Alteration of Allergen Fold of Bos d 5 into a Hypoallergenic Vaccine for Immunotherapy of Cow’s Milk Allergy

<b><i>Background:</i></b> Cow’s milk allergy (CMA) is the most common IgE-mediated food allergy and Bos d 5 is the major allergen in cow’s milk proteins. More than 60% of the patients with CMA are sensitized to this protein. <b><i>Methods and Results:</i></b> A recombinant protein, encoded by a synthetic gene and consisting of reassembled Bos d 5 fragments, was expressed in <i>E. coli</i> strain BL21 (DE3) cells and purified to homogeneity. The B5M lacked relevant IgE-reactivity and allergenic activity compared with Bos d 5 in dot-blot and basophil activation assays. T-cell proliferation experiments demonstrated that B5M preserved the main T cell epitopes of Bos d 5. Immunization of rabbits with B5M induced protective IgG antibodies that blocked the binding of patients’ IgE antibodies to the wild-type allergen and inhibited the degranulation of basophils induced by Bos d 5. <b><i>Conclusion:</i></b> Thus, we developed a new strategy, which was based on rational molecular reassembly for allergen-specific immunotherapy (AIT) of CMA and food allergy.