# Microbial Peptides: Key Players in Reducing Gluten Immunogenicity Through Peptide Degradation
## 3 Treatment for Gluten Elimination: Pros and Cons
Optimization of such bioprocesses could provide a broader range of safer bakery products, enhancing dietary diversity and compliance for gluten-sensitive populations. Beyond enzymatic and fermentation strategies, patient education and awareness remain critical components of successful gluten management. The risk of inadvertent gluten exposure persists due to the prevalence of gluten in processed foods, crosscontact during food preparation, and unclear labeling [9]. Enzymatic treatments should therefore be viewed as complementary tools rather than replacements for strict gluten avoidance. Overreliance on these supplements may engender a false sense of security, potentially leading to increased gluten ingestion and exacerbation of symptoms [45]. Looking forward, the future of gluten elimination therapies lies in the integration of multiple strategies. Combined enzyme formulations, improved delivery systems, and synergistic use of fermentation and biotechnological tools could enhance treatment outcomes. Additionally, personalized medicine approaches, considering individual genetic, immunological, and microbiome profiles, may allow tailored interventions that optimize efficacy and safety. The ultimate goal remains the development of safe, effective, and accessible therapies that improve the quality of life for individuals with gluten-related disorders, reducing the burden of strict dietary restrictions while preventing disease progression. Enzymes Capable of Degrading Immunogenic Peptides and Their Industrial Applications from Microorganisms  
Gluten-related disorders, such as CeD and NCGS, stem from the inability of the human digestive system to fully degrade certain proline and glutamine-rich peptides derived from gluten proteins, leading to the characteristic gluten immune responses in genetically predisposed individuals [42; 43]. Consequently, microbial and plant-derived peptidases that can target these resistant sequences have been extensively investigated for their potential to detoxify gluten and mitigate disease symptoms. Gluten-degrading enzymes are generally classified into three main groups according to their substrate specificity and catalytic mechanisms (Figure 2):  
1. Prolyl Endopeptidaseases (PEP):  
PEPs specifically cleave peptide bonds at the carboxyl side of proline residues, which are highly abundant in immunogenic gluten peptides. Bacterial PEPs have been isolated from species such as _Flavobacterium meningogepictium_ (FM-PEP), _Splingomonas capsulata_ (SC-PEP, also known as ALV002), and _Myxococcus xanthus_ (MX-PEP [54; 55; 56]). These enzymes exhibit robust proteolytic activity against immunotoxic sequences like the 33-mer from \(\alpha\)-gliadin, which is resistant to mammalian gastrointestinal proteases. PEPs from _F. meningogepictum_ and _S. capsulata_ belong to the S9A family of serine proteases and possess a conserved catalytic triad composed of Ser-His-Asp (Ser554, His660, and Asp627 in FM-PEP) [57]. This triad forms a charge relay system critical for the nucleophilic attack on peptide bonds adjacent to internal proline residues. Structural analyses have further revealed that the substrate binding pocket of PEPs accommodates the pyrrolidine ring of proline through hydrophobic interactions, contributing to their substrate specificity [58]. However, their activity is optimal at neutral to slightly alkaline pH and they are sensitive to pepsin degradation, limiting their effectiveness in the acidic stomach environment [38]. Recently, engineered PEP variants with enhanced acid stability and broader substrate specificity have been developed through protein engineering and directed evolution techniques. For example, modified SC-PEP mutants show increased catalytic efficiency in simulated gastric fluid, improving in vivo gluten detoxification potential [59]. Fungal PEPs such as the _Aspergillus niger_ prolyl endopeptidase (AN-PEP) have the advantage of acidic pH optima (around pH 4-5), enabling activity in the stomach before gluten peptides reach the small intestine [60].