Nanopore force spectroscopy tools for analyzing single biomolecular complexes

The time-dependent response of individual biomolecular complexes to an applied force can reveal their mechanical properties, interactions with other biomolecules, and self-interactions. In the past decade, a number of single-molecule methods have been developed and applied to a broad range of biological systems, such as nucleic acid complexes, enzymes and proteins in the skeletal and cardiac muscle sarcomere. Nanopore force spectroscopy (NFS) is an emerging single-molecule method, which takes advantage of the native electrical charge of biomolecule to exert a localized bond-rupture force and measure the biomolecule response. Here, we review the basic principles of the method and discuss two bond breakage modes utilizing either a fixed voltage or a steady voltage ramp. We describe a unified theoretical formalism to extract kinetic information from the NFS data, and illustrate the utility of this formalism by analyzing data from nanopore unzipping of individual DNA hairpin molecules, where the two bond breakage modes were applied.