Structure-Function Study of the Extracellular Domain of the Human IFN-α Receptor (hIFNAR1) Using Blocking Monoclonal Antibodies: The Role of Domains 1 and 2

Abstract

Abstract We have performed a structure-function analysis of extracellular domain regions of the human IFN-α receptor (hIFNAR1) using mAbs generated by immunizing mice with a soluble hIFNAR1-IgG. Five mAbs described in this study recognize different epitopes as determined by a competitive binding ELISA and by alanine substitution mutant analyses of the hIFNAR1-IgG. Two mAbs, 2E1 and 4A7, are able to block IFN-stimulated gene factor 3 (ISGF3) formation and inhibit the antiviral cytopathic effect induced by several IFN-α (IFN-α2/1, -α1, -α2, -α5, and -α8). None of these anti-IFNAR1 mAbs were able to block activity of IFN-β. mAb 4A7 binds to a domain 1-hIFNAR1-IgG but not to a domain 2-hIFNAR1-IgG, which suggests that its binding region is located in domain 1. The binding of the most potent blocking mAb, 2E1, requires the presence of domain 1 and domain 2. The most critical residue for 2E1 binding is a lysine residue at position 249, which is in domain 2. These findings suggest that both domain 1 and domain 2 are necessary to form a functional receptor and that a region in domain 2 is important. IFN-β recognizes regions of the hIFNAR complex that are distinct from those important for the IFN-α.