Enthalpic barriers to the hydrophobic binding of oligosaccharides to phageP22 tailspike protein

The structural thermodynamics of the recognition of complex carbohydrates b y proteins are not well understood, The recognition of O-antigen polysaccha ride by phage P22 tailspike protein is a highly suitable model for advancin g knowledge in this field, The binding to octa- and dodecasaccharides deriv ed from Salmonella enteritidis O-antigen was studied by isothermal titratio n calorimetry and stopped-flow spectrofluorimetry. At room temperature the binding reaction is enthalpically driven with an unfavorable change in entr opy. A large change of -1.8 +/- 0.2 kJ mol(-1) K-1 in heat capacity suggest s that the hydrophobic effect and water reorganization contribute substanti ally to complex formation, As expected from the large heat-capacity change, we found enthalpy-entropy compensation, The calorimetrically measured bind ing enthalpies were identical within error to van't Hoff enthalpies determi ned fi om fluorescence titrations. Binding kinetics were determined:lt temp eratures ranging from 10 to 30 degreesC, The second-order association rate constant varied from 1 x 10(5) M-1 s(-1) for dodecasaccharide at 10 degrees C to 7 x 10(5) M-1 s(-1) for octasaccharids at 30 degreesC. The first-order dissociation rate constants ranged from 0.2 to 3.8 s(-1). The Arrhenius ac tivation energies were close to 50 and 100 kJ mol(-1) for the association a nd dissociation reactions, respectively, indicating mainly enthalpic barrie rs, Despite the fact that this system is quire complex due to the flexibili ty of the saccharide, both the thermodynamic and kinetic data are compatibl e with a simple one-step binding model.