The role of homophilic binding in anti-tumor antibody R24 recognition of molecular surfaces - Demonstration of an intermolecular beta-sheet interaction between V-H domains

The murine antibody R24 and mouse-human Fv-IgG1(kappa) chimeric antibody ch R24 are specific for the cell-surface tumor antigen disialoganglioside GD3. X-ray diffraction and surface plasmon resonance experiments have been empl oyed to study the mechanism of "homophilic binding," in which molecules of R24 recognize and bind to other molecules of R24 though their heavy chain v ariable domains. R24 exhibits strong binding to liposomes containing disial oganglioside GD3; however, the kinetics are unusual in that saturation of b inding is not observed. The binding of chR24 to GD3-bearing liposomes is si gnificantly weaker, suggesting that cooperative interactions involving anti body constant regions contribute to R24 binding of membrane-bound GD3. The crystal structures of the Fabs from R24 and chR24 reveal the mechanism for hemophilic binding and confirm that the hemophilic and antigen-binding idio topes are distinct. The hemophilic binding idiotope is formed largely by an anti-parallel beta-sheet dimerization between the H2 complementarity deter mining region (CDR) loops of two Fabs, while the antigen-binding idiotope i s a pocket formed by the three CDR loops on the heavy chain. The formation of hemophilic dimers requires the presence of a canonical conformation for the H2 CDR in conjunction with participation of side chains. The relative p ositions of the homophilic and antigen-binding sites allows for a lattice o f GD3-specific antibodies to be constructed, which is stabilized by the pre sence of the cell membrane. This model provides for the selective recogniti on by R24 of cells that overexpress GD3 on the cell surface.