An efficient thermostable organophosphate hydrolase and its application in pesticide decontamination
An efficient thermostable organophosphate hydrolase and its application in pesticide decontamination
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DOI：
10.1002/bit.25843
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发表时间：
2016-04-01
影响因子：
3.8
通讯作者：
Manco, Giuseppe
中科院分区：
文献类型：
--
作者：
Del Giudice, Immacolata;Coppolecchia, Rossella;Manco, Giuseppe
酶的体外进化代表了进化新的或改善弱酶功能的强大装置。在目前的工作中，使用半理性工程方法从内酯酶支架（来自硫磺硫化叶菌的 SsoPox）开始设计高效且热稳定的有机磷酸酯水解酶。特别是，通过SsoPox辅助混杂活性的体外进化，获得了三重突变体C258L/I261F/W263A，该突变体在保留其固有稳定性的同时，对氧磷的水解活性增强了300倍，催化效率的绝对值高达10(5) M(-1)s(-1)。对这种增强活性的动力学和结构决定因素进行了彻底研究，并且为了评估其潜在的生物技术应用，在不同溶剂（甲醇或乙醇）或洗涤剂（SDS或商业肥皂）的配方中测试了突变体，以清洁农药污染的表面。生物技术。 (c) 2015 年 Wiley 期刊公司。
In vitro evolution of enzymes represents a powerful device to evolve new or to improve weak enzymatic functions. In the present work a semi-rational engineering approach has been used to design an efficient and thermostable organophosphate hydrolase, starting from a lactonase scaffold (SsoPox from Sulfolobus solfataricus). In particular, by in vitro evolution of the SsoPox ancillary promiscuous activity, the triple mutant C258L/I261F/W263A has been obtained which, retaining its inherent stability, showed an enhancement of its hydrolytic activity on paraoxon up to 300-fold, achieving absolute values of catalytic efficiency up to 10(5) M(-1)s(-1). The kinetics and structural determinants of this enhanced activity were thoroughly investigated and, in order to evaluate its potential biotechnological applications, the mutant was tested in formulations of different solvents (methanol or ethanol) or detergents (SDS or a commercial soap) for the cleaning of pesticide-contaminated surfaces. Biotechnol. (c) 2015 Wiley Periodicals, Inc.