Single-molecule FRET: Technique and applications to the studies of molecular machines

Properties of Molecular Machines. Molecular machines are molecule-based devices, typically on the nanometer scale, that are capable of generating physical motions, for example, translocation, in response to certain inputs from the outside such as a chemical, electrical, or light stimulus. A large number of such sophisticated small devices are found in Nature, including the many biological motors discussed in this chapter, such as helicases and polymerases. These tiny nanomachines work in many ways just like an automobile on the highway, and many consume fuels on a molecular level, for instance, through the hydrolysis of adenosine-5ʹ-triphosphate (ATP) molecules, to power their motions on their tracks. As a result, when lacking the required fuel, these nanomachines tend to slow down and even stop, same as a motor vehicle would. In addition, these biological motors often move in a directional manner with variable speeds, and their processivity characteristics can be described by how far they move on their track of a molecular highway, often formed by a biopolymer such as a nucleic acid or actin filament, before taking off at a later time. Motions of individual components within these protein machines, for example, the ribosome which is discussed in great detail throughout this book, are often nicely coordinated like in any sophisticated, larger-scaled mechanical machines. In recent years, details of the composition, stoichiometry, and three-dimensional arrangement of components within many nanomachines have become available, thanks to the ever-increasing number of high-resolution crystal structures that have been solved, which have provided valuable insights into the mechanisms of how these biological motors accomplish their tasks. In the past two decades, researchers have also brought these machines under scrutiny by a number of novel and powerful methods with ultra-high sensitivity, watching their motions one molecule at a time, and have learned a great deal of previously hidden mechanistic details about their action and dynamics, such as the size of the fundamental steps taken by these motorized nanodevices. In a simplified view of the mechanism of action of biological motors, their strokes of physical translocation are powered by processes such as ATP hydrolysis through a modulation of their conformation, thus converting the chemical energy stored in the molecular fuel, in a stepwise fashion, into directed motions.