菊科植物中倍半萜烯内酯二聚体(+)-ainsliadimer A生物合成途径中Diels-Alder环化反应的酶学机制研究

Diels-Alder环化反应是一类高效且能生成复杂多环结构化合物的反应，是生物合成的研究热点问题。(+)-Ainsliadimer A是来源于菊科植物并通过分子间D-A环化反应生成的倍半萜烯内酯二聚体，不仅结构新颖且生物活性良好，具有进一步开发抗癌、抗炎药物的潜能。因而，植物体内是如何通过酶催化实现该反应是阐明化合物(+)-ainsliadimer A生物合成途径的关键。基于此，本课题拟通过生物信息学和生物化学技术发现候选基因序列，异源表达候选蛋白验证功能，并利用同源建模以及分子对接技术初步阐明酶学机制。前期从桑悬浮培养细胞中成功的发现了首个来源于植物且催化分子间Diels-Alder环化酶，初步阐明了酶学机制，验证了理论以及技术的可行性。本项目将进一步发现菊科植物中的D-A环化酶，拓宽反应种类并加深我们对机制的认识，也为其他D-A环化酶关键基因的寻找和功能鉴定提供借鉴。

Diels–Alder (D–A) cyclisation is a highly efficient reaction that can produce complex multi-ring compounds and is a hotspot of research in the field of biosynthesis. (+)-Ainsliadimer A is a sesquiterpene lactone dimer derived from plants in the Compositae family and is produced via intermolecular D–A cyclisation. It exhibits a novel structure, good biological activity and potential to develop anticancer and anti-inflammatory drugs. Therefore, the mechanism through which this reaction is realised by enzymatic catalysis in plants is key to the elucidation of the biosynthetic pathway of (+)-ainsliadimer A. Therefore, this study intends to explore the sequence of candidate genes through bioinformatics and biochemical analyses, verify the function of candidate proteins with heterologous expression and preliminarily clarify the underlying enzymatic mechanism using homology modelling and molecular docking. In a previous study, the first intermolecular Diels–Alderase from plants and the related catalysis were successfully elucidated via a cell suspension culture of mulberry; the enzymatic mechanism was preliminarily clarified and the feasibility of the hypothesis and technology was verified. This study will further discover Diels–Alderase in Compositae plants, broaden the reaction types and deepen our understanding of the underlying mechanism, thus providing a reference for searching key genes of other Diels–Alderases and for verifying their function.