Structural Basis of Dynamic Instability
The dynamic behavior of microtubules (MTs) is essential to their functions and a large number of cellular factors regulate it during the cell cycle. We aim to define the structural basis of dynamic instability, the essential process that, fueled by GTP hydrolysis, leads to switching between MT growth and depolymerization, and that is inhibited by chemotherapeutics like taxol. Our original studies of tubulin bound to taxol in polymerized, straight protofilaments, obtained by electron crystallography established the structural basis of nucleotide exchange and polymerization-coupled hydrolysis. Our lab later obtained two structures proposed to mimic intermediates in the assembly and disassembly of microtubules that illustrate the conformational consequences of the nucleotide state and how they relate to longitudinal and lateral assembly.
More recently our studies have centered on defining the conformational changes within the microtubule upon GTP hydrolysis. Although cryo-EM is ideal to study MTs, structures had been limited until recently to ~8-10 Å resolution. Through the optimization of data collection and image processing, we produced structures at ~5 Å resolution for three MT states: stable MTs bound to GMPCPP, dynamic MT (where GTP has been hydrolyzed to GDP), and MTs stabilized by taxol. We used Rosetta to generate low energy ensembles to fit each MT map and ultimately generated consensus models that could be compared to define the changes with nucleotide state and taxol binding Our studies showed that GTP hydrolysis results in a compaction at the interdimer longitudinal interface that buries the E-site nucleotide and a conformational change in alpha tubulin that generates strain in the MT lattice. Taxol appears to allosterically inhibit these changes
We are now interested in the study of cellular factors that, by interacting with the dynamic ends of microtubules, are able to modify the behavior of these polymers.
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Eva Nogales, Kenneth H. Downing, Linda A Amos, Jan Löwe. Tubulin and FtsZ form a distinct family of GTPases. Nat Struct Biol. 6, 451-8, 1998.
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Hong-Wei Wang and Eva Nogales. The nucleotide-dependent bending flexibility of tubulin regulates microtubule assembly. (2005) Nature. 435, 911-915.
Gregory M. Alushin, Gabriel C. Lander, Elizabeth H. Kellogg, Rui Zhang, David Baker, and Eva Nogales (2014) High resolution microtubule structures reveal the structural transitions in αβ–tubulin upon GTP hydrolysis Cell, 157(5)1117-29.