发酵
基质(水族馆)
酶
化学
脱氢酶
蛋白质工程
生物化学
突变体
代谢工程
合理设计
立体化学
生物
基因
生态学
遗传学
作者
Dong Liu,Xinglong Wang,Lin Hua,Weizhu Zeng,Jianghua Li,Jingwen Zhou
标识
DOI:10.1021/acs.jafc.3c08365
摘要
The titer of the microbial fermentation products can be increased by enzyme engineering. l-Sorbosone dehydrogenase (SNDH) is a key enzyme in the production of 2-keto-l-gulonic acid (2-KLG), which is the precursor of vitamin C. Enhancing the activity of SNDH may have a positive impact on 2-KLG production. In this study, a computer-aided semirational design of SNDH was conducted. Based on the analysis of SNDH's substrate pocket and multiple sequence alignment, three modification strategies were established: (1) expanding the entrance of SNDH's substrate pocket, (2) engineering the residues within the substrate pocket, and (3) enhancing the electron transfer of SNDH. Finally, mutants S453A, L460V, and E471D were obtained, whose specific activity was increased by 20, 100, and 10%, respectively. In addition, the ability of Gluconobacter oxidans WSH-004 to synthesize 2-KLG was improved by eliminating H2O2. This study provides mutant enzymes and metabolic engineering strategies for the microbial-fermentation-based production of 2-KLG.
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