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Correlated conformational events in EF-G and the ribosome regulate translocation

Nature Structural & Molecular Biology volume 17, pages 14701477 (2010) | Download Citation

Abstract

In bacteria, the translocation of tRNA and mRNA with respect to the ribosome is catalyzed by the conserved GTPase elongation factor-G (EF-G). To probe the rate-determining features in this process, we imaged EF-G–catalyzed translocation from two unique structural perspectives using single-molecule fluorescence resonance energy transfer. The data reveal that the rate at which the ribosome spontaneously achieves a transient, 'unlocked' state is closely correlated with the rate at which the tRNA-like domain IV-V element of EF-G engages the A site. After these structural transitions, translocation occurs comparatively fast, suggesting that conformational processes intrinsic to the ribosome determine the rate of translocation. Experiments conducted in the presence of non-hydrolyzable GTP analogs and specific antibiotics further reveal that allosterically linked conformational events in EF-G and the ribosome mediate rapid, directional substrate movement and EF-G release.

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Acknowledgements

The authors wish to thank C. Walsh (Harvard University) and members of his laboratory for providing the Sfp expression vector and for their assistance with the fluorescent labeling of EF-G and K.Y. Sanbonmatsu and P. Whitford for structural models of the E. coli ribosome. This work was supported by US National Institutes of Health grant GM079238.

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  1. Department of Physiology and Biophysics, Weill Cornell Medical College of Cornell University, New York, New York, USA.

    • James B Munro
    • , Michael R Wasserman
    • , Roger B Altman
    • , Leyi Wang
    •  & Scott C Blanchard

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Contributions

J.B.M. and S.C.B. designed the experiments. J.B.M. and M.R.W. conducted the experiments and analyzed the data. R.B.A. and L.W. prepared reagents. J.B.M., M.R.W., R.B.A. and S.C.B. wrote the paper.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Scott C Blanchard.

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https://doi.org/10.1038/nsmb.1925

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