人工Cu-TM1459金属酶催化不对称迈克尔加成反应

Artificial Cu-TM1459 metalloenzyme catalyzes asymmetric Michael addition reaction

不对称迈克尔加成反应是合成手性化合物的重要反应,手性的构建一般由传统手性金属络合物催化完成,催化剂结构复杂,制备困难。人工金属酶可以利用生物大分子替代过渡金属手性催化剂,成为研究的热点。使用TM1459蛋白质支架,在原有的金属结合基序上理性引入两个组氨酸和一个羧酸盐面部三联基序,配位Cu2+,制备了人工Cu-TM1459金属酶。将其用于催化不对称迈克尔加成反应研究,Cu-H52A/H58E变体金属酶具有中等反应活性和较高对映选择性(e.e.值达58%)。进一步通过分子对接和催化机理研究对金属结合位点附近关键残基进行定点突变,I108A/C106V/K24E突变体催化该反应,产率99%,e.e.值93%。

The asymmetric Michael addition reaction is an important reaction for synthesizing chiral compounds.Traditionally,chiral metal complexes are used as catalysts for chiral induction,but they are complex in structure and difficult to prepare.Artificial metalloenzymes,which utilize biomacromolecules to replace transition metal chiral catalysts,have become a research hotspot.In this study,two histidines and a carboxylate facial triple motif were rationally introduced on the original metal binding motif of TM1459 protein scaffold to coordinate Cu2+,and artificial Cu-TM1459 metalloenzyme was prepared.It was applied to catalyze the asymmetric Michael addition reaction,and the Cu-H52A/H58E variant metalloenzyme exhibited moderate reaction activity and high enantioselectivity(e.e.value up to 58%).Further studies on molecular docking and catalytic mechanism led to site-directed mutations of key residues near the metal binding site.The I108A/C106V/K24E mutant catalyzed the reaction with a yield of 99%and an e.e.value of 93%.