源于酰基转移酶缺失聚酮合酶的天然产物生物合成及基因组挖掘研究

The collection of small molecules with various chemical scaffolds and functionality, representing huge chemical space, has played a crucial role in current drug discovery effort based on high-throughput, high-content and whole-organism screening. Natural products derived from acyltransferase-less polyketide synthases (ATlessPKS) have been found to modulate a variety of remarkable biological functions. There are about 30 known natural products from ATlessPKSs, and three of them, including the antibiotics streptogramin, mupirocin and anticancer drug FK228, are on the markets. Previous studies have shown that there are several unique features in ATlessPKSs, such as the stand-alone trans-acting acyltransferase, and the often associated β-alkylation pathways. We have hypothesized that the discovery of natural products from ATlessPKSs would benefit enormously if the biosynthetic mechanism of ATlessPKSs could be deciphered, and also be used in genome mining of this special class of compounds. The purpose of the current proposal is to learn the underlying mechanism, the interaction and the evolution relationship in the biosynthesis of anticancer antibiotic leinamycin and antibiotic mikamycin A, which are both derived from ATlessPKSs, and produced in Streptomyces atroolivaceous S-140. Several unique genes related to ATlessPKSs, including acyltransferase and HMG-CoA synhase will be used to survey the biosynthetic potential of our unique actinomycete collection, around 5,000 strains collected from Yunan province with one of the greatest biological diversity in the world. Strains with the biosynthetic potential to produce natural products derived from ATlessPKSs, will be used for microbial fermentation, as well as natural products isolation and structure elucidation. The proposed study should result a unique repertoire of natural products for the innovative drug discovery effort , in order to benefit our huge population of 1.3 billion.

源于酰基转移酶缺失聚酮合酶（acyltransferase-less polyketide synthase，ATlessPKS）的天然产物具有优异的生物活性，已有三个上市抗感染和抗肿瘤药物链霉杀阳菌素、假单胞菌酸和FK228。前期研究证实ATlessPKS拥有独立的酰基转移酶和广泛的β-烷基化修饰等特征，是一类非常独特的I型聚酮合酶。 本课题假设对源于ATlessPKS天然产物的生物合成和基因组挖掘的研究将加速这类天然产物的发现。本课题将利用链霉菌S. atroolivaceous S-140中存在的两个ATlessPKSs系统（分别生物合成leinamycin、mikamycin A），深入研究其生物合成机理、相互作用及进化关系；并进一步根据相关的酰基转移酶和羟甲基戊二酸单酰辅酶合酶等基因，对5000株放线菌菌株进行基因扫描，以针对性地进行天然产物分离和结构鉴定。

源于酰基转移酶缺失聚酮合酶（acyltransferase-less type I polyketide synthase，AT-less PKS）的天然产物具有优异的生物活性，已有三个上市抗感染和抗肿瘤药物链霉杀阳菌素、假单胞菌酸和FK228。前期研究证实AT-less PKS拥有独立的酰基转移酶和广泛的β-烷基化修饰等特征，是一类非常独特的I型聚酮合酶。本课题假设对源于AT-less type I PKS天然产物的生物合成和基因组挖掘的研究将加速这类天然产物的发现。项目按预定计划执行。.本课题首先通过生物信息分析，发现了一大类与AT-less PKSs 生物合成密切相关的羟甲基戊二酸单酰辅酶合酶（HCS）；设计并利用特异性克隆HCS的引物，通过对菌种库的基因组扫描，发现了13株具有HCS的菌株；通过全基因组测序，确定了菌株CB01881含有新颖的AT-less PKSs；通过高分辨质谱、紫外光谱等手段，确定了菌株CB02999产生AT-less type I PKSs对应的天然产物，包括一系列bacillaene及其衍生物，以及macrolactin；通过对18份土壤的HCS扫描发现了环境样品中存在大量的AT-less PKSs 特异性的HCS基因；进一步通过对其中的11份样品的酮基合酶的高通量测序，并通过与GenBank中的序列对比，确认了其对应AT-less type I PKSs的存在。上述研究，发现了在菌种库及环境样品中，存在大量的AT-less type I PKSs，这为下一步发现其对其对应的天然产物奠定了基础。