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. 2022 Jul 25;11:e77352. doi: 10.7554/eLife.77352

Figure 4. SWR1 and Swc2 DNA-binding domain (DBD) utilize a combination of sliding and hopping to scan DNA.

(A) Violin plots of diffusion coefficients for SWR1 and Swc2 DBD in increasing potassium chloride concentrations. Medians are shown as white circles and the mean is indicated with a thick horizontal line. 70 mM KCl represents the standard salt condition. (B) 1-CDF plots of SWR1 and Swc2 were fit to exponential decay functions to determine half-lives of binding in varying concentrations of potassium chloride. The number of molecules as well as half-lives determined is printed therein. Dots represent data points, while solid lines represent fits. Half-lives are calculated using the length of all the trajectories in each condition. (C) Upper limits for diffusion of SWR1 and Swc2 predicted using either a helically uncoupled model for hopping diffusion (uppermost solid red line) or a helically coupled model for sliding diffusion (lower dashed red lines). Two dashed lines are shown for helically coupled upper limits because the distance between the helical axis of DNA and the center of mass of either SWR1 or Swc2 is unknown. Markers represent median values. Dcoef values for each condition are shown as horizontal dashes, the number of molecules represented in each condition is as aforementioned. (D) A schematic representation of a model for how SWR1 likely performs 1D diffusion on DNA.

Figure 4—source data 1. Data underlying panels A–C.

Figure 4.

Figure 4—figure supplement 1. Purification and fluorophore labeling of the Swc2 DNA-binding domain (DBD).

Figure 4—figure supplement 1.

Denaturing sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) gel of the Swc2 DBD after nickel his-tag purification and labeling with Cy3-maleimide. A Cy3 scan of the gel (right) reveals that the Swc2 DBD (single band in Coomassie stain on the left) has been labeled with Cy3.
Figure 4—figure supplement 1—source data 1. Gel images (Coomassie and Cy3 scans).
Figure 4—figure supplement 2. Rotation-coupled versus -uncoupled diffusion models and protein size effects on diffusion coefficient.

Figure 4—figure supplement 2.

Apparent diffusion coefficient as a function of two relationships to the radius of SWR1 or Swc2 DNA-binding domain (DBD) where the protein size is printed below the graph and the corresponding model is stated above. A line passing through the origin is fit to both relationships, and the Pearson correlation coefficient is printed within the graph showing a slightly better fit of the measured diffusion coefficients to the rotation-coupled diffusion model.