EF-G-ribosome pre-translocation complex Frealign parameter files

<h3>During protein synthesis, tRNAs and their associated mRNA codons move sequentially on the ribosome from the A (aminoacyl) site to the P (peptidyl) site to the E (exit) site in a process catalyzed by a universally conserved ribosome-dependent GTPase [elongation factor G (EF-G) in prokaryotes and elongation factor 2 (EF-2) in eukaryotes]. Although the high-resolution structure of EF-G bound to the posttranslocation ribosome has been determined, the pretranslocation conformation of the ribosome bound with EF-G and A-site tRNA has evaded visualization owing to the transient nature of this state. Here we use electron cryomicroscopy to determine the structure of the 70S ribosome with EF-G, which is trapped in the pretranslocation state using antibiotic viomycin. Comparison with the posttranslocation ribosome shows that the small subunit of the pretranslocation ribosome is rotated by ∼12° relative to the large subunit. Domain IV of EF-G is positioned in the cleft between the body and head of the small subunit outwardly of the A site and contacts the A-site tRNA. Our findings suggest a model in which domain IV of EF-G promotes the translocation of tRNA from the A to the P site as the small ribosome subunit spontaneously rotates back from the hybrid, rotated state into the nonrotated posttranslocation state.<br></h3><h3>Data description</h3><p>The archive linked to the right contains <a href="https://www.janelia.org/open-science/frealign">Frealign</a> parameter files and scripts that were used to calculate 15 classes from 1.3 million single particle images of an E. coli EF-F-ribosome preparation. The data was collected at 300 kV on an FEI Titan Krios microscope and Falcon I direct electron detector at a calibrated magnification of 133,333, giving a pixel size on the specimen of 1.04 Å. The particle stack has 512 GB (in <a href="http://www.ccp4.ac.uk/html/maplib.html#description">MRC/CCP4 format</a>) and is available upon request as it is too large to be hosted on this web page.</p