title: Peanut defensins: Novel allergens isolated from lipophilic peanut extract.
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.
authors.affiliation: 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.
authors.forename: Arnd
authors.initials: A
authors.lastname: Petersen
authors.affiliation: 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.
authors.forename: Skadi
authors.initials: S
authors.lastname: Kull
authors.affiliation: 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.
authors.forename: Sandra
authors.initials: S
authors.lastname: Rennert
authors.affiliation: 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.
authors.forename: Wolf-Meinhard
authors.initials: WM
authors.lastname: Becker
authors.affiliation: 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.
authors.forename: Susanne
authors.initials: S
authors.lastname: Krause
authors.affiliation: Division of Immune Cell Analytics, Research Center Borstel, Borstel, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Ernst
authors.affiliation: Division of Biophysics, Research Center Borstel, Borstel, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Gutsmann
authors.affiliation: Animal Hygiene, Technische Universität München, Freising-Weihenstephan, Freising, Germany.
authors.forename: Johann
authors.initials: J
authors.lastname: Bauer
authors.affiliation: Division of Bioanalytical Chemistry, Research Center Borstel, Borstel, Germany.
authors.forename: Buko
authors.initials: B
authors.lastname: Lindner
authors.affiliation: 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.
authors.forename: Uta
authors.initials: U
authors.lastname: Jappe
mesh_terms: D000485:Allergens / Q000276:immunology* / Q000302:isolation & purification; D000528:Alternaria / Q000187:drug effects; D000935:Antifungal Agents / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology; D010367:Arachis / Q000276:immunology*; D001491:Basophils / Q000276:immunology*; D023082:Defensins / Q000276:immunology* / Q000302:isolation & purification / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015151:Immunoblotting; D007073:Immunoglobulin E / Q000378:metabolism; D013058:Mass Spectrometry; D021183:Peanut Hypersensitivity / Q000175:diagnosis / Q000276:immunology*; D010936:Plant Extracts / Q000276:immunology* / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid
chemical_list: D000485:Allergens; D000935:Antifungal Agents; D023082:Defensins; D010936:Plant Extracts; D007073:Immunoglobulin E
keywords: Antifungal activity; Ara h 12; Ara h 13; food allergy; lipophilic extraction; peanut allergy; plant defensin