The high affinity receptor for IgE, FcεRI, binds IgE through the second Ig-like domain of the α subunit. The role of the first Ig-like domain is not well understood, but it is required for optimal binding of IgE to FcεRI, either through a minor contact interaction or in a supporting structural capacity. The results reported here demonstrate that domain one of FcεRI plays a major structural role supporting the presentation of the ligand- binding site, by interactions generated within the interdomain interface. Analysis of a series of chimeric receptors and point mutants indicated that specific residues within the A' strand of domain one are crucial to the maintenance of the interdomain interface, and IgE binding. Mutation of the Arg15 and Phe17 residues caused loss in ligand binding, and utilizing a homology model of FcεRI-α based on the solved structure of FcγRIIa, it appears likely that this decrease is brought about by collapse of the interface and consequently the IgE-binding site. In addition discrepancies in results of previous studies using chimeric IgE receptors comprising FcεRIα with either FcγRIIa or FcγRIIIA can be explained by the presence or absence of Arg15 and its influence on the IgE-binding site. The data presented here suggest that the second domain of FcεRI-α is the only domain involved in direct contact with the IgE ligand and that domain one has a structural function of great importance in maintaining the integrity of the interdomain interface and, through it, the ligand-binding site.