Linear and conformational epitopes of vicilin-buried peptides as a model for improved nut allergy diagnostics

Authors: SWIENTONIEWSKI, LAUREN - Oak Ridge Institute For Science And Education (ORISE); RAMBO, IAN - Oak Ridge Institute For Science And Education (ORISE); NESBIT, JACQUELINE - Oak Ridge Institute For Science And Education (ORISE); Cheng, Hsiaopo; GIPSON, STEPHEN - Oak Ridge Institute For Science And Education (ORISE); JONES, STACIE - University Of Arkansas; DRESKIN, STEPHEN - University Of Colorado; MUSTAFA, SHAHZAD - University Of Rochester; MUELLER, GEOFFREY - National Institute Of Environmental Health Sciences (NIEHS, NIH); Maleki, Soheila; SMITH, SCOTT - Vanderbilt University Medical Center; DOAN, DIEU - University Of Arkansas; KULIS, MICHAEL - University Of North Carolina; Rivers, Adam; FOO, ALEXANDER - National Institute Of Environmental Health Sciences (NIEHS, NIH)

Submitted to: Frontiers in Allergy
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/12/2025
Publication Date: 9/21/2025
Citation: Swientoniewski, L., Rambo, I.M., Nesbit, J.B., Cheng, H., Gipson, S.A., Jones, S.M., Dreskin, S.C., Mustafa, S.S., Mueller, G.A., Maleki, S.J., Smith, S.A., Doan, D.T., Kulis, M.D., Rivers, A.R., Foo, A.C. 2025. Linear and conformational epitopes of vicilin-buried peptides as a model for improved nut allergy diagnostics. Frontiers in Allergy. https://doi.org/10.3389/falgy.2025.1648262.
DOI: https://doi.org/10.3389/falgy.2025.1648262

Interpretive Summary: Peanut allergy is one of the most common food allergies and a main cause of food-induced anaphylaxis. Many with peanut allergies are also allergic to tree nuts, and ways to predict what is referred to as cross-reactivity is a large concern for scientists. Reoccurring sections called leader sequences of allergenic proteins in peanut and walnut with a shared 3D structure have been shown to be allergens themselves. Predictions for cross-reactions by an individual to peanut and walnut have been made based on the amino acid sequences of these leader sequence sections. We have analyzed the presence of additional potential cross-reactions to amino acid sequences based on the folded structure of these sections. Statistical modeling has been performed on the data for the sequences and the folded forms to show differences in allergic status. These findings can help predict if a peanut allergic individual may also have a reaction to walnuts or other tree nuts upon ingestion.

Technical Abstract: Introduction: Individuals allergic to peanuts (PN) may show IgE cross-reactivity to tree nuts, especially walnuts (WN), which often complicates diagnosis. Vicilin-buried peptides (VBPs), short segments within the N-terminal vicilin leader sequence (LS), contribute to cross-reactivity due to their ubiquitous, highly conserved and stable a-hairpin structures. The binding patterns of cross-reactive IgE to linear and conformational epitopes of PN and WN LSs and constituent VBPs may serve as a model for understanding clinically symptomatic cross-reactivity. Methods: Serum samples (n'='30) from primarily oral food challenge-positive individuals with PN allergy (PNA, 33%), WN allergy (WNA, 47%), and PN and WN allergies (PWA, 20%) were collected. These sera and a monoclonal IgE antibody (6D12) were examined for IgE binding with microarrays of overlapping peptides from native Ara h 1 LS [AH1LS, Ara h 1.0101 (26–84)] and recombinant Jug r 2 LS [JR2LS, Jug r 2.0101 (1–173)] and via direct and competitive inhibition ELISA with intact LSs and constituent VBPs from PN (AH1.1) and WN (JR2.1, JR2.2, JR2.3). A mixed model analysis assessed the contribution of IgE binding patterns to VBPs in relation to PNA, WNA, or PWA status. Results: All three intact WN VBPs bound IgE at similar frequencies, with individual sera showing varying preferences for specific VBPs. AH1.1 was less recognized by WNA individuals but more frequently recognized by PNA and PWA subjects. WN VBPs were recognized by PNA sera samples at rates comparable to AH1.1. Our data indicates that each VBP can bind to one IgE molecule with high affinity. In a competitive inhibition ELISA, combining VBP competitors did not enhance inhibition compared to the dominant VBP, suggesting that both high- and low-affinity VBPs compete for the same monoclonal IgE in serum. This observation was mimicked by 6D12, a monoclonal IgE against JR2.1. Discussion: Cross-reactivity among VBPs most likely arises from monoclonal IgE binding to a-hairpin structures and their overlapping linear amino acid sequences. The combination of linear and conformational IgE binding patterns enabled us to differentiate between the WNA, PNA, and PWA groups in this study and may assist us in using AH1LS and JR2LS to distinguish PN and WN allergies in the future