摘要
Tailoring of enzyme toward α-tetralones, a class of bulky-bulky ketones, is still a challenge. In this work, the mutants of carbonyl reductase BaSDR1 with improved catalytic performance toward α-tetralone 1 a were obtained by adjusting the steric hindrance and hydrophobicity of the residues that affect the approach of α-tetralone with the catalytic residues. The designed mutants also showed enhanced catalytic performance toward halogenated α-tetralones 2 a–6 a. Remarkably, the activity of the mutant Q237V/I291F toward 7-fluoro-α-tetralone 5 a was 16.3-fold higher than the wildtype enzyme with improved stereoselectivity (98.8 % ee). More notably, the mutants Q139S and Q139S/V187S exhibited decreased or reversed stereoselectivity toward α-tetralone 1 a, 5-bromo-α-tetralone 2 a, 7-fluoro-α-tetralone 5 a and 7-chloro-α-tetralone 6 a, while the relatively high ee values were obtained in the presence of 6-chloro-α-tetralone 3 a and 6-bromo-α-tetralone 4 a as substrates. Further analysis showed the larger size of the substrates was beneficial for the substrates binding to the active cavity with a more specific binding mode, which endows the reaction with higher stereoselectivity. Moreover, the recombinant E. coli expressing the variant Q237V/I291F successfully catalyzed the reduction of a high concentration 7-fluoro-α-tetralone 5 a. These results not only offered a potential tool for chiral α-tetralols, but also provided guiding information for the enzyme engineering toward bulky-bulky ketones.
源语言 | 英语 |
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页(从-至) | 4625-4633 |
页数 | 9 |
期刊 | ChemCatChem |
卷 | 13 |
期 | 21 |
DOI | |
出版状态 | 已出版 - 8 11月 2021 |