X6: A Novel Antibody for Potential Use in Gluten Quantification
Authors: Shatalova, Aleksandrina; Shatalov, Ivan; Lebedin, Yuri
Gliadin is a fraction of gluten, known to trigger celiac disease in susceptible people. To date, the life-long gluten-free diet is used for the prevention of this disease. Hence, methods for gluten control in foods are of significant importance. Being one of the most-used methods used for this purpose, ELISA should use high-affinity antibodies to gliadin peptides involved into celiac process. This study investigates the characteristics of a novel anti-gliadin antibody X6. We found the QXQPFPXP site to be a recognized epitope that provides specific binding of the antibody to cereal prolamins involved in celiac disease manifestation. A specificity study using immunoblotting shows the recognition of wheat, barley and rye proteins-as well as alpha-gliadin homologs from non-edible cereals (Dasypyrum villosum). Reactivity to avenin was less pronounced, as this protein does not contain the PFP motif most critical for antibody recognition. The proteins ofZea maysandSetaria italicawere not recognized by X6. X6-based ELISA highly correlated with R5 and G12, which are Codex Alimentarius standards in the quantitative assessment of gluten content (Pearson's R = 0.86 and 0.87, respectively). Qualitative assessment revealed no significant differences between R5 and G12 and X6.
Gluten hydrolyzing activity ofBacillusspp isolated from sourdough
MICROBIAL CELL FACTORIES
Authors: Rashmi, Bennur Somashekharaiah; Gayathri, Devaraja; Vasudha, Mahanthesh; Prashantkumar, Chakra Siddappa; Swamy, Chidanandamurthy Thippeswamy; Sunil, Kumar S.; Somaraja, Palegar Krishnappa; Prakash, Patil
Background Celiac disease is an intestinal chronic disorder with multifactorial etiology resulting in small intestinal mucosal injuries and malabsorption. In genetically predisposed individuals with HLA DQ2/DQ8 molecules, the gluten domains rich in glutamine and proline present gluten domains to gluten reactive CD4(+)T cells causing injury to the intestine. In the present experimental design, the indigenous bacteria from wheat samples were studied for their gluten hydrolyzing functionality. Results Proteolytic activity ofBacillus spp. was confirmed spectrophotometrically and studied extensively on gliadin-derived synthetic enzymatic substrates, natural gliadin mixture, and synthetic highly immunogenic 33-mer peptide. The degradation of 33-mer peptide and the cleavage specificities of the selected isolates were analyzed by tandem mass spectrometry. The gluten content of the sourdough fermented by the chosen bacterial isolates was determined by R5 antibody based competitive ELISA. All the tested isolates efficiently hydrolyzed Z-YPQ-pNA, Z-QQP-pNA, Z-PPF-pNA, and Z-PFP-pNA and also cleaved 33-mer immunogenic peptide extensively. The gluten content of wheat sourdough was found to be below 110 mg/kg. Conclusion It has been inferred that fourBacillus sppespecially GS 188 could be useful in developing gluten-reduced wheat food product for celiac disease prone individuals.