Figure 5
From: Prokaryotic Argonaute from Archaeoglobus fulgidus interacts with DNA as a homodimer
Dynamics of WT AfAgo-induced DNA loops. (a) A schematic overview of the single-molecule assay using TIRF microscopy. (b,c) Trajectories of donor (green) and acceptor (red) intensity and corresponding proximity ratio, E, of individual DNA fragments without (b) and with 20 nM WT AfAgo (c). (d) Left—an image of 287 pooled time traces of the proximity ratio, E, from the measurement with 20 nM of WT AfAgo. The image is normalized to the maximum image intensity. Right—a section of the image in the left integrated over the first 10 s shown with the two-Gaussian fit. The positions of the Gaussian maxima are 0.09 ± 0.01 and 0.36 ± 0.01. For comparison, a trace- and time-averaged section from the measurement of 227 traces on bare DNA is shown. (e) An example of trajectories of donor (green) and acceptor (red) intensity and corresponding proximity ratio, E, with HMM idealization of an individual DNA fragment with 20 nM WT AfAgo. (f) Cumulative histogram of the looped state durations from 287 E traces with a single-exponential fit with the exponential factor of 33 ± 1 s.
