Skip to main content

Methods for Functional Characterization of the Type IX Secretion System of Porphyromonas gingivalis

  • Protocol
  • First Online:
Periodontal Pathogens

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2210))

  • 1121 Accesses

Abstract

The type IX secretion system (T9SS) is a protein secretion system for gingipain proteases and is found on the cell surface of Porphyromonas gingivalis. Proteins secreted by T9SS contain a signal peptide, functional domains, an immunoglobulin (Ig)-like domain, and a C-terminal domain (CTD). Thirty genes on the P. gingivalis chromosome encode proteins that possess the CTD, which is important for T9SS-mediated translocation to the cell surface across the outer membrane. In T9SS mutant strains, proteins accumulate as precursors in the cell and therefore exhibit a phenotype similar to that of secreted protein-deficient mutants. Black pigment productivity and hemagglutination are phenotypic features of P. gingivalis associated with the activity of gingipains. In P. gingivalis T9SS mutants, unprocessed gingipains with high molecular weights accumulate in the cell, and colony pigmentation and hemagglutination are not observed in the same phenotype as a gingipain null mutant.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Sato K, Yukitake H, Narita Y et al (2013) Identification of Porphyromonas gingivalis proteins secreted by the Por secretion system. FEMS Microbiol Lett 338:68–76

    Article  CAS  Google Scholar 

  2. Kondo Y, Ohara N, Sato K et al (2010) Tetratricopeptide repeat protein-associated proteins contribute to the virulence of Porphyromonas gingivalis. Infect Immun 78:2846–2856

    Article  CAS  Google Scholar 

  3. Shoji M, Sato K, Yukitake H et al (2011) Por secretion system-dependent secretion and glycosylation of Porphyromonas gingivalis hemin-binding protein 35. PLoS One 6:e21372

    Article  CAS  Google Scholar 

  4. Nonaka M, Shoji M, Kadowaki T et al (2014) Analysis of a Lys-specific serine endopeptidase secreted via the type IX secretion system in Porphyromonas gingivalis. FEMS Microbiol Lett 354:60–68

    Article  CAS  Google Scholar 

  5. Veith PD, Nor Muhammad NA, Dashper SG et al (2013) Protein substrates of a novel secretion system are numerous in the Bacteroidetes phylum and have in common a cleavable C-terminal secretion signal, extensive post-translational modification, and cell-surface attachment. J Proteome Res 12:4449–4461

    Article  CAS  Google Scholar 

  6. Seers CA, Slakeski N, Veith PD et al (2006) The RgpB C-terminal domain has a role in attachment of RgpB to the outer membrane and belongs to a novel C-terminal-domain family found in Porphyromonas gingivalis. J Bacteriol 188:6376–6386

    Article  CAS  Google Scholar 

  7. Narita Y, Sato K, Yukitake H et al (2014) Lack of a surface layer in Tannerella forsythia mutants deficient in the type IX secretion system. Microbiology 160:2295–2303

    Article  CAS  Google Scholar 

  8. Kondo Y, Sato K, Nagano K et al (2018) Involvement of PorK, a component of the type IX secretion system, in Prevotella melaninogenica pathogenicity. Microbiol Immunol 62:554–566

    Article  CAS  Google Scholar 

  9. McBride MJ, Zhu Y (2013) Gliding motility and Por secretion system genes are widespread among members of the phylum Bacteroidetes. J Bacteriol 195:270–278

    Article  CAS  Google Scholar 

  10. Nakane D, Sato K, Wada H et al (2013) Helical flow of surface protein required for bacterial gliding motility. Proc Natl Acad Sci U S A 110:11145–11150

    Article  CAS  Google Scholar 

  11. Sato K, Sakai E, Veith PD et al (2005) Identification of a new membrane-associated protein that influences transport/maturation of gingipains and adhesins of Porphyromonas gingivalis. J Biol Chem 280:8668–8677

    Article  CAS  Google Scholar 

  12. Sato K, Naito M, Yukitake H et al (2010) A protein secretion system linked to bacteroidete gliding motility and pathogenesis. Proc Natl Acad Sci U S A 107:276–281

    Article  CAS  Google Scholar 

  13. Saiki K, Konishi K (2007) Identification of a Porphyromonas gingivalis novel protein sov required for the secretion of gingipains. Microbiol Immunol 51:483–491

    Article  CAS  Google Scholar 

  14. Lasica AM, Ksiazek M, Madej M et al (2017) The type IX secretion system (T9SS): highlights and recent insights into its structure and function. Front Cell Infect Microbiol 7:215

    Article  Google Scholar 

  15. Veith PD, Glew MD, Gorasia DG et al (2017) Type IX secretion: the generation of bacterial cell surface coatings involved in virulence, gliding motility and the degradation of complex biopolymers. Mol Microbiol 106:35–53

    Article  CAS  Google Scholar 

  16. Kadowaki T, Yukitake H, Naito M et al (2016) A two-component system regulates gene expression of the type IX secretion component proteins via an ECF sigma factor. Sci Rep 6:23288

    Article  CAS  Google Scholar 

  17. Shah HN, Gharbia SE (1989) Lysis of erythrocytes by the secreted cysteine proteinase of Porphyromonas gingivalis W83. FEMS Microbiol Lett 52:213–217

    Article  CAS  Google Scholar 

  18. Smalley JW, Silver J, Marsh PJ et al (1998) The periodontopathogen Porphyromonas gingivalis binds iron protoporphyrin IX in the mu-oxo dimeric form: an oxidative buffer and possible pathogenic mechanism. Biochem J 331(Pt 3):681–685

    Article  CAS  Google Scholar 

  19. Shi Y, Ratnayake DB, Okamoto K et al (1999) Genetic analyses of proteolysis, hemoglobin binding, and hemagglutination of Porphyromonas gingivalis. Construction of mutants with a combination of rgpA, rgpB, kgp, and hagA. J Biol Chem 274:17955–17960

    Article  CAS  Google Scholar 

  20. Okamoto K, Nakayama K, Kadowaki T et al (1998) Involvement of a lysine-specific cysteine proteinase in hemoglobin adsorption and heme accumulation by Porphyromonas gingivalis. J Biol Chem 273:21225–21231

    Article  CAS  Google Scholar 

  21. Haruyama K, Yoshimura A, Naito M et al (2009) Identification of a gingipain-sensitive surface ligand of Porphyromonas gingivalis that induces Toll-like receptor 2- and 4-independent NF-κB activation in CHO cells. Infect Immun 77:4414–4420

    Article  CAS  Google Scholar 

  22. Gardner RG, Russell JB, Wilson DB et al (1996) Use of a modified Bacteroides-Prevotella shuttle vector to transfer a reconstructed beta-1,4-D-endoglucanase gene into Bacteroides uniformis and Prevotella ruminicola B(1)4. Appl Environ Microbiol 62:196–202

    Article  CAS  Google Scholar 

  23. Fletcher HM, Schenkein HA, Morgan RM et al (1995) Virulence of a Porphyromonas gingivalis W83 mutant defective in the prtH gene. Infect Immun 63:1521–1528

    Article  CAS  Google Scholar 

  24. Simon R, Priefer U, Puhler A (1983) A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in gram negative bacteria. Biotechnology 1:784–791

    Article  CAS  Google Scholar 

  25. Murakami Y, Imai M, Nakamura H et al (2002) Separation of the outer membrane and identification of major outer membrane proteins from Porphyromonas gingivalis. Eur J Oral Sci 110:157–162

    Article  CAS  Google Scholar 

  26. Potempa J, Pike R, Travis J (1995) The multiple forms of trypsin-like activity present in various strains of Porphyromonas gingivalis are due to the presence of either Arg-gingipain or Lys-gingipain. Infect Immun 63:1176–1182

    Article  CAS  Google Scholar 

  27. Sato K, Kakuda S, Yukitake H et al (2018) Immunoglobulin-like domains of the cargo proteins are essential for protein stability during secretion by the type IX secretion system. Mol Microbiol 110:64–81

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Keiko Sato .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2021 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Sato, K. (2021). Methods for Functional Characterization of the Type IX Secretion System of Porphyromonas gingivalis. In: Nagano, K., Hasegawa, Y. (eds) Periodontal Pathogens. Methods in Molecular Biology, vol 2210. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0939-2_12

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-0939-2_12

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0938-5

  • Online ISBN: 978-1-0716-0939-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics