Functional Characterisation of Adaptive Variants in Saccharomyces cerevisiae Evolved under Lignocellulosic Stress. A Proteomics Analysis

Abstract

Waste streams from lignocellulosic biomass are underutilised substrates that can be converted into value added compounds using yeast fermentation. In a previous study a strain of Saccharomyces cerevisiae with increased tolerance to lignocellulosic spruce hydrolysate was developed and mutated genes of interest were identified. This thesis aimed to investigate what functional changes lead to this increase in tolerance. Individual knockouts of the mutated genes were introduced in Wild Type S. cerevisiae (CEN.PK113-7D) to identify gene-specific effects. The strains proteomes were quantified at two stages of growth and compared to a Wild Type using label free quantitative proteomics. Gene Ontology of common biological processes between strains revealed largest differences in expression of proteins related to transmembrane transport, carbohydrate metabolic processes and fatty acid β-oxidation. Uncharacterised upregulated proteins in the strain with increased tolerance suggested a novel pathway potentially connected to stress resistance. The findings demonstrate that increased tolerance to lignocellulosic spruce hydrolysate is achieved through multiple complex proteome adjustments. The identified cellular adjustments indicate a cell level prioritisation towards detoxification of stress-inducing compounds, as well as uptake and utilisation of metabolic substrates. This thesis demonstrated that proteomics can be used as a functional screening tool to characterise promising adaptive variants.