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:

A distinct mechanism for the ABC transporter BtuCD–BtuF revealed by the dynamics of complex formation

Nature Structural & Molecular Biology volume 17, pages 332–338 (2010)Cite this article

Subjects

Abstract

ATP-binding cassette (ABC) transporters are integral membrane proteins that translocate a diverse array of substrates across cell membranes. We present here the dynamics of complex formation of three structurally characterized ABC transporters—the BtuCD vitamin B12 importer and MetNI d/l-methionine importer from Escherichia coli and the Hi1470/1 metal-chelate importer from Haemophilus influenzae—in complex with their cognate binding proteins. Similarly to other ABC importers, MetNI interacts with its binding protein with low affinity (Kd ∼10−4 M). In contrast, BtuCD–BtuF and Hi1470/1–Hi1472 form stable, high-affinity complexes (Kd ∼10−13 and 10−9 M, respectively). In BtuCD–BtuF, vitamin B12 accelerates the complex dissociation rate ∼107-fold, with ATP having an additional destabilizing effect. The findings presented here highlight substantial mechanistic differences between BtuCD–BtuF, and likely Hi1470/1–Hi1472, and the better-characterized maltose and related ABC transport systems, indicating that there is considerable mechanistic diversity within this large protein super-family.

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: Complex formation between ABC transporters and their respective binding proteins.
Figure 2: Dynamics of complex formation.
Figure 3: Substrate binding by components of the vitamin B12 transport system.
Figure 4: Substrate release from the vitamin B12 transport system.
Figure 5: Substrate effects on complex formation in the vitamin B12 transport system.
Figure 6: Effects of nucleotide binding and hydrolysis on complex formation in the vitamin B12 transport system.
Figure 7: Thermodynamics of the BtuCD-F transport cycle and comparison to the maltose system.

Similar content being viewed by others

References

  1. Higgins, C.F. ABC transporters: from microorganisms to man. Annu. Rev. Cell Biol. 8, 67–113 (1992).

    Article  CAS  Google Scholar 

  2. Higgins, C.F. ABC transporters: physiology, structure and mechanism–an overview. Res. Microbiol. 152, 205–210 (2001).

    Article  CAS  Google Scholar 

  3. van der Does, C. & Tampe, R. How do ABC transporters drive transport? Biol. Chem. 385, 927–933 (2004).

    Article  CAS  Google Scholar 

  4. Davidson, A.L., Dassa, E., Orelle, C. & Chen, J. Structure, function, and evolution of bacterial ATP-binding cassette systems. Microbiol. Mol. Biol. Rev. 72, 317–364 (2008).

    Article  CAS  Google Scholar 

  5. Locher, K.P. Review. Structure and mechanism of ATP-binding cassette transporters. Phil. Trans. R. Soc. Lond. B 364, 239–245 (2009).

    Article  CAS  Google Scholar 

  6. Jones, P.M., O'Mara, M.L. & George, A.M. ABC transporters: a riddle wrapped in a mystery inside an enigma. Trends Biochem. Sci. 34, 520–531 (2009).

    Article  CAS  Google Scholar 

  7. Boos, W. & Lucht, J.M. Periplasmic binding protein-dependent ABC transporters. in Escherichia coli and Salmonella typhimurium: Cellular and Molecular Biology Vol. 1 (ed. Neidhardt, F.C.) 1175–1209 (American Society for Microbiology, Washington, DC, 1996).

  8. Rohrbach, M.R., Braun, V. & Koster, W. Ferrichrome transport in Escherichia coli K-12: altered substrate specificity of mutated periplasmic FhuD and interaction of FhuD with the integral membrane protein FhuB. J. Bacteriol. 177, 7186–7193 (1995).

    Article  CAS  Google Scholar 

  9. Doeven, M.K., Abele, R., Tampe, R. & Poolman, B. The binding specificity of OppA determines the selectivity of the oligopeptide ATP-binding cassette transporter. J. Biol. Chem. 279, 32301–32307 (2004).

    Article  CAS  Google Scholar 

  10. Orelle, C., Ayvaz, T., Everly, R.M., Klug, C.S. & Davidson, A.L. Both maltose-binding protein and ATP are required for nucleotide-binding domain closure in the intact maltose ABC transporter. Proc. Natl. Acad. Sci. USA 105, 12837–12842 (2008).

    Article  CAS  Google Scholar 

  11. Oldham, M.L., Khare, D., Quiocho, F.A., Davidson, A.L. & Chen, J. Crystal structure of a catalytic intermediate of the maltose transporter. Nature 450, 515–521 (2007).

    Article  CAS  Google Scholar 

  12. Bassford, P.J. Jr., Bradbeer, C., Kadner, R.J. & Schnaitman, C.A. Transport of vitamin B12 in tonB mutants of Escherichia coli. J. Bacteriol. 128, 242–247 (1976).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Bassford, P.J. Jr. & Kadner, R.J. Genetic analysis of components involved in vitamin B12 uptake in Escherichia coli. J. Bacteriol. 132, 796–805 (1977).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Cadieux, N. et al. Identification of the periplasmic cobalamin-binding protein BtuF of Escherichia coli. J. Bacteriol. 184, 706–717 (2002).

    Article  CAS  Google Scholar 

  15. Borths, E.L., Locher, K.P., Lee, A.T. & Rees, D.C. The structure of Escherichia coli BtuF and binding to its cognate ATP binding cassette transporter. Proc. Natl. Acad. Sci. USA 99, 16642–16647 (2002).

    Article  CAS  Google Scholar 

  16. Hvorup, R.N. et al. Asymmetry in the structure of the ABC transporter binding protein complex BtuCD-BtuF. Science 317, 1387–1390 (2007).

    Article  CAS  Google Scholar 

  17. Goetz, B.A., Perozo, E. & Locher, K.P. Distinct gate conformations of the ABC transporter BtuCD revealed by electron spin resonance spectroscopy and chemical cross-linking. FEBS Lett. 583, 266–270 (2009).

    Article  CAS  Google Scholar 

  18. Dassa, E. & Bouige, E. The ABC of ABCs: a phylogenetic and functional classification of ABC systems in living organisms. Res. Microbiol. 152, 211–229 (2001).

    Article  CAS  Google Scholar 

  19. Rees, D.C., Johnson, E. & Lewinson, O. ABC transporters: the power to change. Nat. Rev. Mol. Cell Biol. 10, 218–227 (2009).

    Article  CAS  Google Scholar 

  20. Borths, E.L., Poolman, B., Hvorup, R.N., Locher, K.P. & Rees, D.C. In vitro functional characterization of BtuCD-F, the Escherichia coli ABC transporters for vitamin B12 uptake. Biochemistry 44, 16301–16309 (2005).

    Article  CAS  Google Scholar 

  21. Chen, J., Sharma, S., Quiocho, F.A. & Davidson, A.L. Trapping the transition state of an ATP-binding cassette transporter: evidence for a concerted mechanism of maltose transport. Proc. Natl. Acad. Sci. USA 98, 1525–1530 (2001).

    Article  CAS  Google Scholar 

  22. Liu, C.E. & Ames, G.F. Characterization of transport through the periplasmic histidine permease using proteoliposomes reconstituted by dialysis. J. Biol. Chem. 272, 859–866 (1997).

    Article  CAS  Google Scholar 

  23. Merino, G., Boos, W., Shuman, H.A. & Bohl, E. The inhibition of maltose transport by the unliganded form of the maltose-binding protein of Escherichia coli: experimental findings and mathematical treatment. J. Theor. Biol. 177, 171–179 (1995).

    Article  CAS  Google Scholar 

  24. Ames, G.F., Liu, C.E., Joshi, A.K. & Nikaido, K. Liganded and unliganded receptors interact with equal affinity with the membrane complex of periplasmic permeases, a subfamily of traffic ATPases. J. Biol. Chem. 271, 14264–14270 (1996).

    Article  CAS  Google Scholar 

  25. van der Does, C., Presenti, C., Schulze, K., Dinkelaker, S. & Tampe, R. Kinetics of the ATP hydrolysis cycle of the nucleotide-binding domain of Mdl1 studied by a novel site-specific labeling technique. J. Biol. Chem. 281, 5694–5701 (2006).

    Article  CAS  Google Scholar 

  26. Pick, U. The interaction of vanadate ions with the Ca-ATPase from sarcoplasmic reticulum. J. Biol. Chem. 257, 6111–6119 (1982).

    CAS  PubMed  Google Scholar 

  27. Austermuhle, M.I., Hall, J.A., Klug, C.S. & Davidson, A.L. Maltose-binding protein is open in the catalytic transition state for ATP hydrolysis during maltose transport. J. Biol. Chem. 279, 28243–28250 (2004).

    Article  CAS  Google Scholar 

  28. James, K.J., Hancock, M.A., Gagnon, J.N. & Coulton, J.W. TonB interacts with BtuF, the Escherichia coli periplasmic binding protein for cyanocobalamin. Biochemistry 48, 9212–9220 (2009).

    Article  CAS  Google Scholar 

  29. Lundrigan, M.D., Koster, W. & Kadner, R.J. Transcribed sequences of the Escherichia coli btuB gene control its expression and regulation by vitamin B12 . Proc. Natl. Acad. Sci. USA 88, 1479–1483 (1991).

    Article  CAS  Google Scholar 

  30. Richter-Dahlfors, A.A., Ravnum, S. & Andersson, D.I. Vitamin B12 repression of the cob operon in Salmonella typhimurium: translational control of the cbiA gene. Mol. Microbiol. 13, 541–553 (1994).

    Article  CAS  Google Scholar 

  31. Van Hove, B., Staudenmaier, H. & Braun, V. Novel two-component transmembrane transcription control: regulation of iron dicitrate transport in Escherichia coli K-12. J. Bacteriol. 172, 6749–6758 (1990).

    Article  CAS  Google Scholar 

  32. Khare, D., Oldham, M.L., Orelle, C., Davidson, A.L. & Chen, J. Alternating access in maltose transporter mediated by rigid-body rotations. Mol. Cell 33, 528–536 (2009).

    Article  CAS  Google Scholar 

  33. Gorbulev, S., Abele, R. & Tampe, R. Allosteric crosstalk between peptide-binding, transport, and ATP hydrolysis of the ABC transporter TAP. Proc. Natl. Acad. Sci. USA 98, 3732–3737 (2001).

    Article  CAS  Google Scholar 

  34. Herget, M. et al. Purification and reconstitution of the antigen transport complex TAP: A prerequisite for determination of peptide stoichiometry and ATP hydrolysis. J. Biol. Chem. 284, 33740–33749 (2009).

    Article  CAS  Google Scholar 

  35. Chang, X.B., Hou, Y.X. & Riordan, J.R. ATPase activity of purified multidrug resistance-associated protein. J. Biol. Chem. 272, 30962–30968 (1997).

    Article  CAS  Google Scholar 

  36. Ozvegy, C., Varadi, A. & Sarkadi, B. Characterization of drug transport, ATP hydrolysis, and nucleotide trapping by the human ABCG2 multidrug transporter. Modulation of substrate specificity by a point mutation. J. Biol. Chem. 277, 47980–47990 (2002).

    Article  CAS  Google Scholar 

  37. Sauna, Z.E., Nandigama, K. & Ambudkar, S.V. Multidrug resistance protein 4 (ABCC4)-mediated ATP hydrolysis: effect of transport substrates and characterization of the post-hydrolysis transition state. J. Biol. Chem. 279, 48855–48864 (2004).

    Article  CAS  Google Scholar 

  38. Quiocho, F.A. & Ledvina, P.S. Atomic structure and specificity of bacterial periplasmic receptors for active transport and chemotaxis: variation of common themes. Mol. Microbiol. 20, 17–25 (1996).

    Article  CAS  Google Scholar 

  39. Eakanunkul, S. et al. Characterization of the periplasmic heme-binding protein shut from the heme uptake system of Shigella dysenteriae. Biochemistry 44, 13179–13191 (2005).

    Article  CAS  Google Scholar 

  40. Ho, W.W. et al. Holo- and apo-bound structures of bacterial periplasmic heme-binding proteins. J. Biol. Chem. 282, 35796–35802 (2007).

    Article  CAS  Google Scholar 

  41. Clarke, T.E., Braun, V., Winkelmann, G., Tari, L.W. & Vogel, H.J. X-ray crystallographic structures of the Escherichia coli periplasmic protein FhuD bound to hydroxamate-type siderophores and the antibiotic albomycin. J. Biol. Chem. 277, 13966–13972 (2002).

    Article  CAS  Google Scholar 

  42. Karpowich, N.K., Huang, H.H., Smith, P.C. & Hunt, J.F. Crystal structures of the BtuF periplasmic-binding protein for vitamin B12 suggest a functionally important reduction in protein mobility upon ligand binding. J. Biol. Chem. 278, 8429–8434 (2003).

    Article  CAS  Google Scholar 

  43. Locher, K.P., Lee, A.T. & Rees, D.C. The E. coli BtuCD structure: a framework for ABC transporter architecture and mechanism. Science 296, 1091–1098 (2002).

    Article  CAS  Google Scholar 

  44. Pinkett, H.W., Lee, A.T., Lum, P., Locher, K.P. & Rees, D.C. An inward-facing conformation of a putative metal-chelate type ABC transporter. Science 315, 373–377 (2007).

    Article  CAS  Google Scholar 

  45. Kadaba, N.S., Kaiser, J.T., Johnson, E., Lee, A. & Rees, D.C. The high-affinity E. coli methionine ABC transporter: structure and allosteric regulation. Science 321, 250–253 (2008).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank H. Pinkett for insightful discussions and critical reading of the manuscript and J. Klein and J. Vielmetter for their help in the initial BiaCore experiments. The work was supported in part by US National Institutes of Health grant GM045162, by the Howard Hughes Medical Institute and by fellowships to O.L. from the Fulbright Foundation and the Jane Coffin Childs Memorial Fund for Medical Research.

Author information

Authors and Affiliations

  1. Division of Chemistry and Chemical Engineering, Pasadena, California, USA

    Oded Lewinson, Allen T Lee & Douglas C Rees

  2. Howard Hughes Medical Institute, California Institute of Technology, Pasadena, California, USA

    Allen T Lee & Douglas C Rees

  3. Institute of Molecular Biology and Biophysics, ETH Zurich, Zurich, Switzerland

    Kaspar P Locher

Authors
  1. Oded Lewinson
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Allen T Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kaspar P Locher
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Douglas C Rees
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

A.T.L. and K.P.L. generated the original constructs used in this work; O.L. and D.C.R. designed the research; O.L. and A.T.L. performed the research; O.L., A.T.L., K.P.L. and D.C.R. analyzed the data; and O.L., K.P.L. and D.C.R. wrote the paper.

Corresponding author

Correspondence to Douglas C Rees.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1 and 2 and Supplementary Methods (PDF 232 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lewinson, O., Lee, A., Locher, K. et al. A distinct mechanism for the ABC transporter BtuCD–BtuF revealed by the dynamics of complex formation. Nat Struct Mol Biol 17, 332–338 (2010). https://doi.org/10.1038/nsmb.1770

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.1770

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