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. Author manuscript; available in PMC: 2015 Jun 12.
Published in final edited form as: Cell Rep. 2014 May 15;7(5):1521–1533. doi: 10.1016/j.celrep.2014.04.033

Figure 6. SecM increases the barrier to translocation but does not hinder EFG binding.

Figure 6

(A) Using Cy5 labeled EF-G and Cy3B/BHQ2 intersubunit FRET, EF-G binding attempts to the rotated state leading to a successful translocation can be tracked as a EF-G binding signal concurrent with a high to low Cy3B intensity transition (Chen et al., 2013b). (B) At 2.5 μM charged total tRNA and 160 nM EF-G-Cy5, the average number of EF-G binding attempts increases from 1.5~2.5 to 3.5~4 after codon 15 (n = 91), indicating an increased energy barrier to translocation. Error bars represent standard error. (C) EF-G binding frequency remains constant at 0.4~0.5 s−1, suggesting that EF-G binding to the rotated state is not inhibited. (D) EF-G dwell times on the ribosome remain constant at 0.10~0.15 s. Frequencies and lifetimes are fitted to single-exponential distributions and error bars represent s.e.m.