酿酒酵母积累黄酮类化合物过程中丙二酰辅酶A供给的动态调控

Flavonoids have extensive applications in food and pharmaceutical industries. Previous researches showed that the bottleneck for the improvement of flavonoids titer in microorganisms was the limited supply of the malonyl-CoA. Current strategies for enhancing the malonyl-CoA for the flavonoids production mainly depended on the stationary regulation. The stationary regulation based metabolic engineering strains require the additional of inducers and antibiotics, and extraordinary multi-phase fermentation processes, which could significantly increase the cost for the final products. In the current project, we want to screen a series of promoters, which could responded to the different concentration of typical flavonoids and their intermediates, to provide the essential units for the engineering Saccharomyces cerevisiae for the dynamic regulation system. Based on the responsible promoters, we would further establish a series of signal amplification systems, which could be used for the dynamic coupled regulation between intracellular malonyl-CoA concentration and cell growth. The results obtained from the current project could not only systematically demonstrate the intracellular regulation process of malonyl-CoA in S. cerevisiae, but could also provide theoretical and technical references for establishing autonomously dynamic regulation systems with the malonyl-CoA as the precursor.

黄酮类化合物在食品、医药等领域具有广泛应用。大量研究表明，微生物胞内丙二酰-CoA含量不足，是导致黄酮类化合物产量提升的瓶颈。目前对微生物积累丙二酰-CoA强化黄酮类化合物的研究策略主要依赖于静态调控策略。基于静态调控策略改造的代谢工程菌株，在发酵过程中通常需要添加诱导剂、抗生素，并进行精细的分阶段培养，极大的增加了生产过程的成本。本项目通过转录组学技术系统筛选酿酒酵母中可感应典型黄酮类化合物及其中间产物的反馈启动子，为改造酿酒酵母动态强化积累丙二酰-CoA提供合成生物学元件。利用筛选得到的系列反馈启动子构建黄酮类化合物感应信号放大系统，实现胞内丙二酰-CoA积累与细胞生长相偶联的动态调控模式。本项目的研究结果将系统解析酿酒酵母胞内丙二酰辅酶A的胞内调控过程，并为在真核微生物中建立自发动态调控过程强化以丙二酰辅酶A为前体的目标产物高效合成过程提供理论与技术参考。

黄酮类化合物是一类普遍存在于植物中的重要天然产物，具有众多重要的生理功能。在食品、药品和化妆品等领域具有重要的应用价值。绝大多数黄酮类化合物可以看作是以柚皮素或生松素为骨架进行的一系列酶催化修饰反应得到的。每合成一分子的黄酮类化合物需要消耗3分子的丙二酰辅酶A，大量消耗丙二酰辅酶A会干扰脂肪酸合成，严重影响细胞生长。而目前基于丙二酰辅酶A的动态调控，严格调控黄酮类产物合成和细胞生长的平衡仍缺乏了解。本项目拟通过动态调控丙二酰辅酶A的合成，通过设计多层次和多基因动态调节网络(DRN)来解决黄酮类化合物的合成和细胞生长之间的平衡，实现黄酮类化合物的高效合成。通过开发一种生长耦合的NCOMB(柚皮素-对香豆酸-丙二酰辅酶-A-平衡)的动态调节网络(DRN)，系统优化了柚皮素和对香豆酸响应的调节途径，用于实时控制细胞内丙二酰辅酶A的供应，阐明丙二酰辅酶A代谢与细胞关键表型、典型的黄酮类化合物(柚皮素)合成的协同调控机制。本项目的研究结果将为类似涉及丙二酰辅酶A衍生物生物合成的动态调控提供理论与技术参考。