Translocation as continuous movement through the ribosome

Belardinelli, Riccardo; Sharma, Heena; Peske, Frank; Wintermeyer, Wolfgang; Rodnina, Marina V.

doi:10.1080/15476286.2016.1240140

Cited by 29 publications

(37 citation statements)

References 62 publications

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“…S2B). Taking into consideration that the shift occurred in a triplet manner and that deacylated tRNA should be cognate to the E-site similarly to reverse translocation in prokaryotes (23,24) we concluded that we observed reverse translocation of the eukaryotic ribosomes.…”

Section: Post Ribosomes Relocate Backwards By Three Nucleotides In Thmentioning

confidence: 71%

“…The role of hydrolysis of GTP by elongation factor 2 in the catalysis of translocation is currently under active discussion (23). The questions that need to be clarified include how hydrolysis is related to the structural rearrangements of the ribosome and the movements of tRNA, and whether the energy released in the reaction directly drives translocation.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Eukaryotic translation elongation factor 2 (eEF2) catalyzes reverse translocation of the eukaryotic ribosome

Susorov

Zakharov

Shuvalova

et al. 2018

Journal of Biological Chemistry

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During protein synthesis, a ribosome moves along the mRNA template and, using aminoacyl-tRNAs, decodes the template nucleotide triplets to assemble a protein amino acid sequence. This movement is accompanied by shifting of mRNA-tRNA complexes within the ribosome in a process called translocation. In living cells, this process proceeds in a unidirectional manner, bringing the ribosome to the 3′-end of mRNA, and is catalyzed by the GTPase translation elongation factor 2 (EF-G in prokaryotes, eEF2 in eukaryotes). Interestingly, the possibility of spontaneous backward translocation has been shown in vitro for bacterial ribosomes, suggesting a potential reversibility of this reaction. However, this possibility has not yet been tested in eukaryotic cells. Here, using a reconstituted mammalian translation system, we show that eukaryotic elongation factor eEF2 catalyzes ribosomal reverse translocation at one mRNA triplet. We found that this process requires a cognate tRNA in the ribosomal E-site and cannot occur spontaneously without eEF2. The efficiency of this reaction depended on the concentrations of eEF2 and cognate tRNAs and increased in the presence of nonhydrolyzable GTP analogues. Of note, ADP-ribosylation of eEF2 domain IV blocked reverse translocation, suggesting a crucial role of interactions of this domain with the ribosome for the catalysis of the reaction. In summary, our findings indicate that eEF2 is able to induce ribosomal translocation in forward and backward directions highlighting the universal mechanism of tRNA-mRNA movements within the ribosome.

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Section: Post Ribosomes Relocate Backwards By Three Nucleotides In Thmentioning

confidence: 71%

Section: Introductionmentioning

confidence: 99%

Eukaryotic translation elongation factor 2 (eEF2) catalyzes reverse translocation of the eukaryotic ribosome

Susorov

Zakharov

Shuvalova

et al. 2018

Journal of Biological Chemistry

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“…After EF-G binding to the ribosome, the SSU head and body domains move in the same counterclockwise (CCW) direction relative to the LSU, which is referred to as forward, because it corresponds with the direction of translocation (Fig. 5) (Guo and Noller 2012;Belardinelli et al 2016b;Wasserman et al 2016). EF-G hydrolyzes GTP, but retains the Pi (Savelsbergh et al 2003(Savelsbergh et al , 2005.…”

Section: Translocationmentioning

confidence: 99%

Translation in Prokaryotes

Rodnina

2018

Cold Spring Harb Perspect Biol

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215

226

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This review summarizes our current understanding of translation in prokaryotes, focusing on the mechanistic and structural aspects of each phase of translation: initiation, elongation, termination, and ribosome recycling. The assembly of the initiation complex provides multiple checkpoints for messenger RNA (mRNA) and start-site selection. Correct codon-anticodon interaction during the decoding phase of elongation results in major conformational changes of the small ribosomal subunit and shapes the reaction pathway of guanosine triphosphate (GTP) hydrolysis. The ribosome orchestrates proton transfer during peptide bond formation, but requires the help of elongation factor P (EF-P) when two or more consecutive Pro residues are to be incorporated. Understanding the choreography of transfer RNA (tRNA) and mRNA movements during translocation helps to place the available structures of translocation intermediates onto the time axis of the reaction pathway. The nascent protein begins to fold cotranslationally, in the constrained space of the polypeptide exit tunnel of the ribosome. When a stop codon is reached at the end of the coding sequence, the ribosome, assisted by termination factors, hydrolyzes the ester bond of the peptidyl-tRNA, thereby releasing the nascent protein. Following termination, the ribosome is dissociated into subunits and recycled into another round of initiation. At each step of translation, the ribosome undergoes dynamic fluctuations between different conformation states. The aim of this article is to show the link between ribosome structure, dynamics, and function.

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“…Structural studies of trapped EF-G-bound translocation intermediates (19)(20)(21)(22), stopped-flow ensemble FRET experiments (23)(24)(25), and single-molecule FRET studies (9) reveal that this movement is coupled to large-scale (∼21°) rotation of the head domain of the 30S subunit ( Fig. 1).…”

mentioning

confidence: 99%