Specificity of IgE antibodies to sequential epitopes of hen’s egg ovomucoid as a marker for persistence of egg allergy (1)
Allergic reactions are more frequently elicited by egg white proteins than egg yolk. Egg white contains 24 different glycoproteins. Ovomucoid (Gal d1), ovalbumin (Gal d2), ovotransferrin (Gal d3), and lysozyme (Gal d4), comprising 10%, 54%, 12%, and 3.5% of total egg white protein, have been identified as major allergens. The importance of ovomucoid may be due to its unique characteristics such as relative stability against heat and digestion with proteinases, and its strong allergenicity. Ovomucoid is a 28 kDa glycoprotein comprised of 186 amino acids (AAs) arranged in three tandem domains containing nine intra-domain, but no inter-domain, disulfide bonds and five carbohydrate side-chains. Evaluation of IgE antibody binding to linearized ovomucoid (i.e. reduced and alkylated) suggested that patients ovomucoid-specific IgE antibodies differed in their ability to bind sequential and conformational epitopes. Sequential epitopes are determined by contiguous AAs, and conformational (discontinuous) epitopes are comprised of AA from different regions of the allergen brought in close proximity by folding of the polypeptide backbone.
Children with persistent allergy had significantly higher levels of IgE against linearized and native ovomucoid than those with transient allergy. Children with persistent egg allergy had a higher level of specific IgE antibodies to linearized and native ovalbumin. However, the level of IgE antibodies against linearized and native ovotransferrin was comparable in patients with persistent and transient egg allergy.
The regions recognized by at least 50% of the patients were defined as major epitopes. Three major IgE-binding epitopes are located in the first domain, and the fourth major epitope was in the second ovomucoid domain. The cumulative OD of IgG binding of egg-allergic patients to each of the peptides, with the number of patients and controls binding to each region shown in parentheses.
None of the patients with transient egg allergy had IgE antibodies against these epitopes, whereas all of the seven patients with persistent allergy possessed such antibodies. Allergenic areas at AA 41–50, 59–68, 131–140 and 151–160, the first two of which were identified as T-cell epitopes, but not IgE-binding epitopes.
IgE antibodies produced by B cells activated following surface-IgM binding to exposed oligopeptides on the native protein may be directed at sequential epitopes determined by contiguous AAs or conformational epitopes, comprised of AA residues from different regions of the allergen brought into proximity by folding. When patients were first found to have clinicahypersensitivity, those with long-lasting egg allergy had relatively larger proportions of IgE antibodies directed at linearized ovomucoid and ovalbumin than to conformational protein compared with the children who subsequently gained clinical tolerance. The more permeable infant gut would allow significant quantities of macromolecular antigen (and more likely conformationally intact food protein) to gain access to local B cells that upon activation generate egg-specific IgE antibodies in genetically predisposed hosts.