title: Directed evolution of biphenyl dioxygenase: emergence of enhanced degradation capacity for benzene, toluene, and alkylbenzenes.
abstract: Biphenyl dioxygenase (Bph Dox) catalyzes the initial oxygenation of biphenyl and related compounds. Bph Dox is a multicomponent enzyme in which a large subunit (encoded by the bphA1 gene) is significantly responsible for substrate specificity. By using the process of DNA shuffling of bphA1 of Pseudomonas pseudoalcaligenes KF707 and Burkholderia cepacia LB400, a number of evolved Bph Dox enzymes were created. Among them, an Escherichia coli clone expressing chimeric Bph Dox exhibited extremely enhanced benzene-, toluene-, and alkylbenzene-degrading abilities. In this evolved BphA1, four amino acids (H255Q, V258I, G268A, and F277Y) were changed from the KF707 enzyme to those of the LB400 enzyme. Subsequent site-directed mutagenesis allowed us to determine the amino acids responsible for the degradation of monocyclic aromatic hydrocarbons.
authors.affiliation: Department of Biosciences and Biotechnology, Faculty of Agriculture, Kyushu University, Fukuoka 812-8581, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Suenaga
authors.forename: M
authors.initials: M
authors.lastname: Mitsuoka
authors.forename: Y
authors.initials: Y
authors.lastname: Ura
authors.forename: T
authors.initials: T
authors.lastname: Watanabe
authors.forename: K
authors.initials: K
authors.lastname: Furukawa
mesh_terms: D000595:Amino Acid Sequence; D001554:Benzene / Q000737:chemistry / Q000378:metabolism*; D001555:Benzene Derivatives / Q000378:metabolism*; D016956:Burkholderia cepacia / Q000201:enzymology / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D019020:Directed Molecular Evolution; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D007506:Iron-Sulfur Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010105:Oxygenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011549:Pseudomonas / Q000201:enzymology / Q000235:genetics; D017422:Sequence Analysis, DNA; D014050:Toluene / Q000378:metabolism*
chemical_list: D001555:Benzene Derivatives; D004269:DNA, Bacterial; D007506:Iron-Sulfur Proteins; D014050:Toluene; D010105:Oxygenases; C074161:biphenyl-2,3-dioxygenase; D001554:Benzene