Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Transition state analogs of 5′-methylthioadenosine nucleosidase disrupt quorum sensing

Nature Chemical Biology volume 5, pages 251–257 (2009)Cite this article

Abstract

5′-Methylthioadenosine/S-adenosylhomocysteine nucleosidase (MTAN) is a bacterial enzyme involved in S-adenosylmethionine–related quorum sensing pathways that induce bacterial pathogenesis factors. Transition state analogs MT-DADMe-Immucillin-A, EtT-DADMe-Immucillin-A and BuT-DADMe-Immucillin-A are slow-onset, tight-binding inhibitors of Vibrio cholerae MTAN (VcMTAN), with equilibrium dissociation constants of 73, 70 and 208 pM, respectively. Structural analysis of VcMTAN with BuT-DADMe-Immucillin-A revealed interactions contributing to the high affinity. We found that in V. cholerae cells, these compounds are potent MTAN inhibitors with IC50 values of 27, 31 and 6 nM for MT-, EtT- and BuT-DADMe-Immucillin-A, respectively; the compounds disrupt autoinducer production in a dose-dependent manner without affecting growth. MT- and BuT-DADMe-Immucillin-A also inhibited autoinducer-2 production in enterohemorrhagic Escherichia coli O157:H7 with IC50 values of 600 and 125 nM, respectively. BuT-DADMe-Immucillin-A inhibition of autoinducer-2 production in both strains persisted for several generations and caused reduction in biofilm formation. These results support MTAN's role in quorum sensing and its potential as a target for bacterial anti-infective drug design.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Role of MTAN in bacterial utilization of SAM.
Figure 2: The reaction catalyzed by MTAN with MTA as substrate.
Figure 3: Crystal structure of VcMTAN in complex with BuT-DADMe-ImmA.
Figure 4: Effect of BuT-DADMe-ImmA on AI-2 production in pathogenic E. coli and V. cholerae upon short-term and long-term inhibitor treatment, and on static biofilm formation.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. Fuqua, W.C., Winans, S.C. & Greenberg, E.P. Quorum sensing in bacteria: the LuxR-LuxI family of cell density-responsive transcriptional regulators. J. Bacteriol. 176, 269–275 (1994).

    Article  CAS  Google Scholar 

  2. Sperandio, V. Novel approaches to bacterial infection therapy by interfering with bacteria-to-bacteria signaling. Expert Rev. Anti Infect. Ther. 5, 271–276 (2007).

    Article  CAS  Google Scholar 

  3. Vendeville, A., Winzer, K., Heurlier, K., Tang, C.M. & Hardie, K.R. Making 'sense' of metabolism: autoinducer-2, LuxS and pathogenic bacteria. Nat. Rev. Microbiol. 3, 383–396 (2005).

    Article  CAS  Google Scholar 

  4. Cegelski, L., Marshall, G.R., Eldridge, G.R. & Hultgren, S.J. The biology and future prospects of antivirulence therapies. Nat. Rev. Microbiol. 6, 17–27 (2008).

    Article  CAS  Google Scholar 

  5. Winzer, K. & Williams, P. Quorum sensing and the regulation of virulence gene expression in pathogenic bacteria. Int. J. Med. Microbiol. 291, 131–143 (2001).

    Article  CAS  Google Scholar 

  6. Stroeher, U.H., Paton, A.W., Ogunniyi, A.D. & Paton, J.C. Mutation of luxS of Streptococcus pneumoniae affects virulence in a mouse model. Infect. Immun. 71, 3206–3212 (2003).

    Article  CAS  Google Scholar 

  7. Winzer, K. et al. Role of Neisseria meningitidis luxS in cell-to-cell signaling and bacteremic infection. Infect. Immun. 70, 2245–2248 (2002).

    Article  CAS  Google Scholar 

  8. Harasawa, H. et al. Chemotherapy targeting methylthioadenosine phosphorylase (MTAP) deficiency in adult T cell leukemia (ATL). Leukemia 16, 1799–1807 (2002).

    Article  CAS  Google Scholar 

  9. Basu, I. et al. A transition state analogue of 5′-methylthioadenosine phosphorylase induces apoptosis in head and neck cancers. J. Biol. Chem. 282, 21477–21486 (2007).

    Article  CAS  Google Scholar 

  10. Singh, V., Lee, J.E., Nunez, S., Howell, P.L. & Schramm, V.L. Transition state structure of 5′-methylthioadenosine/S-adenosylhomocysteine nucleosidase from Escherichia coli and its similarity to transition state analogues. Biochemistry 44, 11647–11659 (2005).

    Article  CAS  Google Scholar 

  11. Singh, V. & Schramm, V.L. Transition-state analysis of S-pneumoniae 5′-methylthioadenosine nucleosidase. J. Am. Chem. Soc. 129, 2783–2795 (2007).

    Article  CAS  Google Scholar 

  12. Singh, V., Luo, M., Brown, R.L., Norris, G.E. & Schramm, V.L. Transition-state structure of Neisseria meningitides 5′-methylthioadenosine/S-adenosylhomocysteine nucleosidase. J. Am. Chem. Soc. 129, 13831–13833 (2007).

    Article  CAS  Google Scholar 

  13. Singh, V. & Schramm, V.L. Transition-state structure of human 5′-methylthioadenosine phosphorylase. J. Am. Chem. Soc. 128, 14691–14696 (2006).

    Article  CAS  Google Scholar 

  14. Balakrishnan, K., Nimmanapalli, R., Ravandi, F., Keating, M.J. & Gandhi, V. Forodesine, an inhibitor of purine nucleoside phosphorylase, induces apoptosis in chronic lymphocytic leukemia cells. Blood 108, 2392–2398 (2006).

    Article  CAS  Google Scholar 

  15. Robak, T., Lech-Maranda, E., Koerycka, A. & Robak, E. Purine nucleoside analogs as immunosuppressive and antineoplastic agents: mechanism of action and clinical activity. Curr. Med. Chem. 13, 3165–3189 (2006).

    Article  CAS  Google Scholar 

  16. Evans, G.B., Furneaux, R.H., Schramm, V.L., Singh, V. & Tyler, P.C. Targeting the polyamine pathway with transition-state analogue inhibitors of 5′-methylthioadenosine phosphorylase. J. Med. Chem. 47, 3275–3281 (2004).

    Article  CAS  Google Scholar 

  17. Evans, G.B. et al. Second generation transition state analogue inhibitors of human 5′-methylthioadenosine phosphorylase. J. Med. Chem. 48, 4679–4689 (2005).

    Article  CAS  Google Scholar 

  18. Singh, V. et al. Femtomolar transition state analogue inhibitors of 5′-methylthioadenosine/S-adenosylhomocysteine nucleosidase from Escherichia coli. J. Biol. Chem. 280, 18265–18273 (2005).

    Article  CAS  Google Scholar 

  19. Singh, V. et al. Picomolar transition state analogue inhibitors of human 5′-methylthioadenosine phosphorylase and X-ray structure with MT-Immucillin-A. Biochemistry 43, 9–18 (2004).

    Article  CAS  Google Scholar 

  20. Singh, V. et al. Structure and inhibition of a quorum sensing target from Streptococcus pneumoniae. Biochemistry 45, 12929–12941 (2006).

    Article  CAS  Google Scholar 

  21. Gutierrez, J.A. et al. Picomolar inhibitors as transition-state probes of 5′-methylthioadenosine nucleosidases. ACS Chem. Biol. 2, 725–734 (2007).

    Article  CAS  Google Scholar 

  22. Lee, J.E. et al. Structural rationale for the affinity of pico- and femtomolar transition state analogues of Escherichia coli 5′-methylthioadenosine/S-adenosylhomocysteine nucleosidase. J. Biol. Chem. 280, 18274–18282 (2005).

    Article  CAS  Google Scholar 

  23. Bassler, B.L., Greenberg, E.P. & Stevens, A.M. Cross-species induction of luminescence in the quorum-sensing bacterium Vibrio harveyi. J. Bacteriol. 179, 4043–4045 (1997).

    Article  CAS  Google Scholar 

  24. Saen-Oon, S., Ghanem, M., Schramm, V.L. & Schwartz, S.D. Remote mutations and active site dynamics correlate with catalytic properties of purine nucleoside phosphorylase. Biophys. J. 94, 4078–4088 (2008).

    Article  CAS  Google Scholar 

  25. Anand, S.K. & Griffiths, M.W. Quorum sensing and expression of virulence in Escherichia coli O157:H7. Int. J. Food Microbiol. 85, 1–9 (2003).

    Article  CAS  Google Scholar 

  26. Sperandio, V., Mellies, J.L., Nguyen, W., Shin, S. & Kaper, J.B. Quorum sensing controls expression of the type III secretion gene transcription and protein secretion in enterohemorrhagic and enteropathogenic Escherichia coli. Proc. Natl. Acad. Sci. USA 96, 15196–15201 (1999).

    Article  CAS  Google Scholar 

  27. Li, J. et al. Quorum sensing in Escherichia coli is signaled by AI-2/LsrR: effects on small RNA and biofilm architecture. J. Bacteriol. 189, 6011–6020 (2007).

    Article  CAS  Google Scholar 

  28. Herzberg, M., Kaye, I.K., Peti, W. & Wood, T.K. YdgG (TqsA) controls biofilm formation in Escherichia coli K-12 through autoinducer 2 transport. J. Bacteriol. 188, 587–598 (2006).

    Article  CAS  Google Scholar 

  29. Zhu, J. & Mekalanos, J.J. Quorum sensing-dependent biofilms enhance colonization in Vibrio cholerae. Dev. Cell 5, 647–656 (2003).

    Article  CAS  Google Scholar 

  30. Matz, C. et al. Biofilm formation and phenotypic variation enhance predation-driven persistence of Vibrio cholerae. Proc. Natl. Acad. Sci. USA 102, 16819–16824 (2005).

    Article  CAS  Google Scholar 

  31. Hammer, B.K. & Bassler, B.L. Quorum sensing controls biofilm formation in Vibrio cholerae. Mol. Microbiol. 50, 101–114 (2003).

    Article  CAS  Google Scholar 

  32. Waters, C.M., Lu, W., Rabinowitz, J.D. & Bassler, B.L. Quorum sensing controls biofilm formation in Vibrio cholerae through modulation of cyclic di-GMP levels and repression of vpsT. J. Bacteriol. 190, 2527–2536 (2008).

    Article  CAS  Google Scholar 

  33. Zhu, J. et al. Quorum-sensing regulators control virulence gene expression in Vibrio cholerae. Proc. Natl. Acad. Sci. USA 99, 3129–3134 (2002).

    Article  CAS  Google Scholar 

  34. Joelsson, A., Liu, Z. & Zhu, J. Genetic and phenotypic diversity of quorum-sensing systems in clinical and environmental isolates of Vibrio cholerae. Infect. Immun. 74, 1141–1147 (2006).

    Article  CAS  Google Scholar 

  35. Gonzalez Barrios, A.F. et al. Autoinducer 2 controls biofilm formation in Escherichia coli through a novel motility quorum-sensing regulator (MqsR, B3022). J. Bacteriol. 188, 305–316 (2006).

    Article  Google Scholar 

  36. Surette, M.G. & Bassler, B.L. Quorum sensing in Escherichia coli and Salmonella typhimurium. Proc. Natl. Acad. Sci. USA 95, 7046–7050 (1998).

    Article  CAS  Google Scholar 

  37. Dunny, G.M. & Leonard, B.A.B. Cell-cell communication in gram-positive bacteria. Annu. Rev. Microbiol. 51, 527–564 (1997).

    Article  CAS  Google Scholar 

  38. Balestrino, D., Haagensen, J.A.J., Rich, C. & Forestier, C. Characterization of type 2 quorum sensing in Klebsiella pneumoniae and relationship with biofilm formation. J. Bacteriol. 187, 2870–2880 (2005).

    Article  CAS  Google Scholar 

  39. Joyce, E.A. et al. LuxS is required for persistent Pneumococcal carriage and expression of virulence and biosynthesis genes. Infect. Immun. 72, 2964–2975 (2004).

    Article  CAS  Google Scholar 

  40. Rader, B.A., Campagna, S.R., Semmelhack, M.F., Bassler, B.L. & Guillemin, K. The quorum-sensing molecule autoinducer 2 regulates motility and flagellar morphogenesis in Helicobacter pylori. J. Bacteriol. 189, 6109–6117 (2007).

    Article  CAS  Google Scholar 

  41. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).

    Article  CAS  Google Scholar 

  42. Potterton, E., Briggs, P., Turkenburg, M. & Dodson, E. A graphical user interface to the CCP4 program suite. Acta Crystallogr. D Biol. Crystallogr. 59, 1131–1137 (2003).

    Article  Google Scholar 

  43. Vagin, A. & Teplyakov, A. MOLREP: an automated program for molecular replacement. J. Appl. Crystallogr. 30, 1022–1025 (1997).

    Article  CAS  Google Scholar 

  44. Murshudov, G.N., Vagin, A.A. & Dodson, E.J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997).

    Article  CAS  Google Scholar 

  45. Emsley, P. & Cowtan, K. Model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  Google Scholar 

  46. DeLano, W.L. The PyMOL Molecular Graphics System (DeLano Scientific, Palo Alto, California, USA, 2002).

    Google Scholar 

  47. Greenberg, E.P., Hastings, J.W. & Ulitzur, S. Induction of luciferase synthesis in Beneckea harveyi by other marine bacteria. Arch. Microbiol. 120, 87–91 (1979).

    Article  CAS  Google Scholar 

  48. O'Toole, G.A. et al. Genetic approaches to study of biofilms. Methods Enzymol. 310, 91–109 (1999).

    Article  CAS  Google Scholar 

  49. Parsek, M.R., Val, D.L., Hanzelka, B.L., Cronan, J.E. & Greenberg, E.P. Acyl homoserine-lactone quorum-sensing signal generation. Proc. Natl. Acad. Sci. USA 96, 4360–4365 (1999).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We acknowledge R.H. Furneaux, G.B. Evans, D.H. Lenz, G.F. Painter and P.C. Tyler of Industrial Research Laboratory, Inc. for supplying the DADMe-Immucillins; M.G. Surette (University of Calgary) for providing V. harveyi strains BB120 and BB170; C. Bradbeer (University of Virginia) for the E. coli MTAN knockout; and the US National Institutes of Health grant GM41916 for funding. Data for this study were collected at beamline X29A of the National Synchrotron Light Source. Financial support comes principally from the Offices of Biological and Environmental Research and of Basic Energy Sciences of the US Department of Energy, and from the National Center for Research Resources of the US National Institutes of Health.

Author information

Authors and Affiliations

  1. Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York, USA

    Jemy A Gutierrez, Tamara Crowder, Agnes Rinaldo-Matthis, Meng-Chiao Ho, Steven C Almo & Vern L Schramm

Authors
  1. Jemy A Gutierrez
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tamara Crowder
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Agnes Rinaldo-Matthis
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Meng-Chiao Ho
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Steven C Almo
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Vern L Schramm
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

J.A.G. performed the inhibition assays on recombinant VcMTAN and on E. coli and V. cholerae cells, and co-wrote the manuscript. T.C. expressed, purified and measured activity of the recombinant VcMTAN. A.R.-M., M.-C.H. and S.C.A. did the structure determination, refinement and characterization for the VcMTAN–BuT-DADMe-ImmA complex. V.L.S. designed the experiments and co-wrote the manuscript.

Corresponding author

Correspondence to Vern L Schramm.

Ethics declarations

Competing interests

V.L.S. is co-scientific founder of Pico Pharmaceuticals Inc., a new startup biotechnology company involved in anticancer and antibiotic development.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–3, Supplementary Tables 1 and 2, and Supplementary Methods (PDF 334 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Gutierrez, J., Crowder, T., Rinaldo-Matthis, A. et al. Transition state analogs of 5′-methylthioadenosine nucleosidase disrupt quorum sensing. Nat Chem Biol 5, 251–257 (2009). https://doi.org/10.1038/nchembio.153

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchembio.153

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing