Characterization of the mammalian DEAD-box protein DDX5 reveals functional conservation with S. cerevisiae ortholog Dbp2 in transcriptional control and glucose metabolism

DDX5 lacks annealing activity in vitro, irrespective of the nucleotide-binding state, and this is partially due to the CTE. (A) DDX5 lacks annealing activity in the presence of ATP. Duplex annealing assays were conducted as above, but with 0.1 nM radiolabeled 16-nt top strand RNA and 0.1 nM cold complementary RNA in the presence of 30 mM NaCl and 2 mM ATP/MgCl₂. (B,C) DDX5 also lacks annealing in the absence of nucleotide or in the presence of ADP, in contrast to Dbp2. Duplex annealing assays using purified recombinant MBP-DDX5-GST, DDX5ΔCTE, Dbp2, or no enzyme, in the absence of ATP (Apo state) (B) or in the presence of 2 mM ADP/MgCl₂ (C). (D) Table of the observed annealing rates of DDX5, DDX5ΔCTE, Dbp2, and the no enzyme control in the presence or absence of different nucleotides. The observed annealing rates were determined using the following equation: Y = 1/(1 + k_obs^Ann × X). n.d., not determined. Data are shown as the mean ± SD of 2–3 independent replicates.