Departamento de Biología Funcional, Universidad de Oviedo, Oviedo, Spain.
Instituto Universitario de Oncología del Principado de Asturias, Universidad de Oviedo, Oviedo, Spain.
Appl Environ Microbiol. 2020 Jan 21;86(3). doi: 10.1128/AEM.02453-19.
The appearance of new infectious diseases, the increase in multidrug-resistant bacteria, and the need for more effective chemotherapeutic agents have oriented the interests of researchers toward the search for metabolites with novel or improved bioactivities. Sipanmycins are disaccharyl glycosylated macrolactams that exert antibiotic and cytotoxic activities. By applying combinatorial biosynthesis and mutasynthesis approaches, we have generated eight new members of the sipanmycin family. The introduction of plasmids harboring genes responsible for the biosynthesis of several deoxysugars into sipanmycin-producing sp. strain CS149 led to the production of six derivatives with altered glycosylation patterns. After structural elucidation of these new metabolites, we conclude that some of these sugars are the result of the combination of the enzymatic machinery encoded by the introduced plasmids and the native enzymes of the d-sipanose biosynthetic pathway of the wild-type CS149 strain. In addition, two analogues of the parental compounds with a modified polyketide backbone were generated by a mutasynthesis approach, feeding cultures of a mutant strain defective in sipanmycin biosynthesis with 3-aminopentanoic acid. The generation of new sipanmycin analogues shown in this work relied on the substrate flexibility of key enzymes involved in sipanmycin biosynthesis, particularly the glycosyltransferase pair SipS9/SipS14 and enzymes SipL3, SipL1, SipL7, and SipL2, which are involved in the incorporation of the polyketide synthase starting unit. Combinatorial biosynthesis has proved its usefulness in generating derivatives of already known compounds with novel or improved pharmacological properties. Sipanmycins are a family of glycosylated macrolactams produced by sp. strain CS149, whose antiproliferative activity is dependent on the sugar moieties attached to the aglycone. In this work, we report the generation of several sipanmycin analogues with different deoxysugars, showing the high degree of flexibility exerted by the glycosyltransferase machinery with respect to the recognition of diverse nucleotide-activated sugars. In addition, modifications in the macrolactam ring were introduced by mutasynthesis approaches, indicating that the enzymes involved in incorporating the starter unit have a moderate ability to introduce different types of β-amino acids. In conclusion, we have proved the substrate flexibility of key enzymes involved in sipanmycin biosynthesis, especially the glycosyltransferases, which can be exploited in future experiments.
新传染病的出现、多药耐药菌的增加以及对更有效化疗药物的需求，使研究人员的兴趣转向寻找具有新型或改善的生物活性的代谢物。Sipanmycins 是一种双糖基化的大环内酯类化合物，具有抗生素和细胞毒性活性。通过应用组合生物合成和突变合成方法，我们生成了 sipanmycin 家族的八个新成员。将负责几种去糖基化糖生物合成的质粒引入 sipanmycin 产生菌 sp. CS149 菌株中，导致了六种糖基化模式改变的衍生物的产生。这些新代谢物的结构阐明后，我们得出结论，其中一些糖是引入质粒编码的酶机制与野生型 CS149 菌株的 d-sipanose 生物合成途径的天然酶相结合的结果。此外，通过突变合成方法，用 3-氨基戊酸喂养突变菌株的培养物，生成了两种母体化合物的类似物，其多酮骨架发生了修饰。本工作中生成的新 sipanmycin 类似物依赖于 sipanmycin 生物合成中关键酶的底物灵活性，特别是糖基转移酶对 SipS9/SipS14 和酶 SipL3、SipL1、SipL7 和 SipL2，它们参与聚酮合酶起始单元的掺入。组合生物合成已证明其在生成具有新型或改善的药理特性的已知化合物衍生物方面的有用性。Sipanmycins 是由 sp. CS149 菌株产生的糖基化大环内酯类化合物家族，其增殖活性取决于连接在糖苷上的糖部分。在本工作中，我们报告了几种具有不同去糖基化糖的 sipanmycin 类似物的生成，表明糖基转移酶对不同核苷酸激活糖的识别具有高度的灵活性。此外，通过突变合成方法引入了大环内酯环的修饰，表明参与起始单元掺入的酶具有引入不同类型β-氨基酸的中等能力。总之，我们已经证明了 sipanmycin 生物合成中关键酶的底物灵活性，特别是糖基转移酶，这可以在未来的实验中得到利用。
