Zhao Houyuan, Wang Liang, Wan Dan, Qi Jianzhao, Gong Rong, Deng Zixin, Chen Wenqing
Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education, School of Pharmaceutical Sciences, Wuhan University, Wuhan, 430071, China.
State Key Laboratory of Microbial Metabolism, School of Life Sciences & Biotechnology, Shanghai Jiao Tong University, Shanghai, 200030, China.
Microb Cell Fact. 2016 Sep 21;15(1):160. doi: 10.1186/s12934-016-0559-7.
Aurantimycin (ATM), produced by Streptomyces aurantiacus JA 4570, is a potent antimicrobial and antitumor antibiotic. Although the chemical structure of ATM is highly distinctive and features a cyclohexadepsipeptide scaffold attached with a C acyl side chain, little is known about its biosynthetic pathway and regulatory mechanism.
In this work, we report the identification and characterization of the ATM biosynthetic gene cluster from S. aurantiacus JA 4570. Targeted inactivation of artG, coding for a NRPS enzyme, completely abolished ATM production, thereof demonstrating the target gene cluster (art) is responsible for ATM biosynthesis. Moreover, four NRPS adenylation (A) domains including a freestanding enzyme ArtC have been characterized in vitro, whose substrate specificities are consistent with in silico analysis. Further genetic analysis of the two regulatory genes artB and artX unambiguously suggested both of them play positive roles in ATM biosynthesis, and ATM-A production was thus rationally enhanced to about 2.5 fold via tandem overexpression of artB and artX in S. aurantiacus JA 4570.
These results will provide the basis for the understanding of precise mechanisms for ATM biosynthesis, and open the way for both rational construction of high-production ATM producer and orient-directed generation of designer ATM derivatives via synthetic biology strategies.
由橙色链霉菌JA 4570产生的橙霉素（Aurantimycin，ATM）是一种强效抗菌和抗肿瘤抗生素。尽管ATM的化学结构高度独特，具有连接C酰基侧链的环己缩肽支架，但其生物合成途径和调控机制却鲜为人知。
在本研究中，我们报道了从橙色链霉菌JA 4570中鉴定和表征ATM生物合成基因簇。编码非核糖体肽合成酶（NRPS）的artG基因的靶向失活完全消除了ATM的产生，从而证明目标基因簇（art）负责ATM的生物合成。此外，已在体外表征了四个NRPS腺苷化（A）结构域，包括一种独立酶ArtC，其底物特异性与计算机分析一致。对两个调控基因artB和artX的进一步遗传分析明确表明它们在ATM生物合成中均起积极作用，因此通过在橙色链霉菌JA 4570中串联过表达artB和artX，ATM-A的产量合理提高至约2.5倍。
这些结果将为理解ATM生物合成的精确机制提供基础，并为通过合成生物学策略合理构建高产ATM生产者以及定向生成设计型ATM衍生物开辟道路。
