Dr. Carien Coetzee
6 December 2022
Yeast strains differ significantly in their ability to release thiols from the conjugated precursors and transform 3MH into 3MHA1. Studies have shown that specific genes are involved in releasing aromatic thiols and could serve as markers to identify yeast strains with high thiol-releasing potential.
Thiol release from precursors
In the grapes, 3MH, 3MHA and 4MMP are not present in their free aromatic form but rather (in the case of 3MH and 4MMP) in the form of non-aromatic, non-volatile conjugated (combined) precursors. The yeast transforms these precursors during alcoholic fermentation to release the free and aromatic volatile thiols from the conjugated compound (thus breaking the compound apart), resulting in intense tropical fruit aromas. Non-odourant cysteinylated and glutathionylated precursors have been identified as conjugated precursors in the grape must2–5. However, studies have also shown that these precursors are not the only source of volatile thiols. Other mechanisms and unidentified precursors also contribute significantly to the final content of fruity thiols in wine (read more here).6,7
3MHA does not have a conjugated precursor in the grape. It is formed during fermentation through the esterification of 3MH with acetic acid, a reaction regulated by specific enzymes produced by the yeast. Therefore, it would seem that 3MH will first have to be formed, after which the reaction between 3MH and acetic acid will yield 3MHA.
The role of yeast
The yeast strain used to perform the fermentation is one of the most important factors affecting the thiol production and concentration in the final wine8. Yeast strains differ significantly in their capability to produce thiols. This ability (or lack thereof) can be exploited by the winemaker to precisely craft the desired wine style by selecting a yeast strain with specific properties and capabilities.
It’s all in the genes
Before thiol liberation, the yeast must transport the conjugated precursor from the juice into the yeast cell. Once in the cell, the glutathionylated precursor is first transformed into the cysteinylated precursor. The yeast then cleaves the precursor, utilising the amino acid portion of the precursor for cell growth while “discarding” the remainder of the precursor (the aromatic thiol). This transformation entails a complex mechanism involving multiple genes9.
For the cleavage of the cysteinylated precursors (and subsequent release of the aromatic thiols), a specific enzyme, namely the carbon-sulphur β-lyase enzyme, is required. Various genes have been identified as responsible for the yeasts’ production capabilities of cleaving enzymes, including BNA3, CYS3, STR3 and GLO1. However, the main gene responsible for the release of thiols is the IRC7 gene. Studies done in a synthetic wine medium with precursors present have shown that a full-length copy of IRC7 is required for the cleavage of precursors. Deleting the gene resulted in most yeast strains being incapable of converting the conjugated precursors into aromatic thiols10. This gene could potentially serve as a molecular marker to predict a yeast’s potential to release thiols.
The esterification of 3MH to form 3MHA also requires a specific gene encoding alcohol acetyltransferase (ATF1).11
Yeast strain variability
Several studies have reported the limited capability of most Saccharomyces cerevisiae yeast strains to release volatile thiols from the corresponding non-volatile precursors (usually less than 5% converted)12. A study13 assessed the ability of 82 different yeast strains (not all commercially available) to produce thiols from a grape-like medium containing realistic concentrations of conjugated thiol precursors.
- a 20-fold difference between the yeasts’ ability to release 3MH
- a 35-fold difference in the yeasts’ ability to release 4MMP
The yeasts were categorised according to the concentration of thiols produced during fermentation. The results showed that most (70%) of the strains tested were considered low releasers of thiols, 20% were moderate releasers, and 10% were considered high releasers. In most cases, the low releasers possessed an inactive IRC7 gene, which limited the ability to release thiols, while the moderate and high releasers had an active or partially active IRC7 gene.
Among the selection of commercial yeast strains tested, the strains recommended by suppliers for the production of white wine produced relatively higher concentrations of volatile thiols when compared to strains that were recommended for the production of red wine only or the production of both red and white wine. This shows some active selection from yeast manufacturers for strains with a higher thiol-releasing capacity for producing fruit-driven white wines such as Sauvignon blanc.
Commercial yeast manufacturers offer a wide range of yeast strains, often with recommendations based on the ability to produce certain attributes. A better understanding of the genes and mechanisms involved will allow winemakers to enquire about the yeast’s ability to release thiols from precursors in order to produce wines in the desired wine style more reliably.
3MHA: 3-mercaptohexyl acetate
(1) Swiegers, J. H.; Willmott, R.; Hill-Ling, A.; Capone, D. L.; Pardon, K. H.; Elsey, G. M.; Howell, K. S.; de Barros Lopes, M. A.; Sefton, M. A.; Lilly, M.; Pretorius, I. S. Modulation of Volatile Thiol and Ester Aromas by Modified Wine Yeast. In Weurman flavour research symposium; Developments in Food Science, Elsevier, Amsterdam, The Netherlands: Roskilde, Denmark, 2005.
(2) Tominaga, T.; Peyrot des Gachons, C.; Dubourdieu, D. A New Type of Flavour Precursors in Vitis Vinifera L. Cv. Sauvignon Blanc: S-Cysteine Conjugates. Journal of Agricultural and Food Chemistry 1998, 46, 5215–5219.
(3) Tominaga, T.; Masneuf, I.; Dubourdieu, D. A S-Cysteine Conjugate, Precursor of Aroma of White Sauvignon. J. Int. Sci. Vigne Vin 1995, 29 (4), 227–232.
(4) Des Gachons, C. P.; Tominaga, T.; Dubourdieu, D. Sulfur Aroma Precursor Present in S-Glutathione Conjugate Form: Identification of S-3-(Hexan-1-Ol)-Glutathione in Must from Vitis Vinifera L. Cv. Sauvignon Blanc. Journal of Agricultural and Food Chemistry 2002, 50 (14), 4076–4079. https://doi.org/10.1021/jf020002y.
(5) Fedrizzi, B.; Pardon, K. H.; Sefton, M. A.; Elsey, G. M.; Jeffery, D. W. First Identification of 4-S-Glutathionyl-4-Methylpentan-2-One, a Potential Precursor of 4-Mercapto-4-Methylpentan-2-One, in Sauvignon Blanc Juice. Journal of Agricultural and Food Chemistry 2009, 57 (3), 991–995.
(6) Pinu, F. R.; Jouanneau, S.; Nicolau, L.; Gardner, R. C.; Villas-Boas, S. G. Concentrations of the Volatile Thiol 3-Mercaptohexanol in Sauvignon Blanc Wines: No Correlation with Juice Precursors. American Journal of Enology and Viticulture 2012, 63 (3), 407–412. https://doi.org/10.5344/ajev.2012.11126.
(7) Harsch, M. J.; Benkwitz, F.; Frost, A.; Colonna-Ceccaldi, B.; Gardner, R. C.; Salmon, J. M. New Precursor of 3-Mercaptohexan-1-Ol in Grape Juice: Thiol-Forming Potential and Kinetics during Early Stages of Must Fermentation. Journal of Agricultural and Food Chemistry 2013, 61 (15), 3703–3713. https://doi.org/10.1021/jf3048753.
(8) Cordente, A. G.; Curtin, C. D.; Varela, C.; Pretorius, I. S. Flavour-Active Wine Yeasts. Applied Microbiology and Biotechnology 2012, 96 (3), 601–618. https://doi.org/10.1007/s00253-012-4370-z.
(9) Belda, I.; Ruiz, J.; Esteban-Fernández, A.; Navascués, E.; Marquina, D.; Santos, A.; Moreno-Arribas, M. V. Microbial Contribution to Wine Aroma and Its Intended Use for Wine Quality Improvement. Molecules 2017, 22 (2). https://doi.org/10.3390/molecules22020189.
(10) Santiago, M.; Gardner, R. C. Yeast Genes Required for Conversion of Grape Precursors to Varietal Thiols in Wine. FEMS Yeast Research 2015, 15 (5), 1–10. https://doi.org/10.1093/femsyr/fov034.
(11) Swiegers, J. H.; Bartowsky, E. J.; Henschke, P. a.; Pretorius, I. S. Yeast and Bacterial Modulation of Wine Aroma and Flavour. In Australian Journal of Grape and Wine Research; Blair Francis, M.E., Pretorius, I.S., R. J., Ed.; The Australian Wine Research Institute: Glen Osmond, Australia, 2005; Vol. 11, pp 139–173. https://doi.org/10.1111/j.1755-0238.2005.tb00285.x.
(12) Ruiz, J.; Kiene, F.; Belda, I.; Fracassetti, D.; Marquina, D.; Navascués, E.; Calderón, F.; Benito, A.; Rauhut, D.; Santos, A.; Benito, S. Effects on Varietal Aromas during Wine Making: A Review of the Impact of Varietal Aromas on the Flavor of Wine. Applied Microbiology and Biotechnology 2019, 103 (18), 7425–7450. https://doi.org/10.1007/s00253-019-10008-9.
(13) Cordente, T.; Schmidt, S.; Curtin, C. Understanding Differences among Wine Yeast Strains in Their Ability to Release “tropical” Thiols. AWRI Technical review 2017, 228, 6–10.
(14) Coetzee, C. Yeast and Its Ability to Release Thiols. Wineland Magazine 2020.
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