Investigation of Protein and Epitope Characteristics of Oats and Its Implications for Celiac Disease

Abstract

The use of pure oats (oats cultivated with special care to avoid gluten contamination from wheat, rye, and barley) in the gluten-free diet (GFD) represents important nutritional benefits for the celiac consumer. However, emerging evidence suggests that some oat cultivars may contain wheat gliadin analog polypeptides. Consequently, it is necessary to screen oats in terms of protein and epitope composition to be able to select safe varieties for gluten-free applications. The overall aim of our study is to investigate the variability of oat protein composition directly related to health-related and techno-functional properties. Elements of an oat sample population representing 162 cultivated varieties from 20 countries and the protein composition of resulting samples have been characterized. Size distribution of the total protein extracts has been analyzed by size exclusion-high performance liquid chromatography (SE-HPLC) while the 70% ethanol-extracted proteins were analyzed by RP-HPLC. Protein extracts separated into three main groups of fractions on the SE-HPLC column: polymeric proteins, avenins (both containing three subgroups based on their size), and soluble proteins, representing respectively 68.79–86.60, 8.86–27.72, and 2.89–11.85% of the total protein content. The ratio of polymeric to monomeric proteins varied between 1.37 and 3.73. Seventy-six reversed phase-HPLC-separated peaks have been differentiated from the ethanol extractable proteins of the entire population. Their distribution among the cultivars varied significantly, 6–23 peaks per cultivar. The number of appearances of peaks also showed large variation: one peak has been found in 107 samples, while 15 peaks have been identified, which appeared in less than five cultivars. An estimation method for ranking the avenin-epitope content of the samples has been developed by using MS spectrometric data of collected RP-HPLC peaks and bioinformatics methods. Using ELISA methodology with the R5 antibody, a high number of the investigated samples were found to be contaminated with wheat, barley, or rye.

Effect of additional water supply during grain filling on protein composition and epitope characteristics of winter oats

“…As it was realized in our recent previous study on oat proteins ( Gell et al, 2021 ), more than 99% of the total amount of oat flour proteins can be extracted in the first step of the extraction procedure of Gupta et al (1993) , without applying sonication. Comparison of samples has been carried out, therefore, using this simplified, one-step procedure.…”

“…1A ): altogether 10 peaks, from which two major peaks - P2 and P6 with retention times: 6.4 and 9.6 min– and some minor ones have been observed on each chromatogram. Following the procedure used by Gell et al (2021) , the amounts of the three main protein groups have been determined by using the analytical data based on the SE-HPLC separation. The polymeric protein fraction consisted of five well-defined peaks (P1–P5) with retention times of 5.2, 6.4, 7.4, 7.9, and 8.3 min, respectively.…”

“…The analytical procedures applied in this study were identical to those used and described in depth by our previous publication ( Gell et al, 2021 ), using the same methods, equipment, and separation procedures used by the same operator.…”

“…The analytical data derived from the RP-HPLC-based separation provides the basis to predict the individual and cumulative amounts of DQ2.5-ave-1a (PYPEQEEPF), DQ2.5-ave-1b (PYPEQEQPF) ( Vader et al, 2003 ; Arentz-Hansen et al, 2004 ), DQ2.5-ave-1c (PYPEQEQPI) ( Hardy et al, 2015 ), and DQ2.5-ave-2 (PYPEQQPF) epitopes of oat using the procedure of Gell et al (2021) based on the detailed proteomic data of Tanner et al (2019) .…”

“…Despite the valid and serious limitations of the prediction method discussed in depth by Gell et al (2021) - with the lack of any other (better) relatively high throughput and cheap method that applies to large sample populations – the method is suitable to be used as a pre-selection screening tool in oat breeding.…”