The force required to remove tubulin from the microtubule lattice by pulling on its α-tubulin C-terminal tail

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. Tubulin, the building blocks of microtubules, can be removed from the microtubule wall by mechanical forces. Using single-molecule methods, the authors show that tubulin partially unfolds prior to its removal and determined the tubulin-extraction force.