Fig. 5.

The ratio of forward to backward steps determines myosin-5B run length and velocity under resistive load. a Force dependence of the average step size. Box plot in Supplementary Fig. 1c. b Step size distributions at ±1.2 pN (left panel) and ±2.4 pN (right panel). For resistive forces close to the stall force (+2.4 pN), the distributions of forward (>0) and backward (<0) steps become nearly symmetric. c Force dependence of the average forward (black squares) and backward (cyan squares) step size. d Force dependence of the ratio of forward to backward steps. The dotted cyan line is the fit of the exponential model equation $R=R_0^{+}\exp \left(-\frac{F\cdot d_{\mathrm{fwd∕bwd}}^{+}}{k_{\mathrm{B}}\cdot T}\right)$ to data for positive forces below the stall force, which gives R₀⁺ = 54 ± 8 and d_fwd/bwd⁺ = 8.2 ± 0.6 nm. e Force dependence of forward (black squares) and backward (cyan squares) stepping rates. Exponential fits $k_{f,b}=k_{0f,b}^{\pm }\exp \left(\frac{F\cdot d_{f,b}^{\pm }}{k_{\mathrm{B}}\cdot T}\right)$ of forward (_f) and backward (_b) stepping rates for positive (⁺) and negative (⁻) forces gave: $k_{0f}^{+}$ = 6.5 ± 0.6 s⁻¹, $d_f^{+}$ = 2.2 ± 0.3 nm; $k_{0b}^{+}$ = 10 ± 2 s⁻¹, $d_b^{+}$ = 2.0 ± 0.4 nm. $k_{0f}^{-}$ = 14.6 ± 0.8 s⁻¹, ${\mathrm{d}}_f^{-}$ = 0.05 ± 0.15 nm; $k_{0b}^{-}$ = 54.1 ± 17.5 s⁻¹; $d_b^{-}$ = 2.4 ± 0.9 nm. Error bars, s.e.m., n_fwd = 2729, n_bwd = 689. [ATP] = 100 μM. Red squares and dotted lines in a–c and e are, respectively, the unloaded step size and stepping rate measured with the single molecule motility assay at [ATP] = 100 μM (n = 42)