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.

Mycobacterium tuberculosis lipoprotein LprG (Rv1411c) binds triacylated glycolipid agonists of Toll-like receptor 2

Abstract

Knockout of lprG results in decreased virulence of Mycobacterium tuberculosis (MTB) in mice. MTB lipoprotein LprG has TLR2 agonist activity, which is thought to be dependent on its N-terminal triacylation. Unexpectedly, here we find that nonacylated LprG retains TLR2 activity. Moreover, we show LprG association with triacylated glycolipid TLR2 agonists lipoarabinomannan, lipomannan and phosphatidylinositol mannosides (which share core structures). Binding of triacylated species was specific to LprG (not LprA) and increased LprG TLR2 agonist activity; conversely, association of glycolipids with LprG enhanced their recognition by TLR2. The crystal structure of LprG in complex with phosphatidylinositol mannoside revealed a hydrophobic pocket that accommodates the three alkyl chains of the ligand. In conclusion, we demonstrate a glycolipid binding function of LprG that enhances recognition of triacylated MTB glycolipids by TLR2 and may affect glycolipid assembly or transport for bacterial cell wall biogenesis.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Figure 1: NA-LprG carries a mycobacterial TLR2 agonist.
Figure 2: Crystal structure of NA-LprG reveals a hydrophobic pocket with the potential to carry a TLR2 agonist.
Figure 3: Site-directed mutagenesis to alter the structure of the NA-LprG pocket and binding of TLR2 agonists.
Figure 4: Triacylated mycobacterial glycolipids are associated with NA-LprG.
Figure 5: Crystal structure of Ac1PIM2 bound to NA-LprG.
Figure 6: LprG binds purified mycobacterial glycolipids and facilitates their recognition by TLR2.

Accession codes

Primary accessions

Protein Data Bank

References

  1. Flynn, J.L. & Chan, J. Immunology of tuberculosis. Annu. Rev. Immunol. 19, 93–129 (2001).

    Article  CAS  PubMed  Google Scholar 

  2. Janeway, C.A. Jr. Approaching the asymptote? Evolution and revolution in immunology. Cold Spring Harb. Symp. Quant. Biol. 54, 1–13 (1989).

    Article  CAS  PubMed  Google Scholar 

  3. Chan, J. et al. Microbial glycolipids: possible virulence factors that scavenge oxygen radicals. Proc. Natl. Acad. Sci. USA 86, 2453–2457 (1989).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Fratti, R.A., Chua, J., Vergne, I. & Deretic, V. Mycobacterium tuberculosis glycosylated phosphatidylinositol causes phagosome maturation arrest. Proc. Natl. Acad. Sci. USA 100, 5437–5442 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Vergne, I. et al. Mycobacterium tuberculosis phagosome maturation arrest: mycobacterial phosphatidylinositol analog phosphatidylinositol mannoside stimulates early endosomal fusion. Mol. Biol. Cell 15, 751–760 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Cole, S.T. et al. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature 393, 537–544 (1998).

    Article  CAS  PubMed  Google Scholar 

  7. Sutcliffe, I.C. & Harrington, D.J. Lipoproteins of Mycobacterium tuberculosis: an abundant and functionally diverse class of cell envelope components. FEMS Microbiol. Rev. 28, 645–659 (2004).

    Article  CAS  PubMed  Google Scholar 

  8. Bigi, F. et al. The knockout of the lprG-Rv1410 operon produces strong attenuation of Mycobacterium tuberculosis . Microbes Infect. 6, 182–187 (2004).

    Article  CAS  PubMed  Google Scholar 

  9. Rengarajan, J., Bloom, B.R. & Rubin, E.J. Genome-wide requirements for Mycobacterium tuberculosis adaptation and survival in macrophages. Proc. Natl. Acad. Sci. USA 102, 8327–8332 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Sassetti, C.M. & Rubin, E.J. Genetic requirements for mycobacterial survival during infection. Proc. Natl. Acad. Sci. USA 100, 12989–12994 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Farrow, M.F. & Rubin, E.J. Function of a mycobacterial major facilitator superfamily pump requires a membrane-associated lipoprotein. J. Bacteriol. 190, 1783–1791 (2008).

    Article  CAS  PubMed  Google Scholar 

  12. Sulzenbacher, G. et al. LppX is a lipoprotein required for the translocation of phthiocerol dimycocerosates to the surface of Mycobacterium tuberculosis . EMBO J. 25, 1436–1444 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Jones, B.W. et al. Different Toll-like receptor agonists induce distinct macrophage responses. J. Leukoc. Biol. 69, 1036–1044 (2001).

    CAS  PubMed  Google Scholar 

  14. Gilleron, M., Quesniaux, V.F. & Puzo, G. Acylation state of the phosphatidyl inositol hexamannosides from Mycobacterium bovis BCG and Mycobacterium tuberculosis H37Rv and its implication in TLR response. J. Biol. Chem. 278, 29880–29889 (2003).

    Article  CAS  PubMed  Google Scholar 

  15. Elass, E. et al. Mycobacterial lipomannan induces matrix metalloproteinase-9 expression in human macrophagic cells through a toll-like receptor 1 (TLR1)/TLR2- and CD14-dependent mechanism. Infect. Immun. 73, 7064–7068 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Tapping, R.I. & Tobias, P.S. Mycobacterial lipoarabinomannan mediates physical interactions between TLR1 and TLR2 to induce signaling. J. Endotoxin Res. 9, 264–268 (2003).

    Article  CAS  PubMed  Google Scholar 

  17. Bhatt, K. & Salgame, P. Host innate immune response to Mycobacterium tuberculosis . J. Clin. Immunol. 27, 347–362 (2007).

    Article  CAS  PubMed  Google Scholar 

  18. Pai, R.K., Convery, M., Hamilton, T.A., Boom, W.H. & Harding, C.V. Inhibition of IFN-γ-induced class II transactivator expression by a 19-kDa lipoprotein from Mycobacterium tuberculosis: a potential mechanism for immune evasion. J. Immunol. 171, 175–184 (2003).

    Article  CAS  PubMed  Google Scholar 

  19. Noss, E.H. et al. Toll-like receptor 2-dependent inhibition of macrophage class II MHC expression and antigen processing by 19 kD lipoprotein of Mycobacterium tuberculosis . J. Immunol. 167, 910–918 (2001).

    Article  CAS  PubMed  Google Scholar 

  20. Brightbill, H.D. et al. Host defense mechanisms triggered by microbial lipoproteins through toll-like receptors. Science 285, 732–736 (1999).

    Article  CAS  PubMed  Google Scholar 

  21. Takeuchi, O. et al. Cutting edge: role of Toll-like receptor 1 in mediating immune response to microbial lipoproteins. J. Immunol. 169, 10–14 (2002).

    Article  CAS  PubMed  Google Scholar 

  22. Nigou, J. et al. Mannan chain length controls lipoglycans signaling via and binding to TLR2. J. Immunol. 180, 6696–6702 (2008).

    Article  CAS  PubMed  Google Scholar 

  23. Pecora, N.D., Gehring, A.J., Canaday, D.H., Boom, W.H. & Harding, C.V. Mycobacterium tuberculosis LprA is a lipoprotein agonist of TLR2 that regulates innate immunity and APC function. J. Immunol. 177, 422–429 (2006).

    Article  CAS  PubMed  Google Scholar 

  24. Gehring, A.J., Dobos, K.M., Belisle, J.T., Harding, C.V. & Boom, W.H. Mycobacterium tuberculosis LprG (Rv1411c): A novel TLR-2 ligand that inhibits human macrophage class II MHC antigen processing. J. Immunol. 173, 2660–2668 (2004).

    Article  CAS  PubMed  Google Scholar 

  25. Jung, S.B. et al. The mycobacterial 38-kilodalton glycolipoprotein antigen activates the mitogen-activated protein kinase pathway and release of proinflammatory cytokines through Toll-like receptors 2 and 4 in human monocytes. Infect. Immun. 74, 2686–2696 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Jin, M.S. et al. Crystal structure of the TLR1–TLR2 heterodimer induced by binding of a tri-acylated lipopeptide. Cell 130, 1071–1082 (2007).

    Article  CAS  PubMed  Google Scholar 

  27. Rezwan, M., Grau, T., Tschumi, A. & Sander, P. Lipoprotein synthesis in mycobacteria. Microbiology 153, 652–658 (2007).

    Article  CAS  PubMed  Google Scholar 

  28. Nigou, J., Gilleron, M. & Puzo, G. Lipoarabinomannans: characterization of the multiacylated forms of the phosphatidyl-myo-inositol anchor by NMR spectroscopy. Biochem. J. 337, 453–460 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Bigi, F. et al. The gene encoding P27 lipoprotein and a putative antibiotic-resistance gene form an operon in Mycobacterium tuberculosis and Mycobacterium bovis . Microbiology 146, 1011–1018 (2000).

    Article  CAS  PubMed  Google Scholar 

  30. Gilleron, M., Nigou, J., Nicolle, D., Quesniaux, V. & Puzo, G. The acylation state of mycobacterial lipomannans modulates innate immunity response through toll-like receptor 2. Chem. Biol. 13, 39–47 (2006).

    Article  CAS  PubMed  Google Scholar 

  31. Kang, J.Y. et al. Recognition of lipopeptide patterns by Toll-like receptor 2-Toll-like receptor 6 heterodimer. Immunity 31, 873–884 (2009).

    Article  CAS  PubMed  Google Scholar 

  32. Berg, S., Kaur, D., Jackson, M. & Brennan, P.J. The glycosyltransferases of Mycobacterium tuberculosis—roles in the synthesis of arabinogalactan, lipoarabinomannan, and other glycoconjugates. Glycobiology 17, 35R–56R (2007).

    Article  CAS  Google Scholar 

  33. Finberg, R.W., Re, F., Popova, L., Golenbock, D.T. & Kurt-Jones, E.A. Cell activation by Toll-like receptors: role of LBP and CD14. J. Endotoxin Res. 10, 413–418 (2004).

    Article  CAS  PubMed  Google Scholar 

  34. Jiang, Z. et al. CD14 is required for MyD88-independent LPS signaling. Nat. Immunol. 6, 565–570 (2005).

    Article  CAS  PubMed  Google Scholar 

  35. Sklar, M.D., Tereba, A., Chen, B.D. & Walker, W.S. Transformation of mouse bone marrow cells by transfection with a human oncogene related to c-myc is associated with the endogenous production of macrophage colony stimulating factor 1. J. Cell. Physiol. 125, 403–412 (1985).

    Article  CAS  PubMed  Google Scholar 

  36. Flo, T.H. et al. Involvement of toll-like receptor (TLR) 2 and TLR4 in cell activation by mannuronic acid polymers. J. Biol. Chem. 277, 35489–35495 (2002).

    Article  CAS  PubMed  Google Scholar 

  37. Latz, E. et al. Lipopolysaccharide rapidly traffics to and from the Golgi apparatus with the toll-like receptor 4-MD-2–CD14 complex in a process that is distinct from the initiation of signal transduction. J. Biol. Chem. 277, 47834–47843 (2002).

    Article  CAS  PubMed  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  39. Vonrhein, C., Blanc, E., Roversi, P. & Bricogne, G. Automated structure solution with autoSHARP. Methods Mol. Biol. 364, 215–230 (2007).

    CAS  PubMed  Google Scholar 

  40. 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  PubMed  Google Scholar 

  41. Adams, P.D. et al. PHENIX: building new software for automated crystallographic structure determination. Acta Crystallogr. D Biol. Crystallogr. 58, 1948–1954 (2002).

    Article  PubMed  Google Scholar 

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

    Article  PubMed  Google Scholar 

  43. Pettersen, E.F. et al. UCSF Chimera–a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004).

    Article  CAS  PubMed  Google Scholar 

  44. Dundas, J. et al. CASTp: computed atlas of surface topography of proteins with structural and topographical mapping of functionally annotated residues. Nucleic Acids Res. 34, W116–8 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank V. Anderson and L. Sweet for advice and assistance with MS, A.G. Hise (Case Western Reserve Univ.) and A. Shizuo (Osaka Univ.) for bone marrow from TLR1−/− TLR6−/− mice, X. Ding, N. Nagy and K. Daniel for technical assistance, T. Musa for comments on the manuscript and K. Dobos-Elder and J. Belisle (Colorado State Univ.) for MTB lysates, antibodies and the pVV16 vector. This work was supported by US National Institutes of Health (NIH) grants AI035726, AI034343 and AI069085 to C.V.H., HL055967 and AI027243 to W.H.B., AI071155 and AI049313 to D.B.M. and AI068135 to J.C.S., the Robert A. Welch Foundation (J.C.S.), the Irvington Institute Fellowship Program of the Cancer Research Institute (C.S.), American Lung Association grant RG48786N (R.E.R.) and the Burroughs Wellcome Fund for Translational Research (D.B.M. and C.S.). Core facilities of the Case Western Reserve University Center for AIDS Research were supported by NIH grant AI067093.

Author information

Authors and Affiliations

Authors

Contributions

M.G.D., H.-C.T., N.D.P., T.-Y.C., A.R.A., S.S., R.E.R. and C.S. designed, performed and interpreted experiments and prepared the manuscript; D.B.M., W.H.B., J.C.S. and C.V.H. designed and interpreted experiments and prepared the manuscript.

Corresponding author

Correspondence to Clifford V Harding.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 and Supplementary Methods (PDF 3949 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Drage, M., Tsai, HC., Pecora, N. et al. Mycobacterium tuberculosis lipoprotein LprG (Rv1411c) binds triacylated glycolipid agonists of Toll-like receptor 2. Nat Struct Mol Biol 17, 1088–1095 (2010). https://doi.org/10.1038/nsmb.1869

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.1869

This article is cited by

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