title: Enlarging the toolbox for allergen epitope definition with an allergen-type model protein.
abstract: BACKGROUND
  Birch pollen-allergic subjects produce polyclonal cross-reactive IgE antibodies that mediate pollen-associated food allergies. The major allergen Bet v 1 and its homologs in plant foods bind IgE in their native protein conformation. Information on location, number and clinical relevance of IgE epitopes is limited. We addressed the use of an allergen-related protein model to identify amino acids critical for IgE binding of PR-10 allergens.
  
  
  METHODS
  Norcoclaurine synthase (NCS) from meadow rue is structurally homologous to Bet v 1 but does not bind Bet v 1-reactive IgE. NCS was used as the template for epitope grafting. NCS variants were tested with sera from 70 birch pollen allergic subjects and with monoclonal antibody BV16 reported to compete with IgE binding to Bet v 1.
  
  
  RESULTS
  We generated an NCS variant (Δ29NCSN57/I58E/D60N/V63P/D68K) harboring an IgE epitope of Bet v 1. Bet v 1-type protein folding of the NCS variant was evaluated by 1H-15N-HSQC NMR spectroscopy. BV16 bound the NCS variant and 71% (50/70 sera) of our study population showed significant IgE binding. We observed IgE and BV16 cross-reactivity to the epitope presented by the NCS variant in a subgroup of Bet v 1-related allergens. Moreover BV16 blocked IgE binding to the NCS variant. Antibody cross-reactivity depended on a defined orientation of amino acids within the Bet v 1-type conformation.
  
  
  CONCLUSIONS
  Our system allows the evaluation of patient-specific epitope profiles and will facilitate both the identification of clinically relevant epitopes as biomarkers and the monitoring of therapeutic outcomes to improve diagnosis, prognosis, and therapy of allergies caused by PR-10 proteins.
authors.affiliation: Department of Biopolymers, University of Bayreuth, Bayreuth, Bavaria, Germany.
authors.forename: Hanna
authors.initials: H
authors.lastname: Berkner
authors.affiliation: Department of Biopolymers, University of Bayreuth, Bayreuth, Bavaria, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Seutter von Loetzen
authors.affiliation: Department of Biopolymers, University of Bayreuth, Bayreuth, Bavaria, Germany.
authors.forename: Maximilian
authors.initials: M
authors.lastname: Hartl
authors.affiliation: Division of Allergology, Paul-Ehrlich-Institut, Langen, Hesse, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Randow
authors.affiliation: Division of Allergology, Paul-Ehrlich-Institut, Langen, Hesse, Germany.
authors.forename: Michaela
authors.initials: M
authors.lastname: Gubesch
authors.affiliation: Division of Allergology, Paul-Ehrlich-Institut, Langen, Hesse, Germany.
authors.forename: Lothar
authors.initials: L
authors.lastname: Vogel
authors.affiliation: Division of Allergology, Paul-Ehrlich-Institut, Langen, Hesse, Germany.
authors.forename: Felix
authors.initials: F
authors.lastname: Husslik
authors.affiliation: Division of Allergology, Paul-Ehrlich-Institut, Langen, Hesse, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Reuter
authors.affiliation: ImmunoDiagnostics Division, Thermo Fisher Scientific Inc., Uppsala, Uppsala län, Sweden.
authors.forename: Jonas
authors.initials: J
authors.lastname: Lidholm
authors.affiliation: Department of Dermatology, Allergy Unit, University Hospital Zürich, Zürich, Zürich, Switzerland.
authors.forename: Barbara
authors.initials: B
authors.lastname: Ballmer-Weber
authors.affiliation: Division of Allergology, Paul-Ehrlich-Institut, Langen, Hesse, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Vieths
authors.affiliation: Department of Biopolymers, University of Bayreuth, Bayreuth, Bavaria, Germany.
authors.forename: Paul
authors.initials: P
authors.lastname: Rösch
authors.affiliation: Division of Allergology, Paul-Ehrlich-Institut, Langen, Hesse, Germany.
authors.forename: Dirk
authors.initials: D
authors.lastname: Schiller
mesh_terms: D000485:Allergens / Q000737:chemistry / Q000276:immunology*; D000595:Amino Acid Sequence; D052179:Antigens, Plant / Q000737:chemistry / Q000276:immunology*; D001665:Binding Sites; D003429:Cross Reactions / Q000276:immunology; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000276:immunology; D010940:Plant Proteins / Q000737:chemistry / Q000276:immunology*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid
chemical_list: D000485:Allergens; D052179:Antigens, Plant; D000939:Epitopes; D007074:Immunoglobulin G; D050505:Mutant Proteins; D010940:Plant Proteins; C061312:Bet v 1 allergen, Betula; D007073:Immunoglobulin E