Engineering of cellobiose phosphorylase for glycoside synthesis

Disaccharide phosphorylases are increasingly applied for glycoside synthesis, since they are very regiospecific and use cheap and easy to obtain donor substrates. A promising enzyme is cellobiose phosphorylase (CP), which was discovered more than 50 years ago. Many other bacterial CP enzymes have since then been characterized, cloned and applied for glycoside synthesis. However, the general application of wild-type CP for glycoside synthesis is hampered by its relatively narrow substrate specificity. Recently we have taken some successful efforts to broaden the substrate specificity of Cellulomonas uda CP by directed evolution and protein engineering. This review will give an overview of the obtained results and address the applicability of the engineered CP enzymes for glycoside synthesis. CP is the first example of an extensively engineered disaccharide phosphorylase, and may provide valuable information for protein engineering of other phosphorylase enzymes.