Citation: Leszczy ´ nska, J.; Szczepankowska, A.K.; Majak, I.; Ma ´ nkowska, D.; Smoli ´ nska, B.; ´ Scieszka, S.; Diowksz, A.; Cukrowska, B.; Aleksandrzak-Piekarczyk, T. Reducing Immunoreactivity of Gluten Peptides by Probiotic Lactic Acid Bacteria for Dietary Management of Gluten-Related Diseases. Nutrients 2024, 16, 976. https://doi.org/ 10.3390/nu16070976 Academic Editors: Magdalena Araya, Juan Rodriguez and Karla A. Bascuñán Received: 19 February 2024 Revised: 24 March 2024 Accepted: 26 March 2024 Published: 27 March 2024 Copyright: © 2024 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). nutrients Article Reducing Immunoreactivity of Gluten Peptides by Probiotic Lactic Acid Bacteria for Dietary Management of Gluten-Related Diseases Joanna Leszczy ´ nska 1 , Agnieszka K. Szczepankowska 2 , Iwona Majak 3 , Dorota Ma ´ nkowska 1 , Beata Smoli ´ nska 1 , Sylwia ´ Scieszka 4 , Anna Diowksz 4 , Bo ˙ zena Cukrowska 5 and Tamara Aleksandrzak-Piekarczyk 2, * 1 Institute of Natural Products and Cosmetics, Faculty of Biotechnology and Food Sciences, Łód´ z University of Technology, Stefanowskiego 2/22, 90-530 Łód´ z, Poland; joanna.leszczynska@p.lodz.pl (J.L.); dorota.mankowska@p.lodz.pl (D.M.); beata.smolinska@p.lodz.pl (B.S.) 2 Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawi ´ nskiego 5a, 02-106 Warsaw, Poland; agaszczep@ibb.waw.pl 3 Institute of Technology and Food Analysis, Faculty of Biotechnology and Food Sciences, Łód´ z University of Technology, Stefanowskiego 2/22, 90-530 Łód´ z, Poland; iwona.majak@p.lodz.pl 4 Institute of Fermentation Technology and Microbiology, Faculty of Biotechnology and Food Sciences, Łód´ z University of Technology, Wólcza ´ nska 171/173, 90-530 Łód´ z, Poland; sylwia.scieszka@p.lodz.pl (S. ´ S.); anna.diowksz@p.lodz.pl (A.D.) 5 Immunology Laboratory, Department of Pathomorphology, The Children’s Memorial Health Institute, Dzieci Polskich 20, 04-760 Warsaw, Poland; b.cukrowska@ipczd.pl * Correspondence: tamara@ibb.waw.pl; Tel.: +48-(22)-592-1213 Abstract: Immunoreactive gluten peptides that are not digested by peptidases produced by humans can trigger celiac disease, allergy and non-celiac gluten hypersensitivity. The aim of this study was to evaluate the ability of selected probiotic strains to hydrolyze immunoreactive gliadin peptides and to identify peptidase-encoding genes in the genomes of the most efficient strains. Residual gliadin immunoreactivity was measured after one- or two-step hydrolysis using commercial enzymes and bacterial peptidase preparations by G12 and R5 immunoenzymatic assays. Peptidase preparations from Lacticaseibacillus casei LC130, Lacticaseibacillus paracasei LPC100 and Streptococcus thermophilus ST250 strains significantly reduced the immunoreactivity of gliadin peptides, including 33-mer, and this effect was markedly higher when a mixture of these strains was used. In silico genome analyses of L. casei LC130 and L. paracasei LPC100 revealed the presence of genes encoding peptidases with the potential to hydrolyze bonds in proline-rich peptides. This suggests that L. casei LC130, L. paracasei LPC100 and S. thermophilus ST250, especially when used as a mixture, have the ability to hydrolyze immunoreactive gliadin peptides and could be administered to patients on a restricted gluten-free diet to help treat gluten-related diseases. Keywords: celiac disease; gluten-related diseases; gluten-free diet; endopeptidase; 33-mer peptide; peptidase-encoding genes; lactobacilli; probiotics 1. Introduction Gluten is the general name for water-insoluble prolamin proteins of cereals, which include gliadin in wheat, secalin in rye and hordein in barley. Gluten can trigger gluten- related diseases such as celiac disease (CD), allergy and non-celiac gluten hypersensitiv- ity [1]. The condition with the best understood pathomechanism associated with gluten intolerance is CD, which affects 0.7% of the world’s population. It is a chronic autoimmune enteropathy of the small intestine that occurs in individuals with a genetic predisposition manifested by the HLA-DQ2 and/or HLA-DQ8 haplotype [2]. Prolamines consist of multiple glutamine residues linked to prolines, making their structure highly complex and resistant to hydrolysis by proteolytic enzymes present in Nutrients 2024, 16, 976. https://doi.org/10.3390/nu16070976 https://www.mdpi.com/journal/nutrients