Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA.
Harvard Medical School, Boston, MA, USA.
Nat Commun. 2022 Jun 25;13(1):3651. doi: 10.1038/s41467-022-31069-x.
Severing enzymes and molecular motors extract tubulin from the walls of microtubules by exerting mechanical force on subunits buried in the lattice. However, how much force is needed to remove tubulin from microtubules is not known, nor is the pathway by which subunits are removed. Using a site-specific functionalization method, we applied forces to the C-terminus of α-tubulin with an optical tweezer and found that a force of ~30 pN is required to extract tubulin from the microtubule wall. Additionally, we discovered that partial unfolding is an intermediate step in tubulin removal. The unfolding and extraction forces are similar to those generated by AAA-unfoldases. Lastly, we show that three kinesin-1 motor proteins can also extract tubulin from the microtubule lattice. Our results provide the first experimental investigation of how tubulin responds to mechanical forces exerted on its α-tubulin C-terminal tail and have implications for the mechanisms of severing enzymes and microtubule stability.
切断酶和分子马达通过对埋藏在晶格中的亚基施加机械力从微管壁中提取微管蛋白。然而，目前尚不清楚从微管中提取微管蛋白需要多大的力，也不知道亚基被去除的途径。我们使用一种特异性的功能化方法，用光镊对 α-微管蛋白的 C 端施加力，发现需要约 30 pN 的力才能从微管壁中提取微管蛋白。此外，我们发现部分展开是微管蛋白去除的中间步骤。展开和提取力与 AAA 展开酶产生的力相似。最后，我们表明三个驱动蛋白-1 马达蛋白也可以从微管晶格中提取微管蛋白。我们的研究结果首次对微管蛋白如何响应施加在其 α-微管蛋白 C 端尾部的机械力进行了实验研究，对切断酶和微管稳定性的机制具有重要意义。
