Abstract
Microtubule-based transport by the kinesin motors, powered by ATP hydrolysis, is essential for a wide range of vital processes in eukaryotes. We obtained insight into this process by developing atomic models for no-nucleotide and ATP states of the monomeric kinesin motor domain on microtubules from cryo-EM reconstructions at 5-6 Å resolution. By comparing these models with existing X-ray structures of ADP-bound kinesin, we infer a mechanistic scheme in which microtubule attachment, mediated by a universally conserved 'linchpin' residue in kinesin (N255), triggers a clamshell opening of the nucleotide cleft and accompanying release of ADP. Binding of ATP re-closes the cleft in a manner that tightly couples to translocation of cargo, via kinesin's 'neck linker' element. These structural transitions are reminiscent of the analogous nucleotide-exchange steps in the myosin and F1-ATPase motors and inform how the two heads of a kinesin dimer 'gate' each other to promote coordinated stepping along microtubules.
Highlights
Conventional kinesin is the founding member of a superfamily of molecular motors that use the energy of ATP hydrolysis to transport cargo along microtubules, serving essential roles in a wide variety of cellular processes, most notably mitosis and neuronal transport
The current work demonstrates that, while existing models of kinesin's ATP state on microtubules appear to be quite accurate (Parke et al, 2010; Sindelar and Downing, 2010; Chang et al, 2013; Gigant et al, 2013), the motor's nucleotide-free conformation exhibits a closed conformation of the switch loops that was not previously anticipated
This feature has lead to the identification of a novel allosteric pathway mediated by the ‘linchpin’ residue N255
Summary
Conventional kinesin is the founding member of a superfamily of molecular motors that use the energy of ATP hydrolysis to transport cargo along microtubules, serving essential roles in a wide variety of cellular processes, most notably mitosis and neuronal transport. The dimerized motor domains take alternating, eight nanometer steps toward the microtubule plus end, tracking along single protofilaments (Gennerich and Vale, 2009). Underlying this behavior, each motor domain cycles between conformations that are strongly attached to the microtubule (no-nucleotide and ATP-bound) and ones that are weakly attached (ADP-bound). The neck linker connects the C-terminus of the motor domain to the stalk, so that docking is
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