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Published on: October 24, 2016
Rational Modification of Carbonyl Reductase from Candida parapsilosis ATCC 7330 to Improve the Yield of 2-Phenethyl
Wei Ye1, Mengjun Chen1, Haotian Huang1
1Key Laboratory of Functional Yeast of China National Light Industry, Hubei Engineering Research Center for Bioenzyme Engineering Technology, College of Biological and Pharmaceutical Sciences, China Three Gorges University, Yichang 443002, China.
Abstract:
Carbonyl reductases (CRs) are a class of oxidoreductases that convert carbonyl groups into hydroxyl groups. The application of CRs in the refined chemical industry is hindered by an unclear catalytic mechanism and low catalytic activity. In this study, CpCR from Candida parapsilosis ATCC 7330 was engineered via error-prone PCR and semirational design. The T171F mutant exhibited 2-fold enhanced catalytic efficiency toward PAAH, with an extended t1/2 by 7.0 h and elevated T50 by 1 °C. Consequently, 2-phenethyl alcohol (2-PE) production achieved a yield of 91.31%, representing a 2.36-fold improvement over wtCpCR. Simultaneously, the P284D/P285W variant displayed 6.2-fold higher kcat/Km toward CPMK and increased T50 by 2 °C. (S)-(4-Chlorophenyl)(pyridin-2-yl)methanol (CPMA) synthesis using P284D/P285W reached 42.71% yield, which is 9.23-fold greater than that of wtCpCR. Molecular dynamics simulations provided mechanistic insights into enhanced catalytic efficiency. This study establishes rational strategies for CR evolution and advances the green biosynthesis of high-value chiral alcohols.
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