RESUMO
Background: Several studies indicate that Der p 7 is an important and clinically relevant allergen of Dermatophagoides pteronyssinus which should be included in vaccines for treatment of house dust mite (HDM) allergy. Aim of this study was to characterize the IgE epitopes of Der p 7. Methods: Recombinant Der p 7 was expressed and purified, analyzed for fold by circular dichroism and tested for its allergenic activity by basophil activation. Seven overlapping, surface-exposed peptides (P1-P7) with a length of 27 to 37 amino acids, which spanned the Der p 7 sequence, were synthesized and tested for IgE reactivity and allergenic activity by basophil activation assay. Carrier-bound peptides were studied for their ability to induce allergen-specific IgG antibodies in rabbits. Peptide-specific antibodies were used to inhibit allergic patients` IgE binding to Der p 7 by ELISA for mapping of IgE epitopes. Results: rDer p 7 showed high allergenic activity comparable with Der p 5, Der p 21, and Der p 23. None of the seven tested peptides showed any IgE reactivity or allergenic activity when tested with HDM- allergic patients indicating lack of sequential IgE epitopes on Der p 7. IgE inhibition experiments using anti-peptide specific IgGs and molecular modeling enabled us to identify discontinuous, conformational IgE epitopes of Der p 7. Conclusion and Clinical Relevance: IgE epitopes of Der p 7 belong to the conformational and discontinuous type whereas sequential Der p 7 peptides lack IgE reactivity. It should thus be possible to construct hypoallergenic vaccines for Der p 7 based on carrier-bound allergen peptides.
Assuntos
Alérgenos/imunologia , Antígenos de Dermatophagoides/imunologia , Proteínas de Artrópodes/imunologia , Epitopos Imunodominantes , Imunoglobulina E/sangue , Pyroglyphidae/imunologia , Hipersensibilidade Respiratória/imunologia , Alérgenos/química , Alérgenos/genética , Animais , Antígenos de Dermatophagoides/química , Antígenos de Dermatophagoides/genética , Proteínas de Artrópodes/química , Proteínas de Artrópodes/genética , Estudos de Casos e Controles , Linhagem Celular Tumoral , Mapeamento de Epitopos , Humanos , Modelos Moleculares , Conformação Proteica , Dobramento de Proteína , Pyroglyphidae/genética , Coelhos , Ratos , Hipersensibilidade Respiratória/sangueRESUMO
Alt a 1 is the major allergen of the fungus Alternaria alternata and can be found in the cell wall of its spores. It is a cysteine linked homodimeric protein with a unique ß-barrel fold as recently revealed by X-ray crystallography. Despite the elucidation of its structure, its biological function remains unknown. For Alternaria-sensitized patients, contact leads to respiratory allergy and in severe cases to asthma-related death. Here we report the sequence-specific Alt a 1 backbone (1)H, (15)N and (13)C chemical shift assignment.