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Vol. 19, Issue 1, 207-215, January 2008

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The Structure of the -Tubulin Small Complex: Implications of Its Architecture and Flexibility for Microtubule Nucleation

Justin M. Kollman^*, Alex Zelter, Eric G.D. Muller, Bethany Fox, Luke M. Rice^*, Trisha N. Davis, and David A. Agard^*

*Department of Biochemistry and Biophysics and the Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158; and Department of Biochemistry, University of Washington, Seattle, WA 98195

Submitted September 10, 2007; Revised October 12, 2007; Accepted October 18, 2007
Monitoring Editor: Tim Stearns

The -tubulin small complex (-TuSC) is an evolutionarily conserved heterotetramer essential for microtubule nucleation. We have determined the structure of the Saccharomyces cerevisiae -TuSC at 25-Å resolution by electron microscopy. -TuSC is Y-shaped, with an elongated body connected to two arms. Gold labeling showed that the two -tubulins are located in lobes at the ends of the arms, and the relative orientations of the other -TuSC components were determined by in vivo FRET. The structures of different subpopulations of -TuSC indicate flexibility in the connection between a mobile arm and the rest of the complex, resulting in variation of the relative positions and orientations of the -tubulins. In all of the structures, the -tubulins are distinctly separated, a configuration incompatible with the microtubule lattice. The separation of the -tubulins in isolated -TuSC likely plays a role in suppressing its intrinsic microtubule-nucleating activity, which is relatively weak until the -TuSC is incorporated into higher order complexes or localized to microtubule-organizing centers. We propose that further movement of the mobile arm is required to bring the -tubulins together in microtubule-like interactions, and provide a template for microtubule growth.

Address correspondence to: David A. Agard (agard{at}msg.ucsf.edu)

Abbreviations used: -TuRC, -tubulin ring complex; -TuSC, -tubulin small complex; FRET, fluorescent resonance energy transfer; SPB, spindle pole body.