Insights into microtubule nucleation from the crystal structure of human γ-tubulin (original) (raw)

Nature volume 435, pages 523–527 (2005)Cite this article

Abstract

Microtubules are hollow polymers of αβ-tubulin that show GTP-dependent assembly dynamics and comprise a critical part of the eukaryotic cytoskeleton. Initiation of new microtubules in vivo requires γ-tubulin, organized as an oligomer within the 2.2-MDa γ-tubulin ring complex (γ-TuRC) of higher eukaryotes1,2,3. Structural insight is lacking regarding γ-tubulin, its oligomerization and how it promotes microtubule assembly. Here we report the 2.7-Å crystal structure of human γ-tubulin bound to GTP-γS (a non-hydrolysable GTP analogue). We observe a ‘curved’ conformation for γ-tubulin–GTPγS, similar to that seen for GDP-bound, unpolymerized αβ-tubulin4. Tubulins are thought to represent a distinct class of GTP-binding proteins, and conformational switching in γ-tubulin might differ from the nucleotide-dependent switching of signalling GTPases. A crystal packing interaction replicates the lateral contacts between α- and β-tubulins in the microtubule5, and this association probably forms the basis for γ-tubulin oligomerization within the γ-TuRC. Laterally associated γ-tubulins in the γ-TuRC might promote microtubule nucleation by providing a template that enhances the intrinsically weak lateral interaction between αβ-tubulin heterodimers. Because they are dimeric, αβ-tubulins cannot form microtubule-like lateral associations in the curved conformation5. The lateral array of γ-tubulins we observe in the crystal reveals a unique functional property of a monomeric tubulin.

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Acknowledgements

We thank S. Murphy for protein purification advice early in the project, the Mullins laboratory for the use of their ultraviolet illuminator, and the Agard laboratory centrosome group for discussions and input. We acknowledge the support of this work by grants from the National Institutes of Health (D.A.A. and T.S.) and the Howard Hughes Medical Institute. H.A. acknowledges support from an NIGMS predoctoral fellowship, and L.M.R. was a Paul Sigler/Agouron Institute fellow of the Helen Hay Whitney Foundation. D.A.A. is a Howard Hughes Medical Institute Investigator.

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Author notes

  1. Hector Aldaz
    Present address: Department of Molecular and Cell Biology, University of California, Berkeley, 16 Barker Hall, Berkeley, California, 94720-3202, USA
  2. Hector Aldaz and Luke M. Rice: *These authors contributed equally to this work

Authors and Affiliations

  1. Howard Hughes Medical Institute and Department of Biochemistry and Biophysics, University of California, San Francisco, 600 16th Street, San Francisco, California, 94143, USA
    Hector Aldaz, Luke M. Rice & David A. Agard
  2. Department of Biological Sciences, Stanford University, Stanford, California, 94305-5020, USA
    Tim Stearns

Authors

  1. Hector Aldaz
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  2. Luke M. Rice
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  3. Tim Stearns
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  4. David A. Agard
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Corresponding author

Correspondence toDavid A. Agard.

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Coordinates and structure factors have been deposited in the Protein Data Bank under accession numbers 1Z5V and 1Z5W. Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

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Aldaz, H., Rice, L., Stearns, T. et al. Insights into microtubule nucleation from the crystal structure of human γ-tubulin.Nature 435, 523–527 (2005). https://doi.org/10.1038/nature03586

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Editorial Summary

Tubulin in the groove

Tubulin proteins have a central role in the life of eukaryotic cells. αβ-Tubulin polymerizes to form the microtubules required for chromosome segregation and organelle positioning. γ-Tubulin initiates microtubule assembly in vivo and is part of a multimeric complex at the centrosome. The crystal structure of human γ-tubulin bound to GTPγS is reported this week, the highest resolution (2.7 Å) structure of any tubulin to date. The structure suggests new ways of thinking about the roles of conformational change and nucleotide binding in microtubule assembly.

The antitumour drug vinblastine is known to target tubulin. Its actual binding site and mechanism of action are unknown but now the X-ray structure of vinblastine bound in a tubulin/protein complex has been determined. Vinblastine introduces a wedge at the junction of two tubulin molecules, thereby interfering with microtubule production and promoting self-association of tubulin molecules into spiral aggregates. A hydrophobic groove on the α-tubulin surface acts both as a binding site for vinblastine and as a point of intermolecular contact in microtubules, so may be an attractive candidate for new microtubule depolymerizing drugs.