Description:
<jats:p>Recent studies revealed an immunoregulatory role of natural IgG-anti-F(ab′)<jats:sub>2</jats:sub>antibodies in both healthy individuals and patients with certain diseases. The implication of anti-F(ab′)<jats:sub>2</jats:sub>antibodies in the pathogenesis of diseases prompted us to study the gene segment structure of their antigen-binding domains and their binding characteristics. cDNA was prepared from the lymphocytes of a patient with a high IgG-anti-F(ab′)<jats:sub>2</jats:sub>serum titer. Variable heavy and light gene segments were amplified by PCR and inserted into a phagemid surface expression vector. Single-chain antibodies displayed on the phage surface were screened for binding to F(ab′)<jats:sub>2</jats:sub>fragments. The subsequent analysis of 95 single clones demonstrated that they all bound specifically to F(ab′)<jats:sub>2</jats:sub>. Sequence analyses of 12 clones showed that 11 were identical and 1 contained a silent point mutation in the heavy chain and three amino acid exchanges in the light chain. The heavy chains belonged to the V<jats:sub>H</jats:sub>3 and the light chains to the V<jats:sub>κ</jats:sub>2 gene family. The 11 identical light-chain genes were completely homologous to a germ-line sequence (DPK-15). Binding assays showed that the single-chain antibodies bind to F(ab′)<jats:sub>2</jats:sub>, but not to Fab, Fc, or intact IgG. This binding pattern was confirmed by surface plasmon resonance studies, which revealed a relatively high affinity (<jats:italic>K</jats:italic><jats:sub>a</jats:sub>= 2.8 × 10<jats:sup>7</jats:sup>M<jats:sup>−1</jats:sup>). The strong binding capacity was further demonstrated by competitive inhibition of the serum anti-IgG antibody’s interaction with antigen. The present study defines for the first time to our knowledge the gene segment structure of the antigen-binding domain of two human IgG-anti-F(ab′)<jats:sub>2</jats:sub>autoantibody clones and describes the binding kinetics of the purified monomeric fragments.</jats:p>