Enlarging the toolbox for allergen epitope definition with an allergen-type model protein.

Hanna Berkner,Andreas Reuter,Stefanie Randow,Paul Rösch,Felix Husslik,Stefan Vieths,Michaela Gubesch,Maximilian Hartl,Barbara Ballmer-Weber,Dirk Schiller,Jonas Lidholm,Lothar Vogel,Christian Seutter Von Loetzen

doi:10.1371/journal.pone.0111691

Hanna Berkner, Andreas Reuter + Show 11 more

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https://doi.org/10.1371/journal.pone.0111691

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Abstract

BackgroundBirch pollen-allergic subjects produce polyclonal cross-reactive IgE antibodies that mediate pollen-associated food allergies. The major allergen Bet v 1 and its homologs in plant foods bind IgE in their native protein conformation. Information on location, number and clinical relevance of IgE epitopes is limited. We addressed the use of an allergen-related protein model to identify amino acids critical for IgE binding of PR-10 allergens.Method Norcoclaurine synthase (NCS) from meadow rue is structurally homologous to Bet v 1 but does not bind Bet v 1-reactive IgE. NCS was used as the template for epitope grafting. NCS variants were tested with sera from 70 birch pollen allergic subjects and with monoclonal antibody BV16 reported to compete with IgE binding to Bet v 1.ResultsWe generated an NCS variant (Δ29NCSN57/I58E/D60N/V63P/D68K) harboring an IgE epitope of Bet v 1. Bet v 1-type protein folding of the NCS variant was evaluated by 1H-15N-HSQC NMR spectroscopy. BV16 bound the NCS variant and 71% (50/70 sera) of our study population showed significant IgE binding. We observed IgE and BV16 cross-reactivity to the epitope presented by the NCS variant in a subgroup of Bet v 1-related allergens. Moreover BV16 blocked IgE binding to the NCS variant. Antibody cross-reactivity depended on a defined orientation of amino acids within the Bet v 1-type conformation.ConclusionOur system allows the evaluation of patient-specific epitope profiles and will facilitate both the identification of clinically relevant epitopes as biomarkers and the monitoring of therapeutic outcomes to improve diagnosis, prognosis, and therapy of allergies caused by PR-10 proteins.

Highlights

Millions of patients with allergies to tree pollen are sensitized to the major allergen of birch (Betula verrucosa) pollen, Bet v 1 [1]
Our system allows the evaluation of patient-specific epitope profiles and will facilitate both the identification of clinically relevant epitopes as biomarkers and the monitoring of therapeutic outcomes to improve diagnosis, prognosis, and therapy of allergies caused by PR-10 proteins
D29NCS variants have Bet v 1-type tertiary structure IgE antibody binding to PR-10 allergens depends on the presence of a native protein fold of the allergen

Summary

Introduction

Millions of patients with allergies to tree pollen are sensitized (produce IgE antibodies) to the major allergen of birch (Betula verrucosa) pollen, Bet v 1 [1]. Bet v 1-binding IgE cross-reacts with Bet v 1-homologous proteins from plant foods, leading to allergy to such foods in a majority of birch pollen-allergic subjects [2]. Structural information on IgE epitopes of Bet v 1 and Bet v 1like allergens in foods is limited. The only structural information on an epitope of Bet v 1 to date was obtained from X-ray crystallography of a complex between the monoclonal Bet v 1-specific mouse IgG BV16 and the major isoform of Bet v 1, Bet v 1 a [20]. Birch pollen-allergic subjects produce polyclonal cross-reactive IgE antibodies that mediate pollen-associated food allergies. The major allergen Bet v 1 and its homologs in plant foods bind IgE in their native protein conformation. We addressed the use of an allergenrelated protein model to identify amino acids critical for IgE binding of PR-10 allergens

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Conclusion