Hydrolysis of β-d-glucopyranosyl fluoride to α-d-glucose catalyzed by Aspergillus niger α-d-glucosidase

Aspergillus niger α-d-glucosidase, crystallized and free of detectable activity for β-d-glucosides, catalyzes the slow hydrolysis of β-d-glucopyranosyl fluoride to form α-d-glucose. Maximal initial rates, V, for the hydrolysis of β-d-glucosyl fluoride, p-nitrophenyl α-d-glucopyranoside, and α-d-glucopyranosyl fluoride are 0.27, 0.75, and 78.5 μmol.min^-1.mg^-1, respectively, with corresponding V/K constants of 0.0068, 1.44, and 41.3. Independent lines of evidence make clear that the reaction stems from β-d-glucosyl fluoride and not from a contaminating trace of α-d-glucosyl fluoride, and is catalyzed by the α-d-glucosidase and not by an accompanying trace of β-d-glucosidase or glucoamylase. Maltotriose competitively inhibits the hydrolysis, and β-d-glucosyl fluoride in turn competitively inhibits the hydrolysis of p-nitrophenyl α-d-glucopyranoside, indicating that β-d-glucosyl fluoride is bound at the same site as known substrates for the α-glucosidase. Present findings provide new evidence that α-glucosidases are not restricted to α-d-glucosylic substrates or to reactions providing retention of configuration. They strongly support the concept that product configuration in glycosylase-catalyzed reactions is primarily determined by enzyme structures controlling the direction of approach of acceptor molecules to the reaction center rather than by the anomeric configuration of the substrate.