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. 2010 Dec 30;286(10):7830–7840. doi: 10.1074/jbc.M110.194407

FIGURE 3.

FIGURE 3.

RecA K72R fails to form extended filaments on cssDNA using wild type RecA as a nucleation factor. Images of RecA filaments bound to cssDNA were obtained at 15,000 × magnification by EM as described under “Experimental Procedures.” Wild type RecA formed filaments on SSB-coated cssDNA, and reaction samples were imaged before and 5, 20, 40, and 60 min after RecA K72R addition. The ratio of wild type RecA to RecA K72R was 1:15. Reactions contained 4 μm M13mp18 cssDNA, 3 mm ATP, 0.4 μm SSB, 0.133 μm wild type RecA, and 2 μm RecA K72R (or an additional 2 μm wild type RecA in the 60-min time point only). The small wild type RecA filaments in the 0 time point disassemble within 5 min of RecA K72R addition and do not reassemble in the presence of RecA K72R. For comparison, panel F shows a 60-min time point for a reaction in which the wild type RecA protein, instead of the mutant protein, was added to the wild type RecA nuclei.