Abstract
In winemaking and brewing, aromatic precursors are found in S-conjugated forms with glutathione, dipeptides or cysteine are cleaved by the intracellular CS lyase (CSL) enzyme activity of yeast during fermentation, releasing polyfunctional thiols. These volatile compounds are key elements in the aroma of these beverages. However, commercially available yeast strains convert on average only 1% of the precursors, which limits the exploitation of the aromatic potential of musts and worts.The main objective of this thesis is to study the functioning of CSL enzymes in order to exploit a possible exogenous addition to increase the revelation of thiols from precursors present before fermentation, during the production of wine and beer.The first part of this work is devoted to the development of a method for characterizing CSL enzymes by spectrophotometry, based on the use of a non-natural substrate, analogous to thiol precursors, whose enzymatic degradation releases a chromophore that can be quantified at UV-visible wavelengths. This functional characterization model was validated by its ability to represent enzymatic behavior towards S-3-mercaptohexan-1-ol-L-cysteine, the most abundantly found thiol precursor in wine and beer.The second part of the project is dedicated to characterizing selected CSL enzymes expressed in recombinant form. Despite the structural differences observed between CSLs expressed by different genes, these enzymes only recognize the simplest form of thiol precursors: the S-conjugated cysteine. CSLs exhibit variable tolerance to the acidic pHs characteristic of wine and beer, and we have shown that their activity is inhibited in the presence of cysteine.Finally, these CSL enzymes were tested on a pilot-scale in winemaking and brewing processes in order to assess the potential of their addition to reveal polyfunctional thiols in these industrial processes.The knowledge acquired in this project highlights the functioning of CSL enzymes towards different precursor forms, the technological barriers to their application due to the composition of the oenological and brewing matrices, and the theoretical solutions that could be envisaged for the application of exogenous CSLs to enhance the release of thiols in these two fermentation processes.