Construction Of Saccharomyces Cerevisiae To Biosynthesize Delta-tocotrienol

Vitamin E is an important fat-soluble antioxidant,which constitutes tocopherol and tocorienol.Recently,tocotrienol,especially delta-tocotrienol,has attracted extenisve attention due to its diverse medical activities,such as anticancer activity.Currently,about 80% of vitamin E products on the market come from chemical synthesis and 20% from plant extracts.With the rapid development and application of gene editing technology,the use of microorganisms to synthesize complex products is a feasible,promising and economic method to replace chemical synthesis and plant extracts.Saccharomyces cerevisiae is an advantageous chassis for the synthesis of naturally plant-derived products due to its eukaryotic transcriptional and translational advantages,strong function of genome recombination and food safety.In this paper,delta-tocotrienol was produced by yeast strain BY4742.Firstly,we introduced three essential heterologous genes(hpd from Pseudomonas putida KT2440,hpt from Synechocystis sp.pcc 6803 and vte1 from Arabidopsis thaliana)for the synthesis of delta-tocotrienol and obtained the recombiant strain VE01.Then,the production of delta-tocotrienol and its precursor geranylgeranyl diphosphate(GGPP)and homogentisic acid(HGA)was detected.The results indicated that the lack of GGPP restricted the synthesis of delta-tocotrienol.Based on that,we increased the supply of precursor GGPP by introducing two genes(thmg1 from Saccharomyces cerevisiae and ggppssa from Sulfolobus acidocaldarius)in the yeast strain VE01 and obtained the recombiant strian VE02,thereby increasing the yield of delta-tocotrienol(1.39 ± 0.01 mg/L).Finally,the response surface methodology(RSM)was used to optimize the fermentation medium of the target strain,and the yield of delta-tocotrienol reached 3.56 ± 0.25 mg/L,which was about2.6 times higher than the original medium without optimization.The fed-batch fermentation in the optimized medium in 2 L fermenter further improved the production titer of delta-tocotrienol to 4.10 ± 0.10 mg/L.This study established an heterologous pathway for delta-tocotrienol synthesis in Saccharomyces cerevisiae and successfully synthesized delta-tocotrienol by improving precursor supply.Finally,the production of delta-tocotrienol reached 4.10± 0.10 mg/L through the medium optimization.