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Time-resolved cryo-EM visualizes ribosomal translocation with EF-G and GTP.

<div class='caption-title'>Time-resolved cryo-EM of translocation with EF-G and GTP.</div>
<div class='caption-title'>Structures of translocation intermediates with EF-G.</div>
<div class='caption-title'>Positions and interactions of EF-G in translocation intermediates.</div>
Carbone CE, Loveland AB, Gamper HB, Hou YM, Demo G, Korostelev AA
OpenAccess logo Nat Commun 12 7236 (2021)
Related entries: 7ss9, 7ssd, 7ssl, 7ssn, 7sso, 7ssw, 7st6, 7st7

Cited: 25 times
EuropePMC logo PMID: 34903725

Abstract

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.

Reviews citing this publication (6)

  1. Electron microscopy holdings of the Protein Data Bank: the impact of the resolution revolution, new validation tools, and implications for the future. Burley SK, Berman HM, Chiu W, Dai W, Flatt JW, Hudson BP, Kaelber JT, Khare SD, Kulczyk AW, Lawson CL, Pintilie GD, Sali A, Vallat B, Westbrook JD, Young JY, Zardecki C. Biophys Rev 14 1281-1301 (2022)
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  4. Frozen in time: analyzing molecular dynamics with time-resolved cryo-EM. Amann SJ, Keihsler D, Bodrug T, Brown NG, Haselbach D. Structure 31 4-19 (2023)
  5. Exploring the Structural Variability of Dynamic Biological Complexes by Single-Particle Cryo-Electron Microscopy. DiIorio MC, Kulczyk AW. Micromachines (Basel) 14 118 (2022)
  6. Mechanisms of ribosome recycling in bacteria and mitochondria: a structural perspective. Seely SM, Gagnon MG. RNA Biol 19 662-677 (2022)

Articles citing this publication (19)

  1. The role of GTP hydrolysis by EF-G in ribosomal translocation. Rexroad G, Donohue JP, Lancaster L, Noller HF. Proc Natl Acad Sci U S A 119 e2212502119 (2022)
  2. Gut colonization by Bacteroides requires translation by an EF-G paralog lacking GTPase activity. Han W, Peng BZ, Wang C, Townsend GE, Barry NA, Peske F, Goodman AL, Liu J, Rodnina MV, Groisman EA. EMBO J 42 e112372 (2023)
  3. Ratchet, swivel, tilt and roll: a complete description of subunit rotation in the ribosome. Hassan A, Byju S, Freitas FC, Roc C, Pender N, Nguyen K, Kimbrough EM, Mattingly JM, Gonzalez RL, de Oliveira RJ, Dunham CM, Whitford PC. Nucleic Acids Res 51 919-934 (2023)
  4. Altered tRNA dynamics during translocation on slippery mRNA as determinant of spontaneous ribosome frameshifting. Poulis P, Patel A, Rodnina MV, Adio S. Nat Commun 13 4231 (2022)
  5. Antibiotic thermorubin tethers ribosomal subunits and impedes A-site interactions to perturb protein synthesis in bacteria. Parajuli NP, Emmerich A, Mandava CS, Pavlov MY, Sanyal S. Nat Commun 14 918 (2023)
  6. Compact IF2 allows initiator tRNA accommodation into the P site and gates the ribosome to elongation. Basu RS, Sherman MB, Gagnon MG. Nat Commun 13 3388 (2022)
  7. Cryo-electron microscopy structure and translocation mechanism of the crenarchaeal ribosome. Wang YH, Dai H, Zhang L, Wu Y, Wang J, Wang C, Xu CH, Hou H, Yang B, Zhu Y, Zhang X, Zhou J. Nucleic Acids Res 51 8909-8924 (2023)
  8. Effects of cryo-EM cooling on structural ensembles. Bock LV, Grubmüller H. Nat Commun 13 1709 (2022)
  9. High-resolution structures of a thermophilic eukaryotic 80S ribosome reveal atomistic details of translocation. Kišonaitė M, Wild K, Lapouge K, Ruppert T, Sinning I. Nat Commun 13 476 (2022)
  10. Hyper-swivel head domain motions are required for complete mRNA-tRNA translocation and ribosome resetting. Nishima W, Girodat D, Holm M, Rundlet EJ, Alejo JL, Fischer K, Blanchard SC, Sanbonmatsu KY. Nucleic Acids Res 50 8302-8320 (2022)
  11. Insights into translocation mechanism and ribosome evolution from cryo-EM structures of translocation intermediates of Giardia intestinalis. Majumdar S, Emmerich A, Krakovka S, Mandava CS, Svärd SG, Sanyal S. Nucleic Acids Res 51 3436-3451 (2023)
  12. Interplay between Inter-Subunit Rotation of the Ribosome and Binding of Translational GTPases. Das A, Adiletta N, Ermolenko DN. Int J Mol Sci 24 6878 (2023)
  13. Microsecond melting and revitrification of cryo samples with a correlative light-electron microscopy approach. Bongiovanni G, Harder OF, Drabbels M, Lorenz UJ. Front Mol Biosci 9 1044509 (2022)
  14. Modulation of translational decoding by m6A modification of mRNA. Jain S, Koziej L, Poulis P, Kaczmarczyk I, Gaik M, Rawski M, Ranjan N, Glatt S, Rodnina MV. Nat Commun 14 4784 (2023)
  15. Molecular basis of the pleiotropic effects by the antibiotic amikacin on the ribosome. Seely SM, Parajuli NP, De Tarafder A, Ge X, Sanyal S, Gagnon MG. Nat Commun 14 4666 (2023)
  16. The cyclic octapeptide antibiotic argyrin B inhibits translation by trapping EF-G on the ribosome during translocation. Wieland M, Holm M, Rundlet EJ, Morici M, Koller TO, Maviza TP, Pogorevc D, Osterman IA, Müller R, Blanchard SC, Wilson DN. Proc Natl Acad Sci U S A 119 e2114214119 (2022)
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  19. mRNA reading frame maintenance during eukaryotic ribosome translocation. Milicevic N, Jenner L, Myasnikov A, Yusupov M, Yusupova G. Nature (2023)


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