Localization and Characterization of α-Glucosidase Activity in Brettanomyces lambicus

Abstract

Brettanomyces lambicus was isolated and identified from a typical overattenuating Belgian Iambic beer and exhibited extracellular and intracellular ax-glucosidase activities. Production of the intracellular enzyme was higher than production of the extracellular enzyme, and localization studies showed that the intracellular a-glucosidase is mostly soluble and partially cell wall bound. Both intracellular and extracellular enzymes were purified by ammonium sulfate precipitation, gel filtration (Sephadex G-150, Sephadex G-200, Ultrogel AcA-44), and ion-exchange chromatography (sulfopropyl-Sephadex C-50, (carboxymethyl-Sephadex C-50). The intracellular a-glucosidase exhibited optimum activity at 39°C and pH 6.2. The extracellular enzyme exhibited optimum catalytic activity at 40°C and pH 6.0. The molecular masses of purified intracellular and extracellular a-glucosidases, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, were 72,500 and 77,250, respectively. For both enzymes there was a decrease in the rate of hydrolysis with an increase in the degree of polymerization, and both enzymes hydrolyzed dextrins isolated from lambic wort (degrees of polymerization, 3 to 9 and more than 9). The Km values for p-nitrophenyl-cv-D-glucopyranoside, maltose, and maltotriose for the intracellular enzyme were 0.9, 3.4, and 3.7 mM, respectively. The Ki values for both enzymes were between 28.5 and 57 ,M for acarbose and between 7.45 and 15.7 mM for Tris. These enzymes are probably involved in the overattenuation of spontaneously fermented Iambic beer.

Characterization of a Maltase from an Early-Diverged Non-Conventional Yeast Blastobotrys adeninivorans

“…In a yeast Brettanomyces bruxellensis (former name Br. lambicus), both extra-and intracellular maltases were inhibited not only by acarbose (K i values between 28.5 and 57 µM) but also by Tris (K i values between 7.45 and 15.7 mM) [57]. Compared to Br.…”

Microbial acidification, alcoholization, and aroma production during spontaneous lambic beer production

“…3). 140 Possibly due to the limited -glucosidase activity at low pH values, the over-attenuation of lambic beers takes a long time. Up to now, Brettanomyces species are the sole yeasts detected in lambic beer production processes capable of performing maltooligosaccharide degradation, although better over-attenuation is obtained when LAB are also present, indicating a synergistic effect.…”

Species-specific identification ofDekkera/Brettanomycesyeasts by fluorescently labeled DNA probes targeting the 26S rRNA

“…Yeasts of the genus Brettanomyces (anamorphic form) and their ascospore-forming type Dekkera (teleomorphic form) are well known to produce off-flavors in wine and must, but have been also isolated from other fermented beverages such as sparkling wine, cider, sherry, and beer (Kumara et al, 1993;Ibeas et al, 1996;Ciani & Ferraro, 1997;Coton et al, 2006). These yeasts are widespread, and have been identified in almost every wine-producing area of the world (Fugelsang, 1997).…”