Inducing the movement of single kinesin molecules by the asymmetric artificial pertarbation.

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Molecular motors move directionally towards the plus or minus end of microtubules or actin filaments. Kinesin moves towards microtubule plus ends while the kinesin-related Ncd motor moves towards the minus ends. The ヤ neck ユ, the region between the stalk and motor domain, is required for Ncd to move to microtubule minus ends 1, 2, but the mechanism underlying directional motor movement is not understood. Here we show that a single amino acid change in the Ncd neck causes the motor to reverse directions and move with wild-type velocities towards the plus or minus end, thus the neck is functional but directionality is defective. Mutation of a motor core residue that touches the neck residue in crystal structures2, 3 also results in movement in both directions, indicating that directed movement to the minus end requires neck-motor core interactions. Low-density laser-trap assays show that a conformational change or working stroke of the Ncd motor is directional and biased towards the minus end, whereas that of the neck mutant occurs in either direction. The directional bias of the working stroke is thus dependent on neck-motor core interactions. Absence of these interactions removes directional constraints and permits movement in either direction.