Crystallographic Analysis of Antigen–Antibody Complexes: End-on Insertion of Ligands in Antibodies—CDR3 Loops as Arbiters

Abstract

As the dominant constituents of the active sites, complementarity-determining regions (CDRs) and particularly the CDR3 loops strongly influence the size and shape of this interdomain space. Six sets of CDR3 loops were extracted from our collection of crystal structures and examined for their modes of association. The CDR3 loops of the NC6.8 Fab face each other across a small crevice that is expanded further by end-on insertion of its high-affinity ligand (a trisubstituted guanidine sweet-tasting compound). This wedging event triggers a series of extensive local and transmitted conformational changes. In the 4-4-20 Fab, the CDR3 loops provide scaffolding for the high-affinity binding of fluorescein and shield the ligand from bulk solvent in the interdomain space above and below the very compact binding slot. Constituents of the CDR2 and CDR3 loops of the BV04-01 Fab interact to form “false floors” over potential cavity-type sites and thereby eliminate end-on insertion. Instead, fragments of single-stranded DNA are bound with low affinity in a groove whose course is altered on complex formation by global movements of VHrelative to VLand by local shifts of HCDR3. Trafficking of even small peptide ligands between the V domains of the Pot Fv is prevented by the collapse of the large HCDR3 segment into the residual interdomain space. This protein is better suited to polyreactive binding of a variety of large protein antigens on its external surfaces. By comparison, the space available between the CDR3 loops of the Mcg light-chain dimer is very large. It has proved to be accessible for end-on insertion of peptides and other ligands ranging over seven orders of magnitude in affinity. Recently, an insect neuropeptide hormone, with pGlu as its penetrating agent, has been found to pierce the entire V dimer interface from the entrance of the traditional active site to the solvent pool between the V and C domains. In an Mcg × Hud heterodimer, the Hud CDR3 plays a role similar to that of the Pot H chain and blocks access to the interdomain space by close interactions with the CDR3 of Mcg.