DIRECTIONAL LOADING OF THE KINESIN MOTOR MOLECULE AS IT BUCKLES A MICROTUBULE

Single kinesin motor molecules were observed to buckle the microtubule s along which they moved in a modified in vitro gliding assay. In this assay a central portion of the microtubule was clamped to the glass s ubstrate via biotin-streptavidin bonds, while the plus end of the micr otubule was free to interact with motors adsorbed at low density to th e substrate. A statistical analysis of the length of microtubules buck led by single motors showed a decreasing probability of buckling for l oads greater than 4-6 pN parallel to the filament. This is consistent with kinesin stalling forces found in other experiments. A detailed an alysis of some buckling events allowed us to estimate both the magnitu de and direction of the loading force as it developed a perpendicular component tending to pull the motor away from the microtubule. We also estimated the motor speed as a function of this changing vector force . The kinesin motors consistently reached unexpectedly high speeds as the force became nonparallel to the direction of motor movement. Our r esults suggest that a perpendicular component of load does not hinder the kinesin motor, but on the contrary causes the motor to move faster against a given parallel load. Because the perpendicular force compon ent speeds up the motor but does no net work, perpendicular force acts as a mechanical catalyst for the reaction. A simple explanation is th at there is a spatial motion of the kinesin molecule during its cycle that is rate-limiting under load; mechanical catalysis results if this motion is oriented away from the surface of the microtubule.