Improving ethanol production and tolerance in Saccharomyces cerevisiae through evolutionary engineering strategy using 1-butanol stress

There are crucial factors in the bioethanol production process that affect production efficiency. Accumulation of ethanol during the fermentation process and the inhibitory effect on growth are inevitable. Increasing ethanol tolerance in Saccharomyces cerevisiae enhances survival and ultimately increases ethanol production. The evolutionary engineering approach is a promising strategy to improve the complex trait of ethanol tolerance in the Saccharomyces cerevisiae. The toxicity mechanism of short-chain alcohols on yeast are similar. In this study, the laboratory strain of Saccharomyces cerevisiae CEN PK113-7D was exposed to 1-butanol stress by evolutionary engineering strategy during a 144-days culture period, after which the specific growth rate (µ) of the evolved strain was boosted from 0.48 h-1 to 0.84 h-1. Increased stress tolerance of 1-butanol led to an increase in ethanol tolerance and also ethanol production in the evolved strain. Ethanol production improved from 68.50 g/L in the parent strain to 87.02 g/L in the evolved strain. The results of the whole genome sequencing of evolved strains and comparing it with the parent strain sequence revealed changes in the single nucleotide polymorphism (SNPs) of the genes involved in this trait. There were changes in genes such as PGM2, MTH1, TCB1, YAP1801, UBP2, FAB1, IAH1 and CIA1. These genes were related to intracellular transport and pathways involved in cytoplasmic membrane composition and structure, cell wall structure, glucose metabolism, and lipids metabolism. So, the significance of this set of genes in the enhancement of ethanol tolerance was reported for the first time.