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Transfer RNA–mediated regulation of ribosome dynamics during protein synthesis

Nature Structural & Molecular Biology volume 18, pages 10431051 (2011) | Download Citation

Abstract

Translocation of tRNAs through the ribosome during protein synthesis involves large-scale structural rearrangement of the ribosome and ribosome-bound tRNAs that is accompanied by extensive and dynamic remodeling of tRNA-ribosome interactions. How the rearrangement of individual tRNA-ribosome interactions influences tRNA movement during translocation, however, remains largely unknown. To address this question, we used single-molecule FRET to characterize the dynamics of ribosomal pretranslocation (PRE) complex analogs carrying either wild-type or systematically mutagenized tRNAs. Our data reveal how specific tRNA-ribosome interactions regulate the rate of PRE complex rearrangement into a critical, on-pathway translocation intermediate and how these interactions control the stability of the resulting configuration. Notably, our results suggest that the conformational flexibility of the tRNA molecule has a crucial role in directing the structural dynamics of the PRE complex during translocation.

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Acknowledgements

We thank U. RajBhandary for providing us with the plasmids and strains necessary to generate, overexpress and purify all tRNAfMet mutants reported here; J. Frank, H. Gao and X. Aguirrezabala for providing us with structural models for GS1 and GS2; K. Schulten and B. Liu for providing us with a quasi-atomic-resolution model of a hybrid P/E–configured tRNA; J. Cate for providing us with an early preprint of ref. 47; D. MacDougall, W. Ning, S. Mitra, C. Perez and J. Cate for valuable discussions and comments on the manuscript; and C. Perez for managing the Gonzalez laboratory. This work was supported by grants to R.L.G. from the Burroughs Wellcome Fund (CABS 1004856), the US National Science Foundation (MCB 0644262) and the US National Institute of General Medical Sciences (GM 084288). A.C.R. was supported, in part, by the Amgen Scholars Program at Columbia University.

Author information

Author notes

    • Jingyi Fei
    • , Arianne C Richard
    •  & Jonathan E Bronson

    Present addresses: Department of Physics, Center for the Physics of Living Cells, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA (J.F.); Immunoregulation Section, Autoimmunity Branch, National Institute of Arthritis and Musculoskeletal and Skin Diseases, Bethesda, Maryland, USA (A.C.R.); Boston Consulting Group, New York, New York, USA (J.E.B.).

Affiliations

  1. Department of Chemistry, Columbia University, New York, New York, USA.

    • Jingyi Fei
    • , Jonathan E Bronson
    •  & Ruben L Gonzalez Jr
  2. Department of Biological Sciences, Columbia University, New York, New York, USA.

    • Arianne C Richard

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Contributions

J.F. and R.L.G. Jr. designed the research; J.F. and A.C.R. carried out the experiments and analyzed the data; J.E.B. helped with the data analysis; J.F., A.C.R. and R.L.G. Jr. wrote the manuscript; all authors approved the final manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Ruben L Gonzalez Jr.

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DOI

https://doi.org/10.1038/nsmb.2098

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