Accidental gluten consumption significantly affects the health and quality of life for individuals who experience immune responses to dietary gluten present in cereals, like wheat, barley or rye. Accordingly, alongside an increased demand for gluten-free food, the gluten-degrading capacity of various enzymes present in fungi, bacteria or plants has also been exploited to develop novel drug-based therapies and digestive supplements.
In this project, digestion efficiency of GluteGuard, an enzyme supplement enriched in caricain, a thiol-proteinase isolated from crude papaya latex, was investigated in combination with gastric and pancreatic enzymes and compared to the gluten digestion efficiency of Aspergillus niger prolyl-endopeptidase (AnPep). Gluten proteins extracted from the wheat cultivar Baxter were digested with the enzyme supplements with and without the presence of pepsin, trypsin and chymotrypsin and monitored in a time course experiment using data dependent acquisition (DDA) and multiple reaction monitoring (MRM) mass spectrometry.
Altogether 215 gluten specific peptides representing six gluten protein sub-types have been identified and monitored across the experimental series. Our results indicate that while AnPep does not digest all the gluten sub-types, caricain digestion is independent of the gluten sub-types and compared to AnPep results in a larger proportion of peptides with decreased abundance levels. Positional mapping of the cleaved peptides shows that most of the epitope dense regions are digested, however similar results obtained both for carician and AnPep indicate the efficiency and rate of digestion is highly variable within the same protein sequence and depends on the simultaneous peptide liberation and digestion caused by the applied enzyme combinations. Results of three highly immunogenic α-gliadin sequences demonstrated both increasing and decreasing peptide abundance patterns of the same protein; therefore, relying on MRM abundance changes without further investigating the position of monitored peptides within the protein sequence can result in false conclusions.