X-ray and Cryo-EM structures reveal mutual conformational changes of Kinesin and GTP-state microtubules upon binding

• Published in: The EMBO journal 2015-05, Vol.34 (9), p.1270-1286
• Main Authors: Morikawa, Manatsu, Yajima, Hiroaki, Nitta, Ryo, Inoue, Shigeyuki, Ogura, Toshihiko, Sato, Chikara, Hirokawa, Nobutaka
• Format: Article
• Language: English
• Subjects: Adenosine diphosphate; Adenosine Diphosphate - metabolism; Amino Acid Sequence; Animals; Binding Sites; cryo-electron microscopy; Cryoelectron Microscopy - methods; Crystal structure; Crystallography, X-Ray; Cytoskeleton; EMBO05; EMBO20; Eukaryotes; Guanosine Triphosphate - analogs & derivatives; Guanosine Triphosphate - chemistry; Guanosine Triphosphate - metabolism; kinesin; Kinesin - chemistry; Kinesin - genetics; Kinesin - metabolism; Mice; microtubule; Microtubules - chemistry; Microtubules - metabolism; Microtubules - ultrastructure; Models, Molecular; Molecular Docking Simulation; Molecular Sequence Data; polarized transport in neuron; Protein Conformation; Proteins; X-ray crystallography
• ISSN: 0261-4189; 1460-2075
• DOI: 10.15252/embj.201490588

The molecular motor kinesin moves along microtubules using energy from ATP hydrolysis in an initial step coupled with ADP release. In neurons, kinesin‐1/KIF5C preferentially binds to the GTP‐state microtubules over GDP‐state microtubules to selectively enter an axon among many processes; however, because the atomic structure of nucleotide‐free KIF5C is unavailable, its molecular mechanism remains unresolved. Here, the crystal structure of nucleotide‐free KIF5C and the cryo‐electron microscopic structure of nucleotide‐free KIF5C complexed with the GTP‐state microtubule are presented. The structures illustrate mutual conformational changes induced by interaction between the GTP‐state microtubule and KIF5C. KIF5C acquires the ‘rigor conformation’, where mobile switches I and II are stabilized through L11 and the initial portion of the neck‐linker, facilitating effective ADP release and the weak‐to‐strong transition of KIF5C microtubule affinity. Conformational changes to tubulin strengthen the longitudinal contacts of the GTP‐state microtubule in a similar manner to GDP‐taxol microtubules. These results and functional analyses provide the molecular mechanism of the preferential binding of KIF5C to GTP‐state microtubules. Synopsis Cryo‐EM and crystal structures of nucleotide‐free KIF5C with and without microtubules reveal their mutual conformational changes, providing insight into the preferential binding of KIF5C to GTP‐state microtubules that is crucial for polarized axonal transport in neurons. Loop L11 of KIF5 recognizes and binds to GTP‐state microtubules, triggering conformational changes in both KIF5 and microtubules. Under these conditions, KIF5 acquires a novel rigor conformation that favours effective ADP release. Reciprocally, KIF5 binding causes a switch in the tubulin conformation and strengthens the microtubule lattice. Graphical Abstract Cryo‐EM and crystal structures of nucleotide‐free KIF5C with and without microtubules reveal their mutual conformational changes, providing insight into the preferential binding of KIF5C to GTP‐state microtubules that is crucial for polarized axonal transport in neurons.