Synthesis of highly branched anionic α-glucans by thermostable phosphorylase-catalyzed α-glucuronylation

Highly branched anionic α-glucans were enzymatically synthesized by thermostable phosphorylase-catalyzed α-glucuronylation of highly branched cyclic dextrin using α-d-glucuronic acid 1-phosphate (GlcA-1-P) as a glycosyl donor. The resulting products were characterized by 1H NMR measurement as well as high performance anion exchange chromatographic and MALDI-TOF MS analyses after treatments with several amylases. α-d-Glucose 1-phosphate was detected in the reaction mixtures, suggesting the occurrence of phosphorolysis in the α-glucuronylation. The glucuronylation ratios of glucuronic acid residues to non-reducing ends were evaluated from quantification of α-d-glucose 1-phosphate and inorganic phosphate in the reaction mixtures, which were relatively in good agreement with those determined by 1H NMR analysis of the products. The glucuronylation ratios increased with increasing feed ratios of GlcA-1-P/non-reducing ends.