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Ribosome dynamics and tRNA movement by time-resolved electron cryomicroscopy


The translocation step of protein synthesis entails large-scale rearrangements of the ribosome–transfer RNA (tRNA) complex. Here we have followed tRNA movement through the ribosome during translocation by time-resolved single-particle electron cryomicroscopy (cryo-EM). Unbiased computational sorting of cryo-EM images yielded 50 distinct three-dimensional reconstructions, showing the tRNAs in classical, hybrid and various novel intermediate states that provide trajectories and kinetic information about tRNA movement through the ribosome. The structures indicate how tRNA movement is coupled with global and local conformational changes of the ribosome, in particular of the head and body of the small ribosomal subunit, and show that dynamic interactions between tRNAs and ribosomal residues confine the path of the tRNAs through the ribosome. The temperature dependence of ribosome dynamics reveals a surprisingly flat energy landscape of conformational variations at physiological temperature. The ribosome functions as a Brownian machine that couples spontaneous conformational changes driven by thermal energy to directed movement.

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Figure 1: Trajectories of tRNA movement during (retro-)translocation.
Figure 2: Dynamics of ribosome–tRNA interactions and of the 30S subunit.
Figure 3: Positions of the E-site tRNA.
Figure 4: Coupling between tRNA movement and 30S dynamics.
Figure 5: Increase of ribosome dynamics with temperature.

Accession codes

Data deposits

The three-dimensional density maps have been deposited in the Electron Microscopy Data Bank at the European Biology Laboratory—European Bioinformatics Institute—under accession codes EMD-1716 to EMD-1720 (pre1–5), EMD-1721 to EMD-1723 (post1–3), EMD-1724 (sub-state of post-translocation complex at 18 °C), and EMD-1725 to EMD-1727 (post-translocation complex at 18 °C, 4 °C and 37 °C), respectively.


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We thank C. Blau for technical help in preparing Fig. 4. Research in the laboratory of H.S. was supported by grants from the Federal Ministry of Education and Research (BMBF), Germany, the Sixth Framework Programme of the European Union via the Integrated Project 3DRepertoire, by the state of Lower-Saxony and the Volkswagen Foundation, Hannover, Germany. N.F. was supported by a Boehringer-Ingelheim fellowship. M.V.R. and W.W. were supported by grants from the Deutsche Forschungsgemeinschaft.

Author information

Authors and Affiliations



N.F. conceived and performed the cryo-EM experiments and data analysis with mentoring by H.S.; A.L.K. prepared complexes for the analysis; N.F., A.L.K., W.W., M.V.R. and H.S. discussed results and wrote the manuscript.

Corresponding author

Correspondence to Holger Stark.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Methods, Supplementary Figures 1- 11 with legends, Supplementary Tables 1 - 4, References and full legends for Supplementary Movies 1- 6 and 3D PDFs. (PDF 3144 kb)

Supplementary Movie 1

GIF-animation visualizing ribosome dynamics and tRNA movement during retro-translocation as revealed by unsupervised 2D classification (see Supplementary Information file for full legend). (GIF 4451 kb)

Supplementary Movie 2

The movie shows a sequence of 20 ribosome 3D structures of sub-states along the (retro-) translocation pathway arranged according to the smallest free energy differences between subsequent sub-states (see Supplementary Information file for full legend). (MPG 1514 kb)

Supplementary Movie 3

GIF-animation of the same sequence of 20 sub-states as in Supplementary Movie 2 (see Supplementary Information file for full legend). (GIF 1847 kb)

Supplementary Movie 4

GIF-animation showing a sequence of 11 different sub-states of the classic pre-translocation state pre1 from top (upper row) and the solvent side of the 30S subunit (lower left) (see Supplementary Information file for full legend). (GIF 1407 kb)

Supplementary Movie 5

GIF-animation similar to Supplementary Movie 2, but depicting only 11 sub states (see Supplementary Information file for full legend). (GIF 1307 kb)

Supplementary Movie 6

Movie visualizing the temperature dependence of ribosome dynamics by interpolating between cryo-EM maps of the same post-translocation complex prepared at different temperatures (see Supplementary Information file for full legend). (MPG 6651 kb)

Supplementary 3D PDFs

Nine isosurface representations of sub-states of the 70S˙fMetVal tRNAVal˙tRNAfMet complex (see Supplementary Information file for full legend). (PDF 24002 kb)

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Fischer, N., Konevega, A., Wintermeyer, W. et al. Ribosome dynamics and tRNA movement by time-resolved electron cryomicroscopy. Nature 466, 329–333 (2010).

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