Abstract
α-Glucosyltransferase ( αGT) from Protaminobacter rubrum catalyzes either an intramolecular transglucosylation of sucrose, giving isomaltulose, or intermolecular transglucosylations from sucrose to versatile acceptors. To obtain insights into this enzyme, especially in connection with the glycosyltransfer mechanism, kinetic studies were carried out using synthetic sucrose analogs. The k cat values obtained for 1′-substituted sucrose analogs exhibited a strong correlation with Hammett substituent constants with a slope of −2, suggesting a rate-limiting glycoside cleavage with a completely protonated transition state. The α-deuterium isotope effect ( k H/ k D=1.20±0.08) indicated an oxocarbenium ion-like transition state. Nojirimycin and 1′-amino-1′-deoxy and 3′-amino-3′-deoxy analogs of 6′-chlorosucrose had relatively strong inhibitory effects toward the intramolecular glucosylation, (inhibition at 10 mM being 97, 71, and 78%, respectively) in contrast to the 4′-amino-4′-deoxy and 6′-amino-6′-deoxy analogs, which showed moderate or no inhibitory activity. This tendency is presumably due to the presence of carboxylates as catalytic groups of the enzyme. These results reveal similarities between αGT and some glycosidases in the glycoside cleavage mechanism.
Published Version
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