Article

Translational arrest by a prokaryotic signal recognition particle is mediated by RNA interactions

  • Nature Structural & Molecular Biology volume 22, pages 767773 (2015)
  • doi:10.1038/nsmb.3086
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Abstract

The signal recognition particle (SRP) recognizes signal sequences of nascent polypeptides and targets ribosome–nascent chain complexes to membrane translocation sites. In eukaryotes, translating ribosomes are slowed down by the Alu domain of SRP to allow efficient targeting. In prokaryotes, however, little is known about the structure and function of Alu domain–containing SRPs. Here, we report a complete molecular model of SRP from the Gram-positive bacterium Bacillus subtilis, based on cryo-EM. The SRP comprises two subunits, 6S RNA and SRP54 or Ffh, and it facilitates elongation slowdown similarly to its eukaryotic counterpart. However, protein contacts with the small ribosomal subunit observed for the mammalian Alu domain are substituted in bacteria by RNA-RNA interactions of 6S RNA with the α-sarcin–ricin loop and helices H43 and H44 of 23S rRNA. Our findings provide a structural basis for cotranslational targeting and RNA-driven elongation arrest in prokaryotes.

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Acknowledgements

We thank C. Ungewickell and O. Berninghausen for support with cryo-EM (sample preparation and data collection), E. van der Sluis and J. Musial for E. coli SRP purification, T. Becker for manual data collection and B. Beatrix for support in eukaryotic SRP purification and discussion. We thank R. Matadeen and S. DeCarlo for data collection at the Netherlands Centre for Electron Nanoscopy facility. We thank B. Dobberstein (Ruprecht-Karls-Universitat Heidelberg) for providing tissues. R.B. is supported by the Deutsche Forschungsgemeinschaft (DFG) through grants SFB646, GRK1721 and FOR1805; the Graduate School of Quantitative Biosciences Munich (QBM); the Center for Integrated Protein Science Munich; and the European Research Council (Advanced Grant CRYOTRANSLATION). D.N.W. is supported by the DFG through grants FOR1805, WI3285/3-1 and GRK1721. B.B. is supported by a European Molecular Biology Organization Long Term Fellowship (ALTF 50-2011).

Author information

Affiliations

  1. Gene Center, Department of Biochemistry, University of Munich, Munich, Germany.

    • Bertrand Beckert
    • , Alexej Kedrov
    • , Daniel Sohmen
    • , Daniel N Wilson
    •  & Roland Beckmann
  2. Center for Integrated Protein Science Munich (CIPSM), Munich, Germany.

    • Bertrand Beckert
    • , Alexej Kedrov
    • , Daniel Sohmen
    • , Daniel N Wilson
    •  & Roland Beckmann
  3. Biochemistry Center, University of Heidelberg, Heidelberg, Germany.

    • Georg Kempf
    • , Klemens Wild
    •  & Irmgard Sinning
  4. Center for Cellular Imaging and NanoAnalytics, Biozentrum, University of Basel, Basel, Switzerland.

    • Henning Stahlberg

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Contributions

B.B. prepared the translation extract, the translation arrest and the SRP complexes, and performed single-particle cryo-EM analysis, model building and figure preparation. A.K. performed the MST analysis. All authors interpreted the data and assisted with manuscript preparation. B.B., A.K., D.N.W. and R.B. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Roland Beckmann.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–7

  2. 2.

    Supplementary Data Set 1

    Cell-extract preparation, raw western blot images

Videos

  1. 1.

    Overview of the Cryo-EM structure of BsSRP–RNC complex

    Cryo-EM reconstruction of the BsSRP–RNC. Small 30S subunit (yellow), large 50S subunit (gray), P-site tRNA (green), 6S RNA (red) and the density corresponding to Ffh M-domain (blue).

  2. 2.

    BsSRP Alu domain interaction with the ribosome

    Molecular model of B. subtilis 6S Alu domain 'locking in' by generating a continuous stacking between the α-sarcin-ricin loop and helix H43 H44 of 23S rRNA.