segment, 335TFNNIRI341, has been replaced by the corresponding segment of BphAEB356, 333GINTIRT339, transforms a broader range of PCB congeners than does either BphAELB400 or BphAEB356, including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 ± 0.5 mM−1 s−1, versus 0.9 ± 0.5 mM−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.

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Centre: Centre INRS-Institut Armand Frappier
Date de dépôt: 24 août 2016 13:50
Dernière modification: 24 août 2016 13:50
URI: http://espace.inrs.ca/id/eprint/4514

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