Metabolic engineering of Saccharomyces cerevisiae to improve farnesol production

Abstract:

[Objective] Farnesol (FOH, C₁₅H₂₆O) is a natural acyclic sesquiterpene alcohol with an aromatic odor. It is widely used in the industrial production of cosmetics and pharmaceuticals and also considered as an aviation jet-fuel substitute. Although food-grade S. cerevisiae can synthesize endogenous FOH, the yield is too low to meet the requirements for industrial production. Therefore, in this work, the FOH synthesis pathway in S. cerevisiae was metabolically engineered to overproduce FOH. [Methods] To improve the intracellular level of farnesyl pyrophosphate (FPP), the direct precursor for FOH synthesis, the mevalonate pathway in S. cerevisiae CEN.PK2-1D was enhanced via constitutive overexpression of genes encoding the two key enzymes whereas the ergosterol pathway was weakened by replacing the EGR9 promoter with the HXT1 promoter. In addition, five genes encoding endogenous phosphatases and two heterologous synthases that could catalyze FOH synthesis, were individually overexpressed to determine the enzyme with optimum performance. [Results] Shake flask culture experiments showed that constitutive overexpression of genes encoding the truncated HMG-CoA reductase (tHMGR1) and the FPP synthase ERG20 in S. cerevisiae CEN.PK2-1D increased the production of FOH by 50.8 fold to reach 5.08 mg/L. Down-regulation of the squalene synthase encoding gene ERG9, by replacing its promoter with the HXT1 promoter, further increased the FOH titer to 239.17 mg/L. On this basis, a maximum yield of FOH (393.13 mg/L) was obtained when the endogenous phosphatase PAH1 was overexpressed. [Conclusion] In this study, metabolic engineering of S. cerevisiae was used to increase the production titer of FOH to 393.13 mg/L. This is the highest reported yield of FOH from S. cerevisiae under shake flask culture conditions.