地杆菌属菌株DPO360中参与二苯并呋喃矿化的三种不同的双加氧酶的特性分析。
Characterization of three distinct extradiol dioxygenases involved in mineralization of dibenzofuran by Terrabacter sp. strain DPO360.
作者信息
Schmid A, Rothe B, Altenbuchner J, Ludwig W, Engesser K H
机构信息
Institut für Mikrobiologie, Universität Stuttgart, Germany.
出版信息
J Bacteriol. 1997 Jan;179(1):53-62. doi: 10.1128/jb.179.1.53-62.1997.
The dibenzofuran-degrading bacterial strain DPO360 represents a new species of the genus Terrabacter together with the previously described dibenzofuran-mineralizing bacterial strain DPO1361 (K.-H. Engesser, V. Strubel, K. Christoglou, P. Fischer, and H. G. Rast, FEMS Microbiol. Lett. 65:205-210, 1989; V. Strubel, Ph.D. thesis, University of Stuttgart, Stuttgart, Germany, 1991; V. Strubel, H. G. Rast, W. Fietz, H.-J. Knackmuss, and K.-H. Engesser, FEMS Microbiol. Lett. 58:233-238, 1989). Two 2,3-dihydroxybiphenyl-1,2-dioxygenases (BphC1 and BphC2) and one catechol-2,3-dioxygenase (C23O) were shown to be expressed in Terrabacter sp. strain DPO360 growing with dibenzofuran as a sole source of carbon and energy. These enzymes exhibited strong sensitivity to oxygen. They were purified to apparent homogeneity as homodimers (BphC and BphC2) and as a homotetrameric catechol-2,3-dioxygenase (C23O). According to their specificity constants kcat/Km, both BphC1 and BphC2 were shown to be responsible for the cleavage of 2,2',3-trihydroxybiphenyl, the first metabolite in dibenzofuran mineralization along the angular dioxygenation pathway. With this substrate, BphC2 exhibited a considerably higher kcat/Km, value (183 microM/min) than BphC1 (29 microM/min). Catechol-2,3-dioxygenase was recognized to be not involved in the ring cleavage of 2,2',3-trihydroxybiphenyl (kcat/Km, 1 microM/min). Analysis of deduced amino acid sequence data of bphC1 revealed 36% sequence identity to nahC from Pseudomonas putida PpG7 (S. Harayama and M. Rekik, J. Biol. Chem. 264:15328-15333, 1989) and about 40% sequence identity to various bphC genes from different Pseudomonas and Rhodococcus strains. In addition, another 2,3-dihydroxybiphenyl-1,2-dioxygenase gene (bphC3) was cloned from the genome of Terrabacter sp. strain DPO360. Expression of this gene, however, could not be detected in Terrabacter sp. strain DPO360 after growth with dibenzofuran.
二苯并呋喃降解细菌菌株DPO360与先前描述的二苯并呋喃矿化细菌菌株DPO1361（K.-H.恩格塞尔、V.施特鲁贝尔、K.克里斯托格卢、P.菲舍尔和H.G.拉斯特，《FEMS微生物学快报》65:205 - 210，1989；V.施特鲁贝尔，博士论文，德国斯图加特大学，斯图加特，1991；V.施特鲁贝尔、H.G.拉斯特、W.菲茨、H.-J.克纳克穆斯和K.-H.恩格塞尔，《FEMS微生物学快报》58:233 - 238，1989）代表地杆菌属的一个新物种。在以二苯并呋喃作为唯一碳源和能源生长的地杆菌属菌株DPO360中，已显示两种2,3 - 二羟基联苯 - 1,2 - 双加氧酶（BphC1和BphC2）和一种儿茶酚 - 2,3 - 双加氧酶（C23O）得以表达。这些酶对氧气表现出强烈的敏感性。它们被纯化至表观均一，分别为同型二聚体（BphC1和BphC2）和同型四聚体儿茶酚 - 2,3 - 双加氧酶（C23O）。根据它们的特异性常数kcat/Km，已表明BphC1和BphC2均负责2,2',3 - 三羟基联苯的裂解，2,2',3 - 三羟基联苯是二苯并呋喃沿角向双加氧途径矿化的首个代谢产物。对于该底物，BphC2表现出比BphC1（29 microM/min）高得多的kcat/Km值（183 microM/min）。已认识到儿茶酚 - 2,3 - 双加氧酶不参与2,2',3 - 三羟基联苯的环裂解（kcat/Km，1 microM/min）。对bphC1推导的氨基酸序列数据的分析显示，其与恶臭假单胞菌PpG7的nahC有36%的序列同一性（S.原山和M.雷基克，《生物化学杂志》264:15328 - 15333，1989），与来自不同假单胞菌和红球菌菌株的各种bphC基因有约40%的序列同一性。此外，从地杆菌属菌株DPO360的基因组中克隆了另一个2,3 - 二羟基联苯 - 1,2 - 双加氧酶基因（bphC3）。然而，在用二苯并呋喃生长后的地杆菌属菌株DPO360中未检测到该基因的表达。
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