Identification and assessment of IgE epitopes of Ara h 1, Jug r 2, Ana o 1 and Pis v 3 leader sequences

Authors: Nesbit, Jacqueline; Hurlburt, Barry; Cheng, Hsiaopo; Maleki, Soheila

Submitted to: Gordon Research Conferences
Publication Type: Abstract Only
Publication Acceptance Date: 12/21/2017
Publication Date: N/A
Citation: N/A

Interpretive Summary: RATIONALE: The vicilins, from peanut, walnut, cashew and pistachio are considered major allergens and are translated with leader sequences (LS) that are cleaved before yielding the mature protein. Structurally these LS resemble the 2S albumins, which are the most immunodominant allergens in nuts. METHODS: Linear IgE epitopes were identified using microarray data generated by printing 15-mer peptides offset by 5 amino acids on glass slides. IgE binding by peanut allergic sera was detected with a fluorescently-labeled antibody. Western blots and mass spectrometry were used to show the presence of the LS in peanut and walnut seeds. SWISS-MODEL protein modeling software was used to model the LS structures and IgE binding sites. RESULTS: For all four leader sequences, the epitopes with the highest degree of IgE binding were clustered within regions that were near cysteine residues. Of the patients tested, 96% showed IgE binding to those epitopes even if they recognized no other epitopes in the vicilins or the LS. Also, based on the molecular models, IgE binding is shown to be located at the junction of the c-terminal region of the alpha helices and the beginning of each flexible loop. CONCLUSIONS: The results indicate that cysteine residues known to confer high structural stability to allergens also confirm previous findings1 that they may coincide with areas of increased IgE binding frequency and intensity in Ara h 1, Jug r 2, Ana o 1 and Pis v 3 LS and Ara h 2. In addition, an unstructured region within close proximity to the cysteine residues may also be essential for IgE binding. The leader sequences contain multiple immunodominant epitopes and may be important components to consider in cross-reactivity and for diagnostic and therapeutic purposes.

Technical Abstract: RATIONALE: The vicilins, from peanut, walnut, cashew and pistachio are considered major allergens and are translated with leader sequences (LS) that are cleaved before yielding the mature protein. Structurally these LS resemble the 2S albumins, which are the most immunodominant allergens in nuts. METHODS: Linear IgE epitopes were identified using microarray data generated by printing 15-mer peptides offset by 5 amino acids on glass slides. IgE binding by peanut allergic sera was detected with a fluorescently-labeled antibody. Western blots and mass spectrometry were used to show the presence of the LS in peanut and walnut seeds. SWISS-MODEL protein modeling software was used to model the LS structures and IgE binding sites. RESULTS: For all four leader sequences, the epitopes with the highest degree of IgE binding were clustered within regions that were near cysteine residues. Of the patients tested, 96% showed IgE binding to those epitopes even if they recognized no other epitopes in the vicilins or the LS. Also, based on the molecular models, IgE binding is shown to be located at the junction of the c-terminal region of the alpha helices and the beginning of each flexible loop. CONCLUSIONS: The results indicate that cysteine residues known to confer high structural stability to allergens also confirm previous findings1 that they may coincide with areas of increased IgE binding frequency and intensity in Ara h 1, Jug r 2, Ana o 1 and Pis v 3 LS and Ara h 2. In addition, an unstructured region within close proximity to the cysteine residues may also be essential for IgE binding. The leader sequences contain multiple immunodominant epitopes and may be important components to consider in cross-reactivity and for diagnostic and therapeutic purposes.