Molecular characterization of the PceA reductive dehalogenase of Desulfitobacterium sp strain Y51
Molecular characterization of the PceA reductive dehalogenase of Desulfitobacterium sp strain Y51
复制标题
DOI：
10.1128/jb.184.13.3419-3425.2002
复制
发表时间：
2002-07-01
影响因子：
3.2
通讯作者：
Furukawa, K
中科院分区：
文献类型：
--
作者：
Suyama, A;Yamashita, M;Furukawa, K
对Desulfitobacterium sp.菌株Y51的四氯乙烯（PCE）还原脱卤酶（pceA基因编码，命名为pceA脱卤酶）进行了纯化和鉴定。PCE和三氯乙烯（TCE）存在时，该酶的表达被高度诱导。纯化后的酶催化PCE经TCE还原脱卤生成顺式-1,2-二氯乙烯，比活性为113.6 nmol。分钟(1)。毫克蛋白质（-1）。PCE和TCE的表观K-m值分别为105.7和535.3 muM。除PCE和TCE以外的氯代乙烯不脱卤。该酶对六氯乙烷、五氯乙烷、1,1,1,2-四氯乙烷和1,1,2,2-四氯乙烷等氯代乙烷均有脱卤活性。在2.8 kb的DNA片段中鉴定出了Desulfitobacterium sp.菌株Y51的pceA基因，并在大肠杆菌中表达该蛋白，制备抗体。免疫印迹分析发现PceA位于细胞的周质中。
The tetrachloroethene (PCE) reductive dehalogenase (encoded by the pceA gene and designated PceA dehalogenase) of Desulfitobacterium sp. strain Y51 was purified and characterized. The expression of the enzyme was highly induced in the presence of PCE and trichloroethene (TCE). The purified enzyme catalyzed the reductive dehalogenation of PCE via TCE to cis-1,2-dichloroethene at a specific activity of 113.6 nmol . min(-1) . mg of protein(-1). The apparent K-m values for PCE and TCE were 105.7 and 535.3 muM, respectively. Chlorinated ethenes other than PCE and TCE were not dehalogenated. However, the enzyme exhibited dehalogenation activity for various chlorinated ethanes such as hexachloroethane, pentachloroethane, 1,1,1,2-tetrachloroethane, and 1,1,2,2-tetrachloroethane. The pceA gene of Desulfitobacterium sp. strain Y51 was identified in a 2.8-kb DNA fragment and used to express the protein in Escherichia coli for the preparation of antibodies. Immunoblot analyses located PceA in the periplasm of the cell.