Skip to main content
. 2017 May 24;12(5):e0177972. doi: 10.1371/journal.pone.0177972

Fig 5. mRNA release from the PoTC and splitting of ribosomal subunits by RRF and EF-G.

Fig 5

(A) PoTC (100 nM) before the reaction and (B) the reaction product in frozen-hydrated states was observed with a transmission electron microscope (JEM1230, JEOL) operated at 120 kV at liquid nitrogen temperature. (B) Reaction products consist mainly reassociated 70S. It is possible to see dissociated 30S on the cryo-EM grid (see particles in white circles). Compared with the PoTC (A), 70S ribosome particles evenly spread over the thin carbon layer due to the release of ribosomes from mRNA by the ribosome recycling reaction. Specimens were observed at 40,000-fold magnification in a -3.5 μm defocused condition. Images were obtained with 1 K X 1 K fast scan CCD (charge coupled device) camera (TVIPS: Tietz Video and Image Processing Systems). (C) Single-particle analysis of cryo-EM images of the PoTC after the RRF reaction. Figure shows vacant 70S ribosomes formed after the recycling reaction (EMD-9532). The 70S ribosome does not have any densities ascribed to released components (tRNA and mRNA), which are clearly observed in the PoTC (Fig 3A and 3B). Color code: 50S, aqua; 30S, light grey. Landmarks of ribosome: CP, central protuberance; L1, L1 stalk; St, L7/12 stalk; Sb, L7/12 stalk base; hd, head; bk, beak; pt, platform; sh, shoulder; sp, spur.