In order to understand biological phenomena in cell systems, we need to determine the detailed mechanisms that express the functions of individual single protein molecules. This is not speculation from static structural information but time-resolved dynamic conformational ones from individual protein molecules. Recent progress in imaging the motions of individual single protein molecules in living cells has been achieved with several single-molecular techniques and systems by using visible light. In particular, single-molecule fluorescence resonance energy transfer (single-molecule FRET) is one of the few methods available for measuring nanometer-scale distance and the intermolecular orientation between two fluorophores. However, measuring intramolecular structural changes of single protein molecules with single-molecule FRET is very difficult due to the lack of both monitoring precision and stability of the signal intensity in physiological conditions. We proposed that one method for improving the positional decision accuracy is to shorten the light wavelength, e.g., x-rays, electrons, neutrons, and other accelerated ion probes (YC SASAKI, PRB 2000, PRL 2001, PRB 2004).
Recently, we succeeded in observing picometer-scale Brownian motions of individual membrane protein (bR, KcsA(Cell 2009), KvAP), antigen-antibody interactions, peptide/ MHC complex for T cell activation, ATP ligand protein (Myosin, Chaperonin) and monitoring super-weak force (pN) field with our single molecular detection system using x-rays, called Diffracted X-ray Tracking (DXT). We observed the rotating motions of individual nanocrystals, each of which was linked to a specific site in an individual protein molecule as shown in the following figure.