Abstract
Wheat gluten, and related prolamin proteins in rye, barley and oats cause the immune-mediated gluten intolerance syndrome, coeliac disease. Foods labelled as gluten-free which can be safely consumed by coeliac patients, must not contain gluten above a level of 20 mg/Kg. Current immunoassay methods for detection of gluten can give conflicting results and may underestimate levels of gluten in foods. Mass spectrometry methods have great potential as an orthogonal method, but require curated protein sequence databases to support method development. The GluPro database has been updated to include avenin-like sequences from bread wheat (n = 685; GluPro v1.1) and genes from the sequenced wheat genome (n = 699; GluPro v 1.2) and Triticum turgidum ssp durum (n = 210; GluPro v 2.1). Companion databases have been developed for prolamin sequences from barley (n = 64; GluPro v 3.0), rye (n = 41; GluPro v 4.0), and oats (n = 27; GluPro v 5.0) and combined to provide a complete cereal prolamin database, GluPro v 6.1 comprising 1,041 sequences. Analysis of the coeliac toxic motifs in the curated sequences showed that they were absent from the minor avenin-like proteins in bread and durum wheat and barley, unlike the related avenin proteins from oats. A comparison of prolamin proteins from the different cereal species also showed α- and γ-gliadins in bread and durum wheat, and the sulphur poor prolamins in all cereals had the highest density of coeliac toxic motifs. Analysis of ion-mobility mass spectrometry data for bread wheat (cvs Chinese Spring and Hereward) showed an increased number of identifications when using the GluPro v1.0, 1.1 and 1.2 databases compared to the limited number of verified sequences bread wheat sequences in reviewed UniProt. This family of databases will provide a basis for proteomic profiling of gluten proteins from all the gluten containing cereals and support identification of specific peptide markers for use in development of new methods for gluten quantitation based on coeliac toxic motifs found in all relevant cereal species.
Highlights
Wheat is one of the most important crop globally, with the combined production with related cereal species (barley (Hordeum vulgare), rye (Secale cereale), and oats (Avena sativa) exceeding ∼95,026 million tonnes in 2017 [1]
Database Construction Sequence sets of seed storage prolamins from T. turgidum ssp durum (GluPro v 2.0), H. vulgare (GluPro v 3.0), S. cereale (GluPro v 4.0) and A. sativa (GluPro v 5.0) were created independently and an update of the bread wheat (T. aestivum) database was undertaken to enrich it with avenin-like sequences (GluPro v 1.1) (Figures S1, S2)
Additional prolamin seed storage protein sequence databases were developed for other cereal species including T. turgidum ssp durum, barley, rye and oats (Figures S1, S2)
Summary
Wheat is one of the most important crop globally, with the combined production with related cereal species (barley (Hordeum vulgare), rye (Secale cereale), and oats (Avena sativa) exceeding ∼95,026 million tonnes in 2017 [1]. The major storage protein fractions in cereal grains are defined as prolamins based on their solubility in mixtures of alcohol and water and their high contents of glutamine and proline These proteins account for up to 80% of total protein content in wheat, barley and rye [2, 3] but are relatively minor components in oats [4]. The gluten proteins of wheat form a visco-elastic network when wheat flour is mixed with water, which enables the production of leavened bread and other products (including pasta and noodles) These properties are not shared by the prolamins in related cereals (barley, rye and oats), restricting the use of these cereals in food processing, their sequences are related to those of wheat gluten proteins. The term gluten strictly applies only to wheat prolamins, it is defined in a regulatory context as; “the protein fraction from wheat, barley, rye, oats or their crossbred varieties and derivatives thereof, to which some persons are intolerant and that is insoluble in water and 0.5 M NaCl” [5]
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