Corpus overview


Overview

MeSH Disease

HGNC Genes

SARS-CoV-2 proteins

NSP14 (18)

ComplexRdRp (4)

NSP5 (4)

ProteinS (3)

NSP16 (1)


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SARS-CoV-2 Proteins
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    Identifying SARS-CoV-2 Antiviral Compounds by Screening for Small Molecule Inhibitors of Nsp14/nsp10 Exoribonuclease PROTEIN

    Authors: Clovis Basier; Souradeep Basu; Rupert Beale; Agustina P Bertolin; Berta Canal; Joseph F Curran; Tom D Deegan; John FX Diffley; Lucy S Drury; Ryo Fujisawa; Michael Howell; Karim Labib; Allison W McClure; Jennifer Milligan; Rachel Ulferts; Florian Weissmann; Mary Wu; Jingkun Zeng

    doi:10.1101/2021.04.07.438812 Date: 2021-04-08 Source: bioRxiv

    SARS-CoV-2 is a coronavirus that emerged in 2019 and rapidly spread across the world causing a deadly pandemic with tremendous social and economic costs. Healthcare systems worldwide are under great pressure, and there is urgent need for effective antiviral treatments. The only currently approved antiviral treatment for COVID-19 MESHD is remdesivir, an inhibitor of viral genome replication. SARS-CoV-2 proliferation relies on the enzymatic activities of the non-structural proteins (nsp), which makes them interesting targets for the development of new antiviral treatments. With the aim to identify novel SARS-CoV-2 antivirals, we have purified the exoribonuclease PROTEIN/methyltransferase (nsp14) and its cofactor (nsp10) and developed biochemical assays compatible with high-throughput approaches to screen for exoribonuclease PROTEIN inhibitors. We have screened a library of over 5000 commercial compounds and identified patulin and aurintricarboxylic acid (ATA) as inhibitors of nsp14 exoribonuclease PROTEIN in vitro. We found that patulin and ATA inhibit replication of SARS-CoV-2 in a VERO E6 cell-culture model. These two new antiviral compounds will be valuable tools for further coronavirus research as well as potentially contributing to new therapeutic opportunities for COVID-19 MESHD.

    Identification of SARS-CoV-2 Antiviral Compounds by Screening for Small Molecule Inhibitors of the nsp14 RNA Cap Methyltransferase

    Authors: Clovis Basier; Souradeep Basu; Rupert Beale; Berta Canal; Victoria H Cowling; Joseph F Curran; Tom D Deegan; John FX Diffley; Lucy S Drury; Ryo Fujisawa; Michael Howell; Karim Labib; Chew Theng Lim; Tiffany Mak; Allison W McClure; Emma Roberts; Kang Wei Tan; Rachel Ulferts; Florian Weissmann; Mary Wu; Theresa U Zeisner

    doi:10.1101/2021.04.07.438810 Date: 2021-04-08 Source: bioRxiv

    The COVID-19 pandemic MESHD has presented itself as one of the most critical public health challenges of the century, with SARS-CoV-2 being the third member of the Coronaviridae family to cause fatal disease MESHD in humans. There is currently only one antiviral compound, remdesivir, that can be used for the treatment of COVID-19 MESHD. In order to identify additional potential therapeutics, we investigated the enzymatic proteins encoded in the SARS-CoV-2 genome. In this study, we focussed on the viral RNA cap methyltransferases, which play a key role in enabling viral protein translation and facilitating viral escape from the immune system. We expressed and purified both the guanine- N7 methyltransferase nsp14 PROTEIN, and the nsp16 2-O-methyltransferase with its activating cofactor, nsp10. We performed an in vitro high-throughput screen for inhibitors of nsp14 using a custom compound library of over 5,000 pharmaceutical compounds that have previously been characterised in either clinical or basic research. We identified 4 compounds as potential inhibitors of nsp14, all of which also show antiviral capacity in a cell based model of SARS-CoV-2 infection MESHD. Three of the 4 compounds also exhibited synergistic effects on viral replication with remdesivir.

    Structure and dynamics of SARS-CoV-2 proofreading exoribonuclease PROTEIN ExoN

    Authors: Nicholas H Moeller; Ke Shi; Özlem Demir; Surajit Banerjee; Lulu Yin; Christopher Belica; Cameron Durfee; Rommie E Amaro; Hideki Aihara

    doi:10.1101/2021.04.02.438274 Date: 2021-04-04 Source: bioRxiv

    High-fidelity replication of the large RNA genome of coronaviruses (CoVs) is mediated by a 3'-to-5' exoribonuclease PROTEIN (ExoN) in non-structural protein 14 PROTEIN (nsp14), which excises nucleotides including antiviral drugs mis-incorporated by the low-fidelity viral RNA-dependent RNA polymerase PROTEIN ( RdRp PROTEIN) and has also been implicated in viral RNA recombination and resistance to innate immunity. Here we determined a 1.6-[A] resolution crystal structure of SARS-CoV-2 ExoN in complex with its essential co-factor, nsp10. The structure shows a highly basic and concave surface flanking the active site, comprising several Lys residues of nsp14 and the N-terminal amino group of nsp10. Modeling suggests that this basic patch binds to the template strand of double-stranded RNA substrates to position the 3' end of the nascent strand in the ExoN active site, which is corroborated by mutational and computational analyses. Molecular dynamics simulations further show remarkable flexibility of multi-domain nsp14 and suggest that nsp10 stabilizes ExoN for substrate RNA-binding to support its exoribonuclease PROTEIN activity. Our high-resolution structure of the SARS-CoV-2 ExoN-nsp10 complex serves as a platform for future development of anti-coronaviral drugs or strategies to attenuate the viral virulence.

    A high-throughput fluorescence polarization assay to discover inhibitors of arenavirus and coronavirus exoribonucleases PROTEIN

    Authors: Sergio Guillermo Hernandez Tapia; Mikael Feracci; Carolina Trajano De Jesus; Priscilla El-Kazzi; Rafik Kaci; Laura Garlatti; Etienne Decroly; Bruno Canard; Francois Ferron; Karine Alvarez

    doi:10.1101/2021.04.02.437736 Date: 2021-04-02 Source: bioRxiv

    Viral exoribonucleases PROTEIN are uncommon in the world of RNA viruses. To date, this activity has been identified only in the Arenaviridae and the Coronaviridae families. These exoribonucleases PROTEIN play important but different roles in both families: for mammarenaviruses the exoribonuclease PROTEIN is involved in the suppression of the host immune response whereas for coronaviruses, exoribonuclease PROTEIN is both involved in a proofreading mechanism ensuring the genetic stability of viral genomes and participating to evasion of the host innate immunity. Because of their key roles, they constitute attractive targets for drug development. Here we present a high-throughput assay using fluorescence polarization to assess the viral exoribonuclease PROTEIN activity and its inhibition. We validate the assay using three different viral enzymes from SARS-CoV-2, lymphocytic choriomeningitis MESHD and Machupo viruses. The method is sensitive, robust, amenable to miniaturization (384 well plates) and allowed us to validate the proof-of-concept of the assay by screening a small focused compounds library (23 metal chelators). We also determined the IC50 of one inhibitor common to the three viruses.

    Synergistic Inhibition of SARS-CoV-2 Replication using Disulfiram/Ebselen and Remdesivir

    Authors: Ting Chen; Cheng-Yin Fei; Yi-Ping Chen; Karen Sargsyan; Chun Ping Chang; Hanna S. Yuan; Carmay Lim

    doi:10.26434/chemrxiv.13604015.v2 Date: 2021-03-11 Source: ChemRxiv

    The SARS-CoV-2 replication and transcription complex (RTC) comprising nonstructural protein (nsp) 2-16 plays crucial roles in viral replication, reducing the efficacy of broad-spectrum nucleoside analog drugs such as remdesivir and in evading innate immune responses. Most studies target a specific viral component of the RTC such as the main protease PROTEIN or the RNA-dependent RNA polymerase PROTEIN. In contrast, our strategy is to target multiple conserved domains of the RTC to prevent SARS-CoV-2 genome replication and to create a high barrier to viral resistance and/or evasion of antiviral drugs. We show that clinically-safe Zn-ejector drugs, disulfiram/ebselen, can target conserved Zn2+-sites in SARS-CoV-2 nsp13 and nsp14 and inhibit nsp13 ATPase and nsp14 exoribonuclease PROTEIN activities. As the SARS-CoV-2 nsp14 domain targeted by disulfiram/ebselen is involved in RNA fidelity control, our strategy allows coupling of the Zn-ejector drug with a broad-spectrum nucleoside analog that would otherwise be excised by the nsp14 proofreading domain. As proof-of-concept, we show that disulfiram/ebselen, when combined with remdesivir, can synergistically inhibit SARS-CoV-2 replication in Vero E6 cells. We present a mechanism of action and the advantages of our multi-targeting strategy, which can be applied to any type of coronavirus with conserved Zn2+-sites.

    New targets for drug design: Importance of nsp14/nsp10 complex formation for the 3'-5' exoribonucleolytic activity on SARS-CoV-2

    Authors: Margarida Saramago; Cátia Bárria; Vanessa Costa; Caio S. Souza; Sandra C. Viegas; Susana Domingues; Diana Lousa; Cláudio M. Soares; Cecília M. Arraiano; Rute G. Matos; Nuria Roca; Nuria Izquierdo-Useros; Julia Blanco; Bonaventura Clotet; Albert Bensaid; Jorge Carrillo; Julia Vergara-Alert; Joaquim Segales

    doi:10.1101/2021.01.07.425745 Date: 2021-01-07 Source: bioRxiv

    Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus has triggered a global pandemic with devastating consequences for health-care and social-economic systems. Thus, the understanding of fundamental aspects of SARS-CoV-2 is of extreme importance. In this work, we have focused our attention on the viral ribonuclease (RNase) nsp14, since this protein was considered one of the most interferon antagonists from SARS-CoV-2, and affects viral replication. This RNase is a multifunctional protein that harbors two distinct activities, an N-terminal 3'-to-5' exoribonuclease PROTEIN (ExoN) and a C-terminal N7-methyltransferase (N7-MTase), both with critical roles in coronaviruses life cycle. Namely, SARS-CoV-2 nsp14 ExoN PROTEIN knockout mutants are non-viable, indicating nsp14 as a prominent target for the development of antiviral drugs. Nsp14 ExoN activity is stimulated through the interaction with the nsp10 protein, which has a pleiotropic function during viral replication. In this study, we have performed the first biochemical characterization of the complex nsp14-nsp10 from SARS-CoV-2. Here we confirm the 3'-5' exoribonuclease PROTEIN and MTase activities of nsp14 in this new Coronavirus, and the critical role of nsp10 in upregulating the nsp14 ExoN PROTEIN activity in vitro. Furthermore, we demonstrate that SARS-CoV-2 nsp14 N7-MTase activity is functionally independent of the ExoN activity. The nsp14 MTase activity also seems to be independent of the presence of nsp10 cofactor, contrarily to nsp14 ExoN PROTEIN. Until now, there is no available structure for the SARS-CoV-2 nsp14-nsp10 complex. As such, we have modelled the SARS-CoV-2 nsp14-nsp10 complex based on the 3D structure of the complex from SARS-CoV MESHD (PDB ID 5C8S). We also have managed to map key nsp10 residues involved in its interaction with nsp14, all of which are also shown to be essential for stimulation of the nsp14 ExoN PROTEIN activity. This reinforces the idea that a stable interaction between nsp10 and nsp14 is strictly required for the nsp14-mediated ExoN activity of SARS-CoV-2, as observed for SARS-CoV MESHD. We have studied the role of conserved DEDD catalytic residues of SARS-CoV-2 nsp14 ExoN PROTEIN. Our results show that motif I of ExoN domain is essential for the nsp14 function contrasting to the functionality of these conserved catalytic residues in SARS-CoV MESHD, and in the Middle East respiratory syndrome coronavirus MESHD (MERS). The differences here revealed can have important implications regarding the specific pathogenesis of SARS-CoV-2. The nsp10-nsp14 interface is a recognized attractive target for antivirals against SARS-CoV-2 and other coronaviruses. This work has unravelled a basis for discovering inhibitors targeting the specific amino acids here reported, in order to disrupt the assembly of this complex and interfere with coronaviruses replication.

    Genomic diversity of SARS-CoV-2 can be accelerated by a mutation in the nsp14 gene

    Authors: Kosuke Takada; Mahoko Takahashi Ueda; Tokiko Watanabe; So Nakagawa

    doi:10.1101/2020.12.23.424231 Date: 2020-12-26 Source: bioRxiv

    Nucleotide substitution rate of severe acute respiratory syndrome coronavirus 2 MESHD (SARS-CoV-2) is relatively low compared to the other RNA viruses because coronaviruses including SARS-CoV-2 encode non-structural protein 14 PROTEIN (nsp14) that is an error-correcting exonuclease protein. In this study, to understand genome evolution of SARS-CoV-2 in the current pandemic, we examined mutations of SARS-CoV-2 nsp14 which could inhibit its error-correcting function. First, to obtain functionally important sites of nsp14, we examined 62 representative coronaviruses belonging to alpha, beta, gamma, delta, and unclassified coronaviruses. As a result, 99 out of 527 amino acid sites of nsp14 were evolutionarily conserved. We then examined nsp14 sequences obtained from 28,082 SARS-CoV-2 genomes and identified 6 amino acid changes in nsp14 mutants that were not detected in the 62 representative coronaviruses. We examined genome substitution rates of these mutants and found that an nsp14 mutant with a proline to leucine change at position 203 (P203L) showed a higher substitution rate (35.9 substitutions/year) than SARS-CoV-2 possessing wild-type nsp14 (19.8 substitutions/year). We confirmed that the substitution rate of the P203L is significantly higher than those of other variants containing mutations in structural proteins. Although the number of SARS-CoV-2 variants containing P203L mutation of nsp14 is limited (26), these mutants appeared at least 10 times independently in the current pandemic. These results indicated that the molecular function of nsp14 is important for survival of various coronaviruses including SARS-CoV-2 and that some mutations such as P203L of nsp14 inhibiting its error-correcting function are removed rapidly due to their deleterious effects.

    SARS-CoV-2 antibody signatures for predicting the outcome of COVID-19 MESHD

    Authors: Qing Lei; Caizheng Yu; Yang Li; Hongyan Hou; Zhaowei Xu; Meian He; Ziyong Sun; Feng Wang; Sheng-ce Tao; Xionglin Fan

    doi:10.1101/2020.11.10.20228890 Date: 2020-11-13 Source: medRxiv

    The COIVD-19 global pandemic is far from ending. There is an urgent need to identify applicable biomarkers for predicting the outcome of COVID-19 MESHD. Growing evidences have revealed that SARS-CoV-2 specific antibodies remain elevated with disease progression and severity in COIVD-19 patients. We assumed that antibodies may serve as biomarkers for predicting disease outcome. By taking advantage of a newly developed SARS-CoV-2 proteome microarray, we surveyed IgM/ IgG responses against 20 SARS-CoV-2 proteins in 1,034 hospitalized COVID-19 MESHD patients on admission, who were followed till 66 days. The microarray results were correlated with clinical information, laboratory test results and patient outcomes. Cox proportional hazards model was used to explore the association between SARS-CoV-2 specific antibodies and COVID-19 MESHD mortality. We found that high level of IgM against ORF7b PROTEIN at the time of hospitalization is an independent predictor of patient survival (p trend = 0.002), while levels of IgG responses to 6 non-structural proteins PROTEIN and 1 accessory protein, i. e PROTEIN., NSP4 HGNC NSP4 PROTEIN, NSP7 PROTEIN, NSP9 PROTEIN, NSP10 PROTEIN, RdRp PROTEIN ( NSP12 PROTEIN), NSP14 PROTEIN, and ORF3b PROTEIN, possess significant predictive power for patient death MESHD, even after further adjustments for demographics, comorbidities, and common laboratory markers for disease severity (all with p trend < 0.05). Spline regression analysis indicated that the correlation between ORF7b PROTEIN IgM, NSP9 PROTEIN IgG, and NSP10 PROTEIN IgG and risk of COVID-19 MESHD mortality is linear (p = 0.0013, 0.0073 and 0.0003, respectively). Their AUCs for predictions, determined by computational cross-validations (validation1), were 0.74 (cut-off = 7.59), 0.66 (cut-off = 9.13), and 0.68 (cut-off = 6.29), respectively. Further validations were conducted in the second and third serial samples of these cases (validation2A, n = 633, validation2B, n = 382), with high accuracy of prediction for outcome. These findings have important implications for improving clinical management, and especially for developing medical interventions and vaccines.

    In silico studies suggest T-cell cross-reactivity between SARS-CoV-2 and less dangerous coronaviruses

    Authors: Marcin Pacholczyk; Piotr Rieske

    doi:10.21203/rs.3.rs-73773/v1 Date: 2020-09-07 Source: ResearchSquare

    So far, it is impossible to explain the diverse individual and population responses to SARS-CoV-2 infection MESHD. Many factors may be involved, including genetics, diet, vaccinations, the innate immune response, viral load, and other phenomena. Further, immune responses raised against pathogens other than SARS-CoV-2 (cross-reactivity) may also be involved. In this work, we analyzed the potential for T-cell cross-reactivity between less contagious coronaviruses (HCoV-OC43, HCoV-HKU1, HCoV-229E, and HCoV-NL63) and SARS-CoV-2. In silico research suggests that SARS-CoV-2 and less dangerous coronaviruses share identical peptides, which can be presented on MHC class I molecules. Those T-cells epitopes belong to several coronavirus proteins localized inside the viral envelope, including helicase HGNC, RNA polymerase, proofreading exoribonuclease PROTEIN, and 2'-O-methyltransferase. Our data suggest that a milder course of COVID-19 MESHD, in some populations, may be related to the cross-reactivity of T cells.

    Characterisation of the SARS-CoV-2 ExoN (nsp14ExoN-nsp10) complex: implications for its role in viral genome stability and inhibitor identification

    Authors: Hannah T Baddock; Sanja Brolih; Yuliana Yosaatmadja; Malitha Ratnaweera; Marcin Bielinski; Lonnie Swift; Abimael Cruz-Migoni; Garrett M Morris; Christopher J Schofield; Opher Gileadi; Peter J McHugh

    doi:10.1101/2020.08.13.248211 Date: 2020-08-13 Source: bioRxiv

    The SARS-CoV-2 coronavirus MESHD (CoV) causes COVID-19 MESHD, a current global pandemic. SARS-CoV-2 belongs to an order of Nidovirales with very large RNA genomes. It is proposed that the fidelity of CoV genome replication is aided by an RNA nuclease complex, formed of non-structural proteins 14 PROTEIN and 10 (nsp14-nsp10), an attractive target for antiviral inhibition. Here, we confirm that the SARS-CoV-2 nsp14-nsp10 complex is an RNase. Detailed functional characterisation reveals nsp14-nsp10 is a highly versatile nuclease capable of digesting a wide variety of RNA structures, including those with a blocked 3-terminus. We propose that the role of nsp14-nsp10 in maintaining replication fidelity goes beyond classical proofreading and purges the nascent replicating RNA strand of a range of potentially replication terminating aberrations. Using the developed assays, we identify a series of drug and drug-like molecules that potently inhibit nsp14-nsp10, including the known Sars-Cov-2 major protease ( Mpro PROTEIN) inhibitor ebselen and the HIV integrase inhibitor raltegravir, revealing the potential for bifunctional inhibitors in the treatment of COVID-19 MESHD.

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MeSH Disease
HGNC Genes
SARS-CoV-2 Proteins


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