Enhancing gluten modification for celiac patients: harnessing microbial transglutaminase, ε-poly-l-lysine, and prolyl endopeptidase.

The immunogenicity of gluten poses a significant challenge for individuals with celiac disease. This study aimed to reduce gluten immunogenicity using a dual treatment strategy: microbial transglutaminase and ε-poly-l-lysine (ε-PL) for structural modification, followed by enzymatic degradation with prolyl endopeptidase. Structural characterization of gluten proteins treated with transglutaminase and ε-PL revealed substantial modification of gliadins, with higher ε-PL concentrations showing more pronounced effects. Low-molecular-weight glutenins were less affected, and no deamidation or impact on digestibility was observed. Although an 80% reduction in R5 epitopes was achieved, gluten levels still exceeded the gluten-free threshold. The application of prolyl endopeptidase successfully reduced gluten content below this threshold, particularly in modified samples. When prolyl endopeptidase was used as a digestive adjuvant, only modified samples reached gluten-free levels. These findings show that combining structural modification and enzymatic degradation can achieve safe gluten-free levels, enabling the development of alternative diets for people with gluten-related disorders.