1887

Abstract

is an opportunistic pathogen, which secretes a wide variety of enzymes and toxins into the extracellular medium. Most exoproteins are exported by the type II secretion machinery, the Xcp system, which encompasses 12 different proteins. One of the core components of the Xcp system is the inner-membrane protein XcpS (GspF), homologues of which can be identified in type II secretion machineries as well as in type IV piliation systems. In this study, XcpS was shown to be stabilized by co-expression of the XcpR (GspE) and XcpY (GspL) components of the machinery, demonstrating an interaction between these three proteins. By replacing segments of XcpS with the corresponding parts of its counterpart, XcpS domains were identified that are important for species-specific functioning and thus represent putative interaction domains. The cytoplasmic loop of XcpS was found to be involved in the stabilization by XcpR and XcpY.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.2006/002840-0
2007-05-01
2019-10-23
Loading full text...

Full text loading...

/deliver/fulltext/micro/153/5/1582.html?itemId=/content/journal/micro/10.1099/mic.0.2006/002840-0&mimeType=html&fmt=ahah

References

  1. Abendroth, J., Murphy, P., Sandkvist, M., Bagdasarian, M. & Hol, W. G. ( 2005; ). The X-ray structure of the type II secretion system complex formed by the N-terminal domain of EpsE and the cytoplasmic domain of EpsL of Vibrio cholerae. J Mol Biol 348, 845–855.[CrossRef]
    [Google Scholar]
  2. Ball, G., Chapon-Hervé, V., Bleves, S., Michel, G. & Bally, M. ( 1999; ). Assembly of XcpR in the cytoplasmic membrane is required for extracellular protein secretion in Pseudomonas aeruginosa. J Bacteriol 181, 382–388.
    [Google Scholar]
  3. Ball, G., Durand, E., Lazdunski, A. & Filloux, A. ( 2002; ). A novel type II secretion system in Pseudomonas aeruginosa. Mol Microbiol 43, 475–485.[CrossRef]
    [Google Scholar]
  4. Bayley, D. P. & Jarrell, K. F. ( 1998; ). Further evidence to suggest that archaeal flagella are related to bacterial type IV pili. J Mol Evol 46, 370–373.
    [Google Scholar]
  5. Bitter, W., Koster, M., Latijnhouwers, M., de Cock, H. & Tommassen, J. ( 1998; ). Formation of oligomeric rings by XcpQ and PilQ, which are involved in protein transport across the outer membrane of Pseudomonas aeruginosa. Mol Microbiol 27, 209–219.[CrossRef]
    [Google Scholar]
  6. Bleves, S., Voulhoux, R., Michel, G., Lazdunski, A., Tommassen, J. & Filloux, A. ( 1998; ). The secretion apparatus of Pseudomonas aeruginosa: identification of a fifth pseudopilin, XcpX (GspK family). Mol Microbiol 27, 31–40.[CrossRef]
    [Google Scholar]
  7. Bleves, S., Gérard-Vincent, M., Lazdunski, A. & Filloux, A. ( 1999; ). Structure-function analysis of XcpP, a component involved in general secretory pathway-dependent protein secretion in Pseudomonas aeruginosa. J Bacteriol 181, 4012–4019.
    [Google Scholar]
  8. Braun, P., de Groot, A., Bitter, W. & Tommassen, J. ( 1998; ). Secretion of elastinolytic enzymes and their propeptides by Pseudomonas aeruginosa. J Bacteriol 180, 3467–3469.
    [Google Scholar]
  9. Chung, Y. S., Breidt, F. & Dubnau, D. ( 1998; ). Cell surface localization and processing of the ComG proteins, required for DNA binding during transformation of Bacillus subtilis. Mol Microbiol 29, 905–913.[CrossRef]
    [Google Scholar]
  10. Cianciotto, N. P. ( 2005; ). Type II secretion: a protein secretion system for all seasons. Trends Microbiol 13, 581–588.[CrossRef]
    [Google Scholar]
  11. Crowther, L. J., Yamagata, A., Craig, L., Tainer, J. A. & Donnenberg, M. S. ( 2005; ). The ATPase activity of BfpD is greatly enhanced by zinc and allosteric interactions with other Bfp proteins. J Biol Chem 280, 24839–24848.[CrossRef]
    [Google Scholar]
  12. de Groot, A., Filloux, A. & Tommassen, J. ( 1991; ). Conservation of xcp genes, involved in the two-step protein secretion process, in different Pseudomonas species and other gram-negative bacteria. Mol Gen Genet 229, 278–284.[CrossRef]
    [Google Scholar]
  13. de Groot, A., Krijger, J. J., Filloux, A. & Tommassen, J. ( 1996; ). Characterization of type II protein secretion (xcp) genes in the plant growth-stimulating Pseudomonas putida, strain WCS358. Mol Gen Genet 250, 491–504.
    [Google Scholar]
  14. de Groot, A., Gerritse, G., Tommassen, J., Lazdunski, A. & Filloux, A. ( 1999; ). Molecular organization of the xcp gene cluster in Pseudomonas putida: absence of an xcpX (gspK) homologue. Gene 226, 35–40.[CrossRef]
    [Google Scholar]
  15. Dente, L., Cesareni, G. & Cortese, R. ( 1983; ). pEMBL: a new family of single stranded plasmids. Nucleic Acids Res 11, 1645–1655.[CrossRef]
    [Google Scholar]
  16. Durand, E., Bernadac, A., Ball, G., Lazdunski, A., Sturgis, J. N. & Filloux, A. ( 2003; ). Type II protein secretion in Pseudomonas aeruginosa: the pseudopilus is a multifibrillar and adhesive structure. J Bacteriol 185, 2749–2758.[CrossRef]
    [Google Scholar]
  17. Durand, E., Michel, G., Voulhoux, R., Kurner, J., Bernadac, A. & Filloux, A. ( 2005; ). XcpX controls biogenesis of the Pseudomonas aeruginosa XcpT-containing pseudopilus. J Biol Chem 280, 31378–31389.[CrossRef]
    [Google Scholar]
  18. Enderle, P. J. & Farwell, M. A. ( 1998; ). Electroporation of freshly plated Escherichia coli and Pseudomonas aeruginosa cells. Biotechniques 25, 954–958.
    [Google Scholar]
  19. Figurski, D. H. & Helinski, D. R. ( 1979; ). Replication of an origin-containing derivative of plasmid RK2 dependent on plasmid function provided in trans. Proc Natl Acad Sci U S A 79, 1648–1652.
    [Google Scholar]
  20. Filloux, A. ( 2004; ). The underlying mechanisms of type II protein secretion. Biochim Biophys Acta 1694, 163–179.[CrossRef]
    [Google Scholar]
  21. Filloux, A., Bally, M., Murgier, M., Wretlind, B. & Lazdunski, A. ( 1989; ). Cloning of the xcp genes located at the 55 min region of the chromosome and involved in protein secretion in Pseudomonas aeruginosa. Mol Microbiol 3, 261–265.[CrossRef]
    [Google Scholar]
  22. Filloux, A., Michel, G. & Bally, M. ( 1998; ). GSP-dependent protein secretion in gram-negative bacteria: the Xcp system of Pseudomonas aeruginosa. FEMS Microbiol Rev 22, 177–198.[CrossRef]
    [Google Scholar]
  23. Fürste, J. P., Pansegrau, W., Frank, R., Blöcker, H., Scholz, P., Bagdasarian, M. & Lanka, E. ( 1986; ). Molecular cloning of the plasmid RP4 primase region in a multi-host-range tacP expression vector. Gene 48, 119–131.[CrossRef]
    [Google Scholar]
  24. Grandori, R., Struck, K., Giovanielli, K. & Carey, J. ( 1997; ). A three-step PCR protocol for construction of chimeric proteins. Protein Eng 10, 1099–1100.[CrossRef]
    [Google Scholar]
  25. Gutierrez, C. & Devedjian, J. C. ( 1989; ). A plasmid facilitating in vitro construction of phoA gene fusions in Escherichia coli. Nucleic Acids Res 17, 3999.[CrossRef]
    [Google Scholar]
  26. Haas, D. & Holloway, B. W. ( 1976; ). R factor variants with enhanced sex factor activity in Pseudomonas aeruginosa. Mol Gen Genet 144, 243–251.[CrossRef]
    [Google Scholar]
  27. Hanahan, D. ( 1983; ). Studies on transformation of Escherichia coli with plasmids. J Mol Biol 166, 557–580.[CrossRef]
    [Google Scholar]
  28. He, S. Y., Schoedel, C., Chatterjee, A. K. & Collmer, A. ( 1991; ). Extracellular secretion of pectate lyase by the Erwinia chrysanthemi out pathway is dependent upon Sec-mediated export across the inner membrane. J Bacteriol 173, 4310–4317.
    [Google Scholar]
  29. Hobbs, M. & Mattick, J. S. ( 1993; ). Common components in the assembly of type 4 fimbriae, DNA transfer systems, filamentous phage and protein-secretion apparatus: a general system for the formation of surface-associated protein complexes. Mol Microbiol 10, 233–243.[CrossRef]
    [Google Scholar]
  30. Kaniga, K., Delor, I. & Cornelis, G. R. ( 1991; ). A wide-host-range suicide vector for improving reverse genetics in gram-negative bacteria: inactivation of the blaA gene of Yersinia enterocolitica. Gene 109, 137–141.[CrossRef]
    [Google Scholar]
  31. Kovach, M. E., Elzer, P. H., Hill, D. S., Robertson, G. T., Farris, M. A., Roop, R. M., II & Peterson, K. M. ( 1995; ). Four new derivatives of the broad-host-range cloning vector pBBR1MCS, carrying different antibiotic-resistance cassettes. Gene 166, 175–176.[CrossRef]
    [Google Scholar]
  32. Mayer, M. P. ( 1995; ). A new set of useful cloning and expression vectors derived from pBlueScript. Gene 163, 41–46.[CrossRef]
    [Google Scholar]
  33. Michel, G., Bleves, S., Ball, G., Lazdunski, A. & Filloux, A. ( 1998; ). Mutual stabilization of the XcpZ and XcpY components of the secretory apparatus in Pseudomonas aeruginosa. Microbiology 144, 3379–3386.[CrossRef]
    [Google Scholar]
  34. Naughton, M. A. & Sanger, F. ( 1961; ). Purification and specificity of pancreatic elastase. Biochem J 78, 156–163.
    [Google Scholar]
  35. Nunn, D. N. & Lory, S. ( 1992; ). Components of the protein-excretion apparatus of Pseudomonas aeruginosa are processed by the type IV prepilin peptidase. Proc Natl Acad Sci U S A 89, 47–51.[CrossRef]
    [Google Scholar]
  36. Peabody, C. R., Chung, Y. J., Yen, M. R., Vidal-Ingigliardi, D., Pugsley, A. P. & Saier, M. H., Jr ( 2003; ). Type II protein secretion and its relationship to bacterial type IV pili and archaeal flagella. Microbiology 149, 3051–3072.[CrossRef]
    [Google Scholar]
  37. Possot, O. M., Vignon, G., Bomchil, N., Ebel, F. & Pugsley, A. P. ( 2000; ). Multiple interactions between pullulanase secreton components involved in stabilization and cytoplasmic membrane association of PulE. J Bacteriol 182, 2142–2152.[CrossRef]
    [Google Scholar]
  38. Py, B., Loiseau, L. & Barras, F. ( 1999; ). Assembly of the type II secretion machinery of Erwinia chrysanthemi: direct interaction and associated conformational change between OutE, the putative ATP-binding component and the membrane protein OutL. J Mol Biol 289, 659–670.[CrossRef]
    [Google Scholar]
  39. Py, B., Loiseau, L. & Barras, F. ( 2001; ). An inner membrane platform in the type II secretion machinery of Gram-negative bacteria. EMBO Rep 2, 244–248.[CrossRef]
    [Google Scholar]
  40. Robert, V., Filloux, A. & Michel, G. P. F. ( 2005; ). Subcomplexes from the Xcp secretion system of Pseudomonas aeruginosa. FEMS Microbiol Lett 252, 43–50.[CrossRef]
    [Google Scholar]
  41. Sambrook, J., Fritsch, E. F. & Maniatis, T. ( 1989; ). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  42. Savvides, S. N., Yeo, H. J., Beck, M. R., Blaesing, F., Lurz, R., Lanka, E., Buhrdorf, R., Fischer, W., Haas, R. & Waksman, G. ( 2003; ). VirB11 ATPases are dynamic hexameric assemblies: new insights into bacterial type IV secretion. EMBO J 22, 1969–1980.[CrossRef]
    [Google Scholar]
  43. Shiue, S. J., Kao, K. M., Leu, W. M., Chen, L. Y., Chan, N. L. & Hu, N. T. ( 2006; ). XpsE oligomerization triggered by ATP binding, not hydrolysis, leads to its association with XpsL. EMBO J 25, 1426–1435.[CrossRef]
    [Google Scholar]
  44. Strom, M. S., Nunn, D. N. & Lory, S. ( 1993; ). A single bifunctional enzyme, PilD, catalyzes cleavage and N-methylation of proteins belonging to the type IV pilin family. Proc Natl Acad Sci U S A 90, 2404–2408.[CrossRef]
    [Google Scholar]
  45. Thanassi, D. G. & Hultgren, S. J. ( 2000; ). Multiple pathways allow protein secretion across the bacterial outer membrane. Curr Opin Cell Biol 12, 420–430.[CrossRef]
    [Google Scholar]
  46. Tommassen, J., van Tol, H. & Lugtenberg, B. ( 1983; ). The ultimate localization of an outer membrane protein of Escherichia coli K-12 is not determined by the signal sequence. EMBO J 2, 1275–1279.
    [Google Scholar]
  47. Turner, L. R., Lara, J. C., Nunn, D. N. & Lory, S. ( 1993; ). Mutations in the consensus ATP-binding sites of XcpR and PilB eliminate extracellular protein secretion and pilus biogenesis in Pseudomonas aeruginosa. J Bacteriol 175, 4962–4969.
    [Google Scholar]
  48. Voulhoux, R., Ball, G., Ize, B., Vasil, M. L., Lazdunski, A., Wu, L. F. & Filloux, A. ( 2001; ). Involvement of the twin-arginine translocation system in protein secretion via the type II pathway. EMBO J 20, 6735–6741.[CrossRef]
    [Google Scholar]
  49. Wretlind, B. & Pavlovskis, O. R. ( 1984; ). Genetic mapping and characterization of Pseudomonas aeruginosa mutants defective in the formation of extracellular proteins. J Bacteriol 158, 801–808.
    [Google Scholar]
  50. Yanisch-Perron, C., Vieira, J. & Messing, J. ( 1985; ). Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene 33, 103–119.[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.2006/002840-0
Loading
/content/journal/micro/10.1099/mic.0.2006/002840-0
Loading

Data & Media loading...

Supplements

vol. , part 5, pp. 1582 - 1592

Oligonucleotides used in this study [ PDF] (24 kb)



PDF
This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error