Gut microbiome-derived prolyl peptidases from Segatella copri and Stenotrophomonas maltophilia degrade immunogenic gliadin peptides and restore intestinal barrier integrity in a Celiac disease model
Celiac disease (CeD) is an autoimmune enteropathy triggered by gluten-derived peptides that resist gastrointestinal digestion, notably the proline-rich 33-mer and 11-mer gliadin epitopes. Here, we describe a rational, data-centric strategy to identify gut microbiome-derived prolyl peptidases capable of degrading these immunogenic peptides. Integrating metagenomic mining with structure-based in silico screening, we identified two novel enzymes PSP692 and PSP464 from Segatella copri and Stenotrophomonas maltophilia, respectively. Recombinant expression and enzymatic characterization confirmed their activity under physiologically relevant conditions: PSP692 efficiently degrades the 33-mer at pH 6, while PSP464 targets the 11-mer at pH 4. Functional assays using CaCo-2 monolayers demonstrated that enzymatic degradation of gliadin epitopes significantly restored expression of tight junction proteins (ZO-1 and Occludin), reduced IL-6 secretion, and improved barrier integrity. These findings establish a foundational strategy for the discovery of microbiome-derived glutenases and provide a compelling case for combinatorial enzyme therapy to mitigate gluten immunotoxicity in CeD and related disorders.