Steric Relationship of Amino-Terminal and Carboxy-Terminal Domains of Murine Interferon-γ as Assessed by Monoclonal Antibodies

Spleen cells from a hamster immunized with murine interferon-γ (IFN-γ) carboxy-terminal (95-133) synthetic peptide [IFN-γ(95-133)] conjugated to keyhole limpet hemocyanin (KLH) were fused with mouse myeloma cells, resulting in the production of an anti-IFN-γ(95-133) monoclonal antibody, which reacted with IFN-γ. This monoclonal antibody bound [125I]IFN-γ in a dose-dependent and reversible fashion, and neutralized the antiviral and macrophage priming activities of IFN-γ. Antibody-antibody competition for [125I]IFN-γ binding, using this carboxy-terminal-specific antibody and two previously described amino-terminal-specific monoclonal antibodies, indicated that the carboxy-terminal-specific monoclonal antibody competed only with itself and that the two amino-terminal-specific monoclonal antibodies similarly competed only with each other for [¹²⁵I]IFN-γ. The data suggest that the amino- and carboxy-terminal IFN-γ domains important for function are sterically distant from one another, and suggest the intriguing possibility that interaction of IFN-γ with its receptor may involve more than one binding site on the IFN-γ molecule.