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Engineering of Corynebacterium glutamicum for high-level γ-aminobutyric acid production from glycerol by dynamic metabolic control

谷氨酸棒杆菌 代谢工程 生物化学 焊剂(冶金) 代谢途径 合成生物学 通量平衡分析 代谢网络 生物 生物合成 甘油 代谢通量分析 柠檬酸循环 新陈代谢 化学 计算生物学 基因 有机化学
作者
Liang Wei,Jinhua Zhao,Yiran Wang,Jinshan Gao,Muhua Du,Yue Zhang,Ning Xu,Huanmin Du,Jiansong Ju,Qingdai Liu,Jun Liu
出处
期刊:Metabolic Engineering [Elsevier BV]
卷期号:69: 134-146 被引量:58
标识
DOI:10.1016/j.ymben.2021.11.010
摘要

Synthetic biology seeks to reprogram microbial cells for efficient production of value-added compounds from low-cost renewable substrates. A great challenge of chemicals biosynthesis is the competition between cell metabolism and target product synthesis for limited cellular resource. Dynamic regulation provides an effective strategy for fine-tuning metabolic flux to maximize chemicals production. In this work, we created a tunable growth phase-dependent autonomous bifunctional genetic switch (GABS) by coupling growth phase responsive promoters and degrons to dynamically redirect the carbon flux for metabolic state switching from cell growth mode to production mode, and achieved high-level GABA production from low-value glycerol in Corynebacterium glutamicum. A ribosome binding sites (RBS)-library-based pathway optimization strategy was firstly developed to reconstruct and optimize the glycerol utilization pathway in C. glutamicum, and the resulting strain CgGly2 displayed excellent glycerol utilization ability. Then, the initial GABA-producing strain was constructed by deleting the GABA degradation pathway and introducing an exogenous GABA synthetic pathway, which led to 5.26 g/L of GABA production from glycerol. In order to resolve the conflicts of carbon flux between cell growth and GABA production, we used the GABS to reconstruct the GABA synthetic metabolic network, in which the competitive modules of GABA biosynthesis, including the tricarboxylic acid (TCA) cycle module and the arginine biosynthesis module, were dynamically down-regulated while the synthetic modules were dynamically up-regulated after sufficient biomass accumulation. Finally, the resulting strain G7-1 accumulated 45.6 g/L of GABA with a yield of 0.4 g/g glycerol, which was the highest titer of GABA ever reported from low-value glycerol. Therefore, these results provide a promising technology to dynamically balance the metabolic flux for the efficient production of other high value-added chemicals from a low-value substrate in C. glutamicum.
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