Ke Zhijian, Zhu Qian, Jiang Wankui, Zhou Yidong, Zhang Mingliang, Jiang Mingli, Hong Qing
Key Laboratory of Agricultural Environmental Microbiology, Ministry of Agriculture, College of Life Sciences, Nanjing Agricultural University, Nanjing, Jiangsu 210095, PR China.
Key Laboratory of Agricultural Environmental Microbiology, Ministry of Agriculture, College of Life Sciences, Nanjing Agricultural University, Nanjing, Jiangsu 210095, PR China.
Ecotoxicol Environ Saf. 2021 Aug 17;224:112666. doi: 10.1016/j.ecoenv.2021.112666.
Carbaryl is the representative of carbamate insecticide. As an acetylcholinesterase inhibitor, it poses potential threat to humans and other non-target organisms. Agrobacterium sp. XWY-2, which could grow with carbaryl as the sole carbon source, was isolated and characterized. The carH gene, encoding a carbaryl hydrolase, was cloned from strain XWY-2 and expressed in Escherichia coli BL21 (DE3). CarH was able to hydrolyze carbamate pesticides including carbaryl, carbofuran, isoprocarb, propoxur and fenobucarb efficiently, while it hydrolyzed oxamyl and aldicarb poorly. The optimal pH of CarH was 8.0 and the optimal temperature was 30 ℃. The apparent K and k values of CarH for carbaryl were 38.01 ± 2.81 μM and 0.33 ± 0.01 s, respectively. The point mutation experiment demonstrated that His341, His343, His346, His416 and D437 are the key sites for CarH to hydrolyze carbaryl.
西维因是氨基甲酸酯类杀虫剂的代表。作为一种乙酰胆碱酯酶抑制剂，它对人类和其他非靶标生物构成潜在威胁。分离并鉴定了能以西维因作为唯一碳源生长的土壤杆菌属菌株XWY-2。从菌株XWY-2中克隆了编码西维因水解酶的carH基因，并在大肠杆菌BL21(DE3)中表达。CarH能够高效水解包括西维因、克百威、异丙威、残杀威和仲丁威在内的氨基甲酸酯类农药，而对杀螟丹和涕灭威的水解效果较差。CarH的最适pH为8.0，最适温度为30℃。CarH对西维因的表观K和k值分别为38.01±2.81μM和0.33±0.01 s⁻¹。点突变实验表明，His341、His343、His346、His416和D437是CarH水解西维因的关键位点。
