Figure 1.
Structural comparison of kinesin-1 and Ncd.
(a) Dimer structures of kinesin-1 (PDB ID 3KIN) in ADP-ADP state in the absence of MT. (b) Dimer structure of the symmetric Ncd dimer (PDB ID 1CZ7). Note that the Ncd motor lacks the neck-linker region. (c) Superposition of the MHs of kinesin-1 (red) and Ncd (blue). Between the MHs of the two motors Cα-rmsd is 1.06 Å, and there is 41% sequence identify. (d) Native contact maps of the two motors, upper left for Ncd and lower right for kinesin-1. Note that the patterns of the contact map between the two MH domains enclosed in black box are very similar, and that the differences are in structural motifs peripheral to MH.
Figure 2.
Conformational transition between symmetric and asymmetric conformers of Ncd dimer in the absence of MTs.
(a) Ncd motor in the symmetric and asymmetric states. (b) Head-stalk contacts present in the symmetric (green) and asymmetric state (red) shown for a monomer. These two sets of contacts cannot be satisfied simultaneously. Residue contact maps for (c) symmetric and (d) asymmetric conformations. The difference in the head-stalk contacts in the asymmetric conformer is highlighted in the contact map by using circles.
Figure 3.
Dynamics between symmetric and asymmetric states of Ncd in solution.
(a) Distribution of the inter-residue distance between E567 of the two monomers calculated from simulation show two peaks corresponding to the symmetric and asymmetric states with representative Ncd structures for each peak position. (b) Two dimensional free energy diagram color-coded in kBT unit. Energetically possible transitions are between symmetric and one of the two asymmetric states (A or B). (c) Low frequency modes from PCA lend support to the distortion dynamics that drives the Ncd dimer from its symmetric to its asymmetric state. Residue-wise vector represents one of the two low frequency modes.
Figure 4.
Correlation between symmetric-asymmetric transition and fluctuation at NT binding region.
(a) Head-stalk contact region and NT binding pocket in Ncd motor are marked on the Ncd structure. (b) Disruption of the head-stalk contact induces disruption of NT binding pocket. Shaded in grey are the anti-correlated signals between the number of head-stalk contacts and the RMSD of NT binding pocket. (c) Different elements of NT binding pocket region in Ncd motor are marked: P-loop (orange), switch I (red) and switch II (yellow). Structural differences of these motifs are compared between the two states. For symmetric state, we use blue color for all elements. Switch I region reveals maximum deformation. (d) RMSD distribution of P-loop, switch I and switch II are shown for both the states. Note the noticeable difference for switch I region.
Figure 5.
ATP dependent population change and minus-end directed power stroke of Ncd on MT track.
(a) Distributions of the angle (θ), among LEU296 in the unbound monomer and D424, E567 in the bound monomer, calculated from simulation for φ-ADP state on MT. The structure of Ncd with θ∼25° (peak position) is depicted on the MTs. (b) The angle (θ) distribution calculated from simulation for the ATP-ADP state. The structure of the Ncd on the MT with θ∼150° (peak position) is shown on the top. Change from φ-ADP to ATP-ADP state (a→b) implies the minus-end-directed power stroke.
Figure 6.
Mechanochemical cycle for Ncd motor.
In solution, Ncd exists in (i) symmetric and two asymmetric states (either (ii) or (ii)’). On the MTs, Ncd in φ-ADP state exists predominantly in (iii) symmetric plus-end directed conformer. ATP binding stabilizes (iv) asymmetric conformer, which gives rise to minus-end-directed stroke. Finally, the ATP hydrolysis returns the Ncd to the ADP-ADP state that has weak MT-binding affinity; thus Ncd dissociates from MT to the solution. The free energy (F(z)) profile of the power stroke step which is calculated by taking stalk tip position (z) on MT as order parameter is also shown in the same figure.