Li Xinjia, Yu Haoran, Liu Shengli, Ma Baodi, Wu Xiaomei, Zheng Xuesong, Xu Yi
School of Chemical and Environmental Engineering, Shanghai Institute of Technology, 100 Haiquan Road, Shanghai, 201418, China.
Xianghu Laboratory, Hangzhou, 311231, China.
Bioresour Bioprocess. 2024 Jun 16;11(1):59. doi: 10.1186/s40643-024-00776-2.
Esterases are crucial biocatalysts in chiral compound synthesis. Herein, a novel esterase EstSIT01 belonging to family V was identified from Microbacterium chocolatum SIT101 through genome mining and phylogenetic analysis. EstSIT01 demonstrated remarkable efficiency in asymmetrically hydrolyzing meso-dimethyl ester [Dimethyl cis-1,3-Dibenzyl-2-imidazolidine-4,5-dicarboxyate], producing over 99% yield and 99% enantiomeric excess (e.e.) for (4S, 5R)-monomethyl ester, a crucial chiral intermediate during the synthesis of d-biotin. Notably, the recombinant E. coli expressing EstSIT01 exhibited over 40-fold higher activity than that of the wild strain. EstSIT01 displays a preference for short-chain p-NP esters. The optimal temperature and pH were 45 °C and 10.0, with K and k values of 0.147 mmol/L and 5.808 s, respectively. Molecular docking and MD simulations suggest that the high stereoselectivity for meso-diester may attribute to the narrow entrance tunnel and unique binding pocket structure. Collectively, EstSIT01 holds great potential for preparing chiral carboxylic acids and esters.
酯酶是手性化合物合成中的关键生物催化剂。在此，通过基因组挖掘和系统发育分析，从巧克力色微杆菌SIT101中鉴定出一种属于V家族的新型酯酶EstSIT01。EstSIT01在不对称水解内消旋二甲基酯[顺式-1,3-二苄基-2-咪唑烷-4,5-二羧酸二甲酯]方面表现出显著效率，(4S, 5R)-单甲酯的产率超过99%，对映体过量(e.e.)超过99%，(4S, 5R)-单甲酯是合成d-生物素过程中的关键手性中间体。值得注意的是，表达EstSIT01的重组大肠杆菌的活性比野生菌株高40多倍。EstSIT01对短链对硝基苯酯表现出偏好。最佳温度和pH分别为45℃和10.0，K和k值分别为0.147 mmol/L和5.808 s⁻¹。分子对接和分子动力学模拟表明，对内消旋二酯的高立体选择性可能归因于狭窄的入口通道和独特的结合口袋结构。总体而言，EstSIT01在制备手性羧酸和酯方面具有巨大潜力。
