Figure 5.

The ATPase DDX55 preferentially binds and is stimulated by double-stranded RNAs. (A) ZZ-DDX55-His₇, ZZ-DDX55_1-403-His₇ and ZZ-DDX55_T60A-His₇ were recombinantly expressed in E. coli and purified via their His tags on a nickel matrix. Purified proteins were separated by SDS-PAGE and visualized by Coomassie staining. (B,C) Fluorescence anisotropy experiments were performed using fluorescently labelled, single-stranded (B) or double-stranded (C) RNAs and different amounts of purified ZZ-DDX55-His₇ and ZZ-DDX55_1-403-His₇. Data from three independent experiments are presented as mean ± standard deviation. (D) In vitro NADH-coupled ATPase assays were performed using no protein (no prot.), DDX55, DDX55_T60A or DDX55_1-403. Samples contained no RNA (-), single- or double-stranded RNA (ss and ds respectively) or a mimic of the cellular RNA sequence crosslinked by DDX55 (H62)