RewiringSaccharomyces cerevisiaemetabolism for optimised Taxol® precursors production

Saccharomyces cerevisiae has been recognised as a convenient host for the production of early precursors to the Taxol® anticancer drug. Recent studies have highlighted the harmful impact of oxidative stress as a result of the activity of Taxol® first cytochrome P450-reductase enzymes ( Taxus spp. CYP725A4-POR). Here, we evolved a new oxidative stress-tolerant yeast strain on galactose, which led to a three-fold higher titre of the CYP725A4 enzyme substrate, taxadiene. We comprehensively analysed the performance of the evolved and parent strain in galactose-limited chemostat cultures before and during oxidative stress induction. Integrating the transcriptomics and metabolite profiling data in an enzyme-constrained genome scale model enabled a more accurate prediction of changes that occurred to biological pathways as a response to/consequence of evolution and oxidative stress. The analyses showed a better performance of the evolved strain with improved respiration and reduced overflow metabolites production. The strain was robust to re-introduction of the oxidative stress, potentially due to the cross-protection mechanism, which contributed to likely better heme, flavin and NADPH availability for an optimal expression of CYP725A4 and POR in yeast. The increased level of taxadiene production has potentially occurred due to the antioxidant properties of taxadiene or as a mechanism to overcome the toxicity of geranylgeranyl diphosphate, the precursor to taxadiene synthase. Highlights The antioxidant properties of taxadiene promotes its production in Saccharomyces cerevisiae S. cerevisiae ALE on H 2 O 2 and galactose regulates Flavin, iron and NADPH metabolism as well as carbon and protein recycling pathways through cross-protection and anticipation mechanisms Abstract Figure Figure 1. Graphical abstract of the study. Figure was created with BioRender.com.