Improving the expression of taxadiene synthase to enhance the titer of taxadiene in Saccharomyces cerevisiae

Abstract

Taxadiene is an important precursor of paclitaxel. However, its low yield in eukaryotic systems limits its biosynthesis. This study found that the low yield of taxadiene is likely due to the degradation of expressed taxadiene synthase (TS) in Saccharomyces cerevisiae. The TS expression degradation problem was improved by the knockout of protease PRB1, and the yield of taxadiene was increased by 97% from 19.8 mg L⁻¹ to 39.2 mg L⁻¹. Furthermore, multi-copy integration of the TS gene and enhancement of the geranylgeranyl diphosphate (GGPP) precursor pathway increased the taxadiene titer to 282.4 mg L⁻¹ in a shake flask. Interestingly, enhancing TS expression also decreased the competitive synthesis of geranylgeraniol (GGOH). Finally, the taxadiene titer reached 878.5 mg L⁻¹ after optimising the fed-batch fermentation, which is the highest taxadiene titer reported for eukaryotic microbes. This study alleviated the problems associated with the expression and degradation of heterologous proteins and provided an efficient and green strategy to produce complex natural compounds.