Abstract
The function of proteins is tightly related to their structures, and thus, the detailed structure of proteins has been studied. However, the obtained structures are essentially static, whereas proteins are dynamic in nature. Dynamic behavior of proteins in action has been studied by single-molecule fluorescence microscopy but protein molecule themselves are invisible in the observations. Hence, the simultaneous assessment of structure and dynamics has long been infeasible, meaning that we have to infer how proteins operate to function from gleaned data with significant resolution gaps. Directly visualizing functioning protein molecules at high spatiotemporal resolution has, therefore, been a “holy grail” for structural biology and single-molecule biophysics. To materialize this long-quested dream, Paul Hansma's group and my group independently commenced around 1993 to develop high-speed atomic force microscopy (HS-AFM). Various devices and techniques, such as small cantilevers, high-speed scanners, and active damping and feedback control techniques, have been developed. After lengthy efforts, HS-AFM with low-invasive performance now comes of age (Ando et al., Prog. Surf. Sci. 83, 337-437, 2008). The imaging studies conducted thus far cover a wide range of targets; myosin V walking on an actin filament (Kodera et al., Nature 468, 72-76, 2010), rotary catalysis of rotorless F(1)-ATPase (Uchihashi et al., Science 333, 755-758, 2011), bacteriorhodopsin in response to light (Shibata et al., Nature Nanotechnol. 5, 208-212, 2010), and so on. The dynamic images captured have clearly demonstrated that this new microscopy is a powerful approach to revealing dynamic process and structure dynamics of protein molecules in great detail. There seems no doubt that continued exploration of countless biological phenomena will be carried out using HS-AFM, while at the same time the next generation of HS-AFM techniques will be developed to visualize dynamic events occurring in live cells.