细胞工厂代谢转换机制及其智能生物制造

Mechanism of Metabolic Transition in Cell Factory and its Intelligent Bio-manufacturing

微生物新药的产业化和高效制造是国家重大需求。然而天然产物作为次级代谢产物并非微生物生存所必需,以生存为目标的自然演化使微生物只在营养困乏期或者应激反应时才精确调控少量产生。尽管大量次级代谢产物的生物合成途径都得到了阐明,但初级代谢和次级代谢之间的转换机制仍不清楚,已成为制约细胞工厂高效生物制造药物的瓶颈。建立合成生物学5M策略和活细胞代谢实时监测工具是解决该科学问题的关键。本文聚焦链霉菌聚酮类化合物代谢转换机制的研究和智能制造,利用开发的系列特异性监测NAD(H)、NADP(H)的遗传编码荧光探针,详细解析了链霉菌胞内三酰甘油(TAGs)的降解机制,通过设计符合原子经济性原则的产物生物合成途径、动态精细调控等策略大幅度提高几种聚酮类药物(actinorhodin,jadomycin B和oxytetracycline)产量,其中阿维菌素B1a的工业生产效价提高了50%以上,达到有史以来最高的效价。

Industrialization and efficient manufacturing of new microbial drugs is a major national interest.However,secondary metabolites are not indispensable for the survival evolution of those microorganisms.Only small amounts of natural products are tightly produced under stress or shortage of food.Pharmaceutically important polyketides such as avermectin are mainly produced as secondary metabolites during the stationary phase growth of Streptomyces species in bioreactors.The source of intracellular metabolites that are funneled into polyketide biosynthesis has proven elusive.We applied 5 M strategy and real time biosensors for living cell metabolism.The intracellular triacylglycerols(TAGs),which accumulates in primary metabolism,are degraded during stationary phase.This process could redirect the carbon flux from both intracellular TAGs and extracellular substrates into polyketide biosynthesis.We devised a strategy named‘dynamic degradation of TAG’(ddTAG)to mobilize the TAG pool and increase polyketide biosynthesis.Using dd TAG we increased the titers of actinorhodin,jadomycin B,oxytetracycline and avermectin B1a in Streptomyces coelicolor,Streptomyces venezuelae,Streptomyces rimosus and Streptomyces avermitilis.Application of dd TAG increased the titer of avermectin B1a by 50%in a 180 m3 industrial-scale fermentation,which is the highest titer ever reported.Our strategy provides new ideas for intelligent biomanufacturing of polyketides and other types of secondary metabolites.Series of work are well received and published in Nat Biotech(article),Nat Methods,Nucleic Acids Res,and etc.