Fig. 5. Multi-state kinetic model and Monte-Carlo simulation of mechanical rupture of XMod-Doc:Coh.

a The multi-state kinetic model postulates that upon molecular collision, the complex has ~80% probability of forming the strong binding mode A (pathways 1 and 2) and 20% probability of forming the weaker binding mode B (pathway 3). Once the binding mode is set, there is no interconversion between the modes. In the strong binding mode, the complex can rupture at high force (pathway 1) with XMod remaining folded, or XMod can unfold prior to complex rupture according to a loading rate-dependent unfolding rate k_u(L) (pathway 2). In pathway 2, XMod unfolding destabilizes the complex, resulting in low force complex rupture. At increased loading rates, the XMod unfolding rate decreases so that pathway 2 is deactivated and the complex has a higher probability of unbinding along pathway 1 (high force rupture). In pathway 3, the complex ruptures at low force without reaching sufficiently high force to unfold XMod. b, c Monte-Carlo simulation results. Force vs. extension curves at constant pulling speed were simulated to obtain the loading rate dependency of complex rupture and XMod unfolding events b, as well as the percentages of the three unbinding pathways at different pulling speeds c. The simulation was carried out at the same pulling speeds as the experiments (100, 400, 1600, 6400 nm/s) and further extended over a range from 1 to 10⁶ nm/s. Simulated force vs. loading rate plots were fitted with the Bell–Evans model to extract k₀ and Δx^‡ values (Supplementary Table 1). The error bars in b represent the standard deviation of rupture forces (n = 192–693).