Figure S5. ATPase activity of ABCE1 is stimulated by 70S and aRF1 during ribosome splitting.

(A) Representative set of two independent TLC radiograms of ³²P-γ-ATP (37.5 μM) showing the stimulation of WT ABCE1 (1 μM) by 70S in concert with aRF1. (B) ATPase activity measured by phosphate release during ribosome splitting. The marked values at 8 and 16 min were excluded from the analysis as they obviously are outside the linear range for this reaction. (C) Total ATP hydrolysis rate in each sample (bars) and the turnover number for ABCE1 after subtraction of control experiments. Interestingly, Although 70S alone does not influence ATPase activity of ABCE1, the presence of aRF1 leads to a reproducibly slight inhibiting effect. Notably, the overall drop in ATPase activity results from the lowered temperature (45°C) and high Mg²⁺ concentration (25 mM) at 70S splitting conditions.