生体分子機能の１分子蛍光イメージング

抄録

Single-molecule imaging is a useful technique for analyzing functions and interactions of protein molecules. I will show some instances how this technique is applied to the biological studies. The first example is the study of chaperonin assisted protein folding. GroEL mediates the folding of nascent or denatured proteins in the E.coli collaborating with co-chaperonin GroES. We visualized GroEL-GroES interaction at the single molecule level [1]. Release of GroES from GroEL occurred after a rag period (–3s), that was not recognized in previous bulk-phase studies. Furthermore, we succeeded in observing the refolding of denatured GFP in the GroEL-GroES complex, and found that GFP could not start to refold for 3s after GroES binding. This observation suggests the presence of a new kinetic intermediate "cis ATP*-complex" in the GroEL-GroES reaction pathway. It is important for the efficient encapsulation of nonnative protein into the GroEL cavity. The second example is the transport of mRNA within a living cell. Fluorescently labeled mRNA was injected into the nuclei of living cells and was visualized by fluorescence microscopy. The injected mRNA molecule were in equilibrium of two states, Brownian motion with diffusion coefficients of 0.2 μm²/s and corralled in a restricted area for 20 s. These results suggested that mRNA travels from the site of synthesis to nuclear pore by diffusion process. REFERENCES [1] Taguchi, H., T. Ueno, H. Tadakuma, M. Yoshida, and T. Funatsu (2001) Nature Biotechnol. 19: 861-865. [2] Ueno T., H. Taguchi, H. Tadakuma, M. Yoshida and T. Funatsu (2004) Molecular Cell, 14: 423-434. [J Physiol Sci. 2006;56 Suppl:S59]