Stereoselective synthesis of (1R, 2S)-norephedrine by recombinant whole-cell biocatalysts coupling acetohydroxyacid synthase I and ω-transaminase
2018
Lee, Yen-Chung | Chen, Yih-Yuan | Lin, Jian-Sin | Chen, Yi-Wun | Li, Jiazhen | Liang, Kun-Xin | Chan, Hsin-Hua | Lin, Wei-De | Kao, Chao-Hung
In this study, a combined whole-cell biotransformation process was used for efficient synthesis of optically pure (1R, 2S)-norephedrine [(1R, 2S)-NE]. The genes encoding R-selective acetohydroxyacid synthase I (AHAS I) from Escherichia coli and S-selective ω-transaminase (ω-TA) from Chromobacterium violaceum BCRC10636 were cloned and over-expressed in E. coli NovaBlue cells. In the first biosynthetic step, l-phenylacetylcarbinol (l-PAC) was produced from benzaldehyde and pyruvate by using recombinant E. coli (pQE-AHAS I) cells, with almost 100% conversion yield and 71.8% purification yield. The purified l-PAC was coupled to l-alanine by using recombinant E. coli (pQE-CvTA) cells to produce (1R, 2S)-NE. This biocatalytic process was optimal at pH 6.5 to 8.0 and 37 °C, with a 1:10 ratio of l-PAC to l-alanine. Under the optimal conditions, the highest conversion yield of (1R, 2S)-NE was 62.2% and the enantiomeric excess value of (1R, 2S)-NE was more than 99%. The recombinant E. coli (pQE-CvTA) cells could be reused for at least 20 cycles, with only a modest reduction in the conversion yield to 76.3% relative to the first cycle. Our results indicate that the combination of AHAS I- and ω-TA-expressing E. coli cells might be a potential biocatalyst for (1R, 2S)-NE production.
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