Laboratory/Faculty

Structural Biology Research Center
Structural Biology Research Center

Leader,Associate Prof.Yoshiyuki Matsuura
Group of Protein Crystallography (Lab for Supra-molecular structures)
*for the research activities of these members of Lab for Supra-molecular Structures, see the pages for the Lab
Leader,Associate Prof.Akihiro Narita
Group of Electron Cryo-microscopy (Lab for Supra-molecular structures)
*for the research activities of these members of Lab for Supra-molecular Structures, see the pages for the Lab
Leader,Visiting Prof.Masatsune Kainosho
Development of techniques of stable isotope labeling for protein NMR study
*for the research activities, see below
Visiting Associate Prof.Yohei Miyanoiri
Development of techniques of stable isotope labeling for protein NMR study
*for the research activities, see below
▶Laboratory HP
Japanese
Masatsune Kainosho Visiting Prof.(left) Yuichiro Maéda Snior Research Fellow

In our research center, by employing various analytical methods in combination (X-ray protein crystallography, electron cryo-microscopy, and protein NMR, among others), we are addressing scientific questions aimed at improving our understanding of the structural dynamics of protein molecules, the structural elucidation of large protein complexes, and the dynamics of protein assembly in the cell. Through these works, we would like to account for the mechanisms of protein function in terms of chemistry and physics, based on protein structure at atomic resolution. Any biological scientists at Nagoya access the leading-edge analytical methods that we have acquired and developed, as well as the large facilities we operate.

Protein NMR Group
Development of techniques of stable isotope labeling for protein NMR study

Because we have obtained atomic-resolution structures by employing X-ray protein crystallography, we tend to view proteins as solid immobile structures. However, in order to understand how proteins work, we need to know how protein structure fluctuates and changes. In the SAIL-NMR methods, developed by Kainosho, minimal numbers of atoms are isotope-labeled, which reduce dramatically the number of NMR peaks, thereby improving the S/N ratio. This enables us then to measure precisely the isomerization of disulfide bonds (Fig. 2a); the rate of rotation of aromatic rings of Tyr or Phe at the 100-1000 msec scale (Fig. 1; Fig. 2b); and other parameters. This rotation is accounted for by the breathing movement of the entire protein molecules, and is sensitive to interactions with other proteins and ligands. We are now able to follow the paths of proton within a protein molecule. This new NMR techniques is now opening up a new field of study, “protein structural dynamics”, and thereby extending our understanding of nature.

Fig.1

Part of the 2D NMR spectrum obtained from a protein that was uniformly isotope-labeled (left) and from a SAIL-protein (right). In the uniformly labeled protein, all the carbon atoms are isotope-labeled, whereas in the SAIL-protein, only Cd’s are labeled. NMR peaks from the SAIL-protein are much sharper and more easily assignable.

Fig.2

Illustrating fluctuations of side chains of a protein molecule. (a) The isomerization of a disulfide bond. (b) The rotation of the aromatic ring.

References

  1. Torizawa et al. (2005) J. Am. Chem. Soc. 127: 12620-6.
  2. Kainosho et al. (2006) Nature 440:52-57.
  3. Ikeya et al. (2006) Magn. Reson. Chem. 44: S152-157.
  4. Takeda et al. (2007) Nat. Protoc. 2: 2896-902.
  5. Takeda et al. (2008) J. Mol. Biol. 380: 608-22.
  6. Takeda et al. (2008) FEBS J. 275: 5873-5884.
  7. Yoshida et al. (2008) J. Mol. Biol. 384: 690-701.
  8. Ikeya et al. (2009) J. Biomol. NMR. 44:261-72.
  9. Takeda et al. (2009) J. Am. Chem. Soc. 131: 18556-18562.
  10. Takeda et al (2010) J. Biomol. NMR 46: 45-9.

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