Comparative study of new α-galactosidases in transglycosylation reactions

Abstract

We have studied the potential of several newly cloned α-galactosidases to catalyze the regioselective synthesis of disaccharides using 4-nitrophenylgalactoside as a donor. The kinetics of the reactions were followed by in situ NMR spectroscopy. The following thermophilic enzymes have been tested: Aga A and an isoenzyme Aga B obtained from the strain KVE39 and Aga 285 from the strain IT285 of Bacillus stearothermophilus; Aga T is an α-galactosidase from Thermus brockianus (strain IT360). Two other non-thermophilic α-galactosidases have also been evaluated: Aga 1 ( Streptococcus mutans, strain Ingbritt) and Raf A ( Escherichia coli, strain D1021). For all of the enzymes studied, high regioselectivity was observed leading to two (1→6)-disaccharides: 4-nitrophenyl α- d-galactopyranosyl-(1→6)-α- d-galactopyranoside and methyl α- d-galactopyranosyl-(1→6)-α- d-galactopyranoside, which were obtained in 54% (Aga B) and 20% (Aga T) yields, respectively.