Definition of Receptor Binding Domains in Interferon-α

Abstract

Earlier studies from this laboratory had identified three regions in interferon-α (IFN-α) that influence the active conformation of the molecule. These domains are associated with the amino acid residues 10–35, 78–107, and 123–166. In this report, we define these domains more accurately by identifying their critical clusters of amino acids. Using a panel of IFN-α_2a variants in antiviral, growth inhibitory, and receptor binding studies, we are able to show that these three domains, defined by residues 29–35, 78–95, and 123–140, are likely located on the surface of the molecule, with domains 29–35 and 123–140 in close spatial proximity. We conclude that the 29–35 and 123–140 domains are responsible for IFN-α receptor binding interactions and constitute receptor recognition sites in IFN-α. Extrapolating from our biological activity data, in the context of a number of predictive algorithms that provide insights into the hydrophobicity/hydrophilicity, surface probability, and flexibility of amino acid clusters, we infer that the residues 29–35 influence the active configuration of IFN-α most significantly. This region likely represents a loop structure that is relatively rigid in configuration. The carboxy-terminally located strategic domain, 123–140, is comprised of two clusters of amino acid residues, one that forms part of a rigid α-helix, the other a more flexible loop structure. Similarly, the 78–95 domain comprises a portion of an α-helical structure that is followed by a loop structure. Close examination of the amino acid sequences in all three regions among the different species of IFN-αs and human IFN-β indicate that the 29–35 and 123–140 domains are most highly conserved, yet some variance is apparent in the 78–95 domain. We propose that the 78–95 region influences species specificity among the murine and human IFN-αs and determines the differential specificity of action between human IFN-α and human IFN-β.