Isothermal titration calorimetric studies on the binding of a family 6 carbohydrate-binding module of Clostridium thermocellum XynA with xlylooligosaccharides

The family 6 carbohydrate-binding module (CBM) of Clostridium thermocellum XynA was expressed, and the binding equilibria of the CBM with xylooligosaccharides (degree of polymerization DP=2−8) were observed by isothermal titration calorimetry (ITC) at pH 8. The association constant, K a , increased with increasing DP from 5×10 3 M −1 (DP=2) to approximately 5×10 5 M −1 (DP=5-8) at 20°C. The K a values at 60°C were about 1/10 of those at 20°C. The binding was found to be an enthalpy-driven reaction. The DP dependence of the thermodynamic parameters of the binding reaction suggested the size of the ligand-binding site to be 5 xylose units long.