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Probing transcription factor dynamics at the single-molecule level in a living cell.

Science (New York, N.Y.) | May 25, 2007

http://www.ncbi.nlm.nih.gov/pubmed/17525339

Transcription factors regulate gene expression through their binding to DNA. In a living Escherichia coli cell, we directly observed specific binding of a lac repressor, labeled with a fluorescent protein, to a chromosomal lac operator. Using single-molecule detection techniques, we measured the kinetics of binding and dissociation of the repressor in response to metabolic signals. Furthermore, we characterized the nonspecific binding to DNA, one-dimensional (1D) diffusion along DNA segments, and 3D translocation among segments through cytoplasm at the single-molecule level. In searching for the operator, a lac repressor spends approximately 90% of time nonspecifically bound to and diffusing along DNA with a residence time of <5 milliseconds. The methods and findings can be generalized to other nucleic acid binding proteins.

Pubmed ID: 17525339 RIS Download

Mesh terms: Bacterial Proteins | DNA, Bacterial | Diffusion | Escherichia coli | Escherichia coli Proteins | Kinetics | Lac Operon | Lac Repressors | Luminescent Proteins | Microscopy, Fluorescence | Operator Regions, Genetic | Protein Binding | Repressor Proteins | Signal Transduction

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Associated grants

  • Agency: NIH HHS, Id: DP1 OD000277
  • Agency: NIH HHS, Id: DP1 OD000277-02

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