Altered IgE epitope presentation: A model for hypoallergenic activity revealed for Bet v 1 trimer
| Autor: | Margarete Focke-Tejkl, Susanne Spitzauer, Josef Thalhamer, Ines Swoboda, Katharina Blatt, Harald Herrmann, Susanne Vrtala, Domen Zafred, Peter Valent, Anna Gieras, Rudolf Valenta, Yuliya Dall’Antonia, Walter Keller, Raffaela Campana, Bernhard Maderegger, Sandra Scheiblhofer, Angela Neubauer |
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| Rok vydání: | 2009 |
| Předmět: |
Circular dichroism
Immunology Trimer Enzyme-Linked Immunosorbent Assay Immunoglobulin E medicine.disease_cause Article law.invention Cell Line Epitopes Allergen law Antibody Specificity medicine Hypersensitivity Animals Humans Pyrophosphatases Protein Structure Quaternary Molecular Biology Antigen Presentation biology Chemistry Phosphoric Diester Hydrolases Models Immunological Antibodies Monoclonal Hypoallergenic Antigens Plant Flow Cytometry Recombinant Proteins beta-N-Acetylhexosaminidases Basophils Rats Up-Regulation Solutions Basophil activation Biochemistry Immunoglobulin G biology.protein Recombinant DNA Antibody |
| Zdroj: | Molecular immunology. 48(4) |
| ISSN: | 1872-9142 |
| Popis: | In order to reduce side effects in the course of allergen specific immunotherapy hypoallergenic allergen derivatives with reduced IgE reactivity have been made by genetic engineering. In contrast to other recombinant hypoallergenic allergen derivatives which showed reduced IgE reactivity, a recombinant trimer of the major birch pollen allergen Bet v 1 showed reduced allergenic activity despite preserved IgE reactivity. We studied rBet v 1 trimer by SDS-PAGE, mass spectrometry, circular dichroism and gel filtration. Furthermore we investigated IgE and IgG reactivity of the rBet v 1 trimer in solid and liquid phase assays and compared its allergenic activity with that of rBet v 1 wildtype using basophil activation assays. In solid phase immunoassays rBet v 1 trimer exhibited even stronger IgE reactivity than the rBet v 1 wildtype, whereas both proteins were equally well recognized by Bet v 1-specific IgG antibody probes. In fluid phase IgE experiments rBet v 1 trimer inhibited IgE reactivity to rBet v 1 wildtype but showed a more than 10-fold reduced allergenic activity compared to the rBet v 1 monomer. By analytical gel filtration it was demonstrated that, despite its monomeric appearance in SDS-PAGE the trimer occurred in fluid phase in the form of defined high molecular weight (>600 kDa) aggregates whereas rBet v 1 wildtype strictly appeared as monomeric protein. The results indicate that the hypoallergenic nature of the rBet v 1 trimer is due to formation of defined high molecular weight aggregates which may be responsible for an altered presentation of IgE epitopes in a form with reduced capacity to crosslink effector-cell bound IgE. We thus provide evidence for a novel mechanism for hypoallergenic activity. |
| Databáze: | OpenAIRE |
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