Figure S2.
3D subtomogram average of a cytoplasmic ribosome positioned for protein import on the OMM shows multiple contact points. (A) FSC plot of the OMM-associated 80S cytoplasmic ribosome reconstruction is shown with resolution reported at 0.143 FSC and the reconstructed subtomogram average (bottom left). The 80S cytoplasmic ribosome was resolved to 19 Å from 1,076 ribosome particles with the color map shows the local resolution (bottom right). (B) The peptide exit tunnel is visible in the subtomogram average (gray density) at higher isosurface volume thresholds, as indicated by the dashed black circle. This was used to mark its position relative to the connecting densities visible in the subtomogram average at lower isosurface volume threshold values (colored density). (C) Representative models of ribosomes positioned within 250 Å of the OMM surface with their exit tunnels facing away from the OMM. 3D refinement of these particles results in a 3D reconstruction of a ribosome that does not contain any distinguishable connecting densities between the 80S ribosome and the OMM, suggesting that these connections are specific to 80S ribosomes optimally positioned for protein import. (D) Density corresponding to the connection labeled #1 in the subtomogram average of mitochondrial-associated ribosomes correlates well with the density corresponding to the expansion segment of eS7La of the 25S rRNA in the large 60S subunit present in the ER-associated ribosome maps from S. cerevisiae (EMD-3764). (E) The density corresponding to the connection labeled #2 correlates well with the density corresponding to the region where the ribosome interacts with the import channel, Ssh1, in the ER-associated ribosome maps from S. cerevisiae (EMD-1667). (F) The density corresponding to the connection labeled #3 correlates well with the density corresponding to the rRNA expansion segment, rpL38, and the TRAP complex in the ER-associated ribosome maps from human (EMD-15884), rabbit (EMD-16232), and canine (EMD-3068). (G) Overlap between the OMM-associated 80S cytoplasmic ribosome structure from this work (transparent blue) with ER-bound ribosome (EMD-14424; green) and mitochondria-bound ribosome (EMD-14423, orange) from S. pombe and mitochondria-bound ribosome from purified S. cerevisiae mitochondria (EMD-3762, pink). Dashed colored lines show the relative orientations of the membranes for the different structures.

3D subtomogram average of a cytoplasmic ribosome positioned for protein import on the OMM shows multiple contact points. (A) FSC plot of the OMM-associated 80S cytoplasmic ribosome reconstruction is shown with resolution reported at 0.143 FSC and the reconstructed subtomogram average (bottom left). The 80S cytoplasmic ribosome was resolved to 19 Å from 1,076 ribosome particles with the color map shows the local resolution (bottom right). (B) The peptide exit tunnel is visible in the subtomogram average (gray density) at higher isosurface volume thresholds, as indicated by the dashed black circle. This was used to mark its position relative to the connecting densities visible in the subtomogram average at lower isosurface volume threshold values (colored density). (C) Representative models of ribosomes positioned within 250 Å of the OMM surface with their exit tunnels facing away from the OMM. 3D refinement of these particles results in a 3D reconstruction of a ribosome that does not contain any distinguishable connecting densities between the 80S ribosome and the OMM, suggesting that these connections are specific to 80S ribosomes optimally positioned for protein import. (D) Density corresponding to the connection labeled #1 in the subtomogram average of mitochondrial-associated ribosomes correlates well with the density corresponding to the expansion segment of eS7La of the 25S rRNA in the large 60S subunit present in the ER-associated ribosome maps from S. cerevisiae (EMD-3764). (E) The density corresponding to the connection labeled #2 correlates well with the density corresponding to the region where the ribosome interacts with the import channel, Ssh1, in the ER-associated ribosome maps from S. cerevisiae (EMD-1667). (F) The density corresponding to the connection labeled #3 correlates well with the density corresponding to the rRNA expansion segment, rpL38, and the TRAP complex in the ER-associated ribosome maps from human (EMD-15884), rabbit (EMD-16232), and canine (EMD-3068). (G) Overlap between the OMM-associated 80S cytoplasmic ribosome structure from this work (transparent blue) with ER-bound ribosome (EMD-14424; green) and mitochondria-bound ribosome (EMD-14423, orange) from S. pombe and mitochondria-bound ribosome from purified S. cerevisiae mitochondria (EMD-3762, pink). Dashed colored lines show the relative orientations of the membranes for the different structures.

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