Enhanced astaxanthin production in yeast via combined mutagenesis and evolution

Astaxanthin exhibiting strong antioxidant activity, is widely used as food and animal feed additives as well as in cosmetics, pharmaceutical and nutraceuticals industries. Microbial production of astaxanthin has received increasing concerns in recent years. A combined strategy consisting of physical mutagenesis by atmospheric and room temperature plasma (ARTP) and adaptive evolution driven by H2O2, was established to expand disturbance in genome of the engineered Saccharomyces cerevisiae. A titer of 65.9 mg/L astaxanthin at flask level was obtained via multiple rounds of crossed ARTP and H2O2 treatment, which was increased nearly 4-fold compared with the starting strain. Moreover, a trend of decreasing intracellular reactive oxygen species (ROS) and increasing chronological lifespan (CLS) was observed in cells with improved titer of astaxanthin. Eventually, a highest reported titer of 404.78 mg/L astaxanthin in S. cerevisiae was achieved in 5-L fermenter by further optimization of fermentation conditions. Our study not only highlights the iterative cycling of the integrated strategy combined by ARTP and adaptive evolution tightly based on the natural properties of the targets provides an efficient route to boost product accumulation, but also demonstrates it can serve as a general strategy for other microbial cell factories.