Modification of peanut allergen Ara h 3: effects on IgE binding and T cell stimulation.
Int Arch Allergy Immunol
; 128(1): 15-23, 2002 May.
Article
en En
| MEDLINE
| ID: mdl-12037397
BACKGROUND: Peanut allergy is a major health concern due to the increased prevalence, potential severity, and chronicity of the reaction. The cDNA encoding a third peanut allergen, Ara h 3, has been previously cloned and characterized. Mutational analysis of the Ara h 3 IgE-binding epitopes with synthetic peptides revealed that single amino acid changes at critical residues could diminish IgE binding. METHODS: Specific oligonucleotides were used in polymerase chain reactions to modify the cDNA encoding Ara h 3 at critical IgE binding sites. Four point mutations were introduced into the Ara h 3 cDNA at codons encoding critical amino acids in epitopes 1, 2, 3 and 4. Recombinant modified proteins were used in SDS-PAGE/Western IgE immunoblot, SDS-PAGE/Western IgE immunoblot inhibition and T cell proliferation assays to determine the effects of these changes on in vitro clinical indicators of peanut hypersensitivity. RESULTS: Higher amounts of modified Ara h 3 were required to compete with the wild-type allergen for peanut-specific serum IgE. Immunoblot analysis with individual serum IgE from Ara-h-3-allergic patients showed that IgE binding to the modified protein decreased approximately 35-85% in comparison to IgE binding to wild-type Ara h 3. Also, the modified Ara h 3 retained the ability to stimulate T cell activation in PBMCs donated by Ara-h-3-allergic patients. CONCLUSIONS: The engineered hypoallergenic Ara h 3 variant displays two characteristics essential for recombinant allergen immunotherapy; it has a reduced binding capacity for serum IgE from peanut-hypersensitive patients and it can stimulate T-cell proliferation and activation.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Alérgenos
Tipo de estudio:
Risk_factors_studies
Límite:
Humans
Idioma:
En
Revista:
Int Arch Allergy Immunol
Asunto de la revista:
ALERGIA E IMUNOLOGIA
Año:
2002
Tipo del documento:
Article
País de afiliación:
Estados Unidos
Pais de publicación:
Suiza