Engineering a Carbonyl Reductase to Simultaneously Increase Activity Toward Bulky Ketone and Isopropanol for Dynamic Kinetic Asymmetric Reduction via Enzymatic Hydrogen Transfer,ACS Catalysis

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Engineering a Carbonyl Reductase to Simultaneously Increase Activity Toward Bulky Ketone and Isopropanol for Dynamic Kinetic Asymmetric Reduction via Enzymatic Hydrogen Transfer
ACS Catalysis ( IF 11.3 ) Pub Date : 2023-07-15 , DOI: 10.1021/acscatal.3c01569
Hongliu Zhang ₁ , Xi Chen _{1,

2} , Tong Lv _{1,

2} , Qian Li ₁ , Weidong Liu _{1,

2} , Jinhui Feng _{1,

2} , Xiangtao Liu ₁ , Peiyuan Yao _{1,

2} , Qiaqing Wu _{1,

2} , Dunming Zhu _{1,

2}

Affiliation

It is very challenging to engineer a carbonyl reductase with simultaneously improved activity toward both a bulky ketone and isopropanol, enabling substrate-coupled NAD(P)H regeneration for the ketone reduction. In this study, a mutant M242F/Q245T of the carbonyl reductase from Sporobolomyces salmonicolor was identified to simultaneously enhance its activity toward both methyl 2-phthalimidomethyl-3-oxobutanoate (1a) and isopropanol and catalyze the dynamic kinetic asymmetric reduction of 1a to a single stereoisomer of the product by hydrogen transfer from isopropanol. Crystal structural and computational analysis suggested that it might be due to the synergic effect of 242F and 245T by reshaping the substrate entrance tunnel and enzyme active site. This study unprecedentedly showed that a carbonyl reductase could be engineered to increase its activity toward both bulky ketones and isopropanol, and the target chiral alcohol can be efficiently prepared via substrate-coupled cofactor regeneration.

中文翻译：

设计羰基还原酶以同时提高对大体积酮和异丙醇的活性，通过酶促氢转移实现动态动力学不对称还原

设计一种羰基还原酶，同时提高对大体积酮和异丙醇的活性，从而实现酮还原的底物偶联 NAD(P)H 再生，是非常具有挑战性的。在这项研究中，来自鲑鱼色孢子酵母的羰基还原酶突变体 M242F/Q245T被鉴定出能够同时增强其对 2-邻苯二甲酰亚胺甲基-3-氧代丁酸甲酯 ( 1a ) 和异丙醇的活性，并催化1a的动态不对称还原。通过从异丙醇进行氢转移而形成产物的单一立体异构体。晶体结构和计算分析表明，这可能是由于 242F 和 245T 通过重塑底物入口通道和酶活性位点的协同效应。这项研究前所未有地表明，可以对羰基还原酶进行改造，以提高其对大酮和异丙醇的活性，并且可以通过底物偶联辅因子再生来有效制备目标手性醇。

更新日期：2023-07-15

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