Arnd Petersen1, Skadi Kull2, Sandra Rennert1, Wolf-Meinhard Becker1, Susanne Krause1, Martin Ernst3, Thomas Gutsmann4, Johann Bauer5, Buko Lindner6, Uta Jappe7. 1. Division of Clinical and Molecular Allergology, Research Center Borstel, Airway Research Center North (ARCN), Member of the German Center for Lung Research (DZL), Borstel, Germany. 2. Division of Clinical and Molecular Allergology, Research Center Borstel, Airway Research Center North (ARCN), Member of the German Center for Lung Research (DZL), Borstel, Germany. Electronic address: skull@fz-borstel.de. 3. Division of Immune Cell Analytics, Research Center Borstel, Borstel, Germany. 4. Division of Biophysics, Research Center Borstel, Borstel, Germany. 5. Animal Hygiene, Technische Universität München, Freising-Weihenstephan, Freising, Germany. 6. Division of Bioanalytical Chemistry, Research Center Borstel, Borstel, Germany. 7. Division of Clinical and Molecular Allergology, Research Center Borstel, Airway Research Center North (ARCN), Member of the German Center for Lung Research (DZL), Borstel, Germany; Department of Dermatology, University of Luebeck, Luebeck, Germany.
Abstract
BACKGROUND: Peanut is one of the most hazardous sources of food allergens. Unknown allergens are still hidden in the complex lipophilic matrix. These allergens need to be discovered to allow estimation of the allergenic risk for patients with peanut allergy and to further improve diagnostic measures. OBJECTIVE: We performed detection, isolation, and characterization of novel peanut allergens from lipophilic peanut extract. METHODS: Extraction of roasted peanuts were performed under defined extraction conditions and examined by means of 2-dimensional PAGE. Subsequently, chromatographic methods were adapted to isolate low-molecular-weight components. Proteins were studied by using SDS-PAGE and immunoblotting with sera from patients with peanut allergy. For allergen identification protein sequencing, homology search and mass spectrometry were applied. Functional characterization for allergenicity was performed by using the basophil activation assay and for antimicrobial activity by using inhibition assays of different bacteria and fungi. RESULTS: IgE-reactive proteins of 12, 11, and 10 kDa were first detected after chloroform/methanol extraction in the flow through of hydrophobic interaction chromatography. The proteins were able to activate basophils of patients with peanut allergy. N-terminal sequencing and homology search in the expressed sequence tag database identified the allergens as peanut defensins, which was confirmed by using mass spectrometry. On microbial cell cultures, the peanut defensins showed inhibitory effects on the mold strains of the genera Cladosporium and Alternaria but none on bacteria. CONCLUSIONS: We identified defensins as novel peanut allergens (Ara h 12 and Ara h 13) that react in particular with IgE of patients with severe peanut allergy. Their antimicrobial activity is solely antifungal.
BACKGROUND:Peanut is one of the most hazardous sources of food allergens. Unknown allergens are still hidden in the complex lipophilic matrix. These allergens need to be discovered to allow estimation of the allergenic risk for patients with peanutallergy and to further improve diagnostic measures. OBJECTIVE: We performed detection, isolation, and characterization of novel peanut allergens from lipophilic peanut extract. METHODS: Extraction of roasted peanuts were performed under defined extraction conditions and examined by means of 2-dimensional PAGE. Subsequently, chromatographic methods were adapted to isolate low-molecular-weight components. Proteins were studied by using SDS-PAGE and immunoblotting with sera from patients with peanutallergy. For allergen identification protein sequencing, homology search and mass spectrometry were applied. Functional characterization for allergenicity was performed by using the basophil activation assay and for antimicrobial activity by using inhibition assays of different bacteria and fungi. RESULTS: IgE-reactive proteins of 12, 11, and 10 kDa were first detected after chloroform/methanol extraction in the flow through of hydrophobic interaction chromatography. The proteins were able to activate basophils of patients with peanutallergy. N-terminal sequencing and homology search in the expressed sequence tag database identified the allergens as peanut defensins, which was confirmed by using mass spectrometry. On microbial cell cultures, the peanut defensins showed inhibitory effects on the mold strains of the genera Cladosporium and Alternaria but none on bacteria. CONCLUSIONS: We identified defensins as novel peanut allergens (Ara h 12 and Ara h 13) that react in particular with IgE of patients with severe peanutallergy. Their antimicrobial activity is solely antifungal.
Authors: Rob C Aalberse; Geoffrey A Mueller; Ninotska I L Derksen; Joost A Aalberse; Lori L Edwards; Anna Pomés; Jonas Lidholm; Theo Rispens; Peter Briza Journal: Clin Exp Allergy Date: 2020-02-07 Impact factor: 5.401
Authors: I Pablos; S Eichhorn; Y Machado; P Briza; A Neunkirchner; B Jahn-Schmid; S Wildner; W T Soh; C Ebner; J-W Park; W F Pickl; N Arora; S Vieths; F Ferreira; G Gadermaier Journal: Allergy Date: 2017-09-27 Impact factor: 13.146
Authors: Uta Jappe; Christian Schwager; Andra B Schromm; Nestor González Roldán; Karina Stein; Holger Heine; Katarzyna A Duda Journal: Front Immunol Date: 2019-02-14 Impact factor: 7.561