Browse
Submit Data
Databases
API
Help

Dataset Information

32 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

An intrinsically disordered nascent protein interacts with specific regions of the ribosomal surface near the exit tunnel

ABSTRACT: The influence of the ribosome on nascent chains is poorly understood, especially in the case of proteins devoid of signal or arrest sequences. Here, we provide explicit evidence for the interaction of specific ribosomal proteins with ribosome-bound nascent chains (RNCs). We target RNCs pertaining to the intrinsically disordered protein PIR and a number of mutants bearing a variable net charge. All the constructs analyzed in this work lack N-terminal signal sequences. By a combination chemical crosslinking and Western-blotting, we find that all RNCs interact with ribosomal protein L23 and that longer nascent chains also weakly interact with L29. The interacting proteins are spatially clustered on a specific region of the large ribosomal subunit, close to the exit tunnel. Based on chain-length-dependence and mutational studies, we find that the interactions with L23 persist despite drastic variations in RNC sequence. Importantly, we also find that the interactions are highly Mg+2-concentration-dependent. This work is significant because it unravels a novel role of the ribosome, which is shown to engage with the nascent protein chain even in the absence of signal or arrest sequences. Guzman-Luna et al. present a crosslinking analysis of the interaction of unstructured nascent chains with ribosomal proteins near the nascent polypeptide exit tunnel of the ribosome. They further analyze the dependence of these interactions on the peptide length, surface charge and ionic strength (including Mg+2 concentration), facilitating the understanding of co-translational protein folding and misfolding/aggregation.

SUBMITTER: Guzman-Luna V

PROVIDER: S-EPMC8556260 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Structural insights into assembly of the ribosomal nascent polypeptide exit tunnel.

Project description:The nascent polypeptide exit tunnel (NPET) is a major functional center of 60S ribosomal subunits. However, little is known about how the NPET is constructed during ribosome assembly. We utilized molecular genetics, biochemistry, and cryo-electron microscopy (cryo-EM) to investigate the functions of two NPET-associated proteins, ribosomal protein uL4 and assembly factor Nog1, in NPET assembly. Structures of mutant pre-ribosomes lacking the tunnel domain of uL4 reveal a misassembled NPET, including an aberrantly flexible ribosomal RNA helix 74, resulting in at least three different blocks in 60S assembly. Structures of pre-ribosomes lacking the C-terminal extension of Nog1 demonstrate that this extension scaffolds the tunnel domain of uL4 in the NPET to help maintain stability in the core of pre-60S subunits. Our data reveal that uL4 and Nog1 work together in the maturation of ribosomal RNA helix 74, which is required to ensure proper construction of the NPET and 60S ribosomal subunits.

| S-EPMC7547690 | biostudies-literature

A multiscale approach to sampling nascent peptide chains in the ribosomal exit tunnel.

Project description:We present a new multiscale method that combines all-atom molecular dynamics with coarse-grained sampling, towards the aim of bridging two levels of physiology: the atomic scale of protein side chains and small molecules, and the huge scale of macromolecular complexes like the ribosome. Our approach uses all-atom simulations of peptide (or other ligand) fragments to calculate local 3D spatial potentials of mean force (PMF). The individual fragment PMFs are then used as a potential for a coarse-grained chain representation of the entire molecule. Conformational space and sequence space are sampled efficiently using generalized ensemble Monte Carlo. Here, we apply this method to the study of nascent polypeptides inside the cavity of the ribosome exit tunnel. We show how the method can be used to explore the accessible conformational and sequence space of nascent polypeptide chains near the ribosome peptidyl transfer center (PTC), with the eventual aim of understanding the basis of specificity for co-translational regulation. The method has many potential applications to predicting binding specificity and design, and is sufficiently general to allow even greater separation of scales in future work.

| S-EPMC4423764 | biostudies-literature

Negamycin binds to the wall of the nascent chain exit tunnel of the 50S ribosomal subunit.

Project description:Negamycin, a small-molecule inhibitor of protein synthesis, binds the Haloarcula marismortui 50S ribosomal subunit at a single site formed by highly conserved RNA nucleotides near the cytosolic end of the nascent chain exit tunnel. The mechanism of antibiotic action and the function of this unexplored tunnel region remain intriguingly elusive.

| S-EPMC2167971 | biostudies-literature

Tertiary interactions within the ribosomal exit tunnel.

Project description:Although tertiary folding of whole protein domains is prohibited by the cramped dimensions of the ribosomal tunnel, dynamic tertiary interactions may permit folding of small elementary units within the tunnel. To probe this possibility, we used a beta-hairpin and an alpha-helical hairpin from the cytosolic N terminus of a voltage-gated potassium channel and determined a probability of folding for each at defined locations inside and outside the tunnel. Minimalist tertiary structures can form near the exit port of the tunnel, a region that provides an entropic window for initial exploration of local peptide conformations. Tertiary subdomains of the nascent peptide fold sequentially, but not independently, during translation. These studies offer an approach for diagnosing the molecular basis for folding defects that lead to protein malfunction and provide insight into the role of the ribosome during early potassium channel biogenesis.

| S-EPMC2670549 | biostudies-literature

Nascent SecM chain interacts with outer ribosomal surface to stabilize translation arrest.

Project description:SecM, a bacterial secretion monitor protein, posttranscriptionally regulates downstream gene expression via translation elongation arrest. SecM contains a characteristic amino acid sequence called the arrest sequence at its C-terminus, and this sequence acts within the ribosomal exit tunnel to stop translation. It has been widely assumed that the arrest sequence within the ribosome tunnel is sufficient for translation arrest. We have previously shown that the nascent SecM chain outside the ribosomal exit tunnel stabilizes translation arrest, but the molecular mechanism is unknown. In this study, we found that residues 57-98 of the nascent SecM chain are responsible for stabilizing translation arrest. We performed alanine/serine-scanning mutagenesis of residues 57-98 to identify D79, Y80, W81, H84, R87, I90, R91, and F95 as the key residues responsible for stabilization. The residues were predicted to be located on and near an ?-helix-forming segment. A striking feature of the ?-helix is the presence of an arginine patch, which interacts with the negatively charged ribosomal surface. A photocross-linking experiment showed that Y80 is adjacent to the ribosomal protein L23, which is located next to the ribosomal exit tunnel when translation is arrested. Thus, the folded nascent SecM chain that emerges from the ribosome exit tunnel interacts with the outer surface of the ribosome to stabilize translation arrest.

| S-EPMC6993859 | biostudies-literature

Ribosomal protein eL39 is important for maturation of the nascent polypeptide exit tunnel and proper protein folding during translation.

Project description:During translation, nascent polypeptide chains travel from the peptidyl transferase center through the nascent polypeptide exit tunnel (NPET) to emerge from 60S subunits. The NPET includes portions of five of the six 25S/5.8S rRNA domains and ribosomal proteins uL4, uL22, and eL39. Internal loops of uL4 and uL22 form the constriction sites of the NPET and are important for both assembly and function of ribosomes. Here, we investigated the roles of eL39 in tunnel construction, 60S biogenesis, and protein synthesis. We show that eL39 is important for proper protein folding during translation. Consistent with a delay in processing of 27S and 7S pre-rRNAs, eL39 functions in pre-60S assembly during middle nucleolar stages. Our biochemical assays suggest the presence of eL39 in particles at these stages, although it is not visualized in them by cryo-electron microscopy. This indicates that eL39 takes part in assembly even when it is not fully accommodated into the body of pre-60S particles. eL39 is also important for later steps of assembly, rotation of the 5S ribonucleoprotein complex, likely through long range rRNA interactions. Finally, our data strongly suggest the presence of alternative pathways of ribosome assembly, previously observed in the biogenesis of bacterial ribosomal subunits.

| S-EPMC9226512 | biostudies-literature

Intrinsically disordered proteins and proteins with intrinsically disordered regions in neurodegenerative diseases.

Project description:Many different intrinsically disordered proteins and proteins with intrinsically disordered regions are associated with neurodegenerative diseases. These types of proteins including amyloid-β, tau, α-synuclein, CHCHD2, CHCHD10, and G-protein coupled receptors are increasingly becoming evaluated as potential drug targets in the pharmaceutical-based treatment approaches. Here, we focus on the neurobiology of this class of proteins, which lie at the center of numerous neurodegenerative diseases, such as Alzheimer's and Parkinson's diseases, Huntington's disease, amyotrophic lateral sclerosis, frontotemporal dementia, Charcot-Marie-Tooth diseases, spinal muscular atrophy, and mitochondrial myopathy. Furthermore, we discuss the current treatment design strategies involving intrinsically disordered proteins and proteins with intrinsically disordered regions in neurodegenerative diseases. In addition, we emphasize that although the G-protein coupled receptors are traditionally investigated using structural biology-based models and approaches, current studies show that these receptors are proteins with intrinsically disordered regions and therefore they require new ways for their analysis.

| S-EPMC9250585 | biostudies-literature

The ribosome modulates folding inside the ribosomal exit tunnel.

Project description:Proteins commonly fold co-translationally at the ribosome, while the nascent chain emerges from the ribosomal exit tunnel. Protein domains that are sufficiently small can even fold while still located inside the tunnel. However, the effect of the tunnel on the folding dynamics of these domains is not well understood. Here, we combine optical tweezers with single-molecule FRET and molecular dynamics simulations to investigate folding of the small zinc-finger domain ADR1a inside and at the vestibule of the ribosomal tunnel. The tunnel is found to accelerate folding and stabilize the folded state, reminiscent of the effects of chaperonins. However, a simple mechanism involving stabilization by confinement does not explain the results. Instead, it appears that electrostatic interactions between the protein and ribosome contribute to the observed folding acceleration and stabilization of ADR1a.

| S-EPMC8100117 | biostudies-literature

Intrinsically disordered regions in autophagy proteins.

Project description:Autophagy is an essential eukaryotic pathway required for cellular homeostasis. Numerous key autophagy effectors and regulators have been identified, but the mechanism by which they carry out their function in autophagy is not fully understood. Our rigorous bioinformatic analysis shows that the majority of key human autophagy proteins include intrinsically disordered regions (IDRs), which are sequences lacking stable secondary and tertiary structure; suggesting that IDRs play an important, yet hitherto uninvestigated, role in autophagy. Available crystal structures corroborate the absence of structure in some of these predicted IDRs. Regions of orthologs equivalent to the IDRs predicted in the human autophagy proteins are poorly conserved, indicating that these regions may have diverse functions in different homologs. We also show that IDRs predicted in human proteins contain several regions predicted to facilitate protein-protein interactions, and delineate the network of proteins that interact with each predicted IDR-containing autophagy protein, suggesting that many of these interactions may involve IDRs. Lastly, we experimentally show that a BCL2 homology 3 domain (BH3D), within the key autophagy effector BECN1 is an IDR. This BH3D undergoes a dramatic conformational change from coil to ?-helix upon binding to BCL2s, with the C-terminal half of this BH3D constituting a binding motif, which serves to anchor the interaction of the BH3D to BCL2s. The information presented here will help inform future in-depth investigations of the biological role and mechanism of IDRs in autophagy proteins.

| S-EPMC3949125 | biostudies-literature

Cysteine oxidation and disulfide formation in the ribosomal exit tunnel.

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.

| S-EPMC7642426 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

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