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A tightly regulated molecular toggle controls AAA+ disaggregase

Nature Structural & Molecular Biology volume 19, pages 1338–1346 (2012)Cite this article

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Abstract

The ring-forming AAA+ protein ClpB cooperates with the DnaK chaperone system to refold aggregated proteins in Escherichia coli. The M domain, a ClpB-specific coiled-coil structure with two wings, motif 1 and motif 2, is essential to disaggregation, but the positioning and mechanistic role of M domains in ClpB hexamers remain unresolved. We show that M domains nestle at the ClpB ring surface, with both M-domain motifs contacting the first ATPase domain (AAA-1). Both wings contribute to maintaining a repressed ClpB activity state. Motif 2 docks intramolecularly to AAA-1 to regulate ClpB unfolding power, and motif 1 contacts a neighboring AAA-1 domain. Mutations that stabilize motif 2 docking repress ClpB, whereas destabilization leads to derepressed ClpB activity with greater unfolding power that is toxic in vivo. Our results underline the vital nature of tight ClpB activity control and elucidate a regulated M-domain toggle control mechanism.

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Figure 1: Structural dynamics of ClpB upon oligomerization and ATP binding.
Figure 2: ATP binding to AAA-1 leads to protection of M domain in HX experiments.
Figure 3: M-domain motif 1 is near the C-terminal subdomain of neighboring AAA-1.
Figure 4: Characterization of inactive and hyperactive ClpB M-domain variants.
Figure 5: ClpB M-domain mutants are arrested in distinct conformational states.
Figure 6: A salt bridge network between AAA-1 and M-domain motif 2 controls ClpB activity.
Figure 7: Hyperactive ClpB M-domain variants are toxic in vivo.
Figure 8: Regulation of ClpB activity through M domains.

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Acknowledgements

This work was supported by a grant from the Deutsche Forschungsgemeinschaft (BU617-17) to B. Bukau and A. Mogk. We thank T. Ruppert and J. Fiaux for help in HX experiments and L. Guilbride for editing the manuscript. E. Kummer and F. Seyffer were supported by the Hartmut Hoffmann-Berling International Graduate School of Molecular and Cellular Biology. Y. Oguchi was supported by a Humboldt fellowship. M.B. and R.C.W. acknowledge the support of the Klaus Tschira Foundation.

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  1. Yuki Oguchi and Eva Kummer: These authors contributed equally to this work.

Authors and Affiliations

  1. Zentrum für Molekulare Biologie der Universität Heidelberg, Heidelberg, Germany

    Yuki Oguchi, Eva Kummer, Fabian Seyffer, Benjamin Anstett, Regina Zahn, Rebecca C Wade, Axel Mogk & Bernd Bukau

  2. Deutsches Krebsforschungszentrum, Heidelberg, Germany

    Yuki Oguchi, Eva Kummer, Fabian Seyffer, Benjamin Anstett, Regina Zahn, Axel Mogk & Bernd Bukau

  3. Heidelberg Institut für Theoretische Studien, Heidelberg, Germany

    Mykhaylo Berynskyy & Rebecca C Wade

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Contributions

Y.O., E.K., F.S., M.B., R.C.W., A.M. and B.B. conceived and designed experiments. Y.O., E.K., F.S., M.B., B.A. and R.Z. carried out experiments. Y.O., E.K., F.S., M.B., B.A., R.Z., R.C.W., A.M. and B.B. analyzed the data. R.C.W., A.M. and B.B. wrote the manuscript.

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Correspondence to Axel Mogk or Bernd Bukau.

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Oguchi, Y., Kummer, E., Seyffer, F. et al. A tightly regulated molecular toggle controls AAA+ disaggregase. Nat Struct Mol Biol 19, 1338–1346 (2012). https://doi.org/10.1038/nsmb.2441

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