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Characterization of hybrid biphenyl dioxygenases obtained by recombining Burkholderia sp. strain LB400 bphA with the homologous gene of Comamonas testosteroni B-356

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The bacterial degradation of polychlorinated biphenyls depends on the ability of the enzyme biphenyl 2,3-dioxygenase (BPDO) to catalyze their oxygenation. Analysis of hybrid BPDOs obtained using common restriction sites to exchange large DNA fragments between LB400 bphA and B-356 bphA showed that the C-terminal portion of LB400 α subunit can withstand extensive structural modifications, and that these modifications can change the catalytic properties of the enzyme. On the other hand, exchanging the C-terminal portion of B-356 BPDO α subunit with that of LB400 α subunit generated inactive chimeras. Data encourage an enzyme engineering approach, consisting of introducing extensive modifications of the C-terminal portion of LB400 bphA to extend BPDO catalytic properties toward polychlorinated biphenyls.Key words: PCB, protein engineering, BphA, BPDO, polychlorinated biphenyl.

La dégradation bactérienne des biphényles polychlorés dépend de la capacité de l'enzyme biphényle 2,3-dioxygénase (BPDO) à catalyser leur oxygénation. L'analyse de BPDO hybrides obtenues en utilisant des sites de restriction communs pour échanger de larges fragments d'ADN entre les gènes bphA de LB400 et de B-356, a montré que la sous unité α de LB400 restait active en dépit de modifications structurelles majeures dans sa portion C-terminale. De plus, ces modifications pouvaient changer les propriétés catalytiques de l'enzyme envers les BPC. Toutefois, le remplacement de la partie C-terminale de la sous unité α de B-356 par celle de LB400 a généré des enzymes hybrides inactives. Nos résultats nous amènent à proposer une approche pour l'ingénierie d'une BPDO potentiel d'activité accrue envers les biphényles polychlorés qui consisterait à introduire de façon aléatoire, d'importantes modifications dans la partie de bphA de LB400 qui code pour la portion C-terminale de la protéine.Mots clés : BPC, ingénierie des protéines, BphA, BPDO, biphényles polychlorés.

Keywords: BPC; BPDO; BphA; PCB; biphényles polychlorés; ingénierie des protéines; polychlorinated biphenyl; protein engineering

Document Type: Research Article

Publication date: 2001-11-01

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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