浅紫灰链霉菌CQ01819及其次级代谢产物的定向发掘

Targeted exploration of Streptomyces lavendulae subsp.CQ01819 and its secondary metabolites

【目的】采用特征次级代谢产物生物合成的保守功能基因探针,定向分离土壤中产生特征次级代谢产物的菌种资源,借助基因转录分析为导向的培养基优化方法,获得目标次生代谢产物。【方法】首先,根据5种特征次级代谢产物保守的合成功能基因设计简并引物,定向从土壤样品中筛选、分离并纯化菌株。然后,以RT-qPCR为指导开展目标产物的发酵培养基优化;最后,对菌株进行发酵,利用多种色谱技术分离纯化目标天然产物,并结合高分辨质谱与核磁共振等技术对所获得的化合物进行结构鉴定。【结果】从土壤中筛选得到了一株AHBA合酶基因和环氧化酶基因均为阳性的链霉菌菌株(编号为CQ01819),根据转录分析优化发酵培养基,最终从该菌株分离纯化得到了含有AHBA结构单元的丝裂霉素C、聚醚类抗生素莫能霉素A和缬吲霉素。【结论】本研究通过菌株的定向分离纯化,筛选得到了产生预期抗生素的浅紫灰链霉菌菌株CQ01819;基于RT-qPCR指导的发酵培养基优化,确定了菌株的发酵条件;获得发酵粗提物后,采用多种色谱整合技术和光谱分析策略,快速分离并鉴定了目标产物。该研究为目标菌种资源的定向筛选、菌株的发酵条件的快速优化和化合物的定向分离提供了较好的参考。

[Objective]To obtain targeted natural products and the bacterial producers based on characteristic biosynthesis genes and transcription level analysis.[Methods]First,we directionally mined the secondary metabolites producers by 5 pair of degenerate primers according to the conserved biosynthesis genes.Then RT-qPCR guided fermentation medium optimization was performed.Finally,the targeted natural products were isolated using different chromatography technologies,and their chemical structures were identified based on High Resolution Mass Spectrometry(HRMS)and Nuclear Magnetic Resonance(NMR).[Results]The AHBA synthase and epoxidase-encoding genes positive Streptomyces lavendulae subsp.CQ01819 was successfully obtained from soil.Targeted compounds purification led to discovering AHBA-derived mitomycin C,polyether antibiotic monensin A and valindolmycin.[Conclusion]Our findings provide a good example for efficiently discovering bacterial resource of expected natural products.