Optimization of a rapid method for studying the cellular location of β-glucosidase activity in wine yeasts
Date
2005
Authors
Arevalo Villena M.
Ubeda Iranzo J.F.
Cordero Otero R.R.
Briones Perez A.I.
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Abstract
Aims: To improve a method for determining β-glucosidase activity and to apply it in yeasts isolated from wine ecosystems from 'La Mancha' region and to know its cellular location. Methods and Results: A total of 82 wine yeasts were identified (PCR/RFLP) and evaluated for their β-glucosidase activity. First, they were qualitatively evaluated by growth on YNB cellobiose, the activity was quantified using different culture media, under aerobic and anaerobic conditions and cells after 24-72 h of growth. To study the location activity, five fractions were obtained (supernatant, whole cell, cell wall, cytosol and cell membrane). The enzymatic assays were optimized, being: growth in YP cellobiose for 72 h in aeration conditions and, after cell removing, enzyme analysis with 128 g 1-1 of cellobiose as substrate, for 30 min at 30°C. The genus that displayed the greatest activity were Pichia, Hanseniaspora and Rhodotorula, and the activity was intracellular. Conclusions: The study showed that β-glucosidase activity was induced by the carbon source and was aerobic dependent. The non-Saccharomyces species displayed the greatest activity, which was intracellular and strain-dependent. Significance and Impact of the Study: This study developed a reliable method for screening β-glucosidase activity in yeasts isolated from wine ecosystems. This activity is very important in the release of monoterpenols from glycoside precursors for the enhancement of wine aromas. © 2005 The Society for Applied Microbiology.
Description
Keywords
beta glucosidase, cellobiose, fermentation, aerobic cell culture, anaerobic cell culture, article, cell growth, cell membrane, cellular distribution, culture medium, cytosol, ecosystem, enzymatic assay, enzyme activity, enzyme analysis, nonhuman, Pichia, polymerase chain reaction, qualitative analysis, restriction fragment length polymorphism, Rhodotorula, whole cell, wine, yeast, beta-Glucosidase, Cell Membrane, Cell Wall, Cellobiose, Culture Media, Cytosol, Food Microbiology, Oxygen, Polymerase Chain Reaction, Polymorphism, Restriction Fragment Length, Wine, Yeasts, Hanseniaspora, Pichia, Saccharomyces
Citation
Journal of Applied Microbiology
99
3
99
3