Construction And Optimization Of Saccharomyces Cerevisiae For Vitamin C Biosynthese

In recent years,with the increasingly prominent role of vitamin C in cancer treatment and pain treatment,the global demand for vitamin C has further increased.At present,Lai’s method and two-step fermentation method are widely used in the industrial synthesis of vitamin C.They are difficult to break through such bottlenecks as high energy consumption,high pollution and difficult molecular operation In this study,the biosynthetic pathway of vitamin C was transferred to Saccharomyces cerevisiae cells with mature genetic manipulation technology,which endowed S.cerevisiae with the ability to synthesize vitamin C in order to improve the stability,efficiency and environmental friendliness of vitamin C production.Firstly,we attempted to introduce the genes encoding proteins of catalytic enzymes of the two-step fermentation pathway which has the highest yield and the most extensive industrial applications into S.cerevisiae cells,but found no synthesis of target products and additional consumption of substrates.Thereafter,the surface display vector p YD1 and GFP fusion technology were used for exploration.Unfortunately,S.cerevisiae failed to reappear the two-step fermentation pathway.Secondly,the catalytic reaction enzyme of plant synthesis of vitamin C lacked in S.cerevisiae were fused with GFP to determine wether they could be heterologous expressed in S.cerevisiae.Then,the heterologous genes were introduced into S.cerevisiae by the strategy of traditional heterogeneous expressions.3.58±0.43 mg/L vitamin C was detected and the construction of S.cerevisiae strain for synthesis of vitamin C was achieved.Finally,we used the inducible promoter and multi-copy plasmids to overexpress the single heterologous gene,and the multi-copy plasmids to overexpress the whole heterologous genes and fuse the key enzymes.We found that the above methods all could improve the production of vitamin C in different degrees.Then we explored one multi-strategy combination optimization method.In this study,a strain of S.cerevisiae with a yield of 19.47±0.44 mg/L was obtained,which is the highest production synthesized by S.cerevisiae reported so far.