The β-glucosidase from Botryodiplodia theobromae Pat. Kinetics of enzyme-catalysed hydrolysis of o-nitrophenyl β-d-glucopyranoside in dioxan/water

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Abstract
The hydrolysis of o-nitrophenyl .beta.-D-glucopyranoside by the high-molecular weight .beta.-glucosidase (.beta.-D-glucoside glucohydrolase, EC 3.2.1.21) from B. theobromae Pat. was studied in the presence of added dioxan. At donor saturation, the maximum rate of hydrolysis in the presence of up to 50% (vol/vol) dioxan was at pH 4.3-4.5 (pH of the buffer system in water) in McIlvaine''s buffer. Increasing dioxan concentrations progressively decreased the maximum rate of hydrolysis. The rate of enzyme-catalyzed reaction was enhanced at high donor concentrations, but inhibited at low donor concentrations, in the presence of glycerol, methanol, fructose or sucrose. The hydrolytic reaction proceeded with retention of configuration at the anomeric C atom. The kinetics of the enzyme-catalyzed process in the presence of added acceptors indicated that water was necessary for the maintenance of the active enzyme conformation apart from its acceptor function.