ABCE1 splits 70S with the canonical aRF1 and the ribosome rescue factor aPelota. ABCE1 splits 70S with the canonical aRF1 and the ribosome rescue factor aPelota. (A) The canonical release factor aRF1 acting on mRNA stop codon and the codon-independent release factor aPelota were purified by C-terminal and N-terminal His6 affinity tag, respectively. Purification steps were the same as for ABCE1 (Fig S1). (B) Both factors eluted in a single peak in SEC (20 mM Tris, pH 8.5, 250 mM NaCl, 5 mM MgCl2, Superdex 200, 24 ml analytical grade, GE Healthcare). (C) The ribosome splitting efficiency by WT ABCE1 does not depend on the supplemented release factor (33% with aRF1 and 34% with aPelota). In the absence of ABCE1 or release factors the splitting rates remained >20%. Representative set of three independent experiments. Elina Nürenberg-Goloub et al. LSA 2018;1:e201800095 © 2018 Nürenberg-Goloub et al.