Structures of kinesin motor proteins

Abstract

Almost 25 years of kinesin research have led to the accumulation of a large body of knowledge about this widespread superfamily of motor and nonmotor proteins present in all eukaryotic cells. This review covers developments in kinesin research with an emphasis on structural aspects obtained by X-ray crystallography and cryoelectron microscopy 3-D analysis on kinesin motor domains complexed to microtubules.

Figure 1

Number of kinesin motor domain structures in the PDB. The bars represent cumulative numbers of structures, and the corresponding years refer to the date of deposition.

Figure 2

Kinesin-1 in x-ray structure analysis and cryo-EM image reconstruction. Red frames outline the dimer structure in each panel (A) Ribbon diagram of the structure of RnK379, a dimeric construct from rat conventional kinesin (PDB-ID 3kin). The construct comprises the motor domain with neck linker and the first half of the neck helix that forms a coiled coil (Kozielski et al. 1997). (B) Negatively stained image of a tubulin sheet decorated with dimeric RnK379 in the presence of AMP-PNP. Both heads touch the tubulin surface along one protofilament as modelled in panel C. In lucky cases these dimers bind in register and form a regular 16-nm repeat. (D) The same binding pattern is also directly visualized by high-resolution shadowing on sparsely decorated tubulin sheets. Left and right panel are identical images with dimers marked in the right panel (Eg5 dimers: Krzysiak et al., 2006). (E) Typically kinesin-1 dimers bind stochastically and therefore helically averaged 3-D reconstructions loose the connection between trailing and leading head (Hoenger et al., 1998 Hoenger et al., 2000). The density of the bridging coiled-coil neck is very low and usually not directly visible in cryo-electron micrographs.

Figure 3

Kinesin-microtubule complex derived from 9Å resolution cryo-EM analysis of microtubules decorated with a monomeric construct (K349) from human conventional kinesin (Figure adapted from Sindelar and Downing [2007]).

Figure 4

Docking and undocking of the neck linker controlled by the switch-2 cluster. Kinesin-1 motor domain structures representing the “ADP state” (A, human KHC, PDB-ID 1bg2) and the “ATP state” (B, rat KHC, PDB-ID 2kin) are shown as cartoon models with semitransparent surface representations of the switch-2 cluster (green) and the “core” domain (motor domain without switch-2 cluster and helix α6; grey). In the “ADP state” (A), the switch-2 cluster prevents binding of the neck linker to the core domain; neck linker and neck are mainly disordered and invisible. In transition to the “ATP state” (B), the switch-2 cluster moves up and opens a binding groove for the neck linker (β9, β10). Note that the rat KHC structure (B) represents the “ATP state” although it has ADP bound, demonstrating that without microtubules the nucleotide is not sufficient to determine the conformation of the motor domain.

Figure 5

Sideview of a microtubule protofilament complexed with a dimeric kinesin rKS379 MD construct from rat conventional kinesin with an SH3 domain inserted between the neck linker and the neck helix of each head (white circles). The SH3 domains serve as a label to identify the position of the neck linkers in cryo-EM image reconstructions (Skiniotis et al. 2003). The trailing head is in an ATP configuration with the neck-linker locked into a plus-end directing position. The leading head shows a nucleotide free configuration with a flexible neck-linker that is pulled backwards by the coiled-coil connection to the trailing head. Nucleotides are displayed as spheres and are GDP in β-tubulin, and GTP in α-tubulin. Taxol is seen as ball-&-stick at the inner tube face of the protofilament.