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Probing Transcription Factor Dynamics at the Single-Molecule Level in a Living Cell
Johan Elf,1*Gene-Wei Li,2*X. Sunney Xie1
Transcription factors regulate gene expression through theirbinding to DNA. In a living Escherichia coli cell, we directlyobserved specific binding of a lac repressor, labeled with afluorescent protein, to a chromosomal lac operator. Using single-moleculedetection techniques, we measured the kinetics of binding anddissociation 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 translocationamong segments through cytoplasm at the single-molecule level.In searching for the operator, a lac repressor spends 90% oftime nonspecifically bound to and diffusing along DNA with aresidence time of <5 milliseconds. The methods and findingscan be generalized to other nucleic acid binding proteins.
1 Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138, USA. 2 Department of Physics, Harvard University, Cambridge, MA 02138, USA.
* These authors contributed equally to this work.
To whom correspondence should be addressed. E-mail: xie{at}chemistry.harvard.edu
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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. 
To whom correspondence should be addressed. E-mail: 
