Epitope recognition patterns of Bet v 1-specific IgE

Birch pollen is one of the most prevalent elicitors of allergy in Europe. Up to 70% of birch pollen allergic patients show adverse reactions to plant foods such as hazelnut, apple, stone fruits, celeriac, carrot and soy milk. In most cases, this is a consequence of sensitization to the major birch pollen allergen, Bet v 1, and subsequent cross-reactivity of immunoglobulin E (IgE) antibodies to Bet v 1- related allergens in plant foods. Most patients, however, tolerate many typical birch pollen-related foods, even if their IgE binds to a broad range of Bet v 1-related food allergens. The varying clinical relevance of IgE reactivity to Bet v 1-related food allergens may be a consequence of patient-specific IgE binding patterns to different sites (epitopes) on the surface of Bet v 1. The aim of this project is the identification of these epitopes and the elucidation of their importance for birch pollen associated food allergy. To this end, chimeric proteins will be created by transferring surface patches of Bet v 1 onto the surface of a non-IgE binding structural Bet v 1 homologue. These proteins, each carrying a single potential epitope, will be tested for binding of IgE and other antibody classes from sera of birch pollen allergic patients. Moreover, they will be used to select epitope-specific recombinant antibodies from a phage library. These antibodies will be used to study the contributions of single epitopes to IgE binding and cross-reactivity of Bet v 1. The contribution of the epitopes to the allergenic activity of Bet v 1 will be examined by measuring the activation of basophil granulocytes by binding of Bet v 1 to cell-bound patients IgE and by inhibiting this interaction with chimeric proteins or epitope-specific antibodies. This project will be the first that aims to obtain a representative overview of the epitope recognition patterns of Bet v 1-specific IgE in a large group of birch pollen-allergic patients with different profiles of food allergies. Results of the project will enable the development of epitope-based diagnostic reagents that may allow clinicians to give birch pollen allergic patients more precise dietary advice.

This project aimed to determine the binding sites of allergen-inducing immunoglobulin E (IgE) antibodies and potentially allergen-inhibiting IgG antibodies on the major birch pollen allergen, Bet v 1. Birch pollen is one of the most prevalent elicitors of respiratory allergy in Europe. Up to 70% of birch pollen allergic patients show adverse reactions to plant foods such as hazelnut, apple, stone fruits, celeriac, carrot and soy milk. In most cases, this is a consequence of induction of IgE antibodies directed to Bet v 1 and subsequent cross-reactions of those antibodies with Bet v 1-related allergens in plant foods. Most patients, however, tolerate many typical birch pollen-related foods, even if their IgE binds to a broad range of Bet v 1-related food allergens. In this project, we determined the binding sites of Bet v 1-specific IgE and IgG on the surface of the allergen. To this end, we produced 13 artificial proteins, each presenting a distinct part of the Bet v 1 surface and then tested sera from birch pollen allergic patients for the presence of specific antibodies. We showed that each patient contains an individual mixture of Bet v 1-specific IgE and IgG antibodies and that the majority of patients contained antibodies specific to several different parts of the allergen. Moreover, in most cases, IgG binds to different sites than IgE from the same patient. The method developed in this project can now be used to analyze serum samples from a large group of birch pollen allergic patients with well-characterized patterns of plant food allergies. This analysis will answer the question whether allergic reactions to different plant foods are associated with different binding patterns of Bet v 1-specific IgE and IgG, which might lead to novel, more precise diagnostic reagents. An additional potential application of the method developed in this project is monitoring the evolution of the Bet v 1-spefic antibody response during birch pollen immunotherapy. This may shed light on the mechanism of allergen immunotherapy and might help in developing clinical markers to predict therapeutic success already in an early stage of the therapy.