结构导向的异戊烯基转移酶AtPT-1底物杂泛性研究及酶法催化合成应用

本研究组前期从土曲霉（Aspergillus terreus）基因组中克隆得到一个芳香类化合物异戊烯基转移酶基因AtPT-1。功能研究表明重组AtPT-1可以高效催化黄酮、香豆素、木脂素和吲哚等多种结构类型的芳香类化合物发生异戊烯基化反应。本项目拟在前期研究的基础上开展以下研究工作：（1）对AtPT-1的底物杂泛性进行深入分析；（2）在蛋白X-射线晶体结构信息的指导下，对AtPT-1的活性位点结合腔进行改造，构建具有广泛底物谱的AtPT-1酶突变体；（3）利用AtPT-1及其突变体催化芳香类化合物发生定向异戊烯基化反应，酶法合成天然/非天然的异戊烯基芳香类药物先导物。本研究将阐明AtPT-1的底物杂泛性与蛋白结构之间的关系，为芳香类化合物异戊烯基转移酶的结构及功能研究提供科学依据；并提供一种高效、环境友好地制备活性异戊烯基芳香类化合物及新颖结构先导化合的新方法，兼具理论及实际应用意义。

In previous study, an aromatic prenyltransferase gene AtPT-1 was cloned from the genome of Aspergillus terreus. Functional assays revealed that recombinant AtPT-1 showed obvious catalytic promiscuity since many kinds of aromatics such as flavonoids, coumarins, lignans and indole derivatives could be accepted as prenyl acceptors. On the basis of these results, this project aims to carry out the following investigations: (1) exhaustive analysis of the substrate promiscuity of AtPT-1; (2) structurally informed site-directed mutagenesis of AtPT-1's active binding pocket to further expand the substrate spectrum under the guidance of X-ray crystallographic structural information; (3) directed and promiscuous prenylation with AtPT-1 and its mutants to afford natural/non-natural active prenylated aromatics. This investigation could not only explore the structural basis for the catalytic promiscuity of AtPT-1 to offer useful hints to the structural-functionally research of aromatic prenylatransrases but also provide a new approach of usefully, effectively and environment-friendly preparation of active prenylated aromatics, with both theoretical and practical significance.