N-phenylglucosylamine hydrolysis: A mechanistic probe of β-glucosidase

The spontaneous hydrolysis of glycosylamines, where the aglycone is either a primary amine or ammonia, is over a hundred million-times faster than that of O- or S-glycosides. The reason for this (as pointed out by Capon and Connett in 1965) is that, in contrast to the mechanism for O- or S-glycoside hydrolysis, hydrolysis of these N-glycosides (e.g., glc-NHR) involves an endocyclic C–O bond cleavage resulting in formation of an imine (iminium ion) which then reacts with water. Since ring-opening is kinetically favored with glycosylamines, compounds such as phenylglucosylamine can be a useful probes of enzymes that have been suggested to possibly follow this mechanism. With β-glucosidase from sweet almonds, the enzyme is highly efficient in catalyzing the hydrolysis of phenyl glucoside (kcat/knon ∼ 1014) and phenyl thioglucoside (kcat/knon ∼ 1010) while with either the almond or the Aspergillus niger enzyme or with yeast α-glucosidase, there is no detectable catalysis of phenylglucosylamine hydrolysis (kcat/knon < 20). These results are consistent with the generally accepted mechanism involving exocyclic bond cleavage by these enzymes.