Characterization of the biosynthetic gene cluster of the polyene macrolide antibiotic reedsmycins from a marine-derived Streptomyces strain

Tingting Yao, Zengzhi Liu, Tong Li, Hui Zhang, Jing Liu, Huayue Li, Qian Che, Tianjiao Zhu, Dehai Li, Wenli Li

Polyene antibiotics are important as antifungal medicines albeit with serious side effects such as nephrotoxicity. Reedsmycin (RDM) A (1), produced by marine-derived Streptomyces youssoufiensis OUC6819, is a non-glycosylated polyene macrolide antibiotic with antifungal activity comparable to that of clinically used nystatin. To elucidate its biosynthetic machinery, herein, the rdm biosynthetic gene cluster was cloned and characterized. The rdm cluster is located within a 104 kb DNA region harboring 21 open reading frames (ORFs), among which 15 ORFs were designated as rdm genes. The assembly line for RDM A is proposed on the basis of module and domain analysis of the polyketide synthetases (PKSs) RdmGHIJ, which catalyze 16 rounds of decarboxylative condensation using malonyl-CoA as the starter unit (loading module), two methylmalonyl-CoA (module 1 and 2), and fourteen malonyl-CoA (module 3-16) as extender units successively. However, the predicted substrate specificity of AT0 in the loading module is methylmalonyl-CoA instead of malonyl-CoA. Interestingly, the rdm cluster contains a five-gene regulation system RdmACDEF, which is different from other reported polyene gene clusters. In vivo experiments demonstrated the XRE family regulator RdmA and the PAS/LuxR family regulator RdmF function in negative and positive manner, respectively. Notably, inactivation of rdmA and overexpression of rdmF led to increased production of RDM A by ~ 2.0-fold and ~ 2.5-fold, reaching yields of 155.3 ± 1.89 and 184.8 ± 9.93 mg/L, respectively. Biosynthesis of RDM A is accomplished on a linear assembly line catalyzed by Rdm PKSs harboring a unique AT0 under the control of a complex regulatory system. These findings enable generation of new biologically active RDM derivatives at high yield and with improved properties by engineered biosynthesis.