Fig. 3: Tension dependencies of real-time DNA unwinding kinetics in the presence of mtSSB.

a Schematic of optical trapping assays with mtSSB. The experimental setup is identical to that described in Fig. 2a, with the addition of mtSSB (5 nM, green shape) in the reaction buffer. Experiments were performed at 4 mM ATP. b Representative unwinding traces with mtSSB. Traces show the activity of WT Twinkle (blue, F = 2 pN), ΔZBD (orange, F = 2 pN) and ΔC-tail (magenta, F = 6 pN) variants in the presence of mtSSB. Under these conditions, only the ΔC-tail exhibited significant rewinding events upon unwinding. Raw data (grey background) was smoothed using a low-pass 10 Hz filter. c–f Tension dependencies of the average unwinding processivity (c), unwinding rate (d), pause-free velocity (e) and pause occupancy (f) for WT Twinkle in the absence (blue empty symbols, N = 45) and presence (solid blue symbols, N = 57) of mtSSB. The shaded box highlights the tension range where mtSSB’s stimulatory effects are most apparent. g–j panels show same metrics as above (c–f) for the ΔZBD variant in the absence (empty orange symbols, N = 32) and presence (full orange symbols, N = 29) of mtSSB. k–n panels show same metrics as above (c–f) for the ΔC-tail variant in the absence (empty magenta symbols, N = 27) and presence (solid magenta symbols, N = 18) of mtSSB. For all panels, data points represent the average of multiple independent measurements, and error bars indicate the s.e. For this figure (c–n) source data are provided as a Source Data file.