Metabolic engineering of Escherichia coli for resveratrol production using food-grade D-xylose as a carbon source.

Resveratrol is a high-value polyphenolic compound widely utilized in nutraceutical, cosmetic, and pharmaceutical applications. However, most microbial production systems rely on glucose as the primary carbon source, which limits flexibility for integrating alternative and renewable feedstocks. In this study, we developed an engineered Escherichia coli platform to investigate resveratrol biosynthesis under xylose-supporting conditions, using food-grade D-xylose as the carbon source. A heterologous pathway consisting of Populus tomentosa 4-coumarate: CoA ligase (Pt4CL) and Arachis hypogaea stilbene synthase (AhSTS) was introduced to convert externally supplied p-coumaric acid (PCA) into resveratrol. To improve precursor availability, intracellular malonyl-CoA supply was enhanced by introducing matB and matC from Streptomyces coelicolor A3(2) and overexpressing the acetyl-CoA carboxylase complex (ACC) from E. coli. Xylose assimilation was further strengthened by expressing xylE, xylA, and xylB, while carbon catabolite repression was alleviated using CRISPR interference (CRISPRi) targeting the glucose transporter gene ptsG. Under shake-flask conditions, the engineered strain produced up to 23.9 mg/L resveratrol from food-grade D-xylose, accompanied by near-complete xylose consumption and 93%-94% precursor conversion. This corresponded to an overall fermentation yield of approximately 12.5 mg resveratrol per g xylose consumed. Similar titers (27 mg/L) were obtained in a 5-L bioreactor, indicating stable pathway performance under controlled fermentation conditions. Overall, these results show that E. coli can be engineered to support efficient precursor-to-product conversion under xylose-supported conditions, providing a useful proof-of-concept framework for integrating alternative carbon sources into microbial production platforms for aromatic compounds. One sentence summary An engineered Escherichia coli system integrates xylose utilization, malonyl-CoA pathway optimization, and CRISPRi regulation to support resveratrol biosynthesis under xylose-supported conditions.