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Using 2D Crystals to Analyze the Structure of Membrane Proteins

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Book cover Membrane Biogenesis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1033))

Abstract

Electron crystallography is a powerful technique for studying the structure and function of membrane proteins, not only in the ground state, but also in active conformations. When combined with high-resolution structures obtained by X-ray crystallography, electron crystallography can provide insights into the mechanism of the protein. In this chapter we discuss obtaining a three-dimensional map of membrane proteins by electron crystallography and how to combine these maps with atomic resolution models in order to study the function of membrane proteins. We argue that this approach is particularly powerful as it combines the high resolution attainable by X-ray crystallography with the visualization of the subject in the near-native environment of the membrane, by electron cryo-microscopy. This point has been illustrated by the analysis of the protein translocation complex SecYEG.

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References

  1. Henderson R, Unwin PNT (1975) Three-dimensional model of purple membrane obtained by electron microscopy. Nature 257: 28–32

    Article  PubMed  CAS  Google Scholar 

  2. Henderson R, Baldwin JM, Ceska TA, Zemlin F, Beckmann E, Downing KH (1990) A model for the structure of bacteriorhodopsin based on high resolution electron cryomicroscopy. J Mol Biol 213:899–929

    Article  PubMed  CAS  Google Scholar 

  3. Subramaniam S, Henderson R (2000) Molecular mechanism of vectorial proton translocation by bacteriorhodopsin. Nature 406:653–657

    Article  PubMed  CAS  Google Scholar 

  4. Vonck J (2000) Structure of the bacteriorhodopsin mutant F219L N-intermediate revealed by electron crystallography. EMBO J 19: 2152–2160

    Article  PubMed  CAS  Google Scholar 

  5. Kühlbrandt W, Wang DN, Fujiyoshi Y (1994) Atomic model of plant light-harvesting complex by electron crystallography. Nature 367:614–621

    Article  PubMed  Google Scholar 

  6. Cheng A, van Hoek AN, Yeager M, Verkman AS, Mitra AK (1997) Three-dimensional organization of a human water channel. Nature 387:627–630

    Article  PubMed  CAS  Google Scholar 

  7. Li H, Lee S, Jap BK (1997) Molecular design of aquaporin-1 water channel as revealed by electron crystallography. Nat Struct Biol 4:263–265

    Article  PubMed  CAS  Google Scholar 

  8. Walz T, Hirai T, Murata K, Heymann JB, Mitsuoka K, Fujiyoshi Y, Smith BL, Agre P, Engel A (1997) The three-dimensional structure of aquaporin-1. Nature 387:624–627

    Article  PubMed  CAS  Google Scholar 

  9. Gonen T, Cheng Y, Sliz P, Hiroaki Y, Fujiyoshi Y, Harrison SC, Walz T (2005) Lipid-protein interactions in double-layered two-dimensional AQP0 crystals. Nature 438:633–638

    Article  PubMed  CAS  Google Scholar 

  10. van den Berg B, Clemons WM Jr, Collinson I, Modis Y, Hartmann E, Harrison SC, Rapoport TA (2004) X-ray structure of a protein-conducting channel. Nature 427:36–44

    Article  PubMed  Google Scholar 

  11. Breyton C, Haase W, Rapoport TA, Kühlbrandt W, Collinson I (2002) Three-dimensional structure of the bacterial protein-translocation complex SecYEG. Nature 418:662–664

    Article  PubMed  CAS  Google Scholar 

  12. Hizlan D, Robson A, Whitehouse S, Gold VA, Vonck J, Mills D, Kühlbrandt W, Collinson I (2012) Structure of the SecY complex unlocked by a preprotein mimic. Cell Rep 1: 21–28

    Article  PubMed  CAS  Google Scholar 

  13. Gipson B, Zeng X, Stahlberg H (2007) 2dx_merge: Data management and merging for 2D crystal images. J Struct Biol 160:375–384

    Article  PubMed  CAS  Google Scholar 

  14. Gipson B, Zeng X, Zhang Z, Stahlberg H (2007) 2dx—User-friendly image processing for 2D crystals. J Struct Biol 157:64–72

    Article  PubMed  CAS  Google Scholar 

  15. Amos LA, Henderson R, Unwin PN (1982) Three-dimensional structure determination by electron microscopy of two-dimensional crystals. Prog Biophys Mol Biol 39:183–231

    Article  PubMed  CAS  Google Scholar 

  16. Henderson R, Baldwin JM, Downing K, Lepault J, Zemlin F (1986) Structure of purple membrane from Halobacterium halobium: recording, measurement and evaluation of electron micrographs at 3.5Å resolution. Ultramicroscopy 19:147–178

    Article  CAS  Google Scholar 

  17. Crowther RA, Henderson R, Smith JM (1996) MRC image processing programs. J Struct Biol 116:9–16

    Article  PubMed  CAS  Google Scholar 

  18. Collaborative Computational Project, N (1994) The CCP4 suite: programs for protein crystallography. Acta Crystallogr D Biol Crystallogr 50:760–763

    Article  Google Scholar 

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

    Article  PubMed  Google Scholar 

  20. Collinson I, Breyton C, Duong F, Tziatzios C, Schubert D, Or E, Rapoport T, Kühlbrandt W (2001) Projection structure and oligomeric properties of a bacterial core protein translocase. EMBO J 20:2462–2471

    Article  PubMed  CAS  Google Scholar 

  21. Glaeser RM, Downing KH (1990) The “specimen flatness” problem in high-resolution electron crystallography of biological macromolecules. In: Peachy LD, Williams DB (eds) XIIth international congress for electron microscopy. San Francisco Press, Inc., Seattle, pp 98–99

    Google Scholar 

  22. Vonck J (2000) Parameters affecting specimen flatness of two-dimensional crystals for electron crystallography. Ultramicroscopy 85:123–129

    Article  PubMed  CAS  Google Scholar 

  23. Downing KH (1991) Spot-scan imaging in transmission electron microscopy. Science 251:53–59

    Article  PubMed  CAS  Google Scholar 

  24. Zeng X, Gipson B, Zheng ZY, Renault L, Stahlberg H (2007) Automatic lattice determination for two-dimensional crystal images. J Struct Biol 160:353–361

    Article  PubMed  CAS  Google Scholar 

  25. Unger VM, Schertler GFX (1995) Low resolution structure of bovine rhodopsin determined by electron cryo-microscopy. Biophys J 68:1776–1786

    Article  PubMed  CAS  Google Scholar 

  26. Williams KA (2000) Three-dimensional structure of the ion-coupled transport protein NhaA. Nature 403:112–115

    Article  PubMed  CAS  Google Scholar 

  27. Vonck J, Krug von Nidda T, Meier T, Matthey U, Mills DJ, Kühlbrandt W, Dimroth P (2002) Molecular architecture of the undecameric rotor of a bacterial Na+-ATP synthase. J Mol Biol 321:307–316

    Article  PubMed  CAS  Google Scholar 

  28. Bostina M, Mohsin B, Kühlbrandt W, Collinson I (2005) Atomic model of the E. coli membrane-bound protein translocation complex SecYEG. J Mol Biol 352:1035–1043

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

We would like to thank Prof. Dr. Werner Kühlbrandt for his support and Dr. Özkan Yildiz for his help with computer programs.

Author information

Authors and Affiliations

  1. School of Biochemistry, University of Bristol, Bristol, UK

    Ian Collinson

  2. Department of Structural Biology, Max Planck Institute of Biophysics, Frankfurt, Germany

    Janet Vonck

  3. Istanbul Sehir University, Istanbul, Turkey

    Dilem Hizlan

Authors
  1. Ian Collinson
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  2. Janet Vonck
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  3. Dilem Hizlan
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Editor information

Editors and Affiliations

  1. Interfaculty Institute of Biochemistry, University of Tuebingen, Tuebingen, Germany

    Doron Rapaport

  2. Cell Biology, University of Kaiserslautern, Kaiserslautern, Germany

    Johannes M. Herrmann

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Collinson, I., Vonck, J., Hizlan, D. (2013). Using 2D Crystals to Analyze the Structure of Membrane Proteins. In: Rapaport, D., Herrmann, J. (eds) Membrane Biogenesis. Methods in Molecular Biology, vol 1033. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-487-6_4

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  • DOI: https://doi.org/10.1007/978-1-62703-487-6_4

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-486-9

  • Online ISBN: 978-1-62703-487-6

  • eBook Packages: Springer Protocols

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