Engineering a Bacillus subtilis esterase for selective hydrolysis of d, l-menthyl acetate in an organic solvent-free system. Issue 16 (3rd April 2023)

• Record Type: Journal Article
• Authors: Qiao, Jingjing; Yang, Duxia; Feng, Yingting; Wei, Wan; Liu, Xun; Zhang, Yinjun; Zheng, Jianyong; Ying, Xiangxian
• Is Part Of: RSC advances. Volume 13:Issue 16(2023)
• Page Start: 10468
• Page End: 10475
• Publication Date: 2023-04-03
• Subjects: Chemistry -- Periodicals; 540.5
• DOI: 10.1039/d3ra00490b
• Languages: English
• ISSNs: 2046-2069

Abstract

Abstract : Esterase/lipase-catalyzed selective hydrolysis of d, l -menthyl esters has become one of the promising approaches for producing l -menthol, one of the most important flavoring chemicals with extensive uses. Abstract : Esterase/lipase-catalyzed selective hydrolysis of d, l -menthyl esters has become one of the promising approaches for producing l -menthol, one of the most important flavoring chemicals with extensive uses. However, the activity and l -enantioselectivity of the biocatalyst are not sufficient for meeting the industrial requirements. Herein, a highly active para -nitrobenzyl esterase from Bacillus subtilis 168 (pnbA-BS) was cloned and then engineered to enhance its l -enantioselectivity. On the basis of the strategy tailoring the steric exclusion effect and structural flexibility of the region adjacent to the substrate, the substitution of Ala400 to Pro caused a remarkable improvement in the E value from 1.0 to 466.6. The variant A400P was purified and further confirmed with strict l -enantioselectivity in the selective hydrolysis of d, l -menthyl acetate, whereas the improved l -enantioselectivity caused decreased activity. To develop an efficient, easy-to-use, and green methodology, organic solvent was omitted and substrate constant feeding was integrated into the whole-cell catalyzed system. During the catalytic process, the selective hydrolysis of 1.0 M d, l -menthyl acetate in 14 h offered a conversion of 48.9%, e.e.p value of >99%, and space-time yield of 160.52 g (l d) −1 .