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Related Experiment Video

Updated: Jun 1, 2026

Combining Single-molecule Manipulation and Imaging for the Study of Protein-DNA Interactions
14:43

Combining Single-molecule Manipulation and Imaging for the Study of Protein-DNA Interactions

Published on: August 27, 2014

Measuring DNA-protein binding affinity on a single molecule using optical tweezers.

Micah J McCauley1, Mark C Williams

  • 1Department of Physics, Northeastern University, Boston, MA, USA.

Methods in Molecular Biology (Clifton, N.J.)
|June 16, 2011
PubMed
Summary

Quantifying DNA-protein interactions is challenging. This study uses a single DNA molecule to measure binding affinity, successfully characterizing high mobility group protein HMGB1 binding to double-stranded DNA.

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Area of Science:

  • Molecular Biology
  • Biophysics
  • Biochemistry

Background:

  • Observing DNA-protein interactions at the single-molecule level is possible with various techniques.
  • Quantifying the binding affinity of these interactions is challenging and typically requires analyzing numerous individual events.

Purpose of the Study:

  • To develop and demonstrate a method for quantifying DNA-protein binding affinity using single DNA molecules.
  • To characterize the interaction between high mobility group protein B1 (HMGB1) and double-stranded DNA (dsDNA).

Main Methods:

  • Utilizing a single λ DNA molecule as a lattice of binding sites.
  • Employing reversible DNA melting by stretching and relaxing the tethered DNA molecule to convert between double-stranded (ds) and single-stranded (ss) forms.

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Last Updated: Jun 1, 2026

Combining Single-molecule Manipulation and Imaging for the Study of Protein-DNA Interactions
14:43

Combining Single-molecule Manipulation and Imaging for the Study of Protein-DNA Interactions

Published on: August 27, 2014

Use of Dual Optical Tweezers and Microfluidics for Single-Molecule Studies
06:53

Use of Dual Optical Tweezers and Microfluidics for Single-Molecule Studies

Published on: November 18, 2022

Optical Tweezers to Study RNA-Protein Interactions in Translation Regulation
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Optical Tweezers to Study RNA-Protein Interactions in Translation Regulation

Published on: February 12, 2022

  • Monitoring DNA property changes with varying protein concentrations and fitting to binding models.

    • Main Results:

    • The method allows for the characterization and quantification of DNA-protein interactions.
    • High mobility group protein HMGB1 (box A + B) was observed to stabilize dsDNA.
    • The equilibrium association constant for HMGB1(box A + B) binding to dsDNA was determined.

    Conclusions:

    • This single-molecule DNA assay provides a quantitative method for determining DNA-protein binding affinities.
    • The technique is effective for characterizing interactions such as HMGB1 binding to dsDNA.