Synthesis Of Hypoallergenic Derivatives Of Fagt1via Sequence Restructuring

Tartary buckwheat is one kind of crop that has high nutritional and medical values. It is a healthy food with lots of nutrients. However, buckwheat could trigger allergy reaction towards ingestion and inhalation, and the symptom including asthma, dermatitis, eczema and even anaphylactic shock. The11S storage protein from buckwheat seeds was identified as one of the major allergens in tartary buckwheat. It belongs to the cupin superfamily, and it is an abundant and stable protein in buckwheat seeds. In current study, we focus on some recombinant hypoallergenic derivatives which constructed by genetic engineering.Fag t1, a legumin-type protein, is the major allergen in tartary buckwheat. In the current study, three recombinant derivatives of Fag t1, designated as Fag t1-rs1, Fag t1-rs2, and Fag t1-rs3, were constructed via rational design and genetic engineering. However, because of the loss of their native-like folds, the Fag t1derivatives failed to bind IgE, and their allergenic activities were reduced. The recombinant hypoallergenic variants are promising vaccine candidates for specific immunotherapy of buckwheat allergy. The unfolding of the Fag t1structure reduced its high resistance to gastrointestinal proteolysis and strongly reduced its IgE reactivity. The derivatives showed a more than90%reduction in allergenic activity compared with rFag t1. These results suggest that the structure-dependent stability of11S seed storage proteins is directly related to digestive stability and allergenic potential. Therefore, the destruction of the native conformation is the appropriate strategy to reduce the allergenicity of the cupin family food allergens.Mail lard reaction commonly occurs in food processing, as one of the main chemical modifications of protein, but few studies exist on the influence of thermal processing on the allergenic potential of buckwheat allergen. Our study focuses on the effects of self-owned plant polysaccharides on the immunoreactivity of Fag t3, the major allergenic protein of buckwheat, during a naturally occurring Maillard reaction. Fag t3and the crude polysaccharides were prepared from tartary buckwheat (Fagopyrum tataricum) flour. Fag t3was covalently linked with polysaccharides at70℃for three days. After heating, the binding of human IgE or rabbit IgG were analyzed by enzyme-linked immunosorbent assay and immuno dot-blotting. Results indicated that the IgE/IgG-binding properties of Fag t3decreased significantly and underwent changes in electrophoretic mobility, secondary structures, solubility, and stability after glycation. The great influence of glycation on the IgE/IgG binding capacity of Fag t3is correlated with the significant change in the structure and epitopes of the protein. This glycation may occur in the processing of buckwheat food, which influences its allergenicity.