Dissecting the Cell Surface

  • Robert Fagan
  • Neil Fairweather
Part of the Methods in Molecular Biology book series (MIMB, volume 646)


The bacterial cell surface is an important structure as it mediates interactions with the external environment. In the case of pathogens like Clostridium difficile, the cell wall and its components also have to mediate interactions with the host cells and their products. In this chapter we discuss the various methods used for dissecting the cell surface and the biochemical and immunological procedures that are commonly used to analyse the properties of the proteins within the cell wall. A major consideration is the S-layer which in C. difficile shows considerable variation in sequence and between strains, a property which is also reflected in its antigenic properties.

Key words

Cell wall S-layer protein purification ELISA immunofluorescence 



We thank Anne Wright, Omar Qazi and Natalie Welsh for developing and optimising some of the procedures described in this chapter.


  1. 1.
    Navarre WW and 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.PubMedGoogle Scholar
  2. 2.
    Calabi E, Ward S, Wren B, et al. (2001) Molecular characterization of the surface layer proteins from Clostridium difficile. Mol Microbiol 40, 1187–1199.PubMedCrossRefGoogle Scholar
  3. 3.
    Qazi O, Brailsford A, Wright A, Faraar J, Campbell J and Fairweather N. (2007) Identification and characterization of the surface-layer protein of Clostridium tetani. FEMS Microbiol Lett 274, 126–131.PubMedCrossRefGoogle Scholar
  4. 4.
    Calabi E, Calabi F, Phillips AD and Fairweather N. (2002) Binding of Clostridium difficile surface layer proteins to gastrointestinal tissues. Infect Immun 70, 5770–5778.PubMedCrossRefGoogle Scholar
  5. 5.
    Sebaihia M, Peck MW, Minton NP, et al. (2007) Genome sequence of a proteolytic (Group I) Clostridium botulinum strain Hall A and comparative analysis of the clostridial genomes. Genome Res 17, 1082–1092.PubMedCrossRefGoogle Scholar
  6. 6.
    Sebaihia M, Wren BW, Mullany P, et al. (2006) The multidrug-resistant human pathogen Clostridium difficile has a highly mobile, mosaic genome. Nat Genet 38, 779–786.PubMedCrossRefGoogle Scholar
  7. 7.
    Bruggemann H, Baumer S, Fricke WF, et al. (2003) The genome sequence of Clostridium tetani, the causative agent of tetanus disease. PNAS 100, 1316–1321.PubMedCrossRefGoogle Scholar
  8. 8.
    Bruggemann H and Gottschalk G. (2008) Comparative genomics of Clostridia: link between the ecological niche and cell surface properties. Ann NY Acad Sci 1125, 73–81.PubMedCrossRefGoogle Scholar
  9. 9.
    Karjalainen T, Waligora-Dupriet A-J, Cerquetti M, et al. (2001) Molecular and genomic analysis of genes encoding surface-anchored proteins from Clostridium difficile. Infect Immun 69, 3442–3446.PubMedCrossRefGoogle Scholar
  10. 10.
    Emerson J and Fairweather NF. (2009) Surface structures of C. difficile and other clostridia. In: Bruggemann H, Gottschalk G, eds., Clostridia – Molecular Biology in the Post-genomic Era, Caister Academic Press, Norfolk.Google Scholar
  11. 11.
    Herbold DR and Glaser L. (1975) Interaction of N-acetylmuramic acid L-alanine amidase with cell wall polymers. J Biol Chem 250, 7231–7238.PubMedGoogle Scholar
  12. 12.
    McCoubrey J and Poxton IR. (2001) Variation in the surface layer proteins of Clostridium difficile. FEMS Immunol Med Microbiol 31, 131–135.PubMedCrossRefGoogle Scholar
  13. 13.
    Masuda K, Itoh M and Kawata T. (1989) Characterization and reassembly of a regular array in the cell wall of Clostridium difficile GA I4131. Microbiol Immunol 33, 287–298.PubMedGoogle Scholar
  14. 14.
    Wright A. Proteomic and bioinformatic analysis of surface proteins from Clostridium difficile, PhD thesis. University of London, 2006.Google Scholar
  15. 15.
    Schneewind O, Model P and Fischetti VA. (1992) Sorting of protein A to the staphylococcal cell wall. Cell 70, 267–281.PubMedCrossRefGoogle Scholar
  16. 16.
    Jonquieres R, Bierne H, Fiedler F, Gounon P and 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.PubMedCrossRefGoogle Scholar
  17. 17.
    Laemmli UK. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685.PubMedCrossRefGoogle Scholar
  18. 18.
    Wittig I, Braun HP and Schagger H. (2006) Blue native PAGE. Nat Prot 1, 418–428.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Robert Fagan
    • 1
  • Neil Fairweather
    • 1
  1. 1.Centre for Molecular Microbiology and Infection, Imperial College LondonLondonUK

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