Asymmetric synthesis of intermediate for (1R,2S)-ethyl 1-amino-2-vinylcyclopropanecarboxylate by desymmetrization using engineered esterase from Bacillus subtilis

(1R,2S)-Ethyl 1-amino-2-vinylcyclopropanecarboxylate (VCPA), is a key intermediate for anti-hepatitis C virus drugs. In this study, we developed an efficient manufacturing method of intermediate for (1R,2S)-VCPA by enzymatic desymmetrization of a malonate diester derivative. In synthesis scheme of V...

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Bibliographic Details
Published in:Journal of bioscience and bioengineering 2021-06, Vol.131 (6), p.599-604
Main Authors: Kawabata, Hiroshi, Miyake, Ryoma, Asada, Kuniko, Dekishima, Yasumasa, Miyaike, Mitsuko, Kato, Ryohei
Format: Article
Language:English
Subjects:
(1R,2S)-Ethyl 1-amino-2-vinylcyclopropanecarboxylate
(1S,2S)-1-(Ethoxycarbonyl)-2-vinylcyclopropanecarboxylic acid
Desymmetrization
Homology model
p-Nitrobenzyl esterase
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Summary:(1R,2S)-Ethyl 1-amino-2-vinylcyclopropanecarboxylate (VCPA), is a key intermediate for anti-hepatitis C virus drugs. In this study, we developed an efficient manufacturing method of intermediate for (1R,2S)-VCPA by enzymatic desymmetrization of a malonate diester derivative. In synthesis scheme of VCPA (1S,2S)-1-(ethoxycarbonyl)-2-vinylcyclopropanecarboxylic acid (VCPME) is the monoester intermediate, which is converted from 2-vinylcyclopropane-1,1-dicarboxylate diethyl ester (VCPDE). As a result of esterase screening for producing (1S,2S)-VCPME from VCPDE by enzymatic desymmetrization, p-nitrobenzyl esterase from Bacillus subtilis NBRC3027 (PNBE3027) showed high enantioselectivity (more than 90% e.e.). Based on the homology model of PNBE3027, a library of mutants with the substitution of L70, L270, L273, and L313 in substrate-binding pocket was created for improvement in enantioselectivity. (1S,2S)-VCPME produced by the best variant harboring L70D, L270Q, L273R, and L313M showed 98.9% e.e. of enanthiopurity. Furthermore, preparative scale production of (1S,2S)-VCPME using the quadruple mutant was achieved. Our investigations present a new efficient process for (1R,2S)-VCPA using esterase and diverse to be applied for the industrial scale production. [Display omitted]
ISSN:1389-1723
1347-4421
DOI:10.1016/j.jbiosc.2021.02.004