Allergy / Distinct epitope structures of defensin\u2010like proteins linked to proline\u2010rich regions give rise to differences in their allergenic activity

Abstract

Background: Art v 1, Amb a 4, and Par h 1 are allergenic defensin\u2010polyproline\u2013linked proteins present in mugwort, ragweed, and feverfew pollen, respectively. We aimed to investigate the physicochemical and immunological features underlying the different allergenic capacities of those allergens. Methods: Recombinant defensin\u2010polyproline\u2013linked proteins were expressed in E. coli and physicochemically characterized in detail regarding identity, secondary structure, and aggregation status. Allergenic activity was assessed by mediator releases assay, serum IgE reactivity, and IgE inhibition ELISA using sera of patients from Austria, Canada, and Korea. Endolysosomal protein degradation and T\u2010cell cross\u2010reactivity were studied in vitro. Results: Despite variations in the proline\u2010rich region, similar secondary structure elements were observed in the defensin\u2010like domains. Seventy\u2010four percent and 52% of the Austrian and Canadian patients reacted to all three allergens, while Korean patients were almost exclusively sensitized to Art v 1. This was reflected by IgE inhibition assays demonstrating high cross\u2010reactivity for Austrian, medium for Canadian, and low for Korean sera. In a subgroup of patients, IgE reactivity toward structurally altered Amb a 4 and Par h 1 was not changed suggesting involvement of linear epitopes. Immunologically relevant endolysosomal stability of the defensin\u2010like domain was limited to Art v 1 and no T\u2010cell cross\u2010reactivity with Art v 125\u201036 was observed. Conclusions: Despite structural similarity, different IgE\u2010binding profiles and proteolytic processing impacted the allergenic capacity of defensin\u2010polyproline\u2013linked molecules. Based on the fact that Amb a 4 demonstrated distinct IgE\u2010binding epitopes, we suggest inclusion in molecule\u2010based allergy diagnosis