Abstract
CAMSAP/Patronins regulate microtubule minus-end dynamics. Their end specificity is mediated by their CKK domains, which we proposed recognise specific tubulin conformations found at minus ends. To critically test this idea, we compared the human CAMSAP1 CKK domain (HsCKK) with a CKK domain from Naegleria gruberi (NgCKK), which lacks minus-end specificity. Here we report near-atomic cryo-electron microscopy structures of HsCKK- and NgCKK-microtubule complexes, which show that these CKK domains share the same protein fold, bind at the intradimer interprotofilament tubulin junction, but exhibit different footprints on microtubules. NMR experiments show that both HsCKK and NgCKK are remarkably rigid. However, whereas NgCKK binding does not alter the microtubule architecture, HsCKK remodels its microtubule interaction site and changes the underlying polymer structure because the tubulin lattice conformation is not optimal for its binding. Thus, in contrast to many MAPs, the HsCKK domain can differentiate subtly specific tubulin conformations to enable microtubule minus-end recognition.
Highlights
Little is known about the native MT ultrastructure of Naegleria, so it is possible that NgCKK could recognise MT minus ends on Naegleria MTs36
To allow a near-atomic resolution investigation of the subtle mechanism(s) at work, we studied NgCKK and human CAMSAP1 CKK domain (HsCKK) MT lattice binding in the context of different MT architectures and used these structures to explain the differences in their MT minus-end recognition properties (Fig. 6)
We found that NgCKK and HsCKK share the same protein fold, they bind at the same intradimer interprotofilament site and, as revealed using a combination of NMR methodologies, share the same intrinsic structural rigidity
Summary
Their end specificity is mediated by their CKK domains, which we proposed recognise specific tubulin conformations found at minus ends. Mutagenesis of residues at the MT-binding interface in the CKK domain disrupted lattice and minus end binding, showing that the same regions of the CKK domain that contact the MT are involved in binding to the minus-end zone Taking these data together, we proposed a model for CAMSAP/ Patronin MT minus end recognition, which is mediated by sensitivity of the CKK domain to a curved sheet-like conformation of tubulin exclusive to MT minus ends. The model suggested that the tighter CKK interaction with β-tubulin disfavours binding at MT plus ends while the looser α-tubulin contacts preferentially accommodate tubulin curvature at minus ends This interaction can occur on the MT lattice, but CKK binding induces distortion of the non-optimal binding site configuration, manifesting as protofilament skew within the polymer
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
