Deinococcus radiodurans
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA386750 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5571032_1.fastq.gz | Fastqsanger.gz | |||
| SRR5571032_2.fastq.gz | Fastqsanger.gz | |||
| SRR5580756_1.fastq.gz | Fastqsanger.gz | |||
| SRR5580756_2.fastq.gz | Fastqsanger.gz |
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Project description:The guided entry of tail-anchored proteins (GET) pathway assists in the proper delivery of tail-anchored (TA) proteins to the ER. Here we uncover how the yeast GET pathway components Get4/5 mediate capture of TA proteins by Sgt2, which interacts with TA sequences and hands them over to the targeting component Get3. Get4/5 binds directly and with high affinity to ribosomes, positions Sgt2 close to the ribosomal tunnel exit, and facilitates the capture of TA proteins by Sgt2. The contact sites of Get4/5 on the ribosome overlap with those of SRP, the factor mediating cotranslational ER-targeting of proteins containing internal TM domains. Exposure of nascent, internal TM domains at the tunnel exit induces high-affinity ribosome-binding of SRP, which in turn prevents ribosome-binding of Get4/5. In this way, the position of TM domains within nascent ER-targeted proteins mediates partitioning into either the GET or SRP pathway directly at the ribosomal tunnel exit.
2020-12-18 | GSE151664 | GEO
Project description:Understanding the conformational sampling of translation-arrested ribosome nascent chain complexes is key to understand co-translational folding. Up to now, coupling of cysteine oxidation, disulfide bond formation and structure formation in nascent chains has remained elusive. Here, we investigate the eye-lens protein γB-crystallin in the ribosomal exit tunnel. Using mass spectrometry, theoretical simulations, dynamic nuclear polarization-enhanced solid-state nuclear magnetic resonance and cryo-electron microscopy, we show that thiol groups of cysteine residues undergo S-glutathionylation and S-nitrosylation and form non-native disulfide bonds. Thus, covalent modification chemistry occurs already prior to nascent chain release as the ribosome exit tunnel provides sufficient space even for disulfide bond formation which can guide protein folding.
2020-11-09 | PXD021574 | Pride
Project description:Continuous translation elongation, irrespective of amino acid sequences, is a prerequisite for living organisms to produce their proteomes. However, the risk of elongation abortion is concealed within nascent polypeptide products. For example, negatively charged sequences with occasional intermittent prolines, termed intrinsic ribosome destabilization (IRD) sequences, destabilizes the translating ribosomal complex. Thus, some nascent chain sequences lead to premature translation cessation. Here, we show that most potential IRD sequences in the middle of open reading frames remain cryptic by two mechanisms: the nascent polypeptide itself that spans the exit tunnel and its bulky amino acid residues that occupy the tunnel entrance region. Thus, nascent polypeptide products have a built-in ability to ensure elongation continuity by serving as a bridge and thus by protecting the large and small ribosomal subunits from dissociation.
2021-12-08 | GSE184331 | GEO
Project description:The cellular environment is critical for efficient protein maturation, but how proteins fold during biogenesis remains poorly understood. We used hydrogen/deuterium exchange (HDX) mass spectrometry (MS) to define, at peptide resolution, the cotranslational chaperone-assisted folding pathway of Escherichia coli dihydrofolate reductase. On the ribosome, the nascent polypeptide folds via structured intermediates not populated during refolding from denaturant. Association with the ribosome allows these intermediates to form, as otherwise destabilizing C-terminal sequences remain confined in the ribosome exit tunnel. We find that partially-folded nascent chains recruit the chaperone Trigger factor, which uses a large composite hydrophobic/hydrophilic interface to engage folding intermediates without disrupting their structure. In addition, we comprehensively mapped dynamic interactions between the nascent chain and ribosomal proteins, tracing the path of the emerging polypeptide during synthesis. Our work provides a high-resolution description of de novo protein folding dynamics, thereby revealing new mechanisms by which cellular factors shape the conformational search for the native state.
2024-07-12 | PXD036945 | Pride
Project description:Cotranslational protein folding depends on general chaperones that engage highly diverse nascent chains at the ribosomes. Here we find that the universal cotranslational machinery adapts to accommodate the challenging biogenesis of abundantly expressed eukaryotic translation elongation factor 1A (eEF1A). During eEF1A synthesis, chaperone Chp1 is recruited to the ribosome with the help of the nascent polypeptide-associated complex (NAC), where it safeguards eEF1A biogenesis. Aberrant eEF1A production triggers instant proteolysis, widespread protein aggregation, activation of Hsf1 stress transcription and compromises cellular fitness. The expression of pathogenic eEF1A2 variants linked to epileptic-dyskinetic encephalopathy is protected by Chp1. Thus, eEF1A is a difficult to fold protein that necessitates dedicated folding factor Chp1 at the ribosomal tunnel exit to protect the eukaryotic cell from proteostasis collapse.
2024-01-16 | GSE221651 | GEO
Project description:The journey of a newly synthesized polypeptide starts in the peptidyltransferase center of the ribosome, from where it traverses the exit tunnel. The interior of the ribosome exit tunnel is neither straight nor smooth. How the ribosome dynamics in vivo is influenced by the exit tunnel is poorly understood. Genome-wide ribosome profiling in mammalian cells reveals elevated ribosome density at the start codon and surprisingly the downstream 5th codon position as well. We found that the highly focused ribosomal pausing shortly after initiation is attributed to the geometry of the exit tunnel, as deletion of the loop region from ribosome protein L4 diminishes translational pausing at the 5th codon position. Unexpectedly, the ribosome variant undergoes translational abandonment shortly after initiation, suggesting that there exists an obligatory step between initiation and elongation commitment. We propose that the post-initiation pausing of ribosomes represents an inherent signature of the translation machinery to ensure productive translation.
2021-02-17 | GSE166874 | GEO
Project description:Macrolides are clinically important antibiotics thought to inhibit bacterial growth by impeding the passage of newly synthesized polypeptides through the nascent peptide exit tunnel of the bacterial ribosome. Recent data challenged this view by showing that macrolide antibiotics can differentially affect synthesis of individual proteins. In order to understand the general mechanism of macrolide action, we used genome-wide ribosome profiling and analyzed the redistribution of ribosomes translating highly expressed genes in bacterial cells treated with high concentrations of macrolide antibiotics. The metagene analysis indicated that inhibition of early rounds of translation, which would be characteristic of the conventional view of macrolide action, occurs only at a limited number of genes. Translation of most genes proceeds past the 5' proximal codons and can be arrested at more distal codons when the ribosome encounters specific short sequence motifs. The sequence motifs enriched in the sites of arrest are confined to the nascent peptide residues in the peptidyl transferase center but not to the peptide segments that contact the antibiotic molecule in the exit tunnel. This led to the conclusion that the general mode of macrolide action involves selective inhibition of peptide bond formation between specific combinations of donor and acceptor substrates. Additional factors operating in the living cell but not during in vitro protein synthesis may modulate site-specific action of macrolide antibiotics. Comparing ribosome distribution in bacterial cells treated with macrolide antibiotics against the control cells.
2014-11-03 | E-GEOD-61619 | biostudies-arrayexpress
Project description:Protein folding is assisted by molecular chaperones that bind nascent polypeptides during mRNA translation. Several structurally-distinct classes of chaperone promote de novo folding, suggesting that their activities are coordinated at the ribosome. We used biochemical reconstitution and structural proteomics to explore the molecular basis for cotranslational chaperone action in bacteria. We found that chaperone binding is disfavoured close to the ribosome, allowing folding to precede chaperone recruitment. Trigger factor recognises compact folding intermediates exposing extensive unfolded surface and dictates DnaJ access to nascent chains. DnaJ uses a large surface to bind structurally diverse intermediates, and recruits DnaK to sequence-diverse solvent-accessible sites. Neither Trigger factor, DnaJ nor DnaK destabilize cotranslational folding intermediates. Instead, the chaperones collaborate to protect incipient structure in the nascent polypeptide well beyond the ribosome exit tunnel. Our findings show how the chaperone network selects and modulates cotranslational folding intermediates.
2024-07-03 | PXD048623 | Pride
Project description:We show that a single change in uL22 in the ribosomal exit tunnel can have a profound impact on gene expression, suggesting further investigation given that changes in the K90 have been clinically isolated and linked to antibiotic resistance in several species.
2021-04-06 | GSE171533 | GEO
Project description:This paper describes the cloning, purification, and characterization of thioredoxin (Trx) and thioredoxin reductase (TrxR) and the structure determination of TrxR from the ionizing radiation-tolerant bacterium Deinococcus radiodurans strain R1. The genes from D. radiodurans encoding Trx and TrxR were amplified by PCR, inserted into a pET expression vector, and overexpressed in Escherichia coli. The overexpressed proteins were purified by metal affinity chromatography, and their activity was demonstrated using well-established assays of insulin precipitation (for Trx), 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) reduction, and insulin reduction (for TrxR). In addition, the crystal structure of oxidized TrxR was determined at 1.9-A resolution. The overall structure was found to be very similar to that of E. coli TrxR and homodimeric with both NADPH- and flavin adenine dinucleotide (FAD)-binding domains containing variants of the canonical nucleotide binding fold, the Rossmann fold. The K(m) (5.7 muM) of D. radiodurans TrxR for D. radiodurans Trx was determined and is about twofold higher than that of the E. coli thioredoxin system. However, D. radiodurans TrxR has a much lower affinity for E. coli Trx (K(m), 44.4 muM). Subtle differences in the surface charge and shape of the Trx binding site on TrxR may account for the differences in recognition. Because it has been suggested that TrxR from D. radiodurans may have dual cofactor specificity (can utilize both NADH and NADPH), D. radiodurans TrxR was tested for its ability to utilize NADH as well. Our results show that D. radiodurans TrxR can utilize only NADPH for activity.
| S-EPMC2805324 | biostudies-literature