Article | Published:

EF-G catalyzes tRNA translocation by disrupting interactions between decoding center and codon–anticodon duplex

Nature Structural & Molecular Biology volume 21, pages 817824 (2014) | Download Citation

Abstract

During translation, elongation factor G (EF-G) catalyzes the translocation of tRNA2–mRNA inside the ribosome. Translocation is coupled to a cycle of conformational rearrangements of the ribosomal machinery, and how EF-G initiates translocation remains unresolved. Here we performed systematic mutagenesis of Escherichia coli EF-G and analyzed inhibitory single-site mutants of EF-G that preserved pretranslocation (Pre)-state ribosomes with tRNAs in A/P and P/E sites (Pre–EF-G). Our results suggest that the interactions between the decoding center and the codon–anticodon duplex constitute the barrier for translocation. Catalysis of translocation by EF-G involves the factor's highly conserved loops I and II at the tip of domain IV, which disrupt the hydrogen bonds between the decoding center and the duplex to release the latter, hence inducing subsequent translocation events, namely 30S head swiveling and tRNA2–mRNA movement on the 30S subunit.

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Acknowledgements

We thank members of the P. Zhu laboratory and N. Gao for help and discussion. Y.Q. is supported by the Institute of Biophysics 135 Goal-oriented project, National Laboratory of Biomacromolecules (Institute of Biophysics, Chinese Academy of Sciences), and the State Key Laboratory of Molecular Biology (Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences). This work was supported by grants from the Ministry of Science and Technology of China (2012CB911000 and 2013CB531200 to Y.Q.), the National Natural Science Foundation of China (31322015, 31170756 and 31270847 to Y.Q.) and the Chinese Academy of Sciences (project KSZD-EW-Z-003 to Y.Q.).

Author information

Affiliations

  1. Laboratory of RNA Biology, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.

    • Guangqiao Liu
    • , Guangtao Song
    • , Danyang Zhang
    • , Dejiu Zhang
    • , Zhikai Li
    • , Jianshu Dong
    • , Weimin Gong
    •  & Yan Qin
  2. Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, College of Chemistry and Molecular Engineering, and Biodynamic Optical Imaging Center, Peking University, Beijing, China.

    • Zhixin Lyu
    •  & Xin Sheng Zhao
  3. NOXXON Pharma AG, Berlin, Germany.

    • John Achenbach
  4. Charité, Institute for Medical Physics and Biophysics, Berlin, Germany.

    • Knud H Nierhaus

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Contributions

G.L., G.S., Danyang Zhang, Dejiu Zhang and Z. Li cloned constructs and performed biochemical assays. Z. Lyu and G.S. collected FRET data and with X.S.Z. analyzed the FRET data. J.D. collected X-ray data and with W.G. resolved the structures. J.A. prepared some figures and analyzed data. Y.Q. and K.H.N. analyzed all data and wrote the manuscript. All authors discussed the results and commented on the manuscript. Y.Q. directed and supervised the project.

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The authors declare no competing financial interests.

Corresponding author

Correspondence to Yan Qin.

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https://doi.org/10.1038/nsmb.2869

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