Synthetic Carbon-Efficient Pathway and Programmable Microenvironment for Sustainable Caffeic Acid Production
Binbing Xu, Qinyu Guo, Qi Cui, Wanqing Wei, Wei Song, Cong Gao, Xiaomin Li, Guangjie Liang, Kaifang Liu, Guipeng Hu, Jing WuAbstract
Microbial production of phenolic acids offers a sustainable alternative to plant extraction, reducing carbon loss and environmental impact. Here, we present a multi-level engineering strategy in Escherichia coli integrating carbon-efficient pathways, enzyme optimization, and subcellular compartmentalization. A synthetic TktA–Rpe–tyrosinase (TRT) pathway increased the theoretical carbon yield from 0.5 to 0.6 mol CaA per mol of glucose. Structure-guided engineering of tyrosinase enhanced catalytic efficiency toward p-coumaric acid by 5.07-fold. Programmable alkaline condensates created intracellular microenvironments optimized for alkaliphilic enzymes. This approach enabled the highest reported space–time yield of 211 mg L–1 h–1 for CaA. This work demonstrates a sustainable, high-efficiency microbial production strategy that maximizes carbon utilization, enhances enzyme performance, and precisely controls intracellular reactions.