Macrowine 2021
IVES 9 IVES Conference Series 9 Proteomic and activity characterization of exocellular laccases from three Botrytis cinerea strains

Proteomic and activity characterization of exocellular laccases from three Botrytis cinerea strains

Abstract

Botrytis cinerea is a fungus that causes common infection in grapes and other fruits. In winemaking, its presence can be both considered desirable in the case of noble rot infection or undesirable when grey rot is developed. This fungus produces an extracellular enzyme known as laccase which is able to cause oxidation of phenolic compounds present in must and wine, causing most of the times a decrease in its quality and problems during the winemaking process [1]. Material and methods: Three B. cinerea strains (B0510, VA612 and RM344) were selected and grown in a liquid medium adapted from one previously described [2]. The enzyme was isolated by tangential ultrafiltration of the culture medium using a QuixStand system equipped with a 30 KDa filtration membrane. The purity of the isolated enzymes was checked using SDS-PAGE. The characteristics (molecular weight, % of glycosylation, specific activity, activity in function of pH of the enzyme isolated from each strain were studied with ABTS as a substrate. Results: The enzymes isolated from the three strains showed the same molecular weight, 97 KDa, in good agreement with the molecular weight previously determined for B. cinerea laccase using SDS-PAGE [2]. The percentage of glycosylation was high, being estimated in 70% on weight, also similar to that described by other authors [3]. Despite similar physical characteristics of the enzymes obtained from different strains, their activity were quite different. The enzymes isolated from B0510 and VA612 strains showed similar specific activity for ABTS oxidation, being 0.3 and 0.21 mM for their Km and their Vmax were 1.28 and 1.45 mM/min per milligram of enzyme respectively. The activity for RM344 enzyme was found much lower, with values of 0.78 mM for Km and 0.13 mM/min per milligram of enzyme for Vmax. The enzyme isolated from the B0510 strain presented its highest activity at pH 2.9 while VA612 and RM344 enzymes showed the maximum activity at pH 3.3. All these values were quite lower compared to previously measured by other authors [4-6]. Those differences in the enzyme activity may be related with differences in the active center of the enzyme and could have important consequences for the winemaking process depending on the strains of the B. cinerea strain involved in the infection of grapes.

[1]P. Ribéreau-Gayon, Y. Glories, A. Maujean, D. Dubourdieu, Handbook of Enology, Volume 2: The chemistry of Wine Stabilization and Treatments, John Wiley & Sons, Ltd, 2000. [2]D. Slomczynski, J.P. Nakas, S.W. Tanenbaum, Applied and Environmental Microbiology, 61 (1995) 907. [3]C. THURSTON, Microbiology-Sgm, 140 (1994) 19. [4]I. MARBACH, E. HAREL, A. MAYER, Phytochemistry, 23 (1984) 2713. [5]I. MARBACH, E. HAREL, A. MAYER, Phytochemistry, 22 (1983) 1535. [6]M. Dubernet, P. Ribereau-Gayon, H.R. Lerner, E. Harel, A.M. Mayer, Phytochemistry, 16 (1977) 191.

Publication date: May 17, 2024

Issue: Macrowine 2016

Type: Poster

Authors

Cédric Saucier*, Anne-Sophie Walker, Christiane Auclair, François Garcia, Francois-Xavier Sauvage, Jullien Drone, Natalia Quijada-Morin, Patrick Chemardin

*Université de Montpellier

Contact the author

Tags

IVES Conference Series | Macrowine | Macrowine 2016

Citation

Related articles…

Simultaneous monitoring of dissolved CO2 and collar from Rosé sparkling wine glasses: the impact of yeast macromolecules

Champagne or sparkling wines elaborated through the same traditional method, which consists in two major yeast-fermented steps, typically hold about 10 to 12 g/L of dissolved CO2 after the second fermentation in a closed bottle. Hundreds of molecules and macromolecules originating from grape and yeast cohabit with dissolved CO2; they are essential compounds contributing to many organoleptic characteristics (effervescence, foam, aroma, taste, colour…). Indeed, the second alcoholic fermentation and the maturation on lees (which may last from 12 months up to several years) both induce various quantitative and qualitative changes in the wine through the action of yeast, as listed hereafter: development of aromas during aging on lees, release of nitrogen compounds during autolysis and release of macromolecules (polysaccharides, lipids, nucleic acids) in wine.

Sensory definition of green aroma concept in red French wines. Evidence for the contribution of novel volatile markers

The aromatic complexity of a wine results from the perception of the association of volatile molecules and each aroma can be categorized into different families. The “green” aromas family in red wines has retained our attention by its close link with the fruity perception. In that study, the “green” olfactory concept of red wines was considered through a strategy combining both sensory analysis and hyphenated chromatographic techniques including HPLC and MDGC (Multidimensional Gas Chromatography). The aromatic space of this concept was specified by lexical generation through a free association task on 22 selected wines by a panel of wine experts. Then, 70 French red wines were scored on the basis of the intensity of their “green” and “fruity” attributes.

Monitoring of Pesticide Residues from Vine to Wine

Those previous years, pesticides are often brought to the forefront by media. Questions arose about their toxicity for growers and consumers. Even if a downward trend is underway, the use of pesticides is required to ensure steady quality and quantity of harvests. A large number of active ingredients are authorized but regarding viticulture, mainly insecticides and fungicides are applied, to control pests and diseases and to increase crop yield. Some phytosanitary products, principally fungicides, applied close to the harvest date may frequently be detected in wines.

New biological tools to control and secure malolactic fermentation in high pH wines

Originally, the role of the malolactic fermentation (MLF) was simply to improve the microbial stability of wine via biological deacidification. However, there is an accumulation of evidence to support the fact that lactic acid bacteria (LAB) also contribute positively to the taste and aroma of wine. Many different LAB enter into grape juice and wine from the surface of grape berries, cluster stems, vine leaves, soil and winery equipment. Due to the highly selective environment of juices and wine, only a few types of LAB are able to grow.

HEAT BERRY : Sensitivity of berries ripening to higher temperature and impact on phenolic compounds in wine

The grapevine is an important economical crop that is very sensitive to climate changes and microclimate. The observations made during the last decades at a vineyard scale all concur to show the impact of climate change on vine physiology, resulting in accelerated phenology and earlier harvest (Jones and Davis 2000). It is well-known that berry content is affected by the ambient temperature. While the first experiences were primarily conducted on the impact of temperature on anthocyanin accumulation in the grape, few studies have focused on others component of phenolic metabolism, such as tannins.