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基于B因子分析通过定点诱变提高脂肪酶的热稳定性

Improving the Thermostability of Lipase Through Site-Directed Mutagenesis Based on B-Factor Analysis.

作者信息

Jiang Zhanbao, Zhang Chengbo, Tang Minyuan, Xu Bo, Wang Lili, Qian Wen, He Jiandong, Zhao Zhihong, Wu Qian, Mu Yuelin, Ding Junmei, Zhang Rui, Huang Zunxi, Han Nanyu

机构信息

School of Life Sciences, Yunnan Normal University, Kunming, China.

Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University, Kunming, China.

出版信息

Front Microbiol. 2020 Mar 3;11:346. doi: 10.3389/fmicb.2020.00346. eCollection 2020.

DOI:10.3389/fmicb.2020.00346
PMID:32194535
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC7063977/
Abstract

In order to improve the thermostability of lipases derived from , we identified lipase (Lipr27RCL) mutagenesis sites that were associated with enhanced flexibility based upon B-factor analysis and multiple sequence alignment. We found that two mutated isoforms (Lipr27RCL-K64N and Lipr27RCL-K68T) exhibited enhanced thermostability and improved residual activity, with respective thermal activity retention values of 37.88% and 48.20% following a 2 h treatment at 50°C relative to wild type Lipr27RCL. In addition, these Lipr27RCL-K64N and Lipr27RCL-K68T isoforms exhibited 2.4- and 3.0-fold increases in enzymatic half-life following a 90 min incubation at 60°C. Together these results indicate that novel mutant lipases with enhanced thermostability useful for industrial applications can be predicted based upon B-factor analysis and constructed via site-directed mutagenesis.

摘要

为了提高源自[具体来源未提及]的脂肪酶的热稳定性，我们基于B因子分析和多序列比对，确定了与增强柔韧性相关的脂肪酶（Lipr27RCL）诱变位点。我们发现，两种突变异构体（Lipr27RCL-K64N和Lipr27RCL-K68T）表现出增强的热稳定性和改善的残余活性，相对于野生型Lipr27RCL，在50°C处理2小时后，其热活性保留值分别为37.88%和48.20%。此外，这些Lipr27RCL-K64N和Lipr27RCL-K68T异构体在60°C孵育90分钟后，酶促半衰期分别增加了2.4倍和3.0倍。这些结果共同表明，可以基于B因子分析预测具有增强热稳定性的新型突变脂肪酶，用于工业应用，并通过定点诱变构建。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/2600939a82d5/fmicb-11-00346-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/7309ada4a078/fmicb-11-00346-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/db149dc4ee9b/fmicb-11-00346-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/6bceeb37d166/fmicb-11-00346-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/bf09ab7ae05d/fmicb-11-00346-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/bf16d4ad350c/fmicb-11-00346-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/2600939a82d5/fmicb-11-00346-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/7309ada4a078/fmicb-11-00346-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/db149dc4ee9b/fmicb-11-00346-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/6bceeb37d166/fmicb-11-00346-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/bf09ab7ae05d/fmicb-11-00346-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/bf16d4ad350c/fmicb-11-00346-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7858/7063977/2600939a82d5/fmicb-11-00346-g006.jpg

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