Structural Changes of Microtubules During GTP Hydrolysis Revealed by X-Ray Fiber-Diffraction

Microtubule (MT) is one of the most essential cytoskeletal elements in the eukaryote cells, which supports the mitosis, cell architecture and motility as well as the intracellular transportation. Tubulin dimers in a GTP-binding state (GTP-tubulin) are assembled onto MT-ends and make them stable. However, the following slow hydrolysis of GTP to GDP occurring in β-tubulin is assumed to weaken the lateral interactions among protofilaments and MTs in an uncapped state (MT-ends without GTP-tubulin) disassemble and shrink quickly. Such features of dynamic instability would be closely related to various MT functions. In the present study, we tested how the GTP-hydrolysis was related to the structural changes of MTs by X-ray fiber diffraction analysis (BL45XU, SPring-8, λ=0.09 nm). By comparing diffraction peaks between GMPCPP- and GTP-taxol-MT, we found that 8 nm meridional reflection intensity (arrow in the figure) appeared only in GTP-taxol-MTs, which are assumed to consist mostly of GDP-tubulin. It would reflect some structural differences between GDP- and GTP-tubulin in situ. The change of tubulin-pitch by GTP-hydrolysis was also found. This is the first demonstration to show structural changes of MTs under physiological conditions.View Large Image | View Hi-Res Image | Download PowerPoint Slide