KINETIC-PARAMETERS OF HYDROLYSIS AND TRANSGLYCOSYLATION CATALYZED BY AN EXO-BETA-(1,4)-GALACTANASE

The action pattern and reaction mechanism of the exo-beta-(1,4)-galact anase from Aspergillus niger var. aculeatus on galactooligosaccharides of degree of polymerization (dp) from 2-6 were investigated. The reac tion was assayed at the optimum pH of both hydrolysis and transglycosy lation, and HPLC analysis of the reaction mixtures was used to obtain kinetic parameters. Hydrolysis of oligosaccharides to galactose follow ed a Michaelis-Menten behavior whatever the dp. Catalytic activity was always higher for hydrolysis than for transglycosylation. At the opti mum pH, the affinity of the enzyme increased for hydrolysis and decrea sed for transglycosylation with increasing dp. Transglycosylation was explained as a dependent binding of two similar molecules of substrate s. The dependence of the kinetic parameters on the degree of polymeriz ation of substrates was used to evaluate the subsite affinities. The a ctive site of exogalactanase was concluded to possess three subsites w ith the catalytic site being located between subsites 1 and 2. (C) 199 7 by Elsevier Science Inc.