Preprint Highlight: Detyrosination enrichment on microtubule subsets is established by the interplay between a stochastically acting enzyme and microtubule stability.

Microtubule detyrosination, the post-translational removal of a tyrosine residue on α-tubulin, marks a stable and rare subpopulation of microtubules. Vasohibin (VASH) is the main enzyme responsible for α-tubulin detyrosination and it binds to α-tubulin on polymerized microtubules. However, the dynamics underlying detyrosination remain unknown. The authors use quantitative super-resolution microscopy, single molecule tracking, and computational modeling to explore microtubule detyrosination. VASH engages with microtubules in a stochastic fashion and detyrosination depends on VASH concentration on single microtubules. As VASH accumulation requires microtubule stability, detyrosination occurs via a stabilization-detyrosination feedback mechanism. This work provides important and novel mechanistic insight into microtubule detyrosination and serves as an example for studying other microtubule post-translational modifications essential for cellular homeostasis.