Selection, characterization and comparison of β-glucosidase from mould and yeasts employable for enological applications

Abstract

A comparison has been made of the main physicochemical and kinetic parameters of the β-glucosidase (βG, EC 3.2.1.21) of a commercial preparation from Aspergillus niger chosen for its high activity and that of 463 yeast strains, isolated and screened from Sicilian musts and wines. A higher percentage of βG-positive strains (30%) was found on Yeast Carbon Base + lysine compared to 18% on Sabouraud + ethanol + SO 2 and just 8% on Sabouraud medium. The positive strains with the highest βG activity were subsequently identified and compared with the purified commercial preparation. Optimum pH values were found to be 4.5 for A. niger βG, with relative activity of about 87% at pH 3.5, that is close to that of wine, whilst amongst the yeasts, AL 41 was the strain with optimum pH closest to that of wine. Optimum temperatures were found to be about 62 and 20–25 °C for A. niger and yeast βG, respectively. Within the latter temperature range, that is potentially the most suitable for the use of the enzymes in enology, mould βG relative activity was found to be 40–50%. Enzyme stability in wine model solution (at 20 °C) was 12 days for mould activity and at least 35–45 for that from yeasts. Finally, the V max values were ranged from 1.75 to 6.94 U mg −l of protein for the yeasts, while the K m values were 0.61–2.53 mM. The behaviour of βG from A. niger could not be explained by the Michaelis–Menten equation. The case was ascribable to the typical behaviour of an unproductive bond with substrate concentration exceeding saturation point and incorrect positioning at the enzyme’s active site. The enzyme from mould would be useful during the final stages of alcoholic fermentation, whilst some yeast strains (AL 112 and AL 41) could be directly added during fermentation in order to enhance wine aroma.