Miao Yingchun, Liu Jiao, Wang Xuanlin, Liu Bo, Liu Weifeng, Tao Yong
CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, State Key Laboratory of Microbial Resources, Institute of Microbiology Chinese Academy of Sciences Beijing China.
College of Life Science University of Chinese Academy of Sciences Beijing China.
mLife. 2022 Jun 19;1(2):171-182. doi: 10.1002/mlf2.12006. eCollection 2022 Jun.
Metabolic engineering to produce tricarboxylic acid (TCA) cycle-derived chemicals is usually associated with problems of low production yield and impaired cellular metabolism. In this work, we found that fatty acid (FA) feedstocks could enable high-yield production of TCA cycle-derived chemicals, while maintaining an efficient and balanced metabolic flux of the glyoxylate-TCA cycle, which is favorable for both product synthesis and cell growth. Here, we designed a novel synthetic pathway for production of β-alanine, an important TCA cycle-derived product, from FAs with a high theortecial yield of 1.391 g/g. By introducing , improving and knocking out , glyoxylate shunt was highly activated in FAs and the yield of β-alanine reached 0.71 g/g from FAs, much higher than from glucose. Blocking the TCA cycle at nodes could increase β-alanine yield in a flask cultivation, but severely reduced cell growth and FA utilization during fed-batch processes. Replenishing oxaloacetate by knocking out and recovering could restore the growth and lead to a titer of 35.57 g/l. After relieving the oxidative stress caused by FA metabolism, β-alanine production could reach 72.05 g/l with a maximum yield of 1.24 g/g, about 86% of the theoretical yield. Our study thus provides a promising strategy for the production of TCA cycle-derived chemicals.
用于生产三羧酸（TCA）循环衍生化学品的代谢工程通常伴随着产量低和细胞代谢受损的问题。在这项工作中，我们发现脂肪酸（FA）原料能够实现TCA循环衍生化学品的高产，同时维持乙醛酸-TCA循环高效且平衡的代谢通量，这对产物合成和细胞生长均有利。在此，我们设计了一条从脂肪酸生产β-丙氨酸（一种重要的TCA循环衍生产物）的新型合成途径，理论产率高达1.391 g/g。通过引入、改进和敲除，乙醛酸分流在脂肪酸中被高度激活，β-丙氨酸从脂肪酸中的产量达到0.71 g/g，远高于从葡萄糖中的产量。在摇瓶培养中，在节点处阻断TCA循环可提高β-丙氨酸产量，但在分批补料过程中会严重降低细胞生长和脂肪酸利用率。通过敲除和恢复来补充草酰乙酸可恢复生长并使滴度达到35.57 g/l。在缓解脂肪酸代谢引起的氧化应激后，β-丙氨酸产量可达72.05 g/l，最大产率为1.24 g/g，约为理论产率的86%。因此，我们的研究为生产TCA循环衍生化学品提供了一种有前景的策略。
