Insights into Api m 10‐Isoforms and Splice Variants: More Than One Major IgE‐Binding Epitope

TL;DR

This study identifies multiple IgE-binding regions in the honey bee venom allergen Api m 10, revealing insights that could improve allergy treatments.

Contribution

The study reveals multiple IgE-binding epitopes in Api m 10 isoforms and splice variants, including a major epitope and potential conformational epitopes.

Findings

Seven linear IgE-binding motifs were identified, with one major epitope shared among multiple Api m 10 isoforms.

Api m 10-specific IgE from patients and IgG from mice showed overlapping binding motifs, suggesting conserved epitopes.

Api m 10 isoforms 1 and 2 exhibited secondary structural elements and aggregation, which may influence allergenicity.

Abstract

Honey bee venom (HBV) often triggers severe IgE‐mediated allergies. The major allergen icarapin (Api m 10) has attracted attention due to low occurrence in some HBV immunotherapy products. Despite being a major allergen, little is known about the Api m 10 structure and IgE‐binding regions. This study aimed to characterize its IgE‐binding epitopes and structure in more detail.

Overlapping Api m 10‐specific peptides covering the sequences of the 11 known Api m 10‐isoforms and variants were synthesized and spotted on microarray slides (15 amino acids (AA), off‐set: 4 AA). Sera from 28 HBV‐allergic patients with detectable Api m 10‐specific IgE were used to characterize the distinct IgE‐binding profiles to all Api m 10‐variants. Sera from ten Api m 10‐immunized BALB/c mice were used to investigate possible shared epitopes between humans and mice. All Api m 10‐variants were investigated for secondary structural elements via circular dichroism spectroscopy and potential aggregation via dynamic light scattering.

We identified 7 different linear IgE‐binding motifs. All 28 patients' sera displayed IgE‐binding to one specific area (present in Api m 10‐isoforms 1 and 2 and putative splice variants 3, 4, 6), indicating a major IgE‐epitope. IgE‐inhibition provided evidence that the major epitope makes up less than 50% of the total IgE‐binding capacity, suggesting that additional (most likely conformational) IgE‐epitopes play an important role in Api m 10‐sensitization. Api m 10‐specific murine IgG and human IgE both predominantly bind to seven different AA‐motifs, of which six are identical between both species. Api m 10‐isoforms 1 and 2 displayed secondary structural elements and appeared to be aggregated.

The structural, clinical and preclinical insights into Api m 10 and its immunodominant epitopes gained in this study provide substantial insights for the future development of active and passive VIT as well as further treatment approaches.

The structural, clinical and preclinical insights into Api m 10 and its immunodominant epitopes gained in this study provide substantial insights for the future development of active and passive VIT as well as further treatment approaches.

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