P450羟化酶的分子改造及其在烯丙位C-H键的不对称羟基化反应中的合成应用

烯丙位C-H键的不对称羟基化反应是合成手性烯丙醇最直接的途径。如何获得高活性、高选择性的P450羟化酶，用于烯丙位C-H键的选择性氧化反应，实现结构多样性手性烯丙醇的精准合成仍是一个具有挑战性的工作。前期研究结果表明，我们获得了S-立体选择性的羟化酶P450DA。因此，本项目拟通过对羟化酶P450DA进行分子改造，基于已建立的高通量筛选方法，快速获得具有高活性、更高S-立体选择性和立体选择性反转的突变酶，用于立体选择性互补的手性烯丙醇的合成，并初步探索P450羟化酶活性位点与催化活性和立体选择性之间的关系。本项目研究将为手性烯丙醇的多样性和立体专一性合成提供一条高效、绿色的途径，对拓展P450酶的催化功能和应用具有重要意义。

Asymmetric hydroxylation of allylic C-H bond is the most direct method to synthesize chiral allylic alcohols. How to obtain highly active and selective P450 hydroxylase for the selective oxidation of allylic C-H bond for precise synthesis of chiral allylic alcohols is still a challenging task. In our previous work, the S-stereoselective hydroxylase P450DA was obtained. Therefore, this project is based on evolution of the obtained hydroxylase P450DA and high-throughput screening method established, to obtain highly active and stereoselective enzyme mutant, and to synthesize stereoselectivity complementary chiral allylic alcohols. In the end, this project is also aimed to explore the relationship between structure and function of P450 hydroxylase. This research will provide an efficient and green method for synthesis of the diversive chiral allylic alcohols, which is of great significance for the study of the catalytic function and application of P450 enzyme.