Browse
Submit Data
Databases
API
Help

Dataset Information

6 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Delicate structural coordination of the Severe Acute Respiratory Syndrome coronavirus Nsp13 upon ATP hydrolysis.

ABSTRACT: To date, an effective therapeutic treatment that confers strong attenuation toward coronaviruses (CoVs) remains elusive. Of all the potential drug targets, the helicase of CoVs is considered to be one of the most important. Here, we first present the structure of the full-length Nsp13 helicase of SARS-CoV (SARS-Nsp13) and investigate the structural coordination of its five domains and how these contribute to its translocation and unwinding activity. A translocation model is proposed for the Upf1-like helicase members according to three different structural conditions in solution characterized through H/D exchange assay, including substrate state (SARS-Nsp13-dsDNA bound with AMPPNP), transition state (bound with ADP-AlF4-) and product state (bound with ADP). We observed that the β19-β20 loop on the 1A domain is involved in unwinding process directly. Furthermore, we have shown that the RNA dependent RNA polymerase (RdRp), SARS-Nsp12, can enhance the helicase activity of SARS-Nsp13 through interacting with it directly. The interacting regions were identified and can be considered common across CoVs, which provides new insights into the Replication and Transcription Complex (RTC) of CoVs.

SUBMITTER: Jia Z

PROVIDER: S-EPMC6614802 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Structural Proteins in Severe Acute Respiratory Syndrome Coronavirus-2.

Project description:What began with a sign of pneumonia-related respiratory disorders in China has now become a pandemic named by WHO as Covid-19 known to be caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). The SARS-CoV-2 are newly emerged β coronaviruses belonging to the Coronaviridae family. SARS-CoV-2 has a positive viral RNA genome expressing open reading frames that code for structural and non-structural proteins. The structural proteins include spike (S), nucleocapsid (N), membrane (M), and envelope (E) proteins. The S1 subunit of S protein facilitates ACE2 mediated virus attachment while S2 subunit promotes membrane fusion. The presence of glutamine, asparagine, leucine, phenylalanine and serine amino acids in SARS-CoV-2 enhances ACE2 binding. The N protein is composed of a serine-rich linker region sandwiched between N Terminal Domain (NTD) and C Terminal Domain (CTD). These terminals play a role in viral entry and its processing post entry. The NTD forms orthorhombic crystals and binds to the viral genome. The linker region contains phosphorylation sites that regulate its functioning. The CTD promotes nucleocapsid formation. The E protein contains a NTD, hydrophobic domain and CTD which form viroporins needed for viral assembly. The M protein possesses hydrophilic C terminal and amphipathic N terminal. Its long-form promotes spike incorporations and the interaction with E facilitates virion production. As each protein is essential in viral functioning, this review describes the insights of SARS-CoV-2 structural proteins that would help in developing therapeutic strategies by targeting each protein to curb the rapidly growing pandemic.

| S-EPMC7247499 | biostudies-literature

The Role of ATP Hydrolysis and Product Release in the Translocation Mechanism of SARS-CoV-2 NSP13.

Project description:In response to the emergence of COVID-19, caused by SARS-CoV-2, there has been a growing interest in understanding the functional mechanisms of the viral proteins to aid in the development of new therapeutics. Non-structural protein 13 (Nsp13) helicase is an attractive target for antivirals because it is essential for viral replication and has a low mutation rate; yet, the structural mechanisms by which this enzyme binds and hydrolyzes ATP to cause unidirectional RNA translocation remain elusive. Using Gaussian accelerated molecular dynamics (GaMD), we generated a comprehensive conformational ensemble of all substrate states along the ATP-dependent cycle. ShapeGMM clustering of the protein yields four protein conformations that describe an opening and closing of both the ATP pocket and RNA cleft. This opening and closing is achieved through a combination of conformational selection and induction along the ATP cycle. Furthermore, three protein-RNA conformations are observed that implicate motifs Ia, IV, and V as playing a pivotal role in an ATP-dependent inchworm translocation mechanism. Finally, based on a linear discriminant analysis of protein conformations, we identify L405 as a pivotal residue for the opening and closing mechanism and propose a L405D mutation as a way of testing our proposed mechanism. This research enhances our understanding of nsp13's role in viral replication and could contribute to the development of antiviral strategies.

| S-EPMC10557736 | biostudies-literature

Coordination of substrate binding and ATP hydrolysis in Vps4-mediated ESCRT-III disassembly.

Project description:ESCRT-III undergoes dynamic assembly and disassembly to facilitate membrane exvagination processes including multivesicular body (MVB) formation, enveloped virus budding, and membrane abscission during cytokinesis. The AAA-ATPase Vps4 is required for ESCRT-III disassembly, however the coordination of Vps4 ATP hydrolysis with ESCRT-III binding and disassembly is not understood. Vps4 ATP hydrolysis has been proposed to execute ESCRT-III disassembly as either a stable oligomer or an unstable oligomer whose dissociation drives ESCRT-III disassembly. An in vitro ESCRT-III disassembly assay was developed to analyze Vps4 function during this process. The studies presented here support a model in which Vps4 acts as a stable oligomer during ATP hydrolysis and ESCRT-III disassembly. Moreover, Vps4 oligomer binding to ESCRT-III induces coordination of ATP hydrolysis at the level of individual Vps4 subunits. These results suggest that Vps4 functions as a stable oligomer that acts upon individual ESCRT-III subunits to facilitate ESCRT-III disassembly.

| S-EPMC2947475 | biostudies-literature

Structural and functional analyses of the severe acute respiratory syndrome coronavirus endoribonuclease Nsp15.

Project description:The severe acute respiratory syndrome (SARS) coronavirus encodes several RNA-processing enzymes that are unusual for RNA viruses, including Nsp15 (nonstructural protein 15), a hexameric endoribonuclease that preferentially cleaves 3' of uridines. We solved the structure of a catalytically inactive mutant version of Nsp15, which was crystallized as a hexamer. The structure contains unreported flexibility in the active site of each subunit. Substitutions in the active site residues serine 293 and proline 343 allowed Nsp15 to cleave at cytidylate, whereas mutation of leucine 345 rendered Nsp15 able to cleave at purines as well as pyrimidines. Mutations that targeted the residues involved in subunit interactions generally resulted in the formation of catalytically inactive monomers. The RNA-binding residues were mapped by a method linking reversible cross-linking, RNA affinity purification, and peptide fingerprinting. Alanine substitution of several residues in the RNA-contacting portion of Nsp15 did not affect hexamer formation but decreased the affinity of RNA binding and reduced endonuclease activity. This suggests a model for Nsp15 hexamer interaction with RNA.

| S-EPMC8740563 | biostudies-literature

Structural characterization of the fusion-active complex of severe acute respiratory syndrome (SARS) coronavirus.

Project description:The causative agent of a recent outbreak of an atypical pneumonia, known as severe acute respiratory syndrome (SARS), has been identified as a coronavirus (CoV) not belonging to any of the previously identified groups. Fusion of coronaviruses with the host cell is mediated by the envelope spike protein. Two regions within the spike protein of SARS-CoV have been identified, showing a high degree of sequence conservation with the other CoV, which are characterized by the presence of heptad repeats (HR1 and HR2). By using synthetic and recombinant peptides corresponding to the HR1 and HR2 regions, we were able to characterize the fusion-active complex formed by this novel CoV by CD, native PAGE, proteolysis protection analysis, and size-exclusion chromatography. HR1 and HR2 of SARS-CoV associate into an antiparallel six-helix bundle, with structural features typical of the other known class I fusion proteins. We have also mapped the specific boundaries of the region, within the longer HR1 domain, making contact with the shorter HR2 domain. Notably, the inner HR1 coiled coil is a stable alpha-helical domain even in the absence of interaction with the HR2 region. Inhibitors binding to HR regions of fusion proteins have been shown to be efficacious against many viruses, notably HIV. Our results may help in the design of anti-SARS therapeutics.

| S-EPMC423260 | biostudies-literature

Crystal Structure of Non-Structural Protein 10 from Severe Acute Respiratory Syndrome Coronavirus-2.

Project description:Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), causing Coronavirus Disease 19 (COVID-19), emerged at the end of 2019 and quickly spread to cause a global pandemic with severe socio-economic consequences. The early sequencing of its RNA genome revealed its high similarity to SARS, likely to have originated from bats. The SARS-CoV-2 non-structural protein 10 (nsp10) displays high sequence similarity with its SARS homologue, which binds to and stimulates the 3'-to-5' exoribonuclease and the 2'-O-methlytransferase activities of nsps 14 and 16, respectively. Here, we report the biophysical characterization and 1.6 Å resolution structure of the unbound form of nsp10 from SARS-CoV-2 and compare it to the structures of its SARS homologue and the complex-bound form with nsp16 from SARS-CoV-2. The crystal structure and solution behaviour of nsp10 will not only form the basis for understanding the role of SARS-CoV-2 nsp10 as a central player of the viral RNA capping apparatus, but will also serve as a basis for the development of inhibitors of nsp10, interfering with crucial functions of the replication-transcription complex and virus replication.

| S-EPMC7583907 | biostudies-literature

Severe acute respiratory syndrome coronavirus envelope protein regulates cell stress responses and apoptosis

Project description:Severe acute respiratory syndrome virus (SARS-CoV) that lacks the envelope (E) gene (rSARS-CoV-ΔE) is attenuated in vivo [1,2]. To identify factors that contribute to rSARS-CoV-ΔE attenuation, gene expression in cells infected by SARS-CoV with or without E gene was compared. Twenty-five stress response genes were preferentially upregulated during infection in the absence of the E gene. In addition, genes involved in signal transduction, transcription, cell metabolism, immunoregulation, inflammation, apoptosis and cell cycle and differentiation were differentially regulated in cells infected with rSARS-CoV with or without the E gene. Administration of E protein in trans reduced the stress response in cells infected with rSARS-CoV-ΔE, with respiratory syncytial virus, or treated with drugs, such as tunicamycin and thapsigarcin that elicit cell stress by different mechanisms. In addition, SARS-CoV E protein down-regulated the signaling pathway inositol-requiring enzyme 1 (IRE-1) of the unfolded protein response, but not the PKR-like ER kinase (PERK) or activating transcription factor 6 (ATF-6) pathways, and reduced cell apoptosis. Overall, the activation of the IRE-1 pathway was not able to restore cell homeostasis, and apoptosis was induced probably as a meassure to protect the host by limiting virus production and dissemination. The expression of proinflammatory cytokines was reduced in rSARS-CoV-ΔE-infected cells compared to rSARS-CoV-infected cells, suggesting that the increase in stress responses and the reduction of inflammation in the absence of the E gene contributed to the attenuation of rSARS-CoV-ΔE. We used Affymetrix microarrays (Human Genome U133 plus 2.0) to compare global gene expression between SARS-CoV-infected, mock-infected and SARS-CoV-ΔE-infected cells. For ech type of sample three hybridizations were carried-out (independent biological replicates).

2012-06-20 | E-GEOD-30589 | biostudies-arrayexpress

Nitric oxide inhibits the replication cycle of severe acute respiratory syndrome coronavirus.

Project description:Nitric oxide (NO) is an important signaling molecule between cells which has been shown to have an inhibitory effect on some virus infections. The purpose of this study was to examine whether NO inhibits the replication cycle of the severe acute respiratory syndrome coronavirus (SARS CoV) in vitro. We found that an organic NO donor, S-nitroso-N-acetylpenicillamine, significantly inhibited the replication cycle of SARS CoV in a concentration-dependent manner. We also show here that NO inhibits viral protein and RNA synthesis. Furthermore, we demonstrate that NO generated by inducible nitric oxide synthase, an enzyme that produces NO, inhibits the SARS CoV replication cycle.

| S-EPMC544093 | biostudies-other

Resolution of primary severe acute respiratory syndrome-associated coronavirus infection requires Stat1.

Project description:Intranasal inhalation of the severe acute respiratory syndrome coronavirus (SARS CoV) in the immunocompetent mouse strain 129SvEv resulted in infection of conducting airway epithelial cells followed by rapid clearance of virus from the lungs and the development of self-limited bronchiolitis. Animals resistant to the effects of interferons by virtue of a deficiency in Stat1 demonstrated a markedly different course following intranasal inhalation of SARS CoV, one characterized by replication of virus in lungs and progressively worsening pulmonary disease with inflammation of small airways and alveoli and systemic spread of the virus to livers and spleens.

| S-EPMC521834 | biostudies-other

CD209L (L-SIGN) is a receptor for severe acute respiratory syndrome coronavirus.

Project description:Angiotensin-converting enzyme 2 (ACE2) is a receptor for SARS-CoV, the novel coronavirus that causes severe acute respiratory syndrome [Li, W. Moore, M. J., Vasilieva, N., Sui, J., Wong, S. K., Berne, M. A., Somasundaran, M., Sullivan, J. L., Luzuriaga, K., Greenough, T. C., et al. (2003) Nature 426, 450-454]. We have identified a different human cellular glycoprotein that can serve as an alternative receptor for SARS-CoV. A human lung cDNA library in vesicular stomatitis virus G pseudotyped retrovirus was transduced into Chinese hamster ovary cells, and the cells were sorted for binding of soluble SARS-CoV spike (S) glycoproteins, S(590) and S(1180). Clones of transduced cells that bound SARS-CoV S glycoprotein were inoculated with SARS-CoV, and increases in subgenomic viral RNA from 1-16 h or more were detected by multiplex RT-PCR in four cloned cell lines. Sequencing of the human lung cDNA inserts showed that each of the cloned cell lines contained cDNA that encoded human CD209L, a C-type lectin (also called L-SIGN). When the cDNA encoding CD209L from clone 2.27 was cloned and transfected into Chinese hamster ovary cells, the cells expressed human CD209L glycoprotein and became susceptible to infection with SARS-CoV. Immunohistochemistry showed that CD209L is expressed in human lung in type II alveolar cells and endothelial cells, both potential targets for SARS-CoV. Several other enveloped viruses including Ebola and Sindbis also use CD209L as a portal of entry, and HIV and hepatitis C virus can bind to CD209L on cell membranes but do not use it to mediate virus entry. Our data suggest that the large S glycoprotein of SARS-CoV may use both ACE2 and CD209L in virus infection and pathogenesis.

| S-EPMC524836 | biostudies-other

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