title: Addressing proteolytic efficiency in enzymatic degradation therapy for celiac disease.
abstract: Celiac disease is triggered by partially digested gluten proteins. Enzyme therapies that complete protein digestion in vivo could support a gluten-free diet, but the barrier to completeness is high. Current options require enzyme amounts on the same order as the protein meal itself. In this study, we evaluated proteolytic components of the carnivorous pitcher plant (Nepenthes spp.) for use in this context. Remarkably low doses enhance gliadin solubilization rates, and degrade gliadin slurries within the pH and temporal constraints of human gastric digestion. Potencies in excess of 1200:1 (substrate-to-enzyme) are achieved. Digestion generates small peptides through nepenthesin and neprosin, the latter a novel enzyme defining a previously-unknown class of prolyl endoprotease. The digests also exhibit reduced TG2 conversion rates in the immunogenic regions of gliadin, providing a twin mechanism for evading T-cell recognition. When sensitized and dosed with enzyme-treated gliadin, NOD/DQ8 mice did not show intestinal inflammation, when compared to mice challenged with only pepsin-treated gliadin. The low enzyme load needed for effective digestion suggests that gluten detoxification can be achieved in a meal setting, using metered dosing based on meal size. We demonstrate this by showing efficient antigen processing at total substrate-to-enzyme ratios exceeding 12,000:1.
authors.affiliation: Department of Biochemistry and Molecular Biology and the Southern Alberta Cancer Research Institute, University of Calgary, Calgary, Alberta, Canada.|Structural Mass Spectrometry and Proteomics Unit, Institut Pasteur, CNRS UMR 3528, Paris, France.
authors.forename: Martial
authors.initials: M
authors.lastname: Rey
authors.affiliation: Department of Biochemistry and Molecular Biology and the Southern Alberta Cancer Research Institute, University of Calgary, Calgary, Alberta, Canada.
authors.forename: Menglin
authors.initials: M
authors.lastname: Yang
authors.affiliation: Department of Biochemistry and Molecular Biology and the Southern Alberta Cancer Research Institute, University of Calgary, Calgary, Alberta, Canada.
authors.forename: Linda
authors.initials: L
authors.lastname: Lee
authors.affiliation: Department of Biochemistry and Molecular Biology and the Southern Alberta Cancer Research Institute, University of Calgary, Calgary, Alberta, Canada.
authors.forename: Ye
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Biochemistry and Molecular Biology and the Southern Alberta Cancer Research Institute, University of Calgary, Calgary, Alberta, Canada.
authors.forename: Joey G
authors.initials: JG
authors.lastname: Sheff
authors.affiliation: Graz University of Technology, Institute of Molecular Biotechnology, Graz, Austria.
authors.forename: Christoph W
authors.initials: CW
authors.lastname: Sensen
authors.affiliation: Institute of Microbiology, Academy of Sciences of the Czech Republic, and Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, Czech Republic.
authors.forename: Hynek
authors.initials: H
authors.lastname: Mrazek
authors.affiliation: Institute of Microbiology, Academy of Sciences of the Czech Republic, and Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Halada
authors.affiliation: Institute of Microbiology, Academy of Sciences of the Czech Republic, and Department of Biochemistry, Faculty of Science, Charles University in Prague, Prague, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Man
authors.affiliation: Farncombe Family Digestive Health Research Institute, McMaster University, Hamilton, ON, Canada.
authors.forename: Justin L
authors.initials: JL
authors.lastname: McCarville
authors.affiliation: Farncombe Family Digestive Health Research Institute, McMaster University, Hamilton, ON, Canada.
authors.forename: Elena F
authors.initials: EF
authors.lastname: Verdu
authors.affiliation: Department of Biochemistry and Molecular Biology and the Southern Alberta Cancer Research Institute, University of Calgary, Calgary, Alberta, Canada.
authors.forename: David C
authors.initials: DC
authors.lastname: Schriemer
mesh_terms: D000818:Animals; D002446:Celiac Disease / Q000201:enzymology / Q000276:immunology / Q000628:therapy*; D055050:Diet, Gluten-Free; D004330:Drosophila / Q000378:metabolism; D057487:Enzyme Therapy; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism*; D005903:Gliadin / Q000378:metabolism*; D005983:Glutens / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007249:Inflammation / Q000276:immunology / Q000378:metabolism / Q000517:prevention & control; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D059748:Proteolysis; D011503:Transglutaminases / Q000378:metabolism*
chemical_list: D005983:Glutens; D005903:Gliadin; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D019204:GTP-Binding Proteins