A Biosynthetic Approach to Studying Multiple Motor Complexes

Abstract

Collective motor dynamics drives important cellular processes ranging from muscle contraction to spindle organization to vesicle trafficking. Although the biomechanical and biochemical properties of individual motors have been widely studied, how motors coordinate their motility when attached to the same cargo is largely unknown. We are developing a biosynthetic approach to generate multi-motor assemblies whose biological properties can be examined in vitro and in cells. To do this, we have assembled a “toolbox” of protein components consisting of scaffolds and linkers. We have characterized scaffold proteins of different lengths that allow for specific separation distances between the components. We have characterized four different linker systems that enable constitutive or regulated attachment of individual motors to scaffolds. Thus, our biosynthetic approach can be used to generate multiple motor complexes with absolute control over motor type, separation, and number. The motility properties of these complexes can then be studied in vitro and in live cells to determine the structural and mechanical features that enable kinesin-1 motors to work collectively. This approach is applicable to other biological questions such as the generation of complex signaling networks as well as the assembly of artificial biological systems for engineering applications.