The force required to remove tubulin from the microtubule lattice

Abstract

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 unknown, as 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. Consistent with this force, we show that several kinesins can also extract tubulin. 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, suggesting that severing proteins such as spastin and katanin use an unfoldase mechanism. Our results reveal the response of tubulin to mechanical force and advance our understanding of severing enzymes and microtubule stability.

Approaches to visualize microtubule dynamics in vitro

“…Кроме того, на основе этого подхода можно легко реализовать одновременное наблюдение отдельных молекул, взаимодействующих с микротрубочкой, с помощью флуоресцентной микроскопии [41]. Этот метод был применён той же группой в работах [42] и [43]. Статья [42] посвящена изучению парадокса, связанного с активностью рассекающие микротрубочки ферментовспастина, катанина, фитжетина.…”

The effect of motor-induced shaft dynamics on microtubule stability and length

“…Only recently it has been discovered that tubulin can exchange all along the microtubule shaft (26) leading to ‘GTP-tubulin islands’ (27). The molecular mechanism responsible for island formation is not yet understood (28), but mechanical forces and severing enzymes like spastin and katanin can enhance tubulin exchange (26, 29–31). Microtubules stop shrinking and resume growth preferentially at such islands (29, 30, 32, 33).…”