Abstract
Single-molecule magnetic tweezers have recently emerged as a powerful technique for measuring the equilibrium dynamics of individual proteins under force. In magnetic tweezers, a single protein is tethered between a glass coverslip and a superparamagnetic bead, and by applying and controlling a magnetic field, the protein is mechanically stretched while force-induced conformational changes are measured by tracking the vertical position of the bead. The soft trap created by the magnetic field provides intrinsic force-clamp conditions, which makes magnetic tweezers particularly well-suited to measure protein conformational dynamics. Traditionally employed to study DNA due to their initially low spatial and temporal resolutions, magnetic tweezers instrumentation has experienced significant progress in recent years. The development of high-speed cameras, stronger illumination sources, advanced image analysis algorithms, and dedicated chemical functionalization strategies, now allow for high-resolution and ultra-stable experiments. Together with their ability to apply and control low forces, magnetic tweezers can capture long-term equilibrium protein folding dynamics, not possible with any other technique. These capabilities have proven particularly valuable in the study of force-sensing protein systems, which often exhibit low mechanical stabilities that are challenging to measure with other techniques. In this review, we will discuss the current status of magnetic tweezers instrumentation for studying protein folding dynamics, focusing on both the instrumental aspects and methodologies to interpret nanomechanical experiments.