Epitope Mapping of Major Ragweed Allergen Amb a 1

Abstract

Ragweed is a prominent cause of seasonal allergies. Thus far, information on IgE-binding sites of major allergen in ragweed pollen, Amb a 1, is very limited. A powerful experimental method to gain insights on the allergen epitopes is the selection of peptides from biological libraries that bind to anti-allergen antibodies. In this work, we aimed to map IgE epitopes of Amb a 1 using epitope-mimicking short peptides-mimotopes that were affinity-selected from phage-displayed random peptide libraries. The peptides weakly aligned with the Amb a 1 primary sequence, thus suggesting that the epitopes are conformational. When the peptides were mapped onto the surface of Amb a 1 homology model, the EpiSearch analysis predicted the location of four potential epitopic sites on surface patches centred at residues K 104 , S 110 , H 214 , and W 312. The peptides matching to the predicted epitopes bound selectively to the IgE from pool of ragweed-allergic patients' sera and therefore represent mimetics of Amb a 1 IgE epitopes. The knowledge of IgE epitopes is a prerequisite for the rational design of molecular-based approaches to diagnosis and immunotherapy of allergic diseases.

Ara h 2‐specific IgE epitope‐like peptides inhibit the binding of IgE to Ara h 2 and suppress lgE‐dependent effector cell activation

“…Biopanning of phage display libraries with anti‐Ara h 2 IgG, screening of phage clones and selection of Ara h 2 epitope‐like peptides were performed previously for Api m 1, Fel d 1 and Amb a 1 28–30 . Binding of peptides (fused with bacteriophage protein pIII as a carrier) to IgE from the sera of patients with peanut allergy was evaluated by immunoblotting 28–30 .…”

“…Biopanning of phage display libraries with anti-Ara h 2 IgG, screening of phage clones and selection of Ara h 2 epitope-like peptides were performed previously for Api m 1, Fel d 1 and Amb a 1. [28][29][30] Binding of peptides (fused with bacteriophage protein pIII as a carrier) to IgE from the sera of patients with peanut allergy was evaluated by immunoblotting. [28][29][30] For all functional experiments (ImmunoCAP inhibition, BAT and MAT, and BAT and MAT inhibition experiments), peptides were synthesized with C-terminal amidation (EZBiolab, Parsippany, NJ, USA).…”

“…[28][29][30] Binding of peptides (fused with bacteriophage protein pIII as a carrier) to IgE from the sera of patients with peanut allergy was evaluated by immunoblotting. [28][29][30] For all functional experiments (ImmunoCAP inhibition, BAT and MAT, and BAT and MAT inhibition experiments), peptides were synthesized with C-terminal amidation (EZBiolab, Parsippany, NJ, USA).…”

“…39 The DHPR motif identified in this study by phage biopanning has not yet been described. Phage biopanning enables discovery of epitopes based on 3D structures, and it has been successfully utilized for various allergens, [28][29][30] including the peanut allergens Ara h 2 and Ara h 6. 45 Recently, Hazebrouck et al 41 showed that the major conformational IgE-binding epitope of Ara h 2 is located in a segment between residues 33 and 81 and that this segment is critical for the capacity of Ara h 2 to induce effector cell degranulation.…”