Engineered Biosynthesis of Phenol Using Acetate as the Carbon Source in <i>Escherichia coli</i>

Acetate is a prospective nonfood carbon source generated from one-carbon feedstocks and lignocellulose in biomanufacturing. This study aimed to efficiently biosynthesize phenol from acetate using engineered Escherichia coli. To achieve this sustainable route, we first constructed ten engineered E. coli strains by introducing the heterogeneous phenol synthesis pathway and enhancing the acetate assimilation pathway, with the strain BL9 exhibiting optimal phenol production. Subsequently, five metabolic engineering strategies were implemented for boosting phenol production, including activating the glyoxylate cycle by deleting iclR, relieving the repression of TyrR, replacing the bottleneck enzyme with AroGfbr, identifying and reinforcing rate-limiting steps in the shikimate pathway, and constructing a precursor phosphoenolpyruvate recycling module. These strategies resulted in a boost in the phenol titer of the engineered strain TPL6 to 0.454 g/L. Finally, a systematic optimization of the fermentation conditions was achieved. Through fed-batch fermentation combined with in situ extraction, the phenol titer was raised to 2.01 g/L, 38 times over the initial flask production. This is the first study to biosynthesize phenol from acetate, and it highlights the great potential of metabolically engineered E. coli.