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. 2009 Dec 24;29(4):770–781. doi: 10.1038/emboj.2009.384

Figure 2.

Figure 2

EF-G(GDPNP) binding at the A-site stabilizes the P/E hybrid state and lowers the activation barriers for L1 stalk motions. Here the dynamics of the EF-G-bound ribosome complex with P-site initiator tRNAfMet are shown. Two unique labelling schemes were used for smFRET imaging: (A) labelled L1 and P-site tRNA in the presence of 10 μM EF-G and 2 mM GDPNP, and (B) labelled P-site tRNA in the presence of 0.1 μM labelled EF-G and 2 mM GDPNP. (Left panels) Cartoon models of the EF-G-bound complex indicating the dynamic elements and the sites of labelling. (Centre panels) Single-molecule fluorescence (Cy3, green; Cy5, red) and FRET (blue) trajectories. The idealization is overlaid in red on the FRET trace. (Right panels) Histograms indicating the occupancies in the observed FRET states as determined by idealization. Highlighted in red/pink are those states that relate to formation of the unlocked ribosome. ‘N' denotes the number of individual smFRET trajectories included in the analysis.