Catalysis of the microtubule on-rate is the major parameter regulating the depolymerase activity of MCAK

Abstract

The kinesin-13, MCAK, is a critical regulator of microtubule dynamics in eukaryotic cells1. We have functionally dissected the structural features responsible for MCAK’s potent microtubule depolymerization activity. MCAK’s positively charged neck enhances its delivery to microtubule ends, not by tethering the molecule to microtubules during diffusion, as commonly thought, but by catalyzing the association of MCAK to microtubules. On the other hand, this same positively charged neck slightly diminishes MCAK’s ability to remove tubulin subunits once at the microtubule end. Conversely, dimerization reduces MCAK delivery but improves MCAK’s ability to remove tubulin subunits. The reported kinetics for these events predict a non-specific binding mechanism that may represent a paradigm for the diffusive interaction of many microtubule binding proteins.

Prime movers: the mechanochemistry of mitotic kinesins

“…Dimerisation increases catastrophase activity but is not required for it 125,176 ; as KIF2 monomers are effective. The proximal part of the N-terminal neck is positively charged and this accelerates the initial recruitment of the motor to the microtubule 54,176 .…”

“…The proximal part of the N-terminal neck is positively charged and this accelerates the initial recruitment of the motor to the microtubule 54,176 . The next part of the N-terminal neck folds around and makes a helix that anneals to this KVD finger 175 .…”

Kif2C Minimal Functional Domain Has Unusual Nucleotide Binding Properties That Are Adapted to Microtubule Depolymerization

“…The MT depolymerization cycle we briefly described above emphasizes the role of direct MT-end binding of Kif2C-ATP to start a productive MT depolymerization process. But because ATP-bound Kif2C-(sNϩM) has submicromolar binding affinity for the MT lattice, it also binds to the lattice (18,49). Diffusion then occurs and assists locating Kif2C-(sNϩM) to MTs ends.…”

“…The average diffusion time of a monomeric Kif2C construct on MTs is 0.25 s (18). The maximum distance traveled during that time may be deduced from the kinesin diffusion coefficient on MTs.…”

“…Remarkably, by contrast with most motile kinesins, monomeric constructs of kinesin-13 proteins are functional. Together with the conserved motor domain, the Neck, a ϳ70-residue extension N-terminal to the motor domain, is sufficient for MT depolymerization (18,19).…”