Single-molecule and single-particle imaging of molecular motors in vitro and in vivo

Motor proteins are multi-potent molecular machines, whose localisation, function and regulation are achieved through tightly controlled processes involving conformational changes and interactions with their tracks, cargos and binding partners. Understanding how these complex machines work requires dissection of these processes both in space and time. Complementing the traditional ensemble measurements, single-molecule assays enable the detection of rare or short-lived intermediates and molecular heterogeneities, and the measurements of subpopulation dynamics. This chapter is focusing on the fluorescence imaging of single motors and their cargo. It discusses what is required in order to achieve single-molecule imaging with high temporal and spatial resolution and how these requirements are met both in vitro and in vivo. It also presents a general overview and applied examples of the major single-molecule imaging techniques and experimental assays which have been used to study motor proteins.