RNA Therapeutics Institute; Department of Biochemistry and Molecular Pharmacology; Graduate School of Biomedical Sciences
Investigative Techniques | Molecular Biology | Nucleic Acids, Nucleotides, and Nucleosides
During translation, a conserved GTPase elongation factor—EF-G in bacteria or eEF2 in eukaryotes—translocates tRNA and mRNA through the ribosome. EF-G has been proposed to act as a flexible motor that propels tRNA and mRNA movement, as a rigid pawl that biases unidirectional translocation resulting from ribosome rearrangements, or by various combinations of motor- and pawl-like mechanisms. Using time-resolved cryo-EM, we visualized GTP-catalyzed translocation without inhibitors, capturing elusive structures of ribosome•EF-G intermediates at near-atomic resolution. Prior to translocation, EF-G binds near peptidyl-tRNA, while the rotated 30S subunit stabilizes the EF-G GTPase center. Reverse 30S rotation releases Pi and translocates peptidyl-tRNA and EF-G by ∼20 Å. An additional 4-Å translocation initiates EF-G dissociation from a transient ribosome state with highly swiveled 30S head. The structures visualize how nearly rigid EF-G rectifies inherent and spontaneous ribosomal dynamics into tRNA-mRNA translocation, whereas GTP hydrolysis and Pi release drive EF-G dissociation.
Molecular Biology, cryo-EM, tRNA, mRNA
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DOI of Published Version
bioRxiv 2021.05.31.446434; doi: https://doi.org/10.1101/2021.05.31.446434. Link to preprint on bioRxiv.
Carbone CE, Loveland AB, Gamper, Jr. H, Hou Y, Demo G, Korostelev AA. (2021). Time-resolved cryo-EM visualizes ribosomal translocation with EF-G and GTP [preprint]. University of Massachusetts Medical School Faculty Publications. https://doi.org/10.1101/2021.05.31.446434. Retrieved from https://escholarship.umassmed.edu/faculty_pubs/2034
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