Reconstitution of the recombinant human γ-tubulin ring complex
Martin Würtz, Anna Böhler, Annett Neuner, Erik Župa, Lukas Rohland, Peng Liu, Bram J. A. Vermeulen, Stefan Pfeffer, Sebastian Eustermann, Elmar Schiebel
Abstract
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.