Possible Intermediate States in the Microtubule Minus-End Directed Movement of the Ncd Stalk

Abstract

The walking step of the microtubule minus-end directed kinesin motor Ncd involves about 75-degree rotation of its coiled-coil stalk. Two crystal structures, 1CZ7 and 1N6M (protein data bank IDs) are related respectively to conformations before (plus-end pointing stalk) and after (minus-end pointing stalk) this movement. Using targeted molecular dynamics we traced the stalk motion between these two structures. We observed that the motor head first rearranges from the 1CZ7 to 1N6M conformation, followed by the travel of the stalk. The 75-degree rotation of the stalk is accompanied by a nearly 20-degree torsion in its coiled coil. Further, the stalk travel can be divided into sub-steps between intermediates at about 17, 38 and 56 degrees from the 1CZ7 conformation that are characterized by the breaking and forming of new salt bridges by Arg335 or Lys336 in the stalk with the charged groups along the alpha-1 domain of the motor head, some of which are not present in the crystal structures. These results suggest that when Ncd is making a step towards the microtubule minus-end, the motor head first changes conformation in a nucleotide-dependent manner that promotes the detachment of the plus-end pointing stalk. This is followed by the stalk rotation that may be broken down into sub-steps between the intermediate states. In case of a diffusive motion, this may be more effective than diffusion over the entire 75-degree range.