HIGH-SPEED ATOMIC FORCE MICROSCOPY FOR DYNAMIC BIOLOGICAL IMAGING - Life at the Nanoscale: Atomic Force Microscopy of Live Cells - page 160

Chapter 8
HIGHSPEED ATOMIC FORCE MICROSCOPY
FOR DYNAMIC BIOLOGICAL IMAGING
Takayuki Uchihashi and Toshio Ando
Department of Physics, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan,
and Core Research for Evolutional Science and Technology (CREST) of the Japan Science and
Technology Agency, Sanban-cho, Chiyoda-ku, Tokyo 102-0075, Japan
[e-mail]
8.1 INTRODUCTION
Proteins are inherently dynamic molecules that undergo structural changes
and interactions with other molecules over a wide timescale range, from
nanoseconds to milliseconds or longer. 1 Protein motions play an important
biological role in the assembly into protein complexes, ligand binding and
enzymatic reactions. Therefore, understanding the dynamic behaviour
of a protein is a requisite for gaining insight into biological processes.
Experimental determination of protein structures has been made using X-ray
crystallography and nuclear magnetic resonance. However, dynamic changes
in protein molecules usually occur spontaneously and unsynchronously and
thus are dificult to detect using these ensemble-average methods.
Recent advances in single-molecule luorescence microscopy have allowed
us to determine the localization of individual protein molecules with high
accuracy. This enables the precise measurement of translational or rotational
motions of individual luorophores attached to biological molecules and, in
some cases, the measurement of the association and dissociation reactions
of biological molecules. Single-molecule luorescence resonance energy
transfer measurement is a powerful approach to analyzing intramolecular
and intermolecular interaction dynamics in proteins. Thus, the “directness”