Velocity Fluctuations in Kinesin-1 Gliding Motility Assays Originate in Motor Attachment Geometry Variations.

Motor proteins such as myosin and kinesin play a major role in cellular cargo transport, muscle contraction, cell division, and engineered nanodevices. Quantifying the collective behavior of coupled motors is critical to our understanding of these systems. An excellent model system is the gliding motility assay, where hundreds of surface-adhered motors propel one cytoskeletal filament such as an actin filament or a microtubule. The filament motion can be observed using fluorescence microscopy, revealing fluctuations in gliding velocity. These velocity fluctuations have been previously quantified by a motional diffusion coefficient, which Sekimoto and Tawada explained as arising from the addition and removal of motors from the linear array of motors propelling the filament as it advances, assuming that different motors are not equally efficient in their force generation. A computational model of kinesin head diffusion and binding to the microtubule allowed us to quantify the heterogeneity of motor efficiency arising from the combination of anharmonic tail stiffness and varying attachment geometries assuming random motor locations on the surface and an absence of coordination between motors. Knowledge of the heterogeneity allows the calculation of the proportionality constant between the motional diffusion coefficient and the motor density. The calculated value (0.3) is within a standard error of our measurements of the motional diffusion coefficient on surfaces with varying motor densities calibrated by landing rate experiments. This allowed us to quantify the loss in efficiency of coupled molecular motors arising from heterogeneity in the attachment geometry.