CHARACTERIZATION OF BIOSYNTHETIC IGG OLIGOMERS RESULTING FROM LIGHT-CHAIN VARIABLE DOMAIN DUPLICATION

We have characterized a human IgG1 monoclonal antibody composed of alt ered light chains. Each light chain consists of two identical variable domains and a kappa constant domain, in association with a normal gam ma chain. This antibody assembled biosynthetically into a mixture of s table oligomers and monomers. Employing gel filtration, PAGE, and elec tron microscopy, we examined the antibody and the nature of the associ ations involved in oligomer formation. By engineering a protease facto r Xa site between the duplicated light chain variable domains and exam ining the fragments produced following factor Xa cleavage, we demonstr ated the association of the IgG monomers occurred through their duplic ated VL domains. Electron microscopy showed the oligomeric antibody to be predominantly dimers and trimers in which the monomeric units were associated through the tips of the Fab portion of the antibody, presu mably through the protuding N-terminal VL domains. Similar examination of monomers demonstrated several molecular forms, including individua l molecules with self-crosslinked Fab arms and others displaying the o pen Y and T shapes typically observed for IgG antibodies. The monomers also displayed distally protruding domain-like structures. The oligom ers produced by this cell line therefore occurred through the nonconva lent interaction between the extra light chain variable domains.