Key Laboratory of Combinatorial Biosynthesis and Drug Discovery, Ministry of Education, and School of Pharmaceutical Sciences, Wuhan University, Wuhan, China.
College of Food Science and Engineering, Ocean University of China, Qingdao, China.
Appl Environ Microbiol. 2020 Jan 7;86(2). doi: 10.1128/AEM.01971-19.
Formycin A (FOR-A) and pyrazofurin A (PRF-A) are purine-related -nucleoside antibiotics in which ribose and a pyrazole-derived base are linked by a -glycosidic bond. However, the logic underlying the biosynthesis of these molecules has remained largely unexplored. Here, we report the discovery of the pathways for FOR-A and PRF-A biosynthesis from diverse actinobacteria and propose that their biosynthesis is likely initiated by a lysine -monooxygenase. Moreover, we show that and (involved in FOR-A and PRF-A biosynthesis, respectively) mutants are correspondingly capable of accumulating the unexpected pyrazole-related intermediates 4-amino-3,5-dicarboxypyrazole and 3,5-dicarboxy-4-oxo-4,5-dihydropyrazole. We also decipher the enzymatic mechanism of ForT/PrfT for -glycosidic bond formation in FOR-A/PRF-A biosynthesis. To our knowledge, ForT/PrfT represents an example of β-RFA-P (β-ribofuranosyl-aminobenzene 5'-phosphate) synthase-like enzymes governing -nucleoside scaffold construction in natural product biosynthesis. These data establish a foundation for combinatorial biosynthesis of related purine nucleoside antibiotics and also open the way for target-directed genome mining of PRF-A/FOR-A-related antibiotics. FOR-A and PRF-A are -nucleoside antibiotics known for their unusual chemical structures and remarkable biological activities. Deciphering the enzymatic mechanism for the construction of a -nucleoside scaffold during FOR-A/PRF-A biosynthesis will not only expand the biochemical repertoire for novel enzymatic reactions but also permit target-oriented genome mining of FOR-A/PRF-A-related -nucleoside antibiotics. Moreover, the availability of FOR-A/PRF-A biosynthetic gene clusters will pave the way for the rational generation of designer FOR-A/PRF-A derivatives with enhanced/selective bioactivity via synthetic biology strategies.
福霉素 A (FOR-A) 和吡嗪呋喃霉素 A (PRF-A) 是嘌呤相关的 -核苷抗生素，其中核糖和吡唑衍生碱基通过 -糖苷键连接。然而，这些分子的生物合成逻辑在很大程度上仍未得到探索。在这里，我们报告了从不同放线菌中发现 FOR-A 和 PRF-A 生物合成途径的情况，并提出它们的生物合成可能是由赖氨酸单加氧酶启动的。此外，我们还表明 和 (分别参与 FOR-A 和 PRF-A 生物合成) 突变体相应地能够积累意想不到的吡唑相关中间体 4-氨基-3,5-二羧酸吡唑和 3,5-二羧酸-4-氧代-4,5-二氢吡唑。我们还破译了 ForT/PrfT 在 FOR-A/PRF-A 生物合成中形成 -糖苷键的酶促机制。据我们所知，ForT/PrfT 代表了一类β-RFA-P（β-核糖呋喃基-氨基苯 5'-磷酸）合酶样酶，负责天然产物生物合成中 -核苷支架的构建。这些数据为相关嘌呤核苷抗生素的组合生物合成奠定了基础，也为 PRF-A/FOR-A 相关抗生素的靶向基因组挖掘开辟了道路。FOR-A 和 PRF-A 是具有不寻常化学结构和显著生物活性的 -核苷抗生素。阐明 FOR-A/PRF-A 生物合成过程中 -核苷支架构建的酶促机制不仅将扩展新型酶促反应的生化谱，还可以通过定向基因组挖掘来获得 FOR-A/PRF-A 相关 -核苷抗生素。此外，FOR-A/PRF-A 生物合成基因簇的可用性将为通过合成生物学策略合理生成具有增强/选择性生物活性的设计 FOR-A/PRF-A 衍生物铺平道路。
