Engineering ω-transaminase by random mutagenesis and semi-rational design for the synthesis of (R)-(+)-1-(1-naphthyl)ethylamine
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National Natural Science Foundation of China (Nos. 31871738, 21776112), National Key Research and Development Program (No. 2018YFA0901700), China Postdoctoral Science Foundation (No. 2017M621631).

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    Abstract:

    (R)-(+)-1-(1-naphthyl)ethylamine is a key chiral intermediate for the synthesis of calcimimetic drug cinacalcet hydrochloride. ω-Transaminase has been considered to be potential for producing (R)-(+)-1-(1-naphthyl)ethylamine by asymmetric reduction of 1-acetonaphthone. Here, ω-transaminase from Arthrobacter sp. was engineered by combinatorial strategies of random mutagenesis and semi-rational design. Variants F225M, C281I, F225M/C281I with improved catalytic efficiency and thermostability were obtained. Compared with WT, variant F225M/C281I showed 85% increased kcat, 56% decreased Km and 3.42-fold kcat/Km. Furthermore, 22% higher conversion rate was achieved by F225M/C281I at 10 mmol/L 1-acetonaphthone after 24 h. Based on molecular docking and molecular dynamics simulation, improved catalytic efficiency of F225M/C281I could be attributed to its increased Pi-Pi T-shaped interaction with substrate 1-acetonaphthone. Additionally, a slightly higher half-life of F225M/C281I was validated by its lower root-mean-square fluctuation (RMSF) value of loop 134–139 compared with WT.

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曹旭东,韩瑞枝,方红辉,倪晔. 随机-半理性组合突变改造ω-转氨酶催化合成(R)-1-(1-萘基)乙胺[J]. Chinese Journal of Biotechnology, 2020, 36(9): 1828-1837

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  • Received:January 15,2020
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  • Online: September 25,2020
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