Reinforced NADPH regeneration in engineered Saccharomyces cerevisiae enhances rosmarinic acid production

Rosmarinic acid (RA) is a valuable natural product for its significant antioxidative activity, which is mainly derived from plants or by chemical synthesis. With the development of biotechnology, the research on the production of RA by microbial cell factory has attracted more attention. In this study, we engineered Saccharomyces cerevisiae to produce RA by constructing a de novo RA synthesis pathway which utilized two cytochrome P450s from Salvia miltiorrhiza Bunge and Coleus scutellarioides (L.) Benth. Through reinforcing NAD(P)H regeneration by overexpression of zwf1 and integration of ARO4^K229L and ARO7^G141S into the genome, the engineered S. cerevisiae produced 4.92 mg/L of RA, 8.2-fold of the control, in shake flask fermentation. The titer of RA reached 11.3 mg/L by fed-batch fermentation in 5 L bioreactor. This study increased the production of RA by combination cofactor and pathway engineering, revealed the diversity of RA synthesis in S. cerevisiae, and also provided a reference for the synthesis and accumulation of other active components in yeast.