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Solid-state NMR and SAXS studies provide a structural basis for the activation of αB-crystallin oligomers

Nature Structural & Molecular Biology volume 17, pages 1037–1042 (2010)Cite this article

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Abstract

The small heat shock protein αB-crystallin (αB) contributes to cellular protection against stress. For decades, high-resolution structural studies on oligomeric αB have been confounded by its polydisperse nature. Here, we present a structural basis of oligomer assembly and activation of the chaperone using solid-state NMR and small-angle X-ray scattering (SAXS). The basic building block is a curved dimer, with an angle of ∼121° between the planes of the β-sandwich formed by α-crystallin domains. The highly conserved IXI motif covers a substrate binding site at pH 7.5. We observe a pH-dependent modulation of the interaction of the IXI motif with β4 and β8, consistent with a pH-dependent regulation of the chaperone function. N-terminal region residues Ser59-Trp60-Phe61 are involved in intermolecular interaction with β3. Intermolecular restraints from NMR and volumetric restraints from SAXS were combined to calculate a model of a 24-subunit αB oligomer with tetrahedral symmetry.

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Figure 1: Structure of the αB-crystallin core domain dimer and its intermolecular interactions.
Figure 2: Curvature of the α-crystallin domain dimer.
Figure 3: A substrate binding site of αB-crystallin.
Figure 4: Molecular organization of the oligomer.
Figure 5: pH-dependent modulation of a substrate binding site.

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Acknowledgements

The authors thank K. Rehbein and A. Diehl for sample preparation and discussions and D. Svergun for fruitful discussions concerning the SAXS experiments. This work was funded by US National Institutes of Health grant 1R01EY017370. SAXS data on αB10.1 was collected at the Stanford Synchrotron Radiation Lightsource, a national user facility operated by Stanford University on behalf of the US Department of Energy, Office of Basic Energy Sciences. The Stanford Synchrotron Radiation Lightsource Structural Molecular Biology Program is supported by the US Department of Energy, Office of Biological and Environmental Research, and by the US National Institutes of Health, National Center for Research Resources, Biomedical Technology Program. SAXS data collection for full-length αB was supported by European Community–European Molecular Biology Laboratory Hamburg Outstation; Deutsches Elektronen-Synchrotron Hamburg X33 beamline.

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  1. Stefan Jehle and Ponni Rajagopal: These authors contributed equally to this work.

Authors and Affiliations

  1. Leibniz-Institut Für Molekulare Pharmakologie, Berlin, Germany

    Stefan Jehle, Benjamin Bardiaux, Stefan Markovic, Ronald Kühne, Victoria A Higman, Barth-Jan van Rossum & Hartmut Oschkinat

  2. Freie Universität Berlin, Berlin, Germany

    Stefan Jehle, Stefan Markovic & Hartmut Oschkinat

  3. Department of Biochemistry, University of Washington, Seattle, Washington, USA

    Ponni Rajagopal, Joseph R Stout & Rachel E Klevit

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Contributions

S.J. contributed to all aspects of the manuscript; P.R. performed solution NMR experiments and helped to write the manuscript; B.B. performed structure calculations; S.M. did solid-state NMR and SAXS measurements as well as data analysis; R.K. contributed to modeling of C-terminal intermolecular interactions; J.R.S. prepared samples; V.A.H. contributed to assignment strategies, was involved in structure calculations and helped write the manuscript; R.E.K. contributed to the interpretation of results and wrote the manuscript; B.J.v.R. contributed to solid-state NMR measurements, discussed the results and helped to write the manuscript; H.O. designed experimental strategies, contributed to the interpretation of results and wrote the manuscript.

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Correspondence to Rachel E Klevit, Barth-Jan van Rossum or Hartmut Oschkinat.

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Jehle, S., Rajagopal, P., Bardiaux, B. et al. Solid-state NMR and SAXS studies provide a structural basis for the activation of αB-crystallin oligomers. Nat Struct Mol Biol 17, 1037–1042 (2010). https://doi.org/10.1038/nsmb.1891

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