Zhen Da, Liu Hong, Wang Shu-Jun, Zhang Jun-Jie, Zhao Fei, Zhou Ning-Yi
Wuhan Institute of Virology, Chinese Academy of Science, Wuhan 430071, People's Republic of China.
Appl Microbiol Biotechnol. 2006 Oct;72(4):797-803. doi: 10.1007/s00253-006-0345-2. Epub 2006 Apr 1.
A strain of Pseudomonas putida ZWL73 was isolated from soil contaminated with chloronitrobenzenes and identified by 16S rDNA sequencing. This bacterium released chloride and ammonia into the medium when grown on 4-chloronitrobenzene (4CNB) as the sole source of carbon, nitrogen and energy. A plasmid designated pZWL73 of approximately 100 kb in this strain was found to be responsible for 4CNB degradation. This was based on the fact that the plasmid-cured strains showed 4CNB- phenotype and the 4CNB+ phenotype could be conjugally transferred. The cell-free extracts of strain ZWL73 exhibited chloronitrobenzene nitroreductase and 2-amino-5-chlorophenol 1, 6-dioxygenase (2A5CPDO) activities, but neither activity was found from that of the plasmid-cured strain. We have also cloned a 4.9-kb EcoRI fragment exhibiting 2A5CPDO activity. Sequencing results revealed beta-subunit (cnbCa) and alpha subunit (cnbCb) of a meta-cleavage dioxygenase, which were subsequently expressed in E. coli with 2A5CPDO activity. The phylogenetic analysis suggested that 2A5CPDO may form a new subgroup in class III meta-cleavage dioxygenase with its close homologs.
从被氯代硝基苯污染的土壤中分离出一株恶臭假单胞菌ZWL73，并通过16S rDNA测序进行鉴定。当该细菌以4-氯硝基苯（4CNB）作为唯一碳源、氮源和能源生长时，会向培养基中释放氯离子和氨。发现该菌株中一个大小约为100 kb的名为pZWL73的质粒负责4CNB的降解。这是基于以下事实：质粒消除菌株表现出4CNB-表型，而4CNB+表型可以通过接合转移。ZWL73菌株的无细胞提取物表现出氯硝基苯硝基还原酶和2-氨基-5-氯苯酚1,6-双加氧酶（2A5CPDO）活性，但在质粒消除菌株中未发现这两种活性。我们还克隆了一个具有2A5CPDO活性的4.9 kb EcoRI片段。测序结果揭示了一种间位裂解双加氧酶的β亚基（cnbCa）和α亚基（cnbCb），随后它们在大肠杆菌中表达并具有2A5CPDO活性。系统发育分析表明，2A5CPDO与其密切同源物可能在III类间位裂解双加氧酶中形成一个新的亚组。
