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1.
Int J Mol Sci ; 21(21)2020 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-33158276

RESUMO

Binding to the host receptor is a critical initial step for the coronavirus SARS-CoV-2 spike protein to enter into target cells and trigger virus transmission. A detailed dynamic and energetic view of the binding mechanisms underlying virus entry is not fully understood and the consensus around the molecular origins behind binding preferences of SARS-CoV-2 for binding with the angiotensin-converting enzyme 2 (ACE2) host receptor is yet to be established. In this work, we performed a comprehensive computational investigation in which sequence analysis and modeling of coevolutionary networks are combined with atomistic molecular simulations and comparative binding free energy analysis of the SARS-CoV and SARS-CoV-2 spike protein receptor binding domains with the ACE2 host receptor. Different from other computational studies, we systematically examine the molecular and energetic determinants of the binding mechanisms between SARS-CoV-2 and ACE2 proteins through the lens of coevolution, conformational dynamics, and allosteric interactions that conspire to drive binding interactions and signal transmission. Conformational dynamics analysis revealed the important differences in mobility of the binding interfaces for the SARS-CoV-2 spike protein that are not confined to several binding hotspots, but instead are broadly distributed across many interface residues. Through coevolutionary network analysis and dynamics-based alanine scanning, we established linkages between the binding energy hotspots and potential regulators and carriers of signal communication in the virus-host receptor complexes. The results of this study detailed a binding mechanism in which the energetics of the SARS-CoV-2 association with ACE2 may be determined by cumulative changes of a number of residues distributed across the entire binding interface. The central findings of this study are consistent with structural and biochemical data and highlight drug discovery challenges of inhibiting large and adaptive protein-protein interfaces responsible for virus entry and infection transmission.


Assuntos
Betacoronavirus/metabolismo , Infecções por Coronavirus/metabolismo , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Infecções por Coronavirus/enzimologia , Infecções por Coronavirus/virologia , Interações entre Hospedeiro e Microrganismos , Humanos , Pandemias , Pneumonia Viral/enzimologia , Pneumonia Viral/virologia , Ligação Proteica , Domínios Proteicos , Receptores Virais/metabolismo , Transdução de Sinais , Internalização do Vírus
2.
Clin Sci (Lond) ; 134(21): 2851-2871, 2020 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-33146371

RESUMO

Angiotensin converting enzyme (ACE) is well-known for its role in blood pressure regulation via the renin-angiotensin aldosterone system (RAAS) but also functions in fertility, immunity, haematopoiesis and diseases such as obesity, fibrosis and Alzheimer's dementia. Like ACE, the human homologue ACE2 is also involved in blood pressure regulation and cleaves a range of substrates involved in different physiological processes. Importantly, it is the functional receptor for severe acute respiratory syndrome (SARS)-coronavirus (CoV)-2 responsible for the 2020, coronavirus infectious disease 2019 (COVID-19) pandemic. Understanding the interaction between SARS-CoV-2 and ACE2 is crucial for the design of therapies to combat this disease. This review provides a comparative analysis of methodologies and findings to describe how structural biology techniques like X-ray crystallography and cryo-electron microscopy have enabled remarkable discoveries into the structure-function relationship of ACE and ACE2. This, in turn, has enabled the development of ACE inhibitors for the treatment of cardiovascular disease and candidate therapies for the treatment of COVID-19. However, despite these advances the function of ACE homologues in non-human organisms is not yet fully understood. ACE homologues have been discovered in the tissues, body fluids and venom of species from diverse lineages and are known to have important functions in fertility, envenoming and insect-host defence mechanisms. We, therefore, further highlight the need for structural insight into insect and venom ACE homologues for the potential development of novel anti-venoms and insecticides.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/enzimologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/enzimologia , Receptores Virais/metabolismo , Internalização do Vírus , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Animais , Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Interações Hospedeiro-Patógeno , Humanos , Pandemias , Peptidil Dipeptidase A/química , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Conformação Proteica , Receptores Virais/química , Relação Estrutura-Atividade
3.
Nat Commun ; 11(1): 5588, 2020 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-33149112

RESUMO

The coronavirus SARS-CoV-2 is the cause of the ongoing COVID-19 pandemic. Therapeutic neutralizing antibodies constitute a key short-to-medium term approach to tackle COVID-19. However, traditional antibody production is hampered by long development times and costly production. Here, we report the rapid isolation and characterization of nanobodies from a synthetic library, known as sybodies (Sb), that target the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein. Several binders with low nanomolar affinities and efficient neutralization activity were identified of which Sb23 displayed high affinity and neutralized pseudovirus with an IC50 of 0.6 µg/ml. A cryo-EM structure of the spike bound to Sb23 showed that Sb23 binds competitively in the ACE2 binding site. Furthermore, the cryo-EM reconstruction revealed an unusual conformation of the spike where two RBDs are in the 'up' ACE2-binding conformation. The combined approach represents an alternative, fast workflow to select binders with neutralizing activity against newly emerging viruses.


Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Pandemias/prevenção & controle , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/prevenção & controle , Anticorpos de Domínio Único/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Microscopia Crioeletrônica , Humanos , Testes de Neutralização , Ligação Proteica , Conformação Proteica , Domínios Proteicos/imunologia , Receptores Virais/metabolismo
4.
Clin Sci (Lond) ; 134(21): 2791-2805, 2020 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-33135725

RESUMO

Angiotensin-converting enzyme II (ACE2) is a homologue of angiotensin-converting enzyme discovered in 2000. From the initial discovery, it was recognized that the kidneys were organs very rich on ACE2. Subsequent studies demonstrated the precise localization of ACE2 within the kidney and the importance of this enzyme in the metabolism of Angiotensin II and the formation of Angiotensin 1-7. With the recognition early in 2020 of ACE2 being the main receptor of severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2), the interest in this protein has dramatically increased. In this review, we will focus on kidney ACE2; its localization, its alterations in hypertension, diabetes, the effect of ACE inhibitors and angiotensin type 1 receptor blockers (ARBs) on ACE2 and the potential use of ACE2 recombinant proteins therapeutically for kidney disease. We also describe the emerging kidney manifestations of COVID-19, namely the frequent development of acute kidney injury. The possibility that binding of SARS-CoV-2 to kidney ACE2 plays a role in the kidney manifestations is also briefly discussed.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/enzimologia , Nefropatias/enzimologia , Rim/enzimologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/enzimologia , Receptores Virais/metabolismo , Lesão Renal Aguda/enzimologia , Lesão Renal Aguda/virologia , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Animais , Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Diabetes Mellitus/enzimologia , Diabetes Mellitus/fisiopatologia , História do Século XXI , Interações Hospedeiro-Patógeno , Humanos , Hipertensão/enzimologia , Hipertensão/fisiopatologia , Rim/fisiopatologia , Nefropatias/tratamento farmacológico , Nefropatias/fisiopatologia , Pandemias , Peptidil Dipeptidase A/história , Peptidil Dipeptidase A/uso terapêutico , Pneumonia Viral/virologia , Receptores Virais/história
5.
Cell Rep ; 33(4): 108322, 2020 10 27.
Artigo em Inglês | MEDLINE | ID: mdl-33091382

RESUMO

Biotin-labeled molecular probes, comprising specific regions of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike, would be helpful in the isolation and characterization of antibodies targeting this recently emerged pathogen. Here, we design constructs incorporating an N-terminal purification tag, a site-specific protease-cleavage site, the probe region of interest, and a C-terminal sequence targeted by biotin ligase. Probe regions include full-length spike ectodomain as well as various subregions, and we also design mutants that eliminate recognition of the angiotensin-converting enzyme 2 (ACE2) receptor. Yields of biotin-labeled probes from transient transfection range from ∼0.5 mg/L for the complete ectodomain to >5 mg/L for several subregions. Probes are characterized for antigenicity and ACE2 recognition, and the structure of the spike ectodomain probe is determined by cryoelectron microscopy. We also characterize antibody-binding specificities and cell-sorting capabilities of the biotinylated probes. Altogether, structure-based design coupled to efficient purification and biotinylation processes can thus enable streamlined development of SARS-CoV-2 spike ectodomain probes.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Infecções por Coronavirus/imunologia , Sondas Moleculares/imunologia , Pneumonia Viral/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Especificidade de Anticorpos/imunologia , Sítios de Ligação de Anticorpos/imunologia , Biotinilação , Microscopia Crioeletrônica , Humanos , Pandemias , Peptidil Dipeptidase A/metabolismo , Receptores Virais/metabolismo
6.
Front Immunol ; 11: 570927, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33123144

RESUMO

The emergence and rapid spread of SARS-CoV-2 in December 2019 has brought the world to a standstill. While less pathogenic than the 2002-2003 SARS-CoV, this novel betacoronavirus presents a global threat due to its high transmission rate, ability to invade multiple tissues, and ability to trigger immunological hyperactivation. The identification of the animal reservoir and intermediate host were important steps toward slowing the spread of disease, and its genetic similarity to SARS-CoV has helped to determine pathogenesis and direct treatment strategies. The exponential increase in cases has necessitated fast and reliable testing procedures. Although RT-PCR remains the gold standard, it is a time-consuming procedure, paving the way for newer techniques such as serologic tests and enzyme immunoassays. Various clinical trials using broad antiviral agents in addition to novel medications have produced controversial results; however, the advancement of immunotherapy, particularly monoclonal antibodies and immune modulators is showing great promise in clinical trials. Non-orthodox medications such as anti-malarials have been tested in multiple institutions but definitive conclusions are yet to be made. Adjuvant therapies have also proven to be effective in decreasing mortality in the disease course. While no formal guidelines have been established, the multitude of ongoing clinical trials as a result of unprecedented access to research data brings us closer to halting the SARS-CoV-2 pandemic.


Assuntos
Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus , Pandemias , Pneumonia Viral , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/uso terapêutico , Alanina/análogos & derivados , Alanina/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Anticorpos Antivirais/uso terapêutico , Betacoronavirus/genética , Betacoronavirus/imunologia , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/patologia , Infecções por Coronavirus/terapia , Reservatórios de Doenças/virologia , Reposicionamento de Medicamentos/métodos , Humanos , Técnicas Imunoenzimáticas , Imunoterapia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/diagnóstico , Pneumonia Viral/patologia , Pneumonia Viral/terapia , Receptores Virais/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Testes Sorológicos/métodos , Glicoproteína da Espícula de Coronavírus/metabolismo
7.
Viruses ; 12(11)2020 10 23.
Artigo em Inglês | MEDLINE | ID: mdl-33114247

RESUMO

Porcine respiratory coronavirus (PRCoV) infects the epithelial cells in the respiratory tract of pigs, causing a mild respiratory disease. We applied air-liquid interface (ALI) cultures of well-differentiated porcine airway cells to mimic the respiratory tract epithelium in vitro and use it for analyzing the infection by PRCoV. As reported for most coronaviruses, virus entry and virus release occurred mainly via the apical membrane domain. A novel finding was that PRCoV preferentially targets non-ciliated and among them the non-mucus-producing cells. Aminopeptidase N (APN), the cellular receptor for PRCoV was also more abundantly expressed on this type of cell suggesting that APN is a determinant of the cell tropism. Interestingly, differentiation-dependent differences were found both in the expression of pAPN and the susceptibility to PRCoV infection. Cells in an early differentiation stage express higher levels of pAPN and are more susceptible to infection by PRCoV than are well-differentiated cells. A difference in the susceptibility to infection was also detected when tracheal and bronchial cells were compared. The increased susceptibility to infection of bronchial epithelial cells was, however, not due to an increased abundance of APN on the cell surface. Our data reveal a complex pattern of infection in porcine differentiated airway epithelial cells that could not be elucidated with immortalized cell lines. The results are expected to have relevance also for the analysis of other respiratory viruses.


Assuntos
Antígenos CD13/metabolismo , Células Epiteliais/metabolismo , Coronavirus Respiratório Porcino/fisiologia , Receptores Virais/metabolismo , Mucosa Respiratória/virologia , Tropismo Viral , Animais , Brônquios/metabolismo , Brônquios/virologia , Diferenciação Celular , Células Cultivadas , Células Epiteliais/citologia , Células Epiteliais/virologia , Suínos , Traqueia/metabolismo , Traqueia/virologia , Internalização do Vírus , Liberação de Vírus , Replicação Viral
8.
Drug Discov Ther ; 14(5): 256-258, 2020 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-33116037

RESUMO

In the ongoing coronavirus diseases-2019 (COVID-19) crisis that caused immense suffering and deaths, the choice of therapy for the prevention and life-saving conditions must be based on sound scientific evidence. Uncertainty and apprehension are exacerbated in people using angiotensin-converting enzyme (ACE) inhibitors to control their comorbidities such as hypertension and diabetes. These drugs are reported to result in unfavorable outcome as they tend to increase the levels of ACE2 which mediates the entry of SARS-CoV-2. Amiloride, a prototypic inhibitor of epithelial sodium channels (ENaC) can be an ideal candidate for COVID-19 patients, given its ACE reducing and cytosolic pH increasing effects. Moreover, its potassium-sparing and anti-epileptic activities make it a promising alternative or a combinatorial agent.


Assuntos
Amilorida/farmacologia , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Bloqueadores do Canal de Sódio Epitelial/farmacologia , Pneumonia Viral/tratamento farmacológico , Mucosa Respiratória/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Células A549 , Betacoronavirus/patogenicidade , Doenças Cardiovasculares/tratamento farmacológico , Doenças Cardiovasculares/enzimologia , Infecções por Coronavirus/enzimologia , Infecções por Coronavirus/virologia , Regulação para Baixo , Interações Hospedeiro-Patógeno , Humanos , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/enzimologia , Pneumonia Viral/virologia , Receptores Virais/metabolismo , Mucosa Respiratória/enzimologia , Mucosa Respiratória/virologia
9.
Sci Rep ; 10(1): 16862, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033344

RESUMO

The prevalence of a novel ß-coronavirus (SARS-CoV-2) was declared as a public health emergency of international concern on 30 January 2020 and a global pandemic on 11 March 2020 by WHO. The spike glycoprotein of SARS-CoV-2 is regarded as a key target for the development of vaccines and therapeutic antibodies. In order to develop anti-viral therapeutics for SARS-CoV-2, it is crucial to find amino acid pairs that strongly attract each other at the interface of the spike glycoprotein and the human angiotensin-converting enzyme 2 (hACE2) complex. In order to find hot spot residues, the strongly attracting amino acid pairs at the protein-protein interaction (PPI) interface, we introduce a reliable inter-residue interaction energy calculation method, FMO-DFTB3/D/PCM/3D-SPIEs. In addition to the SARS-CoV-2 spike glycoprotein/hACE2 complex, the hot spot residues of SARS-CoV-1 spike glycoprotein/hACE2 complex, SARS-CoV-1 spike glycoprotein/antibody complex, and HCoV-NL63 spike glycoprotein/hACE2 complex were obtained using the same FMO method. Following this, a 3D-SPIEs-based interaction map was constructed with hot spot residues for the hACE2/SARS-CoV-1 spike glycoprotein, hACE2/HCoV-NL63 spike glycoprotein, and hACE2/SARS-CoV-2 spike glycoprotein complexes. Finally, the three 3D-SPIEs-based interaction maps were combined and analyzed to find the consensus hot spots among the three complexes. As a result of the analysis, two hot spots were identified between hACE2 and the three spike proteins. In particular, E37, K353, G354, and D355 of the hACE2 receptor strongly interact with the spike proteins of coronaviruses. The 3D-SPIEs-based map would provide valuable information to develop anti-viral therapeutics that inhibit PPIs between the spike protein of SARS-CoV-2 and hACE2.


Assuntos
Betacoronavirus/metabolismo , Biologia Computacional/métodos , Infecções por Coronavirus/epidemiologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/epidemiologia , Mapas de Interação de Proteínas , Glicoproteína da Espícula de Coronavírus/metabolismo , Anticorpos Antivirais/metabolismo , Sítios de Ligação , Infecções por Coronavirus/virologia , Coronavirus Humano NL63/metabolismo , Humanos , Pandemias , Pneumonia Viral/virologia , Prevalência , Domínios Proteicos , Receptores Virais/metabolismo , Vírus da SARS/metabolismo , Síndrome Respiratória Aguda Grave/virologia
10.
Signal Transduct Target Ther ; 5(1): 237, 2020 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-33051445

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an emerging virus that is highly pathogenic and has caused the recent worldwide pandemic officially named coronavirus disease (COVID-19). Currently, considerable efforts have been put into developing effective and safe drugs and vaccines against SARS-CoV-2. Vaccines, such as inactivated vaccines, nucleic acid-based vaccines, and vector vaccines, have already entered clinical trials. In this review, we provide an overview of the experimental and clinical data obtained from recent SARS-CoV-2 vaccines trials, and highlight certain potential safety issues that require consideration when developing vaccines. Furthermore, we summarize several strategies utilized in the development of vaccines against other infectious viruses, such as severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV), with the aim of aiding in the design of effective therapeutic approaches against SARS-CoV-2.


Assuntos
Anticorpos Antivirais/biossíntese , Infecções por Coronavirus/prevenção & controle , Pandemias/prevenção & controle , Peptidil Dipeptidase A/genética , Pneumonia Viral/prevenção & controle , Receptores Virais/genética , Vacinas Virais/biossíntese , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/imunologia , Betacoronavirus/patogenicidade , Ensaios Clínicos como Assunto , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Imunidade Inata/efeitos dos fármacos , Esquemas de Imunização , Imunogenicidade da Vacina , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Coronavírus da Síndrome Respiratória do Oriente Médio/patogenicidade , Segurança do Paciente , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Ligação Proteica , Receptores Virais/antagonistas & inibidores , Receptores Virais/metabolismo , Vírus da SARS/efeitos dos fármacos , Vírus da SARS/imunologia , Vírus da SARS/patogenicidade , Síndrome Respiratória Aguda Grave/imunologia , Síndrome Respiratória Aguda Grave/prevenção & controle , Síndrome Respiratória Aguda Grave/virologia , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/metabolismo , Vacinas Atenuadas , Vacinas de DNA , Vacinas de Subunidades , Vacinas de Partículas Semelhantes a Vírus , Vacinas Virais/administração & dosagem
11.
Mol Med Rep ; 22(5): 4221-4226, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33000221

RESUMO

Infection by the severe acute respiratory syndrome (SARS) coronavirus­2 (SARS­CoV­2) is the cause of the new viral infectious disease (coronavirus disease 2019; COVID­19). Emerging evidence indicates that COVID­19 may be associated with a wide spectrum of neurological symptoms and complications with central nervous system (CNS) involvement. It is now well­established that entry of SARS­CoV­2 into host cells is facilitated by its spike proteins mainly through binding to the angiotensin­converting enzyme 2 (ACE­2). Preclinical studies have suggested that neuropilin­1 (NRP1), which is a transmembrane receptor that lacks a cytosolic protein kinase domain and exhibits high expression in the respiratory and olfactory epithelium, may also be implicated in COVID­19 by enhancing the entry of SARS­CoV­2 into the brain through the olfactory epithelium. In the present study, we expand on these findings and demonstrate that the NRP1 is also expressed in the CNS, including olfactory­related regions such as the olfactory tubercles and paraolfactory gyri. This furthers supports the potential role of NRP1 as an additional SARS­CoV­2 infection mediator implicated in the neurologic manifestations of COVID­19. Accordingly, the neurotropism of SARS­CoV­2 via NRP1­expressing cells in the CNS merits further investigation.


Assuntos
Sistema Nervoso Central/metabolismo , Infecções por Coronavirus/metabolismo , Neuropilina-1/metabolismo , Pneumonia Viral/metabolismo , Receptores Virais/metabolismo , Betacoronavirus/fisiologia , Encéfalo/metabolismo , Encéfalo/virologia , Sistema Nervoso Central/virologia , Bases de Dados Genéticas , Humanos , Pandemias
12.
Signal Transduct Target Ther ; 5(1): 220, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-33024075
13.
Opt Lett ; 45(19): 5428-5431, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-33001920

RESUMO

We show that waveguide sensors can enable a quantitative characterization of coronavirus spike glycoprotein-host-receptor binding-the process whereby coronaviruses enter human cells, causing disease. We demonstrate that such sensors can help quantify and eventually understand kinetic and thermodynamic properties of viruses that control their affinity to targeted cells, which is known to significantly vary in the course of virus evolution, e.g., from SARS-CoV to SARS-CoV-2, making the development of virus-specific drugs and vaccine difficult. With the binding rate constants and thermodynamic parameters as suggested by the latest SARS-CoV-2 research, optical sensors of SARS-CoV-2 spike protein-receptor binding may be within sight.


Assuntos
Betacoronavirus , Técnicas Biossensoriais , Infecções por Coronavirus , Óptica e Fotônica/instrumentação , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral , Receptores Virais/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Sítios de Ligação , Humanos , Ligação Proteica/fisiologia
14.
Vet Q ; 40(1): 243-249, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32921279

RESUMO

Several cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection transmitted from human owners to their dogs have recently been reported. The first ever case of SARS-CoV-2 transmission from a human owner to a domestic cat was confirmed on March 27, 2020. A tiger from a zoo in New York, USA, was also reportedly infected with SARS-CoV-2. It is believed that SARS-CoV-2 was transmitted to tigers from their caretakers, who were previously infected with this virus. On May 25, 2020, the Dutch Minister of Agriculture, Nature and Food Quality reported that two employees were infected with SARS-CoV-2 transmitted from minks. These reports have influenced us to perform a comparative analysis among angiotensin-converting enzyme 2 (ACE2) homologous proteins for verifying the conservation of specific protein regions. One of the most conserved peptides is represented by the peptide "353-KGDFR-357 (H. sapiens ACE2 residue numbering), which is located on the surface of the ACE2 molecule and participates in the binding of SARS-CoV-2 spike receptor binding domain (RBD). Multiple sequence alignments of the ACE2 proteins by ClustalW, whereas the three-dimensional structure of its binding region for the spike glycoprotein of SARS-CoV-2 was assessed by means of Spanner, a structural homology modeling pipeline method. In addition, evolutionary phylogenetic tree analysis by ETE3 was used. ACE2 works as a receptor for the SARS-CoV-2 spike glycoprotein between humans, dogs, cats, tigers, minks, and other animals, except for snakes. The three-dimensional structure of the KGDFR hosting protein region involved in direct interactions with SARS-CoV-2 spike RBD of the mink ACE2 appears to form a loop structurally related to the human ACE2 corresponding protein loop, despite of the reduced available protein length (401 residues of the mink ACE2 available sequence vs 805 residues of the human ACE2). The multiple sequence alignments of the ACE2 proteins shows high homology and complete conservation of the five amino acid residues: 353-KGDFR-357 with humans, dogs, cats, tigers, minks, and other animals, except for snakes. Where the information revealed from our examinations can support precision vaccine design and the discovery of antiviral therapeutics, which will accelerate the development of medical countermeasures, the World Health Organization recently reported on the possible risks of reciprocal infections regarding SARS-CoV-2 transmission from animals to humans.


Assuntos
Betacoronavirus/metabolismo , Infecções por Coronavirus/transmissão , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/transmissão , Receptores Virais/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Sequência de Aminoácidos , Animais , Betacoronavirus/genética , Gatos , Infecções por Coronavirus/prevenção & controle , Cães , Humanos , Vison , Pandemias/prevenção & controle , Peptidil Dipeptidase A/química , Filogenia , Pneumonia Viral/prevenção & controle , Receptores Virais/química , Receptores Virais/genética , Alinhamento de Sequência , Glicoproteína da Espícula de Coronavírus/química , Tigres
15.
Emerg Microbes Infect ; 9(1): 2169-2179, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32969768

RESUMO

Studies on patients with the coronavirus disease-2019 (COVID-19) have implicated that the gastrointestinal (GI) tract is a major site of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. We established a human GI tract cell line model highly permissive to SARS-CoV-2. These cells, C2BBe1 intestinal cells with a brush border having high levels of transmembrane serine protease 2 (TMPRSS2), showed robust viral propagation, and could be persistently infected with SARS-CoV-2, supporting the clinical observations of persistent GI infection in COVID-19 patients. Ectopic expression of viral receptors revealed that the levels of angiotensin-converting enzyme 2 (ACE2) expression confer permissiveness to SARS-CoV-2 infection, and TMPRSS2 greatly facilitates ACE2-mediated SARS-CoV-2 dissemination. Interestingly, ACE2 but not TMPRSS2 expression was significantly promoted by enterocytic differentiation, suggesting that the state of enterocytic differentiation may serve as a determining factor for viral propagation. Thus, our study sheds light on the pathogenesis of SARS-CoV-2 in the GI tract.


Assuntos
Betacoronavirus/fisiologia , Infecções por Coronavirus/virologia , Mucosa Intestinal/virologia , Pneumonia Viral/virologia , Betacoronavirus/genética , Linhagem Celular , Infecções por Coronavirus/genética , Infecções por Coronavirus/metabolismo , Trato Gastrointestinal/metabolismo , Trato Gastrointestinal/virologia , Humanos , Mucosa Intestinal/metabolismo , Pandemias , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/genética , Pneumonia Viral/metabolismo , Receptores Virais/genética , Receptores Virais/metabolismo , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo
16.
PLoS Pathog ; 16(9): e1008828, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32991636

RESUMO

Field isolates of foot-and-mouth disease viruses (FMDVs) utilize integrin-mediated cell entry but many, including Southern African Territories (SAT) viruses, are difficult to adapt to BHK-21 cells, thus hampering large-scale propagation of vaccine antigen. However, FMDVs acquire the ability to bind to cell surface heparan sulphate proteoglycans, following serial cytolytic infections in cell culture, likely by the selection of rapidly replicating FMDV variants. In this study, fourteen SAT1 and SAT2 viruses, serially passaged in BHK-21 cells, were virulent in CHO-K1 cells and displayed enhanced affinity for heparan, as opposed to their low-passage counterparts. Comparative sequence analysis revealed the fixation of positively charged residues clustered close to the icosahedral 5-fold axes of the virus, at amino acid positions 83-85 in the ßD-ßE loop and 110-112 in the ßF-ßG loop of VP1 upon adaptation to cultured cells. Molecular docking simulations confirmed enhanced binding of heparan sulphate to a model of the adapted SAT1 virus, with the region around VP1 arginine 112 contributing the most to binding. Using this information, eight chimeric field strain mutant viruses were constructed with additional positive charges in repeated clusters on the virion surface. Five of these bound heparan sulphate with expanded cell tropism, which should facilitate large-scale propagation. However, only positively charged residues at position 110-112 of VP1 enhanced infectivity of BHK-21 cells. The symmetrical arrangement of even a single amino acid residue in the FMD virion is a powerful strategy enabling the virus to generate novel receptor binding and alternative host-cell interactions.


Assuntos
Vírus da Febre Aftosa/genética , Febre Aftosa/virologia , Modelos Moleculares , Vírion/metabolismo , Animais , Proteínas do Capsídeo/metabolismo , Cricetinae , Heparitina Sulfato/metabolismo , Simulação de Acoplamento Molecular/métodos , Receptores Virais/metabolismo
17.
Cell ; 183(3): 739-751.e8, 2020 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-32991842

RESUMO

The SARS-CoV-2 spike (S) protein variant D614G supplanted the ancestral virus worldwide, reaching near fixation in a matter of months. Here we show that D614G was more infectious than the ancestral form on human lung cells, colon cells, and on cells rendered permissive by ectopic expression of human ACE2 or of ACE2 orthologs from various mammals, including Chinese rufous horseshoe bat and Malayan pangolin. D614G did not alter S protein synthesis, processing, or incorporation into SARS-CoV-2 particles, but D614G affinity for ACE2 was reduced due to a faster dissociation rate. Assessment of the S protein trimer by cryo-electron microscopy showed that D614G disrupts an interprotomer contact and that the conformation is shifted toward an ACE2 binding-competent state, which is modeled to be on pathway for virion membrane fusion with target cells. Consistent with this more open conformation, neutralization potency of antibodies targeting the S protein receptor-binding domain was not attenuated.


Assuntos
Betacoronavirus/fisiologia , Betacoronavirus/ultraestrutura , Glicoproteína da Espícula de Coronavírus/fisiologia , Glicoproteína da Espícula de Coronavírus/ultraestrutura , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Betacoronavirus/patogenicidade , Células Cultivadas , Infecções por Coronavirus/virologia , Feminino , Variação Genética , Células HEK293 , Humanos , Masculino , Modelos Moleculares , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/virologia , Conformação Proteica , Processamento de Proteína Pós-Traducional , Receptores Virais/metabolismo , Especificidade da Espécie
18.
PLoS Pathog ; 16(9): e1008920, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32997730

RESUMO

The virions of enteroviruses such as poliovirus undergo a global conformational change after binding to the cellular receptor, characterized by a 4% expansion, and by the opening of holes at the two and quasi-three-fold symmetry axes of the capsid. The resultant particle is called a 135S particle or A-particle and is thought to be on the pathway to a productive infection. Previously published studies have concluded that the membrane-interactive peptides, namely VP4 and the N-terminus of VP1, are irreversibly externalized in the 135S particle. However, using established protocols to produce the 135S particle, and single particle cryo-electron microscopy methods, we have identified at least two unique states that we call the early and late 135S particle. Surprisingly, only in the "late" 135S particles have detectable levels of the VP1 N-terminus been trapped outside the capsid. Moreover, we observe a distinct density inside the capsid that can be accounted for by VP4 that remains associated with the genome. Taken together our results conclusively demonstrate that the 135S particle is not a unique conformation, but rather a family of conformations that could exist simultaneously.


Assuntos
Capsídeo/ultraestrutura , Poliomielite/metabolismo , RNA Viral/ultraestrutura , Vírion/ultraestrutura , Capsídeo/metabolismo , Proteínas do Capsídeo/metabolismo , Microscopia Crioeletrônica , Humanos , Modelos Moleculares , RNA Viral/metabolismo , Receptores Virais/metabolismo , Vírion/metabolismo , Internalização do Vírus
19.
Viruses ; 12(10)2020 09 26.
Artigo em Inglês | MEDLINE | ID: mdl-32993136

RESUMO

In a short time, the COVID-19 pandemic has left the world with over 25 million cases and staggering death tolls that are still rising. Treatments for SARS-CoV-2 infection are desperately needed as there are currently no approved drug therapies. With limited knowledge of viral mechanisms, a network controllability method of prioritizing existing drugs for repurposing efforts is optimal for quickly moving through the drug approval pipeline using limited, available, virus-specific data. Based on network topology and controllability, 16 proteins involved in translation, cellular transport, cellular stress, and host immune response are predicted as regulators of the SARS-CoV-2 infected cell. Of the 16, eight are prioritized as possible drug targets where two, PVR and SCARB1, are previously unexplored. Known compounds targeting these genes are suggested for viral inhibition study. Prioritized proteins in agreement with previous analysis and viral inhibition studies verify the ability of network controllability to predict biologically relevant candidates.


Assuntos
Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Reposicionamento de Medicamentos/métodos , Pneumonia Viral/tratamento farmacológico , Betacoronavirus/isolamento & purificação , Betacoronavirus/fisiologia , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Aprovação de Drogas , Sistemas de Liberação de Medicamentos , Interações Hospedeiro-Patógeno , Humanos , Pandemias , Pneumonia Viral/metabolismo , Pneumonia Viral/virologia , Mapas de Interação de Proteínas/efeitos dos fármacos , Receptores Virais/genética , Receptores Virais/metabolismo , Receptores Depuradores Classe B/metabolismo , Integração Viral
20.
OMICS ; 24(11): 634-644, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32940573

RESUMO

In the first quarter of the 21st century, we are already facing the third emergence of a coronavirus outbreak, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) responsible for the coronavirus disease 2019 (COVID-19) pandemic. Comparative genomics can inform a deeper understanding of the pathogenesis of COVID-19. Previous strains of coronavirus, SARS-CoV, and Middle-East respiratory syndrome-coronavirus (MERS-CoV), have been known to cause acute lung injuries in humans. SARS-CoV-2 shares genetic similarity with SARS-CoV with some modification in the S protein leading to their enhanced binding affinity toward the angiotensin-converting enzyme 2 (ACE2) receptors of human lung cells. This expert review examines the features of all three coronaviruses through a conceptual lens of comparative genomics. In particular, the life cycle of SARS-CoV-2 that enables its survival within the host is highlighted. Susceptibility of humans to coronavirus outbreaks in the 21st century calls for comparisons of the transmission history, hosts, reservoirs, and fatality rates of these viruses so that evidence-based and effective planetary health interventions can be devised to prevent future zoonotic outbreaks. Comparative genomics offers new insights on putative and novel viral targets with an eye to both therapeutic innovation and prevention. We conclude the expert review by (1) articulating the lessons learned so far, whereas the research is still being actively sought after in the field, and (2) the challenges and prospects in deciphering the linkages among multiomics biological variability and COVID-19 pathogenesis.


Assuntos
Betacoronavirus/patogenicidade , Infecções por Coronavirus/epidemiologia , Genômica/métodos , Pandemias , Pneumonia Viral/epidemiologia , Síndrome Respiratória Aguda Grave/epidemiologia , Animais , Betacoronavirus/genética , Quirópteros/virologia , Infecções por Coronavirus/mortalidade , Infecções por Coronavirus/virologia , Eutérios/virologia , Saúde Global/tendências , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Coronavírus da Síndrome Respiratória do Oriente Médio/patogenicidade , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/mortalidade , Pneumonia Viral/virologia , Ligação Proteica , Receptores Virais/genética , Receptores Virais/metabolismo , Vírus da SARS/genética , Vírus da SARS/patogenicidade , Síndrome Respiratória Aguda Grave/mortalidade , Síndrome Respiratória Aguda Grave/virologia , Índice de Gravidade de Doença , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/metabolismo , Análise de Sobrevida
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