Fig. 4: Absence of Nse1-3-4 or Nse5-6 alters Smc5/6’s junction binding behavior.
From: Molecular determinants of Smc5/6 association with DNA junctions
a SDS-PAGE gel picture of purified Smc5/6 5-mer (lacking Nse1-3-4) and 6-mer (lacking Nse5-6) complexes. Coomassie blue stained bands correspond to the subunits as indicated. b Representative kymograph of A555-labeled Smc5/6 5-mer (green) binding behavior on the gapped substrate in the presence of LD655-labeled RPA (red) and ATP. Black arrowheads indicate the positions of the 3’ and 5’ ss-dsDNA junctions. c Dot plot showing the distributions of junction dwell times for Smc5/6 8-mer (black) or 5-mer (gold) at 3’ and 5’ junctions in the presence of RPA and ATP. Plots and statistical analyses are presented as in Fig. 1d. Sample sizes: 3’ 8-mer (N = 301), 3’ 5-mer (N = 156), 5’ 8-mer (N = 513), 5’ 5-mer (N = 141). d Representative kymograph of A555-labeled Smc5/6 6-mer (green) binding behavior on the gapped substrate in the presence of LD655-labeled RPA (red) and ATP. Black arrowheads indicate the positions of the 3’ and 5’ ss-dsDNA junctions. e Dot plot showing the numbers of Smc5/6 binding events at the 3’ or 5’ junctions per 100 s in the presence of ATP for the 8-mer Smc5/6 (black) versus the 6-mer Smc5/6 (magenta). Plots and statistical analyses are presented as in Fig. 1d. Sample sizes are: 3’ 8-mer (N = 37), 3’ 6-mer (N = 44), 5’ 8-mer (N = 37), 5’ 6-mer (N = 46). f Dot plot showing the distributions of junction dwell times for Smc5/6 8-mer (black) and 6-mer (magenta) in the presence of RPA and ATP. Plots and statistical analyses are presented as in Fig. 1d. Sample sizes are: 3’ 8-mer (N = 301), 3’ 6-mer (N = 1,443), 5’ 8-mer (N = 513), 5’ 6-mer (N = 1,468).
