Regioselective synthesis of p-nitrophenyl glycosides of β-d-galactopyranosyl-disaccharides by transglycosylation with β-d-galactosidases Original Research Article

The β-d-galactosidase from porcine liver induced regiospecific transglycosylation of β-d-galactose from β-d-Gal-OC6H4NO2-o to OH-6 of, respectively, p-nitrophenyl glycoside acceptors of Gal, GlcNAc and GalNAc to afford β-Gal-(1→6)-α-Gal-OC6H4NO2-p, β-Gal-(1→6)-β-Gal-OC6H4NO2-p, β-Gal-(1→6)-α-GalNAc-OC6H4NO2-p, β-Gal-(1→6)-β-GalNAc-OC6H4NO2-p, β-Gal-(1→6)-α-GlcNAc-OC6H4NO2-p, and β-Gal-(1→6)-β-GlcNAc-OC6H4NO2-p. The enzyme showed much higher transglycosylation activity for the α-glycoside acceptors than the corresponding β-glycoside acceptors. The regioselectivity of the β-d-galactosidase from Bacillus circulans ATCC 31382 greatly depended on the nature of the acceptor. When α-d-GalNAc-OC6H4NO2-p and α-d-GlcNAc-OC6H4NO2-p were used as acceptors, the enzyme showed high potency for regioselective synthesis of β-Gal-(1→3)-α-GalNAc-OC6H4NO2-p and β-Gal-(1→3)-α-GlcNAc-OC6H4NO2-p in high respective yields of 75.9 and 79.3% based on the acceptors added. However, replacement of β-d-Gal-OC6H4NO2-p by β-d-GalNAc-OC6H4NO2-p did change the direction of galactosylation. The enzyme formed regioselectively β-Gal-(1→6)-β-Gal-OC6H4NO2-p with (β-Gal-1→(6-β-Gal-1→)n6-β-Gal-OC6H4NO2-p, n=1–4). No β-(1→3)-linked product was detected during the reaction. Use of the two readily available β-d-galactosidases facilitates the preparation of (1→3)- and (1→6)-linked disaccharide glycosides of β-d-Gal-GalNAc and β-d-Gal-GlcNAc.