Figure 1. CHD5 is a nucleosome stimulated ATPase.
(A) Schematic drawing of CHD3, CHD4 and CHD5 proteins showing conserved domains and the CHD5 specific sequence. (B) Table showing sequence identity between different domains of CHD3, CHD4 and CHD5. (C) Expression of recombinant CHD5. Recombinant CHD5-Flag was purified from extracts of baculovirus-infected Sf9 cells. Extracts were incubated with anti-Flag affinity resin, washed and eluted with an excess of Flag peptide. Purified protein was seperated by SDS-PAGE followed by Western blotting with CHD5 antibody (lane 1) or silver staining (lane 2). Asterisk indicates degradation products with intact anti-CHD5 epitope. (D) CHD5 is a nucleosome-stimulated ATPase. Purified recombinant CHD5-Flag was subjected to ATPase assay as described in Materials and Methods. Different amounts of recombinant human CHD5-Flag, naked DNA or in vitro assembled nucleosomes were used as indicated at the bottom of the panel. Reaction products were separated by thin layer chromatography and the amount of released 32Pi was quantitated. The percentage of ATP hydrolysis was calculated for each reaction. The percentage of ATP hydrolysis of control reactions (without enzyme) was set to one. Error bars depict the standard deviation of 3 independent experiments.