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1.
PLoS Pathog ; 16(9): e1008825, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32886709

RESUMO

Most alphaviruses (family Togaviridae) including Sindbis virus (SINV) and other human pathogens, are transmitted by arthropods. The first open reading frame in their positive strand RNA genome encodes for the non-structural polyprotein, a precursor to four separate subunits of the replicase. The replicase interacts with cis-acting elements located near the intergenic region and at the ends of the viral RNA genome. A trans-replication assay was developed and used to analyse the template requirements for nine alphavirus replicases. Replicases of alphaviruses of the Semliki Forest virus complex were able to cross-utilize each other's templates as well as those of outgroup alphaviruses. Templates of outgroup alphaviruses, including SINV and the mosquito-specific Eilat virus, were promiscuous; in contrast, their replicases displayed a limited capacity to use heterologous templates, especially in mosquito cells. The determinants important for efficient replication of template RNA were mapped to the 5' region of the genome. For SINV these include the extreme 5'- end of the genome and sequences corresponding to the first stem-loop structure in the 5' untranslated region. Mutations introduced in these elements drastically reduced infectivity of recombinant SINV genomes. The trans-replicase tools and approaches developed here can be instrumental in studying alphavirus recombination and evolution, but can also be applied to study other viruses such as picornaviruses, flaviviruses and coronaviruses.


Assuntos
Alphavirus , Genoma Viral , Conformação de Ácido Nucleico , RNA Replicase , RNA Viral , Proteínas Virais , Alphavirus/química , Alphavirus/genética , Alphavirus/metabolismo , Linhagem Celular Tumoral , Células HEK293 , Humanos , RNA Replicase/química , RNA Replicase/genética , RNA Replicase/metabolismo , RNA Viral/química , RNA Viral/genética , RNA Viral/metabolismo , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais/metabolismo
2.
Proc Natl Acad Sci U S A ; 117(39): 24384-24391, 2020 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-32913053

RESUMO

An improved understanding of human T cell-mediated immunity in COVID-19 is important for optimizing therapeutic and vaccine strategies. Experience with influenza shows that infection primes CD8+ T cell memory to peptides presented by common HLA types like HLA-A2, which enhances recovery and diminishes clinical severity upon reinfection. Stimulating peripheral blood mononuclear cells from COVID-19 convalescent patients with overlapping peptides from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to the clonal expansion of SARS-CoV-2-specific CD8+ and CD4+ T cells in vitro, with CD4+ T cells being robust. We identified two HLA-A*02:01-restricted SARS-CoV-2-specfic CD8+ T cell epitopes, A2/S269-277 and A2/Orf1ab3183-3191 Using peptide-HLA tetramer enrichment, direct ex vivo assessment of A2/S269 +CD8+ and A2/Orf1ab3183 +CD8+ populations indicated that A2/S269 +CD8+ T cells were detected at comparable frequencies (∼1.3 × 10-5) in acute and convalescent HLA-A*02:01+ patients. These frequencies were higher than those found in uninfected HLA-A*02:01+ donors (∼2.5 × 10-6), but low when compared to frequencies for influenza-specific (A2/M158) and Epstein-Barr virus (EBV)-specific (A2/BMLF1280) (∼1.38 × 10-4) populations. Phenotyping A2/S269 +CD8+ T cells from COVID-19 convalescents ex vivo showed that A2/S269 +CD8+ T cells were predominantly negative for CD38, HLA-DR, PD-1, and CD71 activation markers, although the majority of total CD8+ T cells expressed granzymes and/or perforin. Furthermore, the bias toward naïve, stem cell memory and central memory A2/S269 +CD8+ T cells rather than effector memory populations suggests that SARS-CoV-2 infection may be compromising CD8+ T cell activation. Priming with appropriate vaccines may thus be beneficial for optimizing CD8+ T cell immunity in COVID-19.


Assuntos
Betacoronavirus/imunologia , Linfócitos T CD8-Positivos/imunologia , Infecções por Coronavirus/imunologia , Antígeno HLA-A2/imunologia , Pneumonia Viral/imunologia , Linfócitos T CD4-Positivos/imunologia , Epitopos de Linfócito T , Feminino , Humanos , Memória Imunológica , Imunofenotipagem , Leucócitos Mononucleares/imunologia , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Pandemias , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/imunologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Proteínas Virais/química , Proteínas Virais/imunologia
3.
Structure ; 28(8): 874-878, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32755569

RESUMO

During global pandemics, the spread of information needs to be faster than the spread of the virus in order to ensure the health and safety of human populations worldwide. In our current crisis, the demand for SARS-CoV-2 drugs and vaccines highlights the importance of biological targets and their three-dimensional shape. In particular, structural biology as a field was poised to quickly respond to crises due to previous experience and expertise and because of its early adoption of open access practices.


Assuntos
Betacoronavirus/química , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/virologia , Proteínas Virais/química , Cisteína Endopeptidases/química , Bases de Dados de Proteínas , Humanos , Modelos Moleculares , Biologia Molecular , Conformação Proteica , RNA Replicase/química , Glicoproteína da Espícula de Coronavírus/química , Proteínas não Estruturais Virais/química
4.
Sci Rep ; 10(1): 14179, 2020 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-32843695

RESUMO

A novel coronavirus (SARS-CoV-2) emerged from China in late 2019 and rapidly spread across the globe, infecting millions of people and generating societal disruption on a level not seen since the 1918 influenza pandemic. A safe and effective vaccine is desperately needed to prevent the continued spread of SARS-CoV-2; yet, rational vaccine design efforts are currently hampered by the lack of knowledge regarding viral epitopes targeted during an immune response, and the need for more in-depth knowledge on betacoronavirus immunology. To that end, we developed a computational workflow using a series of open-source algorithms and webtools to analyze the proteome of SARS-CoV-2 and identify putative T cell and B cell epitopes. Utilizing a set of stringent selection criteria to filter peptide epitopes, we identified 41 T cell epitopes (5 HLA class I, 36 HLA class II) and 6 B cell epitopes that could serve as promising targets for peptide-based vaccine development against this emerging global pathogen. To our knowledge, this is the first study to comprehensively analyze all 10 (structural, non-structural and accessory) proteins from SARS-CoV-2 using predictive algorithms to identify potential targets for vaccine development.


Assuntos
Betacoronavirus/imunologia , Biologia Computacional , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito T/imunologia , Interações Hospedeiro-Patógeno/imunologia , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Proteínas Virais/imunologia , Sequência de Aminoácidos , Linfócitos B/imunologia , Linfócitos B/metabolismo , Betacoronavirus/classificação , Betacoronavirus/genética , Betacoronavirus/metabolismo , Biologia Computacional/métodos , Infecções por Coronavirus/metabolismo , Epitopos de Linfócito B/química , Epitopos de Linfócito T/química , Genoma Viral , Genômica/métodos , Humanos , Modelos Moleculares , Pandemias , Peptídeos/química , Peptídeos/imunologia , Filogenia , Pneumonia Viral/metabolismo , Relação Estrutura-Atividade , Linfócitos T/imunologia , Linfócitos T/metabolismo , Vacinas de Subunidades/imunologia , Proteínas Virais/química , Vacinas Virais/imunologia
5.
J Enzyme Inhib Med Chem ; 35(1): 1539-1544, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32746637

RESUMO

Coronavirus disease 2019 (COVID-19) has been a pandemic disease of which the termination is not yet predictable. Currently, researches to develop vaccines and treatments is going on globally to cope with this disastrous disease. Main protease (3CLpro) from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is one of the good targets to find antiviral agents before vaccines are available. Some flavonoids are known to inhibit 3CLpro from SARS-CoV which causes SARS. Since their sequence identity is 96%, a similar approach was performed with a flavonoid library. Baicalin, herbacetin, and pectolinarin have been discovered to block the proteolytic activity of SARS-CoV-2 3CLpro. An in silico docking study showed that the binding modes of herbacetin and pectolinarin are similar to those obtained from the catalytic domain of SARS-CoV 3CLpro. However, their binding affinities are different due to the usage of whole SARS-CoV-2 3CLpro in this study. Baicalin showed an effective inhibitory activity against SARS-CoV-2 3CLpro and its docking mode is different from those of herbacetin and pectolinarin. This study suggests important scaffolds to design 3CLpro inhibitors to develop antiviral agents or health-foods and dietary supplements to cope with SARS-CoV-2.


Assuntos
Infecções por Coronavirus/tratamento farmacológico , Flavonoides/química , Pneumonia Viral/tratamento farmacológico , Proteínas Virais/antagonistas & inibidores , Proteínas Virais/química , Antivirais/química , Betacoronavirus , Desenho de Fármacos , Transferência Ressonante de Energia de Fluorescência , Humanos , Simulação de Acoplamento Molecular , Pandemias , Inibidores de Proteases/química , Ligação Proteica , Conformação Proteica , Espectrofotometria , Triptofano/química
6.
Methods Mol Biol ; 2203: 187-204, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32833213

RESUMO

Biotin-based proximity labeling circumvents major pitfalls of classical biochemical approaches to identify protein-protein interactions. It consists of enzyme-catalyzed biotin tags ubiquitously apposed on proteins located in close proximity of the labeling enzyme, followed by affinity purification and identification of biotinylated proteins by mass spectrometry. Here we outline the methods by which the molecular microenvironment of the coronavirus replicase/transcriptase complex (RTC), i.e., proteins located within a close perimeter of the RTC, can be determined by different proximity labeling approaches using BirAR118G (BioID), TurboID, and APEX2. These factors represent a molecular signature of coronavirus RTCs and likely contribute to the viral life cycle, thereby constituting attractive targets for the development of antiviral intervention strategies.


Assuntos
Coronavirus/patogenicidade , Enzimas/genética , Interações Hospedeiro-Patógeno/fisiologia , Proteômica/métodos , Proteínas Virais/metabolismo , Animais , Ascorbato Peroxidases/genética , Biotinilação , Carbono-Nitrogênio Ligases/genética , Linhagem Celular , Coronavirus/genética , Enzimas/metabolismo , Proteínas de Escherichia coli/genética , Imunofluorescência , Microrganismos Geneticamente Modificados , Proteínas Repressoras/genética , Proteínas Virais/química , Proteínas Virais/genética
7.
Virol J ; 17(1): 131, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32854725

RESUMO

BACKGROUND: The Covid19 infection is caused by the SARS-CoV-2 virus, a novel member of the coronavirus (CoV) family. CoV genomes code for a ORF1a / ORF1ab polyprotein and four structural proteins widely studied as major drug targets. The genomes also contain a variable number of open reading frames (ORFs) coding for accessory proteins that are not essential for virus replication, but appear to have a role in pathogenesis. The accessory proteins have been less well characterized and are difficult to predict by classical bioinformatics methods. METHODS: We propose a computational tool GOFIX to characterize potential ORFs in virus genomes. In particular, ORF coding potential is estimated by searching for enrichment in motifs of the X circular code, that is known to be over-represented in the reading frames of viral genes. RESULTS: We applied GOFIX to study the SARS-CoV-2 and related genomes including SARS-CoV and SARS-like viruses from bat, civet and pangolin hosts, focusing on the accessory proteins. Our analysis provides evidence supporting the presence of overlapping ORFs 7b, 9b and 9c in all the genomes and thus helps to resolve some differences in current genome annotations. In contrast, we predict that ORF3b is not functional in all genomes. Novel putative ORFs were also predicted, including a truncated form of the ORF10 previously identified in SARS-CoV-2 and a little known ORF overlapping the Spike protein in Civet-CoV and SARS-CoV. CONCLUSIONS: Our findings contribute to characterizing sequence properties of accessory genes of SARS coronaviruses, and especially the newly acquired genes making use of overlapping reading frames.


Assuntos
Betacoronavirus/genética , Genoma Viral , Fases de Leitura Aberta , Vírus da SARS/genética , Proteínas Virais Reguladoras e Acessórias/genética , Animais , Códon , Biologia Computacional , Evolução Molecular , Genes Virais , Humanos , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Proteínas da Matriz Viral/genética , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais Reguladoras e Acessórias/química
8.
Cell Stress Chaperones ; 25(5): 737-741, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32754823

RESUMO

Severe acute respiratory syndrome corona virus 2 (SARS-CoV-2), the cause of COVID-19 disease, has the potential to elicit autoimmunity because mimicry of human molecular chaperones by viral proteins. We compared viral proteins with human molecular chaperones, many of which are heat shock proteins, to determine if they share amino acid-sequence segments with immunogenic-antigenic potential, which can elicit cross-reactive antibodies and effector immune cells with the capacity to damage-destroy human cells by a mechanism of autoimmunity. We identified the chaperones that can putatively participate in molecular mimicry phenomena after SARS-CoV-2 infection, focusing on those for which endothelial cell plasma-cell membrane localization has already been demonstrated. We also postulate that post-translational modifications, induced by physical (shear) and chemical (metabolic) stress caused respectively by the risk factors hypertension and diabetes, might have a role in determining plasma-cell membrane localization and, in turn, autoimmune-induced endothelial damage.


Assuntos
Betacoronavirus/metabolismo , Infecções por Coronavirus/virologia , Proteínas de Choque Térmico , Pneumonia Viral/virologia , Proteínas Virais , Sequência de Aminoácidos , Autoantígenos , Autoimunidade , Bases de Dados de Proteínas , Células Endoteliais/metabolismo , Proteínas de Choque Térmico/química , Proteínas de Choque Térmico/imunologia , Humanos , Epitopos Imunodominantes , Mimetismo Molecular , Pandemias , Proteínas Virais/química , Proteínas Virais/imunologia
9.
J Phys Chem Lett ; 11(17): 7021-7027, 2020 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-32787330

RESUMO

The novel coronavirus (2019-nCoV) spike protein is a smart molecular machine that instigates the entry of coronavirus to the host cell causing the COVID-19 pandemic. In this study, a symmetry-information-loaded structure-based Hamiltonian is developed using recent Cryo-EM structural data to explore the complete conformational energy landscape of the full-length prefusion spike protein. The study finds the 2019-nCoV prefusion spike to adopt a unique strategy by undertaking a dynamic conformational asymmetry that results in two prevalent asymmetric structures of spike where one or two spike heads rotate up to provide better exposure to the host-cell receptor. A few unique interchain interactions are identified at the interface of closely associated N-terminal domain (NTD) and receptor binding domain (RBD) playing a crucial role in the thermodynamic stabilization of the up conformation of the RBD in the case of the 2019-nCoV spike. The interaction-level information decoded in this study may provide deep insight into developing effective therapeutic targets.


Assuntos
Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Proteínas Virais/química , Proteínas Virais/metabolismo , Modelos Moleculares , Domínios Proteicos
10.
Arch Virol ; 165(10): 2177-2191, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32748179

RESUMO

The canonical frameworks of viral evolution describe viruses as cellular predecessors, reduced forms of cells, or entities that escaped cellular control. The discovery of giant viruses has changed these standard paradigms. Their genetic, proteomic and structural complexities resemble those of cells, prompting a redefinition and reclassification of viruses. In a previous genome-wide analysis of the evolution of structural domains in proteomes, with domains defined at the fold superfamily level, we found the origins of viruses intertwined with those of ancient cells. Here, we extend these data-driven analyses to the study of fold families confirming the co-evolution of viruses and ancient cells and the genetic ability of viruses to foster molecular innovation. The results support our suggestion that viruses arose by genomic reduction from ancient cells and validate a co-evolutionary 'symbiogenic' model of viral origins.


Assuntos
Evolução Biológica , DNA Viral/genética , Genoma Viral , Vírus Gigantes/genética , Filogenia , Proteínas Virais/genética , Archaea/genética , Archaea/virologia , Bactérias/genética , Bactérias/virologia , DNA Viral/química , Eucariotos/genética , Eucariotos/virologia , Tamanho do Genoma , Vírus Gigantes/classificação , Proteogenômica/métodos , Proteoma/genética , Proteínas Virais/química
11.
Comput Biol Med ; 124: 103936, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32738628

RESUMO

Virtual screening of phytochemicals was performed through molecular docking, simulations, in silico ADMET and drug-likeness prediction to identify the potential hits that can inhibit the effects of SARS-CoV-2. Considering the published literature on medicinal importance, 154 phytochemicals with analogous structure from limonoids and triterpenoids were selected to search potential inhibitors for the five therapeutic protein targets of SARS-CoV-2, i.e., 3CLpro (main protease), PLpro (papain-like protease), SGp-RBD (spike glycoprotein-receptor binding domain), RdRp (RNA dependent RNA polymerase) and ACE2 (angiotensin-converting enzyme 2). The in silico computational results revealed that the phytochemicals such as glycyrrhizic acid, limonin, 7-deacetyl-7-benzoylgedunin, maslinic acid, corosolic acid, obacunone and ursolic acid were found to be effective against the target proteins of SARS-CoV-2. The protein-ligand interaction study revealed that these phytochemicals bind with the amino acid residues at the active site of the target proteins. Therefore, the core structure of these potential hits can be used for further lead optimization to design drugs for SARS-CoV-2. Also, the medicinal plants containing these phytochemicals like licorice, neem, tulsi, citrus and olives can be used to formulate suitable therapeutic approaches in traditional medicines.


Assuntos
Antivirais/farmacologia , Betacoronavirus/química , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Limoninas/farmacologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Triterpenos/farmacologia , Antivirais/química , Antivirais/farmacocinética , Sítios de Ligação , Biologia Computacional , Simulação por Computador , Avaliação Pré-Clínica de Medicamentos , Interações entre Hospedeiro e Microrganismos/efeitos dos fármacos , Humanos , Limoninas/química , Limoninas/farmacocinética , Simulação de Acoplamento Molecular , Pandemias , Peptidil Dipeptidase A/química , Peptidil Dipeptidase A/efeitos dos fármacos , Compostos Fitoquímicos/química , Compostos Fitoquímicos/farmacocinética , Compostos Fitoquímicos/farmacologia , RNA Replicase/química , RNA Replicase/efeitos dos fármacos , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/efeitos dos fármacos , Triterpenos/química , Triterpenos/farmacocinética , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/efeitos dos fármacos , Proteínas Virais/química , Proteínas Virais/efeitos dos fármacos
12.
Virology ; 548: 236-249, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32791352

RESUMO

Species-specific guinea pig cytomegalovirus (GPCMV) causes congenital CMV and the virus encodes homolog glycoprotein complexes to human CMV, including gH-based trimer (gH/gL/gO) and pentamer-complex (PC). Platelet-derived growth factor receptor alpha (gpPDGFRA), only present on fibroblast cells, was identified via CRISPR as the putative receptor for PC-independent GPCMV infection. Immunoprecipitation assays demonstrated direct interaction of gH/gL/gO with gpPDGFRA but not in absence of gO. Expression of viral gB also resulted in precipitation of gB/gH/gL/gO/gpPDGFRA complex. Cell-cell fusion assays determined that expression of gpPDGFRA and gH/gL/gO in adjacent cells enabled cell fusion, which was not enhanced by gB. N-linked gpPDGFRA glycosylation inhibition had limited effect and blocking tyrosine kinase (TK) transduction had no impact on infection. Ectopically expressed gpPDGFRA or TK-domain mutant in trophoblast or epithelial cells previously non-susceptible to GPCMV(PC-) enabled viral infection. In contrast, transient human PDGFRA expression did not complement GPCMV(PC-) infection, a potential basis for viral species specificity.


Assuntos
Betaherpesvirinae/fisiologia , Infecções por Herpesviridae/veterinária , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Doenças dos Roedores/metabolismo , Proteínas Virais/metabolismo , Animais , Betaherpesvirinae/genética , Fusão Celular , Cobaias , Infecções por Herpesviridae/genética , Infecções por Herpesviridae/metabolismo , Ligação Proteica , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Doenças dos Roedores/genética , Doenças dos Roedores/virologia , Proteínas Virais/química , Proteínas Virais/genética , Internalização do Vírus
13.
Virology ; 548: 136-151, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32838935

RESUMO

Bovine herpesvirus envelope glycoprotein E (gE) and, in particular, the gE cytoplasmic tail (CT) is a virulence determinant in cattle. Also, the gE CT contributes to virus cell-to-cell spread and anterograde neuronal transport. In this study, our goal was to map the gE CT sub-domains that contribute to virus cell-to-cell spread property. A panel of gE-CT specific mutant viruses was constructed and characterized, in vitro, with respect to their plaque phenotypes, gE recycling and gE basolateral membrane targeting. The results revealed that disruption of the tyrosine-based motifs, 467YTSL470 and 563YTVV566, individually produced smaller plaque phenotypes than the wild type. However, they were slightly larger than the gE CT-null virus plaques. The Y467A mutation affected the gE endocytosis, gE trans-Golgi network (TGN) recycling, and gE virion incorporation properties. However, the Y563A mutation affected only the gE basolateral cell-surface redistribution function. Notably, the simultaneous Y467A/Y563A mutations produced gE CT-null virus-like plaque phenotypes.


Assuntos
Doenças dos Bovinos/virologia , Citoplasma/virologia , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 1/metabolismo , Proteínas Virais/química , Proteínas Virais/metabolismo , Motivos de Aminoácidos , Animais , Bovinos , Endocitose , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 1/genética , Proteínas Virais/genética , Rede trans-Golgi/virologia
14.
Biomolecules ; 10(8)2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32752270

RESUMO

Posttranslational modifications of cellular proteins by covalent conjugation of ubiquitin and ubiquitin-like polypeptides regulate numerous cellular processes that are captured by viruses to promote infection, replication, and spreading. The importance of these protein modifications for the viral life cycle is underscored by the discovery that many viruses encode deconjugases that reverse their functions. The structural and functional characterization of these viral enzymes and the identification of their viral and cellular substrates is providing valuable insights into the biology of viral infections and the host's antiviral defense. Given the growing body of evidence demonstrating their key contribution to pathogenesis, the viral deconjugases are now recognized as attractive targets for the design of novel antiviral therapeutics.


Assuntos
Antivirais/farmacologia , Enzimas/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Ubiquitina/metabolismo , Proteínas Virais/metabolismo , Viroses/metabolismo , Adenoviridae/enzimologia , Coronavirus/enzimologia , Enzimas/química , Herpesviridae/enzimologia , Humanos , Processamento de Proteína Pós-Traducional , Proteínas Virais/química , Viroses/tratamento farmacológico
15.
Int J Mol Sci ; 21(14)2020 Jul 21.
Artigo em Inglês | MEDLINE | ID: mdl-32708196

RESUMO

(1) Background: Virtual screening studies on the therapeutically relevant proteins of the severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) require a detailed characterization of their druggable binding sites, and, more generally, a convenient pocket mapping represents a key step for structure-based in silico studies; (2) Methods: Along with a careful literature search on SARS-CoV-2 protein targets, the study presents a novel strategy for pocket mapping based on the combination of pocket (as performed by the well-known FPocket tool) and docking searches (as performed by PLANTS or AutoDock/Vina engines); such an approach is implemented by the Pockets 2.0 plug-in for the VEGA ZZ suite of programs; (3) Results: The literature analysis allowed the identification of 16 promising binding cavities within the SARS-CoV-2 proteins and the here proposed approach was able to recognize them showing performances clearly better than those reached by the sole pocket detection; and (4) Conclusions: Even though the presented strategy should require more extended validations, this proved successful in precisely characterizing a set of SARS-CoV-2 druggable binding pockets including both orthosteric and allosteric sites, which are clearly amenable for virtual screening campaigns and drug repurposing studies. All results generated by the study and the Pockets 2.0 plug-in are available for download.


Assuntos
Antivirais/química , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Proteínas Virais/química , Sítios de Ligação/efeitos dos fármacos , Reposicionamento de Medicamentos , Humanos , Simulação de Acoplamento Molecular , Pandemias , Ligação Proteica/efeitos dos fármacos , Conformação Proteica
16.
Arch Virol ; 165(9): 2123-2126, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32617763

RESUMO

In this study, we report the complete genome sequence of a novel luteovirus detected in almond using high-throughput sequencing. The genome of the new luteovirus comprises 5,047 nucleotides, and its genomic organization is similar to that of the recently described nectarine stem pitting associated virus (NSPaV), with only four open reading frames, encoding replication-related proteins, the coat protein (CP), and a CP readthrough protein involved in the aphid transmission of luteovirids. Phylogenic and pairwise distance analyses showed that this virus shares 79% and 57.8% amino acid identity in the P1-P2 fusion protein and the P3-P5 protein, respectively, with the most closely related luteovirus, NSPaV, suggesting that it represents a novel species, for which the name "Almond associated luteovirus 1" is proposed. To our knowledge, this is the first report of an almond-infecting luteovirus.


Assuntos
Genoma Viral , Luteovirus/genética , Doenças das Plantas/virologia , Prunus dulcis/virologia , Sequência de Aminoácidos , Sequência de Bases , Luteovirus/classificação , Luteovirus/isolamento & purificação , Fases de Leitura Aberta , Filogenia , Alinhamento de Sequência , Proteínas Virais/química , Proteínas Virais/genética , Sequenciamento Completo do Genoma
17.
Int J Mol Sci ; 21(13)2020 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-32645951

RESUMO

The SARS-CoV-2 virus is a recently-emerged zoonotic pathogen already well adapted to transmission and replication in humans. Although the mutation rate is limited, recently introduced mutations in SARS-CoV-2 have the potential to alter viral fitness. In addition to amino acid changes, mutations could affect RNA secondary structure critical to viral life cycle, or interfere with sequences targeted by host miRNAs. We have analysed subsets of genomes from SARS-CoV-2 isolates from around the globe and show that several mutations introduce changes in Watson-Crick pairing, with resultant changes in predicted secondary structure. Filtering to targets matching miRNAs expressed in SARS-CoV-2-permissive host cells, we identified ten separate target sequences in the SARS-CoV-2 genome; three of these targets have been lost through conserved mutations. A genomic site targeted by the highly abundant miR-197-5p, overexpressed in patients with cardiovascular disease, is lost by a conserved mutation. Our results are compatible with a model that SARS-CoV-2 replication within the human host is constrained by host miRNA defences. The impact of these and further mutations on secondary structures, miRNA targets or potential splice sites offers a new context in which to view future SARS-CoV-2 evolution, and a potential platform for engineering conditional attenuation to vaccine development, as well as providing a better understanding of viral tropism and pathogenesis.


Assuntos
Betacoronavirus/genética , Genoma Viral , MicroRNAs/metabolismo , RNA Viral/química , Regiões 3' não Traduzidas , Sequência de Bases , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Bases de Dados Genéticas , Humanos , MicroRNAs/química , MicroRNAs/genética , Mutação , Conformação de Ácido Nucleico , Pandemias , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Sítios de Splice de RNA , Processamento de RNA , Alinhamento de Sequência , Proteínas não Estruturais Virais/genética , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais/metabolismo
18.
Virol J ; 17(1): 117, 2020 07 29.
Artigo em Inglês | MEDLINE | ID: mdl-32727485

RESUMO

Coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2 infection has spread rapidly across the world and become an international public health emergency. Both SARS-CoV-2 and SARS-CoV belong to subfamily Coronavirinae in the family Coronaviridae of the order Nidovirales and they are classified as the SARS-like species while belong to different cluster. Besides, viral structure, epidemiology characteristics and pathological characteristics are also different. We present a comprehensive survey of the latest coronavirus-SARS-CoV-2-from investigating its origin and evolution alongside SARS-CoV. Meanwhile, pathogenesis, cardiovascular disease in COVID-19 patients, myocardial injury and venous thromboembolism induced by SARS-CoV-2 as well as the treatment methods are summarized in this review.


Assuntos
Betacoronavirus , Infecções por Coronavirus , Pandemias , Pneumonia Viral , Antivirais/uso terapêutico , Infecções Assintomáticas , Betacoronavirus/química , Betacoronavirus/classificação , Betacoronavirus/patogenicidade , Betacoronavirus/fisiologia , Comorbidade , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/patologia , Infecções por Coronavirus/terapia , Suscetibilidade a Doenças , Evolução Molecular , Genoma Viral , Humanos , Imunização Passiva , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/epidemiologia , Pneumonia Viral/imunologia , Pneumonia Viral/patologia , Pneumonia Viral/terapia , Receptores Virais/metabolismo , Vírus da SARS/química , Vírus da SARS/classificação , Vírus da SARS/patogenicidade , Vírus da SARS/fisiologia , Proteínas Virais/química
19.
Crit Rev Clin Lab Sci ; 57(6): 400-414, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32662342

RESUMO

Since Coronavirus Disease 2019 (COVID-19) first emerged in December 2019, the disease has rapidly evolved into a pandemic that threatens societies around the world. As soon as the causative pathogen, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was identified and its genome sequence determined, a laboratory diagnosis based on nucleic acid amplification technologies was quickly established and has played essential roles in the confirmation of a clinical diagnosis. Serological testing for antibodies against SARS-CoV-2 is becoming available for complementary diagnosis, identification of convalescent plasma, and epidemiologic studies. Additional laboratory biochemical tests, including monitoring the change in blood cells, blood gas, coagulation, liver function, cardiac markers, and inflammatory responses such as cytokine levels in plasma, are also critical in combating COVID-19. Nevertheless, with overwhelming numbers of patients and potentially large numbers of asymptomatic cases, clinical laboratories encounter enormous challenges in diagnostic approaches that can rapidly and accurately identify infected persons. Strategies that can effectively detect disease progression in order to stratify patients for appropriate care, and that can thereby prevent exacerbation of the disease, are urgently needed. This review discusses the laboratory's role and challenges in combating COVID-19.


Assuntos
Betacoronavirus/fisiologia , Serviços de Laboratório Clínico , Infecções por Coronavirus/terapia , Pneumonia Viral/terapia , Sequência de Aminoácidos , Infecções por Coronavirus/sangue , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/epidemiologia , Humanos , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/diagnóstico , Pneumonia Viral/epidemiologia , Proteínas Virais/química
20.
BMC Bioinformatics ; 21(Suppl 12): 305, 2020 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-32703190

RESUMO

BACKGROUND: Horizontal gene transfer, i.e. the acquisition of genetic material from nonparent organism, is considered an important force driving species evolution. Many cases of horizontal gene transfer from prokaryotes to eukaryotes have been registered, but no transfer mechanism has been deciphered so far, although viruses were proposed as possible vectors in several studies. In agreement with this idea, in our previous study we discovered that in two eukaryotic proteins bacteriophage recombination site (AttP) was adjacent to the regions originating via horizontal gene transfer. In one of those cases AttP site was present inside the introns of cysteine-rich repeats. In the present study we aimed to apply computational tools for finding multiple horizontal gene transfer events in large genome databases. For that purpose we used a sequence of cysteine-rich repeats to identify genes potentially acquired through horizontal transfer. RESULTS: HMMER remote similarity search significantly detected 382 proteins containing cysteine-rich repeats. All of them, except 8 sequences, belong to eukaryotes. In 124 proteins the presence of conserved structural domains was predicted. In spite of the fact that cysteine-rich repeats are found almost exclusively in eukaryotic proteins, many predicted domains are most common for prokaryotes or bacteriophages. Ninety-eight proteins out of 124 contain typical prokaryotic domains. In those cases proteins were considered as potentially originating via horizontal transfer. In addition, HHblits search revealed that two domains of the same fungal protein, Glycoside hydrolase and Peptidase M15, have high similarity with proteins of two different prokaryotic species, hinting at independent horizontal gene transfer events. CONCLUSIONS: Cysteine-rich repeats in eukaryotic proteins are usually accompanied by conserved domains typical for prokaryotes or bacteriophages. These proteins, containing both cysteine-rich repeats, and characteristic prokaryotic domains, might represent multiple independent horizontal gene transfer events from prokaryotes to eukaryotes. We believe that the presence of bacteriophage recombination site inside cysteine-rich repeat coding sequence may facilitate horizontal genes transfer. Thus computational approach, described in the present study, can help finding multiple sequences originated from horizontal transfer in eukaryotic genomes.


Assuntos
Bacteriófagos/genética , Transferência Genética Horizontal/genética , Genes Virais , Recombinação Genética/genética , Proteínas Virais/química , Sequência de Aminoácidos , Sequência de Bases , Sequência Conservada , Domínios Proteicos , Proteínas Virais/classificação
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