Skip to main content
SpringerLink
Log in
Book cover

Cystic Fibrosis pp 321–327Cite as

Introduction to Section IV: Biophysical Methods to Approach CFTR Structure

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology ((MIMB,volume 741))

Abstract

Inefficient folding of CFTR into a functional three-dimensional structure is the basic pathophysiologic mechanism leading to most cases of cystic fibrosis. Knowledge of the structure of CFTR and placement of these mutations into a structural context would provide information key for developing targeted therapeutic approaches for cystic fibrosis. As a large polytopic membrane protein containing disordered regions, intact CFTR has been refractory to efforts to solve a high-resolution structure using X-ray crystallography. The following chapters summarize current efforts to circumvent these obstacles by utilizing NMR, electron microscopy, and computational methodologies and by development of experimental models of the relevant domains of CFTR.

This is a preview of subscription content, log in via an institution.

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Cutting, G. R. (1993) Spectrum of mutations in cystic fibrosis. J. Bioenerg. Biomembr. 25, 7–10.

    Article  PubMed  CAS  Google Scholar 

  2. Riordan, J. R., Rommens, J. M., Kerem, B., Alon, N., Rozmahel, R., Grzelczak, Z., et al. (1989) Identification of the cystic fibrosis gene: Cloning and characterization of complementary DNA. Science 245, 1066–1073.

    Article  PubMed  CAS  Google Scholar 

  3. Mendoza, J. L., and Thomas, P. J. (2007) Building an understanding of cystic fibrosis on the foundation of ABC transporter structures. J. Bioenerg. Biomembr. 39, 499–505.

    Article  PubMed  CAS  Google Scholar 

  4. Riordan, J. R. (2008) CFTR function and prospects for therapy. Annu. Rev. Biochem. 77, 701–726.

    Article  PubMed  CAS  Google Scholar 

  5. Choi, J. Y., Joo, N. S., Krouse, M. E., Wu, J. V., Robbins, R. C., Ianowski, J. P., et al. (2007) Synergistic airway gland mucus secretion in response to vasoactive intestinal peptide and carbachol is lost in cystic fibrosis. J. Clin. Invest. 117, 3118–3127.

    Article  PubMed  CAS  Google Scholar 

  6. Devor, D. C., Bridges, R. J., and Pilewski, J. M. (2000) Pharmacological modulation of ion transport across wild-type and DeltaF508 CFTR-expressing human bronchial epithelia. Am. J. Physiol. 279, C461–479.

    Google Scholar 

  7. Quinton, P. M., and Reddy, M. M. (1992) Control of CFTR chloride conductance by ATP levels through non-hydrolytic binding. Nature 360, 79–81.

    Article  PubMed  CAS  Google Scholar 

  8. Ko, S. B., Zeng, W., Dorwart, M. R., Luo, X., Kim, K. H., Millen, L., et al. (2004) Gating of CFTR by the STAS domain of SLC26 transporters. Nat. Cell. Biol. 6, 343–350.

    Article  PubMed  CAS  Google Scholar 

  9. Kunzelmann, K., Kiser, G. L., Schreiber, R., and Riordan, J. R. (1997) Inhibition of epithelial Na+ currents by intracellular domains of the cystic fibrosis transmembrane conductance regulator. FEBS Lett. 400, 341–344.

    Article  PubMed  CAS  Google Scholar 

  10. Mall, M., Bleich, M., Kuehr, J., Brandis, M., Greger, R., and Kunzelmann, K. (1999) CFTR-mediated inhibition of epithelial Na+ conductance in human colon is defective in cystic fibrosis. Am. J. Physiol. 277, G709–716.

    Google Scholar 

  11. Schreiber, R., Hopf, A., Mall, M., Greger, R., and Kunzelmann, K. (1999) The first-nucleotide binding domain of the cystic-fibrosis transmembrane conductance regulator is important for inhibition of the epithelial Na+ channel. Proc. Natl. Acad. Sci. USA 96, 5310–5315.

    Article  PubMed  CAS  Google Scholar 

  12. Linsdell, P., Zheng, S. X., and Hanrahan, J. W. (1998) Non-pore lining amino acid side chains influence anion selectivity of the human CFTR Cl– channel expressed in mammalian cell lines. J. Physiol. 512, 1–16.

    Article  PubMed  CAS  Google Scholar 

  13. McCarty, N. A. (2000) Permeation through the CFTR chloride channel. J. Exp. Biol. 203, 1947–1962.

    PubMed  CAS  Google Scholar 

  14. Sheppard, D. N., Rich, D. P., Ostedgaard, L. S., Gregory, R. J., Smith, A. E., and Welsh, M. J. (1993) Mutations in CFTR associated with mild-disease-form Cl– channels with altered pore properties. Nature 362, 160–164.

    Article  PubMed  CAS  Google Scholar 

  15. Fulmer, S. B., Schwiebert, E. M., Morales, M. M., Guggino, W. B., and Cutting, G. R. (1995) Two cystic fibrosis transmembrane conductance regulator mutations have different effects on both pulmonary phenotype and regulation of outwardly rectified chloride currents. Proc. Natl. Acad. Sci. USA 92, 6832–6836.

    Article  PubMed  CAS  Google Scholar 

  16. Qu, B. H., Strickland, E., and Thomas, P. J. (1997) Cystic fibrosis: A disease of altered protein folding. J. Bioenerg. Biomembr. 29, 483–490.

    Article  PubMed  CAS  Google Scholar 

  17. Thomas, P. J., Ko, Y. H., and Pedersen, P. L. (1992) Altered protein folding may be the molecular basis of most cases of cystic fibrosis. FEBS Lett. 312, 7–9.

    Article  PubMed  CAS  Google Scholar 

  18. Gregory, R. J., Rich, D. P., Cheng, S. H., Souza, D. W., Paul, S., Manavalan, P., et al. (1991) Maturation and function of cystic fibrosis transmembrane conductance regulator variants bearing mutations in putative nucleotide-binding domains 1 and 2. Mol. Cell. Biol. 11, 3886–3893.

    PubMed  CAS  Google Scholar 

  19. Hutt, D. M., Herman, D., Rodrigues, A. P., Noel, S., Pilewski, J. M., Matteson, J., et al. (2010) Reduced histone deacetylase 7 activity restores function to misfolded CFTR in cystic fibrosis. Nat. Chem. Biol. 6, 25–33.

    Article  PubMed  CAS  Google Scholar 

  20. Wang, X., Venable, J., LaPointe, P., Hutt, D. M., Koulov, A. V., Coppinger, J., et al. (2006) Hsp90 cochaperone Aha1 downregulation rescues misfolding of CFTR in cystic fibrosis. Cell 127, 803–815.

    Article  PubMed  CAS  Google Scholar 

  21. Younger, J. M., Fan, C. Y., Chen, L., Rosser, M. F., Patterson, C., and Cyr, D. M. (2005) Cystic fibrosis transmembrane conductance regulator as a model substrate to study endoplasmic reticulum protein quality control in mammalian cells. Methods Mol. Biol. 301, 293–303.

    PubMed  CAS  Google Scholar 

  22. Awayn, N. H., Rosenberg, M. F., Kamis, A. B., Aleksandrov, L. A., Riordan, J. R., and Ford, R. C. (2005) Crystallographic and single-particle analyses of native- and nucleotide-bound forms of the cystic fibrosis transmembrane conductance regulator (CFTR) protein. Biochem. Soc. Trans. 33, 996–999.

    Article  PubMed  CAS  Google Scholar 

  23. Kleizen, B., van Vlijmen, T., de Jonge, H. R., and Braakman, I. (2005) Folding of CFTR is predominantly cotranslational. Mol. Cell 20, 277–287.

    Article  PubMed  CAS  Google Scholar 

  24. Cui, L., Aleksandrov, L., Chang, X. B., Hou, Y. X., He, L., Hegedus, T., et al. (2007) Domain interdependence in the biosynthetic assembly of CFTR. J. Mol. Biol. 365, 981–994.

    Article  PubMed  CAS  Google Scholar 

  25. Zhang, L., Aleksandrov, L. A., Zhao, Z., Birtley, J. R., Riordan, J. R., and Ford, R. C. (2009) Architecture of the cystic fibrosis transmembrane conductance regulator protein and structural changes associated with phosphorylation and nucleotide binding. J. Struct. Biol. 167, 242–251.

    Article  PubMed  CAS  Google Scholar 

  26. Rosenberg, M. F., Kamis, A. B., Aleksandrov, L. A., Ford, R. C., and Riordan, J. R. (2004) Purification and crystallization of the cystic fibrosis transmembrane conductance regulator (CFTR). J. Biol. Chem. 279, 39051–39057.

    Article  PubMed  CAS  Google Scholar 

  27. Aller, S. G., Yu, J., Ward, A., Weng, Y., Chittaboina, S., Zhuo, R., et al. (2009) Structure of P-glycoprotein reveals a molecular basis for poly-specific drug binding. Science 323, 1718–1722.

    Article  PubMed  CAS  Google Scholar 

  28. Dawson, R. J., and Locher, K. P. (2006) Structure of a bacterial multidrug ABC transporter. Nature 443, 180–185.

    Article  PubMed  CAS  Google Scholar 

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

    Article  PubMed  CAS  Google Scholar 

  30. Huang, S. Y., Bolser, D., Liu, H. Y., Hwang, T. C., and Zou, X. (2009) Molecular modeling of the heterodimer of human CFTR’s nucleotide-binding domains using a protein-protein docking approach. J. Mol. Graph. 27, 822–828.

    Article  CAS  Google Scholar 

  31. Moran, O. (2007) Model of the cAMP activation of chloride transport by CFTR channel and the mechanism of potentiators. J. Theor. Biol. 262, 73–79.

    Article  Google Scholar 

  32. Mornon, J. P., Lehn, P., and Callebaut, I. (2009) Molecular models of the open and closed states of the whole human CFTR protein. Cell. Mol. Life Sci. 66, 3469–3486.

    Article  PubMed  CAS  Google Scholar 

  33. Serohijos, A. W., Hegedus, T., Aleksandrov, A. A., He, L., Cui, L., Dokholyan, N. V., et al. (2008) Phenylalanine-508 mediates a cytoplasmic-membrane domain contact in the CFTR 3D structure crucial to assembly and channel function. Proc. Natl. Acad. Sci. USA 105, 3256–3261.

    Article  PubMed  CAS  Google Scholar 

  34. Atwell, S., Brouillette, C. G., Conners, K., Emtage, S., Gheyi, T., Guggino, W. B., et al. (2010) Structures of a minimal human CFTR first nucleotide-binding domain as a monomer, head-to-tail homodimer, and pathogenic mutant. Protein Eng. Des. Sel. 23, 375–384.

    Article  PubMed  CAS  Google Scholar 

  35. Lewis, H. A., Buchanan, S. G., Burley, S. K., Conners, K., Dickey, M., Dorwart, M., et al. (2004) Structure of nucleotide-binding domain 1 of the cystic fibrosis transmembrane conductance regulator. EMBO J. 23, 282–293.

    Article  PubMed  CAS  Google Scholar 

  36. Lewis, H. A., Wang, C., Zhao, X., Hamuro, Y., Conners, K., Kearins, M. C., et al. (2010) Structure and dynamics of NBD1 from CFTR characterized using crystallography and hydrogen/deuterium exchange mass spectrometry. J. Mol. Biol. 396, 406–430.

    Article  PubMed  CAS  Google Scholar 

  37. Thibodeau, P. H., Brautigam, C. A., Machius, M., and Thomas, P. J. (2005) Side chain and backbone contributions of Phe508 to CFTR folding. Nat. Struct. Mol. Biol. 12, 10–16.

    Article  PubMed  CAS  Google Scholar 

  38. Baker, J. M., Hudson, R. P., Kanelis, V., Choy, W. Y., Thibodeau, P. H., Thomas, P. J., et al. (2007) CFTR regulatory region interacts with NBD1 predominantly via multiple transient helices. Nat. Struct. Mol. Biol. 14, 738–745.

    Article  PubMed  CAS  Google Scholar 

  39. Kanelis, V., Hudson, R. P., Thibodeau, P. H., Thomas, P. J., and Forman-Kay, J. D. (2010) NMR evidence for differential phosphorylation-dependent interactions in WT and DeltaF508 CFTR. EMBO J. 29, 263–277.

    Article  PubMed  CAS  Google Scholar 

  40. Brown, C. R., Hong-Brown, L. Q., Biwersi, J., Verkman, A. S., and Welch, W. J. (1996) Chemical chaperones correct the mutant phenotype of the delta F508 cystic fibrosis transmembrane conductance regulator protein. Cell Stress Chaperones 1, 117–125.

    Article  PubMed  CAS  Google Scholar 

  41. Denning, G. M., Anderson, M. P., Amara, J. F., Marshall, J., Smith, A. E., and Welsh, M. J. (1992) Processing of mutant cystic fibrosis transmembrane conductance regulator is temperature-sensitive. Nature 358, 761–764.

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The authors are grateful for the support provided by the NIH-NIDDK, Welch Foundation, and the CF Foundation for much of the work summarized. Also we would like to thank the many investigators that have contributed to these studies and our thinking regarding the utility of structural approaches including the authors of Chapters 22 , http://23 , and http://25 , the members of the CFTR folding consortium, the SGX-CFFT joint research committee, Chad Brautigam, Hanoch Senderowitz, Martin Mense, and former members of the laboratory at UT Southwestern, including Bao-He Qu, Elizabeth Strickland, Michael Dorwart, Patrick Thibodeau, John Richardson, Jarod Watson, and Emmanuel Caspa.

Author information

Authors and Affiliations

  1. Department of Physiology, University of Texas Southwestern Medical Center, Dallas, TX, 75390-9040, USA

    Juan L. Mendoza, André Schmidt & Philip J. Thomas

Authors
  1. Juan L. Mendoza
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. André Schmidt
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Philip J. Thomas
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Faculty of Sciences, Centre for Biodiversity & Functional, University of Lisboa, Campo Grande, Lisboa, 1749-016, Portugal

    Margarida D. Amaral

  2. , Department of Physiology, University of Regensburg, Universitätsstr. 31, Regensburg, 93053, Germany

    Karl Kunzelmann

Rights and permissions

Reprints and permissions

Copyright information

© 2011 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Mendoza, J.L., Schmidt, A., Thomas, P.J. (2011). Introduction to Section IV: Biophysical Methods to Approach CFTR Structure. In: Amaral, M., Kunzelmann, K. (eds) Cystic Fibrosis. Methods in Molecular Biology, vol 741. Humana Press. https://doi.org/10.1007/978-1-61779-117-8_21

Download citation

  • DOI: https://doi.org/10.1007/978-1-61779-117-8_21

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-116-1

  • Online ISBN: 978-1-61779-117-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation