High-Level De Novo Production of (2 S )-Eriodictyol in Yarrowia Lipolytica by Metabolic Pathway and NADPH Regeneration Engineering.

(2 S )-Eriodictyol, a polyphenolic flavonoid, has found widespread applications in health supplements and food additives. However, the limited availability of plant-derived (2 S )-eriodictyol cannot meet the market demand. Microbial production of (2 S )-eriodictyol faces challenges, including the low catalytic efficiency of flavone 3'-hydroxylase/cytochrome P450 reductase (F3'H/CPR), insufficient precursor supplementation, and inadequate NADPH regeneration. This study systematically engineered Yarrowia lipolytica for high-level (2 S )-eriodictyol production. In doing this, the expression of F3'H/CPR was balanced, and the supply of precursors was enhanced by relieving feedback inhibition of the shikimate pathway, promoting fatty acid β-oxidation, and increasing the copy number of synthetic pathway genes. These strategies, combined with NADPH regeneration, achieved an (2 S )-eriodictyol titer of 423.6 mg/L. Finally, in fed-batch fermentation, a remarkable 6.8 g/L (2 S )-eriodictyol was obtained, representing the highest de novo microbial titer reported to date and paving the way for industrial production.