Cooperative Movement Of Wild-type Kinesin And Velocity-deficient Mutants

Abstract

In the classic sliding filament assay, the molecular motor kinesin exhibits processive movement on microtubules with a velocity that is invariant over a large range of motor concentrations. This indicates that kinesin motors move synchronously at high density, but studies examining the motility of 2–3 kinesin motors have shown a surprising lack of synchronization. These results together led us to believe that kinesin motors under high density conditions can pull one another off the microtubule track, accelerating dissociation. Using a computational model, we can demonstrate that this would enable synchronization of MT movement without complete motor to motor synchronization. To test this experimentally, we combined kinesin dimers containing a mutation in the neck-linker (termed VKN) that elicits a 3-fold reduction in velocity when compared to wild-type motor (5.4μm/min vs. 16.2 μm/min) with wild type motors in the sliding filament assay. No significant amount of microtubule buckling was observed for any mixture of wild-type and mutant motors; even at limiting dilutions, and speckled microtubules moved at the same velocities throughout their length, indicating that the motors behave cooperatively, coordinating their movement through a shared interaction with the microtubule. We plan to examine whether this cooperativity is positive (WT motors accelerating VKN mutant movement) or negative (VKN slowing down WT).