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. Author manuscript; available in PMC: 2022 Mar 4.
Published in final edited form as: Mol Cell. 2021 Jan 8;81(5):1043–1057.e8. doi: 10.1016/j.molcel.2020.12.019

Figure 4. ATP hydrolysis by Rad55-Rad57 is required for its release from Rad51-ssDNA filaments.

Figure 4.

(A) Experimental schematic.

(B) Wide-field views of RPA-mCherry (magenta)-bound ssDNA at the indicated time points. Rad51 and Rad55-Rad57 (green) were injected with 2 mM AMP-PNP.

(C) Representative kymographs showing loss of mCherry-RPA and retention of GFP-Rad55-Rad57. Time of injection is indicated by a white dashed line.

(D) Mean normalized RPA-mCherry and GFP-Rad55-Rad57 intensity; the shaded area represents 95% CI. Data were derived using three flow cells per reaction condition. Rad51 assembly kinetics are plotted as [1 – RPA] signal for comparison.

(E) Representative kymographs showing RPA-mCherry loss at the indicated concentrations of Rad55-Rad57 co-injected with 2 μM Rad51 in AMP-PNP. Time of injection is indicated by a white dashed line.

(F) Quantification of the experiment described in (E), showing kinetics of RPA-mCherry loss under the indicated conditions. Data are represented as mean normalized intensity; the shaded area represents 95% CI. Data were derived using three flow cells per reaction condition.

See also Figures S4 and S5.