Carboxylic acid reductase-dependent biosynthesis of trans-anethole in engineered Escherichia coli

Trans-anethole stands as a significant natural compound, finding extensive application as a flavoring ingredient across the food, cosmetics, perfumery, and pharmaceutical sectors. Trans-anethole also offers a variety of positive health benefits to humans, including anti-cancer, anti-inflammatory, anti-diabetic, immunomodulatory, and neuroprotective effects. In this study, we constructed a de novo trans-anethole pathway in E. coli by exploring the reduction of p-coumaric acid to p-coumaryl alcohol through carboxylate reductase and endogenous aldehyde reductase. First, an engineered Escherichia coli was created that could biosynthesise trans-anethole from glucose by the co-expression of aroG^fbr, tyrA^fbr,TAL, CAR, Sfp, ATF1, LtPPS1 and AIMT1. Second, by increasing the availability of SAM, we further increased the production of trans-anethole to 125 mg/L. Subsequently, the genes for transketolase I tktA and phosphoenolpyruvate synthase ppsA were further overexpressed to increase the availability of erythrose-4-phosphate and phosphoenolpyruvate, resulting in an increase in trans-anethole production to 223 mg/L. Finally, by knocking out the pheA gene to block the competitive pathway, the titer of trans-anethole was increased to 356 mg/L.