Building Well-defined Mixed Motor Ensembles With DNA Origami

Cytoplasmic dynein is an essential protein for creating intracellular organization and facilitating intracellular transport of molecules and organelles. During transport, multiple motor proteins work together in ensembles to transport a single load. Motor protein transport has been well studied for individual motors, but force integration and cooperativity of multiple motors on a single load is less understood, despite the proteins’ importance in intracellular transport. DNA origami (“chassis”) structures and total internal reflectance microscopy have allowed us to build synthetic cytoskeletal motor ensembles and observe their motility when hauling a resistive load. These chassis structures allow for motor position and quantity, as well as for resistive forces, on the cargo to be controlled, enabling studies of specific cargo-motor geometries and architectures. In this study, I aim to compare motor position and quantity of an ensemble to the force integration observed when pulling a resistive load. I hypothesize that “in-parallel” arrangements reduce interference when pulling a resistive load when compared to “in-series” arrangements. This thesis sought to improve upon existing methods and establish new techniques for essential steps when investigating motility and force integration of synthetic motor ensembles.