Structural evidence for entropic contribution of salt bridge formation to a protein antigen-antibody interaction - The case of hen lysozyme-HyHEL-10 Fv complex

A structural and thermodynamic study of the entropic contribution of salt b ridge formation to the interaction between hen egg white lysozyme (HEL) and the variable domain fragment (Fv) of anti-MEL antibody, HyMEL-10, was carr ied out. Three Fv mutants (HD32A, HD96A, and MD32AD96A) were prepared, and the interactions between the mutant Fvs and HEL were investigated. Crystall ography revealed that the overall structures of these mutant complexes were almost identical to that of wildtype Fv, Little structural changes were ob served in the HD32AD96A mutant-MEL complex, and two water molecules were in troduced into the mutation site, indicating that the two water molecules st ructurally compensated for the complete removal of the salt bridges. This r esult suggests that the entropic contribution of the salt bridge originates from dehydration. In the singly mutated complexes, one water molecule was also introduced into the mutated site, bridging the antigen-antibody interf ace. However, a local structural difference was observed in the MD32A FV-HE L complex, and conformational changes occurred due to changes in the relati ve orientation of the heavy chain to the light chain upon complexation in H D96A Fv-HEL complexes. The reduced affinity of these single mutants for the antigen originates from the increase in entropy loss, indicating that thes e structural changes also introduced an increase in entropy loss. These res ults suggest that salt bridge formation makes an entropic contribution to t he protein antigen-antibody interaction through reduction of entropy loss d ue to dehydration and structural changes.