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Crystal structure of the FimD usher bound to its cognate FimC–FimH substrate


Type 1 pili are the archetypal representative of a widespread class of adhesive multisubunit fibres in Gram-negative bacteria. During pilus assembly, subunits dock as chaperone-bound complexes to an usher, which catalyses their polymerization and mediates pilus translocation across the outer membrane. Here we report the crystal structure of the full-length FimD usher bound to the FimC–FimH chaperone–adhesin complex and that of the unbound form of the FimD translocation domain. The FimD–FimC–FimH structure shows FimH inserted inside the FimD 24-stranded β-barrel translocation channel. FimC–FimH is held in place through interactions with the two carboxy-terminal periplasmic domains of FimD, a binding mode confirmed in solution by electron paramagnetic resonance spectroscopy. To accommodate FimH, the usher plug domain is displaced from the barrel lumen to the periplasm, concomitant with a marked conformational change in the β-barrel. The amino-terminal domain of FimD is observed in an ideal position to catalyse incorporation of a newly recruited chaperone–subunit complex. The FimD–FimC–FimH structure provides unique insights into the pilus subunit incorporation cycle, and captures the first view of a protein transporter in the act of secreting its cognate substrate.

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Figure 1: Structure of the FimD–FimC–FimH complex.
Figure 2: Channel conformations in apo and activated (FimC–FimH-engaged) FimD usher.
Figure 3: FimC–FimH interactions with FimD in the FimD–FimC–FimH complex.
Figure 4: Chaperone–subunit incorporation cycle at the FimD usher.

Accession codes

Primary accessions

Protein Data Bank


  1. Mulvey, M. A. et al. Induction and evasion of host defenses by type 1-piliated uropathogenic Escherichia coli . Science 282, 1494–1497 (1998)

    CAS  Article  Google Scholar 

  2. Sauer, F. G., Remaut, H., Hultgren, S. J. & Waksman, G. Fiber assembly by the chaperone-usher pathway. Biochim. Biophys. Acta 1694, 259–267 (2004)

    CAS  Article  Google Scholar 

  3. Waksman, G. & Hultgren, S. J. Structural biology of the chaperone-usher pathway of pilus biogenesis. Nature Rev. Microbiol. 7, 765–774 (2009)

    CAS  Article  Google Scholar 

  4. Jones, C. H. et al. FimH adhesin of type 1 pili is assembled into a fibrillar tip structure in the Enterobacteriaceae. Proc. Natl Acad. Sci. USA 92, 2081–2085 (1995)

    CAS  ADS  Article  Google Scholar 

  5. Hahn, E. et al. Exploring the 3D molecular architecture of Escherichia coli type 1 pili. J. Mol. Biol. 323, 845–857 (2002)

    CAS  Article  Google Scholar 

  6. Le Trong, I. et al. Structural basis for mechanical force regulation of the adhesin FimH via finger trap-like β sheet twisting. Cell 141, 645–655 (2010)

    CAS  Article  Google Scholar 

  7. Vetsch, M. et al. Pilus chaperones represent a new type of protein-folding catalyst. Nature 431, 329–333 (2004)

    CAS  ADS  Article  Google Scholar 

  8. Barnhart, M. M. et al. PapD-like chaperones provide the missing information for folding of pilin proteins. Proc. Natl Acad. Sci. USA 97, 7709–7714 (2000)

    CAS  ADS  Article  Google Scholar 

  9. Jacob-Dubuisson, F., Striker, R. & Hultgren, S. J. Chaperone-assisted self-assembly of pili independent of cellular energy. J. Biol. Chem. 269, 12447–12455 (1994)

    CAS  Google Scholar 

  10. Nishiyama, M., Ishikawa, T., Rechsteiner, H. & Glockshuber, R. Reconstitution of pilus assembly reveals a bacterial outer membrane catalyst. Science 320, 376–379 (2008)

    CAS  ADS  Article  Google Scholar 

  11. Sauer, F. G. et al. Structural basis of chaperone function and pilus biogenesis. Science 285, 1058–1061 (1999)

    CAS  Article  Google Scholar 

  12. Choudhury, D. et al. X-ray structure of the FimC-FimH chaperone-adhesin complex from uropathogenic Escherichia coli . Science 285, 1061–1066 (1999)

    CAS  Article  Google Scholar 

  13. Zavialov, A. V. et al. Structure and biogenesis of the capsular F1 antigen from Yersinia pestis: preserved folding energy drives fiber formation. Cell 113, 587–596 (2003)

    CAS  Article  Google Scholar 

  14. Sauer, F. G., Pinkner, J. S., Waksman, G. & Hultgren, S. J. Chaperone priming of pilus subunits facilitates a topological transition that drives fiber formation. Cell 111, 543–551 (2002)

    CAS  Article  Google Scholar 

  15. Remaut, H. et al. Fiber formation across the bacterial outer membrane by the chaperone/usher pathway. Cell 133, 640–652 (2008)

    CAS  Article  Google Scholar 

  16. Nishiyama, M., Vetsch, M., Puorger, C., Jelesarov, I. & Glockshuber, R. Identification and characterization of the chaperone-subunit complex-binding domain from the type 1 pilus assembly platform FimD. J. Mol. Biol. 330, 513–525 (2003)

    CAS  Article  Google Scholar 

  17. Nishiyama, M. et al. Structural basis of chaperone-subunit complex recognition by the type 1 pilus assembly platform FimD. EMBO J. 24, 2075–2086 (2005)

    CAS  Article  Google Scholar 

  18. Ng, T. W., Akman, L., Osisami, M. & Thanassi, D. G. The usher N terminus is the initial targeting site for chaperone-subunit complexes and participates in subsequent pilus biogenesis events. J. Bacteriol. 186, 5321–5331 (2004)

    CAS  Article  Google Scholar 

  19. Shu Kin So, S. & Thanassi, D. G. Analysis of the requirements for pilus biogenesis at the outer membrane usher and the function of the usher C-terminus. Mol. Microbiol. 60, 364–375 (2006)

    Article  Google Scholar 

  20. Ford, B. et al. Structural homology between the C-terminal domain of the PapC usher and its plug. J. Bacteriol. 192, 1824–1831 (2010)

    CAS  Article  Google Scholar 

  21. Munera, D., Hultgren, S. & Fernandez, L. A. Recognition of the N-terminal lectin domain of FimH adhesin by the usher FimD is required for type 1 pilus biogenesis. Mol. Microbiol. 64, 333–346 (2007)

    CAS  Article  Google Scholar 

  22. Saulino, E. T., Thanassi, D. G., Pinkner, J. S. & Hultgren, S. J. Ramifications of kinetic partitioning on usher-mediated pilus biogenesis. EMBO J. 17, 2177–2185 (1998)

    CAS  Article  Google Scholar 

  23. Li, Q. et al. The differential affinity of the usher for chaperone-subunit complexes is required for assembly of complete pili. Mol. Microbiol. 76, 159–172 (2010)

    CAS  Article  Google Scholar 

  24. Lawrence, M. C. & Colman, P. M. Shape complementarity at protein/protein interfaces. J. Mol. Biol. 234, 946–950 (1993)

    CAS  Article  Google Scholar 

  25. Eidam, O., Dworkowski, F. S., Glockshuber, R., Grutter, M. G. & Capitani, G. Crystal structure of the ternary FimC–FimFt–FimDN complex indicates conserved pilus chaperone–subunit complex recognition by the usher FimD. FEBS Lett. 582, 651–655 (2008)

    CAS  Article  Google Scholar 

  26. Remaut, H. et al. Donor-strand exchange in chaperone-assisted pilus assembly proceeds through a concerted β strand displacement mechanism. Mol. Cell 22, 831–842 (2006)

    CAS  Article  Google Scholar 

  27. Verger, D., Miller, E., Remaut, H., Waksman, G. & Hultgren, S. Molecular mechanism of P pilus termination in uropathogenic Escherichia coli . EMBO Rep. 7, 1228–1232 (2006)

    CAS  Article  Google Scholar 

  28. Rose, R. J. et al. Unraveling the molecular basis of subunit specificity in P pilus assembly by mass spectrometry. Proc. Natl Acad. Sci. USA 105, 12873–12878 (2008)

    CAS  ADS  Article  Google Scholar 

  29. Verger, D. et al. Structural determinants of polymerization reactivity of the P pilus adaptor subunit PapF. Structure 16, 1724–1731 (2008)

    CAS  Article  Google Scholar 

  30. Pinkner, J. S. et al. Rationally designed small compounds inhibit pilus biogenesis in uropathogenic bacteria. Proc. Natl Acad. Sci. USA 103, 17897–17902 (2006)

    CAS  ADS  Article  Google Scholar 

  31. McCoy, A. J. Solving structures of protein complexes by molecular replacement with Phaser . Acta Crystallogr. D 63, 32–41 (2007)

    CAS  Article  Google Scholar 

  32. Vagin, A. & Teplyakov, A. Molecular replacement with MOLREP . Acta Crystallogr. D 66, 22–25 (2010)

    CAS  Article  Google Scholar 

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

    Article  Google Scholar 

  34. Murshudov, G. N. et al. REFMAC5 for the refinement of macromolecular crystal structures. Acta Crystallogr. D 67, 355–367 (2011)

    CAS  Article  Google Scholar 

  35. Collaborative Computational Project, number 4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D 50, 760–763 (1994)

  36. Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D 66, 213–221 (2010)

    CAS  Article  Google Scholar 

  37. Pannier, M., Veit, S., Godt, A., Jeschke, G. & Spiess, H. W. Dead-time free measurement of dipole-dipole interactions between electron spins. J. Magn. Reson. 142, 331–340 (2000)

    CAS  ADS  Article  Google Scholar 

  38. Jeschke, G. et al. DeerAnalysis2006—a comprehensive software package for analyzing pulsed ELDOR data. Appl. Magn. Reson. 30, 473–498 (2006)

    CAS  Article  Google Scholar 

  39. Polyhach, Y., Bordignon, E. & Jeschke, G. Rotamer libraries of spin labelled cysteines for protein studies. Phys. Chem. Chem. Phys. 13, 2356–2366 (2011)

    CAS  Article  Google Scholar 

  40. Chiu, J., Tillett, D., Dawes, I. W. & March, P. E. Site-directed, Ligase-Independent Mutagenesis (SLIM) for highly efficient mutagenesis of plasmids greater than 8kb. J. Microbiol. Methods 73, 195–198 (2008)

    CAS  Article  Google Scholar 

  41. Leslie, A. G. The integration of macromolecular diffraction data. Acta Crystallogr. D 62, 48–57 (2006)

    Article  Google Scholar 

  42. Evans, P. Scaling and assessment of data quality. Acta Crystallogr. D 62, 72–82 (2006)

    Article  Google Scholar 

  43. Otwinowski, Z. & Minor, W. in Methods in Enzymology Vol. 276 (ed. Carter, C. W. Jr) Ch. 20, 307–326 (Elsevier, 1997)

    Google Scholar 

  44. Vagin, A. A. & Isupov, M. N. Spherically averaged phased translation function and its application to the search for molecules and fragments in electron-density maps. Acta Crystallogr. D 57, 1451–1456 (2001)

    CAS  Article  Google Scholar 

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This work was funded by Medical Research Council grant 85602 to G.W., NIH grant GM62987 to D.G.T., NIH grants 49950, 29549 and 48689 to S.J.H., and NIH grant GM74985 and BNL LDRD grant 10-16 to H.L.; H.R. is supported by a VIB Young PI project grant and the Odysseus program of the FWO-Vlaanderen. K.F.P. is supported by a Schrödinger Fellowship from the Austrian Science Fund, project J 2959-N17. We thank the staff of beamlines X25 and X29 at NSLS, the staff of beamline ID23-1 at ESRF, N. Cronin for technical assistance during data collection, and H. Saibil and E. Orlova for comments on the manuscript.

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Authors and Affiliations



Author contribution G.P. produced the FimD–FimC–FimH complex, grew the crystals of this complex, collected X-ray crystallographic data, and initiated the determination of the structure by molecular replacement, and participated in the building and refinement of the structure. H.R. produced the FimD–FimC–FimH complex, trained G.P., supervised the work, analysed the structures and wrote the paper. T.W. grew crystals of the FimD translocation domain, collected X-ray crystallographic data, and determined the structure. W.J.A. set up the DSE assay and prepared the samples for EPR. K.F.P. carried out the EPR experiments, which were analysed by K.F.P., M.B.A.K. and C.W.M.K.; A.L. completed the structure determination of the FimD–FimC–FimH complex, built and refined the structure. N.S.H., E.V., J.S.P. and B.F. cloned and purified the translocation domain of FimD, and cloned and analysed the FimD CTD mutants. S.G. participated in the building and refinement of the FimD–FimC–FimH structure and analysed the structure. J.Y. carried out the native mass spectrometry experiments on the FimD–FimC–FimH complex. C.W.M.K supervised the EPR work. H.L., S.J.H. and D.G.T. supervised the work on apo-FimD, analysed the structures, and wrote the paper. G.W. supervised the work on FimD–FimC–FimH, analysed the structures, and wrote the paper.

Corresponding authors

Correspondence to David G. Thanassi or Gabriel Waksman.

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The authors declare no competing financial interests.

Additional information

Structure factors and coordinates have been deposited in the Protein Data Bank (entry codes 3RFZ and 3OHN for coordinates and structure factors of the FimD–FimC–FimHcomplex and the translocation domain of FimD, respectively).

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Supplementary Information

The file contains Supplementary Tables 1-3, Supplementary Figures 1-10 with legends and additional references. (PDF 24878 kb)

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Phan, G., Remaut, H., Wang, T. et al. Crystal structure of the FimD usher bound to its cognate FimC–FimH substrate. Nature 474, 49–53 (2011).

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