Shiroishi M, Yokota A, Tsumoto K, Kondo H, Nishimiya Y, Horii K, Matsushima M, Ogasahara K, Yutani K, Kumagai I
Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University, Aoba-yama 07, Aoba-ku, Sendai 980-8579, Japan.
J Biol Chem. 2001 Jun 22;276(25):23042-50. doi: 10.1074/jbc.M100480200. Epub 2001 Apr 10.
A structural and thermodynamic study of the entropic contribution of salt bridge formation to the interaction between hen egg white lysozyme (HEL) and the variable domain fragment (Fv) of anti-HEL antibody, HyHEL-10, was carried out. Three Fv mutants (HD32A, HD96A, and HD32AD96A) were prepared, and the interactions between the mutant Fvs and HEL were investigated. Crystallography revealed that the overall structures of these mutant complexes were almost identical to that of wild-type Fv. Little structural changes were observed in the HD32AD96A mutant-HEL complex, and two water molecules were introduced into the mutation site, indicating that the two water molecules structurally compensated for the complete removal of the salt bridges. This result 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 interface. However, a local structural difference was observed in the HD32A Fv-HEL complex, and conformational changes occurred due to changes in the relative orientation of the heavy chain to the light chain upon complexation in HD96A Fv-HEL complexes. The reduced affinity of these single mutants for the antigen originates from the increase in entropy loss, indicating that these structural changes also introduced an increase in entropy loss. These results suggest that salt bridge formation makes an entropic contribution to the protein antigen-antibody interaction through reduction of entropy loss due to dehydration and structural changes.
对盐桥形成对鸡蛋清溶菌酶（HEL）与抗HEL抗体HyHEL-10的可变结构域片段（Fv）之间相互作用的熵贡献进行了结构和热力学研究。制备了三个Fv突变体（HD32A、HD96A和HD32AD96A），并研究了突变体Fv与HEL之间的相互作用。晶体学研究表明，这些突变体复合物的整体结构与野生型Fv几乎相同。在HD32AD96A突变体-HEL复合物中观察到的结构变化很小，并且在突变位点引入了两个水分子，这表明这两个水分子在结构上补偿了盐桥的完全去除。该结果表明盐桥的熵贡献源于脱水。在单突变复合物中，也有一个水分子被引入突变位点，桥接抗原-抗体界面。然而，在HD32A Fv-HEL复合物中观察到局部结构差异，并且在HD96A Fv-HEL复合物中，由于复合物形成时重链与轻链相对取向的变化而发生了构象变化。这些单突变体对抗原亲和力的降低源于熵损失的增加，这表明这些结构变化也导致了熵损失的增加。这些结果表明，盐桥的形成通过减少脱水和结构变化引起的熵损失，对蛋白质抗原-抗体相互作用做出了熵贡献。
