A bifunctional enzyme belonging to cytochrome P450 family involved in the O-dealkylation and N-dealkoxymethylation toward chloroacetanilide herbicides in Rhodococcus sp. B2.
A bifunctional enzyme belonging to cytochrome P450 family involved in the O-dealkylation and N-dealkoxymethylation toward chloroacetanilide herbicides in Rhodococcus sp. B2.
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属于细胞色素 P450 家族的双功能酶，参与红球菌属氯乙酰苯胺除草剂的 O-脱烷基化和 N-脱烷氧基甲基化。
DOI：
10.1186/s12934-021-01544-z
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发表时间：
2021-03-04
影响因子：
6.4
通讯作者：
Sun LN
中科院分区：
文献类型：
--
作者：
Liu HM;Yuan M;Liu AM;Ren L;Zhu GP;Sun LN
氯乙酰胺类除草剂丙草胺是一种新兴的污染物。由于大量使用，其在环境中的存在威胁着人类健康。然而，丙草胺降解的分子机制仍然未知。目前，从稻田中分离到一株具有降解丙草胺能力的红球菌B2。在30.1 °C、初始底物浓度50 mg/L、pH 6.98、培养时间5 d时，丙草胺的降解率最高，为86.1%。通过气相色谱-质谱联用技术（GC-MS）鉴定了一种新的代谢产物N-羟乙基-2-氯-N-（2，6-二乙基苯基）-乙酰胺。基因组草图比较表明，一个32，147-bp的DNA片段，窝藏基因簇（EthRABCDB 2），从突变株TB 2不能降解丙草胺。Eth基因簇编码AraC/XylS家族转录调节因子（EthRB 2）、铁氧还蛋白还原酶（EthAB 2）、细胞色素P450单加氧酶（EthBB 2）、铁氧还蛋白（EthCB 2）和功能未知的10-kDa蛋白（EthDB 2）。与EthABCDB 2和EthABDB 2互补，但不与菌株TB 2中的EthABCB 2互补，恢复了其降解氯乙酰胺除草剂的能力。随后，进行EthABCDB 2的密码子优化，之后优化的组分在大肠杆菌中分别表达，并使用Ni亲和层析纯化。EthABCDB 2或EthABDB 2的混合物催化甲草胺、乙草胺、丁草胺和异丙草胺的N-脱烷氧基甲基化和丙草胺的O-脱烷基化，而EthABCB 2不催化，表明EthDB 2在菌株B2中充当铁氧还蛋白。EthABDB 2在30 °C和pH 7.5下显示最大活性。这是首次报道的P450家族加氧酶催化丙草胺和异丙草胺的O-脱烷基化和N-脱烷氧基甲基化。研究结果为氯乙酰胺类农药污染场地的修复提供了微生物资源。
The chloroacetamide herbicides pretilachlor is an emerging pollutant. Due to the large amount of use, its presence in the environment threatens human health. However, the molecular mechanism of pretilachlor degradation remains unknown. Now, Rhodococcus sp. B2 was isolated from rice field and shown to degrade pretilachlor. The maximum pretilachlor degradation efficiency (86.1%) was observed at a culture time of 5 d, an initial substrate concentration 50 mg/L, pH 6.98, and 30.1 °C. One novel metabolite N-hydroxyethyl-2-chloro-N-(2, 6-diethyl-phenyl)-acetamide was identified by gas chromatography-mass spectrometry (GC–MS). Draft genome comparison demonstrated that a 32,147-bp DNA fragment, harboring gene cluster (EthRABCDB2), was absent from the mutant strain TB2 which could not degrade pretilachlor. The Eth gene cluster, encodes an AraC/XylS family transcriptional regulator (EthRB2), a ferredoxin reductase (EthAB2), a cytochrome P450 monooxygenase (EthBB2), a ferredoxin (EthCB2) and a 10-kDa protein of unknown function (EthDB2). Complementation with EthABCDB2 and EthABDB2, but not EthABCB2 in strain TB2 restored its ability to degrade chloroacetamide herbicides. Subsequently, codon optimization of EthABCDB2 was performed, after which the optimized components were separately expressed in Escherichia coli, and purified using Ni-affinity chromatography. A mixture of EthABCDB2 or EthABDB2 but not EthABCB2 catalyzed the N-dealkoxymethylation of alachlor, acetochlor, butachlor, and propisochlor and O-dealkylation of pretilachlor, revealing that EthDB2 acted as a ferredoxin in strain B2. EthABDB2 displayed maximal activity at 30 °C and pH 7.5. This is the first report of a P450 family oxygenase catalyzing the O-dealkylation and N-dealkoxymethylation of pretilachlor and propisochlor, respectively. And the results of the present study provide a microbial resource for the remediation of chloroacetamide herbicides-contaminated sites.
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影响因子：
4.3
作者：
Jiang, Jinhua;Chen, Yanhong;Cai, Leiming
通讯作者：
Cai, Leiming
DOI：
10.1016/s1383-5742(99)00019-8
发表时间：
1999-07-15
影响因子：
1.9
作者：
Dearfield, KL;McCarroll, NE;Waters, MD
通讯作者：
Waters, MD
影响因子：
2.4
作者：
Hou, Y.;Dong, W.;Cui, Z.
通讯作者：
Cui, Z.
DOI：
10.1080/03601230600851141
发表时间：
2006-01-01
影响因子：
2
作者：
Pal, R.;Das, P.;Chowdhury, A.
通讯作者：
Chowdhury, A.
DOI：
10.1007/s001280000197
发表时间：
2001-01-01
影响因子：
2.7
作者：
Chiang, HC;Duh, JR;Wang, YS
通讯作者：
Wang, YS