Magnetic Tweezers to Observe the Ultrafast Downhill Folding of a Protein in the Millisecond and Nanometer Scale

Recent advances in magnetic tweezers set them as a promising tool to unveil the dynamics of proteins and nucleic acids by means of hours to days-long stable constant force experiments. Archetypal folds play a crucial role in our means to uncover the intricacies of protein folding. One prominent case is the λ-phage cI transcription repressor fragment λ6-85; a five-helix bundle protein were single point mutations shift its folding time from fast 200 μs to ultrafast downhill-like 2.3 μs kinetics. Can we resolve ultrafast kinetics with the milliseconds time-resolution of the magnetic tweezers? Here, we report the observation of ultrafast protein folding in magnetic tweezers. Small pN forces slow down the folding to tens of milliseconds; ∼10-fold bigger compared to optical tweezers measurements. We developed an analytic model to describe folding kinetics in magnetic tweezers and extract folding and unfolding rates at a broad range of dynamics ranging from milliseconds to seconds. We decipher free energy landscapes of three different variants of λ6-85, namely λWt, λHA and λYA, and compare transition state positions and barrier heights as small as few kBT measured with magnetic tweezers and optical tweezers.