Structural conformation and self‐assembly process of p31‐43 gliadin peptide in aqueous solution. Implications for celiac disease

Abstract

Celiac disease (CeD) is a highly prevalent chronic immune-mediated enteropathy developed in genetically predisposed individuals after ingestion of a group of wheat proteins (called gliadins and glutenins). The 13mer agliadin peptide, p31-43, induces proinflammatory responses, observed by in vitro assays and animal models, that may contribute to innate immune mechanisms of CeD pathogenesis. Since a cellular receptor for p31-43 has not been identified, this raises the question of whether this peptide could mediate different biological effects. In this work, we aimed to characterize the p31-43 secondary structure by different biophysical and in silico techniques. By dynamic light scattering and using an oligomer/fibril-sensitive fluorescent probe, we showed the presence of oligomers of this peptide in solution. Furthermore, atomic force microscopy analysis showed p31-43 oligomers with different height distribution. Also, peptide concentration had a very strong influence on peptide self-organization process. Oligomers gradually increased their size at lower concentration. Whereas, at higher ones, oligomers increased their complexity, forming branched structures. By CD, we observed that p31-43 self-organized in a polyproline II conformation in equilibrium with b-sheets-like structures, whose pH remained stable in the range of 3-8. In addition, these findings were supported by molecular dynamics simulation. The formation of p31-43 nanostructures with increased b-sheet structure may help to explain the molecular etiopathogenesis in the induction of proinflammatory effects and subsequent damage at the intestinal mucosa in CeD.

Evaluation of Peptide/Protein Self-Assembly and Aggregation by Spectroscopic Methods

“…Additionally, it can interconvert into other forms such as β-turns and β-strands because of the proximity of the corresponding dihedral angles [ 44 ]. PPII plays a vital role in protein–protein interactions [ 45 ] and aggregation, as it was detected as the intermediate motif during the self-aggregation of lysozyme [ 46 ], the Aβ-peptide [ 47 ] and gliadin peptides [ 48 , 49 ].…”

“…Recently, a Triazole-BODIPY-based probe was shown to detect, in a time-dependent manner, the first stages of Aβ-peptide oligomerization before fibril formation [ 104 ]. Additionally, the same BODIPY derivative identified the presence of oligomers of the toxic p31–43 peptide related to celiac disease [ 49 ]. This probe has proved to be a helpful tool in the rational screening of drugs that reduce peptide aggregation before the onset of disease.…”

“…The p31–43 peptide has been shown to aggregate into oligomers, but in this case, the peptide presents a stable PPII structure. As the concentration increases, a hypochromic displacement of the negative is detected [ 49 ]. In the case of proteins, the β-lactoglobulin, which is an α, β structured protein, have been shown to self-aggregate in a concentration-dependent manner which was detected by CD.…”

“…This effect is observed in the case of proline-rich peptides, given that proline is s a helix breaker. For example, in the case of the 33-mer and p31–43 gliadin peptides rich in prolines, both have a PPII structure, which in the presence of increasing concentrations of TFE converts to a β-sheet-like one [ 48 , 49 ]. This equilibrium occurs due to the propensity of PPII to interconvert into β turns and β-strands [ 44 ].…”

Coeliac Disease Pathogenesis: The Uncertainties of a Well-Known Immune Mediated Disorder

“…p31-43 was shown to inhibit the subunit of the chloride channel (CFTR) and thereby cause NF-kB activation, induction of IL-15 and TG2 activation, with a range of inflammatory consequences ( 88 ). Moreover, p31-43 is able to self-assemble in oligomers with potential important effects ( 89 ). In vivo analysis in a murine model showed that activation of the NLRP3 inflammasome, either by direct detection of oligomers or indirectly by sensing danger signals, is required for histological changes in the small intestinal mucosa ( 90 ).…”