Acceptor specificity of cellobiose phosphorylase from Cellvibrio gilvus: synthesis of three branched trisaccharides

Cellobiose phosphorylase from Cellvibrio gilvus was examined for its acceptor specificity in the synthetic reaction with glucose-1-phosphate, using substrates in which the C-6 substituent of d-Glc had been altered. A range of disaccharides were also tested for acceptor specificity but only those with (1⃗6)-linkages were successful acceptors. Melibiose, gentiobiose, isomaltose and also the monosaccharide glucuronamide were found to react with cellobiose phosphorylase and glucose-1-phosphate giving β-d-Glcp-(1⃗4)-[α-d-Galp-(1⃗6)]-d-Glcp, β-d-Glcp-(1⃗4)-[β-d-Glcp-(1⃗6)]-d-Glcp, β-d-Glcp-(1⃗4)-[α-d-Glcp-(1⃗6)]-d-Glcp and β-d-Glcp-(1⃗4)-d-GlcUNp, respectively. These products were purified using a range of chromatographic methods and characterised by NMR and FAB-MS. This is the first time cellobiose phosphorylase has been shown to synthesise trisaccharides.