Purification and biochemical characterization of an atypical β-glucosidase from Stachybotrys microspora

Stachybotrys microspora is a filamentous fungus secreting various β-glucosidases. The current work undertakes purification and biochemical characterization of the most particular one, named bglG, which is the only one to be highly produced on glucose and fairly on cellulose-based medium. Although produced on glucose, bglG activity continues to be highly inhibited by this sugar. After two chromatographic steps, bglG was purified to homogeneity and shown to be a monomeric protein with the molecular mass of 225 kDa. The highest bglG activity was obtained at pH 5 and a temperature range of 50–60 °C. This enzyme was shown to act through a retaining-enzyme mechanism. The N-terminal sequence analysis did not reveal any homology with all available sequences in the database. BglG is somehow atypical for multiple reasons: (1) BglG is insensitive to the conventional Coomassie staining protocol and CuCl2 method was applied to reveal the protein; (2) the bglG activity is strongly enhanced by ferrous ion (Fe2+) to 161% at 5 and 10 mM of Fe2+. Flame spectrometry analysis showed that iron was stoechiometrically and strongly bound to bglG; (3) besides cellobiose, BglG is active on sucrose (114%); a rarely described property among β-glucosidases and (4) bglG is significantly stimulated by xylose. BglG could be considered as very original, since all known β-glucosidases, did not share these properties.