Biochemical Journal

Research article

Mutational epitope analysis of Pru av 1 and Api g 1, the major allergens of cherry (Prunus avium) and celery (Apium graveolens): correlating IgE reactivity with three-dimensional structure

Philipp NEUDECKER, Katrin LEHMANN, Jörg NERKAMP, Tanja HAASE, Andrea WANGORSCH, Kay FÖTISCH, Silke HOFFMANN, Paul RÖSCH, Stefan VIETHS, Stephan SCHEURER

Abstract

Birch pollinosis is often accompanied by adverse reactions to food due to pollen-allergen specific IgE cross-reacting with homologous food allergens. The tertiary structure of Pru av 1, the major cherry (Prunus avium) allergen, for example, is nearly identical with Bet v 1, the major birch (Betula verrucosa) pollen allergen. In order to define cross-reactive IgE epitopes, we generated and analysed mutants of Pru av 1 and Api g 1.0101, the major celery (Apium graveolens) allergen, by immunoblotting, EAST (enzyme allergosorbent test), CD and NMR spectroscopy. The mutation of Glu45 to Trp45 in the P-loop region, a known IgE epitope of Bet v 1, significantly reduced IgE binding to Pru av 1 in a subgroup of cherry-allergic patients. The backbone conformation of Pru av 1 wild-type is conserved in the three-dimensional structure of Pru av 1 Trp45, demonstrating that the side chain of Glu45 is involved in a cross-reactive IgE epitope. Accordingly, for a subgroup of celery-allergic patients, IgE binding to the homologous celery allergen Api g 1.0101 was enhanced by the mutation of Lys44 to Glu. The almost complete loss of IgE reactivity to the Pru av 1 Pro112 mutant is due to disruption of its tertiary structure. Neither the mutation Ala112 nor deletion of the C-terminal residues 155–159 influenced IgE binding to Pru av 1. In conclusion, the structure of the P-loop partially explains the cross-reactivity pattern, and modulation of IgE-binding by site-directed mutagenesis is a promising approach to develop hypo-allergenic variants for patient-tailored specific immunotherapy.

  • allergen structure
  • allergy
  • cross-reactivity
  • hypoallergenic mutants
  • IgE-epitope analysis

Footnotes

  • 2 Present address: JASCO, Robert-Bosch-Strasse 11, 64823 Gross-Umstadt, Germany.

  • 3 Present address: Forschungszentrum Juelich, IBI-2, 52425 Juelich, Germany, and Institut fuer Physikalische Biologie, Heinrich-Heine-Universitaet, 40225 Duesseldorf, Germany.

  • Abbreviations used: 1D/2D/3D, one-/two-/three-dimensional; EAST, enzyme allergosorbent test; HMQC, heteronuclear multiple quantum correlation spectroscopy; HSQC, heteronuclear single-quantum coherence; mAb, monoclonal antibody; OAS, oral allergy syndrome; RMSD, root-mean-square deviation; SIT, specific immunotherapy.