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Following the intersubunit conformation of the ribosome during translation in real time

Nature Structural & Molecular Biology volume 17, pages 793–800 (2010)Cite this article

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Abstract

We report the direct observation of conformational rearrangements of the ribosome during multiple rounds of elongation. Using single-molecule fluorescence resonance energy transfer, we monitored the intersubunit conformation of the ribosome in real time as it proceeds from codon to codon. During each elongation cycle, the ribosome unlocks upon peptide bond formation, then reverts to the locked state upon translocation onto the next codon. Our data reveal both the specific and cumulative effects of antibiotics on individual steps of translation and uncover the processivity of the ribosome as it elongates. Our approach interrogates the precise molecular events occurring at each codon of the mRNA within the full context of ongoing translation.

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Figure 1: Intersubunit FRET reports on the elongation cycle at each codon during translation.
Figure 2: The number of complete FRET cycles is controlled by coding length.
Figure 3: Each FRET cycle reports on one round of elongation at a distinct codon.
Figure 4: Erythromycin stalls single translating ribosomes by blocking the nascent chain.
Figure 5: The precise mechanisms and cumulative effects of ribosome-targeting antibiotics observed at codon resolution.
Figure 6: Monitoring the elongation cycle during multiple rounds of translation reveals increased translocation rates beyond initial codons.
Figure 7: A general model for ribosome dynamics and function.

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Acknowledgements

This work was funded by grants to J.D.P. from the US National Institutes of Health.

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Author notes

  1. Colin Echeverría Aitken

    Present address: Present address: Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.,

Authors and Affiliations

  1. Biophysics Program, Stanford University, Stanford, California, USA

    Colin Echeverría Aitken

  2. Department of Structural Biology, Stanford University School of Medicine, Stanford, California, USA

    Joseph D Puglisi

  3. Stanford Magnetic Resonance Laboratory, Stanford University School of Medicine, Stanford, California, USA

    Joseph D Puglisi

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C.E.A. performed experiments and data analysis. C.E.A. and J.D.P. discussed results and wrote the manuscript.

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Correspondence to Joseph D Puglisi.

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Aitken, C., Puglisi, J. Following the intersubunit conformation of the ribosome during translation in real time. Nat Struct Mol Biol 17, 793–800 (2010). https://doi.org/10.1038/nsmb.1828

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