ω-转氨酶对映体识别的分子基础及其改造

项目名称： ω-转氨酶对映体识别的分子基础及其改造

项目编号： No.21476199

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 有机化学

项目作者： 杨立荣

作者单位： 浙江大学

项目金额： 90万元

中文摘要： 生物催化过程高效绿色，已成为当前热点研究领域之一。生物催化剂-酶是生物催化的核心。但是现有催化用酶在工业应用过程中，面对非天分子然底物和非天然环境，存在性能差、效率低、应用难等问题。问题的关键在于缺乏对酶催化过程分子识别机制的深入研究和解析，酶分子性能改造困难，来源有限。本项目拟选择来源于C. violaceum DSM3019的S-转氨酶作为研究对象，以丙氨酸和苯乙酮的转氨反应作为模型，基于晶体结构，采用分子动力学/量子力学方法研究ω-转氨酶和底物间相互作用和动态结构变化，在分子水平上解析转氨酶对映体识别机制和调控原理；利用ω-转氨酶对映体识别的混乱性，理性改造S-转氨酶，构建高选择性R-转氨酶；深入研究催化过程热/动力学，综合运用和系统评价底物工程、介质工程和多酶偶联在线去除副产物等技术手段，实现过程强化，改善反应效率。本项目研究不仅具有重要的理论价值，同时其应用前景也非常明确和广阔。

中文关键词： 生物催化；ω-转氨酶；对映体识别；分子机制；过程强化

英文摘要： Biocatalysis technology has become one of the hot research fields due to its advantages of high efficiency and green characteristic. The core of biocatalysis is the biocatalyst-enzyme. But the poor performance and low efficiency of the existing enzymes limited their applications in industrial process with non-natural substrate and environment. The key problem is that the molecular recognition mechanism of the enzyme catalysis process have not been resolved. It is very difficult to reform the performance of enzyme, thus the industrial enzyme source is limited. S-aminotransferase from C. violaceum DSM3019 was selected as the target biocatalyst and the transamination between alanine and acetophenone was selected as the model reaction. Based on the crystal structure, the interaction between the ω-aminotransferase and the substrate and their dynamic structure changes will be studied by using the methods of molecular dynamics/quantum mechanics. The molecular recognition mechanism and regulation principles of the enantioselectivity of ω-aminotransferase will be resolved at the molecular level. Employing the enantiomer recognition promiscuity of ω-aminotransferase, the construction of highly selective R-aminotransferase will be realized via rational transformation of S-aminotransferase. Based on the thermodynamics/kinetics studies, the substrate engineering, medium engineering and online removing of by-products via multiple-enzyme coupling process will be comprehensively used and systematically evaluated to intensify the process and improve the reaction efficiency. This project is not only a very important supplement to the existing theory of aminotransferase with significant theoretical value, but also has very clear and broad application prospects.

英文关键词： biocatalysis;ω-aminotransferase;enantiomeric recognition;molecular mechanism;process intensification