Browse
Submit Data
Databases
API
Help

Dataset Information

14 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Interaction strengths between the ribosome and tRNA at various steps of translocation.

ABSTRACT: Transfer RNA (tRNA) translocates inside the ribosome during translation. We studied the interaction strengths between the ribosome and tRNA at various stages of translocation. We utilized an optical trap to measure the mechanical force to rupture tRNA from the ribosome. We measured the rupture forces of aminoacyl tRNA or peptidyl tRNA mimic from the ribosome in a prepeptidyl transfer state, the pretranslocational state, and the posttranslocational state. In addition, we measured the interaction strength between the ribosome and aminoacyl-tRNA in presence of viomycin. Based on the interaction strengths between the ribosome and tRNA under these conditions, 1), we concluded that tRNA interaction with the 30S subunit is far more important than the interaction with the 50S subunit in the mechanism of translocation; and 2), we propose a mechanism of translocation where the ribosomal ratchet motion, with the aid of EF-G, drives tRNA translocation.

SUBMITTER: Liu CY

PROVIDER: S-EPMC3149268 | biostudies-other | 2011 May

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Publications

Interaction strengths between the ribosome and tRNA at various steps of translocation.

Liu Chen-Yu CY Qureshi Mohd Tanvir MT Lee Tae-Hee TH

Biophysical journal 20110501 9

Transfer RNA (tRNA) translocates inside the ribosome during translation. We studied the interaction strengths between the ribosome and tRNA at various stages of translocation. We utilized an optical trap to measure the mechanical force to rupture tRNA from the ribosome. We measured the rupture forces of aminoacyl tRNA or peptidyl tRNA mimic from the ribosome in a prepeptidyl transfer state, the pretranslocational state, and the posttranslocational state. In addition, we measured the interaction ...[more]

PMID: 21539788

Similar Datasets

Reverse translocation of tRNA in the ribosome.

Project description:A widely held view is that directional movement of tRNA in the ribosome is determined by an intrinsic mechanism and driven thermodynamically by transpeptidation. Here, we show that, in certain ribosomal complexes, the pretranslocation (PRE) state is thermodynamically favored over the posttranslocation (POST) state. Spontaneous and efficient conversion from the POST to PRE state is observed when EF-G is depleted from ribosomes in the POST state or when tRNA is added to the E site of ribosomes containing P-site tRNA. In the latter assay, the rate of tRNA movement is increased by streptomycin and neomycin, decreased by tetracycline, and not affected by the acylation state of the tRNA. In one case, we provide evidence that complex conversion occurs by reverse translocation (i.e., direct movement of the tRNAs from the E and P sites to the P and A sites, respectively). These findings have important implications for the energetics of translocation.

| S-EPMC2661759 | biostudies-literature

Fluctuations between multiple EF-G-induced chimeric tRNA states during translocation on the ribosome.

Project description:The coupled translocation of transfer RNA and messenger RNA through the ribosome entails large-scale structural rearrangements, including step-wise movements of the tRNAs. Recent structural work has visualized intermediates of translocation induced by elongation factor G (EF-G) with tRNAs trapped in chimeric states with respect to 30S and 50S ribosomal subunits. The functional role of the chimeric states is not known. Here we follow the formation of translocation intermediates by single-molecule fluorescence resonance energy transfer. Using EF-G mutants, a non-hydrolysable GTP analogue, and fusidic acid, we interfere with either translocation or EF-G release from the ribosome and identify several rapidly interconverting chimeric tRNA states on the reaction pathway. EF-G engagement prevents backward transitions early in translocation and increases the fraction of ribosomes that rapidly fluctuate between hybrid, chimeric and posttranslocation states. Thus, the engagement of EF-G alters the energetics of translocation towards a flat energy landscape, thereby promoting forward tRNA movement.

| S-EPMC4490557 | biostudies-literature

Interaction of tRNA with eukaryotic ribosome.

Project description:This paper is a review of currently available data concerning interactions of tRNAs with the eukaryotic ribosome at various stages of translation. These data include the results obtained by means of cryo-electron microscopy and X-ray crystallography applied to various model ribosomal complexes, site-directed cross-linking with the use of tRNA derivatives bearing chemically or photochemically reactive groups in the CCA-terminal fragment and chemical probing of 28S rRNA in the region of the peptidyl transferase center. Similarities and differences in the interactions of tRNAs with prokaryotic and eukaryotic ribosomes are discussed with concomitant consideration of the extent of resemblance between molecular mechanisms of translation in eukaryotes and bacteria.

| S-EPMC4425011 | biostudies-literature

Conserved 5-methyluridine tRNA modification modulates ribosome translocation

Project description:While the centrality of post-transcriptional modifications to RNA biology has long been acknowledged, the function of the vast majority of modified sites remains to be discovered. Illustrative of this, there is not yet a discrete biological role assigned for one the most highly conserved modifications, 5-methyluridine at position 54 in tRNAs (m5U54). Here, we uncover contributions of m5U54 to both tRNA maturation and protein synthesis. Our mass spectrometry analyses demonstrate that cells lacking the enzyme that installs m5U in the T-loop (TrmA inE. coli, Trm2 inS. cerevisiae) exhibit altered tRNA modifications patterns. Furthermore, m5U54 deficient tRNAs are desensitized to small molecules that prevent translocationin vitro.This finding is consistent with our observations that, relative to wild-type cells,trm2Δ cell growth and transcriptome-wide gene expression are less perturbed by translocation inhibitors. Together our data suggest a model in which m5U54 acts as an important modulator of tRNA maturation and translocation of the ribosome during protein synthesis.

2024-08-08 | GSE273506 | GEO

Connecting the kinetics and energy landscape of tRNA translocation on the ribosome.

Project description:Functional rearrangements in biomolecular assemblies result from diffusion across an underlying energy landscape. While bulk kinetic measurements rely on discrete state-like approximations to the energy landscape, single-molecule methods can project the free energy onto specific coordinates. With measures of the diffusion, one may establish a quantitative bridge between state-like kinetic measurements and the continuous energy landscape. We used an all-atom molecular dynamics simulation of the 70S ribosome (2.1 million atoms; 1.3 microseconds) to provide this bridge for specific conformational events associated with the process of tRNA translocation. Starting from a pre-translocation configuration, we identified sets of residues that collectively undergo rotary rearrangements implicated in ribosome function. Estimates of the diffusion coefficients along these collective coordinates for translocation were then used to interconvert between experimental rates and measures of the energy landscape. This analysis, in conjunction with previously reported experimental rates of translocation, provides an upper-bound estimate of the free-energy barriers associated with translocation. While this analysis was performed for a particular kinetic scheme of translocation, the quantitative framework is general and may be applied to energetic and kinetic descriptions that include any number of intermediates and transition states.

| S-EPMC3605090 | biostudies-literature

Conformational changes of elongation factor G on the ribosome during tRNA translocation.

Project description:The universally conserved GTPase elongation factor G (EF-G) catalyzes the translocation of tRNA and mRNA on the ribosome after peptide bond formation. Despite numerous studies suggesting that EF-G undergoes extensive conformational rearrangements during translocation, high-resolution structures exist for essentially only one conformation of EF-G in complex with the ribosome. Here, we report four atomic-resolution crystal structures of EF-G bound to the ribosome programmed in the pre- and posttranslocational states and to the ribosome trapped by the antibiotic dityromycin. We observe a previously unseen conformation of EF-G in the pretranslocation complex, which is independently captured by dityromycin on the ribosome. Our structures provide insights into the conformational space that EF-G samples on the ribosome and reveal that tRNA translocation on the ribosome is facilitated by a structural transition of EF-G from a compact to an elongated conformation, which can be prevented by the antibiotic dityromycin.

| S-EPMC4297320 | biostudies-literature

tRNA Translocation by the Eukaryotic 80S Ribosome and the Impact of GTP Hydrolysis.

Project description:Translocation moves the tRNA2?mRNA module directionally through the ribosome during the elongation phase of protein synthesis. Although translocation is known to entail large conformational changes within both the ribosome and tRNA substrates, the orchestrated events that ensure the speed and fidelity of this critical aspect of the protein synthesis mechanism have not been fully elucidated. Here, we present three high-resolution structures of intermediates of translocation on the mammalian ribosome where, in contrast to bacteria, ribosomal complexes containing the translocase eEF2 and the complete tRNA2?mRNA module are trapped by the non-hydrolyzable GTP analog GMPPNP. Consistent with the observed structures, single-molecule imaging revealed that GTP hydrolysis principally facilitates rate-limiting, final steps of translocation, which are required for factor dissociation and which are differentially regulated in bacterial and mammalian systems by the rates of deacyl-tRNA dissociation from the E site.

| S-EPMC6314685 | biostudies-literature

Cryoelectron microscopy structures of the ribosome complex in intermediate states during tRNA translocation.

Project description:mRNA-tRNA translocation is a central and highly regulated process during translational elongation. Along with the mRNA, tRNA moves through the ribosome in a stepwise fashion. Using cryoelectron microscopy on ribosomes with a P-loop mutation, we have identified novel structural intermediates likely to exist transiently during translocation. Our observations suggest a mechanism by which the rate of translocation can be regulated.

| S-EPMC3064370 | biostudies-literature

eEF3 promotes late stages of tRNA translocation on the ribosome

Project description:In addition to the conserved translation elongation factors eEF1A and eEF2, fungi require a third essential elongation factor, eEF3. While eEF3 has been implicated in tRNA binding and release at the A and E sites, its exact mechanism of action is unclear. Here we show that eEF3 acts at the mRNA–tRNA translocation step by promoting the dissociation of the tRNA from the E site, but independent of aminoacyl-tRNA recruitment to the A site. Depletion of eEF3 in vivo leads to a general slow-down in translation elongation due to accumulation of ribosomes with an occupied A site. Cryo-EM analysis of native eEF3-ribosome complexes shows that eEF3 facilitates late steps of translocation by favoring non-rotated ribosomal states as well as by opening the L1 stalk to release the E-site tRNA. Additionally, our analysis provides structural insights into novel translation elongation states, enabling presentation of a revised yeast translation elongation cycle.

2020-10-28 | GSE160206 | GEO

Domain movements of elongation factor eEF2 and the eukaryotic 80S ribosome facilitate tRNA translocation.

Project description:An 11.7-A-resolution cryo-EM map of the yeast 80S.eEF2 complex in the presence of the antibiotic sordarin was interpreted in molecular terms, revealing large conformational changes within eEF2 and the 80S ribosome, including a rearrangement of the functionally important ribosomal intersubunit bridges. Sordarin positions domain III of eEF2 so that it can interact with the sarcin-ricin loop of 25S rRNA and protein rpS23 (S12p). This particular conformation explains the inhibitory action of sordarin and suggests that eEF2 is stalled on the 80S ribosome in a conformation that has similarities with the GTPase activation state. A ratchet-like subunit rearrangement (RSR) occurs in the 80S.eEF2.sordarin complex that, in contrast to Escherichia coli 70S ribosomes, is also present in vacant 80S ribosomes. A model is suggested, according to which the RSR is part of a mechanism for moving the tRNAs during the translocation reaction.

| S-EPMC380967 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data