Abstract Text: In food allergy, polyclonal IgE recognition of sequential epitopes is highly prevalent; however, little is known about the specific clonal recognition of these epitopes. Greater diversity of sequential epitope recognition by allergen-specific IgE is associated with increased severity and persistence of food allergy. For peanut allergy, the sequential epitope 63DPYSPOHSQDPYS72, located on a loop region of Ara h 2 and is identified in the serum of over 50% of peanut-allergic patients. We hypothesize that antibodies recognize multiple patterns of binding within this region of the immunodominant peanut allergen Ara h 2. To investigate this hypothesis, we utilized twelve monoclonal antibodies cloned from peanut-allergic, oral immunotherapy-treated patients to characterize the binding patterns of antibodies to synthetic variants of 63DPYSPOHSQDPYS72. Using alanine substitutions, we were able to identify three distinct patterns of amino acid residues engaged in epitope-paratope interactions within the 63DPYSPOHSQDPYS72 loop region of Ara h 2. Of these three patterns of binding, the most common binding pattern involves the hydroxyproline. Not only do these three groups of antibodies have distinct epitope-paratope binding patterns, but they also inhibit IgE binding to polyclonal serum differently on ELISA. In conclusion, we have identified three distinct binding patterns within the loop region of Ara h 2 which influences their ability to functionally disrupt IgE-allergen interactions. Therefore, previously identified sequential epitopes instead contain multiple overlapping epitopes. These variations in epitope-paratope interactions and their functional relevance is an essential component of antibody-mediated tolerance in IgE-mediated food allergy.
