title: Structure-function analysis reveals that the Pseudomonas aeruginosa Tps4 two-partner secretion system is involved in CupB5 translocation.
abstract: Pseudomonas aeruginosa is a Gram-negative opportunistic bacterium, synonymous with cystic fibrosis patients, which can cause chronic infection of the lungs. This pathogen is a model organism to study biofilms: a bacterial population embedded in an extracellular matrix that provide protection from environmental pressures and lead to persistence. A number of Chaperone-Usher Pathways, namely CupA-CupE, play key roles in these processes by assembling adhesive pili on the bacterial surface. One of these, encoded by the cupB operon, is unique as it contains a nonchaperone-usher gene product, CupB5. Two-partner secretion (TPS) systems are comprised of a C-terminal integral membrane β-barrel pore with tandem N-terminal POTRA (POlypeptide TRansport Associated) domains located in the periplasm (TpsB) and a secreted substrate (TpsA). Using NMR we show that TpsB4 (LepB) interacts with CupB5 and its predicted cognate partner TpsA4 (LepA), an extracellular protease. Moreover, using cellular studies we confirm that TpsB4 can translocate CupB5 across the P. aeruginosa outer membrane, which contrasts a previous observation that suggested the CupB3 P-usher secretes CupB5. In support of our findings we also demonstrate that tps4/cupB operons are coregulated by the RocS1 sensor suggesting P. aeruginosa has developed synergy between these systems. Furthermore, we have determined the solution-structure of the TpsB4-POTRA1 domain and together with restraints from NMR chemical shift mapping and in vivo mutational analysis we have calculated models for the entire TpsB4 periplasmic region in complex with both TpsA4 and CupB5 secretion motifs. The data highlight specific residues for TpsA4/CupB5 recognition by TpsB4 in the periplasm and suggest distinct roles for each POTRA domain.
authors: Department of Life Sciences, Centre for Structural Biology, Imperial College London, South Kensington Campus, London, SW7 2AZ, United Kingdom; Department of Life Sciences, MRC Centre for Molecular Bacteriology and Infection, Imperial College London, South Kensington Campus, London, SW7 2AZ, United Kingdom.
authors: James A
authors: JA
authors: Garnett
authors: Daniela
authors: D
authors: Muhl
authors: Christopher H
authors: CH
authors: Douse
authors: Kailyn
authors: K
authors: Hui
authors: Andreas
authors: A
authors: Busch
authors: Ayodele
authors: A
authors: Omisore
authors: Yi
authors: Y
authors: Yang
authors: Peter
authors: P
authors: Simpson
authors: Jan
authors: J
authors: Marchant
authors: Gabriel
authors: G
authors: Waksman
authors: Steve
authors: S
authors: Matthews
authors: Alain
authors: A
authors: Filloux
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D018832:Molecular Chaperones / Q000737:chemistry* / Q000235:genetics; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000737:chemistry* / Q000235:genetics; D013329:Structure-Activity Relationship; D000069320:Type V Secretion Systems / Q000737:chemistry
chemical_list: D001426:Bacterial Proteins; D018832:Molecular Chaperones; D000069320:Type V Secretion Systems
keywords: NMR; POTRA; Pseudomonas aeruginosa; biofilm; chaperone-usher; two-partner secretion; virulence factor
references: Curr Protein Pept Sci. 2012 Dec;13(8):739-55
references: 23305361
references: Biochim Biophys Acta. 2002 May 3;1562(1-2):6-31
references: 11988218
references: J Biomol NMR. 1999 Mar;13(3):289-302
references: 10212987
references: Curr Opin Struct Biol. 2012 Aug;22(4):500-6
references: 22402496
references: J Biol Chem. 2010 Jun 4;285(23):18016-24
references: 20348103
references: Biomol NMR Assign. 2008 Dec;2(2):119-21
references: 19636884
references: Structure. 2009 Sep 9;17(9):1187-94
references: 19748339
references: PLoS One. 2009;4(6):e6018
references: 19547710
references: Microbiol Mol Biol Rev. 2007 Dec;71(4):551-75
references: 18063717
references: PLoS One. 2013;8(3):e58769
references: 23516555
references: Science. 2007 Aug 17;317(5840):957-61
references: 17702945
references: Med Sci (Paris). 2003 Jan;19(1):77-83
references: 12836195
references: Structure. 2008 Dec 10;16(12):1873-81
references: 19081063
references: Infect Immun. 2005 Aug;73(8):4960-71
references: 16041011
references: Mol Microbiol. 1998 Feb;27(3):617-30
references: 9489673
references: Mol Microbiol. 2006 Jul;61(2):368-82
references: 16771844
references: J Biol Chem. 2012 Jan 20;287(4):2591-9
references: 22134917
references: Proc Natl Acad Sci U S A. 2001 Jun 5;98(12):6911-6
references: 11381121
references: Infect Immun. 2011 Jan;79(1):211-9
references: 21041488
references: Biology (Basel). 2013 Jun 26;2(3):841-60
references: 24833049
references: EMBO J. 2008 Oct 22;27(20):2669-80
references: 18833195
references: Res Microbiol. 2013 Jul-Aug;164(6):583-95
references: 23542425
references: Cell Microbiol. 2008 Jul;10(7):1491-504
references: 18331590
references: Nature. 2013 Apr 11;496(7444):243-6
references: 23579681
references: Environ Microbiol. 2011 Jul;13(7):1666-81
references: 21554516
references: Science. 2007 Aug 17;317(5840):961-4
references: 17702946
references: J Mol Biol. 2011 Mar 11;406(5):659-66
references: 21236263
references: J Biol Chem. 2005 Apr 29;280(17):17339-45
references: 15728184
references: EMBO J. 2011 Sep 14;30(18):3864-74
references: 21841773
references: Mol Microbiol. 2001 Apr;40(2):306-13
references: 11309114
references: J Bacteriol. 2004 May;186(9):2880-90
references: 15090530
references: Mol Microbiol. 2013 Aug;89(3):450-63
references: 23750818
references: J Biomol NMR. 1994 Sep;4(5):603-14
references: 22911360
references: Mol Microbiol. 2005 Jan;55(2):368-80
references: 15659157
references: Nat Protoc. 2010 Apr;5(4):725-38
references: 20360767
references: Mol Microbiol. 1994 Jan;11(2):337-47
references: 8170396
references: Mol Microbiol. 2010 Feb;75(4):827-42
references: 20088866
references: Gene. 1991 Dec 20;109(1):137-41
references: 1756974
references: J Bacteriol. 2007 May;189(9):3547-55
references: 17293418
references: Mol Microbiol. 2014 Aug;93(3):415-25
references: 24913916
references: Mol Microbiol. 2014 Jun;92(6):1164-76
references: 24646315
references: Infect Immun. 2008 Oct;76(10):4649-58
references: 18678657
references: Philos Trans R Soc Lond B Biol Sci. 2012 Apr 19;367(1592):1088-101
references: 22411980
references: Mol Microbiol. 1998 Aug;29(3):763-74
references: 9723916
references: J Bacteriol. 1997 Feb;179(3):775-83
references: 9006033
references: J Biol Chem. 2014 Jul 11;289(28):19799-809
references: 24872418
references: Mol Microbiol. 2011 Mar;79(5):1353-66
references: 21205015
references: Nature. 2009 Jan 29;457(7229):594-8
references: 19060885
references: J Biol Chem. 1991 Jul 25;266(21):13640-5
references: 1856199
references: Mol Biosyst. 2010 Mar;6(3):494-8
references: 20174678
references: FEMS Microbiol Rev. 2010 May;34(3):317-78
references: 20070375
references: J Bacteriol. 2013 Feb;195(4):788-97
references: 23222722
references: Environ Microbiol. 2011 Mar;13(3):666-83
references: 21091863
references: FEBS J. 2010 Nov;277(22):4755-65
references: 20955520
references: Science. 2003 Jan 10;299(5604):262-5
references: 12522254
references: Bioinformatics. 2007 Feb 1;23(3):381-2
references: 17121777
references: Mol Microbiol. 2011 Jul;81(1):99-112
references: 21542859
references: Mol Microbiol. 2008 Jun;68(5):1216-27
references: 18430136
references: Bioinformatics. 2007 Nov 1;23(21):2947-8
references: 17846036
references: PLoS One. 2011;6(1):e16583
references: 21304995
references: J Am Chem Soc. 2003 Feb 19;125(7):1731-7
references: 12580598
references: Infect Immun. 2003 May;71(5):2591-7
references: 12704133