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1.
Viruses ; 15(1)2023 Jan 12.
Статья в английский | MEDLINE | ID: mdl-36680253

Реферат

The coronavirus disease (COVID-19) is a pandemic that started in the City of Wuhan, Hubei Province, China, caused by the spread of coronavirus (SARS-CoV-2). Drug discovery teams around the globe are in a race to develop a medicine for its management. It takes time for a novel molecule to enter the market, and the ideal way is to exploit the already approved drugs and repurpose them therapeutically. We have attempted to screen selected molecules with an affinity towards multiple protein targets in COVID-19 using the Schrödinger suit for in silico predictions. The proteins selected were angiotensin-converting enzyme-2 (ACE2), main protease (MPro), and spike protein. The molecular docking, prime MM-GBSA, induced-fit docking (IFD), and molecular dynamics (MD) simulations were used to identify the most suitable molecule that forms a stable interaction with the selected viral proteins. The ligand-binding stability for the proteins PDB-IDs 1ZV8 (spike protein), 5R82 (Mpro), and 6M1D (ACE2), was in the order of nintedanib > quercetin, nintedanib > darunavir, nintedanib > baricitinib, respectively. The MM-GBSA, IFD, and MD simulation studies imply that the drug nintedanib has the highest binding stability among the shortlisted. Nintedanib, primarily used for idiopathic pulmonary fibrosis, can be considered for repurposing for us against COVID-19.


Тема - темы
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/metabolism , Molecular Docking Simulation , Angiotensin-Converting Enzyme 2/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Molecular Dynamics Simulation , Antiviral Agents/therapeutic use , Antiviral Agents/chemistry , Drug Repositioning
2.
Indian J Pathol Microbiol ; 66(1): 19-23, 2023.
Статья в английский | MEDLINE | ID: mdl-36656205

Реферат

Context: Researchers throughout the world devote enormous efforts to reveal the peculiarities of the pathogenesis of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus, however, it continues to surprise and cause the death of millions of people. Aims: This article aims to study the molecular mechanisms provoked by SARS-CoV-2, the virus-induced changes in Angiotensin-converting enzyme 2 (ACE2) functionality, in the vascular homeostasis through CD34 expression, B-cell immunity through the expression of CD20 and CD79α, and adhesion molecules through E-cadherin. Settings and Design: This was a prospective, descriptive, and observational study. Methods and Material: A total of 15 autopsies of patients deceased by COVID-19 infection, confirmed by PCR, were performed. The lungs of all patients were examined histologically and immunohistochemically for ACE2, E-cadherin, CD34, CD20, and CD79α. Results: Immunohistological analysis showed increased ACE2 expression in all lung autopsy material affected by COVID-19 infection and we found a higher intensity of ACE2 expression than that of a healthy lung. CD20 examination reveals total deficiency of B-cells in the pulmonary parenchyma and CD79α is also absent. E-Cadherin is not expressed in the basal cellular sections where the contact elements are missing. CD34 demonstrates a desquamation of the endothelial cells, which indicates a direct damage of the vascular walls. Conclusions: We found that patients who died after severe COVID-19 had high immune deficiency and impaired intercellular communication in the parenchyma and endothelium of lung tissue, leading to severe thromboembolic complications in patients with multiple diseases.


Тема - темы
COVID-19 , SARS-CoV-2 , Humans , COVID-19/pathology , Angiotensin-Converting Enzyme 2/metabolism , Autopsy , Endothelial Cells , Prospective Studies , Peptidyl-Dipeptidase A/metabolism , Lung/pathology , Cadherins/metabolism
3.
Sci Rep ; 13(1): 881, 2023 Jan 17.
Статья в английский | MEDLINE | ID: mdl-36650223

Реферат

We recently reported exacerbated endotoxic signs of neuroinflammation and autonomic defects in offspring of preeclamptic (PE) dams. Here, we investigated whether PE programming similarly modifies hemodynamic and renal vasoconstrictor responsiveness to endotoxemia in PE offspring and whether this interaction is modulated by gestational angiotensin 1-7 (Ang1-7). Preeclampsia was induced by gestational treatment with L-NAME. Adult offspring was challenged with lipopolysaccharides (LPS, 5 mg/kg) and systolic blood pressure (SBP) and renal vasoconstrictions were assessed 4 h later. Male, but not female, offspring of PE rats exhibited SBP elevations that were blunted by LPS. Renal vasoconstrictions induced by angiotensin II (Ang II), but not phenylephrine, were intensified in perfused kidneys of either sex. LPS blunted the heightened Ang II responses in male, but not female, kidneys. While renal expressions of AT1-receptors and angiotensin converting enzyme (ACE) were increased in PE offspring of both sexes, ACE2 was upregulated in female offspring only. These molecular effects were diminished by LPS in male offspring. Gestational Ang1-7 caused sex-unrelated attenuation of phenylephrine vasoconstrictions and preferentially downregulated Ang II responses and AT1-receptor and nuclear factor-kB (NFkB) expressions in females. Together, endotoxemia and Ang1-7 offset in sexually-related manners imbalances in renal vasoconstriction and AT1/ACE/ACE2 signaling in PE offspring.


Тема - темы
Endotoxemia , Pre-Eclampsia , Animals , Female , Male , Rats , Angiotensin II/metabolism , Angiotensin-Converting Enzyme 2/metabolism , Endotoxemia/chemically induced , Endotoxemia/metabolism , Endotoxins/metabolism , Kidney/metabolism , Lipopolysaccharides/pharmacology , Pre-Eclampsia/metabolism , Renin-Angiotensin System , Vasoconstriction
4.
Nat Commun ; 14(1): 132, 2023 Jan 10.
Статья в английский | MEDLINE | ID: mdl-36627352

Реферат

As an inherited disorder characterized by severe pulmonary disease, cystic fibrosis could be considered a comorbidity for coronavirus disease 2019. Instead, current clinical evidence seems to be heading in the opposite direction. To clarify whether host factors expressed by the Cystic Fibrosis epithelia may influence coronavirus disease 2019 progression, here we describe the expression of SARS-CoV-2 receptors in primary airway epithelial cells. We show that angiotensin converting enzyme 2 (ACE2) expression and localization are regulated by Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) channel. Consistently, our results indicate that dysfunctional CFTR channels alter susceptibility to SARS-CoV-2 infection, resulting in reduced viral entry and replication in Cystic Fibrosis cells. Depending on the pattern of ACE2 expression, the SARS-CoV-2 spike (S) protein induced high levels of Interleukin 6 in healthy donor-derived primary airway epithelial cells, but a very weak response in primary Cystic Fibrosis cells. Collectively, these data support that Cystic Fibrosis condition may be at least partially protecting from SARS-CoV-2 infection.


Тема - темы
Angiotensin-Converting Enzyme 2 , COVID-19 , Cystic Fibrosis , SARS-CoV-2 , Virus Internalization , Humans , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Cystic Fibrosis/genetics , Cystic Fibrosis Transmembrane Conductance Regulator/genetics , Cystic Fibrosis Transmembrane Conductance Regulator/metabolism , Down-Regulation , Receptors, Virus/genetics , Receptors, Virus/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/metabolism , Virus Replication
5.
Int J Mol Sci ; 24(1)2022 Dec 21.
Статья в английский | MEDLINE | ID: mdl-36613540

Реферат

Myocardial damage caused by the newly emerged coronavirus (SARS-CoV-2) infection is one of the key determinants of COVID-19 severity and mortality. SARS-CoV-2 entry to host cells is initiated by binding with its receptor, angiotensin-converting enzyme (ACE) 2, and the ACE2 abundance is thought to reflect the susceptibility to infection. Here, we report that ibudilast, which we previously identified as a potent inhibitor of protein complex between transient receptor potential canonical (TRPC) 3 and NADPH oxidase (Nox) 2, attenuates the SARS-CoV-2 spike glycoprotein pseudovirus-evoked contractile and metabolic dysfunctions of neonatal rat cardiomyocytes (NRCMs). Epidemiologically reported risk factors of severe COVID-19, including cigarette sidestream smoke (CSS) and anti-cancer drug treatment, commonly upregulate ACE2 expression level, and these were suppressed by inhibiting TRPC3-Nox2 complex formation. Exposure of NRCMs to SARS-CoV-2 pseudovirus, as well as CSS and doxorubicin (Dox), induces ATP release through pannexin-1 hemi-channels, and this ATP release potentiates pseudovirus entry to NRCMs and human iPS cell-derived cardiomyocytes (hiPS-CMs). As the pseudovirus entry followed by production of reactive oxygen species was attenuated by inhibiting TRPC3-Nox2 complex in hiPS-CMs, we suggest that TRPC3-Nox2 complex formation triggered by panexin1-mediated ATP release participates in exacerbation of myocardial damage by amplifying ACE2-dependent SARS-CoV-2 entry.


Тема - темы
COVID-19 , NADPH Oxidase 2 , TRPC Cation Channels , Animals , Humans , Rats , Adenosine Triphosphate/metabolism , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/metabolism , Myocytes, Cardiac/metabolism , NADPH Oxidase 2/metabolism , Protein Binding , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Up-Regulation , TRPC Cation Channels/metabolism
6.
Cells ; 12(1)2022 Dec 25.
Статья в английский | MEDLINE | ID: mdl-36611877

Реферат

Protein microarray screenings identified fungal natural products from the azaphilone family as potent inhibitors of SARS-CoV-2 spike protein binding to host ACE2 receptors. Cohaerin F, as the most potent substance from the cohaerin group, led to more than 50% less binding of ACE2 and SARS-CoV-2 spike protein. A survey for structurally related azaphilones yielded the structure elucidation of six new multiformins E-J (10-15) and the revision of the stereochemistry of the multiformins. Cohaerin and multiformin azaphilones (1-5, 8, 12) were assessed for their activity in a cell-based infection assay. Calu-3 cells expressing human ACE2 receptor showed more than 75% and 50% less infection by SARS-CoV-2 pseudotyped lentivirus particles after treatment with cohaerin C (1) and cohaerin F (4), respectively. Multiformin C (8) and G (12) that nearly abolished the infection of cells. Our data show that multiformin-type azaphilones prevent the binding of SARS-CoV-2 to the cell entry receptor ACE2.


Тема - темы
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/metabolism , Angiotensin-Converting Enzyme 2/metabolism , Protein Binding
7.
Kidney360 ; 3(12): 2086-2094, 2022 12 29.
Статья в английский | MEDLINE | ID: mdl-36591353

Реферат

Background: ACE2 is a key enzyme in the renin-angiotensin system (RAS) capable of balancing the RAS by metabolizing angiotensin II (AngII). First described in cardiac tissue, abundance of ACE2 is highest in the kidney, and it is also expressed in several extrarenal tissues. Previously, we reported an association between enhanced susceptibility to hypertension and elevated renal AngII levels in global ACE2-knockout mice. Methods: To examine the effect of ACE2 expressed in the kidney, relative to extrarenal expression, on the development of hypertension, we used a kidney crosstransplantation strategy with ACE2-KO and WT mice. In this model, both native kidneys are removed and renal function is provided entirely by the transplanted kidney, such that four experimental groups with restricted ACE2 expression are generated: WT→WT (WT), KO→WT (KidneyKO), WT→KO (SystemicKO), and KO→KO (TotalKO). Additionally, we used nanoscale mass spectrometry-based proteomics to identify ACE2 fragments in early glomerular filtrate of mice. Results: Although significant differences in BP were not detected, a major finding of our study is that shed or soluble ACE2 (sACE2) was present in urine of KidneyKO mice that lack renal ACE2 expression. Detection of sACE2 in the urine of KidneyKO mice during AngII-mediated hypertension suggests that sACE2 originating from extrarenal tissues can reach the kidney and be excreted in urine. To confirm glomerular filtration of ACE2, we used micropuncture and nanoscale proteomics to detect peptides derived from ACE2 in the Bowman's space. Conclusions: Our findings suggest that both systemic and renal tissues may contribute to sACE2 in urine, identifying the kidney as a major site for ACE2 actions. Moreover, filtration of sACE2 into the lumen of the nephron may contribute to the pathophysiology of kidney diseases characterized by disruption of the glomerular filtration barrier.


Тема - темы
Angiotensin-Converting Enzyme 2 , Hypertension , Kidney , Renin-Angiotensin System , Animals , Mice , Angiotensin II/metabolism , Angiotensin II/pharmacology , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Hypertension/genetics , Hypertension/metabolism , Kidney/metabolism , Mice, Knockout , Peptidyl-Dipeptidase A/genetics , Peptidyl-Dipeptidase A/metabolism , Peptidyl-Dipeptidase A/pharmacology , Renin-Angiotensin System/genetics , Renin-Angiotensin System/physiology
8.
Egypt J Immunol ; 30(1): 73-86, 2023 Jan.
Статья в английский | MEDLINE | ID: mdl-36592120

Реферат

The worldwide medical systems are still being severely impacted by the coronavirus disease-2019 (COVID-19) pandemic, which is responsible for catastrophic mortality and morbidity. It becomes more and more obvious that this unique respiratory virus's impacts go beyond the respiratory system as time goes on and our comprehension of it deepens. The transmembrane serine protease 2 (TMPRSS2) protein is necessary for the severe acute respiratory syndrome coronavirus 2, which is the cause of COVID-19, to gain cellular entry through the angiotensin-converting enzyme 2 (ACE2) receptor. Most endocrine glands exhibit high levels of expression for ACE2 and TMPRSS2. This pays the attention to the effect of COVID-19 on the endocrine system. Besides its capability to pass to the central nervous system especially the hypothalamus inducing a lot of functional disorders in COVID-19 individuals. Although effective vaccines became widely available, and mortality declined but attention is shifting more and more to the lengthy health impacts on COVID-19 survivors. To inform suitable research and effective management, this review provides an overview of the data examining the impacts of COVID-19 on the endocrine glands besides the hypothalamus. In addition, we reported if the endocrinal and thalamic disorders could affect the incidence and progress of COVID-19.


Тема - темы
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/metabolism , Angiotensin-Converting Enzyme 2/metabolism , Peptidyl-Dipeptidase A , Hypothalamus/metabolism
9.
Eur Rev Med Pharmacol Sci ; 26(24): 9230-9239, 2022 12.
Статья в английский | MEDLINE | ID: mdl-36591835

Реферат

OBJECTIVE: Serum ACE2 level in the acute phase of ST-segment elevation myocardial infarction may be an indicator of heart failure, however, limited studies have reported conflicting results. Therefore, in our study, we aimed to evaluate the relationship between serum ACE2 level and infarct size in the acute phase of ST-segment elevation myocardial infarction and compare the predictive value of ACE2 level with classical biomarkers. PATIENTS AND METHODS: Sixty-six patients after the primary percutaneous coronary intervention were included in the study. For the measurement of serum ACE2 levels, blood samples were taken twice from the patients: in the first 24 hours and on the 5th day of the infarction, and once from 30 healthy volunteers. hs-cTnT, BNP, and CRP levels were measured daily, and their peak values were taken. On the 7th day of ST-segment elevation myocardial infarction, gSPECT was used with the 99mTc-MIBI method for assessment of infarct size. RESULTS: Baseline ACE2 values were found to be higher in patients compared to controls, and ACE2 values obtained on the 5th day were found to be higher than the baseline values in the patients. There was no significant correlation between serum ACE2 levels and the RSS (%), while peak levels of hs-cTnT, BNP, and CRP were assessed as predictive factors for the RSS (%). CONCLUSIONS: Although serum ACE2 levels increased in the acute phase of ST-segment elevation myocardial infarction, this increase was not associated with infarct size. Serum ACE2 level did not provide additional benefit to classical biomarkers for infarct size-related prognosis prediction.


Тема - темы
Angiotensin-Converting Enzyme 2 , ST Elevation Myocardial Infarction , Humans , Angiotensin-Converting Enzyme 2/blood , Angiotensin-Converting Enzyme 2/metabolism , Biomarkers/blood , Biomarkers/metabolism , Percutaneous Coronary Intervention , Prognosis , ST Elevation Myocardial Infarction/blood , ST Elevation Myocardial Infarction/diagnosis , ST Elevation Myocardial Infarction/metabolism , ST Elevation Myocardial Infarction/therapy , Treatment Outcome
10.
FASEB J ; 37(2): e22741, 2023 02.
Статья в английский | MEDLINE | ID: mdl-36583713

Реферат

The SARS-CoV-2 life cycle is strictly dependent on the environmental redox state that influences both virus entry and replication. A reducing environment impairs the binding of the spike protein (S) to the angiotensin-converting enzyme 2 receptor (ACE2), while a highly oxidizing environment is thought to favor S interaction with ACE2. Moreover, SARS-CoV-2 interferes with redox homeostasis in infected cells to promote the oxidative folding of its own proteins. Here we demonstrate that synthetic low molecular weight (LMW) monothiol and dithiol compounds induce a redox switch in the S protein receptor binding domain (RBD) toward a more reduced state. Reactive cysteine residue profiling revealed that all the disulfides present in RBD are targets of the thiol compounds. The reduction of disulfides in RBD decreases the binding to ACE2 in a cell-free system as demonstrated by enzyme-linked immunosorbent and surface plasmon resonance (SPR) assays. Moreover, LMW thiols interfere with protein oxidative folding and the production of newly synthesized polypeptides in HEK293 cells expressing the S1 and RBD domain, respectively. Based on these results, we hypothesize that these thiol compounds impair both the binding of S protein to its cellular receptor during the early stage of viral infection, as well as viral protein folding/maturation and thus the formation of new viral mature particles. Indeed, all the tested molecules, although at different concentrations, efficiently inhibit both SARS-CoV-2 entry and replication in Vero E6 cells. LMW thiols may represent innovative anti-SARS-CoV-2 therapeutics acting directly on viral targets and indirectly by inhibiting cellular functions mandatory for viral replication.


Тема - темы
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/metabolism , Angiotensin-Converting Enzyme 2/metabolism , Viral Proteins/metabolism , HEK293 Cells , Protein Binding , Sulfhydryl Compounds/pharmacology
11.
Virol J ; 19(1): 227, 2022 12 30.
Статья в английский | MEDLINE | ID: mdl-36581924

Реферат

BACKGROUND: The paucity of SARS-CoV-2-specific virulence factors has greatly hampered the therapeutic management of patients with COVID-19 disease. Although available vaccines and approved therapies have shown tremendous benefits, the continuous emergence of new variants of SARS-CoV-2 and side effects of existing treatments continue to challenge therapy, necessitating the development of a novel effective therapy. We have previously shown that our developed novel single-stranded DNA aptamers not only target the trimer S protein of SARS-CoV-2, but also block the interaction between ACE2 receptors and trimer S protein of Wuhan origin, Delta, Delta plus, Alpha, Lambda, Mu, and Omicron variants of SARS-CoV-2. We herein performed in vivo experiments that administer the aptamer to the lungs by intubation as well as in vitro studies utilizing PBMCs to prove the efficacy and safety of our most effective aptamer, AYA2012004_L. METHODS: In vivo studies were conducted in transgenic mice expressing human ACE2 (K18hACE2), C57BL/6J, and Balb/cJ. Flow cytometry was used to check S-protein expressing pseudo-virus-like particles (VLP) uptake by the lung cells and test the immuogenicity of AYA2012004_L. Ames test was used to assess mutagenicity of AYA2012004_L. RT-PCR and histopathology were used to determine the biodistribution and toxicity of AYA2012004_L in vital organs of mice. RESULTS: We measured the in vivo uptake of VLPs by lung cells by detecting GFP signal using flow cytometry. AYA2012004_L specifically neutralized VLP uptake and also showed no inflammatory response in mice lungs. In addition, AYA2012004_L did not induce inflammatory response in the lungs of Th1 and Th2 mouse models as well as human PBMCs. AYA2012004_L was detectable in mice lungs and noticeable in insignificant amounts in other vital organs. Accumulation of AYA2012004_L in organs decreased over time. AYA2012004_L did not induce degenerative signs in tissues as seen by histopathology and did not cause changes in the body weight of mice. Ames test also certified that AYA2012004_L is non-mutagenic and proved it to be safe for in vivo studies. CONCLUSIONS: Our aptamer is safe, effective, and can neutralize the uptake of VLPs by lung cells when administered locally suggesting that it can be used as a potential therapeutic agent for COVID-19 management.


Тема - темы
Aptamers, Nucleotide , COVID-19 , Humans , Mice , Animals , COVID-19/therapy , SARS-CoV-2/genetics , Aptamers, Nucleotide/therapeutic use , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Tissue Distribution , Antibodies, Viral , Mice, Inbred C57BL , Mice, Transgenic , Spike Glycoprotein, Coronavirus/genetics , Antibodies, Neutralizing
12.
Nat Commun ; 13(1): 7926, 2022 12 24.
Статья в английский | MEDLINE | ID: mdl-36566234

Реферат

Recent waves of COVID-19 correlate with the emergence of the Delta and the Omicron variant. We report that the Spike trimer acts as a highly dynamic molecular caliper, thereby forming up to three tight bonds through its RBDs with ACE2 expressed on the cell surface. The Spike of both Delta and Omicron (B.1.1.529) Variant enhance and markedly prolong viral attachment to the host cell receptor ACE2, as opposed to the early Wuhan-1 isolate. Delta Spike shows rapid binding of all three Spike RBDs to three different ACE2 molecules with considerably increased bond lifetime when compared to the reference strain, thereby significantly amplifying avidity. Intriguingly, Omicron (B.1.1.529) Spike displays less multivalent bindings to ACE2 molecules, yet with a ten time longer bond lifetime than Delta. Delta and Omicron (B.1.1.529) Spike variants enhance and prolong viral attachment to the host, which likely not only increases the rate of viral uptake, but also enhances the resistance of the variants against host-cell detachment by shear forces such as airflow, mucus or blood flow. We uncover distinct binding mechanisms and strategies at single-molecule resolution, employed by circulating SARS-CoV-2 variants to enhance infectivity and viral transmission.


Тема - темы
Angiotensin-Converting Enzyme 2 , COVID-19 , SARS-CoV-2 , Single Molecule Imaging , Spike Glycoprotein, Coronavirus , Humans , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/virology , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/metabolism , Virus Attachment
13.
Biomacromolecules ; 24(1): 141-149, 2023 Jan 09.
Статья в английский | MEDLINE | ID: mdl-36562668

Реферат

The coronavirus disease 2019 (COVID-19) pandemic has threatened the stability of global healthcare, which is becoming an endemic issue. Despite the development of various treatment strategies to fight COVID-19, the currently available treatment options have shown varied efficacy. Herein, we have developed an avidity-based SARS-CoV-2 antagonist using dendrimer-peptide conjugates (DPCs) for effective COVID-19 treatment. Two different peptide fragments obtained from angiotensin-converting enzyme 2 (ACE2) were integrated into a single sequence, followed by the conjugation to poly(amidoamine) (PAMAM) dendrimers. We hypothesized that the strong multivalent binding avidity endowed by dendrimers would help peptides effectively block the interaction between SARS-CoV-2 and ACE2, and this antagonist effect would be dependent upon the generation (size) of the dendrimers. To assess this, binding kinetics of the DPCs prepared from generation 4 (G4) and G7 PAMAM dendrimers to spike protein of SARS-CoV-2 were quantitatively measured using surface plasmon resonance. The larger dendrimer-based DPCs exhibited significantly enhanced binding strength by 3 orders of magnitude compared to the free peptides, whereas the smaller one showed a 12.8-fold increase only. An in vitro assay using SARS-CoV-2-mimicking microbeads also showed the improved SARS-CoV-2 blockade efficiency of the G7-peptide conjugates compared to G4. In addition, the interaction between the DPCs and SARS-CoV-2 was analyzed using molecular dynamics (MD) simulation, providing an insight into how the dendrimer-mediated multivalent binding effect can enhance the SARS-CoV-2 blockade. Our findings demonstrate that the DPCs having strong binding to SARS-CoV-2 effectively block the interaction between ACE2 and SARS-CoV-2, providing a potential as a high-affinity drug delivery system to direct anti-COVID payloads to the virus.


Тема - темы
COVID-19 , Dendrimers , Humans , Angiotensin-Converting Enzyme 2/metabolism , Dendrimers/pharmacology , Peptides/pharmacology , Peptides/metabolism , Peptidyl-Dipeptidase A/chemistry , Peptidyl-Dipeptidase A/metabolism , Protein Binding , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
14.
Viruses ; 14(12)2022 Dec 09.
Статья в английский | MEDLINE | ID: mdl-36560757

Реферат

Because of the interface between coagulation and the immune response, it is expected that COVID-19-associated coagulopathy occurs via activated protein C signaling. The objective was to explore putative changes in the expression of the protein C signaling network in the liver, peripheral blood mononuclear cells, and nasal epithelium of patients with COVID-19. Single-cell RNA-sequencing data from patients with COVID-19 and healthy subjects were obtained from the COVID-19 Cell Atlas database. A functional protein-protein interaction network was constructed for the protein C gene. Patients with COVID-19 showed downregulation of protein C and components of the downstream protein C signaling cascade. The percentage of hepatocytes expressing protein C was lower. Part of the liver cell clusters expressing protein C presented increased expression of ACE2. In PBMC, there was increased ACE2, inflammatory, and pro-coagulation transcripts. In the nasal epithelium, PROC, ACE2, and PROS1 were expressed by the ciliated cell cluster, revealing co-expression of ACE-2 with transcripts encoding proteins belonging to the coagulation and immune system interface. Finally, there was upregulation of coagulation factor 3 transcript in the liver and PBMC. Protein C could play a mechanistic role in the hypercoagulability syndrome affecting patients with severe COVID-19.


Тема - темы
COVID-19 , Thrombophilia , Humans , COVID-19/genetics , Leukocytes, Mononuclear/metabolism , SARS-CoV-2/genetics , Protein C/genetics , Protein C/metabolism , Down-Regulation , Transcriptome , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Peptidyl-Dipeptidase A/metabolism , Thrombophilia/genetics
15.
Viruses ; 14(12)2022 Dec 14.
Статья в английский | MEDLINE | ID: mdl-36560795

Реферат

The spike protein is key to SARS-CoV-2 high infectivity because it facilitates the receptor binding domain (RBD) encounter with ACE2. As targeting subunit S1 has not yet delivered an ACE2-binding inhibitor, we have assessed the druggability of the conserved segment of the spike protein stalk within subunit S2 by means of an integrated computational approach that combines the molecular docking of an optimized library of fragments with high-throughput molecular dynamics simulations. The high propensity of the spike protein to mutate in key regions that are responsible for the recognition of the human angiotensin-converting enzyme 2 (hACE2) or for the recognition of antibodies, has made subunit S1 of the spike protein difficult to target. Despite the inherent flexibility of the stalk region, our results suggest two hidden interhelical binding sites, whose accessibility is only partially hampered by glycan residues.


Тема - темы
COVID-19 , Humans , Spike Glycoprotein, Coronavirus/metabolism , SARS-CoV-2/metabolism , Angiotensin-Converting Enzyme 2/metabolism , Molecular Docking Simulation , Protein Domains , Protein Binding , Molecular Dynamics Simulation
16.
Nat Commun ; 13(1): 7907, 2022 12 23.
Статья в английский | MEDLINE | ID: mdl-36564389

Реферат

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic. Angiotensin-converting enzyme 2 (ACE2) is an entry receptor for SARS-CoV-2. The full-length membrane form of ACE2 (memACE2) undergoes ectodomain shedding to generate a shed soluble form (solACE2) that mediates SARS-CoV-2 entry via receptor-mediated endocytosis. Currently, it is not known how the physiological regulation of ACE2 shedding contributes to the etiology of COVID-19 in vivo. The present study identifies Membrane-type 1 Matrix Metalloproteinase (MT1-MMP) as a critical host protease for solACE2-mediated SARS-CoV-2 infection. SARS-CoV-2 infection leads to increased activation of MT1-MMP that is colocalized with ACE2 in human lung epithelium. Mechanistically, MT1-MMP directly cleaves memACE2 at M706-S to release solACE218-706 that binds to the SARS-CoV-2 spike proteins (S), thus facilitating cell entry of SARS-CoV-2. Human solACE218-706 enables SARS-CoV-2 infection in both non-permissive cells and naturally insusceptible C57BL/6 mice. Inhibition of MT1-MMP activities suppresses solACE2-directed entry of SARS-CoV-2 in human organoids and aged mice. Both solACE2 and circulating MT1-MMP are positively correlated in plasma of aged mice and humans. Our findings provide in vivo evidence demonstrating the contribution of ACE2 shedding to the etiology of COVID-19.


Тема - темы
Angiotensin-Converting Enzyme 2 , COVID-19 , Host-Pathogen Interactions , Matrix Metalloproteinase 14 , SARS-CoV-2 , Animals , Humans , Mice , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/metabolism , COVID-19/virology , Mice, Inbred C57BL , Peptidyl-Dipeptidase A/metabolism , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism
17.
Biomolecules ; 12(12)2022 11 23.
Статья в английский | MEDLINE | ID: mdl-36551170

Реферат

Since its first appearance in April 2021, B.1.617.2, also termed variant Delta, catalyzed one major worldwide wave dominating the second year of coronavirus disease 2019 (COVID-19) pandemic. Despite its quick disappearance worldwide, the strong virulence caused by a few point mutations remains an unsolved problem largely. Along with the other two sublineages, the Delta variant harbors an accumulation of Spike protein mutations, including the previously identified L452R, E484Q, and the newly emerged T478K on its receptor binding domain (RBD). We used molecular dynamics (MD) simulations, in combination with free energy perturbation (FEP) calculations, to examine the effects of two combinative mutation sets, L452R + E484Q and L452R + T478K. Our dynamic trajectories reveal an enhancement in binding affinity between mutated RBD and the common receptor protein angiotensin converting enzyme 2 (ACE2) through a net increase in the buried molecular surface area of the binary complex. This enhanced binding, mediated through Gln493, sets the same stage for all three sublineages due to the presence of L452R mutation. The other mutation component, E484Q or T478K, was found to impact the RBD-ACE2 binding and help the variant to evade several monoclonal antibodies (mAbs) in a distinct manner. Especially for L452R + T478K, synergies between mutations are mediated through a complex residual and water interaction network and further enhance its binding to ACE2. Taking together, this study demonstrates that new variants of SARS-CoV-2 accomplish both "attack" (infection) and "defense" (antibody neutralization escape) with the same "polished sword" (mutated Spike RBD).


Тема - темы
Angiotensin-Converting Enzyme 2 , COVID-19 , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Humans , Angiotensin-Converting Enzyme 2/metabolism , Antibodies, Monoclonal/immunology , COVID-19/genetics , COVID-19/immunology , Protein Binding/genetics , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , Virulence/genetics , Point Mutation , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Allosteric Regulation , Protein Domains/genetics
18.
Nanoscale ; 15(3): 1076-1085, 2023 Jan 19.
Статья в английский | MEDLINE | ID: mdl-36546457

Реферат

COVID-19, caused by the severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2), originated a global health crisis, causing over 2 million casualties and altering human daily life all over the world. This pandemic emergency revealed the limitations of current diagnostic tests, highlighting the urgency to develop faster, more precise and sensitive sensors. Graphene field effect transistors (GFET) are analytical platforms that enclose all these requirements. However, the design of a sensitive and robust GFET is not a straightforward objective. In this work, we report a GFET array biosensor for the detection of SARS-CoV-2 spike protein using the human membrane protein involved in the virus internalisation: angiotensin-converting enzyme 2 (ACE2). By finely controlling the graphene functionalisation, by tuning the Debye length, and by deeply characterising the ACE2-spike protein interactions, we have been able to detect the target protein with an extremely low limit of detection (2.94 aM). This work set the basis for a new class of analytical platforms, based on human membrane proteins, with the potential to detect a broad variety of pathogens, even before their isolation, being a powerful tool in the fight against future pandemics.


Тема - темы
COVID-19 , Graphite , Humans , COVID-19/diagnosis , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/metabolism , Angiotensin-Converting Enzyme 2/metabolism , Protein Binding
19.
ACS Appl Bio Mater ; 6(1): 182-190, 2023 Jan 16.
Статья в английский | MEDLINE | ID: mdl-36550079

Реферат

SARS-CoV-2 has been shown to enter and infect human cells via interactions between spike protein (S glycoprotein) and angiotensin-converting enzyme 2 (ACE2). As such, it may be possible to suppress the infection of the virus via the blocking of this binding interaction through the use of specific peptides that can mimic the human ACE 2 peptidase domain (PD) α 1-helix. Herein, we report the use of competitive assays along with surface plasmon resonance (SPR) to investigate the effect of peptide sequence and length on spike protein inhibition. The characterization of these binding interactions helps us understand the mechanisms behind peptide-based viral blockage and develop SPR methodologies to quickly screen disease inhibitors. This work not only helps further our understanding of the important biological interactions involved in viral inhibition but will also aid in future studies that focus on the development of therapeutics and drug options. Two peptides of different sequence lengths, [30-42] and [22-44], based on the α 1-helix of ACE2 PD were selected for this fundamental investigation. In addition to characterizing their inhibitory behavior, we also identified the critical amino acid residues of the RBD/ACE2-derived peptides by combining experimental results and molecular docking modeling. While both investigated peptides were found to effectively block the RBD residues known to bind to ACE2 PD, our investigation showed that the shorter peptide was able to reach a maximal inhibition at lower concentrations. These inhibition results matched with molecular docking models and indicated that peptide length and composition are key in the development of an effective peptide for inhibiting biophysical interactions. The work presented here emphasizes the importance of inhibition screening and modeling, as longer peptides are not always more effective.


Тема - темы
COVID-19 , SARS-CoV-2 , Humans , Surface Plasmon Resonance , Angiotensin-Converting Enzyme 2/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Molecular Docking Simulation , Peptides/pharmacology
20.
PLoS One ; 17(12): e0276115, 2022.
Статья в английский | MEDLINE | ID: mdl-36538516

Реферат

Human-based organ models can provide strong predictive value to investigate the tropism, virulence, and replication kinetics of viral pathogens. Currently, such models have received widespread attention in the study of SARS-CoV-2 causing the COVID-19 pandemic. Applicable to a large set of organoid models and viruses, we provide a step-by-step work instruction for the infection of human alveolar-like organoids with SARS-CoV-2 in this protocol collection. We also prepared a detailed description on state-of-the-art methodologies to assess the infection impact and the analysis of relevant host factors in organoids. This protocol collection consists of five different sets of protocols. Set 1 describes the protein extraction from human alveolar-like organoids and the determination of protein expression of angiotensin-converting enzyme 2 (ACE2), transmembrane serine protease 2 (TMPRSS2) and FURIN as exemplary host factors of SARS-CoV-2. Set 2 provides detailed guidance on the extraction of RNA from human alveolar-like organoids and the subsequent qPCR to quantify the expression level of ACE2, TMPRSS2, and FURIN as host factors of SARS-CoV-2 on the mRNA level. Protocol set 3 contains an in-depth explanation on how to infect human alveolar-like organoids with SARS-CoV-2 and how to quantify the viral replication by plaque assay and viral E gene-based RT-qPCR. Set 4 provides a step-by-step protocol for the isolation of single cells from infected human alveolar-like organoids for further processing in single-cell RNA sequencing or flow cytometry. Set 5 presents a detailed protocol on how to perform the fixation of human alveolar-like organoids and guides through all steps of immunohistochemistry and in situ hybridization to visualize SARS-CoV-2 and its host factors. The infection and all subsequent analytical methods have been successfully validated by biological replications with human alveolar-like organoids based on material from different donors.


Тема - темы
COVID-19 , Humans , COVID-19/metabolism , SARS-CoV-2 , Furin/metabolism , Angiotensin-Converting Enzyme 2/metabolism , Pandemics , Lung/metabolism , Organoids
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