Chemistry Reference
In-Depth Information
12
High-speed Atomic Force Microscopy for Nano-visualization
of Biomolecular Processes
Toshio Ando, Takayuki Uchihashi, Noriyuki Kodera, Daisuke Yamamoto,
Masaaki Taniguchi, Atsushi Miyagi, and Hayato Yamashita
12.1
Introduction
Single-molecule observations by fluorescence microscopy provide us with transla-
tional or rotational information about individual fluorescent spots emitted from
fluorophores attached to molecules. Such information is valuable for dissecting
the dynamic behavior of the labeled biomolecules at work. However, it must be
inferred from the observed
fluorescent spots how the labeled molecules are in fact
behaving. In order to understand biomolecular behavior, we should directly observe
single molecules at nanometer spatial and millisecond temporal resolution.
The atomic forcemicroscope (AFM) [1] made it possible to observe the nanometer-
scale world in liquids. Although it can visualize the structure of unstained
biomolecules under physiological solution conditions, it takes minutes to get an
image, far too slow to observe dynamic biomolecular processes. This slow imaging
rate is due to the fact that AFM employs mechanical scanning to detect the sample
height at each pixel. It is quite dif cult to quickly move a mechanical device of
macroscopic size with sub-nanometer accuracy without producing unwanted
vibrations. Various efforts carried out in the past decade have improved the imaging
rate of AFM (e.g. [2 - 8]). Current high-speed AFMcan capture images on video at
30
frames/s with a scan range of
100, without signi cantly
disturbing weak biomolecular interactions [9]. Recent studies have demonstrated
that this new microscope can reveal biomolecular processes such as myosin
V walking along actin tracks (N. Kodera et al., unpublished data) and association/
dissociation dynamics of chaperonin GroEL-GroES that occurs in a negatively
cooperative manner (D. Yamamoto et al., unpublished data). These studies clearly
indicate that high-speed AFM has great potential to reveal how and what structural
changes in individual molecules occur when exercising their physiological functions.
However, high-speed AFM technology is still immature. The imaging rate needs to
250 nm and scan lines of
 
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