Open Biology (Feb 2021)

Reconstitution of the recombinant human γ-tubulin ring complex

  • Martin Würtz,
  • Anna Böhler,
  • Annett Neuner,
  • Erik Zupa,
  • Lukas Rohland,
  • Peng Liu,
  • Bram J. A. Vermeulen,
  • Stefan Pfeffer,
  • Sebastian Eustermann,
  • Elmar Schiebel

DOI
https://doi.org/10.1098/rsob.200325
Journal volume & issue
Vol. 11, no. 2

Abstract

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Cryo-electron microscopy recently resolved the structure of the vertebrate γ-tubulin ring complex (γ-TuRC) purified from Xenopus laevis egg extract and human cells to near-atomic resolution. These studies clarified the arrangement and stoichiometry of γ-TuRC components and revealed that one molecule of actin and the small protein MZT1 are embedded into the complex. Based on this structural census of γ-TuRC core components, we developed a recombinant expression system for the reconstitution and purification of human γ-TuRC from insect cells. The recombinant γ-TuRC recapitulates the structure of purified native γ-TuRC and has similar functional properties in terms of microtubule nucleation and minus end capping. This recombinant system is a central step towards deciphering the activation mechanisms of the γ-TuRC and the function of individual γ-TuRC core components.

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