Fig. 2: Single molecule kinetic measurements of utrn CH1–CH2 mutants in vitro.
From: Biased localization of actin binding proteins by actin filament conformation
a Single molecule binding assay to measure the kinetic properties. Images in the example shown are for utrnWT. Maximum intensity projection through time displays the filament backbones and a kymograph the kinetics of binding. Scale bars are 5 µm. The kymograph is a 12 µm section of filament backbone. b Average binding dwell times for the different CH1–CH2 mutants. c Binding on-rates for the different utrophin mutants evaluated by fitting the binding event frequency over a range of different concentrations. Error bars are the mean ± SEM for each concentration measured from more than 12 fields of view collected from two imaging chambers. d Cumulative distribution function for utrnWT (green) and utrnLAM (magenta). e Cumulative distribution function for utrnWT (green) and utrnΔN (orange). f Cumulative distribution function for K121A (red) and G125A L132A (blue). g Comparisons of the first timescale τ1, h second timescale τ2, and i relative amplitude of events belonging to each timescale from a double exponential fit to the cumulative distribution functions for the different constructs. Error bars are the standard deviation from the mean of 3 technical replicates. Conditions were compared using a two-tailed students t-test, with p < 0.05 denoted by a star (*).
