Fan Xinjiong, Zhao Meng, Wen Huamei, Zhang Yanyu, Zhang Yixin, Zhang Jing, Liu Xiaolong
College & Hospital of Stomatology, Anhui Medical University, Key Lab. of Oral Diseases Research of Anhui Province, Hefei, 230032, China; School of Basic Medical Sciences, Anhui Medical University, 81 Meishan Rd, Hefei, 230032, Anhui, China.
School of Basic Medical Sciences, Anhui Medical University, 81 Meishan Rd, Hefei, 230032, Anhui, China; Minhang Branch, Zhongshan Hospital, Fudan University,170 Xinsong Rd, Shanghai 200000, China.
Chemosphere. 2023 Apr;319:138021. doi: 10.1016/j.chemosphere.2023.138021. Epub 2023 Jan 30.
The pervasive use of pyrethroids is seriously hazardous to the environment and even human health. Enzymatic bioremediation is potentially a rapid and environmentally friendly technology to combat the pollution of pyrethroid pesticides. The hydrolysis of ester linkages is the initial and critical enzymatic step in microbial degradation pathways. Here, the versatile and thermostable esterase Est816 was cloned and its new function, pyrethroid-hydrolysis activity, was expanded. To further improve its pyrethroid-hydrolysis ability, Est816 was modified by rational design. After two rounds of mutation, the best-performing mutant, Est816 was obtained, which could completely degrade 1 mg/L λ-cyhalothrin, cypermethrin, and deltamethrin within 20 min, and efficiently degrade fenvalerate, reaching over 80% conversion. Degradation activity analyses showed that three substitutions (A216V, K238 N and M97V) were beneficial for enhancing the activity of Est816. Enzymatic characterization showed that Est816 inherited broad substrate specificity and possessed excellent stability and adaptability over wide ranges of temperature and pH, which is essential for bioremediation in frequently changing conditions. Furthermore, Est816 had the best degradation effect on all four pyrethroid residues in Panax notoginseng root, with more than 87% conversion after 24 h. Pyrethroid residues in tea, cucumber, and soil were reduced by more than 76%, 80%, and 76%, respectively. Taken together, these findings highlight the great potential of Est816 in the bioremediation of pyrethroid-contaminated soil and agricultural products.
拟除虫菊酯的广泛使用对环境甚至人类健康都有严重危害。酶促生物修复可能是一种快速且环保的技术，用于对抗拟除虫菊酯类农药污染。酯键的水解是微生物降解途径中的初始关键酶促步骤。在此，克隆了多功能且热稳定的酯酶Est816，并拓展了其新功能——拟除虫菊酯水解活性。为进一步提高其拟除虫菊酯水解能力，通过合理设计对Est816进行了改造。经过两轮突变，获得了性能最佳的突变体Est816，它能在20分钟内完全降解1毫克/升的氯氟氰菊酯、氯氰菊酯和溴氰菊酯，并能有效降解氰戊菊酯，转化率超过80%。降解活性分析表明，三个取代位点（A216V、K238N和M97V）有利于增强Est816的活性。酶学特性表明，Est816继承了广泛的底物特异性，在较宽的温度和pH范围内具有出色的稳定性和适应性，这对于在频繁变化的条件下进行生物修复至关重要。此外，Est816对三七根中所有四种拟除虫菊酯残留的降解效果最佳，24小时后转化率超过87%。茶叶、黄瓜和土壤中的拟除虫菊酯残留分别减少了76%以上、80%以上和76%以上。综上所述，这些发现突出了Est816在拟除虫菊酯污染土壤和农产品生物修复中的巨大潜力。
