STRUCTURAL IMMUNO-ANALYSIS OF HUMAN AND PORCINE INTERFERON-GAMMA - IDENTIFICATION OF SHARED ANTIGENIC DOMAIN

We examined the antigenic resemblance between human (h) and porcine (p ) interferon (IFN)-gamma by binding (ELISA) and neutralization assays. The murine polyclonal antisera and sets of murine monoclonal antibodi es (mAbs) raised against either IFN were tested in confrontation with recombinant IFNs of either species, and with site-specific mutants of hIFN-gamma. Several of the mAbs raised against pIFN-gamma cross-reacte d in ELISA with hIFN-gamma. In contrast, none of the anti-hIFN-gamma m Abs cross-reacted. By employing site-specific mutants of recombinant h IFN-gamma as antigens in ELISA we succeeded in identifying the C-termi nal portion 97-111 as the antigenic site in hIFN-gamma recognized by t he cross-reactive anti-pIFN-gamma mAbs. None of the mAbs recognizing t he common antigenic structure had neutralizing potency, although His11 1 was determined by others as the residue important for bioactivy of h IFN-gamma. Mutations in the domain 97-111 had no or little influence o n homospecific reactivity of anti-hIFN-gamma mAbs, indicating that thi s domain, while being mouse-immunodominant in the case of pIFN-gamma w as poorly immunogenic in the case of hIFN-gamma. The epitopes of three out of five anti-hIFN-gamma mAbs mapped in the N-terminal region 1-23 , indicating immunodominance of this region in hIFN-gamma. Another mAb (D9D10), also directed to the N-terminus of hIFN-gamma, apparently re cognized a conformational epitope. This antibody lacked ELISA-reactivi ty with the wild-type hIFN-gamma but strongly bound mutant protein wit h an engineered disulfide bridge Cys7-Cys69. Surprisingly, D9D10 showe d high reactivity also with the wild type hIFN-gamma produced by bacul ovirus construct coding for the mature protein with signal sequence or with the wild type protein possessing residues Cys-Tyr-Cys from the s ignal sequence.