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α-Helical nascent polypeptide chains visualized within distinct regions of the ribosomal exit tunnel

Nature Structural & Molecular Biology volume 17, pages 313–317 (2010)Cite this article

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Abstract

As translation proceeds, the nascent polypeptide chain passes through a tunnel in the large ribosomal subunit. Although this ribosomal exit tunnel was once thought only to be a passive conduit for the growing nascent chain, accumulating evidence suggests that it may in fact play a more active role in regulating translation and initial protein folding events. Here we have determined single-particle cryo–electron microscopy reconstructions of eukaryotic 80S ribosomes containing nascent chains with high α-helical propensity located within the exit tunnel. The maps enable direct visualization of density for helices as well as allowing the sites of interaction with the tunnel wall components to be elucidated. In particular regions of the tunnel, the nascent chain adopts distinct conformations and establishes specific contacts with tunnel components, both ribosomal RNA and proteins, that have been previously implicated in nascent chain–ribosome interaction.

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Figure 1: Cryo–electron microscopy reconstructions of RNCs.
Figure 2: Comparison of tRNA–nascent chains from RNCs.
Figure 3: Helix–nascent chain interactions with tunnel components.
Figure 4: Implications of helix formation within the ribosomal exit tunnel.

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References

  1. Frank, J. et al. A model of protein synthesis based on cryo–electron microscopy of the E. coli ribosome. Nature 376, 441–444 (1995).

    Article  CAS  Google Scholar 

  2. Ban, N., Nissen, P., Hansen, J., Moore, P.B. & Steitz, T.A. The complete atomic structure of the large ribosomal subunit at 2.4 Å resolution. Science 289, 905–920 (2000).

    Article  CAS  Google Scholar 

  3. Morgan, D.G. et al. A comparison of the yeast and rabbit 80 S ribosome reveals the topology of the nascent chain exit tunnel, inter-subunit bridges and mammalian rRNA expansion segments. J. Mol. Biol. 301, 301–321 (2000).

    Article  CAS  Google Scholar 

  4. Halic, M. et al. Signal recognition particle receptor exposes the ribosomal translocon binding site. Science 312, 745–747 (2006).

    Article  CAS  Google Scholar 

  5. Lu, J. & Deutsch, C. Electrostatics in the ribosomal tunnel modulate chain elongation rates. J. Mol. Biol. 384, 73–86 (2008).

    Article  CAS  Google Scholar 

  6. Lu, J., Kobertz, W.R. & Deutsch, C. Mapping the electrostatic potential within the ribosomal exit tunnel. J. Mol. Biol. 371, 1378–1391 (2007).

    Article  CAS  Google Scholar 

  7. Tenson, T. & Ehrenberg, M. Regulatory nascent peptides in the ribosomal tunnel. Cell 108, 591–594 (2002).

    Article  CAS  Google Scholar 

  8. Gong, F. & Yanofsky, C. Instruction of translating ribosome by nascent peptide. Science 297, 1864–1867 (2002).

    Article  CAS  Google Scholar 

  9. Nakatogawa, H. & Ito, K. The ribosomal exit tunnel functions as a discriminating gate. Cell 108, 629–636 (2002).

    Article  CAS  Google Scholar 

  10. Vazquez-Laslop, N., Thum, C. & Mankin, A.S. Molecular mechanism of drug-dependent ribosome stalling. Mol. Cell 30, 190–202 (2008).

    Article  CAS  Google Scholar 

  11. Voss, N.R., Gerstein, M., Steitz, T.A. & Moore, P.B. The geometry of the ribosomal polypeptide exit tunnel. J. Mol. Biol. 360, 893–906 (2006).

    Article  CAS  Google Scholar 

  12. Hardesty, B. & Kramer, G. Folding of a nascent peptide on the ribosome. Prog. Nucleic Acid Res. Mol. Biol. 66, 41–66 (2001).

    Article  CAS  Google Scholar 

  13. Woolhead, C.A., McCormick, P.J. & Johnson, A.E. Nascent membrane and secretory proteins differ in FRET-detected folding far inside the ribosome and in their exposure to ribosomal proteins. Cell 116, 725–736 (2004).

    Article  CAS  Google Scholar 

  14. Kosolapov, A., Tu, L., Wang, J. & Deutsch, C. Structure acquisition of the T1 domain of Kv1.3 during biogenesis. Neuron 44, 295–307 (2004).

    Article  CAS  Google Scholar 

  15. Lu, J. & Deutsch, C. Folding zones inside the ribosomal exit tunnel. Nat. Struct. Mol. Biol. 12, 1123–1129 (2005).

    Article  CAS  Google Scholar 

  16. Lu, J. & Deutsch, C. Secondary structure formation of a transmembrane segment in Kv channels. Biochemistry 44, 8230–8243 (2005).

    Article  CAS  Google Scholar 

  17. Woolhead, C.A., Johnson, A.E. & Bernstein, H.D. Translation arrest requires two-way communication between a nascent polypeptide and the ribosome. Mol. Cell 22, 587–598 (2006).

    Article  CAS  Google Scholar 

  18. Halic, M. et al. Structure of the signal recognition particle interacting with the elongation-arrested ribosome. Nature 427, 808–814 (2004).

    Article  CAS  Google Scholar 

  19. Halic, M. et al. Following the signal sequence from ribosomal tunnel exit to signal recognition particle. Nature 444, 507–511 (2006).

    Article  CAS  Google Scholar 

  20. Marqusee, S. & Baldwin, R.L. Helix stabilization by Glu−.Lys+ salt bridges in short peptides of de novo design. Proc. Natl. Acad. Sci. USA 84, 8898–8902 (1987).

    Article  CAS  Google Scholar 

  21. Arai, R., Ueda, H., Kitayama, A., Kamiya, N. & Nagamune, T. Design of the linkers which effectively separate domains of a bifunctional fusion protein. Protein Eng. 14, 529–532 (2001).

    Article  CAS  Google Scholar 

  22. Sicheri, F. & Yang, D.S. Ice-binding structure and mechanism of an antifreeze protein from winter flounder. Nature 375, 427–431 (1995).

    Article  CAS  Google Scholar 

  23. Liepinsh, E. et al. Solution structure of a hydrophobic analogue of the winter flounder antifreeze protein. Eur. J. Biochem. 269, 1259–1266 (2002).

    Article  CAS  Google Scholar 

  24. Schmeing, T.M., Huang, K.S., Strobel, S.A. & Steitz, T.A. An induced-fit mechanism to promote peptide bond formation and exclude hydrolysis of peptidyl-tRNA. Nature 438, 520–524 (2005).

    Article  CAS  Google Scholar 

  25. Cruz-Vera, L.R., Rajagopal, S., Squires, C. & Yanofsky, C. Features of ribosome-peptidyl-tRNA interactions essential for tryptophan induction of tna operon expression. Mol. Cell 19, 333–343 (2005).

    Article  CAS  Google Scholar 

  26. Kosolapov, A. & Deutsch, C. Tertiary interactions within the ribosomal exit tunnel. Nat. Struct. Mol. Biol. 16, 405–411 (2009).

    Article  CAS  Google Scholar 

  27. Liao, S., Lin, J., Do, H. & Johnson, A.E. Both lumenal and cytosolic gating of the aqueous ER translocon pore are regulated from inside the ribosome during membrane protein integration. Cell 90, 31–41 (1997).

    Article  CAS  Google Scholar 

  28. Berndt, U., Oellerer, S., Zhang, Y., Johnson, A.E. & Rospert, S. A signal-anchor sequence stimulates signal recognition particle binding to ribosomes from inside the exit tunnel. Proc. Natl. Acad. Sci. USA 106, 1398–1403 (2009).

    Article  CAS  Google Scholar 

  29. Mingarro, I., Nilsson, I., Whitley, P. & von Heijne, G. Different conformations of nascent polypeptides during translocation across the ER membrane. BMC Cell Biol. 1, 3 (2000).

    Article  CAS  Google Scholar 

  30. Erickson, A.H. & Blobel, G. Cell-free translation of messenger RNA in a wheat germ system. Methods Enzymol. 96, 38–50 (1983).

    Article  CAS  Google Scholar 

  31. Wagenknecht, T., Frank, J., Boublik, M., Nurse, K. & Ofengand, J. Direct localization of the tRNA-anticodon interaction site on the Escherichia coli 30 S ribosomal subunit by electron microscopy and computerized image averaging. J. Mol. Biol. 203, 753–760 (1988).

    Article  CAS  Google Scholar 

  32. Mindell, J.A. & Grigorieff, N. Accurate determination of local defocus and specimen tilt in electron microscopy. J. Struct. Biol. 142, 334–347 (2003).

    Article  Google Scholar 

  33. Frank, J. Three-dimensional electron microscopy of macromolecular assemblies. in Three-dimensional Electron Microscopy of Macromolecular Assemblies (Academic Press, 1996).

  34. Jossinet, F. & Westhof, E. Sequence to structure (S2S): display, manipulate and interconnect RNA data from sequence to structure. Bioinformatics 21, 3320–3321 (2005).

    Article  CAS  Google Scholar 

  35. Selmer, M. et al. Structure of the 70S ribosome complexed with mRNA and tRNA. Science 313, 1935–1942 (2006).

    Article  CAS  Google Scholar 

  36. Schuwirth, B.S. et al. Structures of the bacterial ribosome at 3.5 Å resolution. Science 310, 827–834 (2005).

    Article  CAS  Google Scholar 

  37. Massire, C. & Westhof, E. MANIP: an interactive tool for modelling RNA. J. Mol. Graph. Model. 16, 197–205, 255–197 (1998).

    Article  CAS  Google Scholar 

  38. Yang, H. et al. Tools for the automatic identification and classification of RNA base pairs. Nucleic Acids Res. 31, 3450–3460 (2003).

    Article  CAS  Google Scholar 

  39. Trabuco, L.G., Villa, E., Mitra, K., Frank, J. & Schulten, K. Flexible fitting of atomic structures into electron microscopy maps using molecular dynamics. Structure 16, 673–683 (2008).

    Article  CAS  Google Scholar 

  40. Notredame, C., Higgins, D.G. & Heringa, J. T-Coffee: a novel method for fast and accurate multiple sequence alignment. J. Mol. Biol. 302, 205–217 (2000).

    Article  CAS  Google Scholar 

  41. Eswar, N. et al. Comparative protein structure modeling using Modeller. Curr. Protoc. Bioinformatics 5, 5.6 (2006).

    Google Scholar 

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

    Article  Google Scholar 

  43. Humphrey, W., Dalke, A. & Schulten, K. VMD: visual molecular dynamics. J. Mol. Graph. 14, 33–38 (1996).

    Article  CAS  Google Scholar 

  44. Schmeing, T.M., Huang, K.S., Kitchen, D.E., Strobel, S.A. & Steitz, T.A. Structural insights into the roles of water and the 2′ hydroxyl of the P site tRNA in the peptidyl transferase reaction. Mol. Cell 20, 437–448 (2005).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

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Acknowledgements

We would like to thank B. Beatrix for help with the wheat germ translation system and J. Buerger for help with the electron microscopy. This research was supported by Federation of European Biochemical Societies and Knut och Alice Wallenbergs Stiftelse postdoctoral fellowships (to S.B.), grants from the Deutsche Forschungsgemeinschaft SFB594 and SFB646 (to R.B.), SFB740 (to T.M.) and WI3285/1-1 (to D.N.W.) and by the European Union and Senatsverwaltung für Wissenschaft, Forschung und Kultur Berlin.

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Author notes

  1. Mario Halic

    Present address: Present address: Harvard Medical School, Department of Cell Biology, Boston, Massachusetts, USA.,

  2. Shashi Bhushan and Marco Gartmann: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry, Gene Center and Center for Integrated Protein Science Munich, University of Munich, Munich, Germany

    Shashi Bhushan, Marco Gartmann, Mario Halic, Jean-Paul Armache, Alexander Jarasch, Otto Berninghausen, Daniel N Wilson & Roland Beckmann

  2. UltraStrukturNetzwerk, Max Planck Institute for Molecular Genetics, Berlin, Germany

    Thorsten Mielke

  3. Institut für Medizinische Physik und Biophysik, Charité, Berlin, Germany

    Thorsten Mielke

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  1. Shashi Bhushan
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Contributions

S.B., M.G. and R.B. designed research; S.B. prepared the complexes and collected data; O.B. and T.M. helped with data collection; S.B., M.G. and M.H. processed datasets; J.-P.A., A.J. and D.N.W. prepared models; S.B., D.N.W. and R.B. analyzed results, prepared figures and wrote the paper.

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Correspondence to Roland Beckmann.

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Bhushan, S., Gartmann, M., Halic, M. et al. α-Helical nascent polypeptide chains visualized within distinct regions of the ribosomal exit tunnel. Nat Struct Mol Biol 17, 313–317 (2010). https://doi.org/10.1038/nsmb.1756

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