Institute of Applied Ecology, Chinese Academy of Sciences, Shenyang, China; University of Chinese Academy of Sciences, Beijing, China.
Lett Appl Microbiol. 2014 Jan;58(1):53-9. doi: 10.1111/lam.12155. Epub 2013 Sep 30.
Methyl parathion hydrolase (MPH) can degrade a wide range of organophosphorus compounds, but its efficiency in hydrolysing chlorpyrifos, one of the most popular pesticides applied for crop protection, is much lower than that in hydrolysing the preferred substrate methyl parathion. In this study, random mutagenesis was adopted to improve MPH to enhance its efficiency in hydrolysing the poorly hydrolysed substrate chlorpyrifos. Rapid screening of the improved MPH variants was carried out using Bacillus subtilis WB800 secretory expression system to investigate the distribution of improved MPH variants based on the size of clear haloes as a result of chlorpyrifos hydrolysis. Four improved MPH variants were isolated, and one variant K3, in particular, showed a 5-fold increase in kcat value for chlorpyrifos hydrolysis. Furthermore, most of the MPH variants obtained in this study possessed enhanced thermostability and pH stability. The approaches adopted in this study could be extended to create other MPH variants with increased activity for hydrolysing other poorly hydrolysed substrates.
Chlorpyrifos is one of the toxic organophosphorus compounds (OP compounds) widely used for insecticides control. Water, soil and foodstuff have been contaminated seriously by chlorpyrifos in some areas. It is urgent to find effective methods to remove its contamination. This work contributes to improve methyl parathion hydrolase (MPH) to enhance its efficiency in hydrolysing the poorly hydrolysed substrate chlorpyrifos. Our study brings new insights for enzymatic strategy for the decontamination of toxic OP compounds.
甲基对硫磷水解酶（MPH）可以降解广泛的有机磷化合物，但它对氯吡噁磷的水解效率，比其对首选基质甲基对硫磷的水解效率要低得多。在这项研究中，采用随机突变来提高 MPH 的效率，以增强其对水解效率较低的底物氯吡噁磷的水解能力。利用枯草芽孢杆菌 WB800 分泌表达系统对改良后的 MPH 变体进行快速筛选，根据氯吡噁磷水解产生的清晰晕圈大小，研究改良后的 MPH 变体的分布。分离出四个改良后的 MPH 变体，其中一个变体 K3 对氯吡噁磷水解的 kcat 值增加了 5 倍。此外，本研究中获得的大多数 MPH 变体具有增强的热稳定性和 pH 稳定性。本研究中采用的方法可以扩展到创建其他具有增加水解其他水解效率较低的基质的活性的 MPH 变体。
氯吡噁磷是广泛用于杀虫剂控制的有毒有机磷化合物（OP 化合物）之一。在一些地区，水、土壤和食物已受到严重污染。迫切需要找到有效的方法来去除其污染。这项工作有助于提高甲基对硫磷水解酶（MPH）的效率，以增强其对水解效率较低的底物氯吡噁磷的水解能力。我们的研究为酶法去除有毒 OP 化合物提供了新的见解。
