Shotgun proteomics of Starmerella bacillaris and S. cerevisiae in sequential fermentation with a focus on glycans biosynthesis and metabolism

Introduction: The use of Starmerella bacillaris in sequential fermentation with Saccharomyces cerevisiae has been previously linked to improved wine quality due to an increased release of mannoproteins into the wine matrix. However, metabolic pathways involved in this process during sequential fermentation are still underexplored. This study aimed at investigating cellular response by exploring the proteomes of two strains (S. bacillaris FR1751 and S. cerevisiae EC1118) in single and sequential wine fermentation. Material and methods: Three fermentation trials were performed with synthetic must (MS300) at 20°C using single yeast inoculum (2x106 cells/mL), and sequential fermentation starting with the inoculum S. bacillaris FRI751 and adding S. cerevisiae EC1118 after 48h. The yeast pellets were collected after 168 hours for the single fermentation of S. bacillaris FRI751 and sequential fermentation (SEQ), and 120 hours for the S. cerevisiae EC1118. The proteins were prepared by suspension trapping (STrap) method using C18 and micro-quartz fiber paper tip-column, digested with trypsin, and sequenced by Liquid Chromatography with tandem mass Spectrometry. The proteins were identified and quantified by MaxQuant software (Version 1.6.17.0) with the MaxLFQ algorithm against the global proteome database of S. cerevisiae EC1118, and S. bacillaris FRI751. The differentially abundant proteins (DAPs) were analyzed following S. cerevisiae EC1118 single versus SEQ, and FRI751 single versus SEQ by the DEP R package. Results: A total of 1907 proteins were obtained after removing contaminants and reverses, and 277 of these proteins were DAPs. Among them, six proteins involved in the glycan biosynthesis and metabolism in the sequential fermentation group compared to single fermentation of S. cerevisiae EC1118 (reaching up to 5.42 log2 fold change). Conclusion: The presence of S. bacillaris in sequential fermentation with S. cerevisiae up-regulated DAPs involved in glycan biosynthesis and metabolism, indicating a yeast-to-yeast interaction. Glycans are involved in cell wall composition, and they can lead to modifications in yeast cell wall structure, mainly beta-glucans and mannoproteins. These compounds released into the wine matrix can impact improving wine instability and quality.