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[Preprint]. 2024 May 11:2024.05.10.593625. [Version 1] doi: 10.1101/2024.05.10.593625

Figure 3. Arginines 482, 488 and 496 of Arp5 are important for Arp5 binding to the acid pocket and for INO80 mobilizing nucleosomes.

Figure 3.

(A) The graph shows the extent of nucleosomes mobilized by INO80 with wild type (WT) and mutant Arp5 (R3) that has arginine 482, 488 and 496 mutated to alanine using EMSA (see Figure S3B). 50nM of nucleosomes were incubated with 75nM of INO80 complex with addition of 80 μM ATP (B) The ATPase activity of wild type and mutant INO80 was measured using thin layer chromatography under similar condition to that in (A). (C) The interactions of Arp5 with the histone octamer face of nucleosomes are probed for WT and R3 mutant by site-specific histone crosslinking. The relative efficiency of crosslinking Arp5, Ies2 and Ies6 is shown for three replicates. (D-E) The extent of DNA movement on the (D) exit and (E) entry side is plotted versus time using a photoreactive probe attached to residue 53 of histone H2B for wild type and R3 mutant Arp5 containing INO80 complex. (E) The net loss of DNA crosslinking on the entry side is plotted versus time. Three replicates are performed for each experiment and error bars represent the mean ± SD.