Tension-regulated VWF self-association. (A) Average time course of VWF (10 μg mL⁻¹) binding to single tethered VWF concatemers at 960 dyn cm⁻². Curves show association (solid lines) and dissociation (dashed lines) at 80 dyn cm⁻² and are color coded by tension bin as shown in the key (the triangle shows the tether point). Data are averaged from 62 tethered VWF molecules used in Figure 1C-E that are large enough to include a 40-to-60 pN tension bin at 960 dyn cm⁻². (B) Initial VWF binding rate and (C) final bound density of VWF from data in panel A. In panels B and C, the maximum binding rate and total binding density are estimated as the average of the values in the 20-to-30 and 30-to-40 pN bins. The force at half-maximal binding, f₅₀, was determined as the intercept of the half maxima with the lines drawn through the data points on either side of the half maxima. (D) Initial binding rate of VWF (10 μg mL⁻¹) at 480 dyn cm⁻² using smaller (5 pN) tension bins. In panels B-D, red lines connect neighboring points. (E-F) Fits of the data in panels B and D to a 2-state model (red lines) in which the observed initial binding rate $kobs=kobs,max/(1+exp ((ΔG-f·Δx)/kBT)),$ where $kobs,max$ is the maximum observed initial binding rate, ΔG is the free energy difference between the 2 states (fast binding and slow or no binding), and Δx is the displacement along the force f direction between the 2 states. Data in panels A-F are from the same set of tethered VWF concatemers. In panels B-D, error bars represent standard deviation. In panels E-F, error bars represent standard deviation for tension and 95% confidence intervals for the other variables.