title: Generation of food-grade recombinant Lactobacillus casei delivering Myxococcus xanthus prolyl endopeptidase.
abstract: Prolyl endopeptidases (PEP) (EC 3.4.21.26), a family of serine proteases with the ability to hydrolyze the peptide bond on the carboxyl side of an internal proline residue, are able to degrade immunotoxic peptides responsible for celiac disease (CD), such as a 33-residue gluten peptide (33-mer). Oral administration of PEP has been suggested as a potential therapeutic approach for CD, although delivery of the enzyme to the small intestine requires intrinsic gastric stability or advanced formulation technologies. We have engineered two food-grade Lactobacillus casei strains to deliver PEP in an in vitro model of small intestine environment. One strain secretes PEP into the extracellular medium, whereas the other retains PEP in the intracellular environment. The strain that secretes PEP into the extracellular medium is the most effective to degrade the 33-mer and is resistant to simulated gastrointestinal stress. Our results suggest that in the future, after more studies and clinical trials, an engineered food-grade Lactobacillus strain may be useful as a vector for in situ production of PEP in the upper small intestine of CD patients.
authors.affiliation: Instituto de Productos Lácteos de Asturias (IPLA-CSIC), 33300, Villaviciosa, Asturias, Spain.
authors.forename: Patricia
authors.initials: P
authors.lastname: Alvarez-Sieiro
authors.forename: Maria Cruz
authors.initials: MC
authors.lastname: Martin
authors.forename: Begoña
authors.initials: B
authors.lastname: Redruello
authors.forename: Beatriz
authors.initials: B
authors.lastname: Del Rio
authors.forename: Victor
authors.initials: V
authors.lastname: Ladero
authors.forename: Brad A
authors.initials: BA
authors.lastname: Palanski
authors.forename: Chaitan
authors.initials: C
authors.lastname: Khosla
authors.forename: Maria
authors.initials: M
authors.lastname: Fernandez
authors.forename: Miguel A
authors.initials: MA
authors.lastname: Alvarez
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism* / Q000627:therapeutic use; D002446:Celiac Disease / Q000188:drug therapy* / Q000378:metabolism; D016503:Drug Delivery Systems; D057487:Enzyme Therapy; D041981:Gastrointestinal Tract / Q000378:metabolism / Q000382:microbiology; D015870:Gene Expression; D005983:Glutens / Q000378:metabolism; D006801:Humans; D007780:Lacticaseibacillus casei / Q000235:genetics* / Q000378:metabolism; D016941:Myxococcus xanthus / Q000201:enzymology* / Q000235:genetics; D000085822:Prolyl Oligopeptidases; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism* / Q000627:therapeutic use
chemical_list: D001426:Bacterial Proteins; D005983:Glutens; D012697:Serine Endopeptidases; C000705287:PREPL protein, human; D000085822:Prolyl Oligopeptidases
references.citation: Expert Opin Drug Discov. 2013 Apr;8(4):445-57
references.pmid: 23293929
references.citation: Appl Microbiol Biotechnol. 2006 Jul;71(4):394-406
references.pmid: 16628446
references.citation: Am J Clin Nutr. 2012 Mar;95(3):670-7
references.pmid: 22258271
references.citation: Nature. 1970 Aug 15;227(5259):680-5
references.pmid: 5432063
references.citation: Aliment Pharmacol Ther. 2012 Feb;35(4):403-13
references.pmid: 22225517
references.citation: J Chromatogr A. 2010 Jun 18;1217(25):4167-83
references.pmid: 20181349
references.citation: Trends Biotechnol. 2011 Oct;29(10):499-508
references.pmid: 21665301
references.citation: Gene. 1993 Feb 14;124(1):133-4
references.pmid: 8382652
references.citation: Br J Nutr. 2013 Nov;110(10):1828-36
references.pmid: 23657133
references.citation: Vaccine. 2010 Sep 24;28(41):6714-22
references.pmid: 20699130
references.citation: Res Microbiol. 2012 Jun;163(5):357-65
references.pmid: 22564557
references.citation: Biotechnol Bioeng. 2005 Dec 20;92(6):674-84
references.pmid: 16136593
references.citation: Immunol Lett. 2008 Aug 15;119(1-2):78-83
references.pmid: 18547649
references.citation: Arch Biochem Biophys. 1998 Oct 1;358(1):141-8
references.pmid: 9750174
references.citation: World J Gastroenterol. 2012 Nov 14;18(42):6036-59
references.pmid: 23155333
references.citation: Crit Rev Microbiol. 2012 Aug;38(3):185-90
references.pmid: 22168378
references.citation: J Appl Microbiol. 1998 May;84(5):759-68
references.pmid: 9674129
references.citation: J Bacteriol. 2010 May;192(10):2647-8
references.pmid: 20348264
references.citation: Lett Appl Microbiol. 2009 May;48(5):618-22
references.pmid: 19416463
references.citation: Clin Vaccine Immunol. 2008 Sep;15(9):1429-35
references.pmid: 18632920
references.citation: Appl Environ Microbiol. 2002 Dec;68(12):6172-81
references.pmid: 12450842
references.citation: J Appl Microbiol. 2004;97(4):749-56
references.pmid: 15357724
references.citation: Am J Physiol Gastrointest Liver Physiol. 2002 Oct;283(4):G996-G1003
references.pmid: 12223360
references.citation: Methods Enzymol. 2012;502:241-71
references.pmid: 22208988
references.citation: Appl Environ Microbiol. 1989 Sep;55(9):2130-7
references.pmid: 2679379
references.citation: J Biochem. 1991 Dec;110(6):873-8
references.pmid: 1840588
references.citation: J Biol Chem. 1980 May 25;255(10):4786-92
references.pmid: 6768725
references.citation: Drug Discov Today. 2003 Aug 1;8(15):692-700
references.pmid: 12927512
references.citation: Biochem J. 2004 Oct 15;383(Pt 2):311-8
references.pmid: 15245330
references.citation: J Dairy Sci. 1997 Jun;80(6):1031-7
references.pmid: 9201571
references.citation: Semin Immunopathol. 2012 Jul;34(4):497-511
references.pmid: 22572887
references.citation: Cell Mol Life Sci. 2014 Jan;71(2):183-203
references.pmid: 23516017
references.citation: Lancet. 2003 Aug 2;362(9381):383-91
references.pmid: 12907013
references.citation: PLoS One. 2008 Feb 20;3(2):e1614
references.pmid: 18286171
references.citation: Nat Rev Microbiol. 2008 May;6(5):349-62
references.pmid: 18345021
references.citation: J Pharmacol Exp Ther. 2004 Oct;311(1):213-9
references.pmid: 15143130
references.citation: J Bacteriol. 2003 Jun;185(11):3288-96
references.pmid: 12754226
references.citation: Clin Gastroenterol Hepatol. 2005 Jul;3(7):687-94
references.pmid: 16206502
references.citation: J Dairy Sci. 1987 Jan;70(1):1-12
references.pmid: 3106442
references.citation: Crit Rev Microbiol. 2002;28(4):281-370
references.pmid: 12546196
references.citation: J Pharmacol Exp Ther. 2005 Jan;312(1):19-26
references.pmid: 15358813
references.citation: Lancet. 2001 Aug 4;358(9279):356-61
references.pmid: 11502314
references.citation: Appl Environ Microbiol. 2011 Mar;77(6):2174-9
references.pmid: 21257814
references.citation: Gene. 1991 Apr;100:189-94
references.pmid: 1905257
references.citation: FEMS Microbiol Lett. 2004 Aug 15;237(2):385-91
references.pmid: 15321687
references.citation: Virology. 1998 Oct 10;250(1):185-93
references.pmid: 9770432
references.citation: Science. 2002 Sep 27;297(5590):2275-9
references.pmid: 12351792
references.citation: Eur J Gastroenterol Hepatol. 1997 Nov;9(11):1097-9
references.pmid: 9431901
references.citation: Gastroenterology. 2009 Dec;137(6):1912-33
references.pmid: 19766641
references.citation: Scand J Immunol. 2011 Oct;74(4):335-41
references.pmid: 21615450
references.citation: J Immunol. 2004 Aug 1;173(3):1757-62
references.pmid: 15265905
references.citation: Appl Environ Microbiol. 2000 Jun;66(6):2599-604
references.pmid: 10831443