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A conserved proline switch on the ribosome facilitates the recruitment and binding of trGTPases

Nature Structural & Molecular Biology volume 19, pages 403–410 (2012)Cite this article

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Abstract

When elongation factor G (EF-G) binds to the ribosome, it first makes contact with the C-terminal domain (CTD) of L12 before interacting with the N-terminal domain (NTD) of L11. Here we have identified a universally conserved residue, Pro22 of L11, that functions as a proline switch (PS22), as well as the corresponding center of peptidyl-prolyl cis-trans isomerase (PPIase) activity on EF-G that drives the cis-trans isomerization of PS22. Only the cis configuration of PS22 allows direct contact between the L11 NTD and the L12 CTD. Mutational analyses of both PS22 and the residues of the EF-G PPIase center reveal their function in translational GTPase (trGTPase) activity, protein synthesis and cell survival in Escherichia coli. Finally, we demonstrate that all known universal trGTPases contain an active PPIase center. Our observations suggest that the cis-trans isomerization of the L11 PS22 is a universal event required for an efficient turnover of trGTPases throughout the translation process.

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Figure 1: The conserved PS22 is necessary for protein synthesis.
Figure 2: PS22 isomerization for the L11-L12 connection.
Figure 3: Identification of PS22 as the interaction site of the L11 NTD with EF-G.
Figure 4: EF-G has in vitro PPIase activity with the catalytic center facing PS22.
Figure 5: Physiological effects of PPIase activity on EF-G and total polypeptide synthesis.
Figure 6: PPIase activity is common among trGTPases with a conserved catalytic center.
Figure 7: Switch-and-latch model.

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Acknowledgements

We are very grateful to J. Frank and Y. Hashem for their critical discussions and great efforts in polishing the manuscript. We thank J. Wang, J. Zhou, J. Lin, Y. Tao and J. Lou for their technical support and help. We thank Y. Feng for NMR analysis, G. Liu and J. Xue for preparing some of the proteins and W. Zhang for creating the movie. This work was supported by grants from the Major State Basic Research Development Program of China (2012CB911001 and 2010CB834201), the National Natural Science Foundation of China (31170756) and the Novo Nordisk-Chinese Academy of Sciences Research Foundation.

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Authors and Affiliations

  1. National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China

    Li Wang, Fang Yang, Dejiu Zhang, Zhi Chen, Rui-Ming Xu, Weimin Gong & Yan Qin

  2. School of Life Sciences, University of Science and Technology of China, Hefei, China

    Li Wang, Weimin Gong & Yan Qin

  3. School of Life Sciences, University of Science and Technology of China, Hefei, China

    Knud H Nierhaus

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Contributions

Y.Q., K.H.N., W.G. and R.-M.X. designed the experiments. L.W., F.Y. and D.Z. conducted the experiments. Z.C. developed the molecular dynamics simulation. Y.Q. and L.W. analyzed the data. Y.Q., L.W. and K.H.N. wrote the manuscript.

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Correspondence to Yan Qin.

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Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–3, Supplementary Tables 1–6 and Supplementary Methods (PDF 657 kb)

Supplementary Movie

The Switch-and-Latch model. The movie demonstrates the conformational change of L11–L12-CTD complex between the open and the closed states. Open state (L11 in magenta, L12 in yellow, the proline switch trans-PS22 in green and loop 62 in cyan) is represented by the L11 on the ribosome•RF2 complex (PDB 2X9S, chain K) and L12-CTD in its isolated form (PDB 1CTF). Closed state (L11 in red, L12 in yellow, cis-PS22 in green and loop 62 in cyan) is presented by the L11 on the ribosome•EF-G•GDP•fusidic acid complex (PDB 2WRL, chain K) and L12 from the same complex (PDB 2WRL, chain L). The model was built based on MD simulation of L11-NTD. The animation is generated by the Morph Server and PyMol. (WMV 1970 kb)

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Wang, L., Yang, F., Zhang, D. et al. A conserved proline switch on the ribosome facilitates the recruitment and binding of trGTPases. Nat Struct Mol Biol 19, 403–410 (2012). https://doi.org/10.1038/nsmb.2254

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