Directional loading of the kinesin motor molecule as it buckles a microtubule.
Single kinesin motor molecules were observed to buckle the microtubules 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 substrate via biotin-streptavidin bonds, while the plus end of the microtubule was free to interact with motors adsorbed at low density to the substrate. A statistical analysis of the length of microtubules buckled by single motors showed a decreasing probability of buckling for loads greater than 4-6 pN parallel to the filament. This is consistent with kinesin stalling forces found in other experiments. A detailed analysis of some buckling events allowed us to estimate both the magnitude 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 results 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 component speeds up the motor but does no net work, perpendicular force acts as a mechanical catalyst for the reaction. A simple explanation is that 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.
Gittes
F
F
Department of Physiology and Biophysics, University of Washington, Seattle 98195-7290, USA.
Meyhöfer
E
E
Baek
S
S
Howard
J
J
D000818:Animals; D001696:Biomechanical Phenomena; D055592:Biophysical Phenomena; D001703:Biophysics; D002417:Cattle; D004548:Elasticity; D066298:In Vitro Techniques; D016547:Kinesins / Q000502:physiology*; D008870:Microtubules / Q000502:physiology* / Q000648:ultrastructure; D008954:Models, Biological; D009068:Movement / Q000502:physiology; D013816:Thermodynamics
D016547:Kinesins