1887

Abstract

Autotransporters of Gram-negative bacteria consist of an N-terminal signal sequence, a C-terminal translocator domain and the secreted passenger domain in between. The autotransporter NalP of includes a protease domain that facilitates the release of several immunogenic proteins from the cell surface into the extracellular milieu. Rather exceptionally among autotransporters, NalP is a lipoprotein. We investigated the role of lipidation in the biogenesis and function of the protein. To this end, the N-terminal cysteine, which is lipidated in the wild-type protein, was substituted by alanine. Like the wild-type protein, the mutant protein was secreted into the medium, demonstrating that lipidation is not required for biogenesis of the protein. However, the non-lipidated NalP variant had a drastically reduced capacity to cleave its substrate proteins from the cell surface, suggesting that the lipid moiety is important for function. Kinetic experiments demonstrated that the autocatalytic processing of the non-lipidated protein at the cell surface was much faster than that of the wild-type protein. Thus, the lipid moiety delays the release of NalP from the cell surface, thereby allowing it to release other surface-exposed proteins into the milieu.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.063982-0
2013-02-01
2024-04-20
Loading full text...

Full text loading...

/deliver/fulltext/micro/159/2/286.html?itemId=/content/journal/micro/10.1099/mic.0.063982-0&mimeType=html&fmt=ahah

References

  1. Ashgar S. S. A., Oldfield N. J., Wooldridge K. G., Jones M. A., Irving G. J., Turner D. P. J., Ala’Aldeen D. A. A. ( 2007). CapA, an autotransporter protein of Campylobacter jejuni, mediates association with human epithelial cells and colonization of the chicken gut. J Bacteriol 189:1856–1865 [View Article][PubMed]
    [Google Scholar]
  2. Bendtsen J. D., Nielsen H., von Heijne G., Brunak S. ( 2004). Improved prediction of signal peptides: SignalP 3.0. J Mol Biol 340:783–795[PubMed] [CrossRef]
    [Google Scholar]
  3. Benz I., Schmidt M. A. ( 2011). Structures and functions of autotransporter proteins in microbial pathogens. Int J Med Microbiol 301:461–468[PubMed] [CrossRef]
    [Google Scholar]
  4. Bos M. P., Tommassen J. ( 2005). Viability of a capsule- and lipopolysaccharide-deficient mutant of Neisseria meningitidis . Infect Immun 73:6194–6197 [View Article][PubMed]
    [Google Scholar]
  5. Claus H., Frosch M., Vogel U. ( 1998). Identification of a hotspot for transformation of Neisseria meningitidis by shuttle mutagenesis using signature-tagged transposons. Mol Gen Genet 259:363–371[PubMed] [CrossRef]
    [Google Scholar]
  6. Coutte L., Antoine R., Drobecq H., Locht C., Jacob-Dubuisson F. ( 2001). Subtilisin-like autotransporter serves as maturation protease in a bacterial secretion pathway. EMBO J 20:5040–5048 [View Article][PubMed]
    [Google Scholar]
  7. Coutte L., Alonso S., Reveneau N., Willery E., Quatannens B., Locht C., Jacob-Dubuisson F. ( 2003a). Role of adhesin release for mucosal colonization by a bacterial pathogen. J Exp Med 197:735–742 [View Article][PubMed]
    [Google Scholar]
  8. Coutte L., Willery E., Antoine R., Drobecq H., Locht C., Jacob-Dubuisson F. ( 2003b). Surface anchoring of bacterial subtilisin important for maturation function. Mol Microbiol 49:529–539 [View Article][PubMed]
    [Google Scholar]
  9. Dautin N., Bernstein H. D. ( 2007). Protein secretion in gram-negative bacteria via the autotransporter pathway. Annu Rev Microbiol 61:89–112 [View Article][PubMed]
    [Google Scholar]
  10. Driessen A. J. M., Nouwen N. ( 2008). Protein translocation across the bacterial cytoplasmic membrane. Annu Rev Biochem 77:643–667 [View Article][PubMed]
    [Google Scholar]
  11. Henderson I. R., Navarro-Garcia F., Nataro J. P. ( 1998). The great escape: structure and function of the autotransporter proteins. Trends Microbiol 6:370–378[PubMed] [CrossRef]
    [Google Scholar]
  12. Johnston D. M., Cannon J. G. ( 1999). Construction of mutant strains of Neisseria gonorrhoeae lacking new antibiotic resistance markers using a two gene cassette with positive and negative selection. Gene 236:179–184 [View Article][PubMed]
    [Google Scholar]
  13. Jong W. S. P., Luirink J. ( 2008). The conserved extension of the Hbp autotransporter signal peptide does not determine targeting pathway specificity. Biochem Biophys Res Commun 368:522–527 [View Article][PubMed]
    [Google Scholar]
  14. Leyton D. L., de Luna M., Sevastsyanovich Y. R., Tveen Jensen K., Browning D. F., Scott-Tucker A., Henderson I. R. ( 2010). The unusual extended signal peptide region is not required for secretion and function of an Escherichia coli autotransporter. FEMS Microbiol Lett 311:133–139 [View Article][PubMed]
    [Google Scholar]
  15. Odenbreit S., Till M., Hofreuter D., Faller G., Haas R. ( 1999). Genetic and functional characterization of the alpAB gene locus essential for the adhesion of Helicobacter pylori to human gastric tissue. Mol Microbiol 31:1537–1548[PubMed] [CrossRef]
    [Google Scholar]
  16. Okuda S., Tokuda H. ( 2011). Lipoprotein sorting in bacteria. Annu Rev Microbiol 65:239–259 [View Article][PubMed]
    [Google Scholar]
  17. Oomen C. J., van Ulsen P., van Gelder P., Feijen M., Tommassen J., Gros P. ( 2004). Structure of the translocator domain of a bacterial autotransporter. EMBO J 23:1257–1266[PubMed] [CrossRef]
    [Google Scholar]
  18. Pettersson A., Prinz T., Umar A., van der Biezen J., Tommassen J. ( 1998). Molecular characterization of LbpB, the second lactoferrin-binding protein of Neisseria meningitidis . Mol Microbiol 27:599–610[PubMed] [CrossRef]
    [Google Scholar]
  19. Pettersson A., Kortekaas J., Weynants V. E., Voet P., Poolman J. T., Bos M. P., Tommassen J. ( 2006). Vaccine potential of the Neisseria meningitidis lactoferrin-binding proteins LbpA and LbpB. Vaccine 24:3545–3557 [View Article][PubMed]
    [Google Scholar]
  20. Roussel-Jazédé V., Jongerius I., Bos M. P., Tommassen J., van Ulsen P. ( 2010). NalP-mediated proteolytic release of lactoferrin-binding protein B from the meningococcal cell surface. Infect Immun 78:3083–3089 [View Article][PubMed]
    [Google Scholar]
  21. Serruto D., Spadafina T., Ciucchi L., Lewis L. A., Ram S., Tontini M., Santini L., Biolchi A., Seib K. L. & other authors ( 2010). Neisseria meningitidis GNA2132, a heparin-binding protein that induces protective immunity in humans. Proc Natl Acad Sci U S A 107:3770–3775 [View Article][PubMed]
    [Google Scholar]
  22. Stork M., Bos M. P., Jongerius I., de Kok N., Schilders I., Weynants V. E., Poolman J. T., Tommassen J. ( 2010). An outer membrane receptor of Neisseria meningitidis involved in zinc acquisition with vaccine potential. PLoS Pathog 6:e1000969 [View Article][PubMed]
    [Google Scholar]
  23. Szabady R. L., Peterson J. H., Skillman K. M., Bernstein H. D. ( 2005). An unusual signal peptide facilitates late steps in the biogenesis of a bacterial autotransporter. Proc Natl Acad Sci U S A 102:221–226[PubMed] [CrossRef]
    [Google Scholar]
  24. van Dam V., Bos M. P. ( 2012). Generating knock-out and complementation strains of Neisseria meningitidis . Methods Mol Biol 799:55–72 [View Article][PubMed]
    [Google Scholar]
  25. van Ulsen P., van Alphen L., Hopman C. T. P., van der Ende A., Tommassen J. ( 2001). In vivo expression of Neisseria meningitidis proteins homologous to the Haemophilus influenzae Hap and Hia autotransporters. FEMS Immunol Med Microbiol 32:53–64[PubMed] [CrossRef]
    [Google Scholar]
  26. van Ulsen P., van Alphen L., ten Hove J., Fransen F., van der Ley P., Tommassen J. ( 2003). A neisserial autotransporter NalP modulating the processing of other autotransporters. Mol Microbiol 50:1017–1030[PubMed] [CrossRef]
    [Google Scholar]
  27. van Ulsen P., Adler B., Fassler P., Gilbert M., van Schilfgaarde M., van der Ley P., van Alphen L., Tommassen J. ( 2006). A novel phase-variable autotransporter serine protease, AusI, of Neisseria meningitidis . Microbes Infect 8:2088–2097[PubMed] [CrossRef]
    [Google Scholar]
  28. Vidarsson G., Overbeeke N., Stemerding A. M., van den Dobbelsteen G., van Ulsen P., van der Ley P., Kilian M., van de Winkel J. G. J. ( 2005). Working mechanism of immunoglobulin A1 (IgA1) protease: cleavage of IgA1 antibody to neisseria meningitidis PorA requires de novo synthesis of IgA1 protease. Infect Immun 73:6721–6726 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.063982-0
Loading
/content/journal/micro/10.1099/mic.0.063982-0
Loading

Data & Media loading...

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