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

摘要

pBirch 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 (iPrunus avium/i) allergen, for example, is nearly identical with Bet v 1, the major birch (iBetula verrucosa/i) 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 (iApium graveolens/i) allergen, by immunoblotting, EAST (enzyme allergosorbent test), CD and NMR spectroscopy. The mutation of Glusup45/sup to Trpsup45/sup 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 Trpsup45/sup, demonstrating that the side chain of Glusup45/sup 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 Lyssup44/sup to Glu. The almost complete loss of IgE reactivity to the Pru av 1 Prosup112/sup mutant is due to disruption of its tertiary structure. Neither the mutation Alasup112/sup 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./p