Unlocking the secrets of peptide transport in wine yeast: insights into oligopeptide transporter functions and nitrogen source preferences.

IMPORTANCE: Limited nitrogen supply can prevent the completion of alcoholic fermentation. Supplementation through peptides as an alternative, natural source of nitrogen for yeast offers an interesting solution for this issue. In this work, the S. cerevisiae peptide transporters of the Opt and Fot families were studied. We demonstrated that Fot and Opt2 have a broader peptide length preference than previously reported, enabling yeasts to acquire sufficient nitrogen from peptides without requiring additional ammonia or amino acids to complete fermentation. On the contrary, Opt1 was unable to consume any peptide in the given conditions, whereas it has been described elsewhere as the main peptide transporter for peptides longer than three amino acid residues in experiments in laboratory conditions. This controversy signifies the need in applied sciences for approaching experimental conditions to those prevalent in the industry for its more accurate characterization. Altogether, this work provides further evidence of the importance of peptides as a nitrogen source for yeast and their consequent positive impact on fermentation kinetics.

Yeast Plasma Membrane Fungal Oligopeptide Transporters Display Distinct Substrate Preferences despite Their High Sequence Identity.

Fungal Oligopeptide Transporters (Fot) Fot1, Fot2 and Fot3 have been found in Saccharomyces cerevisiae wine strains, but not in strains from other environments. In the S. cerevisiae wine strain EC1118, Fot1 and Fot2 are responsible for a broader range of oligopeptide utilization in comparison with strains not containing any Fot. This leads to better fermentation efficiency and an increased production of desirable organoleptic compounds in wine. Despite the benefits associated with Fot activity in S. cerevisiae within the wine environment, little is known about this family of transporters in yeast. The presence of Fot1, Fot2 and Fot3 in S. cerevisiae wine strains is due to horizontal gene transfer from the yeast Torulaspora microellipsoides, which harbors Fot2Tm, FotX and FotY proteins. Sequence analyses revealed that Fot family members have a high sequence identity in these yeast species. In this work, we aimed to further characterize the different Fot family members in terms of subcellular localization, gene expression in enological fermentation and substrate specificity. Using CRISPR/Cas9, we constructed S. cerevisiae wine strains containing each different Fot as the sole oligopeptide transporter to analyze their oligopeptide preferences by phenotype microarrays. The results of oligopeptide consumption show that Fot counterparts have different di-/tripeptide specificities, suggesting that punctual sequence divergence between FOT genes can be crucial for substrate recognition, binding and transport activity. FOT gene expression levels in different S. cerevisiae wine strains during enological fermentation, together with predicted binding motifs for transcriptional regulators in nitrogen metabolism, indicate that these transporters may be under the control of the Nitrogen Catabolite Repression (NCR) system. Finally, we demonstrated that Fot1 is located in the yeast plasma membrane. This work contributes to a better understanding of this family of oligopeptide transporters, which have demonstrated a key role in the utilization of oligopeptides by S. cerevisiae in enological fermentation.

Diversity of Oligopeptide Transport in Yeast and Its Impact on Adaptation to Winemaking Conditions.

Nitrogen is an essential nutrient for yeasts and its relative abundance is an important modulator of fermentation kinetics. The main sources of nitrogen in food are ammonium and free amino acids, however, secondary sources such as oligopeptides are also important contributors to the nitrogen supply. In yeast, oligopeptide uptake is driven by different families of proton-coupled transporters whose specificity depends on peptide length. Proton-dependent Oligopeptide Transporters (POT) are specific to di- and tri-peptides, whereas the Oligopeptide Transport (OPT) family members import tetra- and pentapeptides. Recently, the novel family of Fungal Oligopeptide Transporters (FOT) has been identified in Saccharomyces cerevisiae wine strains as a result of a horizontal gene transfer from Torulaspora microellipsoides. In natural grape must fermentations with S. cerevisiae, Fots have a broader range of oligopeptide utilization in comparison with non-Fot strains, leading to higher biomass production and better fermentation efficiency. In this review we present the current knowledge on the diversity of oligopeptide transporters in yeast, also discussing how the consumption of oligopeptides provides an adaptive advantage to yeasts within the wine environment.

Glycation affects differently the main soybean Bowman-Birk isoinhibitors, IBB1 and IBBD2, altering their antiproliferative properties against HT29 colon cancer cells.

Naturally-occurring serine protease inhibitors of the Bowman-Birk family, particularly abundant in legume seeds, exert their potential chemopreventive and/or therapeutic properties via protease inhibition. Processing of legume seeds, including soybeans, has been proposed as a major cause for their loss of bioactivity due to glycation. In order to assess how glycation affected the protease inhibitory activities of major soybean Bowman-Birk isoinhibitors (BBI) and their antiproliferative properties, IBB1 and IBBD2 were purified and subjected to glycation under controlled conditions using glucose at high temperature. Both soybean isoinhibitors showed remarkable heat stability. In the presence of glucose, IBBD2 lost most of its trypsin inhibitory activity while IBB1 maintains similar trypsin and chymotrypsin inhibitory activities as in the absence of sugar. Glycation patterns of both BBI proteins were assessed by MALDI-TOF spectrometry. Our results show that the glycation process affects IBBD2, losing partially its antiproliferative activity against HT29 colon cancer cells, while glycated-IBB1 was unaffected.