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1.
Int J Antimicrob Agents ; 56(2): 106020, 2020 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-32862840

RESUMEN

The emergence of SARS-coronavirus-2 (SARS-CoV-2) has led to a global pandemic disease referred to as coronavirus disease 19 (COVID-19). Hydroxychloroquine (CLQ-OH)/azithromycin (ATM) combination therapy is currently being tested for the treatment of COVID-19, with promising results. However, the molecular mechanism of action of this combination is not yet established. Using molecular dynamics (MD) simulations, this study shows that the drugs act in synergy to prevent any close contact between the virus and the plasma membrane of host cells. Unexpected molecular similarity is shown between ATM and the sugar moiety of GM1, a lipid raft ganglioside acting as a host attachment cofactor for respiratory viruses. Due to this mimicry, ATM interacts with the ganglioside-binding domain of SARS-CoV-2 spike protein. This binding site shared by ATM and GM1 displays a conserved amino acid triad Q-134/F-135/N-137 located at the tip of the spike protein. CLQ-OH molecules are shown to saturate virus attachment sites on gangliosides in the vicinity of the primary coronavirus receptor, angiotensin-converting enzyme-2 (ACE-2). Taken together, these data show that ATM is directed against the virus, whereas CLQ-OH is directed against cellular attachment cofactors. We conclude that both drugs act as competitive inhibitors of SARS-CoV-2 attachment to the host-cell membrane. This is consistent with a synergistic antiviral mechanism at the plasma membrane level, where therapeutic intervention is likely to be most efficient. This molecular mechanism may explain the beneficial effects of CLQ-OH/ATM combination therapy in patients with COVID-19. Incidentally, the data also indicate that the conserved Q-134/F-135/N-137 triad could be considered as a target for vaccine strategies.


Asunto(s)
Azitromicina/farmacología , Infecciones por Coronavirus/tratamiento farmacológico , Gangliósido G(M1)/metabolismo , Hidroxicloroquina/farmacología , Neumonía Viral/tratamiento farmacológico , Glicoproteína de la Espiga del Coronavirus/metabolismo , Acoplamiento Viral/efectos de los fármacos , Secuencia de Aminoácidos , Antivirales/farmacología , Betacoronavirus/efectos de los fármacos , Betacoronavirus/metabolismo , Sitios de Unión/efectos de los fármacos , Sinergismo Farmacológico , Quimioterapia Combinada/métodos , Interacciones Huésped-Patógeno/efectos de los fármacos , Humanos , Simulación de Dinámica Molecular , Pandemias , Peptidil-Dipeptidasa A/metabolismo , Unión Proteica/efectos de los fármacos , Dominios Proteicos/efectos de los fármacos , Alineación de Secuencia
2.
BMC Infect Dis ; 20(1): 565, 2020 Aug 03.
Artículo en Inglés | MEDLINE | ID: mdl-32746807

RESUMEN

BACKGROUND: Patients coinfected with HBV and hepatitis D virus (HDV) have a greater risk of HCC and cirrhosis. The current study was undertaken to assess HDV genotype distribution and determine clinical characteristics of hepatitis delta virus (HDV) among HBsAg positive individuals in Shanghai. METHOD: This retrospective study involved 225 serum samples from HBsAg positive hospitalized patients from October 2010 to April 2013. HDV-specific RT-nested PCR was used to amplify HDV RNA. HDV genotypes were characterized by Next-generation sequencing (NGS), followed by phylogenetic analyses. HDV/HBV co-infected patients and HBV mono-infected patients were compared clinically and virologically. RESULTS: Out of the 225 HBsAg-positive serum samples with elevated transaminases, HDV-RNA was identified in 11 (4.9%) patients. The HBV loads in the HDV positive group were significantly lower than the HDV negative HBV-infected patients. The aminotransferase enzymes were significantly higher in HDV/HBV co-infected compared to HDV negative patients (P < 0.05). Phylogenetic analyses indicated that HDV-2 genotype being the predominant genotype, other HDV genotypes were not observed. HDV/HBV patients were significantly associated with a rather unfavourable clinical outcome. CONCLUSION: In summary, the prevalence of HDV infection in patients with elevated transaminases is not low and the predominance of HDV genotype 2 infection in Shanghai. This finding helps us to better understand the correlation of HDV/HBV co-infection. Moreover, Next-generation sequencing (NGS) technologies provide a rapid, precise method for generating HDV genomes to define infecting genotypes.


Asunto(s)
Hepatitis D/diagnóstico , Virus de la Hepatitis Delta/genética , Adulto , Secuencia de Aminoácidos , China/epidemiología , Coinfección , Femenino , Genotipo , Hepatitis B/diagnóstico , Hepatitis B/virología , Antígenos de Superficie de la Hepatitis B/sangre , Hepatitis D/epidemiología , Hepatitis D/virología , Virus de la Hepatitis Delta/clasificación , Virus de la Hepatitis Delta/aislamiento & purificación , Humanos , Masculino , Persona de Mediana Edad , Filogenia , Prevalencia , ARN Viral/química , ARN Viral/metabolismo , Estudios Retrospectivos , Factores de Riesgo , Carga Viral , Adulto Joven
3.
BMC Bioinformatics ; 21(1): 340, 2020 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-32738892

RESUMEN

BACKGROUND: Ribosome profiling has been widely used for studies of translation under a large variety of cellular and physiological contexts. Many of these studies have greatly benefitted from a series of data-mining tools designed for dissection of the translatome from different aspects. However, as the studies of translation advance quickly, the current toolbox still falls in short, and more specialized tools are in urgent need for deeper and more efficient mining of the important and new features of the translation landscapes. RESULTS: Here, we present RiboMiner, a bioinformatics toolset for mining of multi-dimensional features of the translatome with ribosome profiling data. RiboMiner performs extensive quality assessment of the data and integrates a spectrum of tools for various metagene analyses of the ribosome footprints and for detailed analyses of multiple features related to translation regulation. Visualizations of all the results are available. Many of these analyses have not been provided by previous methods. RiboMiner is highly flexible, as the pipeline could be easily adapted and customized for different scopes and targets of the studies. CONCLUSIONS: Applications of RiboMiner on two published datasets did not only reproduced the main results reported before, but also generated novel insights into the translation regulation processes. Therefore, being complementary to the current tools, RiboMiner could be a valuable resource for dissections of the translation landscapes and the translation regulations by mining the ribosome profiling data more comprehensively and with higher resolution. RiboMiner is freely available at https://github.com/xryanglab/RiboMiner and https://pypi.org/project/RiboMiner .


Asunto(s)
Biología Computacional/métodos , Biosíntesis de Proteínas , Ribosomas/metabolismo , Programas Informáticos , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Aminoácidos/genética , Codón/genética , Análisis de Datos , Minería de Datos
4.
Nat Commun ; 11(1): 3895, 2020 08 04.
Artículo en Inglés | MEDLINE | ID: mdl-32753588

RESUMEN

The mussel byssus has long been a source of inspiration for the adhesion community. Recently, adhesive synergy between flanking lysine (Lys, K) and 3,4-Dihydroxyphenylalanine (DOPA, Y) residues in the mussel foot proteins (Mfps) has been highlighted. However, the complex topological relationship of DOPA and Lys as well as the interfacial adhesive roles of other amino acids have been understudied. Herein, we study adhesion of Lys and DOPA-containing peptides to organic and inorganic substrates using single-molecule force spectroscopy (SMFS). We show that a modest increase in peptide length, from KY to (KY)3, increases adhesion strength to TiO2. Surprisingly, further increase in peptide length offers no additional benefit. Additionally, comparison of adhesion of dipeptides containing Lys and either DOPA (KY) or phenylalanine (KF) shows that DOPA is stronger and more versatile. We furthermore demonstrate that incorporating a nonadhesive spacer between (KY) repeats can mimic the hidden length in the Mfp and act as an effective strategy to dissipate energy.


Asunto(s)
Adhesivos/química , Dihidroxifenilalanina/química , Lisina/química , Secuencia de Aminoácidos , Animales , Bivalvos , Dipéptidos , Péptidos/síntesis química , Propiedades de Superficie , Titanio/química
5.
J Phys Chem Lett ; 11(16): 6655-6663, 2020 Aug 20.
Artículo en Inglés | MEDLINE | ID: mdl-32787225

RESUMEN

The COVID-19 pandemic is an urgent global health emergency, and the presence of Furin site in the SARS-CoV-2 spike glycoprotein alters virulence and warrants further molecular, structural, and biophysical studies. Here we report the structure of Furin in complex with SARS-CoV-2 spike glycoprotein, demonstrating how Furin binds to the S1/S2 region of spike glycoprotein and eventually cleaves the viral protein using experimental functional studies, molecular dynamics, and docking. The structural studies underline the mechanism and mode of action of Furin, which is a key process in host cell entry and a hallmark of enhanced virulence. Our whole-exome sequencing analysis shows the genetic variants/alleles in Furin were found to alter the binding affinity for viral spike glycoprotein and could vary in infectivity in humans. Unravelling the mechanisms of Furin action, binding dynamics, and the genetic variants opens the growing arena of bona fide antibodies and development of potential therapeutics targeting the blockage of Furin cleavage.


Asunto(s)
Betacoronavirus/química , Furina/metabolismo , Glicoproteína de la Espiga del Coronavirus/metabolismo , Virulencia/fisiología , Secuencia de Aminoácidos , Animales , Betacoronavirus/patogenicidad , Células CHO , Dominio Catalítico , Cricetulus , Furina/química , Furina/genética , Expresión Génica/fisiología , Hexosaminas/metabolismo , Humanos , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Unión Proteica , Proteolisis , Inhibidores de Serina Proteinasa/metabolismo , Glicoproteína de la Espiga del Coronavirus/química
6.
Int J Mol Sci ; 21(16)2020 Aug 13.
Artículo en Inglés | MEDLINE | ID: mdl-32823591

RESUMEN

While SARS-CoV-2 uses angiotensin converting enzyme 2 (ACE2) as the receptor for cell entry, it is important to examine other potential interactions between the virus and other cell receptors. Based on the clinical observation of low prevalence of smoking among hospitalized COVID-19 patients, we examined and identified a "toxin-like" amino acid (aa) sequence in the Receptor Binding Domain of the Spike Glycoprotein of SARS-CoV-2 (aa 375-390), which is homologous to a sequence of the Neurotoxin homolog NL1, one of the many snake venom toxins that are known to interact with nicotinic acetylcholine receptors (nAChRs). We present the 3D structural location of this "toxin-like" sequence on the Spike Glycoprotein and the superposition of the modelled structure of the Neurotoxin homolog NL1 and the SARS-CoV-2 Spike Glycoprotein. We also performed computational molecular modelling and docking experiments using 3D structures of the SARS-CoV-2 Spike Glycoprotein and the extracellular domain of the nAChR α9 subunit. We identified a main interaction between the aa 381-386 of the SARS-CoV-2 Spike Glycoprotein and the aa 189-192 of the extracellular domain of the nAChR α9 subunit, a region which forms the core of the "toxin-binding site" of the nAChRs. The mode of interaction is very similar to the interaction between the α9 nAChR and α-bungarotoxin. A similar interaction was observed between the pentameric α7 AChR chimera and SARS-CoV-2 Spike Glycoprotein. The findings raise the possibility that SARS-CoV-2 may interact with nAChRs, supporting the hypothesis of dysregulation of the nicotinic cholinergic system being implicated in the pathophysiology of COVID-19. Nicotine and other nicotinic cholinergic agonists may protect nAChRs and thus have therapeutic value in COVID-19 patients.


Asunto(s)
Betacoronavirus/metabolismo , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/metabolismo , Secuencia de Aminoácidos/genética , Biología Computacional , Infecciones por Coronavirus/fisiopatología , Humanos , Simulación del Acoplamiento Molecular , Neurotoxinas/genética , Neurotoxinas/metabolismo , Pandemias , Neumonía Viral/fisiopatología , Estructura Terciaria de Proteína/genética , Alineación de Secuencia , Venenos de Serpiente/genética
7.
mBio ; 11(4)2020 08 20.
Artículo en Inglés | MEDLINE | ID: mdl-32820005

RESUMEN

We assessed various newly generated compounds that target the main protease (Mpro) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and various previously known compounds reportedly active against SARS-CoV-2, employing RNA quantitative PCR (RNA-qPCR), cytopathicity assays, and immunocytochemistry. Here, we show that two indole-chloropyridinyl-ester derivatives, GRL-0820 and GRL-0920, exerted potent activity against SARS-CoV-2 in cell-based assays performed using VeroE6 cells and TMPRSS2-overexpressing VeroE6 cells. While GRL-0820 and the nucleotide analog remdesivir blocked SARS-CoV-2 infection, viral breakthrough occurred. No significant anti-SARS-CoV-2 activity was found for several compounds reportedly active against SARS-CoV-2 such as lopinavir, nelfinavir, nitazoxanide, favipiravir, and hydroxychroloquine. In contrast, GRL-0920 exerted potent activity against SARS-CoV-2 (50% effective concentration [EC50] = 2.8 µM) and dramatically reduced the infectivity, replication, and cytopathic effect of SARS-CoV-2 without significant toxicity as examined with immunocytochemistry. Structural modeling shows that indole and chloropyridinyl of the derivatives interact with two catalytic dyad residues of Mpro, Cys145 and His41, resulting in covalent bonding, which was verified using high-performance liquid chromatography-mass spectrometry (HPLC/MS), suggesting that the indole moiety is critical for the anti-SARS-CoV-2 activity of the derivatives. GRL-0920 might serve as a potential therapeutic for coronavirus disease 2019 (COVID-19) and might be optimized to generate more-potent anti-SARS-CoV-2 compounds.IMPORTANCE Targeting the main protease (Mpro) of SARS-CoV-2, we identified two indole-chloropyridinyl-ester derivatives, GRL-0820 and GRL-0920, active against SARS-CoV-2, employing RNA-qPCR and immunocytochemistry and show that the two compounds exerted potent activity against SARS-CoV-2. While GRL-0820 and remdesivir blocked SARS-CoV-2 infection, viral breakthrough occurred as examined with immunocytochemistry. In contrast, GRL-0920 completely blocked the infectivity and cytopathic effect of SARS-CoV-2 without significant toxicity. Structural modeling showed that indole and chloropyridinyl of the derivatives interacted with two catalytic dyad residues of Mpro, Cys145 and His41, resulting in covalent bonding, which was verified using HPLC/MS. The present data should shed light on the development of therapeutics for COVID-19, and optimization of GRL-0920 based on the present data is essential to develop more-potent anti-SARS-CoV-2 compounds for treating COVID-19.


Asunto(s)
Antivirales/farmacología , Betacoronavirus/efectos de los fármacos , Infecciones por Coronavirus/tratamiento farmacológico , Indoles/farmacología , Neumonía Viral/tratamiento farmacológico , Secuencia de Aminoácidos , Animales , Betacoronavirus/enzimología , Chlorocebus aethiops , Cloroquina/farmacología , Infecciones por Coronavirus/virología , Cisteína Endopeptidasas/química , Cisteína Endopeptidasas/genética , Cisteína Endopeptidasas/metabolismo , Indoles/química , Indoles/uso terapéutico , Modelos Moleculares , Pandemias , Neumonía Viral/virología , Células Vero , Proteínas no Estructurales Virales/antagonistas & inhibidores , Proteínas no Estructurales Virales/química , Proteínas no Estructurales Virales/genética , Proteínas no Estructurales Virales/metabolismo
8.
PLoS Negl Trop Dis ; 14(8): e0008510, 2020 08.
Artículo en Inglés | MEDLINE | ID: mdl-32760059

RESUMEN

Serine protease inhibitors (serpins) regulate proteolytic events within diverse biological processes, including digestion, coagulation, inflammation and immune responses. The presence of serpins in Fasciola hepatica excretory-secretory products indicates that the parasite exploits these to regulate proteases encountered during its development within vertebrate hosts. Interrogation of the F. hepatica genome identified a multi-gene serpin family of seven members that has expanded by gene duplication and divergence to create an array of inhibitors with distinct specificities. We investigated the molecular properties and functions of two representatives, FhSrp1 and FhSrp2, highly expressed in the invasive newly excysted juvenile (NEJ). Consistent with marked differences in the reactive centre loop (RCL) that executes inhibitor-protease complexing, the two recombinant F. hepatica serpins displayed distinct inhibitory profiles against an array of mammalian serine proteases. In particular, rFhSrp1 efficiently inhibited kallikrein (Ki = 40 nM) whilst rFhSrp2 was a highly potent inhibitor of chymotrypsin (Ki = 0.07 nM). FhSrp1 and FhSrp2 are both expressed on the NEJ surface, predominantly around the oral and ventral suckers, suggesting that these inhibitors protect the parasites from the harmful proteolytic effects of host proteases, such as chymotrypsin, during invasion. Furthermore, the unusual inhibition of kallikrein suggests that rFhSrp1 modulates host responses such as inflammation and vascular permeability by interfering with the kallikrein-kinin system. A vaccine combination of rFhSrp1 and rFhSrp2 formulated in the adjuvant Montanide ISA 206VG elicited modest but non-significant protection against a challenge infection in a rat model, but did induce some protection against liver pathogenesis when compared to a control group and a group vaccinated with two well-studied vaccine candidates, F. hepatica cathepsin L2 and L3. This work highlights the importance of F. hepatica serpins to regulate host responses that enables parasite survival during infection and, coupled with the vaccine data, encourages future vaccine trials in ruminants.


Asunto(s)
Fasciola hepatica/metabolismo , Inhibidores de Serina Proteinasa/metabolismo , Serpinas/metabolismo , Secuencia de Aminoácidos , Animales , Fasciola hepatica/genética , Regulación de la Expresión Génica , Interacciones Huésped-Parásitos
9.
Nat Commun ; 11(1): 3969, 2020 08 07.
Artículo en Inglés | MEDLINE | ID: mdl-32769976

RESUMEN

Mevalonate diphosphate decarboxylases (MDDs) catalyze the ATP-dependent-Mg2+-decarboxylation of mevalonate-5-diphosphate (MVAPP) to produce isopentenyl diphosphate (IPP), which is essential in both eukaryotes and prokaryotes for polyisoprenoid synthesis. The substrates, MVAPP and ATP, have been shown to bind sequentially to MDD. Here we report crystals in which the enzyme remains active, allowing the visualization of conformational changes in Enterococcus faecalis MDD that describe sequential steps in an induced fit enzymatic reaction. Initial binding of MVAPP modulates the ATP binding pocket with a large loop movement. Upon ATP binding, a phosphate binding loop bends over the active site to recognize ATP and bring the molecules to their catalytically favored configuration. Positioned substrates then can chelate two Mg2+ ions for the two steps of the reaction. Closure of the active site entrance brings a conserved lysine to trigger dissociative phosphoryl transfer of γ-phosphate from ATP to MVAPP, followed by the production of IPP.


Asunto(s)
Carboxiliasas/metabolismo , Enterococcus faecalis/enzimología , Secuencia de Aminoácidos , Sitios de Unión , Biocatálisis , Carboxiliasas/química , Secuencia Conservada , Cristalografía por Rayos X , Ligandos , Metales/metabolismo , Modelos Moleculares , Estructura Secundaria de Proteína , Especificidad por Sustrato
10.
Nat Commun ; 11(1): 4031, 2020 08 12.
Artículo en Inglés | MEDLINE | ID: mdl-32788582

RESUMEN

Calcium (Ca2+) influx into mitochondria occurs through a Ca2+-selective uniporter channel, which regulates essential cellular processes in eukaryotic organisms. Previous evolutionary analyses of its pore-forming subunits MCU and EMRE, and gatekeeper MICU1, pinpointed an evolutionary paradox: the presence of MCU homologs in fungal species devoid of any other uniporter components and of mt-Ca2+ uptake. Here, we trace the mt-Ca2+ uniporter evolution across 1,156 fully-sequenced eukaryotes and show that animal and fungal MCUs represent two distinct paralogous subfamilies originating from an ancestral duplication. Accordingly, we find EMRE orthologs outside Holoza and uncover the existence of an animal-like uniporter within chytrid fungi, which enables mt-Ca2+ uptake when reconstituted in vivo in the yeast Saccharomyces cerevisiae. Our study represents the most comprehensive phylogenomic analysis of the mt-Ca2+ uptake system and demonstrates that MCU, EMRE, and MICU formed the core of the ancestral opisthokont uniporter, with major implications for comparative structural and functional studies.


Asunto(s)
Canales de Calcio/genética , Evolución Molecular , Proteínas Fúngicas/genética , Secuencia de Aminoácidos , Calcio/metabolismo , Canales de Calcio/química , Quitridiomicetos/genética , Proteínas Fúngicas/química , Células HeLa , Humanos , Funciones de Verosimilitud , Filogenia , Especificidad de la Especie
11.
Nat Commun ; 11(1): 4027, 2020 08 12.
Artículo en Inglés | MEDLINE | ID: mdl-32788676

RESUMEN

Programmed cell death or apoptosis is a central biological process that is dysregulated in many diseases, including inflammatory conditions and cancer. The detection and quantification of apoptotic cells in vivo is hampered by the need for fixatives or washing steps for non-fluorogenic reagents, and by the low levels of free calcium in diseased tissues that restrict the use of annexins. In this manuscript, we report the rational design of a highly stable fluorogenic peptide (termed Apo-15) that selectively stains apoptotic cells in vitro and in vivo in a calcium-independent manner and under wash-free conditions. Furthermore, using a combination of chemical and biophysical methods, we identify phosphatidylserine as a molecular target of Apo-15. We demonstrate that Apo-15 can be used for the quantification and imaging of drug-induced apoptosis in preclinical mouse models, thus creating opportunities for assessing the in vivo efficacy of anti-inflammatory and anti-cancer therapeutics.


Asunto(s)
Apoptosis , Imagenología Tridimensional , Péptidos Cíclicos/farmacología , Secuencia de Aminoácidos , Animales , Apoptosis/efectos de los fármacos , Línea Celular , Femenino , Humanos , Ratones Endogámicos C57BL , Microscopía Fluorescente , Neutrófilos/citología , Neutrófilos/efectos de los fármacos , Péptidos Cíclicos/administración & dosificación , Péptidos Cíclicos/síntesis química , Péptidos Cíclicos/química , Fagocitosis/efectos de los fármacos , Fosfatidilserinas/metabolismo
12.
Sci Rep ; 10(1): 14004, 2020 08 19.
Artículo en Inglés | MEDLINE | ID: mdl-32814791

RESUMEN

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), a novel evolutionary divergent RNA virus, is responsible for the present devastating COVID-19 pandemic. To explore the genomic signatures, we comprehensively analyzed 2,492 complete and/or near-complete genome sequences of SARS-CoV-2 strains reported from across the globe to the GISAID database up to 30 March 2020. Genome-wide annotations revealed 1,516 nucleotide-level variations at different positions throughout the entire genome of SARS-CoV-2. Moreover, nucleotide (nt) deletion analysis found twelve deletion sites throughout the genome other than previously reported deletions at coding sequence of the ORF8 (open reading frame), spike, and ORF7a proteins, specifically in polyprotein ORF1ab (n = 9), ORF10 (n = 1), and 3´-UTR (n = 2). Evidence from the systematic gene-level mutational and protein profile analyses revealed a large number of amino acid (aa) substitutions (n = 744), demonstrating the viral proteins heterogeneous. Notably, residues of receptor-binding domain (RBD) showing crucial interactions with angiotensin-converting enzyme 2 (ACE2) and cross-reacting neutralizing antibody were found to be conserved among the analyzed virus strains, except for replacement of lysine with arginine at 378th position of the cryptic epitope of a Shanghai isolate, hCoV-19/Shanghai/SH0007/2020 (EPI_ISL_416320). Furthermore, our results of the preliminary epidemiological data on SARS-CoV-2 infections revealed that frequency of aa mutations were relatively higher in the SARS-CoV-2 genome sequences of Europe (43.07%) followed by Asia (38.09%), and North America (29.64%) while case fatality rates remained higher in the European temperate countries, such as Italy, Spain, Netherlands, France, England and Belgium. Thus, the present method of genome annotation employed at this early pandemic stage could be a promising tool for monitoring and tracking the continuously evolving pandemic situation, the associated genetic variants, and their implications for the development of effective control and prophylaxis strategies.


Asunto(s)
Betacoronavirus/clasificación , Betacoronavirus/genética , Infecciones por Coronavirus/epidemiología , Heterogeneidad Genética , Genoma Viral/genética , Estudio de Asociación del Genoma Completo/métodos , Salud Global , Neumonía Viral/epidemiología , Secuencia de Aminoácidos/genética , Anticuerpos Neutralizantes/inmunología , Disparidad de Par Base , Secuencia de Bases/genética , Clima , Infecciones por Coronavirus/virología , Humanos , Sistemas de Lectura Abierta/genética , Pandemias , Peptidil-Dipeptidasa A/metabolismo , Filogenia , Neumonía Viral/virología , Dominios Proteicos/genética , Dominios Proteicos/inmunología , Eliminación de Secuencia , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/metabolismo
13.
Infect Dis Poverty ; 9(1): 88, 2020 Jul 10.
Artículo en Inglés | MEDLINE | ID: mdl-32741372

RESUMEN

BACKGROUND: An outbreak of infection caused by SARS-CoV-2 recently has brought a great challenge to public health. Rapid identification of immune epitopes would be an efficient way to screen the candidates for vaccine development at the time of pandemic. This study aimed to predict the protective epitopes with bioinformatics methods and resources for vaccine development. METHODS: The genome sequence and protein sequences of SARS-CoV-2 were retrieved from the National Center for Biotechnology Information (NCBI) database. ABCpred and BepiPred servers were utilized for sequential B-cell epitope analysis. Discontinuous B-cell epitopes were predicted via DiscoTope 2.0 program. IEDB server was utilized for HLA-1 and HLA-2 binding peptides computation. Surface accessibility, antigenicity, and other important features of forecasted epitopes were characterized for immunogen potential evaluation. RESULTS: A total of 63 sequential B-cell epitopes on spike protein were predicted and 4 peptides (Spike315-324, Spike333-338, Spike648-663, Spike1064-1079) exhibited high antigenicity score and good surface accessibility. Ten residues within spike protein (Gly496, Glu498, Pro499, Thr500, Leu1141, Gln1142, Pro1143, Glu1144, Leu1145, Asp1146) are forecasted as components of discontinuous B-cell epitopes. The bioinformatics analysis of HLA binding peptides within nucleocapsid protein produced 81 and 64 peptides being able to bind MHC class I and MHC class II molecules respectively. The peptides (Nucleocapsid66-75, Nucleocapsid104-112) were predicted to bind a wide spectrum of both HLA-1 and HLA-2 molecules. CONCLUSIONS: B-cell epitopes on spike protein and T-cell epitopes within nucleocapsid protein were identified and recommended for developing a protective vaccine against SARS-CoV-2.


Asunto(s)
Betacoronavirus/genética , Betacoronavirus/inmunología , Biología Computacional/métodos , Infecciones por Coronavirus/prevención & control , Epítopos de Linfocito B/inmunología , Epítopos de Linfocito T/inmunología , Pandemias/prevención & control , Neumonía Viral/prevención & control , Vacunas Virales/inmunología , Secuencia de Aminoácidos , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/virología , Diseño de Fármacos , Epítopos de Linfocito B/química , Epítopos de Linfocito T/química , Humanos , Inmunogenicidad Vacunal/inmunología , Modelos Moleculares , Neumonía Viral/inmunología , Neumonía Viral/virología , Alineación de Secuencia , Análisis de Secuencia , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunología , Proteínas del Envoltorio Viral/inmunología
14.
Med Sci (Paris) ; 36(8-9): 783-796, 2020.
Artículo en Francés | MEDLINE | ID: mdl-32773024

RESUMEN

SARS-CoV-2 is a new human coronavirus (CoV), which emerged in People's Republic of China at the end of 2019 and is responsible for the global Covid-19 pandemic that caused more than 540 000 deaths in six months. Understanding the origin of this virus is an important issue and it is necessary to determine the mechanisms of its dissemination in order to be able to contain new epidemics. Based on phylogenetic inferences, sequence analysis and structure-function relationships of coronavirus proteins, informed by the knowledge currently available, we discuss the different scenarios evoked to account for the origin - natural or synthetic - of the virus. On the basis of currently available data, it is impossible to determine whether SARS-CoV-2 is the result of a natural zoonotic emergence or an accidental escape from experimental strains. Regardless of its origin, the study of the evolution of the molecular mechanisms involved in the emergence of this pandemic virus is essential to develop therapeutic and vaccine strategies.


Asunto(s)
Betacoronavirus/genética , Enfermedades Transmisibles Emergentes/virología , Infecciones por Coronavirus/virología , Coronavirus/clasificación , Evolución Molecular , Pandemias , Filogenia , Neumonía Viral/virología , ARN Viral/genética , Secuencia de Aminoácidos , Animales , Betacoronavirus/clasificación , Betacoronavirus/aislamiento & purificación , Derrame de Material Biológico , China/epidemiología , Infecciones por Coronaviridae/transmisión , Infecciones por Coronaviridae/veterinaria , Infecciones por Coronaviridae/virología , Infecciones por Coronavirus/epidemiología , Infecciones por Coronavirus/transmisión , Reservorios de Enfermedades , Mutación con Ganancia de Función , Genoma Viral , VIH/genética , Especificidad del Huésped , Humanos , Mamíferos/virología , Neumonía Viral/epidemiología , Neumonía Viral/transmisión , Virus Reordenados/genética , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/fisiología , Zoonosis
15.
J Comput Chem ; 41(24): 2158-2161, 2020 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-32779780

RESUMEN

D614G spike glycoprotein (sgp) mutation in rapidly spreading severe acute respiratory syndrome coronavirus-2 (SARS-COV-2) is associated with enhanced fitness and higher transmissibility in new cases of COVID-19 but the underlying mechanism is unknown. Here, using atomistic simulation, a plausible mechanism has been delineated. In G614 sgp but not wild type, increased D(G)614-T859 Cα-distance within 65 ns is interpreted as S1/S2 protomer dissociation. Overall, ACE2-binding, post-fusion core, open-state and sub-optimal antibody-binding conformations were preferentially sampled by the G614 mutant, but not wild type. Furthermore, in the wild type, only one of the three sgp chains has optimal communication route between residue 614 and the receptor-binding domain (RBD); whereas, two of the three chains communicated directly in G614 mutant. These data provide evidence that D614G sgp mutant is more available for receptor binding, cellular invasion and reduced antibody interaction; thus, providing framework for enhanced fitness and higher transmissibility in D614G SARS-COV-2 mutant.


Asunto(s)
Betacoronavirus/metabolismo , Simulación por Computador , Infecciones por Coronavirus/virología , Modelos Químicos , Neumonía Viral/virología , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Secuencia de Aminoácidos , Sitios de Unión , Humanos , Modelos Moleculares , Mutación , Pandemias , Unión Proteica , Conformación Proteica , Dominios Proteicos
16.
Bioorg Med Chem Lett ; 30(17): 127377, 2020 09 01.
Artículo en Inglés | MEDLINE | ID: mdl-32738988

RESUMEN

The unprecedented pandemic of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is threatening global health. The virus emerged in late 2019 and can cause a severe disease associated with significant mortality. Several vaccine development and drug discovery campaigns are underway. The SARS-CoV-2 main protease is considered a promising drug target, as it is dissimilar to human proteases. Sequence and structure of the main protease are closely related to those from other betacoronaviruses, facilitating drug discovery attempts based on previous lead compounds. Covalently binding peptidomimetics and small molecules are investigated. Various compounds show antiviral activity in infected human cells.


Asunto(s)
Antivirales/farmacología , Betacoronavirus/efectos de los fármacos , Inhibidores de Cisteína Proteinasa/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Secuencia de Aminoácidos , Animales , Antivirales/farmacocinética , Betacoronavirus/enzimología , Línea Celular , Cisteína Endopeptidasas/química , Inhibidores de Cisteína Proteinasa/farmacocinética , Descubrimiento de Drogas , Humanos , Peptidomiméticos/farmacocinética , Peptidomiméticos/farmacología , Especificidad por Sustrato , Proteínas no Estructurales Virales/química
17.
Front Immunol ; 11: 1663, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32754160

RESUMEN

A recent pandemic caused by a single-stranded RNA virus, COVID-19, initially discovered in China, is now spreading globally. This poses a serious threat that needs to be addressed immediately. Genome analysis of SARS-CoV-2 has revealed its close relation to SARS-coronavirus along with few changes in its spike protein. The spike protein aids in receptor binding and viral entry within the host and therefore represents a potential target for vaccine and therapeutic development. In the current study, the spike protein of SARS-CoV-2 was explored for potential immunogenic epitopes to design multi-epitope vaccine constructs. The S1 and S2 domains of spike proteins were analyzed, and two vaccine constructs were prioritized with T-cell and B-cell epitopes. We adapted a comprehensive predictive framework to provide novel insights into immunogenic epitopes of spike proteins, which can further be evaluated as potential vaccine candidates against COVID-19. Prioritized epitopes were then modeled using linkers and adjuvants, and respective 3D models were constructed to evaluate their physiochemical properties and their possible interactions with ACE2, HLA Superfamily alleles, TLR2, and TLR4.


Asunto(s)
Betacoronavirus/inmunología , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/prevención & control , Epítopos de Linfocito B/inmunología , Epítopos de Linfocito T/inmunología , Pandemias/prevención & control , Neumonía Viral/inmunología , Neumonía Viral/prevención & control , Vacunas Virales/inmunología , Secuencia de Aminoácidos , Infecciones por Coronavirus/virología , Epítopos de Linfocito B/química , Epítopos de Linfocito T/química , Antígenos HLA/química , Antígenos HLA/inmunología , Humanos , Modelos Químicos , Simulación del Acoplamiento Molecular , Peptidil-Dipeptidasa A/química , Peptidil-Dipeptidasa A/inmunología , Neumonía Viral/virología , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Glicoproteína de la Espiga del Coronavirus/inmunología , Receptor Toll-Like 2/química , Receptor Toll-Like 2/inmunología , Receptor Toll-Like 4/química , Receptor Toll-Like 4/inmunología , Vacunas Virales/química
18.
Parasit Vectors ; 13(1): 409, 2020 Aug 10.
Artículo en Inglés | MEDLINE | ID: mdl-32778178

RESUMEN

The coronavirus disease 19 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected millions of people worldwide. Recent evidence raised the question about the possibility that cats may be a domestic host for SARS-CoV-2 with unknown implications in disease dissemination. Based on the fact that the domestic cat flea, Ctenocephalides felis, are abundant ectoparasites infesting humans, companion animals and wildlife and that coronavirus-like agents have been identified in the ectoparasite tick vector, Ixodes uriae of seabirds, herein we considered the presence of coronaviruses in general and SARS-CoV-2 in particular in C. felis. We identified coronavirus-derived and cell receptor angiotensin-converting enzyme RNA/proteins in C. felis. Although current evidence suggests that pets are probably dead-end-hosts with small risk of transmission to humans, our results suggested that cat flea may act as biological and/or mechanical vectors of SARS-CoV. Although preliminary, these results indicate a possibility of ectoparasites acting as reservoirs and vectors of SARS-CoV and related beta-coronavirus although with little disease risk due to systemic transmission route, low viremia, virus attenuation or other unknown factors. These results support the need to further study the role of animal SARS-CoV-2 hosts and their ectoparasite vectors in COVID-19 disease spread.


Asunto(s)
Infecciones por Coronavirus/veterinaria , Coronavirus/aislamiento & purificación , Ctenocephalides/virología , Insectos Vectores/virología , Neumonía Viral/veterinaria , Secuencia de Aminoácidos , Animales , Betacoronavirus/genética , Betacoronavirus/aislamiento & purificación , Infecciones por Coronavirus/transmisión , Infecciones por Coronavirus/virología , Humanos , Pandemias , Peptidil-Dipeptidasa A/genética , Neumonía Viral/transmisión , Neumonía Viral/virología
19.
Nat Commun ; 11(1): 3830, 2020 07 31.
Artículo en Inglés | MEDLINE | ID: mdl-32737313

RESUMEN

The mammalian mitochondrial ribosome (mitoribosome) and its associated translational factors have evolved to accommodate greater participation of proteins in mitochondrial translation. Here we present the 2.68-3.96 Å cryo-EM structures of the human 55S mitoribosome in complex with the human mitochondrial elongation factor G1 (EF-G1mt) in three distinct conformational states, including an intermediate state and a post-translocational state. These structures reveal the role of several mitochondria-specific (mito-specific) mitoribosomal proteins (MRPs) and a mito-specific segment of EF-G1mt in mitochondrial tRNA (tRNAmt) translocation. In particular, the mito-specific C-terminal extension in EF-G1mt is directly involved in translocation of the acceptor arm of the A-site tRNAmt. In addition to the ratchet-like and independent head-swiveling motions exhibited by the small mitoribosomal subunit, we discover significant conformational changes in MRP mL45 at the nascent polypeptide-exit site within the large mitoribosomal subunit that could be critical for tethering of the elongating mitoribosome onto the inner-mitochondrial membrane.


Asunto(s)
Mitocondrias/metabolismo , Proteínas Mitocondriales/química , Extensión de la Cadena Peptídica de Translación , Factor G de Elongación Peptídica/química , ARN Mitocondrial/química , ARN de Transferencia/química , Proteínas Ribosómicas/química , Ribosomas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Microscopía por Crioelectrón , Células HEK293 , Humanos , Mitocondrias/ultraestructura , Membranas Mitocondriales/metabolismo , Membranas Mitocondriales/ultraestructura , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Modelos Moleculares , Conformación de Ácido Nucleico , Factor G de Elongación Peptídica/genética , Factor G de Elongación Peptídica/metabolismo , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , ARN Mitocondrial/genética , ARN Mitocondrial/metabolismo , ARN de Transferencia/genética , ARN de Transferencia/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/metabolismo , Ribosomas/ultraestructura , Alineación de Secuencia , Homología de Secuencia de Aminoácido
20.
Nat Commun ; 11(1): 3797, 2020 07 30.
Artículo en Inglés | MEDLINE | ID: mdl-32732998

RESUMEN

Receptor-mediated perception of surface-exposed carbohydrates like lipo- and exo-polysaccharides (EPS) is important for non-self recognition and responses to microbial associated molecular patterns in mammals and plants. In legumes, EPS are monitored and can either block or promote symbiosis with rhizobia depending on their molecular composition. To establish a deeper understanding of receptors involved in EPS recognition, we determined the structure of the Lotus japonicus (Lotus) exopolysaccharide receptor 3 (EPR3) ectodomain. EPR3 forms a compact structure built of three putative carbohydrate-binding modules (M1, M2 and LysM3). M1 and M2 have unique ßαßß and ßαß folds that have not previously been observed in carbohydrate binding proteins, while LysM3 has a canonical ßααß fold. We demonstrate that this configuration is a structural signature for a ubiquitous class of receptors in the plant kingdom. We show that EPR3 is promiscuous, suggesting that plants can monitor complex microbial communities though this class of receptors.


Asunto(s)
Lipopolisacáridos/metabolismo , Lotus/microbiología , Lotus/fisiología , Mesorhizobium/metabolismo , Proteínas de Plantas/metabolismo , Secuencia de Aminoácidos , Mesorhizobium/genética , Fijación del Nitrógeno/fisiología , Proteínas de Plantas/genética , Pliegue de Proteína , Nódulos de las Raíces de las Plantas/microbiología , Nódulos de las Raíces de las Plantas/fisiología , Simbiosis/fisiología
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