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Sequence comparison of omega-5 gliadins encoded by the B and D genomes in Chinese Spring with the full-length omega-5 gliadin BAE20328. N-and C-terminal sequences are shown in black and blue boxes, respectively. Within each protein sequence, the dominant WDEIA epitopes QQFPQQQ and QQIPQQQ are shown in red and blue, respectively. Lines above the protein sequence highlight peptides unique to omegaD4 that were identified by MS/MS in protein spots from DH20 reacting with sera from WDEIA patients. Red lines highlight peptides identified in the more basic protein while blue lines highlight peptides identified in the more acidic protein. MS/MS data are shown in Additional file 1
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Background:
Omega-5 gliadins are a group of highly repetitive gluten proteins in wheat flour encoded on the 1B chromosome of hexaploid wheat. These proteins are the major sensitizing allergens in a severe form of food allergy called wheat-dependent exercise-induced anaphylaxis (WDEIA). The elimination of omega-5 gliadins from wheat flour through b...
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Context 1
... amino acid sequences of the omega-5 gliadin pro- teins encoded by the B genome in Chinese Spring (ome- ga-B3 and omega-B6) were compared to that of omega-D4 (Fig. 6). The only other full-length omega-5 gliadin sequence in NCBI, BAE20328, was also included in the analysis. Omega-B3 differs from BAE20328 by two amino acid substitutions and a 28 amino acid dele- tion while omega-B6 has six amino acid substitutions and a five amino acid insertion. In comparison, the se- quence of omega-D4 is quite ...
Context 2
... overlap and the regions of overlap are shorter. BAE20328 and omega-B3 each have only two regions where two epitopes overlap, two regions where 3 epitopes overlap and one region where four epitopes overlap while omega-B6 has four regions where two epitopes overlap, one region where three epitopes overlap and one region where four epitopes overlap (Fig. 6, Table ...
Context 3
... excised from 2-D gels, digested with chymotrypsin, thermolysin or trypsin and analyzed by MS/MS. For analysis of spectral data, the se- quences of gluten protein genes from Chinese Spring were added to the database used for interrogation. Pep- tides unique to omega-D4 were identified in spots corre- sponding to both of the reactive proteins ( Fig. 6, Additional file 1), suggesting that these proteins are encoded by the D genome. PCR analysis of genomic DNA with primers specific for the Chinese Spring omega-D4 gene (Table 2) demon- strated the presence of omega-D4 in both of the parental lines as well as in the mutant DH20 (Fig. 7). Amplifica- tion of aneuploid lines of Chinese ...
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Citations
... Notably, ω5-gliadin and high-molecular-weight glutenin have been identified as major and minor allergens in WDEIA, respectively [4]. Enzymolysis/ion exchanger deamidation, thioredoxin treatment, and wheat strains lacking allergen-coding genes have been used to obtain hypoallergenic wheat [5]. The wheat products obtained using these approaches substantially reduce the reactivity of serum IgE in patients with WDEIA; however, they are either found to be ineffective in some patient populations or low levels of IgE reactivity are still observed for these products [5]. ...
... Enzymolysis/ion exchanger deamidation, thioredoxin treatment, and wheat strains lacking allergen-coding genes have been used to obtain hypoallergenic wheat [5]. The wheat products obtained using these approaches substantially reduce the reactivity of serum IgE in patients with WDEIA; however, they are either found to be ineffective in some patient populations or low levels of IgE reactivity are still observed for these products [5]. ...
Wheat-dependent, exercise-induced anaphylaxis has no fundamental cure and requires patients to refrain from wheat consumption or to rest after eating. Although hypoallergenic wheat production by enzymatic degradation or thioredoxin treatment has been investigated, challenges still exist in terms of labor and efficacy. We investigated a hypoallergenic wheat product manufacturing technology that takes advantage of the property of tannins to bind tightly to proteins. Commercially available bread wheat (BW) and hypoallergenic wheat (1BS-18 “Minaminokaori”, 1BS-18M) were used. Chestnut inner skin (CIS) was selected as a tannin material based on the screening of breads with added unused parts of persimmon and chestnut. Hypoallergenicity was evaluated using Western blotting. The effect of CIS addition on the antioxidative properties of bread was also measured. For both BW and 1BS-18M, CIS addition reduced the immunoreactivity of wheat allergens. Antioxidant activities increased with increasing CIS substitution. However, 10% CIS-substituted breads were substantially less puffy. Five percent CIS substitution was optimal for achieving low allergenicity, while maintaining bread quality. The strategy investigated herein can reduce allergies related to wheat bread consumption. In this study, the evaluation of hypoallergenicity was limited to instrumental analysis. In the future, we will evaluate hypoallergenicity through clinical trials in humans.
... In the present study, NFA had no significant effect on the albumin and globulin concentrations in the wheat grains, which was because they were structural proteins that were less affected by environmental factors [15]. Gliadin and glutenin, also known as gluten proteins, were closely related to the wheat processing quality and were controlled by genetic and environmental factors [32]. A previous study showed that the application of N fertilizer increased the content of alcoholsoluble protein and gluten [33], and the same results were obtained in this study. ...
Crop quality tends to decrease with an increasing grain yield. Nitrogen is an important nutrient and moderate nitrogen foliage application (NFA) can significantly improve the wheat yield and quality. The objective of this study was to investigate the effect of NFA on the grain yield and quality of wheat and its genotype-dependent variation. Eighteen wheat cultivars were used, and two NFA levels (N1 and N2; 10.70 and 21.40 kg N ha−1 two day−1, respectively) were applied. Significant genotypic differences in the yield and quality were observed among the 18 varieties, and their responses to NFA differed. For nine varieties in the experiment, N1 increased the grain yield, but N2 did not. In contrast, high concentrations of NFA had no effect on the grain yield in the other nine varieties. The protein content and composition and trace element (Fe, Zn, etc.) are all nutrient elements that notably affect the wheat grain quality and yield. NFA significantly increased the grain prolamin and glutelin concentrations in the grains, thereby increasing the total protein concentration. The prolamin, glutelin, and total protein concentrations in the grains of the lower-protein cultivars were more sensitive to NFA than those of the higher-protein cultivars. In addition, NFA significantly decreased the amylose concentration in the grains. By affecting the prolamin, glutelin, and amylose concentrations in the grains, NFA significantly increased the development and stability times of the corresponding wheat flour dough, thereby improving the dough quality. Moreover, NFA reduced the molar ratio of phytic acid to Fe and Zn, increasing the bioavailability of trace elements. The judicious application of nitrogen fertilizer resulted in the synergistic improvement in the yield and quality.
... Skinprick and specific immunoglobulin E (IgE) tests are also used to diagnose WDEIA, but they provide unreliable results [11]. An additional approach to diagnose WDEIA in vitro is to measure immune response to wheat by analyzing the patient's serum [12,13]. However, this approach is not ideal both because obtaining a patient's serum is costly and time consuming and because some patients' immune responses may be too low to detect. ...
... Omega-5 gliadins represent the major class of gluten antigens that cause WDEIA. Omega-5 gliadins are encoded by Gli-B1 and Gli-D1 genes on the short arm of chromosome 1 [12,14]. Their primary structure consists of an N-terminus starting with SRL or TRQ residues, followed by repeat sequences of FPQQQ and QQIPQQ and then a C-terminus [12,[15][16][17]. ...
... Omega-5 gliadins are encoded by Gli-B1 and Gli-D1 genes on the short arm of chromosome 1 [12,14]. Their primary structure consists of an N-terminus starting with SRL or TRQ residues, followed by repeat sequences of FPQQQ and QQIPQQ and then a C-terminus [12,[15][16][17]. Omega-5 gliadins do not contain any cysteine (Cys) residues and therefore do not form disulfide bonds [18,19]. ...
In allergic individuals, ingestion of wheat can lead to wheat-dependent exercise-induced anaphylaxis (WDEIA). Many studies have been conducted to find WDEIA allergen–deficient wheat, including by generating omega-5 gliadin antibodies. However, the reported antibodies have not been specific enough to detect omega-5 gliadins encoded on the 1B chromosome. In this study, we generated monoclonal antibodies against the major allergens causing WDEIA, omega-5 gliadins. Using these antibodies (mono-O5B-1C10), we assessed accumulation of omega-5 gliadins in wild-type and nullisomic-tetrasomic (NT) lines of the wheat ( Triticum aestivum ) varieties Chinese Spring (CS) by one- and two-dimensional gel electrophoresis, followed by Coomassie blue staining or immunoblotting with mono-O5B-1C10. We also tested mono-O5B-1C10 for major omega-5 gliadins in various wheat germplasms. Our results thus demonstrate the specificity of mono-O5B-1C10 for major omega-5 gliadins and potentially useful for identifying of omega-5 gliadin–deficient wheat varieties that should not cause WDEIA.
... No oral challenge test was performed in human. [46][47][48][49] [ [60][61][62][63][64] SDS-PAGE: sodium dodecyl sulfate-polyacrylamide gel electrophoresis, WDEIA: wheat-dependent exerciseinduced anaphylaxis, RP-HPLC: reversed phase-high performance liquid chromatography, ELISA: enzyme-linked immunosorbent assay, AD: atopic dermatitis, CAP-FEIA: fluorescence enzyme immunoassay using ImmunoCAP. ...
... Notably, two-dimensional immunoblot analysis revealed the existence of several minor but highly immunogenic ω5-gliadin proteins in a wheat mutant line, DH20, with a deletion of 5.8 Mb, including the Gli-B1 locus of Chromosome 1B [48]. These proteins were found to be encoded in the ω5-gliadin gene of Chromosome 1D and to have a high homology, including repetitive IgE binding epitopes to the ω5-gliadin of the Gli-B1 locus, although they have a TRQ N-terminal amino acid sequence and altered C-terminal amino acid sequence compared with the Gli-B1 ω5-gliadin [49]. ...
Immunoglobulin E (IgE)-mediated food allergies to wheat that develop after school age typically shows a type of wheat-dependent exercise-induced anaphylaxis (WDEIA). At present, avoidance of wheat products or postprandial rest after ingesting wheat is recommended for patients with WDEIA, depending on the severity of the allergy symptoms. ω5-Gliadin has been identified as the major allergen in WDEIA. In addition, α/β-, γ-, and ω1,2-gliadins, high and low molecular weight-glutenins, and a few water-soluble wheat proteins have been identified as IgE-binding allergens in a small proportion of patients with IgE-mediated wheat allergies. A variety of approaches have been manufactured to develop hypoallergenic wheat products that can be consumed by patients with IgE-mediated wheat allergies. In order to analyze such approaches, and to contribute to the further improvement, this study outlined the current status of these hypoallergenic wheat productions, including wheat lines with a reduced allergenicity that are mostly constructed for the patients sensitized to ω5-gliadin, hypoallergenic wheat by enzymic degradation/ion exchanger deamidation, and hypoallergenic wheat by thioredoxin treatment. The wheat products obtained by these approaches significantly reduced the reactivity of Serum IgE in wheat-allergic patients. However, either these were not effective on some populations of the patients, or low-level IgE-reactivity to some allergens of the products was observed in the patients. These results highlight some of the difficulties faced in creating hypoallergenic wheat products or hypoallergenic wheat lines through either traditional breeding or biotechnology approaches in developing hypoallergenic wheat completely safe for all the patients allergic to wheat.
... Омега-5-гліадини, контрольовані локусом Gli-B1, виявилися основними алергенами, що спричиняють анафілактичний шок у чутливих людей у випадку, коли вживання продуктів із пшениці супроводжується фізичними навантаженнями (WDEIA, wheat-dependent exercise-induced anaphylaxis), тому цю хворобу також називають алергія на омега-5-гліадин [4]. Крім того, WDEIA теж можуть викликати білки, кодовані гліадиновими генами локусу Gli-D1 [5]. Омега-5-гліадини також є основним алергеном у дітей з атопічним дерматитом, які мають алергію на пшеницю з реакцією гіперчутливості з швидким розвитком симптомів [6]. ...
... Копусем на основі сорту Безоста 1 [10]. Лінія B3 відрізняється від сорту Безоста 1 присутністю пшенично-житньої 1BL.1RS транслокації як у сорту Кавказ, маркером якої є характерний блок секалінів, зазначений у каталозі гліадинових алелей як алель Gli-B1l [5]. Лінія D4 відрізняється від сорту Безоста 1 присутністю алелі Gli-D1j замість Gli-D1b. ...
Aim. The study was aimed at identification and selection of winter common wheat genotypes with null-alleles at the gliadin loci Gli-B1 and Gli-D1. Methods. The search for spontaneous mutants with the absence of synthesis of gliadin blocks was made in winter common wheat hybrid material and cultivars. To identify mutations, APAG electrophoresis and SDS-electrophoresis of seed storage proteins were performed. Results. The frequency of spontaneous mutations resulting in the null-allele at the loci Gli-B1 and Gli-D1 in F2 was 0,1 % and 0,05 %, respectively, in the cross Odesska chervonokolosa × B-16. Via sowing the material with those mutations and marker selection, F6 lines were developed: the line OB-Bnull with the null-allele at the Gli-B1 locus and the line OB-Dnull with the null-allele at the Gli-D1 locus. Another line with the null-allele at Gli-B1 was produced from the cross B3 × D4 by marker selection, as well as a biotype with the null-allele at the Gli-D1 locus was selected from the cultivar Slaven. Conclusions. The developed lines with null-alleles at the gliadin locus Gli-B1 (without omega-5 gliadins) and Gli-D1 (without omega-1,2 gliadins) are initial material for hypoallergenic wheat breeding.
... Herman et al. produced a hypoallergenic soybean lacking the major soybean allergen Gly m Bd 30 K using transgene-induced gene-silencing technology [9]. Lee et al. developed hypoallergic cow's milk with reduced levels of bovine α-casein and β-lactoglobulin by irradiation with gamma rays [10]. Regarding wheat cultivars, Altenbach et al. produced transgenic wheat with a reduced content of ω5-gliadin using RNA interference technology [11][12][13][14]. However, these hypoallergic foods may be unacceptable to consumers, especially in Japan, because they were developed using transgenic technology [15]. ...
... Thus, we consider that its slight detection in 1BS-18H gluten may be related to the cross-reactivity of anti-ω5-gliadin Abs to other gluten components in ELISA. Altenbach et al. reported that a mutant wheat line lacking the major ω5-gliadin encoded on chromosome 1B had several minor ω5-gliadin proteins located on chromosome 1D that possessed several IgE-binding epitopes for WDEIA patients [12]. This report suggests that 1BS-18H wheat may also contain minor ω5-gliadin encoded on chromosome 1D. ...
The early ingestion of food can prevent the onset of food allergy related to inducing oral tolerance (OT). We developed the Hokushin wheat line as a hypoallergenic wheat (1BS-18H) lacking ω5-gliadin, a major allergen of wheat-dependent exercise-induced anaphylaxis (WDEIA). The 1BS-18H wheat had lower ability of sensitization for ω5-gliadin compared with Hokushin wheat. Here, we evaluated the induction of OT to gluten and ω5-gliadin by the early consecutive ingestion of 1BS-18H gluten using a rat model of wheat allergy. Rats were subcutaneously immunized with commercial gluten or native ω5-gliadin following the daily oral administration of gluten. The daily oral administration of 1BS-18H gluten for 5 days before immunization suppressed the increase in gluten- or ω5-gliadin-specific IgE and IgG1 antibodies induced by immunization to a level similar to Hokushin gluten. Intravenous challenge with gluten or ω5-gliadin did not decrease the rectal temperature in rats with OT induced by 1BS-18H or Hokushin gluten, although it was decreased in non-OT rats. In conclusion, the early consecutive ingestion of 1BS-18H wheat before sensitization induced OT to gluten and ω5-gliadin. These findings support the benefit of 1BS-18H wheat to prevent wheat allergy including WDEIA by consecutive ingestion in humans.
... However, it still expresses the α/β-, ω-1/2, some γ-gliadins, and a small amount of ω-5 gliadins from the 1D chromosome, as well as HMW and some LMW glutenin subunits. 9,11 In this study, we evaluated the allergenicity of a wheat cultivar deficient in the ω-5 gliadin and especially m-type LMW glutenin in WDEIA patients using in vitro immunoglobulin E (IgE) immunoblot and inhibition IgE immunoassays with enzyme-linked immunosorbent assay (ELISA) and ImmunoCAP. ...
... This ω5D cultivar has missing Glu-B3 and closely linked Gli-B1 loci, which results in deficiency of ω-5, γ-gliadins, s-type-, i-type-, and m-type-LMW glutenins. 9,11,12 We extracted gliadins and glutenins from the ω5D and wild-type bread flour (T. aestivum ssp. ...
... Rather than using genetic engineering, our ω5D cultivar was bred from double-haploid lines derived from two preexisting wheat cultivars: the hard white Keumkang and soft red Olgeru cultivars. 9,11,12 So, there is no risk of gaining new allergenic epitopes in this ω5D cultivar. Although the quantity of ω-5 gliadins is about one-fourth of the parent cultivars, there was no difference in the percentages of the α/β, ω-1/2, and albumin between the two cultivars. ...
Purpose:
ω-5 gliadin is the major allergen that causes wheat-dependent exercise-induced anaphylaxis (WDEIA). Recently, a missing mutant wheat cultivar at 1B chromosome Glu-B3 and closely linked Gli-B1 loci was bred. This cultivar (ω5D) has a deficiency in ω-5 and γ-gliadins as well as some low-molecular-weight glutenins. We evaluated specific immunoglobulin E (sIgE) reactivity of the ω5D in WDEIA patients compared to wild-type cultivar.
Methods:
Serum samples from 14 WDEIA and 7 classic wheat allergy patients were used to compare the allergenicity of ω5D and wild-type cultivars using immunoglobulin E immunoblotting, enzyme-linked immunosorbent assay (ELISA), and ImmunoCAP inhibition assays.
Results:
Immunoblotting revealed that ω5D extracts had less sIgE binding to gliadins and glutenins in WDEIA sera than wild-type extracts. Immunoblot inhibition assay for gliadin sIgE reactivity also showed that ω5D gliadins had less allergenicity than wild-type gliadins. ELISA inhibition assay showed stronger allergenicity of gliadins than glutenins, although they had cross-reactivity. This assay also showed that the 50% inhibitory concentrations (IC50) of ω5D extracts against gliadin- or glutenin-sIgE reactivity were approximately 4-fold higher in WDEIA patients than those of wild-type extracts. The inhibition capacity of ω5D gliadins against recombinant ω-5 gliadin-sIgE reactivity was also lower in WDEIA patients than that of wild-type.
Conclusions:
The allergenicity of the ω5D cultivar is markedly lower for WDEIA patients in the sIgE inhibition tests. These results suggest that the ω5D cultivar may be a safe alternative for WDEIA patients.
... It should be kept in mind that almost any food group (like cereals, seafood, vegetables, legumes, fruits, tree nuts, meat, etc) can trigger this condition. [65] The occurrence of wheat-dependent exercise-induced anaphylaxis (WDEIA) is related to the ingestion of wheat followed by physical exercise. [66] It has been suggested that FDEIA is a primary IgE-mediated food allergy augmented by cofactors. ...
Gluten is the most common protein in wheat and is related to different pathogeneses. There are three main gluten-related disorders: celiac disease (CD), wheat allergy (WA), and non-celiac gluten sensitivity (NCGS). Clinical manifestations are strictly related in all of them, such as abdominal pain and distention, however the development pathways may differ. The diagnosis criteria are also different, in CD features immunological markers such as IgA, while WA presents IgE markers, and NCGS does not have any specific immunological marker. The consumption of wheat, and consequently gluten-related products, is historical and it has grown over human evolution, which leads us to question what role this component plays in other diseases, and the consequence of a gluten-free diet (GFD). GFD is the treatment with major efficacy in CD and, likewise, is related with improvements in neuropathologies (autism spectrum disorder, anxiety, depression and schizophrenia), obesity, and the gut microbiome. Further investigation is required. GFD studies in people who do not present CD seem to be safe, especially when products containing gluten are not replaced by processed gluten-free products but by minimally processed foods, and also when the food strategy is accompanied by a qualified professional.
... In particular, T-cells from coeliac patients recognize epitopes containing the repetitive sequences present in omega-gliadins (Sollid et al., 2012). Yet, omega-gliadins are only a minor group among prolamins, amount-wise, and have been until recently less well characterized than other prolamin classes for their polymorphism and contribution to gluten properties (Altenbach et al., 2018). ...
Among wheat storage proteins, omega-gliadins display a singular amino acid sequence only composed by repetitive sequences. They have been described as a major wheat allergen and also contain T-cell epitopes implicated in coeliac disease. To study the structural and biophysical properties of omega-gliadins and other gliadins (e.g., gluten network formation, allergic response), highly purified and concentrated fractions are needed. In the present work, we used chromatography media screening to improve their fractionation. A S Ceramic Hyper D (Pall) resin was selected for its ability to concentrate omega-gliadins in the first elution peaks. Gamma- and beta-gliadins were then separated by hydrophobic interaction chromatography and eluted with an ethanol gradient. Alpha-gliadins were purified by gel filtration. Each fraction was characterized by electrophoresis and reverse phase HPLC. The developed preparative protocol enabled the purification of several grams of highly purified gliadins to the detriment of the yield.
... The ability of fermented gluten to degranulate basophile cells was significantly reduced (p = 0.0001) by the chimeric anti-ω IgE, which recognizes PQQPFPQQ, a motif that occurs twenty times in ω5-type gliadin sequences [25]. The ω5 gliadin is considered a significant allergen in wheat allergic adults suffering from WDEIA despite the low amount in wheat gluten, around 5-10% [61,62]. The ω5 gliadins molecular weight is around 53 kDa, and the aminoacid sequence contains a large repetitive domain flanked by a short C-ter region. ...
Wheat is a worldwide staple food, yet some people suffer from strong immunological reactions after ingesting wheat-based products. Lactic acid bacteria (LAB) constitute a promising approach to reduce wheat allergenicity because of their proteolytic system. In this study, 172 LAB strains were screened for their proteolytic activity on gluten proteins and α-amylase inhibitors (ATIs) by SDS-PAGE and RP-HPLC. Gliadins, glutenins, and ATI antigenicity and allergenicity were assessed by Western blot/Dot blot and by degranulation assay using RBL-SX38 cells. The screening resulted in selecting 9 high gluten proteolytic strains belonging to two species: Enterococcus faecalis and Lactococcus lactis. Proteomic analysis showed that one of selected strains, Lc. lactis LLGKC18, caused degradation of the main gluten allergenic proteins. A significant decrease of the gliadins, glutenins, and ATI antigenicity was observed after fermentation of gluten by Lc. lactis LLGKC18, regardless the antibody used in the tests. Also, the allergenicity as measured by the RBL-SX38 cell degranulation test was significantly reduced. These results indicate that Lc. lactis LLGKC18 gluten fermentation can be deeply explored for its capability to hydrolyze the epitopes responsible for wheat allergy.
