Autotransporter domain-dependent enzymatic analysis of a novel extremely thermostable carboxylesterase with high biodegradability towards pyrethroid pesticides.
Autotransporter domain-dependent enzymatic analysis of a novel extremely thermostable carboxylesterase with high biodegradability towards pyrethroid pesticides.
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对拟除虫菊酯农药具有高生物降解性的新型极其耐热的羧酸酯酶的自转运蛋白结构域依赖性酶分析
DOI：
10.1038/s41598-017-03561-8
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发表时间：
2017-06-14
影响因子：
4.6
通讯作者：
Wei D
中科院分区：
文献类型：
--
作者：
Cai X;Wang W;Lin L;He D;Huang G;Shen Y;Wei W;Wei D
克隆并测序了EstPS1基因，该基因编码从石油井生产的水中分离出的Pseudomonas synxanthaPS1的新型羧酸酯酶。EstPS1具有1923 bp的开放阅读框，编码640个氨基酸的羧酸酯酶（EstPS1），其包含自转运蛋白（AT）结构域（357-640个氨基酸）。同源性分析显示，EstPS1 与荧光假单胞菌 A506（NCBI 数据库）的 EstA 具有最高的同源性（88%），并且属于羧酸酯酶家族（EC 3.1.1.1）。重组EstPS1的最适pH和温度分别为8.0和60°C。 EstPS1表现出较高的热稳定性，在60、70、80、90和100℃下的半衰期（T1/2热失活）分别为14小时、2小时、31分钟、10分钟和1分钟。为了了解 AT 结构域在羧酸酯酶中的作用，生成了 AT 结构域截短的羧酸酯酶 (EstPS1ΔAT)。 EstPS1ΔAT 显示出明显降低的分泌率，因为 AT 结构域大大提高了异质系统中的分泌表达。 EstPS1 可降解多种拟除虫菊酯农药，水解效率取决于拟除虫菊酯分子结构。 EstPS1降解了所有测试的拟除虫菊酯农药和水解的中短链脂肪酸对硝基苯酯，表明EstPS1是一种具有广泛特异性的酯酶。
TheEstPS1 gene, which encodes a novel carboxylesterase ofPseudomonas synxanthaPS1 isolated from oil well-produced water, was cloned and sequenced.EstPS1 has an open reading frame of 1923 bp and encodes the 640-amino acid carboxylesterase (EstPS1), which contains an autotransporter (AT) domain (357–640 amino acids). Homology analysis revealed that EstPS1 shared the highest identity (88%) with EstA fromPseudomonas fluorescensA506 (NCBI database) and belonged to the carboxylesterase family (EC 3.1.1.1). The optimum pH and temperature of recombinant EstPS1 were found to be 8.0 and 60 °C, respectively. EstPS1 showed high thermostability, and the half-lives (T1/2thermal inactivation) at 60, 70, 80, 90, and 100 °C were 14 h, 2 h, 31 min, 10 min, and 1 min, respectively. To understand the role of the AT domain in carboxylesterase, AT domain-truncated carboxylesterase (EstPS1ΔAT) was generated. EstPS1ΔAT showed a clearly decreased secretion rate, owing to the AT domain strongly improved secretory expression in the heterogeneous system. EstPS1 degraded various pyrethroid pesticides, and hydrolysis efficiencies were dependent on the pyrethroid molecular structure. EstPS1 degraded all the tested pyrethroid pesticides and hydrolysed thep-nitrophenyl esters of medium-short-chain fatty acids, indicating that EstPS1 is an esterase with broad specificity.