VASH1–SVBP and VASH2–SVBP generate different detyrosination profiles on microtubules

The detyrosination/tyrosination cycle of α-tubulin is critical for proper cell functioning. VASH1–SVBP and VASH2–SVBP are ubiquitous enzymes involved in microtubule detyrosination, whose mode of action is little known. Here, we show in reconstituted systems and cells that VASH1–SVBP and VASH2–SVBP drive the global and local detyrosination of microtubules, respectively. We solved the cryo-electron microscopy structure of VASH2–SVBP bound to microtubules, revealing a different microtubule-binding configuration of its central catalytic region compared to VASH1–SVBP. We show that the divergent mode of detyrosination between the two enzymes is correlated with the microtubule-binding properties of their disordered N- and C-terminal regions. Specifically, the N-terminal region is responsible for a significantly longer residence time of VASH2–SVBP on microtubules compared to VASH1–SVBP. We suggest that this VASH region is critical for microtubule detachment and diffusion of VASH–SVBP enzymes on lattices. Our results suggest a mechanism by which VASH1–SVBP and VASH2–SVBP could generate distinct microtubule subpopulations and confined areas of detyrosinated lattices to drive various microtubule-based cellular functions.