昆那霉素（kinamycin）属于非典型角蒽环聚酮化合物。研究表明目前已知的四类非典型角蒽环化合物均是从苯并蒽类中间体出发，经过复杂的氧化开环和重排反应合成。氧化开环及重排反应是此类化合物发生结构分化的关键节点。昆那霉素A环氧化和乙酰化以及B环重氮修饰在已知聚酮化合物中很少见，其生物合成过程也不清楚。本项目以Streptomyces ambofaciens中昆那霉素合成基因簇alp为研究对象，对其生物合成过程中的特殊后修饰反应展开研究，通过体内基因敲除、异源表达及生物转化、蛋白表达纯化、晶体结构及体外活性研究，深入剖析氧化开环的反应机制以及决定产物结构的关键因素，鉴定参与A环修饰及重氮基团形成的关键酶，以期阐明昆那霉素的生物合成途径。对这些特殊后修饰反应进行深入研究，不仅有助于我们理解天然产物的合成机理，也为合成生物学与酶催化有机合成提供新的工具，具有重要理论意义和潜在应用价值。

Kinamycins represent a series of atypical angucycline compounds produced by Streptomyces murayamaensis and Streptomyces ambofaciens, which contain unique post-PKS modifications including the formation of a five-membered B ring, a highly hydroxylated and acetylated A ring and a diazo group on the B ring. Atypical angucyclines are biosynthesized from benz[b]anthracene intermediates via oxidative ring-opening and rearrangement reactions. Four types of atypical angucyclines could be formed due to different post ring-opening and rearrangement reactions, thus these reactions are critical for the structural diversity of atypical angucyclines. Hydroxylation of A ring is uncommon in atypical angucyclines. Diazo group is rare in natural products, nothing is known about its biosynthesis. In this study, we will characterize the unique post-PKS tailoring reactions in kinamycin biosynthesis using the producer S. ambofaciens. These reactions will be studied by in vivo gene knock-out, heterologous gene expression and bioconversion experiments, the key enzymes will be further characterized by in vitro enzymatic assay and crystallography. Fast separation and identification techniques such as LC-MS will be used to analyze reaction intermediates or products. The insights gained from these studies will not only be helpful for our understanding about the biosynthesis of antibiotics, but also offer enzymes with novel functions for synthetic biology and enzyme-catalyzed organic synthesis.