Targeting a Cross-Reactive Gly m 5 Soy Peptide as Responsible for Hypersensitivity Reactions in a Milk Allergy Mouse Model

Paola L Smaldini,Ivana Barrios,Silvana Petruccelli,Carlos A Fossati,Virginia González,Guillermo H Giambartolomei,Guillermo H Docena,Ana Cauerhff,Gustavo Parisi,Luis Bruno Blanch,Angela M Candreva,Renata Curciarello

doi:10.1371/journal.pone.0082341

Abstract

BackgroundCross-reactivity between soybean allergens and bovine caseins has been previously reported. In this study we aimed to map epitopes of the major soybean allergen Gly m 5 that are co-recognized by casein specific antibodies, and to identify a peptide responsible for the cross-reactivity.MethodsCow's milk protein (CMP)-specific antibodies were used in different immunoassays (immunoblotting, ELISA, ELISA inhibition test) to evaluate the in vitro recognition of soybean proteins (SP). Recombinant Gly m 5 (α), a truncated fragment containing the C-terminal domain (α-T) and peptides of α-T were obtained and epitope mapping was performed with an overlapping peptide assay. Bioinformatics tools were used for epitope prediction by sequence alignment, and for modelling the cross-recognized soy proteins and peptides. The binding of SP to a monoclonal antibody was studied by surface Plasmon resonance (SPR). Finally, the in vivo cross-recognition of SP was assessed in a mouse model of milk allergy.ResultsBoth α and α-T reacted with the different CMP-specific antibodies. α-T contains IgG and IgE epitopes in several peptides, particularly in the peptide named PA. Besides, we found similar values of association and dissociation constants between the α-casein specific mAb and the different milk and soy components. The food allergy mouse model showed that SP and PA contain the cross-reactive B and T epitopes, which triggered hypersensitivity reactions and a Th2-mediated response on CMP-sensitized mice.ConclusionsGly m 5 is a cross-reactive soy allergen and the α-T portion of the molecule contains IgG and IgE immunodominant epitopes, confined to PA, a region with enough conformation to be bound by antibodies. These findings contribute to explain the intolerance to SP observed in IgE-mediated CMA patients, primarily not sensitised to SP, as well as it sets the basis to propose a mucosal immunotherapy for milk allergy using this soy peptide.

Highlights

Food allergy is an increasing health care concern
Different Cow’s milk protein (CMP)-specific antibodies were used to assess the cross-reactivity of these soy components (Figure 1)
We found that all sera recognized CMP (100%), while most of the milk allergic patient sera presented IgE antibodies that recognized a (86.6%) and a-T (93.3%) (Figure 1A)

Summary

Introduction

Cow’s milk protein (CMP) is the leading cause of food allergy in children younger than 3 years in many regions, including our country [1,2]. This is reflected by most of the position papers and guidance with statements and recommendations for food allergy that are focused on CMP allergy (CMA) [3]. Cross-reactivity between soybean allergens and bovine caseins has been previously reported. In this study we aimed to map epitopes of the major soybean allergen Gly m 5 that are co-recognized by casein specific antibodies, and to identify a peptide responsible for the cross-reactivity

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Discussion

Conclusion