title: Listeria monocytogenes aguA1, but not aguA2, encodes a functional agmatine deiminase: biochemical characterization of its catalytic properties and roles in acid tolerance.
abstract: Listeria monocytogenes is adaptable to low pH environments and therefore crosses the intestinal barrier to establish systemic infections. L. monocytogenes aguA1 and aguA2 encode putative agmatine deiminases (AgDIs) AguA1 and AguA2. Transcription of aguA1 and aguA2 was significantly induced at pH 5.0. Deletion of aguA1 significantly impaired its survival both in gastric fluid at pH 2.5 and in mouse stomach, whereas aguA2 deletion did not show significant defect of survival in gastric fluid. With agmatine as the sole substrate, AguA1 expressed in Escherichia coli was optimal at 25 °C and over a wide range of pH from 3.5 to 10.5. Recombinant AguA2 showed no deiminase activity. Site-directed mutagenesis revealed that all nine AguA1 mutants completely lost enzymatic activity. AguA2 acquired AgDI activity only when Cys-157 was mutated to glycine. AguA1 mutation at the same site, G157C, also inactivated the enzyme. Thus, we have discovered Gly-157 as a novel residue other than the known catalytic triad (Cys-His-Glu/Asp) in L. monocytogenes that is critical for enzyme activity. Of the two putative AgDIs, we conclude that only AguA1 functionally participates in the AgDI pathway and mediates acid tolerance in L. monocytogenes.
authors.affiliation: From the Zhejiang University Institute of Preventive Veterinary Medicine, Zhejiang Provincial Key Laboratory of Preventive Veterinary Medicine, and Key Laboratory of Molecular Animal Nutrition of the Ministry of Education, Hangzhou, Zhejiang 310058, China.
authors.forename: Changyong
authors.initials: C
authors.lastname: Cheng
authors.forename: Jianshun
authors.initials: J
authors.lastname: Chen
authors.forename: Chun
authors.initials: C
authors.lastname: Fang
authors.forename: Ye
authors.initials: Y
authors.lastname: Xia
authors.forename: Ying
authors.initials: Y
authors.lastname: Shan
authors.forename: Yuan
authors.initials: Y
authors.lastname: Liu
authors.forename: Guilan
authors.initials: G
authors.lastname: Wen
authors.forename: Houhui
authors.initials: H
authors.lastname: Song
authors.forename: Weihuan
authors.initials: W
authors.lastname: Fang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001667:Binding, Competitive; D002384:Catalysis; D019295:Computational Biology; D005260:Female; D005816:Genetic Complementation Test; D005998:Glycine / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism*; D020128:Inhibitory Concentration 50; D008089:Listeria monocytogenes / Q000201:enzymology*; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017386:Sequence Homology, Amino Acid; D013270:Stomach / Q000382:microbiology; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D006867:Hydrolases; C019626:agmatine deiminase; D005998:Glycine
keywords: Acid Tolerance; Agmatine Deiminase; Biochemical Characterization; Enzyme Kinetics; Enzyme Mechanisms; Kinetic Properties; Listeria monocytogenes; Microbiology; Site-directed Mutagenesis; Stress Response
references.citation: Microbiol Rev. 1986 Sep;50(3):314-52
references.pmid: 3534538
references.citation: Foodborne Pathog Dis. 2009 Apr;6(3):365-76
references.pmid: 19278345
references.citation: J Bacteriol. 2006 Feb;188(3):834-41
references.pmid: 16428386
references.citation: Protein Expr Purif. 2003 Jul;30(1):88-93
references.pmid: 12821325
references.citation: FEBS Lett. 2003 Jun 5;544(1-3):258-61
references.pmid: 12782327
references.citation: Mol Microbiol. 2001 Apr;40(2):465-75
references.pmid: 11309128
references.citation: Appl Environ Microbiol. 1996 May;62(5):1693-8
references.pmid: 8633868
references.citation: Methods Mol Biol. 2012;857:107-36
references.pmid: 22323219
references.citation: Microbiology (Reading). 2011 Nov;157(Pt 11):3150-3161
references.pmid: 21835877
references.citation: Int J Food Microbiol. 2000 Sep 25;60(2-3):137-46
references.pmid: 11016603
references.citation: J Bacteriol. 1998 Jul;180(14):3650-6
references.pmid: 9658010
references.citation: Methods. 2001 Dec;25(4):402-8
references.pmid: 11846609
references.citation: Nat Protoc. 2009;4(1):1-13
references.pmid: 19131951
references.citation: Cell Microbiol. 2009 May;11(5):703-9
references.pmid: 19191786
references.citation: Int J Food Microbiol. 2008 Jan 15;121(1):53-9
references.pmid: 18045718
references.citation: Environ Microbiol. 2009 Feb;11(2):432-45
references.pmid: 19196274
references.citation: Trends Biochem Sci. 2001 Aug;26(8):465-8
references.pmid: 11504612
references.citation: J Bacteriol. 2007 Feb;189(4):1254-65
references.pmid: 17028272
references.citation: Proteins. 2006 Sep 1;64(4):1010-23
references.pmid: 16779844
references.citation: J Bacteriol. 1988 Oct;170(10):4522-7
references.pmid: 3139630
references.citation: Infect Immun. 2006 May;74(5):2505-12
references.pmid: 16622185
references.citation: Microbiology (Reading). 2003 Mar;149(Pt 3):707-714
references.pmid: 12634339
references.citation: Biochemistry. 2005 Aug 9;44(31):10570-82
references.pmid: 16060666
references.citation: Bioorg Chem. 2010 Apr;38(2):62-73
references.pmid: 20036411
references.citation: Anal Biochem. 2000 Nov 15;286(2):257-64
references.pmid: 11067748
references.citation: Science. 2001 Oct 26;294(5543):849-52
references.pmid: 11679669
references.citation: Clin Microbiol Rev. 2001 Jul;14(3):584-640
references.pmid: 11432815
references.citation: Appl Environ Microbiol. 2009 May;75(9):2629-37
references.pmid: 19270124
references.citation: Biochemistry. 2010 Nov 2;49(43):9413-23
references.pmid: 20939536
references.citation: Mol Microbiol. 1993 Apr;8(1):143-57
references.pmid: 8388529
references.citation: J Med Microbiol. 2013 Jun;62(Pt 6):813-821
references.pmid: 23518652
references.citation: J Biol Chem. 2005 Oct 7;280(40):34080-7
references.pmid: 16091358
references.citation: Proteins. 2007 Feb 15;66(3):740-50
references.pmid: 17080455
references.citation: Biochemistry. 2006 Jan 31;45(4):1162-72
references.pmid: 16430212
references.citation: Microbiol Mol Biol Rev. 2003 Sep;67(3):429-53, table of contents
references.pmid: 12966143
references.citation: J Bacteriol. 2004 Mar;186(6):1902-4
references.pmid: 14996823
references.citation: Biochem Biophys Res Commun. 1991 Jan 15;174(1):32-6
references.pmid: 1989611
references.citation: J Microbiol Biotechnol. 2009 Mar;19(3):238-49
references.pmid: 19349748
references.citation: Phytochemistry. 2001 Apr;56(7):643-7
references.pmid: 11314948
references.citation: Bioinformatics. 2006 Jan 15;22(2):195-201
references.pmid: 16301204
references.citation: J Biol Chem. 2004 Apr 2;279(14):14001-8
references.pmid: 14701825
references.citation: Int J Food Microbiol. 1989 Jul;8(4):299-316
references.pmid: 2518321
references.citation: J Food Prot. 2001 Sep;64(9):1362-8
references.pmid: 11563513
references.citation: Appl Environ Microbiol. 2008 Jul;74(13):3921-34
references.pmid: 18441118
references.citation: Structure. 2004 Apr;12(4):657-67
references.pmid: 15062088
references.citation: Microbiology (Reading). 2007 Jul;153(Pt 7):2221-2230
references.pmid: 17600066