Effect of Structural Targeted Modifications on the Potential Allergenicity of Peanut Allergen Ara h 2

Protein structure affects allergenicity, and critical structural elements, especially conformational epitopes that determine allergenicity, have attracted a great deal of interest. In this study, we aimed to identify the localized structure that affects the potential allergenicity of protein by making targeted modifications of Ara h 2 and comparing the structure and allergenicity of mutants with those of the wide-type allergen. The structures of the allergen and its mutants were characterized by circular dichroism and ultraviolet absorption spectroscopy and simulated by molecular dynamics. The allergenicity was assessed by Western blotting, an indirect competitive enzyme-linked immunosorbent assay, a cell model, and a mouse model. Then, the structures that affect allergenicity were analyzed and screened. Our results showed that mutations in amino acids changed the nearby localized structure and the overall structures. The structural changes affected the IgE binding capacity of the allergen and reduced its potential allergenicity. The solvent accessible surface area (SASA) of aromatic residues was positively correlated with the IgE binding capacity. The integrity of the disulfide bond is also critical for the binding of IgE to allergens. Interestingly, different mutations induced similar electrostatic potential and allergenicity changes, such as localized structure R62DPYSPSQDPYSPS75. In conclusion, the disulfide bond and the SASA of aromatic residues are important for the allergenicity of Ara h 2. The localized structure R62DPYSPSQDPYSPS75 is also crucial for the allergenicity of Ara h 2.