IDENTIFICATION OF CONTACT RESIDUES IN THE IGE BINDING-SITE OF HUMAN FC-EPSILON-RI-ALPHA

The high-affinity receptor for immunoglobulin E (IgE), Fc epsilon RI, is an alpha beta gamma(2) tetramer found on mast cells, basophils, and several other types of immune effector cells. The interaction of IgE with the alpha-subunit of Fc epsilon RI is central to the pathogenesis of allergy. Detailed knowledge of the mode of interaction of Fc epsil on RI with IgE may facilitate the development of inhibitors for genera l use in the treatment of allergic disease. To this end we have perfor med site-directed mutagenesis on a soluble form of the Fc epsilon RI a lpha-chain (sFc epsilon RI alpha). The effects of four mutations in th e second immunoglobulin-like domain of sFc epsilon RI alpha upon the k inetics of binding to IgE and fragments of IgE have been analyzed usin g surface plasmon resonance. As described in the preceding paper of th is issue [Henry, A. J., et al. (1997) Biochemistry 36, 15568-15578], b iphasic binding kinetics was observed. Two of the mutations had signif icant effects on binding: K117D reduced the affinity of sFc epsilon RI alpha for IgE by a factor of 30, while D159K increased the affinity f or IgE by a factor of 7, both principally through changes in the rates of dissociation of the slower phase of the interaction. Circular dich roism spectra of sFc epsilon RI alpha incorporating either of these mu tations were indistinguishable from those of wild-type sFc epsilon RI alpha, demonstrating that the native conformation had not been disrupt ed. Our results, together with those from site-directed mutagenesis on fragments of IgE presented in the accompanying paper, define the cont act surfaces in the IgE:sFc epsilon RI alpha complex.