疏棉状嗜热丝孢菌(Thermomyces lanuginosus)脂肪酶的理性设计提高其活性和温度稳定性  

Improving the Activity and Thermal Stability of Thermomyces lanuginosus Lipase by Rational Design

作　　者：魏子翔 张柳群 雷磊 韩正刚 杨江科 WEI Zi-xiang;ZHANG Liu-qun;LEI Lei;HAN Zheng-gang;YANG Jiang-ke(College of Biology and Pharmaceutical Engineering,Wuhan Polytechnic University,Wuhan 430023,China)

机构地区：[1]武汉轻工大学生物与制药工程学院,武汉430023

出　　处：《中国生物工程杂志》2021年第2期63-69,共7页China Biotechnology

摘　　要：目的:通过对疏棉状嗜热丝孢菌(Thermomyces lanuginosus)脂肪酶的理性设计,获得高酶活与耐高温的脂肪酶品种,为脂肪酶在饲料、油脂加工和生物柴油等领域的应用奠定基础。方法:对脂肪酶典型结构域lid和loop区域的系统发育分析,找到候选的位点,理性设计并通过实验验证,获得脂肪酶活性和耐高温特性显著提高的脂肪酶重组子,并构建多拷贝载体,完成50L发酵罐中进行产酶能力评价。结果:经过设计的脂肪酶高温稳定性显著提升。其在80℃下放置12h后仍保留78.94%的酶活。脂肪酶重组子在50L发酵罐中发酵诱导168h,其上清液酶活达到29000U/m L。结论:成功设计并获得了一种新型的高活性、耐高温的脂肪酶品种TLL,并实现了高效表达,为其产业化和工业应用奠定了基础。Objective:In order to obtain a lipase with high enzyme activity and high temperature resistance,the Thermomyces lanuginosus lipase was modified by rational design,which laid the foundation for the application of lipase in the feed industry,oil processing and biodiesel.Methods:A candidate site was found by phylogenetic analysis of the lid and loop regions of the typical domain of lipase.A lipase recombinant with significantly improved lipase activity and high temperature resistance was obtained through rational design and experimental verification,and a multi-copy vector was constructed to complete the evaluation of the enzyme production ability in a 50 L fermentor.Results:The thermal stability of lipase was significantly improved after designing.The crude enzyme solution still retained 78.94%of the enzyme activity after being placed at 80℃for 12 h.After fermentation for 168 h in a 50 L fermentor,the enzyme activity of the supernatant reached 29000 U/m L.Conclusion:A novel high activity and high thermal stable lipase was successfully designed and obtained,which facilitates its large-scale production and industrial application.

关 键 词：脂肪酶 理性设计 高活性 高温稳定性 

分 类 号：Q814.9[生物学—生物工程]