Fig. 3. Event-oriented analysis of AgTx2 binding.

(A) Representative data of height transitions and definitions for the event-oriented analysis. The AgTx2-bound and AgTx2-unbound states are indicated by white and black arrowheads, respectively. Red and blue arrowheads indicate binding and dissociation events, respectively. For an arbitrary dissociation event (blue arrowhead), t_persist was defined from the instance of dissociation (blue). The time origin of Δt was the end of t_persist. The time variable definitions are the same as for the binding event (red arrowhead). (B and C) A representative dataset and the event-oriented analysis. The AgTx2-bound and AgTx2-unbound states are indicated by white and black arrowheads, respectively. All dissociation events were detected from the long-lasting height transition data, and strips of data before and after the event were collected and aligned at the dissociation events (Δt = 0) (blue arrowhead). Five examples are shown. For t_persist = 0 s (B), all the strips were ensemble-averaged, providing P_bound(Δt) for t_persist = 0 s. For t_persist = 1 s (C), strips of data persisting in the unbound state longer than t_persist after the dissociation event were used for ensemble averaging. (D) Time course of the binding probability after dissociation. P_bound(Δt) for different t_persist values ranging from immediately after dissociation (t_persist = 0 s) to 1 s from dissociation (t_persist = 1 s) are shown as a function of Δt measured from the end of t_persist. As t_persist increased, the time course of P_bound(Δt) could be expressed by a single-exponential function. (E) Time course of the binding probability after binding. P_bound(Δt) for different t_persist values from immediately after binding (t_persist = 0 s) to 1 s (t_persist = 1 s) after binding are shown. The binding transitions were measured in 10 mM Hepes (pH 7.5) containing 200 mM KCl and 20 nM AgTx2.