Identification of major wheat allergens by means of the Escherichia coli expression system

Abstract

Wheat proteins were fractionated into salt-soluble, glutenin-rich, and gliadin-rich fractions. Reactivities of these protein fractions with sera of patients with wheat-associated allergies were examined under various conditions. The relative reactivity of the fractions was generally in the order glutenin-rich Ͼ gliadin-rich ӷ salt-soluble fractions, although their reactivities were variable among patients and among the reaction conditions, indicating that the kind, the number and the epitope of allergens were variable among patients. To identify major allergens, A-, γ-and ω-gliadin, and low-molecular-mass (LMM)-and high-molecular-mass (HMM)-glutenin genes were expressed in Escherichia coli by means of a pET vector. Recombinant gliadins and glutenins were partially purfied on the basis of the solubilities of prolamin and glutelin. The partially purified recombinant proteins were reacted with the patients' sera. LMM glutenin containing many Gln-Gln-Gln-Pro-Pro motifs, which was identified to be IgE-binding epitope [Tanabe, S., Arai, S., Yanagihara, Y., Mita, H., Takahashi, K. & Watanabe, M. (1996) Biochem. Biophys. Res. Commun. 219, 290Ϫ293], exhibited the highest reactivity. The next highest reactivities were observed on A-gliadin and γ-gliadin, which had not been identified as allergens.

Antibody reactivity in patients with IgE-mediated wheat allergy to various subunits and fractions of gluten and non-gluten proteins from ω-gliadin-free wheat genotypes

“…The current work is also consistent with the results obtained by other authors who reported allergenicity of α, β and γ gliadins in atopic dermatitis, and LMW glutenins in baker's asthma [20,29,30]. The study of Maruyama et al [31] determined their relative reactivity in the following order: glutenins>gliadins>NGP. However, in the present study, the specificity of the obtained IgE-binding profiles was observed at the level of individual subunits and fractions of GP and NGP, which allowed to consider patterns of IgE binding as fingerprints characteristic for individual patients.…”

“…Comparison of immunoreactivity of native and reduced proteins allow to hypothesize whether conformational or linear epitopes play more important role in the IgE binding [30,31]. Our findings evidently demonstrated no IgE detection of non-reduced gliadins, which indicated that α, β and γ fractions gained immunoreactivity only after the disruption of SS bonds and unfolding of protein molecules.…”

Presence of ferulic acid in wheat glutenin fraction and its enzymatic hydrolysates - a short report

“…Ground wheat grains (10 g) were suspended in 100 ml 10% NaCl, stirred for 45 min and centrifuged at 5000 g for 10 min (Maruyama et al 1998). The extraction was repeated twice more.…”

Analysis of Food Allergen Structures and Development of Foods for Allergic Patients

“…Recombinant low-molecular-weight glutenins with many Gln-Gln-Gln-Pro-Pro motifs have been expressed in Escherichia coli by a pET vector system, and were confirmed to possess IgE-binding ability. 29) As other allergens, thioredoxin, 19) lipid transfer protein, 20) acyl-CoA oxidase, 30) peroxidase, 31) and fructose-bisphosphate aldolase 32) were identified. 8,21) In addition, the epitope structures of gliadin in patients with FDEIA 24) and celiac disease 33) have been recently reported.…”