海洋真菌CPA类多环吲哚生物碱的分子内双环化机制研究

天然多环吲哚生物碱具有高度多样的化学结构和生物活性。自然通过复杂的多酶催化机制可由单一前体化合物创造出数量人惊人的衍生物。我们推测自然同时也进化出了高度精炼的生物催化机制来完成复杂多环生物碱的环系构造化学。基于我们发现自海洋米曲霉的吲哚生物碱CPAs的结构类型，提出了其三种环系骨架均由CpaO酶单独催化完成的科学假设。即CpaO酶不仅能够高度精炼地催化分子内双环化反应构建复杂环系结构，还可通过自身的底物杂泛性保持了创造化学多样性的能力。为了验证这一假设，我们将通过开展体内基因功能验证、体外生化表征及催化机制探索等研究内容达到揭示CPA类化合物分子内双环化机制的目标。这对于阐明CPA不同环系骨架的来源、揭示其生合成途径中缺失的环节均具有直接的科学意义。而且，本研究还将为多环吲哚生物碱相关的仿生合成及合成生物学等研究中的环系构建化学提供更精炼的研究策略和生物催化工具。

Nature creates vast chemical diversity of polycyclic indole alkaloids from a single precursor using multiplex enzymatic machineries. We believe that besides the highly diverting and sophisticated strategy, nature also develops highly integrated and laconic biosynthetic machinery to generate polycyclic skeletons of indole alkaloids. Therefore, we isolated an array of polycyclic indole natural products from marine-derived microbes. Among them, we found that the cyclopiazonic acids (CPAs) produced by a sponge-associated Aspergillus oryzae HMP-F28 show intriguing skeletal divergence and cyclization chemistry. We propose that the three distinct ring systems are generated by an elegant intramolecular double cyclization catalyzed by only one enzyme, CpaO, a flavin dependent oxidoreductase. To test the hypothesis, we’ll fully characterized CpaO and its catalytic mechanism through in vivo gene inactivation, in vitro biochemical characterization, substrate promiscuity assays and structure biology as well. The significance of this study is especially profound regarding the discovery of a highly integrated biosynthetic machinery and biocatalyst for generation of complex ring system of indole alkaloids. This project will also reveal the critical missing link of intramolecular cyclization and the TEII shunt pathway in the CPA biosynthesis pathway. Nature’s highly integrated biosynthetic machinery for polycyclic scaffold construction is expecting to inspire future work in natural product discovery, biomimetic synthesis and synthetic biology.