Development of glutathione-coupled cantilever for the single-molecule force measurement by scanning force microscopy.
The accuracy and the fidelity of a single-molecule force measurement largely rely on how the molecule of interest is attached to the solid substrate surface (bead, cantilever, cover glass and etc.). A site-specific attachment of a protein without affecting its structure and enzymatic function has been a major concern. Here, we established a glutathione-coupled cantilever to which any glutathione S-transferase (GST)-fused proteins can be attached in a desired direction. The rupture force between glutathione and GST was approximately 100 pN on average. By using this cantilever, we succeeded in measuring the interaction force between importin alpha and importin beta.
Yoshimura
Shige H
SH
Laboratory of Plasma Membrane and Nuclear Signaling, Graduate School of Biostudies, Kyoto University, Kitashirakawa-oiwake-cho, Sakyo-ku, Kyoto 606-8502, Japan. yoshimura@lif.kyoto-u.ac.jp
Takahashi
Hirohide
H
Otsuka
Shotaro
S
Takeyasu
Kunio
K
D001696:Biomechanical Phenomena; D005978:Glutathione / Q000737:chemistry*; D005982:Glutathione Transferase / Q000737:chemistry; D018625:Microscopy, Atomic Force / Q000379:methods*; D028901:alpha Karyopherins / Q000737:chemistry; D028961:beta Karyopherins / Q000737:chemistry
D028901:alpha Karyopherins; D028961:beta Karyopherins; D005982:Glutathione Transferase; D005978:Glutathione