Definition of receptor binding domains in interferon-alpha.
Earlier studies from this laboratory had identified three regions in interferon-alpha (IFN-alpha) 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-alpha 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-alpha receptor binding interactions and constitute receptor recognition sites in IFN-alpha. 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-alpha 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 alpha-helix, the other a more flexible loop structure. Similarly, the 78-95 domain comprises a portion of an alpha-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-alpha s and human IFN-beta 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-alpha s and determines the differential specificity of action between human IFN-alpha and human IFN-beta.
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Department of Microbiology, University of Toronto, Ontario, Canada.
D000595:Amino Acid Sequence; D001665:Binding Sites / Q000502:physiology; D002455:Cell Division / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D006801:Humans; D016898:Interferon-alpha / Q000737:chemistry*; D007455:Iodine; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017471:Receptors, Interferon / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014407:Tumor Cells, Cultured
D016898:Interferon-alpha; D017471:Receptors, Interferon; D007455:Iodine