Two-step computational redesign of Bacillus subtilis cellulase and β-glucanase for enhanced thermostability and activity.

Autor: Zhang H; College of Food Science and Biology, Hebei University of Science & Technology, Shijiazhuang 050018, China; AIM center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China., Zhu T; AIM center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China., Zhai Q; AIM center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China., Chen Q; Zhengzhou Tobacco Research Institute of CNTC, Zhengzhou 450001, China., Zhang X; AIM center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China., Chen Y; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., He W; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., Li J; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., Fan J; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., Tao J; Zhengzhou Tobacco Research Institute of CNTC, Zhengzhou 450001, China., Hu X; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., Qi L; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., Wang C; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., Liao K; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China., Chen Y; AIM center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China., Cui Y; AIM center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China., Chen S; China Tobacco Fujian Industrial Co., Ltd., Xiamen 361021, China. Electronic address: csy23174@fjtic.cn., Wu B; AIM center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China. Electronic address: wub@im.ac.cn.
Jazyk: angličtina
Zdroj: International journal of biological macromolecules [Int J Biol Macromol] 2024 Dec 01, pp. 138274. Date of Electronic Publication: 2024 Dec 01.
DOI: 10.1016/j.ijbiomac.2024.138274
Abstrakt: The growing demand for biocatalysts in biomass processing highlights the necessity of enhancing the thermostability of glycoside hydrolases. However, improving both thermostability and activity is often hindered by trade-offs between backbone rigidity and the flexibility of substrate-binding regions. In this study, Bacillus subtilis cellulase and β-glucanase were engineered using a two-step process incorporating the computational tools Pythia and ESM-2, which were found complementary in improving stability and activity. The engineered cellulase and β-glucanase exhibited increases in their apparent melting temperatures (5.8 °C and 8.4 °C), accompanied by up to a 1.5-fold increase in initial activities. At 50 °C, while the wild-type cellulase lost 60% of its activity after 24 h and wild-type β-glucanase lost activity completely in 2 h, the engineered cellulase-M5 retained its initial activity, and β-glucanase-M7 displayed a 2.2-fold increase in its half-life. Structural analysis indicated that Pythia-identified mutations likely enhanced backbone robustness through refined polar and hydrophobic interactions, while beneficial mutations from ESM-2 appeared to affect polysaccharide-binding regions. This two-step computational redesign offers a promising approach for optimizing both thermostability and activity in glycoside hydrolases and other enzyme families with extensive sequence diversity.
Competing Interests: Declaration of competing interest The authors declare no competing financial interests or personal relationships that could have influenced the work reported in this paper.
(Copyright © 2024. Published by Elsevier B.V.)
Databáze: MEDLINE
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