Enhanced Biosynthesis of Dihydromyricetin in <i>Saccharomyces cerevisiae</i> by Coexpression of Multiple Hydroxylases

Abstract

Dihydromyricetin (DHM) is a traditional plant-extracted flavonoid with some health benefits. This study aimed to metabolically engineer the strains for DHM bioproduction. Two strains of BK-11 and BQ-21 were integrated with flavonoid 3-hydroxylase (F3H) or both F3H and flavonoid 3'-hydroxylase (F3'H). The resulting strains have expressed the enzymes of <i>Gm</i>CPR and <i>Sl</i>F3'5'H, and then, the promoters of INO1p and TDH1p were used to enhance further the DHM production from naringenin in <i>Saccharomyces cerevisiae</i>. Through multiple-copy integration, 709.6 mg/L DHM was obtained by adding 2.5 g/L naringenin in a 5 L bioreactor, implying that the synergistic effect between F3'H and flavonoid 3'5'-hydroxylase is likely to promote the DHM production. An yield of 246.4 mg/L DHM was obtained from glucose by deleting genes for branch pathways and integrating <i>Ph</i>CHS, <i>Ms</i>CHI, <i>Pc</i>4CL, and <i>Fj</i>TAL. To our knowledge, this is the highest production reported for the <i>de novo</i> biosynthesis of DHM.