1887

Microbiology Society logo

Volume 149, Issue 10

Identification and characterization of a fibronectin-binding protein from Free

Abstract

A 68 kDa fibronectin-binding protein (Fbp68) from displaying significant homology to several established or putative Fbps from other bacteria was identified. The one-copy gene is highly conserved in isolates. Fbp68 was expressed in in fusion with glutathione -transferase; the fusion protein and the native Fbp68 were purified. Immunoblot analysis and cell fractionation experiments revealed that Fbp68 is present on the surface of the bacteria. Far-immuno dot-blotting demonstrated that Fbp68 was capable of fixing fibronectin. Indirect immunofluorescence and ELISA were employed to demonstrate that could bind both soluble and immobilized fibronectin. With competitive adherence inhibition assays it was shown that antibodies raised against Fbp68 partially inhibited attachment of to fibronectin and Vero cells. Furthermore, Vero cells could fix purified membrane-immobilized Fbp68. Thus Fbp68 appears to be one of the several adhesins identified to date in .

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): BCIP, 5-bromo-4-chloro-3-indolyl phosphate , Fbp, fibronectin-binding protein and NBT, nitro blue tetrazolium
SGM
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.26145-0
2003-10-01
2024-04-25
Loading full text...

Full text loading...

/deliver/fulltext/micro/149/10/mic1492779.html?itemId=/content/journal/micro/10.1099/mic.0.26145-0&mimeType=html&fmt=ahah

References

  1. Bartlett J. G., Chang T. W., Gurwith M., Gorbach S. L., Onderdonk A. B. 1978; Antibiotic-associated pseudomembranous colitis due to toxin-producing clostridia. N Engl J Med 298:531–534
     [Google Scholar]
  2. Calabi E., Ward S., Wren B., Paxton T., Panico M., Morris H., Dell A., Dougan G., Fairweather N. 2001; Molecular characterization of the surface layer proteins from Clostridium difficile . Mol Microbiol 40:1187–1199
     [Google Scholar]
  3. Calabi E., Calabi F., Phillips A. D., Fairweather N. 2002; Binding of Clostridium difficile surface layer proteins to gastrointestinal tissues. Infect Immun 70:5770–5778
     [Google Scholar]
  4. Cerquetti M., Molinari A., Sebastianelli A., Diociaiuti M., Petruzzelli R., Capo C., Mastrantonio P. 2000; Characterization of surface layer proteins from different Clostridium difficile clinical isolates. Microb Pathog 28:363–372
     [Google Scholar]
  5. Cerquetti M., Serafino A., Sebastianelli A., Mastrantonio O. 2002; Binding of Clostridium difficile to Caco-2 epithelial cell line and to extracellular matrix proteins. FEMS Immunol Med Microbiol 32:211–218
     [Google Scholar]
  6. Chia J. S., Yeh C. Y., Chen J. Y. 2000; Identification of a fibronectin binding protein from Streptococcus mutans . Infect Immun 68:1864–1870
     [Google Scholar]
  7. Corpet F., Servant F., Gouzy J., Kahn D. 2000; ProDom and ProDom-CG: tools for protein domain analysis and whole genome comparisons. Nucleic Acids Res 28:267–269
     [Google Scholar]
  8. Courtney H. S., Li Y., Dale J. B., Hasty D. L. 1994; Cloning, sequencing, and expression of a fibronectin/fibrinogen-binding protein from group A streptococci. Infect Immun 62:3937–3946
     [Google Scholar]
  9. Davies H. A., Borriello S. P. 1990; Detection of capsule in strains of Clostridium difficile of varying virulence and toxigenicity. Microb Pathog 9:141–146
     [Google Scholar]
  10. dos Santos A. R. Jr, Wada M. L. 1999; Foetal calf serum and dexamethasone effects on Vero cell growth and differentiation. Cytobios 99:159–171
     [Google Scholar]
  11. Drudy D., O'Donoghue D. P., Baird A., Fenelon L., O'Farrelly C. 2001; Flow cytometric analysis of Clostridium difficile adherence to human intestinal epithelial cells. J Med Microbiol 50:526–534
     [Google Scholar]
  12. Fischetti V. A., Pancholi V., Schneewind O. 1990; Conservation of a hexapeptide sequence in the anchor region of surface proteins from gram-positive cocci. Mol Microbiol 4:1603–1605
     [Google Scholar]
  13. Gibson C. M., Caparon M. G. 2002; Alkaline phosphatase reporter transposon for identification of genes encoding secreted proteins in gram-positive microorganisms. Appl Environ Microbiol 68:928–932
     [Google Scholar]
  14. Hennequin C., Collignon A., Karjalainen T. 2001a; Analysis of expression of GroEL (Hsp60) of Clostridium difficile in response to stress. Microb Pathog 31:255–260
     [Google Scholar]
  15. Hennequin C., Porcheray F., Waligora-Dupriet A., Collignon A., Barc M. C., Bourlioux P., Karjalainen T. 2001b; GroEL (Hsp60) of Clostridium difficile is involved in cell adherence. Microbiology 147:87–96
     [Google Scholar]
  16. Holmes A. R., McNab R., Millsap K. W., Rohde M., Hammerschmidt S., Mawdsley J. L., Jenkinson H. F. 2001; The pavA gene of Streptococcus pneumoniae encodes a fibronectin-binding protein that is essential for virulence. Mol Microbiol 41:1395–1408
     [Google Scholar]
  17. Jonquières R, Bierne H., Fiedler F., Gounon P., Cossart P. 1999; Interaction between the protein InlB of Listeria monocytogenes and lipoteichoic acid: a novel mechanism of protein association at the surface of Gram-positive bacteria. Mol Microbiol 34:902–914
     [Google Scholar]
  18. Karjalainen T., Barc M. C., Collignon A., Trolle S., Boureau H., Cotte-Laffitte J., Bourlioux P. 1994; Cloning of a genetic determinant from Clostridium difficile involved in adherence to tissue culture cells and mucus. Infect Immun 62:4347–4355
     [Google Scholar]
  19. Karjalainen T., Waligora-Dupriet A. J., Cerquetti M., Spigaglia P., Maggioni A., Mauri P., Mastrantonio P. 2001; Molecular and genomic analysis of genes encoding surface-anchored proteins from Clostridium difficile . Infect Immun 69:3442–3446
     [Google Scholar]
  20. Karjalainen T., Saumier N., Barc M. C., Delmée M., Collignon A. 2002; Clostridium difficile genotyping based on slpA variable region in S-layer gene sequence: an alternative to serotyping. J Clin Microbiol 40:2452–2458
     [Google Scholar]
  21. Lupas A., Van Dyke M., Stock J. 1991; Predicting coiled coils from protein sequences. Science 252:1162–1164
     [Google Scholar]
  22. Manganelli R., van de Rijn I. 1999; Characterization of emb , a gene encoding the major adhesin of Streptococcus defectivus . Infect Immun 67:50–56
     [Google Scholar]
  23. Meehan M., Muldowney D. A., Watkins N. J., Owen P. 2000; Localization and characterization of the ligand-binding domain of the fibrinogen-binding protein (FgBP) of Streptococcus equi subsp. equi . Microbiology 146:1187–1194
     [Google Scholar]
  24. Navarre W. W., Schneewind O. 1999; Surface proteins of gram-positive bacteria and mechanisms of their targeting to the cell wall envelope. Microbiol Mol Biol Rev 63:174–229
     [Google Scholar]
  25. Ozeri V., Rosenshine I., Mosher D. F., Fassler R., Hanski E. 1998; Roles of integrins and fibronectin in the entry of Streptococcus pyogenes into cells via protein F1. Mol Microbiol 30:625–637
     [Google Scholar]
  26. Poilane I., Karjalainen T., Barc M. C., Bourlioux P., Collignon A. 1998; Protease activity of Clostridium difficile strains. Can J Microbiol 44:157–161
     [Google Scholar]
  27. Sakurai T., Hajiro K., Takakuwa H., Nishi A., Aihara M., Chiba T. 2001; Liver abscess caused by Clostridium difficile . Scand J Infect Dis 33:69–70
     [Google Scholar]
  28. Seddon S. V., Borriello S. P. 1992; Proteolytic activity of Clostridium difficile . J Med Microbiol 36:307–311
     [Google Scholar]
  29. Tasteyre A., Barc M. C., Karjalainen T., Dodson P., Hyde S., Bourlioux P., Borriello P. 2000a; A Clostridium difficile gene encoding flagellin. Microbiology 146:957–966
     [Google Scholar]
  30. Tasteyre A., Karjalainen T., Avesani V., Delmee M., Collignon A., Bourlioux P., Barc M. C. 2000b; Phenotypic and genotypic diversity of the flagellin gene ( fliC ) among Clostridium difficile isolates from different serogroups. J Clin Microbiol 38:3179–3186
     [Google Scholar]
  31. Tasteyre A., Karjalainen T., Avesani V., Delmee M., Collignon A., Bourlioux P., Barc M. C. 2001a; Molecular characterization of fliD gene encoding flagellar cap and its expression among Clostridium difficile isolates from different serogroups. J Clin Microbiol 39:1178–1183
     [Google Scholar]
  32. Tasteyre A., Barc M. C., Collignon A., Boureau H., Karjalainen T. 2001b; Role of FliC and FliD flagellar proteins of Clostridium difficile in adherence and gut colonization. Infect Immun 69:7937–7940
     [Google Scholar]
  33. Thompson J. D., Higgins D. G., Gibson T. J. 1994; CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680
     [Google Scholar]
  34. van der Flier M., Chhun N., Wizemann T. M., Min J., McCarthy J. B., Tuomanen E. I. 1995; Adherence of Streptococcus pneumoniae to immobilized fibronectin. Infect Immun 63:4317–4322
     [Google Scholar]
  35. Vercellotti G. M., McCarthy J. B., Lindholm P., Peterson P. K., Jacob H. S., Furcht L. T. 1985; Extracellular matrix proteins (fibronectin, laminin, and type IV collagen) bind and aggregate bacteria. Am J Pathol 120:13–21
     [Google Scholar]
  36. von Heijne G. 1985; Structural and thermodynamic aspects of the transfer of proteins into and across membranes. Curr Top Membr Transp 242:151–179
     [Google Scholar]
  37. Waligora A. J., Hennequin C., Mullany P., Bourlioux P., Collignon A., Karjalainen T. 2001; Characterization of a cell surface protein of Clostridium difficile with adhesive properties. Infect Immun 69:2144–2153
     [Google Scholar]
  38. Whatmore A. M. 2001; Streptococcus pyogenes sclB encodes a putative hypervariable surface protein with a collagen-like repetitive structure. Microbiology 147:419–429
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.26145-0
Loading
Identification and characterization of a fibronectin-binding protein from Clostridium difficile
Microbiology 149, 2779 (2003); https://doi.org/10.1099/mic.0.26145-0
/content/journal/micro/10.1099/mic.0.26145-0
/content/journal/micro/10.1099/mic.0.26145-0
Loading

Data & Media loading...

Most cited Most Cited RSS feed

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