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Purification and immunoglobulin E-binding properties of peanut allergen Ara h 6: Evidence for cross-reactivity with Ara h 2

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Author: Koppelman, S.J. · Jong, G.A.H.de · Laaper-Ertmann, M. de · Peeters, K.A.B.M. · Knulst, A.C. · Hefle, S.L. · Knol, E.F.
Type:article
Date:2005
Institution: TNO Kwaliteit van Leven
Source:Clinical and Experimental Allergy, 4, 35, 490-497
Identifier: 238425
doi: doi:10.1111/j.1365-2222.2005.02204.x
Keywords: Nutrition Safety · Innovation Policy · Allergen · Basophil · IgE · Peanut · Protein purification · Skin prick test · allergen · Ara h 2 allergen · Ara h 6 allergen · epitope · immunoglobulin E · monomer · peanut agglutinin · unclassified drug · adult · amino acid sequence · amino terminal sequence · antigen antibody reaction · antigen binding · article · basophil degranulation · carboxy terminal sequence · clinical article · clinical trial · controlled clinical trial · controlled study · cross reaction · double blind procedure · human · human cell · immunoblotting · matrix assisted laser desorption ionization time of flight mass spectrometry · molecular weight · peanut allergy · prick test · priority journal · protein analysis · protein purification · sequence homology · Adult · Albumins · Allergens · Amino Acid Sequence · Antibody Specificity · Basophils · Cross Reactions · Glycoproteins · Humans · Hypersensitivity · Immunoglobulin E · Molecular Weight · Plant Proteins · Recombinant Proteins

Abstract

Background: IgE-binding peanut proteins smaller than 15 kDa were previously identified as potential allergens in the majority of our peanut allergic population. Objective: To characterize the novel allergen in order to determine whether it was similar to one of the thus far identified recombinant peanut allergens (Ara h 1-7). Methods: An IgE-binding protein of <15 kDa was purified and identified via N-terminal sequencing. Its IgE-binding properties were investigated using immunoblotting, basophil degranulation, and skin prick testing. Possible cross-reacting epitopes with other peanut allergens were studied using IgE-immunoblotting inhibition. Results: The purified protein is a monomeric protein with a molecular weight of 14981 Da as determined using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectroscopy. The amino acid sequence of the first 39 N-terminal residues is identical to that of Ara h 6, indicating that the allergen is Ara h 6. It is recognized by 20 out of 29 peanut-allergic patients on IgE-immunoblot, and its potent biological functionality is demonstrated by the degranulation of basophils, even at concentrations below 10 pg/mL, and by positive skin prick reactions. Ara h 6 has homology to Ara h 2, especially in the middle part and at the C-terminal part of the protein. Almost complete inhibition of IgE-Ara h 6 interaction with Ara h 2 demonstrates that at least part of the epitopes of Ara h 6 are cross-reactive with epitopes on Ara h 2. Conclusions: Peanut-derived Ara h 6 is a biologically active allergen recognized by the majority of our peanut-allergic patient population and can be considered a clinically relevant peanut allergen. © 2005 Blackwell Publishing Ltd. Chemicals / CAS: immunoglobulin E, 37341-29-0; Albumins; Allergens; Ara h 2 allergen, Arachis hypogaea; Ara h 6 allergen, Arachis hypogaea; Glycoproteins; Immunoglobulin E, 37341-29-0; Plant Proteins; Recombinant Proteins