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Structure of the human 80S ribosome

Nature volume 520, pages 640–645 (2015)Cite this article

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Abstract

Ribosomes are translational machineries that catalyse protein synthesis. Ribosome structures from various species are known at the atomic level, but obtaining the structure of the human ribosome has remained a challenge; efforts to address this would be highly relevant with regard to human diseases. Here we report the near-atomic structure of the human ribosome derived from high-resolution single-particle cryo-electron microscopy and atomic model building. The structure has an average resolution of 3.6 Å, reaching 2.9 Å resolution in the most stable regions. It provides unprecedented insights into ribosomal RNA entities and amino acid side chains, notably of the transfer RNA binding sites and specific molecular interactions with the exit site tRNA. It reveals atomic details of the subunit interface, which is seen to remodel strongly upon rotational movements of the ribosomal subunits. Furthermore, the structure paves the way for analysing antibiotic side effects and diseases associated with deregulated protein synthesis.

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Figure 1: Overall structure of the human 80S ribosome.
Figure 2: Local resolution and electron density maps.
Figure 3: Conformational changes of the human ribosome.
Figure 4: Ribosomal inter-subunit bridges.
Figure 5: Molecular details of characteristic inter-subunit contacts.
Figure 6: Molecular recognition by the human ribosome at the level of the E-site tRNA.

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Accession codes

Primary accessions

Electron Microscopy Data Bank

Protein Data Bank

Data deposits

The cryo-EM map and atomic coordinates have been deposited in the EMDB and Protein Data Bank under accession codes EMD-2938 and 4ug0, respectively.

References

  1. Anger, A. M. et al. Structures of the human and Drosophila 80S ribosome. Nature 497, 80–85 (2013)

    Article  ADS  CAS  PubMed  Google Scholar 

  2. Ben-Shem, A. et al. The structure of the eukaryotic ribosome at 3.0 Å resolution. Science 334, 1524–1529 (2011)

    Article  ADS  CAS  PubMed  Google Scholar 

  3. Rabl, J., Leibundgut, M., Ataide, S. F., Haag, A. & Ban, N. Crystal structure of the eukaryotic 40S ribosomal subunit in complex with initiation factor 1. Science 331, 730–736 (2011)

    Article  ADS  CAS  PubMed  Google Scholar 

  4. Klinge, S., Voigts-Hoffmann, F., Leibundgut, M., Arpagaus, S. & Ban, N. Crystal structure of the eukaryotic 60S ribosomal subunit in complex with initiation factor 6. Science 334, 941–948 (2011)

    Article  ADS  CAS  PubMed  Google Scholar 

  5. Spahn, C. M. et al. Cryo-EM visualization of a viral internal ribosome entry site bound to human ribosomes: the IRES functions as an RNA-based translation factor. Cell 118, 465–475 (2004)

    Article  CAS  PubMed  Google Scholar 

  6. Boehringer, D., Thermann, R., Ostareck-Lederer, A., Lewis, J. D. & Stark, H. Structure of the hepatitis C virus IRES bound to the human 80S ribosome: remodeling of the HCV IRES. Structure 13, 1695–1706 (2005)

    Article  CAS  PubMed  Google Scholar 

  7. Chandramouli, P. et al. Structure of the mammalian 80S ribosome at 8.7 Å resolution. Structure 16, 535–548 (2008)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Voorhees, R. M., Fernandez, I. S., Scheres, S. H. & Hegde, R. S. Structure of the mammalian ribosome-Sec61 complex to 3.4 Å resolution. Cell 157, 1632–1643 (2014)

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Amunts, A. et al. Structure of the yeast mitochondrial large ribosomal subunit. Science 343, 1485–1489 (2014)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  10. Greber, B. J. et al. The complete structure of the large subunit of the mammalian mitochondrial ribosome. Nature 515, 283–286 (2014)

    Article  ADS  CAS  PubMed  Google Scholar 

  11. Brown, A. et al. Structure of the large ribosomal subunit from human mitochondria. Science 346, 718–722 (2014)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  12. Li, X., Zheng, S. Q., Egami, K., Agard, D. A. & Cheng, Y. Influence of electron dose rate on electron counting images recorded with the K2 camera. J. Struct. Biol. 184, 251–260 (2013)

    Article  CAS  PubMed  Google Scholar 

  13. Li, X. et al. Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM. Nature Methods 10, 584–590 (2013)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Bai, X. C., Fernandez, I. S., McMullan, G. & Scheres, S. H. Ribosome structures to near-atomic resolution from thirty thousand cryo-EM particles. eLife 2, e00461 (2013)

    Article  PubMed  PubMed Central  Google Scholar 

  15. Khatter, H. et al. Purification, characterization and crystallization of the human 80S ribosome. Nucleic Acids Res. 42, e49 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. DeLaBarre, B. & Brunger, A. T. Considerations for the refinement of low-resolution crystal structures. Acta Crystallogr. D 62, 923–932 (2006)

    Article  CAS  PubMed  Google Scholar 

  17. Afonine, P. V. et al. FEM: feature-enhanced map. Acta Crystallogr. D 71, 646–666 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Ben-Shem, A., Jenner, L., Yusupova, G. & Yusupov, M. Crystal structure of the eukaryotic ribosome. Science 330, 1203–1209 (2010)

    Article  ADS  CAS  PubMed  Google Scholar 

  19. Fei, J., Kosuri, P., MacDougall, D. D. & Gonzalez, R. L., Jr Coupling of ribosomal L1 stalk and tRNA dynamics during translation elongation. Mol. Cell 30, 348–359 (2008)

    Article  CAS  PubMed  Google Scholar 

  20. Cornish, P. V. et al. Following movement of the L1 stalk between three functional states in single ribosomes. Proc. Natl Acad. Sci. USA 106, 2571–2576 (2009)

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  21. Feng, S., Chen, Y. & Gao, Y. G. Crystal structure of 70S ribosome with both cognate tRNAs in the E and P sites representing an authentic elongation complex. PLoS ONE 8, e58829 (2013)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  22. Valle, M. et al. Locking and unlocking of ribosomal motions. Cell 114, 123–134 (2003)

    Article  CAS  PubMed  Google Scholar 

  23. Frank, J. & Agrawal, R. K. A ratchet-like inter-subunit reorganization of the ribosome during translocation. Nature 406, 318–322 (2000)

    Article  ADS  CAS  PubMed  Google Scholar 

  24. Budkevich, T. V. et al. Regulation of the mammalian elongation cycle by subunit rolling: a eukaryotic-specific ribosome rearrangement. Cell 158, 121–131 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Budkevich, T. et al. Structure and dynamics of the mammalian ribosomal pretranslocation complex. Mol. Cell 44, 214–224 (2011)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Chen, J., Petrov, A., Tsai, A., O’Leary, S. E. & Puglisi, J. D. Coordinated conformational and compositional dynamics drive ribosome translocation. Nature Struct. Mol. Biol. 20, 718–727 (2013)

    Article  CAS  Google Scholar 

  27. Leontis, N. B. & Westhof, E. Geometric nomenclature and classification of RNA base pairs. RNA 7, 499–512 (2001)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Klaholz, B. & Moras, D. C.-H. O hydrogen bonds in the nuclear receptor RARgamma–a potential tool for drug selectivity. Structure 10, 1197–1204 (2002)

    Article  CAS  PubMed  Google Scholar 

  29. Liu, A. et al. NMR detection of bifurcated hydrogen bonds in large proteins. J. Am. Chem. Soc. 130, 2428–2429 (2008)

    Article  CAS  PubMed  Google Scholar 

  30. Jiang, L. & Lai, L. C. H. O hydrogen bonds at protein-protein interfaces. J. Biol. Chem. 277, 37732–37740 (2002)

    Article  CAS  PubMed  Google Scholar 

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

    Article  ADS  CAS  PubMed  Google Scholar 

  32. Cate, J. H., Yusupov, M. M., Yusupova, G. Z., Earnest, T. N. & Noller, H. F. X-ray crystal structures of 70S ribosome functional complexes. Science 285, 2095–2104 (1999)

    Article  CAS  PubMed  Google Scholar 

  33. Yusupov, M. M. et al. Crystal structure of the ribosome at 5.5 Å resolution. Science 292, 883–896 (2001)

    Article  ADS  CAS  PubMed  Google Scholar 

  34. Shenvi, C. L., Dong, K. C., Friedman, E. M., Hanson, J. A. & Cate, J. H. Accessibility of 18S rRNA in human 40S subunits and 80S ribosomes at physiological magnesium ion concentrations–implications for the study of ribosome dynamics. RNA 11, 1898–1908 (2005)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Moore, M. N. & Spremulli, L. L. Effects of cations and cosolvents on eukaryotic ribosomal subunit conformation. Biochemistry 24, 191–196 (1985)

    Article  CAS  PubMed  Google Scholar 

  36. Sperrazza, J. M., Russell, D. W. & Spremulli, L. L. Reversible dissociation of wheat germ ribosomal subunits: cation-dependent equilibria and thermodynamic parameters. Biochemistry 19, 1053–1058 (1980)

    Article  CAS  PubMed  Google Scholar 

  37. Zhang, W., Dunkle, J. A. & Cate, J. H. Structures of the ribosome in intermediate states of ratcheting. Science 325, 1014–1017 (2009)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  38. Dunkle, J. A. et al. Structures of the bacterial ribosome in classical and hybrid states of tRNA binding. Science 332, 981–984 (2011)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  39. Tourigny, D. S., Fernandez, I. S., Kelley, A. C. & Ramakrishnan, V. Elongation factor G bound to the ribosome in an intermediate state of translocation. Science 340, 1235490 (2013)

    Article  CAS  PubMed  Google Scholar 

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

    Article  ADS  CAS  PubMed  Google Scholar 

  41. Garreau de Loubresse, N. et al. Structural basis for the inhibition of the eukaryotic ribosome. Nature 513, 517–522 (2014)

    Article  ADS  CAS  PubMed  Google Scholar 

  42. Jenner, L., Rees, B., Yusupov, M. & Yusupova, G. Messenger RNA conformations in the ribosomal E site revealed by X-ray crystallography. EMBO Rep. 8, 846–850 (2007)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Rhodin, M. H. & Dinman, J. D. A flexible loop in yeast ribosomal protein L11 coordinates P-site tRNA binding. Nucleic Acids Res. 38, 8377–8389 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Yamamoto, H. et al. Structure of the mammalian 80S initiation complex with initiation factor 5B on HCV-IRES RNA. Nature Struct. Mol. Biol. 21, 721–727 (2014)

    Article  CAS  Google Scholar 

  45. Hinnebusch, A. G. The scanning mechanism of eukaryotic translation initiation. Annu. Rev. Biochem. 83, 779–812 (2014)

    Article  CAS  PubMed  Google Scholar 

  46. Pestova, T. V. et al. Molecular mechanisms of translation initiation in eukaryotes. Proc. Natl Acad. Sci. USA 98, 7029–7036 (2001)

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  47. Demeshkina, N., Jenner, L., Westhof, E., Yusupov, M. & Yusupova, G. A new understanding of the decoding principle on the ribosome. Nature 484, 256–259 (2012)

    Article  ADS  CAS  PubMed  Google Scholar 

  48. Ludtke, S. J., Baldwin, P. R. & Chiu, W. EMAN: semiautomated software for high-resolution single-particle reconstructions. J. Struct. Biol. 128, 82–97 (1999)

    Article  CAS  PubMed  Google Scholar 

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

    Article  PubMed  Google Scholar 

  50. Scheres, S. H. RELION: implementation of a Bayesian approach to cryo-EM structure determination. J. Struct. Biol. 180, 519–530 (2012)

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Chen, S. et al. High-resolution noise substitution to measure overfitting and validate resolution in 3D structure determination by single particle electron cryomicroscopy. Ultramicroscopy 135, 24–35 (2013)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Rosenthal, P. B. & Henderson, R. Optimal determination of particle orientation, absolute hand, and contrast loss in single-particle electron cryomicroscopy. J. Mol. Biol. 333, 721–745 (2003)

    Article  CAS  PubMed  Google Scholar 

  53. van Heel, M. & Schatz, M. Fourier shell correlation threshold criteria. J. Struct. Biol. 151, 250–262 (2005)

    Article  CAS  PubMed  Google Scholar 

  54. Kucukelbir, A., Sigworth, F. J. & Tagare, H. D. Quantifying the local resolution of cryo-EM density maps. Nature Methods 11, 63–65 (2014)

    Article  CAS  PubMed  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  56. Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D 66, 486–501 (2010)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Ban, N. et al. A new system for naming ribosomal proteins. Curr. Opin. Struct. Biol. 24, 165–169 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Afonine, P. V. et al. Towards automated crystallographic structure refinement with phenix.refine. Acta Crystallogr. D 68, 352–367 (2012)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  ADS  CAS  PubMed  Google Scholar 

  60. DeLano, W. L. The PyMOL Molecular Graphic System. (DeLano Scientiic, 2002).

Download references

Acknowledgements

We thank A. Urzhumtsev and P. Afonine for discussions on FEM maps; R. Fritz and J. Michalon for IT support; R. Drui for constant high-quality engineers support; J.-F. Ménétret for technical support; H. Stark for discussions on Cs-correction; D. Moras and J.-C. Thierry for continuous encouragement in pushing structural biology frontiers; the IGBMC cell culture facilities for HeLa cell production; and the High Performance Computing Center of the University of Strasbourg funded by the Equipex Equip@Meso project. This long-term project (since 2003) was supported by the CNRS and the European Research Council (ERC Starting Grant N_243296 TRANSLATIONMACHINERY), and the electron microscope facility was supported by the Alsace Region, the FRM, the IBiSA platform program, INSERM, CNRS and the Association pour la Recherche sur le Cancer (ARC) and by the French Infrastructure for Integrated Structural Biology (FRISBI) ANR-10-INSB-05-01, and Instruct as part of the European Strategy Forum on Research Infrastructures (ESFRI).

Author information

Author notes

  1. Heena Khatter, Alexander G. Myasnikov and S. Kundhavai Natchiar: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Integrated Structural Biology, Centre for Integrative Biology (CBI), IGBMC (Institute of Genetics and of Molecular and Cellular Biology), 1 rue Laurent Fries, Illkirch, 67404, France

    Heena Khatter, Alexander G. Myasnikov, S. Kundhavai Natchiar & Bruno P. Klaholz

  2. Centre National de la Recherche Scientifique (CNRS), UMR 7104, Illkirch, 67404, France

    Heena Khatter, Alexander G. Myasnikov, S. Kundhavai Natchiar & Bruno P. Klaholz

  3. Institut National de la Santé et de la Recherche Médicale (INSERM) U964, Illkirch, 67404, France

    Heena Khatter, Alexander G. Myasnikov, S. Kundhavai Natchiar & Bruno P. Klaholz

  4. Université de Strasbourg, 67081, Strasbourg, France

    Heena Khatter, Alexander G. Myasnikov, S. Kundhavai Natchiar & Bruno P. Klaholz

Authors
  1. Heena Khatter
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  2. Alexander G. Myasnikov
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  3. S. Kundhavai Natchiar
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  4. Bruno P. Klaholz
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Contributions

H.K. conducted purification, optimization of samples for cryo-EM and cryo-EM data processing. A.G.M. performed cryo-EM data acquisition, image processing, structure refinement and model building. S.K.N. performed structure refinement and model building. B.P.K supervised the study. All authors analysed the data. B.P.K and H.K. wrote the manuscript, with input from A.G.M. and S.K.N.

Corresponding author

Correspondence to Bruno P. Klaholz.

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The authors declare no competing financial interests.

Extended data figures and tables

Extended Data Figure 1 Resolution estimation.

The average resolution of the cryo-EM map as estimated from Fourier shell correlation according to the 0.143 criterion.

Extended Data Figure 2 Features of the refined cryo-EM 3D reconstruction.

Final cryo-EM map of the peptidyl-transferase centre region illustrating the high level of details observed.

Extended Data Figure 3 Typical electron density map regions of the human ribosome structure.

These final electron density maps were obtained by combining experimental amplitudes derived from the cryo-EM map and phases calculated from the iteratively refined atomic model, as done in standard refinement procedures in X-ray crystallography (see main text).

Extended Data Figure 4 Comparison of maps determined by cryo-EM and X-ray crystallography.

Top and middle, cryo-EM map and Phenix map of the human 80S ribosome (this study, A4546 region); bottom, crystal structure of a 70S ribosome at 2.8 Å resolution59 (corresponding A2600 region).

Extended Data Figure 5 Ribosomal protein structures.

a, b, Structure of ribosomal proteins uS2 and eS17.

Extended Data Figure 6 The decoding centre on the 40S.

a, Contact regions of the E-site tRNA on the 40S subunit. b, Catalytic PTC region of the ribosome, highlighting the partial structural disorder of a region of protein uL16 (loop), and disorder of residue U4548 (28S rRNA) in the absence of P-site tRNA, while all other residues are well-ordered. This suggests that the P-site pocket is largely pre-defined while U4548 and the loop participate in tRNA accommodation.

Extended Data Figure 7 Particle sorting scheme.

Particle sorting was done by 3D classification using six classes starting from 75,000 particles, resulting in two dominant classes with 10,000 and 45,000 particles, in rotated and non-rotated ribosome conformations respectively; the highest-resolution close-to-focus data set was refined using movie processing (see Methods).

Extended Data Table 1 Inter-subunit bridges and 60S and 40S residues involved in inter-subunit interactions
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Khatter, H., Myasnikov, A., Natchiar, S. et al. Structure of the human 80S ribosome. Nature 520, 640–645 (2015). https://doi.org/10.1038/nature14427

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