Figure 4.
HrpB nucleic acid cofactor dependence. (A) Polyribonucleotide specificity. Reaction mixtures (15 µl) containing 50 mM Tris-HCl (pH 8.0), 1 mM DTT, 2 mM MgCl2, 1 mM [γ-32P] ATP, 250 ng/µl of the indicated polyribonucleotide (or No RNA), and HrpB as specified were incubated for 15 min at 37°C. Pi release was plotted as a function of input protein. (B) RNA preferences. Reaction mixtures (15 µl) containing 50 mM Tris-HCl (pH 8.0), 1 mM DTT, 2 mM MgCl2, 1 mM [γ-32P] ATP, nucleic acids (either 80 ng/µl MS2, 100 ng/µl P. aeruginosa rRNA, 5 µM R41, or 5 µM D41), and HrpB (as specified) were incubated for 15 min at 37°C. Pi release was plotted as a function of input protein. Data are the average ± SEMs from three independent experiments.