文献类型：期刊文献

英文题名：Colorimetric High-Throughput Screening Method for Directed Evolution of Prazole Sulfide Monooxygenase

作者：Liu, Feng[1];Geng, Qiang[1];Zhao, Chen[1];Ren, Shi-Miao[1];Yu, Hui-Lei[1];Xu, Jian-He[1]

机构：[1]East China Univ Sci & Technol, Shanghai Collaborat Innovat Ctr Biomfg, Sch Biotechnol, State Key Lab Bioreactor Engn, 130 Meilong Rd, Shanghai 200237, Peoples R China

年份：2022

卷号：23

期号：16

外文期刊名：CHEMBIOCHEM

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000817203400001)】；

基金：This work was financially supported by the National Key Research and Development Program of China (grant number 2021YFC2102900 & 2019YFA0905000), the National Natural Science Foundation of China (grant numbers 21922804 and 21871085), Program of Shanghai Academic Research Leader (21XD1400800), and the Fundamental Research Funds for the Central Universities (grant number 22221818014).

语种：英文

外文关键词：Baeyer-Villiger monooxygenase; biocatalysis; directed evolution; high-throughput screening; proton pump inhibitors

摘要：Baeyer-Villiger monooxygenases (BVMOs) are important biocatalysts for the enzymatic synthesis of chiral sulfoxides, including chiral sulfoxide-type proton pump inhibitors for the treatment of gastrointestinal diseases. However, native BVMOs are not yet suitable for practical application due to their unsatisfactory activity and thermostability. Although protein engineering approaches can help address these issues, few feasible high-throughput methods are available for the engineering of such enzymes. Herein, a colorimetric detection method to distinguish sulfoxides from sulfides and sulfones was developed for prazole sulfide monooxygenases. Directed evolution enabled by this method has identified a prazole sulfide monooxygenase CbBVMO variant with improved activity and thermostability that catalyzes the asymmetric oxidation of lansoprazole sulfide. A 71.3 % increase in conversion and 6 degrees C enhancement in the melting point were achieved compared with the wild-type enzyme. This new method is feasible for high-throughput screening of prazole sulfide monooxygenase variants with improved activity, thermostability, and/or substrate specificity.