Enhanced Heat Tolerance and Ethanol Production of Industrial Saccharomyces cerevisiae via Replacement of the CDC15 Gene

Background CDC15 is an important gene involved in mitotic exit and cytokinesis in S.cerevisiae . However, its additional functions remain unclear. Comparative genomics analysis between the fast-growing strain MC15 and the high-ethanol-producing strain MF01 revealed CDC15 as a significant sequence-divergent gene. To validate the hypothesis that replacing this gene could enhance MF01's growth rate, this study engineered the recombinant strain MF01- CDC15 (MT) by substituting the CDC15 in MF01 (WT) with that from MC15. Rapid growth is one of the desirable traits required for production strains. Achieving sufficient yeast cell density not only reduces contamination by contaminating microbes but also shortens the ethanol production cycle and improves production efficiency. Results The CDC15 mutation significantly improved cell viability on solid medium at 50–54°C. In liquid culture, the MT entered the stationary phase 8 h earlier than WT. Furthermore, the maximum cell densities achieved by MT at 30°C, 37°C and 41°C increased by 37%, 57%, 155%, respectively, compared to WT. Unexpectedly, CDC15 mutation caused ethanol yields to increase by 26.41% using sucrose and 29.56% using molasses as carbon sources at 30°C. Notably, MT cells maintained normal morphology at 41°C, whereas WT displayed cell adhesion. The intracellular trehalose content in MT was 4.8 times higher than that in WT at 41°C ( p  = 0.014). Homology modeling indicated that the Phe626→Leu626 mutation in Cdc15p of MT may contribute to structural stability. The combination of this structural change and the trehalose accumulation likely contributes to MT’s superior heat resistance. Conclusions This study provides more detailed information about the function of CDC15 and suggest some clues to efficiently improve the performance of industrial yeast strains.