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. Author manuscript; available in PMC: 2020 May 25.
Published in final edited form as: Nat Struct Mol Biol. 2019 Nov 25;26(12):1132–1140. doi: 10.1038/s41594-019-0331-x

Fig. 2. Structure of the ribosome stalled during poly(A) translation.

Fig. 2

(A) Schematic of the construct and ribosome-nascent chain complex (RNC) used for structure determination. (B) Characterization of the RNC purification strategy for structure determination. Rabbit reticulocyte lysate translation reactions containing or lacking the mRNA depicted in panel A were subjected to affinity purification via the N-terminal Twin-Strep-tag and eluted sequentially with biotin and SDS. Aliquots of the total translation reaction (input) and 32-fold excess of each eluate were analyzed by SDS-PAGE and staining with Coomassie blue. The sample subjected to cryo-EM analysis is indicated in red. (C) Overview of the poly(A)-stalled RNC structure with key elements indicated in the inset.