Feng Jiacheng, Ye Huiru, Lu Changxin, Pan Linyan, Chen Hanchi, Zhu Linjiang, Chen Xiaolong
Institute of Fermentation Engineering, Zhejiang University of Technology, Hangzhou, China.
College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, China.
Crit Rev Biotechnol. 2025 May;45(3):665-682. doi: 10.1080/07388551.2024.2382957. Epub 2024 Aug 12.
Ene-reductase (ER) has been widely applied for asymmetrical synthesis of chiral intermediates due to its substrate promiscuity, photoexcited reactivity, and excellent property with producing two chiral centers at a time. Natural ERs often exhibit the same stereoselectivity, and they need to be engineered for opposite configuration of chiral compounds. The hydrogenation process toward activated alkenes by ERs is composed of reductive half reaction and oxidative half reaction, which are dependent upon two cofactors NAD(P)H and flavin mononucleotide. The catalytic activity of ERs will be affected by the size of the substrate, the activating strength of the electron-withdrawing groups, redox potential of cofactors, and the loop flexibility around catalytic cavity. Currently, protein engineering to ERs has been successfully employed to enhance various catalytic properties, including photoexcited asymmetric synthesis. This review summarizes the approaches to reverse the stereoselectivity and enhance catalytic activity of ERs and new applications of the engineered ERs in photobiocatalytic asymmetric synthesis, besides the discussion with the existing molecular mechanisms of mutants regarding the improved catalytic performance.
烯还原酶（ER）因其底物宽泛性、光激发反应性以及一次生成两个手性中心的优异特性，已被广泛应用于手性中间体的不对称合成。天然的烯还原酶通常表现出相同的立体选择性，若要获得手性化合物的相反构型，则需要对其进行改造。烯还原酶对活化烯烃的氢化过程由还原半反应和氧化半反应组成，这两个过程依赖于两种辅因子NAD(P)H和黄素单核苷酸。烯还原酶的催化活性会受到底物大小、吸电子基团的活化强度、辅因子的氧化还原电位以及催化腔周围环的柔韧性的影响。目前，对烯还原酶进行蛋白质工程改造已成功用于增强其各种催化性能，包括光激发不对称合成。本综述总结了逆转烯还原酶立体选择性和增强其催化活性的方法，以及工程化烯还原酶在光生物催化不对称合成中的新应用，此外还讨论了现有突变体关于改善催化性能的分子机制。
