Article

Identification and analysis of multivalent proteolytically resistant peptides from gluten: Implications for Celiac Sprue

University of Oslo, Kristiania (historical), Oslo, Norway
Journal of Proteome Research (Impact Factor: 5). 08/2005; 4(5):1732-41. DOI: 10.1021/pr050173t
Source: PubMed

ABSTRACT Dietary gluten proteins from wheat, rye, and barley are the primary triggers for the immuno-pathogenesis of Celiac Sprue, a widespread immune disease of the small intestine. Recent molecular and structural analyses of representative gluten proteins, most notably alpha- and gamma-gliadin proteins from wheat, have improved our understanding of these pathogenic mechanisms. In particular, based on the properties of a 33-mer peptide, generated from alpha-gliadin under physiological conditions, a link between digestive resistance and inflammatory character of gluten has been proposed. Here, we report three lines of investigation in support of this hypothesis. First, biochemical and immunological analysis of deletion mutants of alpha-2 gliadin confirmed that the DQ2 restricted T cell response to the alpha-2 gliadin are directed toward the epitopes clustered within the 33-mer. Second, proteolytic analysis of a representative gamma-gliadin led to the identification of another multivalent 26-mer peptide that was also resistant to further gastric, pancreatic and intestinal brush border degradation, and was a good substrate of human transglutaminase 2 (TG2). Analogous to the 33-mer, the synthetic 26-mer peptide displayed markedly enhanced T cell antigenicity compared to monovalent control peptides. Finally, in silico analysis of the gluten proteome led to the identification of at least 60 putative peptides that share the common characteristics of the 33-mer and the 26-mer peptides. Together, these results highlight the pivotal role of physiologically generated, proteolytically stable, TG2-reactive, multivalent peptides in the immune response to dietary gluten in Celiac Sprue patients. Prolyl endopeptidase treatment was shown to abolish the antigenicity of both the 33-mer and the 26-mer peptides, and was also predicted to have comparable effects on other proline-rich putatively immunotoxic peptides identified from other polypeptides within the gluten proteome.

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    • "Sequence of 300 amino acids usually forms hordeins or gliadins however, it is known that 8e17 amino acid residues peptides can still demonstrate toxicity for celiac patients (Shan et al., 2005; Tye- Din et al., 2010). Malting and fermentation processes alter the barley/wheat prolamins into short peptide fragments and amino acids (Colgrave et al., 2012; Comino et al., 2013) with immunization properties and hinder their further separation. "
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    • "This method revealed the presence of a 33-mer peptide that is highly resistant to gastrointestinal digestion, deamidated at multiple Gln residues by TG2, and immunogenic toward most celiac patient biopsy-derived intestinal T cell lines when presented on DQ2 + antigen-presenting cells (Figure 2) (Shan et al., 2002). Similar mock digestion of another representative gluten protein, γ5-gliadin, revealed a 26-mer peptide with these same properties (Shan et al., 2005b). As such, simulated gastrointestinal digestion of gluten is an effective method for identifying disease-relevant peptide substrates for preclinical testing of glutenases. "
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    • "Among the different gluten epitopes that have been identified, the a-gliadin epitopes are considered the most immunogenic (Arentz-Hansen et al., 2000a,b, 2002; Camarca et al., 2009; Janatuinen et al., 2002; Maiuri et al., 2003; Molberg et al., 2003; Schuppan et al., 2003; Vader et al., 2002) of which the Glia-a9 is a major immunodominant epitope. The Glia-a9 epitope sequence (aI) is part of the proteolytic resistant 33-mer identified by Shan et al. (2002, 2005) that has a high T-cell stimulatory effect. The 33-mer sequence (LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF) is only present in D-genome protein sequences of a/b-gliadins (Salentijn, personal communication). "
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