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. Author manuscript; available in PMC: 2010 Nov 27.
Published in final edited form as: J Mol Biol. 2009 Sep 15;394(2):251–267. doi: 10.1016/j.jmb.2009.09.020

Figure 5. Effect of RF-3 and/or RRF overexpression on nascent peptide-induced ribosome pausing.

Figure 5

(a) Peptidyl-tRNA turnover in tmRNA+ cells overexpressing RF-3 and/or RRF. FLAG-(m)YbeL-PP nascent chains were pulse labeled with [35S]-methionine/cysteine, then chased with unlabeled amino acids for the indicated time. The T1/2 of peptidyl prolyl-tRNA2Pro turnover was 15 ± 0.3 s during RF-3 overexpression, 25 ± 3.2 s during RRF overexpression, and 20 ± 1.8 s during RF-3/RRF co-overexpression. (b) RF-3 and RRF overexpression and RF-ribosome interaction. Lysates from tmRNA+ cells co-expressing YbeL-PP (UAA) along with RF-3 and/or RRF were fractionated by density ultracentrifugation, and analyzed by Western blot. Expression of RF-3 and/or RRF (+ L-Ara) had no effect on the RF-1-ribosome interaction, compared to control cells without RF-3/RRF induction (+ D-Glu).