Zhang Rui, Alushin Gregory M, Brown Alan, Nogales Eva
Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
Biophysics Graduate Program, University of California, Berkeley, Berkeley, CA 94720, USA.
Cell. 2015 Aug 13;162(4):849-59. doi: 10.1016/j.cell.2015.07.012. Epub 2015 Jul 30.
Microtubule (MT) dynamic instability is driven by GTP hydrolysis and regulated by microtubule-associated proteins, including the plus-end tracking end-binding protein (EB) family. We report six cryo-electron microscopy (cryo-EM) structures of MTs, at 3.5 Å or better resolution, bound to GMPCPP, GTPγS, or GDP, either decorated with kinesin motor domain after polymerization or copolymerized with EB3. Subtle changes around the E-site nucleotide during hydrolysis trigger conformational changes in α-tubulin around an "anchor point," leading to global lattice rearrangements and strain generation. Unlike the extended lattice of the GMPCPP-MT, the EB3-bound GTPγS-MT has a compacted lattice that differs in lattice twist from that of the also compacted GDP-MT. These results and the observation that EB3 promotes rapid hydrolysis of GMPCPP suggest that EB proteins modulate structural transitions at growing MT ends by recognizing and promoting an intermediate state generated during GTP hydrolysis. Our findings explain both EBs end-tracking behavior and their effect on microtubule dynamics.
微管（MT）的动态不稳定性由GTP水解驱动，并受微管相关蛋白调控，包括正端追踪末端结合蛋白（EB）家族。我们报告了六种MT的冷冻电子显微镜（cryo-EM）结构，分辨率达到3.5埃或更高，这些结构与GMPCPP、GTPγS或GDP结合，聚合后用驱动蛋白运动结构域修饰或与EB3共聚。水解过程中E位点核苷酸周围的细微变化会触发α-微管蛋白围绕“锚定点”的构象变化，导致整体晶格重排和应变产生。与GMPCPP-MT的延伸晶格不同，EB3结合的GTPγS-MT具有紧凑的晶格，其晶格扭曲与同样紧凑的GDP-MT不同。这些结果以及EB3促进GMPCPP快速水解的观察结果表明，EB蛋白通过识别和促进GTP水解过程中产生的中间状态来调节生长中的MT末端的结构转变。我们的发现解释了EBs的末端追踪行为及其对微管动力学的影响。
