Extracting kinetics from single-molecule force spectroscopy: nanopore unzipping of DNA hairpins

Biophys J. 2007 Jun 15;92(12):4188-95. doi: 10.1529/biophysj.106.102855. Epub 2007 Mar 23.


Single-molecule force experiments provide powerful new tools to explore biomolecular interactions. Here, we describe a systematic procedure for extracting kinetic information from force-spectroscopy experiments, and apply it to nanopore unzipping of individual DNA hairpins. Two types of measurements are considered: unzipping at constant voltage, and unzipping at constant voltage-ramp speeds. We perform a global maximum-likelihood analysis of the experimental data at low-to-intermediate ramp speeds. To validate the theoretical models, we compare their predictions with two independent sets of data, collected at high ramp speeds and at constant voltage, by using a quantitative relation between the two types of measurements. Microscopic approaches based on Kramers theory of diffusive barrier crossing allow us to estimate not only intrinsic rates and transition state locations, as in the widely used phenomenological approach based on Bell's formula, but also free energies of activation. The problem of extracting unique and accurate kinetic parameters of a molecular transition is discussed in light of the apparent success of the microscopic theories in reproducing the experimental data.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural

MeSH terms

  • Computer Simulation
  • DNA / chemistry*
  • DNA / ultrastructure*
  • Elasticity
  • Mechanics
  • Micromanipulation / methods
  • Microscopy, Atomic Force / methods*
  • Models, Chemical*
  • Models, Molecular*
  • Nanostructures / chemistry*
  • Nanostructures / ultrastructure*
  • Nucleic Acid Conformation
  • Porosity
  • Stress, Mechanical


  • DNA