Improved xylose utilization and ethanol fermentation performance with ultracentrifugation-assisted adaptive evolution of Saccharomyces cerevisiae.

Saccharomyces cerevisiae is the most widely used microorganism for industrial ethanol fermentation, but its limited xylose assimilation capacity remains a major bottleneck for efficient cellulosic ethanol production. In this study, repeated ultracentrifugation was incorporated as a physical perturbation into an adaptive evolution protocol to obtain the evolved strain S. cerevisiae X100. Compared with the parental strain, the respective utilization rates increased from 0.35g/L/h to 0.80g/L/h for xylose and from 4.67g/L/h to 7.77g/L/h for glucose, corresponding to increases of 128.6% and 66.4%. After 48h of fermentation, the final ethanol titer increased from 48.14g/L in S. cerevisiae Z100 to 63.15g/L in S. cerevisiae X100, corresponding to a 31.2% increase. The fed-batch fermentation of S. cerevisiae X100 achieved 94.96g/L ethanol with a yield of 0.50g/g mixed sugars, while 95.66% of xylose was consumed. Transcriptomic and whole-genome resequencing analyses identified changes associated with stress adaptation, cellular recovery, and metabolic remodeling involving ethanol biosynthesis, NAD(H) metabolism, and mitochondrial function. In this study, repeated ultracentrifugation perturbation was incorporated into an adaptive evolution protocol to improve glucose and xylose utilization and ethanol fermentation performance in S. cerevisiae.