Optical tweezers have become the method of choice in single-molecule manipulation
studies. In this Primer, we first review the physical principles of optical tweezers and the …

Manipulation of individual molecules with optical tweezers provides a powerful means of
interrogating the structure and folding of proteins. Mechanical force is not only a relevant …

Proteins have dynamic structures that undergo chain motions on time scales spanning from
picoseconds to seconds. Resolving the resultant conformational heterogeneity is essential …

Folding may be described conceptually in terms of trajectories over a landscape of free
energies corresponding to different molecular configurations. In practice, energy landscapes …

Transition paths are the trajectories followed during the fleeting moments when molecular
structure changes during folding reactions. Because they provide a direct look at the …

The use of force probes to induce unfolding and refolding of single molecules through the
application of mechanical tension, known as single-molecule force spectroscopy (SMFS) …

Statistical mechanics can describe the major conformational ensembles determining the
equilibrium free-energy landscape of a folding protein. The challenge is to capture the full …

Protein folding is described conceptually in terms of diffusion over a configurational free-
energy landscape, typically reduced to a one-dimensional profile along a reaction …

Single-molecule force spectroscopy sheds light onto the free energy landscapes governing
protein folding and molecular recognition. Since only a single molecule or single molecular …

Highlights•Optical tweezers provide a sensitive tool to probe structural dynamics.•Native and
non-native folding can be characterized in detail.•Effects of intermolecular interactions on …