文献类型：期刊文献

英文题名：Identification, Characterization, and Computer-Aided Rational Design of a Novel Thermophilic Esterase from Geobacillus subterraneus, and Application in the Synthesis of Cinnamyl Acetate

作者：Zhang, Jin[1];Lin, Lin[2,3];Wei, Wei[1];Wei, Dongzhi[1]

机构：[1]East China Univ Sci & Technol, Newworld Inst Biotechnol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[2]Shanghai Inst Technol, Sch Chem & Environm Engn, Shanghai 201418, Peoples R China;[3]Natl Engn Res Ctr Nanotechnol, Res Lab Funct Nanomat, Shanghai 200241, Peoples R China

年份：2024

卷号：196

期号：6

起止页码：3553

外文期刊名：APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY

收录：;EI(收录号：20233814742002);WOS:【SCI-EXPANDED(收录号:WOS:001068678700004)】；

基金：This research was supported by the Shanghai Natural Science Foundation (No. 20ZR1415400), the Shanghai outstanding technical leaders plan 19XD1431900, 19XD1431800, the National Natural Science Foundation of China (Grant No. 81830052, 81530053), and Shanghai Key Laboratory of Molecular Imaging (18DZ2260400).

语种：英文

外文关键词：Thermophilic esterase; Rational design; Thermal stability; ADAP strategy; Cinnamyl acetate

摘要：Investigation of a novel thermophilic esterase gene from Geobacillus subterraneus DSMZ 13552 indicated a high amino acid sequence similarity of 25.9% to a reported esterase from Geobacillus sp. A strategy that integrated computer-aided rational design tools was developed to select mutation sites. Six mutants were selected from four criteria based on the simulated saturation mutation (including 19 amino acid residues) results. Of these, the mutants Q78Y and G119A were found to retain 87% and 27% activity after incubation at 70 degrees C for 20 min, compared with the 19% activity for the wild type. Subsequently, a double-point mutant (Q78Y/G119A) was obtained and identified with optimal temperature increase from 65 to 70 degrees C and a 41.51% decrease in Km. The obtained T1/2 values of 42.2 min (70 degrees C) and 16.9 min (75 degrees C) for Q78Y/G119A showed increases of 340% and 412% compared with that in the wild type. Q78Y/G119A was then employed as a biocatalyst to synthesize cinnamyl acetate, for which the conversion rate reached 99.40% with 0.3 M cinnamyl alcohol at 60 degrees C. The results validated the enhanced enzymatic properties of the mutant and indicated better prospects for industrial application as compared to that in the wild type. This study reported a method by which an enzyme could evolve to achieve enhanced thermostability, thereby increasing its potential for industrial applications, which could also be expanded to other esterases.