CRYSTAL-STRUCTURE OF A DIABODY, A BIVALENT ANTIBODY FRAGMENT

Background: Diabodies are dimeric antibody fragments. In each polypeptide, a heavy-chain variable domain (V-H) is linked to a light-chain variable domain (V-L) but unlike single-chain Fv fragments, each antigen-binding site is formed by pairing of one V-H and one V-L domain from the two different polypeptides. Diabodies thus have two antigen-binding sites, and can be bispecific. Direct structural evidence is lacking for the connections and dimeric interactions between the two polypeptides of the diabody. Results: The 2.6 Angstrom resolution structure has been determined for a bivalent diabody with a flexible five-residue polypeptide linker between the (amino-terminal) V-H and (carboxy-terminal) V-L domains. The asymmetric unit of the crystal consists of four polypeptides comprising two diabodies; for one of these polypeptides the linker can be traced between the V-H and V-L domains. Within each diabody the two associated V-H and V-L domains make back-to-back interactions through the V-H domains, and there is an extensive V-L-V-L interface between the two diabodies in the asymmetric unit. Conclusions: The structure of the diabody is very similar to that which had been predicted by molecular modelling. Diabodies directed against cell-surface antigens should be capable of bringing together two cells, such as in cell-targeted therapy, because the two antigen-binding sites of the diabody are at opposite ends of the molecule and separated by similar to 65 Angstrom.