The assembly of microtubules and drug-induced tubulin polymers : an X-ray diffraction and cryo-electron microscopy study

• Main Author: Morena, Evangelina Nogales de la
• Published: Keele University 1992
• Subjects: 571.4; Biophysics
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.317597

Synchrotron Radiation (SR) X-ray solution scattering/diffraction and cryo-electron microscopy have been used to characterise the structure of tubulin polymers assembled in a variety of biochemical conditions. Cryo-microscopy has been used to image individual structures and to interpret the X-ray diffraction profiles from the protein structures in solution. The time-resolved X-ray data provided by SR has been analysed using correlation techniques in order to elucidate the structural pathway of reactions triggered by temperature changes. The results can be summarised as follows: • The structure and temperature-induced assembly of microtubules polymerised from microtubule protein (MTP), and purified tubulin (TB) preparations are very similar. Differences in the assembly process concern the existence of prenucleation events, linked to the presence of cold structured aggregates, only present in the MTP solutions. The only structural difference is the presence of MAPs attached to the wall of MTP microtubules. When TB microtubules were assembled in the presence of taxol the surface lattice was preserved but the diameter was smaller, corresponding to an average loss of one proto filament. • The vinblastine-induced self-assembly of TB can give rise to both, double-coiled spirals that aggregate in a non-ordered fashion, and paracrystalline structures where the repeating motif is a single-coiled spiral. The type of polymer is mainly detennined by the amount of magnesium ion present in the preparation. Concentrations higher than 7.5 mM result in double coils similar to those found in MAPs-containing preparations, pointing to a similarity of effect possibly due to the positive charge of both MAPs and magnesium ions. At any magnesium concentration, a temperature change induced a reversible change in the pitch of the spirals, reflecting a temperature-induced confonnational change in the tubulin subunit.