高効率スチルベノイド生合成を可能にする統合動的制御と酵素共局在化戦略

Stilbenoids are plant-derived polyphenols with potent bioactivities, which makes them promising for pharmaceutical and nutraceutical use. Their production remains challenging due to their low natural abundance in plants and the inherent toxicity associated with chemical synthesis. Here, we developed an integrated strategy to engineer Escherichia coli for efficient biosynthesis of multiple stilbenoids. A p-coumaric acid-responsive CRISPRi system dynamically regulated malonyl-CoA allocation, balancing host metabolism and stilbenoid production. We systematically explored enzyme co-localization strategies and used structure-informed in silico analysis to support dual-enzyme fusion design. Orthogonal SpyTag/SpyCatcher and SnoopTag/SnoopCatcher systems enabled optimal intermediate channeling and significantly enhanced pathway flux. As a result, we achieved the highest reported titers of piceatannol (494.30 mg/L from glycerol, increased to 583.10 mg/L with l-tyrosine supplementation) and pterostilbene (563.89 mg/L from glycerol, reaching 1110.92 mg/L with l-tyrosine feeding). Techno-economic analysis indicated that combined optimization of metabolic regulation, enzyme co-localization, and process design contributes substantially to the overall feasibility of stilbenoid manufacturing. Altogether, this study demonstrates the potential of stilbenoid biosynthesis and provides a valuable paradigm for the efficient synthesis of other complex natural products.