Solid-State NMR Reveals a Bifunctional Disordered Linker That Couples Katanin Microtubule Anchoring to Condensate-Activated Severing

Microtubule severing by katanin drives rapid cytoskeletal remodeling for cell fate decision, yet how the catalytic subunit p60 (Kp60) recognizes the microtubule lattice to achieve efficient severing remains unresolved. Here, we report that the N-terminal intrinsically disordered region (IDR) of Kp60 is an essential bifunctional module for severing activity. Using solid-state NMR (ssNMR), we identified a microtubule-binding domain (MTBD) within the IDR which directly engages the lattice to guide substrate anchoring. Surprisingly, the same IDR also enables Kp60 phase separation. Our ssNMR analyses of the condensed phase pinpointed three discrete interaction regions within the IDR which cooperatively orchestrate this multivalent assembly. Importantly, Kp60 phase separation was associated with markedly increased ATPase activity, supporting a functional link between condensation and enzymatic activation. Using function-separation-mutation analyses, our studies show both microtubule binding and condensation are essential for severing: microtubule targeting provides spatial specificity, while condensation provides the catalytic threshold through local enrichment. Thus, the Kp60 IDR constitutes a bifunctional interface that integrates substrate recognition with enzymatic robustness, thereby establishing phase separation as a fundamental and efficient regulator of microtubule dynamics and cytoskeletal remodeling.