Analytical and Bioanalytical Chemistry

, Volume 387, Issue 2, pp 497–511 | Cite as

Comprehensive profiling of N-acylhomoserine lactones produced by Yersinia pseudotuberculosis using liquid chromatography coupled to hybrid quadrupole–linear ion trap mass spectrometry

  • Catharine A. Ortori
  • Steve Atkinson
  • Siri Ram Chhabra
  • Miguel Cámara
  • Paul Williams
  • David A. Barrett
Original Paper

Abstract

A method for the comprehensive profiling of the N-acylhomoserine lactone (AHL) family of bacterial quorum-sensing molecules is presented using liquid chromatography (LC) coupled to hybrid quadrupole–linear ion trap (QqQLIT) mass spectrometry. Information-dependent acquisition (IDA), using triggered combinations of triple-quadrupole and linear ion trap modes in the same LC-MS/MS run, was used to simultaneously screen, quantify and identify multiple AHLs in a single sample. This MS method uses common AHL fragment ions attributed to the homoserine moiety and the 3-oxo-, 3-hydroxy- or unsubstituted acyl side chains, to identify unknown AHLs in cell-free culture supernatants in an unbiased manner. This LC-MS technique was applied to determine the relative molar ratios of AHLs produced by Yersinia pseudotuberculosis and the consequences of inactivating by mutation either or both of the AHL synthase genes (ypsI and ytbI) on AHL profile and concentration. The Y. pseudotuberculosis wild type but not the ypsI ytbI double mutant produced at least 24 different AHLs with acyl chains ranging from C4 to C15 with or without 3-oxo or 3-hydroxy substituents. YtbI, in contrast to YpsI, could direct the synthesis of all of the AHLs identified. The most abundant and hence most biologically relevant Y. pseudotuberculosis AHLs were found to be the 3-oxo-substituted C6, C7 and C8 AHLs and the unsubstituted C6 and C8 compounds. The LC-QqQLIT methodology is broadly applicable to quorum-sensing signal molecule analysis and can provide comprehensive AHL profiles and concentrations from a single sample and simultaneously collect confirmatory spectra for each AHL identified.

Keywords

Yersinia Quorum sensing N-Acylhomoserine lactones LC-MS Quadrupole linear ion trap 

Supplementary material

216_2006_710_MOESM1_ESM.doc (390 kb)
(DOC 399 kb)

References

  1. 1.
    Swift S, Downie JA, Whitehead N, Barnard AML, Salmond, GPC Williams P (2001) Adv Microb Physiol 45:199–270Google Scholar
  2. 2.
    Williams P, Winzer K, Chan W, Cámara M (2006) Phil Trans Roy Soc B (in press)Google Scholar
  3. 3.
    Milton D, Chalker V, Hardman A, Cámara M, Williams P (2001) J Bact 183:3537–3547CrossRefGoogle Scholar
  4. 4.
    Chhabra SR, Philipp B, Eberl L, Givskov M, Williams P, Camara M (2005) In: Schulz S (eds) Chemistry of pheromones and other semiochemicals, vol 240. Springer-Verlag, pp 279–315Google Scholar
  5. 5.
    Swift S, Williams P, Stewart GSAB (1999) In: Winans S, Dunny G (eds) Cell-cell signalling in bacteria. American Society for Microbiology Press, pp 291–313Google Scholar
  6. 6.
    Swift S, Winson MK, Chan PF, Bainton NJ, Birdsall M, Reeves PJ, Rees CED, Chhabra SR, Hill PH, Throup JP, Bycroft BW, Salmond GPC, Williams P, Stewart GSAB (1993) Mol Microbiol 10:511–520CrossRefGoogle Scholar
  7. 7.
    Wren BW (2003) Nat Rev Microbiol 1:55–64CrossRefGoogle Scholar
  8. 8.
    Achtman M, Zurth K, Morelli G, Torrea G, Guiyoule A, Carniel E (1999) Proc Natl Acad Sci USA 96:14043–14048CrossRefGoogle Scholar
  9. 9.
    Atkinson S, Throup JP, Stewart GSAB, Williams P (1999) Mol Microbiol 33:1267–1277CrossRefGoogle Scholar
  10. 10.
    Yates EA, Philipp B, Buckley C, Atkinson S, Chhabra SR, Sockett RE, Goldner M, Dessaux Y, Camara M, Smith H, Williams P (2002) Infect Immun 70:5635–5646CrossRefGoogle Scholar
  11. 11.
    Bainton NJ, Stead P, Chhabra SR, Bycroft BW, Salmond GPC, Stewart GSAB, Williams P (1992) Biochem J 288:997–1004Google Scholar
  12. 12.
    McClean KH, Winson MK, Fish L, Taylor A, Chhabra SR, Cámara M, Daykin M, Swift S, Bycroft BW, Stewart GSAB, Williams P (1997) Microbiology-SGM 143:3703–3711CrossRefGoogle Scholar
  13. 13.
    Winson MK, Swift S, Fish L, Throup JP, Jorgensen F, Chhabra SR, Bycroft, BW, Williams P, Stewart GSAB (1998) FEMS Letts 163:185–192CrossRefGoogle Scholar
  14. 14.
    Shaw PD, Gao P, Daly SL, Cha C, Cronan JE Jr, Rinehaert KL, Farrand SK (1997) Proc Nat Acad Sci USA 94:6036–6041CrossRefGoogle Scholar
  15. 15.
    Andersen JB, Heydorn A, Hentzer M, Eberl L, Geisenberger O, Christensen BB, Molin S, Givskov M (2001) Appl Environ Microbiol 67:575–585CrossRefGoogle Scholar
  16. 16.
    Morin D, Grasland B, Vallee-Rehel K, Dufau C, Hara D (2003) J Chromatogr A 1002:79–82CrossRefGoogle Scholar
  17. 17.
    Hager JW (2002) Rapid Commun Mass Spectrom 16:512–526CrossRefGoogle Scholar
  18. 18.
    Hager JW, Yves Le Blanc JC (2003) Rapid Commun Mass Spectrom 17:1056–1064CrossRefGoogle Scholar
  19. 19.
    Chhabra SR, Stead P, Bainton NJ, Salmond GPC, Stewart GSAB, Williams P, Bycroft BW (1993) J Antibiotics 46:441–454Google Scholar
  20. 20.
    Chhabra SR, Harty C, Hooi DSW, Daykin M, Williams P, Pritchard DI, Bycroft BW (2003) J Med Chem 46:97–104CrossRefGoogle Scholar
  21. 21.
    Throup JK, Cámara M, Briggs GS, Winson MK, Bycroft BW, Williams P, Stewart GSAB (1995) Mol Microbiol 17:345–356CrossRefGoogle Scholar
  22. 22.
    Atkinson S, Sockett, RE, Camara M, Williams P (2006) Curr Issues Mol Biol 8:1–10Google Scholar
  23. 23.
    Kirwan JP, Gould TA, Schweizer HP, Bearden SW, Murphy RC, Churchill MEA (2006) J Bacteriol 188:784–788CrossRefGoogle Scholar
  24. 24.
    Zhang L, Murphy PJ, Kerr A, Tate ME (1993) Nature 362:446–448CrossRefGoogle Scholar
  25. 25.
    Toth IK, Newton JA, Hyman LJ, Lees AK, Daykin M, Otori C, Williams P, Fray RG (2004) Mol Plant Microb Interact 8:880–887CrossRefGoogle Scholar
  26. 26.
    Angel-Picard C, Faure D, Penot I, Dessaux Y (2005) Environ Microbiol 7:1796–1808CrossRefGoogle Scholar
  27. 27.
    Lithgow JK, Wilkinson A, Hardman A, Rodelas B, Wisniewski-Dye F, Williams P, Downie JA (2000) Mol Microbiol 37:81–97CrossRefGoogle Scholar
  28. 28.
    Horng YT, Deng SC, Daykin M, Soo PC, Wei JR, Luh KT, Ho SW, Swift S, Lai HC, Williams P (2002) Mol Microbiol 45:1655–1671CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Catharine A. Ortori
    • 1
  • Steve Atkinson
    • 2
  • Siri Ram Chhabra
    • 2
  • Miguel Cámara
    • 2
  • Paul Williams
    • 2
  • David A. Barrett
    • 1
  1. 1.Centre for Analytical Bioscience, School of PharmacyUniversity of NottinghamNottinghamUK
  2. 2.Institute of Infection, Immunity and Inflammation, Centre for Biomolecular SciencesUniversity of NottinghamNottinghamUK

Personalised recommendations