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Structural characterization of a eukaryotic chaperone—the ribosome-associated complex

Nature Structural & Molecular Biology volume 20, pages 23–28 (2013)Cite this article

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Abstract

Ribosome-associated chaperones act in early folding events during protein synthesis. Structural information is available for prokaryotic chaperones (such as trigger factor), but structural understanding of these processes in eukaryotes lags far behind. Here we present structural analyses of the eukaryotic ribosome-associated complex (RAC) from Saccharomyces cerevisiae and Chaetomium thermophilum, consisting of heat-shock protein 70 (Hsp70) Ssz1 and the Hsp40 Zuo1. RAC is an elongated complex that crouches over the ribosomal tunnel exit and seems to be stabilized in a distinct conformation by expansion segment ES27. A unique α-helical domain in Zuo1 mediates ribosome interaction of RAC near the ribosomal proteins L22e and L31e and ribosomal RNA helix H59. The crystal structure of the Ssz1 ATPase domain bound to ATP-Mg2+ explains its catalytic inactivity and suggests that Ssz1 may act before the RAC-associated chaperone Ssb. Our study offers insights into the interplay between RAC, the ER membrane–integrated Hsp40-type protein ERj1 and the signal-recognition particle.

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Figure 1: Structural analyses of RAC.
Figure 2: RAC interaction with the 80S ribosome.
Figure 3: ES27 might contact RAC at the ribosome.
Figure 4: RAC function at the ribosomal tunnel exit.

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Acknowledgements

We thank A. Hendricks (BZH) for excellent technical assistance, C. Siegmann from the BZH Cluster of Excellence CellNetworks crystallization platform for technical support, and M. Rakwalska and R. Zimmermann for stimulating discussion. We thank O. Berninghausen and C. Ungewickell from the Ludwig Maximilians University Munich EM facility for their support, A. Bracher (Max Planck Institute for Biochemistry) for the Zuo1-N plasmid and S. Rospert for help with the ScRAC purification. X-ray data collection was performed at the European Synchrotron Radiation Facility (ESRF), and SAXS data were collected on the X33 beamline at the Deutsches Elektronen Synchotron–European Molecular Biology Laboratory (DESY/EMBL). We thank the staffs of both synchrotrons for excellent support. E.H. and I.S. are investigators of the Cluster of Excellence CellNetworks. G.B. is supported by the Peter and Traudl Engelhorn Foundation. R.B. and G.W. are funded by German Research Foundation (DFG) grant GRK1721. This work was supported by DFG grants to R.B. (FOR967 and SFB594) and I.S. (FOR967 and SFB638).

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  1. Christoph Leidig and Gert Bange: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Chemistry and Biochemistry, Gene Center and Center of Integrated Protein Science Munich (CiPS-M), University of Munich, Munich, Germany

    Christoph Leidig, Stephan Wickles, Gregor Witte & Roland Beckmann

  2. Heidelberg University Biochemistry Center (BZH), Heidelberg, Germany

    Gert Bange, Jürgen Kopp, Stefan Amlacher, Ajay Aravind, Ed Hurt & Irmgard Sinning

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Contributions

C.L., G.B., R.B. and I.S. designed the experiments, analyzed the data and wrote the manuscript. G.B., A.A. and J.K. performed crystallographic analysis. C.L. and S.W. performed cryo-EM analysis. C.L. and G.W. performed SAXS analysis. S.A. and E.H. provided Ct80S ribosomes and cDNA of C. thermophilum. All authors commented on the manuscript.

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Correspondence to Roland Beckmann or Irmgard Sinning.

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Leidig, C., Bange, G., Kopp, J. et al. Structural characterization of a eukaryotic chaperone—the ribosome-associated complex. Nat Struct Mol Biol 20, 23–28 (2013). https://doi.org/10.1038/nsmb.2447

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