Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 21.257
Filtrar
1.
BMC Mol Cell Biol ; 22(1): 23, 2021 Apr 23.
Artículo en Inglés | MEDLINE | ID: mdl-33892639

RESUMEN

BACKGROUND: The SARS-CoV-2 virus, the causative agent of COVID-19, consists of an assembly of proteins that determine its infectious and immunological behavior, as well as its response to therapeutics. Major structural biology efforts on these proteins have already provided essential insights into the mode of action of the virus, as well as avenues for structure-based drug design. However, not all of the SARS-CoV-2 proteins, or regions thereof, have a well-defined three-dimensional structure, and as such might exhibit ambiguous, dynamic behaviour that is not evident from static structure representations, nor from molecular dynamics simulations using these structures. MAIN: We present a website ( https://bio2byte.be/sars2/ ) that provides protein sequence-based predictions of the backbone and side-chain dynamics and conformational propensities of these proteins, as well as derived early folding, disorder, ß-sheet aggregation, protein-protein interaction and epitope propensities. These predictions attempt to capture the inherent biophysical propensities encoded in the sequence, rather than context-dependent behaviour such as the final folded state. In addition, we provide the biophysical variation that is observed in homologous proteins, which gives an indication of the limits of their functionally relevant biophysical behaviour. CONCLUSION: The https://bio2byte.be/sars2/ website provides a range of protein sequence-based predictions for 27 SARS-CoV-2 proteins, enabling researchers to form hypotheses about their possible functional modes of action.


Asunto(s)
/química , Proteínas Virales/química , Bases de Datos de Proteínas , Humanos , Acceso a Internet , Alineación de Secuencia , Análisis de Secuencia de Proteína , Programas Informáticos , Proteínas Virales/metabolismo
2.
Viruses ; 13(4)2021 03 26.
Artículo en Inglés | MEDLINE | ID: mdl-33810401

RESUMEN

The risk posed by Severe Acute Respiratory Syndrome Coronavirus -2 (SARS-CoV-2) dictates that live-virus research is conducted in a biosafety level 3 (BSL3) facility. Working with SARS-CoV-2 at lower biosafety levels can expedite research yet requires the virus to be fully inactivated. In this study, we validated and compared two protocols for inactivating SARS-CoV-2: heat treatment and ultraviolet irradiation. The two methods were optimized to render the virus completely incapable of infection while limiting the destructive effects of inactivation. We observed that 15 min of incubation at 65 °C completely inactivates high titer viral stocks. Complete inactivation was also achieved with minimal amounts of UV power (70,000 µJ/cm2), which is 100-fold less power than comparable studies. Once validated, the two methods were then compared for viral RNA quantification, virion purification, and antibody detection assays. We observed that UV irradiation resulted in a 2-log reduction of detectable genomes compared to heat inactivation. Protein yield following virion enrichment was equivalent for all inactivation conditions, but the quality of resulting viral proteins and virions were differentially impacted depending on inactivation method and time. Here, we outline the strengths and weaknesses of each method so that investigators might choose the one which best meets their research goals.


Asunto(s)
/virología , Desinfección/métodos , Virión/efectos de la radiación , Inactivación de Virus/efectos de la radiación , Desinfección/instrumentación , Calor , Humanos , /genética , Rayos Ultravioleta , Proteínas Virales/genética , Proteínas Virales/metabolismo , Virión/química , Virión/genética , Virión/fisiología
3.
J Gen Virol ; 102(4)2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33855951

RESUMEN

SARS-CoV-2 is thought to have originated in the human population from a zoonotic spillover event. Infection in humans results in a variety of outcomes ranging from asymptomatic cases to the disease COVID-19, which can have significant morbidity and mortality, with over two million confirmed deaths worldwide as of January 2021. Over a year into the pandemic, sequencing analysis has shown that variants of SARS-CoV-2 are being selected as the virus continues to circulate widely within the human population. The predominant drivers of genetic variation within SARS-CoV-2 are single nucleotide polymorphisms (SNPs) caused by polymerase error, potential host factor driven RNA modification, and insertion/deletions (indels) resulting from the discontinuous nature of viral RNA synthesis. While many mutations represent neutral 'genetic drift' or have quickly died out, a subset may be affecting viral traits such as transmissibility, pathogenicity, host range, and antigenicity of the virus. In this review, we summarise the current extent of genetic change in SARS-CoV-2, particularly recently emerging variants of concern, and consider the phenotypic consequences of this viral evolution that may impact the future trajectory of the pandemic.


Asunto(s)
Adaptación Fisiológica/genética , /genética , Animales , Antígenos Virales/genética , Antígenos Virales/inmunología , /inmunología , Humanos , Evasión Inmune/genética , Mutación , ARN Viral/biosíntesis , ARN Viral/genética , /patogenicidad , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/metabolismo , Proteínas Virales/genética , Proteínas Virales/metabolismo , Internalización del Virus , Replicación Viral , Zoonosis/transmisión , Zoonosis/virología
4.
Arch Immunol Ther Exp (Warsz) ; 69(1): 10, 2021 Apr 03.
Artículo en Inglés | MEDLINE | ID: mdl-33811524

RESUMEN

The review discusses a new approach to the prevention and treatment of viral infections based on the use of pine needles polyprenyl phosphate (PPP) and associated with the infringement of prenylation process-the attachment of farnesol or geranyl geraniol to the viral protein. Currently, prenylation has been detected in type 1 adenovirus, hepatitis C virus, several herpes viruses, influenza virus, HIV. However, this list is far from complete, given that prenylated proteins play an extremely important role in the activity of the virus. We assume that the interferon produced in response to PPP may suppress expression of the SREBP2 transcription factor. As a result, the mevalonic acid pathway is violated and, as a result, the formation of early polyprenols precursors (geraniol, geranyl geraniol, farnesol), which are necessary for the prenylation of viral proteins, is blocked and the formation of mature, virulent virus particles is broken. As a consequence, the maturation of viral particles is inhibited, and defective particles are formed. Polyprenol was extracted from greenery (pine, fir and spruce needles, mulberry leaves, etc.), purified by chromatography, phosphorylated and identified by HPLC and NMR. Obtained PPP was used as antiviral in some experimental models in vitro and in vivo. During numerous studies, it was found that PPP manifested versatile antiviral effects, both in vitro and in vivo. The maximum effect was observed with viruses in which the presence of prenylated proteins was established, namely influenza A virus, HIV-1, tick-borne encephalitis virus, hepatitis A and C viruses, herpes simplex viruses type 1 and 2, some coronavirus. The available data obtained both in the experimental conditions and during clinical trials allow us to regard PPPs as safe and effective medicine for prevention and treatment of viral diseases.


Asunto(s)
Antivirales/farmacología , Pinus/química , Fosfatos de Poliisoprenilo/farmacología , Prenilación de Proteína/efectos de los fármacos , Virosis/tratamiento farmacológico , Animales , Antivirales/uso terapéutico , Ensayos Clínicos como Asunto , Modelos Animales de Enfermedad , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/inmunología , Humanos , Interferones/metabolismo , Microscopía Electrónica , Fosfatos de Poliisoprenilo/uso terapéutico , Proteína 2 de Unión a Elementos Reguladores de Esteroles/metabolismo , Resultado del Tratamiento , Proteínas Virales/metabolismo , Virión/efectos de los fármacos , Virión/ultraestructura , Virosis/inmunología , Virosis/prevención & control , Replicación Viral/efectos de los fármacos , Replicación Viral/inmunología
5.
Proc Natl Acad Sci U S A ; 118(17)2021 04 27.
Artículo en Inglés | MEDLINE | ID: mdl-33827988

RESUMEN

In order to understand the transmission and virulence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), it is necessary to understand the functions of each of the gene products encoded in the viral genome. One feature of the SARS-CoV-2 genome that is not present in related, common coronaviruses is ORF10, a putative 38-amino acid protein-coding gene. Proteomic studies found that ORF10 binds to an E3 ubiquitin ligase containing Cullin-2, Rbx1, Elongin B, Elongin C, and ZYG11B (CRL2ZYG11B). Since CRL2ZYG11B mediates protein degradation, one possible role for ORF10 is to "hijack" CRL2ZYG11B in order to target cellular, antiviral proteins for ubiquitylation and subsequent proteasomal degradation. Here, we investigated whether ORF10 hijacks CRL2ZYG11B or functions in other ways, for example, as an inhibitor or substrate of CRL2ZYG11B While we confirm the ORF10-ZYG11B interaction and show that the N terminus of ORF10 is critical for it, we find no evidence that ORF10 is functioning to inhibit or hijack CRL2ZYG11B Furthermore, ZYG11B and its paralog ZER1 are dispensable for SARS-CoV-2 infection in cultured cells. We conclude that the interaction between ORF10 and CRL2ZYG11B is not relevant for SARS-CoV-2 infection in vitro.


Asunto(s)
/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas Cullin/metabolismo , Complejos Multiproteicos/metabolismo , Sistemas de Lectura Abierta , Proteínas Virales/metabolismo , /genética , Proteínas de Ciclo Celular/genética , Proteínas Cullin/genética , Células HEK293 , Humanos , Complejos Multiproteicos/genética , Proteínas Virales/genética
6.
Nat Commun ; 12(1): 2061, 2021 04 06.
Artículo en Inglés | MEDLINE | ID: mdl-33824318

RESUMEN

Stress can induce cell surface expression of MHC-like ligands, including MICA, that activate NK cells. Human cytomegalovirus (HCMV) glycoprotein US9 downregulates the activating immune ligand MICA*008 to avoid NK cell activation, but the underlying mechanism remains unclear. Here, we show that the N-terminal signal peptide is the major US9 functional domain targeting MICA*008 to proteasomal degradation. The US9 signal peptide is cleaved with unusually slow kinetics and this transiently retained signal peptide arrests MICA*008 maturation in the endoplasmic reticulum (ER), and indirectly induces its degradation via the ER quality control system and the SEL1L-HRD1 complex. We further identify an accessory, signal peptide-independent US9 mechanism that directly binds MICA*008 and SEL1L. Collectively, we describe a dual-targeting immunoevasin, demonstrating that signal peptides can function as protein-integral effector domains.


Asunto(s)
Evasión Inmune , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/metabolismo , Señales de Clasificación de Proteína , Proteínas Virales/química , Proteínas Virales/metabolismo , Línea Celular , Citomegalovirus/inmunología , Citomegalovirus/fisiología , Infecciones por Citomegalovirus/inmunología , Retículo Endoplásmico/metabolismo , Degradación Asociada con el Retículo Endoplásmico , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Células Asesinas Naturales/inmunología , Cinética , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Unión Proteica , Dominios Proteicos , Proteínas/metabolismo , Proteolisis , Solubilidad
7.
J Bioinform Comput Biol ; 19(2): 2150006, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33866960

RESUMEN

Binding site prediction for new proteins is important in structure-based drug design. The identified binding sites may be helpful in the development of treatments for new viral outbreaks in the world when there is no information available about their pockets with COVID-19 being a case in point. Identification of the pockets using computational methods, as an alternative method, has recently attracted much interest. In this study, the binding site prediction is viewed as a semantic segmentation problem. An improved 3D version of the U-Net model based on the dice loss function is utilized to predict the binding sites accurately. The performance of the proposed model on the independent test datasets and SARS-COV-2 shows the segmentation model could predict the binding sites with a more accurate shape than the recently published deep learning model, i.e. DeepSite. Therefore, the model may help predict the binding sites of proteins and could be used in drug design for novel proteins.


Asunto(s)
Biología Computacional/métodos , Proteínas/química , Proteínas/metabolismo , /química , Algoritmos , Sitios de Unión , Bases de Datos de Proteínas , Modelos Moleculares , Proteínas Virales/química , Proteínas Virales/metabolismo
8.
Nat Commun ; 12(1): 2420, 2021 04 23.
Artículo en Inglés | MEDLINE | ID: mdl-33893297

RESUMEN

Bacteriophages have long been known to use modified bases in their DNA to prevent cleavage by the host's restriction endonucleases. Among them, cyanophage S-2L is unique because its genome has all its adenines (A) systematically replaced by 2-aminoadenines (Z). Here, we identify a member of the PrimPol family as the sole possible polymerase of S-2L and we find it can incorporate both A and Z in front of a T. Its crystal structure at 1.5 Å resolution confirms that there is no structural element in the active site that could lead to the rejection of A in front of T. To resolve this contradiction, we show that a nearby gene is a triphosphohydolase specific of dATP (DatZ), that leaves intact all other dNTPs, including dZTP. This explains the absence of A in S-2L genome. Crystal structures of DatZ with various ligands, including one at sub-angstrom resolution, allow to describe its mechanism as a typical two-metal-ion mechanism and to set the stage for its engineering.


Asunto(s)
2-Aminopurina/análogos & derivados , Adenina/química , Bacteriófagos/genética , Cianobacterias/virología , ADN Viral/química , Synechococcus/virología , 2-Aminopurina/química , 2-Aminopurina/metabolismo , Adenina/metabolismo , Bacteriófagos/metabolismo , Sitios de Unión/genética , Biocatálisis , ADN Primasa/química , ADN Primasa/genética , ADN Primasa/metabolismo , ADN Viral/genética , ADN Viral/metabolismo , ADN Polimerasa Dirigida por ADN/química , ADN Polimerasa Dirigida por ADN/genética , ADN Polimerasa Dirigida por ADN/metabolismo , Enlace de Hidrógeno , Modelos Moleculares , Estructura Molecular , Dominios Proteicos , Proteínas Virales/química , Proteínas Virales/genética , Proteínas Virales/metabolismo
9.
Biomolecules ; 11(5)2021 04 27.
Artículo en Inglés | MEDLINE | ID: mdl-33925394

RESUMEN

The SARS-CoV-2 outbreak was declared a worldwide pandemic in 2020. Infection triggers the respiratory tract disease COVID-19, which is accompanied by serious changes in clinical biomarkers such as hemoglobin and interleukins. The same parameters are altered during hemolysis, which is characterized by an increase in labile heme. We present two computational-experimental approaches aimed at analyzing a potential link between heme-related and COVID-19 pathophysiologies. Herein, we performed a detailed analysis of the common pathways induced by heme and SARS-CoV-2 by superimposition of knowledge graphs covering heme biology and COVID-19 pathophysiology. Focus was laid on inflammatory pathways and distinct biomarkers as the linking elements. In a second approach, four COVID-19-related proteins, the host cell proteins ACE2 and TMPRSS2 as well as the viral proteins 7a and S protein were computationally analyzed as potential heme-binding proteins with an experimental validation. The results contribute to the understanding of the progression of COVID-19 infections in patients with different clinical backgrounds and may allow for a more individual diagnosis and therapy in the future.


Asunto(s)
/metabolismo , Hemo/metabolismo , /fisiología , /metabolismo , Biología Computacional , Hemólisis , Interacciones Huésped-Patógeno , Humanos , Inflamación/metabolismo , Inflamación/patología , Modelos Biológicos , Modelos Moleculares , Unión Proteica , Mapas de Interacción de Proteínas , Serina Endopeptidasas/metabolismo , Proteínas Virales/metabolismo
10.
mBio ; 12(2)2021 04 13.
Artículo en Inglés | MEDLINE | ID: mdl-33849972

RESUMEN

RNA viruses that replicate in the cytoplasm often disrupt nucleocytoplasmic transport to preferentially translate their own transcripts and prevent host antiviral responses. The Sarbecovirus accessory protein ORF6 has previously been shown to be a major inhibitor of interferon production in both severe acute respiratory syndrome coronavirus (SARS-CoV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we show SARS-CoV-2-infected cells display an elevated level of nuclear mRNA accumulation compared to mock-infected cells. We demonstrate that ORF6 is responsible for this nuclear imprisonment of host mRNA, and using a cotransfected reporter assay, we show this nuclear retention of mRNA blocks expression of newly transcribed mRNAs. ORF6's nuclear entrapment of host mRNA is associated with its ability to copurify with the mRNA export factors, Rae1 and Nup98. These protein-protein interactions map to the C terminus of ORF6 and can be abolished by a single amino acid mutation in Met58. Overexpression of Rae1 restores reporter expression in the presence of SARS-CoV-2 ORF6. SARS-CoV ORF6 also interacts with Rae1 and Nup98. However, SARS-CoV-2 ORF6 more strongly copurifies with Rae1 and Nup98 and results in significantly reduced expression of reporter proteins compared to SARS-CoV ORF6, a potential mechanism for the delayed symptom onset and presymptomatic transmission uniquely associated with the SARS-CoV-2 pandemic. We also show that both SARS-CoV and SARS-CoV-2 ORF6 block nuclear import of a broad range of host proteins. Together, these data support a model in which ORF6 clogs the nuclear pore through its interactions with Rae1 and Nup98 to prevent both nuclear import and export, rendering host cells incapable of responding to SARS-CoV-2 infection.IMPORTANCE SARS-CoV-2, the causative agent of coronavirus disease 2019 (COVID-19), is an RNA virus with a large genome that encodes multiple accessory proteins. While these accessory proteins are not required for growth in vitro, they can contribute to the pathogenicity of the virus. We demonstrate that SARS-CoV-2-infected cells accumulate poly(A) mRNA in the nucleus, which is attributed to the accessory protein ORF6. Nuclear entrapment of mRNA and reduced expression of newly transcribed reporter proteins are associated with ORF6's interactions with the mRNA export proteins Rae1 and Nup98. SARS-CoV ORF6 also shows the same interactions with Rae1 and Nup98. However, SARS-CoV-2 ORF6 more strongly represses reporter expression and copurifies with Rae1 and Nup98 compared to SARS-CoV ORF6. Both SARS-CoV ORF6 and SARS-CoV-2 ORF6 block nuclear import of a wide range of host factors through interactions with Rae1 and Nup98. Together, our results suggest ORF6's disruption of nucleocytoplasmic transport prevents infected cells from responding to the invading virus.


Asunto(s)
Núcleo Celular/metabolismo , Proteínas Asociadas a Matriz Nuclear/metabolismo , Proteínas de Complejo Poro Nuclear/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Proteínas Virales/metabolismo , Transporte Activo de Núcleo Celular , Sitios de Unión , /virología , Línea Celular , Regulación de la Expresión Génica , Humanos , Mutación , Proteínas Asociadas a Matriz Nuclear/genética , Proteínas de Complejo Poro Nuclear/genética , Proteínas de Transporte Nucleocitoplasmático/genética , Unión Proteica , ARN Mensajero/metabolismo , Proteínas Virales/química , Proteínas Virales/genética
11.
Nat Commun ; 12(1): 2149, 2021 04 12.
Artículo en Inglés | MEDLINE | ID: mdl-33846319

RESUMEN

Reovirus infection requires the concerted action of viral and host factors to promote cell entry. After interaction of reovirus attachment protein σ1 with cell-surface carbohydrates and proteinaceous receptors, additional host factors mediate virus internalization. In particular, ß1 integrin is required for endocytosis of reovirus virions following junctional adhesion molecule A (JAM-A) binding. While integrin-binding motifs in the surface-exposed region of reovirus capsid protein λ2 are thought to mediate integrin interaction, evidence for direct ß1 integrin-reovirus interactions and knowledge of how integrins function to mediate reovirus entry is lacking. Here, we use single-virus force spectroscopy and confocal microscopy to discover a direct interaction between reovirus and ß1 integrins. Comparison of interactions between reovirus disassembly intermediates as well as mutants and ß1 integrin show that λ2 is the integrin ligand. Finally, using fluidic force microscopy, we demonstrate a functional role for ß1 integrin interaction in promoting clathrin recruitment to cell-bound reovirus. Our study demonstrates a direct interaction between reovirus and ß1 integrins and offers insights into the mechanism of reovirus cell entry. These results provide new perspectives for the development of efficacious antiviral therapeutics and the engineering of improved viral gene delivery and oncolytic vectors.


Asunto(s)
Clatrina/metabolismo , Interacciones Huésped-Patógeno , Integrina beta1/metabolismo , Reoviridae/fisiología , Animales , Sitios de Unión , Cápside/metabolismo , Cationes , Línea Celular , Membrana Celular/metabolismo , Endocitosis , Cinética , Ratones , Ácido N-Acetilneuramínico/metabolismo , Mutación Puntual/genética , Unión Proteica , Termodinámica , Proteínas Virales/metabolismo , Virión/metabolismo
12.
Proc Natl Acad Sci U S A ; 118(19)2021 05 11.
Artículo en Inglés | MEDLINE | ID: mdl-33906951

RESUMEN

The COVID-19 pandemic has highlighted the need to quickly and reliably prioritize clinically approved compounds for their potential effectiveness for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections. Here, we deployed algorithms relying on artificial intelligence, network diffusion, and network proximity, tasking each of them to rank 6,340 drugs for their expected efficacy against SARS-CoV-2. To test the predictions, we used as ground truth 918 drugs experimentally screened in VeroE6 cells, as well as the list of drugs in clinical trials that capture the medical community's assessment of drugs with potential COVID-19 efficacy. We find that no single predictive algorithm offers consistently reliable outcomes across all datasets and metrics. This outcome prompted us to develop a multimodal technology that fuses the predictions of all algorithms, finding that a consensus among the different predictive methods consistently exceeds the performance of the best individual pipelines. We screened in human cells the top-ranked drugs, obtaining a 62% success rate, in contrast to the 0.8% hit rate of nonguided screenings. Of the six drugs that reduced viral infection, four could be directly repurposed to treat COVID-19, proposing novel treatments for COVID-19. We also found that 76 of the 77 drugs that successfully reduced viral infection do not bind the proteins targeted by SARS-CoV-2, indicating that these network drugs rely on network-based mechanisms that cannot be identified using docking-based strategies. These advances offer a methodological pathway to identify repurposable drugs for future pathogens and neglected diseases underserved by the costs and extended timeline of de novo drug development.


Asunto(s)
/tratamiento farmacológico , Reposicionamiento de Medicamentos/métodos , Biología de Sistemas/métodos , Animales , Antivirales/administración & dosificación , Antivirales/farmacología , Antivirales/uso terapéutico , Chlorocebus aethiops , Bases de Datos Farmacéuticas , Humanos , Redes Neurales de la Computación , Unión Proteica , Células Vero , Proteínas Virales/metabolismo
13.
FASEB J ; 35(5): e21573, 2021 05.
Artículo en Inglés | MEDLINE | ID: mdl-33913206

RESUMEN

Coronavirus (CoV) 3-chymotrypsin (C)-like cysteine protease (3CLpro ) is a target for anti-CoV drug development and drug repurposing because along with papain-like protease, it cleaves CoV-encoded polyproteins (pp1a and pp1ab) into nonstructural proteins (nsps) for viral replication. However, the cleavage sites of 3CLpro and their relevant nsps remain unclear, which is the subject of this perspective. Here, we address the subject from three standpoints. First, we explore the inconsistency in the cleavage sites and relevant nsps across CoVs, and investigate the function of nsp11. Second, we consider the nsp16 mRNA overlapping of the spike protein mRNA, and analyze the effect of this overlapping on mRNA vaccines. Finally, we study nsp12, whose existence depends on ribosomal frameshifting, and investigate whether 3CLpro requires a large number of inhibitors to achieve full inhibition. This perspective helps us to clarify viral replication and is useful for developing anti-CoV drugs with 3CLpro as a target in the current coronavirus disease 2019 (COVID-19) pandemic.


Asunto(s)
/metabolismo , Proteínas Virales/metabolismo , Antivirales/farmacología , Desarrollo de Medicamentos/métodos , Poliproteínas/química , Poliproteínas/genética , Poliproteínas/metabolismo , Inhibidores de Proteasas/farmacología , Vacunas Sintéticas/metabolismo , Proteínas Virales/química , Proteínas Virales/genética
14.
PLoS Comput Biol ; 17(3): e1008805, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33730015

RESUMEN

Thrombosis is a recognized complication of Coronavirus disease of 2019 (COVID-19) and is often associated with poor prognosis. There is a well-recognized link between coagulation and inflammation, however, the extent of thrombotic events associated with COVID-19 warrants further investigation. Poly(A) Binding Protein Cytoplasmic 4 (PABPC4), Serine/Cysteine Proteinase Inhibitor Clade G Member 1 (SERPING1) and Vitamin K epOxide Reductase Complex subunit 1 (VKORC1), which are all proteins linked to coagulation, have been shown to interact with SARS proteins. We computationally examined the interaction of these with SARS-CoV-2 proteins and, in the case of VKORC1, we describe its binding to ORF7a in detail. We examined the occurrence of variants of each of these proteins across populations and interrogated their potential contribution to COVID-19 severity. Potential mechanisms, by which some of these variants may contribute to disease, are proposed. Some of these variants are prevalent in minority groups that are disproportionally affected by severe COVID-19. Therefore, we are proposing that further investigation around these variants may lead to better understanding of disease pathogenesis in minority groups and more informed therapeutic approaches.


Asunto(s)
Coagulación Sanguínea , Proteínas Sanguíneas/genética , Proteína Inhibidora del Complemento C1/genética , Proteínas de Unión a Poli(A)/genética , Vitamina K Epóxido Reductasas/genética , Anticoagulantes/administración & dosificación , Proteínas Sanguíneas/metabolismo , /virología , Proteína Inhibidora del Complemento C1/metabolismo , Estudio de Asociación del Genoma Completo , Humanos , Modelos Moleculares , Mutación , Proteínas de Unión a Poli(A)/metabolismo , Unión Proteica , Índice de Severidad de la Enfermedad , Proteínas Virales/metabolismo , Vitamina K Epóxido Reductasas/metabolismo , Warfarina/administración & dosificación
15.
Gene ; 784: 145596, 2021 Jun 05.
Artículo en Inglés | MEDLINE | ID: mdl-33766711

RESUMEN

The SARS-CoV-2 Variant of Concern 202012/01 (VOC-202012/01) is rapidly spreading worldwide owing to its substantial transmission advantage. The variant has changes in critical sites of the spike protein with potential biological significance. Moreover, VOC-202012/01 has a mutation that inactivates the ORF8 protein, whose absence can change the clinical features of the infection. Why VOC-202012/01 is more transmissible remains unclear, but spike mutations and ORF8 inactivation stand out by their known phenotypic effects. Here I show that variants combining relevant spike mutations and the absence of ORF8 occurred in SARS-CoV-2 and related viruses circulating in other host species. A truncated ORF8 (Q23stop) occurred in a SARS-CoV-2-related virus from a pangolin seized in China in 2017, also with several mutations in critical spike sites. Strikingly, I found that variants without ORF8 (E19stop) and with the N501T spike mutation circulated in farmed mink and humans from Denmark. Although with differences to VOC-202012/01, the identification of these variants highlights the danger of having reservoirs of SARS-CoV-2 and related viruses where more transmissible variants may occur and spill over to humans.


Asunto(s)
/veterinaria , Visón/virología , /genética , Glicoproteína de la Espiga del Coronavirus/genética , Proteínas Virales/genética , Animales , /virología , China , Codón sin Sentido , Dinamarca , Reservorios de Enfermedades/veterinaria , Reservorios de Enfermedades/virología , Especificidad del Huésped , Humanos , Proteínas Virales/metabolismo
16.
Cell Rep ; 34(13): 108916, 2021 03 30.
Artículo en Inglés | MEDLINE | ID: mdl-33765414

RESUMEN

The presence of an ORF6 gene distinguishes sarbecoviruses such as severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2 from other betacoronaviruses. Here we show that ORF6 inhibits induction of innate immune signaling, including upregulation of type I interferon (IFN) upon viral infection as well as type I and III IFN signaling. Intriguingly, ORF6 proteins from SARS-CoV-2 lineages are more efficient antagonists of innate immunity than their orthologs from SARS-CoV lineages. Mutational analyses identified residues E46 and Q56 as important determinants of the antagonistic activity of SARS-CoV-2 ORF6. Moreover, we show that the anti-innate immune activity of ORF6 depends on its C-terminal region and that ORF6 inhibits nuclear translocation of IRF3. Finally, we identify naturally occurring frameshift/nonsense mutations that result in an inactivating truncation of ORF6 in approximately 0.2% of SARS-CoV-2 isolates. Our findings suggest that ORF6 contributes to the poor IFN activation observed in individuals with coronavirus disease 2019 (COVID-19).


Asunto(s)
/metabolismo , Interferón Tipo I/metabolismo , Proteínas Virales/metabolismo , Animales , Chlorocebus aethiops , Células HEK293 , Humanos , Inmunidad Innata/inmunología , Transducción de Señal/inmunología , Células Vero , Proteínas Virales/genética
17.
Viruses ; 13(3)2021 02 26.
Artículo en Inglés | MEDLINE | ID: mdl-33652634

RESUMEN

Ubiquitination of proteins is a post-translational modification process with many different cellular functions, including protein stability, immune signaling, antiviral functions and virus replication. While ubiquitination of viral proteins can be used by the host as a defense mechanism by destroying the incoming pathogen, viruses have adapted to take advantage of this cellular process. The ubiquitin system can be hijacked by viruses to enhance various steps of the replication cycle and increase pathogenesis. Emerging viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), flaviviruses like Zika and dengue, as well as highly pathogenic viruses like Ebola and Nipah, have the ability to directly use the ubiquitination process to enhance their viral-replication cycle, and evade immune responses. Some of these mechanisms are conserved among different virus families, especially early during virus entry, providing an opportunity to develop broad-spectrum antivirals. Here, we discuss the mechanisms used by emergent viruses to exploit the host ubiquitin system, with the main focus on the role of ubiquitin in enhancing virus replication.


Asunto(s)
Ubiquitina/metabolismo , Virosis/metabolismo , Replicación Viral , Virus/metabolismo , Evasión Inmune , Ubiquitinación , Proteínas Virales/metabolismo , Ensamble de Virus , Virosis/inmunología , Virosis/virología , Internalización del Virus , Liberación del Virus , Virus/clasificación , Virus/inmunología , Virus/patogenicidad
18.
Molecules ; 26(4)2021 Feb 14.
Artículo en Inglés | MEDLINE | ID: mdl-33673017

RESUMEN

Influenza A virus (IAV) encodes a polymerase composed of three subunits: PA, with endonuclease activity, PB1 with polymerase activity and PB2 with host RNA five-prime cap binding site. Their cooperation and stepwise activation include a process called cap-snatching, which is a crucial step in the IAV life cycle. Reproduction of IAV can be blocked by disrupting the interaction between the PB2 domain and the five-prime cap. An inhibitor of this interaction called pimodivir (VX-787) recently entered the third phase of clinical trial; however, several mutations in PB2 that cause resistance to pimodivir were observed. First major mutation, F404Y, causing resistance was identified during preclinical testing, next the mutation M431I was identified in patients during the second phase of clinical trials. The mutation H357N was identified during testing of IAV strains at Centers for Disease Control and Prevention. We set out to provide a structural and thermodynamic analysis of the interactions between cap-binding domain of PB2 wild-type and PB2 variants bearing these mutations and pimodivir. Here we present four crystal structures of PB2-WT, PB2-F404Y, PB2-M431I and PB2-H357N in complex with pimodivir. We have thermodynamically analysed all PB2 variants and proposed the effect of these mutations on thermodynamic parameters of these interactions and pimodivir resistance development. These data will contribute to understanding the effect of these missense mutations to the resistance development and help to design next generation inhibitors.


Asunto(s)
Farmacorresistencia Viral/efectos de los fármacos , Virus de la Influenza A/enzimología , Subunidades de Proteína/antagonistas & inhibidores , Piridinas/química , Piridinas/farmacología , Pirimidinas/química , Pirimidinas/farmacología , Pirroles/química , Pirroles/farmacología , Proteínas Virales/antagonistas & inhibidores , Cristalografía por Rayos X , Virus de la Influenza A/efectos de los fármacos , Modelos Moleculares , Proteínas Mutantes/metabolismo , Mutación/genética , Dominios Proteicos , Subunidades de Proteína/química , Subunidades de Proteína/metabolismo , Teoría Cuántica , /química , Termodinámica , Proteínas Virales/química , Proteínas Virales/metabolismo
19.
Mikrochim Acta ; 188(4): 137, 2021 03 25.
Artículo en Inglés | MEDLINE | ID: mdl-33763734

RESUMEN

The novel corona (SARS-CoV-2) virus causes a global pandemic, which motivates researchers to develop reliable and effective methods for screening and detection of SARS-CoV-2. Though there are several methods available for the diagnosis of SARS-CoV-2 such as RT-PCR and ELSIA, nevertheless, these methods are time-consuming and may not apply at the point of care. In this study, we have developed a specific, sensitive, quantitative and fast detection method for SARS-CoV-2 by fluorescence resonance energy transfer (FRET) assay. The total extracellular protease proteolytic activity from the virus has been used as the biomarker. The specific peptide sequences from the library of 115 dipeptides were identified via changes in the fluorescence signal. The fluorogenic dipeptide substrates have the fluorophore and a quencher at the N- and the C- terminals, respectively. When the protease hydrolyzes the peptide bond between the two specific amino acids, it leads to a significant increase in the fluorescence signals. The specific fluorogenic peptide (H-d) produces a high fluorescence signal. A calibration plot was obtained from the changes in the fluorescence intensity against the different concentrations of the viral protease. The lowest limit of detection of this method was 9.7 ± 3 pfu/mL. The cross-reactivity of the SARS-CoV-2-specific peptide was tested against the MERS-CoV which does not affect the fluorescence signal. A significant change in the fluorescence signal with patient samples indicates that this FRET-based assay might be applied for the diagnosis of SARS-CoV-2 patients. Graphical abstract.


Asunto(s)
/métodos , /metabolismo , Colorantes Fluorescentes/metabolismo , Péptidos/metabolismo , Proteínas Virales/metabolismo , Animales , Bioensayo , Chlorocebus aethiops , Transferencia Resonante de Energía de Fluorescencia , Humanos , Biblioteca de Péptidos , /metabolismo , Células Vero , Ensayo de Placa Viral
20.
Aging (Albany NY) ; 13(7): 9160-9185, 2021 03 19.
Artículo en Inglés | MEDLINE | ID: mdl-33744846

RESUMEN

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this study, we collected open access data to analyze the mechanisms associated with SARS-CoV-2 infection. Gene set enrichment analysis (GSEA) revealed that apoptosis-related pathways were enriched in the cells after SARS-CoV-2 infection, and the results of differential expression analysis showed that biological functions related to endoplasmic reticulum stress (ERS) and lipid metabolism were disordered. TMBIM6 was identified as a potential target for SARS-CoV-2 in host cells through weighted gene coexpression network analysis (WGCNA) of the time course of expression of host and viral proteins. The expression and related functions of TMBIM6 were subsequently analyzed to illuminate how viral proteins interfere with the physiological function of host cells. The potential function of viral proteins was further analyzed by GEne Network Inference with Ensemble of trees (GENIE3). This study identified TMBIM6 as a target protein associated with the pathogenesis of SARS-CoV-2, which might provide a novel therapeutic approach for COVID-19 in the future.


Asunto(s)
Proteínas Reguladoras de la Apoptosis/metabolismo , Interacciones Huésped-Patógeno , Proteínas de la Membrana/metabolismo , Proteínas Virales/metabolismo , Células A549 , Proteínas Reguladoras de la Apoptosis/genética , Células CACO-2 , Redes Reguladoras de Genes , Genómica , Humanos , Proteínas de la Membrana/genética , Mapas de Interacción de Proteínas , Proteínas Virales/genética
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA
...