Design And Construction Of Saccharomyces Cerevisiae For Taxifolin Production

Flavonoids are a large class of natural products synthesized by successive multi-step biochemical reactions based on phenol amino acids precursors.They have important functions such as anti-oxidation,anti-tumor,anti-viral,anti-bacterial and anti-inflammatory properties,offering beneficial values in fields of food and medicine.At present,flavonoids are mainly extracted from plants and this process is usually complicated,detrimental to environment and low-yield.Recent years synthetic biology shows power in engineering microbial chassis to produce natural products and also offers opportunity for production of flavonoids by similar approach.In this study,two key nodal compounds in the synthesis pathway of flavonoids,naringenin and taxifolin,were selected as representatives.Their respective biosynthetic pathways were constructed in Saccharomyces cerevisiae strains.For naringenin,the effects of two combinations of multiple genes on naringenin production were researched.The optimal pathway led to production of 41.22 mg/L naringenin.Further,the upstream module for biosynthesis of phenol amino acids were distributed to an engineered E.coli.Using L-tyrosine as intermediate,yeast cells could merely operate downstream heterologous modules.Based on optimization of co-culture conditions,a condition of initial OD_{600 nm} of 7,addition of 0.5 g/L L-tyrosine and 40 g/L xylose was used,getting production of naringenin as 103.3 mg/L.For the synthesis of taxifolin from naringenin as precursor,different flavanone-3-hydroxylase?F3H?genes derived from four species were combined with the same flavonoid 3'-hydroxylase gene?F3'H?and cytochrome P450 reductase gene?CPR?,and these combinations were tested in yeast cells.After test,we selected the optimal combination where the F3H from Camellia sinensis participated and achieved production of taxifolin as 5.18 mg/L.Meanwhile,we also found that the efficiency of collaboration of heterologous upstream and downstream modules still required improvement,considering the downstream module still did not work well.This study mainly focused on synthesis of flavonoids in different synthetic biological systems,namely engineered single yeast or co-culture system consisting of engineered yeast and E.coli.Our work made a primary trial to investigate an optimal mode of metabolic division and collaboration for long pathway of production synthesis.