title: The D3 bacteriophage α-polymerase inhibitor (Iap) peptide disrupts O-antigen biosynthesis through mimicry of the chain length regulator Wzz in Pseudomonas aeruginosa.
abstract: Lysogenic bacteriophage D3 causes seroconversion of Pseudomonas aeruginosa PAO1 from serotype O5 to O16 by inverting the linkage between O-specific antigen (OSA) repeat units from α to β. The OSA units are polymerized by Wzy to modal lengths regulated by Wzz1 and Wzz2. A key component of the D3 seroconversion machinery is the inhibitor of α-polymerase (Iap) peptide, which is able to solely suppress α-linked long-chain OSA production in P. aeruginosa PAO1. To establish the target specificity of Iap for Wzyα, changes in OSA phenotypes were examined via Western immunoblotting for wzz1 and wzz2 single-knockout strains, as well as a wzz1 wzz2 double knockout, following the expression of iap from a tuneable vector. Increased induction of Iap expression completely abrogated OSA production in the wzz1 wzz2 double mutant, while background levels of OSA production were still observed in either of the single mutants. Therefore, Iap inhibition of OSA biosynthesis was most effective in the absence of both Wzz proteins. Sequence alignment analyses revealed a high degree of similarity between Iap and the first transmembrane segment (TMS) of either Wzz1 or Wzz2. Various topology prediction analyses of the Iap sequence consistently predicted the presence of a single TMS, suggesting a propensity for Iap to insert itself into the inner membrane (IM). The compromised ability of Iap to abrogate Wzyα function in the presence of Wzz1 or Wzz2 provides compelling evidence that inhibition occurs after Wzyα inserts itself into the IM and is achieved through mimicry of the first TMS from the Wzz proteins of P. aeruginosa PAO1.
authors: Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON, Canada.
authors: Véronique L
authors: VL
authors: Taylor
authors: Molly L
authors: ML
authors: Udaskin
authors: Salim T
authors: ST
authors: Islam
authors: Joseph S
authors: JS
authors: Lam
mesh_terms: D000595:Amino Acid Sequence; D000951:Antigens, Neoplasm; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001435:Bacteriophages / Q000502:physiology*; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D008958:Models, Molecular; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000145:classification / Q000235:genetics* / Q000378:metabolism / Q000821:virology*; D012703:Serotyping
chemical_list: D000951:Antigens, Neoplasm; D001426:Bacterial Proteins; C061613:osteosarcoma-associated antigen
references: J Bacteriol. 1983 Jul;155(1):203-12
references: 6190794
references: J Bacteriol. 2007 Jun;189(11):4141-52
references: 17384183
references: FEBS Lett. 2000 Dec 15;486(3):267-9
references: 11119716
references: J Biol Chem. 2012 May 4;287(19):15696-705
references: 22437828
references: Microbiologyopen. 2013 Apr;2(2):350-64
references: 23408725
references: Environ Microbiol. 2002 Dec;4(12):883-97
references: 12534470
references: Annu Rev Biochem. 2006;75:39-68
references: 16756484
references: J Biol Chem. 2009 Mar 13;284(11):7395-403
references: 19129185
references: Virology. 1958 Feb;5(1):68-91
references: 13519750
references: mBio. 2010 Aug 24;1(3):
references: 20824106
references: J Bacteriol. 1997 Mar;179(5):1482-9
references: 9045803
references: J Mol Biol. 2001 Jan 19;305(3):567-80
references: 11152613
references: Microbiology (Reading). 2009 Oct;155(Pt 10):3260-3269
references: 19556292
references: Science. 1967 Dec 22;158(3808):1536-42
references: 6060356
references: J Mol Biol. 1998 Oct 23;283(2):489-506
references: 9769220
references: J Bacteriol. 2006 Jul;188(14):5124-35
references: 16816184
references: J Bacteriol. 2000 Nov;182(21):6066-74
references: 11029426
references: J Biol Chem. 2011 Sep 16;286(37):32464-74
references: 21778229
references: J Biol Chem. 2011 Dec 23;286(51):44277-44284
references: 22069314
references: Bioinformatics. 2007 Nov 1;23(21):2947-8
references: 17846036
references: Trends Microbiol. 1995 May;3(5):178-85
references: 7542987
references: J Bacteriol. 2005 May;187(9):3002-12
references: 15838026
references: EMBO J. 1986 Nov;5(11):3021-7
references: 16453726
references: Environ Microbiol. 2013 Apr;15(4):1001-15
references: 23016929
references: J Bacteriol. 1962 Dec;84:1321-4
references: 13961373
references: Nat Struct Mol Biol. 2008 Feb;15(2):130-8
references: 18204465
references: J Mol Biol. 1992 May 20;225(2):487-94
references: 1593632
references: Mol Microbiol. 2007 Sep;65(5):1345-59
references: 17697256
references: Nucleic Acids Res. 2009 Jul;37(Web Server issue):W465-8
references: 19429891
references: Nat Protoc. 2010 Apr;5(4):725-38
references: 20360767
references: PLoS Comput Biol. 2008 Nov;4(11):e1000213
references: 18989393
references: J Bacteriol. 1992 Apr;174(7):2178-84
references: 1372599
references: J Bacteriol. 2011 Aug;193(15):3710-21
references: 21642455
references: Gene. 1994 Oct 11;148(1):81-6
references: 7926843
references: J Mol Biol. 2004 May 14;338(5):1027-36
references: 15111065
references: Mol Microbiol. 1999 Oct;34(1):181-94
references: 10540296
references: J Biol Chem. 2011 May 13;286(19):16658-68
references: 21454677
references: J Bacteriol. 1999 Oct;181(20):6564-8
references: 10515954
references: Mol Microbiol. 1998 Jun;28(6):1103-19
references: 9680202
references: J Mol Biol. 1969 Jun 14;42(2):237-46
references: 5803297
references: J Bacteriol. 2010 Jul;192(13):3385-93
references: 20453100
references: Biol Chem. 2013 Apr;394(4):541-59
references: 23241669
references: J Bacteriol. 1979 Dec;140(3):902-10
references: 118160
references: J Biol Chem. 2011 Jun 10;286(23):20600-5
references: 21498511
references: J Med Microbiol. 1987 Jun;23(4):289-302
references: 2438412
references: Protein Eng Des Sel. 2012 Nov;25(11):797-802
references: 23100544
references: Front Microbiol. 2011 Jun 01;2:118
references: 21687428
references: Infect Immun. 1987 May;55(5):1051-7
references: 2437030
references: Mol Microbiol. 1996 Nov;22(3):481-95
references: 8939432
references: Appl Environ Microbiol. 2008 Dec;74(23):7422-6
references: 18849445
references: Acta Microbiol Hung. 1988;35(1):3-24
references: 3134788
references: Mol Microbiol. 1995 May;16(3):565-74
references: 7565115
references: Mol Microbiol. 2001 Mar;39(5):1237-47
references: 11251840
references: Gene. 1991 Oct 30;107(1):43-52
references: 1720755
references: Nucleic Acids Res. 1994 Nov 11;22(22):4673-80
references: 7984417
references: Nucleic Acids Res. 2004 Jul 1;32(Web Server issue):W321-6
references: 15215403
references: J Bacteriol. 1983 Apr;154(1):269-77
references: 6187729
references: Mol Microbiol. 2012 Jun;84(6):1165-76
references: 22554073
references: Innate Immun. 2009 Oct;15(5):261-312
references: 19710102
references: BMC Microbiol. 2013 Feb 17;13:39
references: 23414301