Transglutaminase-mediated cross-linking of a peptic fraction of ω-5 gliadin enhances IgE reactivity in wheat-dependent, exercise-induced anaphylaxis

Abstract Background: Patients with wheat-dependent, exercise-induced anaphylaxis (WDEIA) experience recurrent anaphylactic reactions when exercising after ingestion of wheat products. We have identified ω-5 gliadin (Tri a 19) as a major allergen in WDEIA, but the role of exercise in eliciting the symptoms remains obscure. Objective: The aim was to examine whether tissue transglutaminase (tTG)-mediated cross-linking could be involved in modulating the IgE-binding ability and in vivo reactivity of digested ω-5 gliadin peptides in WDEIA. Methods: Purified ω-5 gliadin was digested with pepsin or with pepsin and trypsin and treated with tTG. The binding of IgE antibodies in pooled sera from 10 patients with WDEIA was studied by means of immunoblotting before and after tTG treatment of the digested peptides. The peptides derived from pepsin digestion were separated by means of gel-filtration chromatography, and IgE reactivity of 4 different peptide fractions was studied by immunoblotting before and after tTG treatment. The fraction showing the greatest degree of cross-linking by tTG was further studied by means of IgE ELISA, ELISA inhibition, and skin prick testing. Results: The IgE-binding ability of ω-5 gliadin was retained after pepsin and pepsin-trypsin digestion. tTG treatment of the whole peptic digest formed large peptide complexes, with molecular weights ranging from 40 to greater than 200 kd. These cross-linked aggregates bound IgE antibodies in immunoblotting more intensely than untreated, pepsin-digested, or pepsin-trypsin-digested ω-5 gliadin. A gel-filtration fraction of the whole peptic digest corresponding to the highest peak of the chromatogram and showing the greatest degree of tTG-mediated cross-linking showed an increase in serum IgE reactivity in ELISA after tTG treatment, as well as a shift of reactivity to cross-linked complexes. In the 20 patients with WDEIA, the mean skin prick test wheal elicited by this tTG-treated peptic fraction was 77% larger ( P P