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Studying 3D Subdomains of Proteins at the Nanometer Scale Using Fluorescence Spectroscopy

  • Pierre M. Viallet
  • Tuan Vo-Dinh
Part of the Methods in Molecular Biology™ book series (MIMB, volume 300)

Summary

Databases devoted to the crystal structure of proteins have dramatically increased in size during the last two decades. Moreover, X-ray and NMR technology studies have shown that proteins belonging to the same family generally share the same global 3D architecture. These results suggest that the need for experimental determination of protein structure will be reduced to those that are suspected to have sufficiently novel structures. Furthermore, NMR and other techniques have demonstrated that a protein in solution experiences constant random thermal motions that occur over large time scales, ranging from picoseconds to seconds and perhaps hours. Such changes may have important functional consequences, but identifying which changes are functionally relevant remains a difficult task even if this problem has been addressed both with experimental and computational methods. For that specific purpose, there is a need for methods allowing a fast and accurate monitoring of conformation changes (that occur at specific sub-domains of proteins. Fluorescence resonance energy transfer (FRET) is a suitable tool for monitoring conformational changes at the nanoscale level. This chapter describes the various FRET methods that are used for monitoring the 3D sub-domain conformation of proteins in solution, in single living cells and at the single molecular level.

Key Words

Fluorescence spectroscopy fluorescence resonance energy transfer fluorescence lifetime imaging measurements lifetime measurements energy transfer near-field scanning optical microscope protein subdomain 

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Copyright information

© Humana Press Inc., Totowa, NJ 2005

Authors and Affiliations

  • Pierre M. Viallet
    • 1
    • 2
  • Tuan Vo-Dinh
    • 3
  1. 1.Laboratory of Physicochemical Biology of Integrated SystemsUniversity of PerpignanPerpignanFrance
  2. 2.Advanced Biomedical Science and Technology GroupOak Ridge National LaboratoryOak Ridge
  3. 3.Center for Advanced Biomedical Photonics, Life Sciences DivisionOak Ridge National LaboratoryOak Ridge

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