A lipase mutant and its use in the preparation of (s)-3-cyclohexene-1-carboxylic acid

By performing site-directed amino acid mutations on the lipase CALB, a recombinant lipase CALB mutant was constructed, solving the problem of low enzyme selectivity in existing technologies and realizing the industrial production of (S)-3-cyclohexene-1-carboxylic acid with high conversion rate and optical purity.

CN119876088BActive Publication Date: 2026-07-14ZHEJIANG UNIV OF TECH +1
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
ZHEJIANG UNIV OF TECH
Filing Date
2025-01-08
Publication Date
2026-07-14

AI Technical Summary

Technical Problem

Existing methods for preparing (S)-3-cyclohexene-1-carboxylic acid by bio-enzyme catalysis suffer from low enzyme selectivity and low catalytic efficiency, making them difficult to apply to industrial production. Furthermore, there is a lack of reports on the synthesis of (S)-3-cyclohexene-1-carboxylic acid catalyzed by the lipase CALB.

Method used

By site-directed amino acid mutation of Candida antarctica lipase B (CALB), a recombinant lipase CALB mutant was constructed. Its enantiomeric resolution ability of racemic 3-cyclohexene-1-carboxylic acid methyl ester was optimized. (S)-3-cyclohexene-1-carboxylic acid was prepared by fermentation and catalytic reaction using recombinant genetically engineered bacteria.

Benefits of technology

It improves the stereoselectivity and catalytic activity of CALB lipase, reduces production costs, and is suitable for industrial production. It enables the efficient preparation of (S)-3-cyclohexene-1-carboxylic acid with a conversion rate of up to 52% and an optical purity of up to 72%.

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Abstract

The application belongs to the field of biopharmacy and bioconversion, and particularly relates to a lipase mutant and application thereof in preparation of (S)-3-cyclohexene-1-carboxylic acid. The lipase CALB mutant of the application is obtained by adopting single-point mutation on position 281 of the amino acid sequence shown in SEQ ID NO. 1. The stereoselectivity of the mutant in catalyzing the reaction of producing (S)-3-cyclohexene-1-carboxylic acid from racemic 3-cyclohexene-1-carboxylic acid methyl ester is greatly improved compared with the wild type. Compared with the traditional chemical method for preparing (S)-3-cyclohexene-1-carboxylic acid, the lipase CALB mutant provided by the application has high stereoselectivity, mild reaction conditions, is friendly to the environment, and has reduced requirements on equipment, greatly reduces the production cost, and shows wide application prospect in industrial application.
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