Performance enhancement of chemically modified lipase in the organic phase guided by molecular simulation

doi:10.16085/j.issn.1000-6613.2015.10.032

Abstract

Abstract: Lipase is one of the most important biocatalysts because of its stereoselectivity towards chiral substrates and reasonable adaptiveness of non-conventional reaction media, including hydrophobic organic solvents. Increasing the selectivity of lipase in such non-conventional environments has been a continuous hotspot. However, current researches on improving lipase selectivity usually cause critical decrease of catalytic activity. In this work, a novel computation- directed approach to enhance lipase enantioselectivity in organic media without losing activity was introduced. Stereoselective acylation of chiral secondary alcohols catalyzed by PcL(Pseudomonas cepacia lipase) in n-hexane was taken as the model reaction. According to the results from molecular dynamics simulation, when the Tyr²⁹ inside the PcL catalytic cavity was modified by N-acetylimidazole (NAI), the enantioselectivity of the lipase was significantly increased. The simulation result was agreed with the experimental data. MALDI-TOF-MS confirmed the Tyr²⁹ modified position, and kinetic resolution of chiral secondary alcohols by NAI-modified PcL displayed an increase of enantioselectivity ratio from 12.6 to 48.1 and higher activity.

Key words: chemical modification, simulation, chiral secondary alcohol, enantioselectivity, lipase

摘要： 脂肪酶是一种应用广泛的重要生物催化剂,它对手性底物具有良好的立体选择性,而且能适应包括疏水有机溶剂在内的多种非天然反应介质。提高脂肪酶在非天然环境中的立体选择性已经成为关于脂肪酶的研究热点。然而已有研究在提高选择性的同时,往往伴随催化活力的降低。本文提出一种基于计算机模拟的理性改造方法,能够在不降低脂肪酶活力的情况下大幅提高其在有机介质中的立体选择性。以洋葱假单胞菌脂肪酶(Pseudomonas cepacia lipase,PcL)在正己烷中催化手性仲醇的转酯化反应为模型,借助分子动力学模拟,预测 N-乙酰咪唑(NAI)修饰 PcL的效果。蛋白质谱证实产生主要影响的是位于 PcL活性口袋内的 Tyr²⁹ 残基。进一步通过实验验证,发现经 NAI 修饰酪氨酸残基(Tyr)后 PcL催化拆分手性仲醇的对映体选择性最高从 E = 12.6 提高到 48.1,同时催化活力也有所提高。

关键词: 化学修饰, 模拟, 手性仲醇, 对映体选择性, 脂肪酶

CLC Number:

O643

GUO Li, XU Gang, WU Jianping, YANG Lirong. Performance enhancement of chemically modified lipase in the organic phase guided by molecular simulation[J]. Chemical Industry and Engineering Progree, 2015, 34(10): 3725-3730.

郭丽, 徐刚, 吴坚平, 杨立荣. 分子模拟指导化学修饰对有机相中脂肪酶的性能强化[J]. 化工进展, 2015, 34(10): 3725-3730.

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