Amino acid residues that influence FcεRI-mediated effector functions of human immunoglobulin E

Ian Sayers, Stuart A. Cain, Jillian R M Swan, Mark A. Pickett, Peter J. Watt, Stephen T. Holgate, Eduardo A. Padlan, Peter Schuck, Birgit A. Helm

    Research output: Contribution to journalArticlepeer-review

    Abstract

    Immunoglobulin E (IgE) mediates its effector functions via the Fc region of the molecule. IgE binding to and subsequent aggregation of the high- affinity receptor (FcεRI) by allergen plays a pivotal role in type I hypersensitivity responses. Earlier studies implicated the Cε2 and 3 interface and the A-B loop in Cε3 in the IgE-FcεRI interaction. These regions and glycosylation sites in Cε3 were now targeted by site-specific mutagenesis. IgE binding to FcεRI was compared with surface plasmon resonance (SPR) measurements, which assessed the binding of the soluble extracellular domain of FcεRI to IgE. Kinetic analysis based on a pseudo- first-order model agrees with previous determinations. A more refined SPR- based kinetic analysis suggests a biphasic interaction. A model-free empirical analysis, comparing the binding strength and kinetics of native and mutant forms of IgE, identified changes in the kinetics of IgE-FcεRI interaction. Conservative substitutions introduced into the A-B loop have a small effect on binding, suggesting that the overall conformation of the loop is important for the complementary interaction, but multiple sites across the Cε3 domain may influence IgE-FcεRI interactions. Asn394 is essential for the generation of a functional IgE molecule in mammalian cells. A role of Pro333 in the maintenance of a constrained conformation at the interface between Ce2-3 emerged by studying the functional consequences of replacing this residue by Ala and Gly. These substitutions cause a dramatic decrease in the ability of the ligand to mediate stimulus secretion coupling, although only small changes in the association and dissociation rates are observed. Understanding the molecular basis of this phenomenon may provide important information for the design of inhibitors of mast cell degranulation.
    Original languageEnglish
    Pages (from-to)16152-16164
    Number of pages12
    JournalBiochemistry
    Volume37
    Issue number46
    DOIs
    Publication statusPublished - 17 Nov 1998

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