Characterization of the aurantimycin biosynthetic gene cluster and enhancing its production by manipulating two pathway-specific activators in Streptomyces aurantiacus JA 4570

Houyuan Zhao, Liang Wang, Dan Wan, Jianzhao Qi, Rong Gong, Zixin Deng, Wenqing Chen

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 C14 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.