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Single-Molecule FRET Analysis of Protein-DNA Complexes

  • Mike Heilemann
  • Ling Chin Hwang
  • Konstantinos Lymperopoulos
  • Achillefs N. Kapanidis
Protocol
Part of the Methods in Molecular Biology™ book series (MIMB, volume 543)

Summary

We present a single-molecule method for studying protein–DNA interactions based on fluorescence resonance energy transfer (FRET) and alternating-laser excitation (ALEX) of single diffusing molecules. An application of this method to the study of a bacterial transcription initiation complex is presented.

Key words

Single-molecule fluorescence spectroscopy Fluorescence resonance energy transfer (FRET) Alternating-laser excitation (ALEX) Protein–DNA interactions 

Notes

The help of the rest of the members of the Gene Machines’ group at Physics Department, University of Oxford, is greatly acknowledged. We would also like to thank the groups of Richard Ebright and Shimon Weiss for a fruitful collaboration on transcription-related projects. Funding was provided by the UK IRC in Bionanotechnology, the European Union (MIRG-CT-2005-031079), and the National Institutes of Health (NIH R01 GM069709). MH was supported by a fellowship from the German Academic Exchange Service (DAAD).

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Mike Heilemann
    • 1
  • Ling Chin Hwang
    • 1
  • Konstantinos Lymperopoulos
    • 1
  • Achillefs N. Kapanidis
    • 1
  1. 1.Department of Physics and IRC in BionanotechnologyClarendon Laboratory, University of OxfordOxford

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