STUDIES ON ANTIGEN-BINDING BY INTACT AND HINGE-DELETED CHIMERIC ANTIBODIES

A matched set of chimeric IgG1 and IgG4 antibodies were used to invest igate the role of the IgG hinge in binding to Ag with differing space between the epitopes. Antibodies bearing identical V regions and eithe r IgG1 or IgG4 C regions were engineered with and without hinges. We m easured the binding of these antibodies to the peptide CYYYEEEEY and t o CYYYEEEEY-BSA conjugates with decreasing numbers of peptides per BSA molecule. We earlier showed that V region differences in antibodies c ould affect Ag binding patterns in solid-phase but not solution-phase assays; however, both types of assay yielded similar results for the h inge-deleted antibodies. Binding of CYYYEEEEY-BSA by hinge-deleted and intact IgG1 was similar, but intact IgG1 bound free peptide better th an did hinge-deleted IgG1. Intact IgG4 antibody bound less well to CYY YEEEEY and CYYYEEEEY-BSA than did IgG1 but, surprisingly, hinge-delete d IgG4 showed better binding than did intact IgG4 and was more like th e IgG1 antibodies in binding affinity. Thus, the IgG4 hinge may impart a structural constraint that prevents high affinity binding to Ag. Th e hinge-deleted IgG4 antibody did not activate C, although it bound Ag similarly to IgG1. This study is the first to address the effect of t he IgG hinge on Ag binding by using well defined Ag with different epi tope densities. Our results may provide an explanation for the apparen t low affinity of IgG4 antibodies.