Establishment of an Efficient Expression and Regulation System in Streptomyces for Economical and High-Level Production of the Natural Blue Pigment Indigoidine

Indigoidine, as a kind of natural blue pigment, is widely used in textiles, food, and pharmaceuticals and is mainly synthesized from l-glutamine via a condensation reaction by indigoidine synthetases, most of which originates from Streptomyces species. However, due to the complex metabolic switches of Streptomyces, most of the researchers choose to overexpress indigoidine synthetases in the heterologous host to achieve high-level production of indigoidine. Considering the advantages of low-cost culture medium and simple culture conditions during the large-scale culture of Streptomyces, here, an updated regulation system derived from the Streptomyces self-sustaining system, constructed in our previous study, was established for the highly efficient production of indigoidine in Streptomyces lividans TK24. The updated system was constructed via promoter mining and σhrdB expression optimization, and this system was applied to precisely and continuously regulate the expression of indigoidine synthetase IndC derived from Streptomyces albus J1704. Finally, the engineered strain was cultured with cheap industrial glycerol as a supplementary carbon source, and 14.3 and 46.27 g/L indigoidine could be achieved in a flask and a 4 L fermentor, respectively, reaching the highest level of microbial synthesis of indigoidine. This study will lay a foundation for the industrial application of Streptomyces cell factories to produce indigoidine.

靛苷素（Indigoidine）作为一类天然蓝色色素，被广泛应用于纺织、食品及医药领域，其主要由靛苷素合成酶（indigoidine synthetases）以L-谷氨酰胺为底物通过缩合反应合成，该类合成酶多数源自链霉菌属（Streptomyces）菌种。然而由于链霉菌的代谢调控机制复杂，多数研究者选择在异源宿主中过量表达靛苷素合成酶，以实现靛苷素的高水平生产。考虑到链霉菌大规模培养时培养基成本低廉、培养条件简便，本研究基于前期构建的链霉菌自主调控系统，开发了一套更新版调控体系，并在变铅青链霉菌TK24（Streptomyces lividans TK24）中实现了靛苷素的高效生产。该更新体系通过启动子挖掘及σ^HrdB表达优化构建而成，被用于精准且持续调控源自白色链霉菌J1704（Streptomyces albus J1704）的靛苷素合成酶IndC的表达。最终，工程菌株以廉价工业甘油作为补充碳源进行培养，在摇瓶及4L发酵罐中分别获得了14.3 g/L与46.27 g/L的靛苷素产量，达到了微生物合成靛苷素的最高水平。本研究为链霉菌细胞工厂工业化生产靛苷素奠定了坚实基础。