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1.
J Biomol Struct Dyn ; 39(16): 6218-6230, 2021 10.
Article in English | MEDLINE | ID: covidwho-1409840

ABSTRACT

A novel severe acute respiratory syndrome coronavirus (SARS-CoV-2) has emerged as the causative agent behind the coronavirus disease 2019 (COVID-19) pandemic. Treatment efforts have been severely impeded due to the lack of specific effective antiviral drugs for the treatment of COVID-associated pathologies. In the present research endeavour the inhibitory prospects of cyanobacterial metabolites were assessed at the active binding pockets of the two vital SARS-CoV-2 proteases namely, main protease (Mpro) and the papain-like protease (PLpro) that proteolytically process viral polyproteins and facilitate viral replication, employing an in silico molecular interaction-based approach. It was evident from our analysis based on the binding energy scores that the metabolites cylindrospermopsin, deoxycylindrospermopsin, carrageenan, cryptophycin 52, eucapsitrione, tjipanazole, tolyporphin and apratoxin A exhibited promising inhibitory potential against the SARS-CoV-2 Mpro. The compounds cryptophycin 1, cryptophycin 52 and deoxycylindrospermopsin were observed to display encouraging binding energy scores with the PLpro of SARS-CoV-2. Subsequent estimation of physicochemical properties and potential toxicity of the metabolites followed by robust molecular dynamics simulations and analysis of MM-PBSA energy scoring function established deoxycylindrospermopsin as the most promising inhibitory candidate against both SARS-CoV-2 proteases. Present research findings bestow ample scopes to further exploit the potential of deoxycylindrospermopsin as a successful inhibitor of SARS-CoV-2 in vitro and in vivo and pave the foundation for the development of novel effective therapeutics against COVID-19.Communicated by Ramaswamy H. Sarma.


Subject(s)
COVID-19 , Pharmaceutical Preparations , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Protease Inhibitors/pharmacology , SARS-CoV-2
2.
Inform Med Unlocked ; 24: 100617, 2021.
Article in English | MEDLINE | ID: covidwho-1409424

ABSTRACT

The high pathogenic nature of the Middle East Respiratory coronavirus (MER) and the associated high fatality rate demands an urgent attention from researchers. Because there is currently no approved drug for the management of the disease, research efforts have been intensified towards the discovery of a potent drug for the treatment of the disease. Papain Like protease (PLpro) is one of the key proteins involved in the viral replication. We therefore docked forty-six compounds already characterized from Azadirachta indica, Xylopia aethipica and Allium cepa against MERS-CoV-PLpro. The molecular docking analysis was performed with AutoDock 1.5.6 and compounds which exhibit more negative free energy of binding, and low inhibition constant (Ki) with the protein (MERS-CoV-PLpro) were considered potent. The physicochemical and pharmacokinetic properties of the compounds were predicted using the Swissadme web server. Twenty-two of the compounds showed inhibition potential similar to dexamethasone and remdesvir, which had binding affinity of -6.8 and -6.3 kcal/mol respectively. The binding affinity of the compounds ranged between -3.4 kcal/mol and -7.7 kcal/mol whereas; hydroxychloroquine had a binding affinity of -4.5 kcal/mol. Among all the compounds, nimbanal and verbenone showed drug likeliness, they did not violate the Lipinski rule neither were they inhibitors of drug-metabolizing enzymes. Both nimbanal and verbenone were further post-scored with MM/GBSA and the binding free energy of nimbanal (-25.51 kcal/mol) was comparable to that of dexamethasone (-25.46 kcal/mol). The RMSD, RMSF, torsional angle, and other analysis following simulation further substantiate the efficacy of nimbanal as an effective drug candidate. In conclusion, our study showed that nimbanal is a more promising therapeutic agent and could be a lead for the discovery of a new drug that may be useful in the management of severe respiratory coronavirus syndrome.

3.
Nat Commun ; 12(1): 668, 2021 01 28.
Article in English | MEDLINE | ID: covidwho-1387328

ABSTRACT

Except remdesivir, no specific antivirals for SARS-CoV-2 infection are currently available. Here, we characterize two small-molecule-compounds, named GRL-1720 and 5h, containing an indoline and indole moiety, respectively, which target the SARS-CoV-2 main protease (Mpro). We use VeroE6 cell-based assays with RNA-qPCR, cytopathic assays, and immunocytochemistry and show both compounds to block the infectivity of SARS-CoV-2 with EC50 values of 15 ± 4 and 4.2 ± 0.7 µM for GRL-1720 and 5h, respectively. Remdesivir permitted viral breakthrough at high concentrations; however, compound 5h completely blocks SARS-CoV-2 infection in vitro without viral breakthrough or detectable cytotoxicity. Combination of 5h and remdesivir exhibits synergism against SARS-CoV-2. Additional X-ray structural analysis show that 5h forms a covalent bond with Mpro and makes polar interactions with multiple active site amino acid residues. The present data suggest that 5h might serve as a lead Mpro inhibitor for the development of therapeutics for SARS-CoV-2 infection.


Subject(s)
COVID-19/drug therapy , Coronavirus Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Viral Proteases/drug effects , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , Animals , Antiviral Agents/pharmacology , Cell Line , Chlorocebus aethiops , Humans , Indoles/pharmacology , Pyridines/pharmacology , Vero Cells , Viral Proteases/metabolism
4.
J Chem Inf Model ; 61(6): 3058-3073, 2021 06 28.
Article in English | MEDLINE | ID: covidwho-1313532

ABSTRACT

ß-coronavirus (CoVs) alone has been responsible for three major global outbreaks in the 21st century. The current crisis has led to an urgent requirement to develop therapeutics. Even though a number of vaccines are available, alternative strategies targeting essential viral components are required as a backup against the emergence of lethal viral variants. One such target is the main protease (Mpro) that plays an indispensable role in viral replication. The availability of over 270 Mpro X-ray structures in complex with inhibitors provides unique insights into ligand-protein interactions. Herein, we provide a comprehensive comparison of all nonredundant ligand-binding sites available for SARS-CoV2, SARS-CoV, and MERS-CoV Mpro. Extensive adaptive sampling has been used to investigate structural conservation of ligand-binding sites using Markov state models (MSMs) and compare conformational dynamics employing convolutional variational auto-encoder-based deep learning. Our results indicate that not all ligand-binding sites are dynamically conserved despite high sequence and structural conservation across ß-CoV homologs. This highlights the complexity in targeting all three Mpro enzymes with a single pan inhibitor.


Subject(s)
COVID-19 , Peptide Hydrolases , Antiviral Agents , Binding Sites , Humans , Ligands , Protease Inhibitors , RNA, Viral , SARS-CoV-2
5.
J Virol ; 94(20)2020 09 29.
Article in English | MEDLINE | ID: covidwho-1271852

ABSTRACT

The 3C-like protease (3CLpro) of nidovirus plays an important role in viral replication and manipulation of host antiviral innate immunity, which makes it an ideal antiviral target. Here, we characterized that porcine torovirus (PToV; family Tobaniviridae, order Nidovirales) 3CLpro autocatalytically releases itself from the viral precursor protein by self-cleavage. Site-directed mutagenesis suggested that PToV 3CLpro, as a serine protease, employed His53 and Ser160 as the active-site residues. Interestingly, unlike most nidovirus 3CLpro, the P1 residue plays a less essential role in N-terminal self-cleavage of PToV 3CLpro Substituting either P1 or P4 residue of substrate alone has little discernible effect on N-terminal cleavage. Notably, replacement of the two residues together completely blocks N-terminal cleavage, suggesting that N-terminal self-cleavage of PToV 3CLpro is synergistically affected by both P1 and P4 residues. Using a cyclized luciferase-based biosensor, we systematically scanned the polyproteins for cleavage sites and identified (FXXQ↓A/S) as the main consensus sequences. Subsequent homology modeling and biochemical experiments suggested that the protease formed putative pockets S1 and S4 between the substrate. Indeed, mutants of both predicted S1 (D159A, H174A) and S4 (P62G/L185G) pockets completely lost the ability of cleavage activity of PToV 3CLpro In conclusion, the characterization of self-processing activities and substrate specificities of PToV 3CLpro will offer helpful information for the mechanism of nidovirus 3C-like proteinase's substrate specificities and the rational development of the antinidovirus drugs.IMPORTANCE Currently, the active-site residues and substrate specificities of 3C-like protease (3CLpro) differ among nidoviruses, and the detailed catalytic mechanism remains largely unknown. Here, porcine torovirus (PToV) 3CLpro cleaves 12 sites in the polyproteins, including its N- and C-terminal self-processing sites. Unlike coronaviruses and arteriviruses, PToV 3CLpro employed His53 and Ser160 as the active-site residues that recognize a glutamine (Gln) at the P1 position. Surprisingly, mutations of P1-Gln impaired the C-terminal self-processing but did not affect N-terminal self-processing. The "noncanonical" substrate specificity for its N-terminal self-processing was attributed to the phenylalanine (Phe) residue at the P4 position in the N-terminal site. Furthermore, a double glycine (neutral) substitution at the putative P4-Phe-binding residues (P62G/L185G) abolished the cleavage activity of PToV 3CLpro suggested the potential hydrophobic force between the PToV 3CLpro and P4-Phe side chains.


Subject(s)
Coronavirus 3C Proteases/metabolism , Protein Processing, Post-Translational , Proteolysis , Torovirus Infections/embryology , Torovirus/enzymology , Animals , Coronavirus 3C Proteases/genetics , HEK293 Cells , Humans , Substrate Specificity , Swine , Torovirus/genetics , Torovirus Infections/genetics
6.
J Chem Inf Model ; 61(6): 3058-3073, 2021 06 28.
Article in English | MEDLINE | ID: covidwho-1267986

ABSTRACT

ß-coronavirus (CoVs) alone has been responsible for three major global outbreaks in the 21st century. The current crisis has led to an urgent requirement to develop therapeutics. Even though a number of vaccines are available, alternative strategies targeting essential viral components are required as a backup against the emergence of lethal viral variants. One such target is the main protease (Mpro) that plays an indispensable role in viral replication. The availability of over 270 Mpro X-ray structures in complex with inhibitors provides unique insights into ligand-protein interactions. Herein, we provide a comprehensive comparison of all nonredundant ligand-binding sites available for SARS-CoV2, SARS-CoV, and MERS-CoV Mpro. Extensive adaptive sampling has been used to investigate structural conservation of ligand-binding sites using Markov state models (MSMs) and compare conformational dynamics employing convolutional variational auto-encoder-based deep learning. Our results indicate that not all ligand-binding sites are dynamically conserved despite high sequence and structural conservation across ß-CoV homologs. This highlights the complexity in targeting all three Mpro enzymes with a single pan inhibitor.


Subject(s)
COVID-19 , Peptide Hydrolases , Antiviral Agents , Binding Sites , Humans , Ligands , Protease Inhibitors , RNA, Viral , SARS-CoV-2
7.
Science ; 371(6536): 1374-1378, 2021 03 26.
Article in English | MEDLINE | ID: covidwho-1255508

ABSTRACT

The COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continually poses serious threats to global public health. The main protease (Mpro) of SARS-CoV-2 plays a central role in viral replication. We designed and synthesized 32 new bicycloproline-containing Mpro inhibitors derived from either boceprevir or telaprevir, both of which are approved antivirals. All compounds inhibited SARS-CoV-2 Mpro activity in vitro, with 50% inhibitory concentration values ranging from 7.6 to 748.5 nM. The cocrystal structure of Mpro in complex with MI-23, one of the most potent compounds, revealed its interaction mode. Two compounds (MI-09 and MI-30) showed excellent antiviral activity in cell-based assays. In a transgenic mouse model of SARS-CoV-2 infection, oral or intraperitoneal treatment with MI-09 or MI-30 significantly reduced lung viral loads and lung lesions. Both also displayed good pharmacokinetic properties and safety in rats.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Protease Inhibitors/pharmacology , Animals , Antiviral Agents/chemistry , Antiviral Agents/therapeutic use , COVID-19/pathology , COVID-19/virology , Cell Line , Cell Survival/drug effects , Chemokine CXCL10/metabolism , Disease Models, Animal , Drug Design , Humans , Interferon-beta/metabolism , Lung/immunology , Lung/pathology , Lung/virology , Mice , Mice, Transgenic , Oligopeptides , Proline/analogs & derivatives , Protease Inhibitors/chemistry , Protease Inhibitors/therapeutic use , Protease Inhibitors/toxicity , Rats , Rats, Sprague-Dawley , Viral Load/drug effects , Virus Replication
8.
J Biomol Struct Dyn ; : 1-16, 2021 Feb 27.
Article in English | MEDLINE | ID: covidwho-1246561

ABSTRACT

Repurposing of antivirals is an attractive therapeutic option for the treatment of COVID-19. Main protease (Mpro), also called 3 C-like protease (3CLpro) is a key protease of SARS-CoV-2 involved in viral replication, and is a promising drug target for antivirals. A major challenge to test the efficacy of antivirals is the conformational plasticity of Mpro and its future mutation prone flexibility. Suitable choice of drugs in catalytic and allosteric pockets appear to be essential for combination therapy. Current study, based on docking and extensive set of MD simulations, finds the combination of Elbasvir, Glecaprevir and Ritonavir to be a viable candidate for further experimental drug testing/pharmacophore design for Mpro.Communicated by Ramaswamy H. Sarma.

9.
J Mol Struct ; 1242: 130733, 2021 Oct 15.
Article in English | MEDLINE | ID: covidwho-1240516

ABSTRACT

Throughout the history, natural products always give new paths to develop new drugs. As with many other diseases, natural compounds can be helpful in the treatment of COVID-19. SARS-CoV-2 main protease enzyme has an important role in viral replication and transcription. Therefore, inhibiting this enzyme may be helpful in the treatment of COVID-19. In this study, it is aimed to investigate eight natural compounds which have recently entered the literature, computationally for their potential use against SARS-CoV-2. For this purpose, first, density functional theory (DFT) calculations were performed on the investigated compounds, and energy minimizations, geometry optimizations, vibrational analyses, molecular electrostatic potential map calculations were carried out. After DFT calculations, geometry optimized structures were subjected to molecular docking calculations with the use of SARS-CoV-2 main protease (pdb id: 5r80) and top-scoring ligand-receptor complexes were obtained. In the next part of the study, molecular dynamics (MD) simulations were performed on the top-scoring ligand-receptor complexes to investigate the stability of the ligand-receptor complexes and the interactions between ligands and receptor in more detail. Additionally, in this part of the study, binding free energies are calculated with the use of molecular mechanics with Poisson-Boltzmann surface area (MM-PBSA) method. Results showed that, all ligand-receptor complexes remain stable during the MD simulations and most of the investigated compounds but especially two of them showed considerably high binding affinity to SARS-CoV-2 main protease. Finally, in the study, ADME (adsorption, desorption, metabolism, excretion) predictions and drug-likeness analyses were performed on the investigated compounds.

10.
J Biomol Struct Dyn ; : 1-14, 2021 May 21.
Article in English | MEDLINE | ID: covidwho-1238092

ABSTRACT

The SARS-CoV-2 3CL protease (3CLpro) shows a high similarity with 3CL proteases of other beta-coronaviruses, such as SARS and MERS. It is the main enzyme involved in generating various non-structural proteins that are important for viral replication and is one of the most important proteins responsible for SARS-CoV-2 virulence. In this study, we have conducted an ensemble docking of molecules from the DrugBank database using both the crystallographic structure of the SARS-CoV-2 3CLpro, as well as five conformations obtained after performing a cluster analysis of a 300 ns molecular dynamics (MD) simulation. This procedure elucidated the inappropriateness of the active site for non-covalent inhibitors, but it has also shown that there exists an additional, more favorable, allosteric binding site, which could be a better target for non-covalent inhibitors, as it could prevent dimerization and activation of SARS-CoV-2 3CLpro. Two such examples are radotinib and nilotinib, tyrosine kinase inhibitors already in use for treatment of leukemia and which binding to the newly found allosteric binding site was also confirmed using MD simulations. Communicated by Ramaswamy H. Sarma.

11.
J Biomol Struct Dyn ; : 1-15, 2021 May 05.
Article in English | MEDLINE | ID: covidwho-1216503

ABSTRACT

In March 2020, the World Health Organization (WHO) declared coronavirus disease-19 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a pandemic. Since then, the search for a vaccine or drug for COVID-19 treatment has started worldwide. In this regard, a fast approach is the repurposing of drugs, primarily antiviral drugs. Herein, we performed a virtual screening using 22 antiviral drugs retrieved from the DrugBank repository, azithromycin (antibiotic), ivermectin (antinematode), and seven non-structural proteins (Nsps) of SARS-CoV-2, which are considered important targets for drugs, via molecular docking and molecular dynamics simulations. Drug-receptor binding energy was employed as the main descriptor. Based on the results, paritaprevir was predicted as a promising multi-target drug that favorably bound to all tested Nsps, mainly adipose differentiation-related protein (ADRP) (-36.2 kcal mol-1) and coronavirus main proteinase (Mpro) (-32.2 kcal mol-1). Moreover, the results suggest that simeprevir is a strong inhibitor of Mpro (-37.2 kcal mol-1), which is an interesting finding because Mpro plays an important role in viral replication. In addition to drug-receptor affinity, hot spot residues were characterized to facilitate the design of new drug derivatives with improved biological responses.

12.
Sheng Wu Gong Cheng Xue Bao ; 37(4): 1334-1345, 2021 Apr 25.
Article in Chinese | MEDLINE | ID: covidwho-1209675

ABSTRACT

The main protease (Mpro) of SARS-CoV-2 is a highly conserved and mutation-resistant coronaviral enzyme, which plays a pivotal role in viral replication, making it an ideal target for the development of novel broad-spectrum anti-coronaviral drugs. In this study, a codon-optimized Mpro gene was cloned into pET-21a and pET-28a expression vectors. The recombinant plasmids were transformed into E. coli Rosetta(DE3) competent cells and the expression conditions were optimized. The highly expressed recombinant proteins, Mpro and Mpro-28, were purified by HisTrapTM chelating column and its proteolytic activity was determined by a fluorescence resonance energy transfer (FRET) assay. The FRET assay showed that Mpro exhibits a desirable proteolytic activity (25 000 U/mg), with Km and kcat values of 11.68 µmol/L and 0.037/s, respectively. The specific activity of Mpro is 25 times that of Mpro-28, a fusion protein carrying a polyhistidine tag at the N and C termini, indicating additional residues at the N terminus of Mpro, but not at the C terminus, are detrimental to its proteolytic activity. The preparation of active SARS-CoV-2 Mpro through codon-optimization strategy might facilitate the development of the rapid screening assays for the discovery of broad-spectrum anti-coronaviral drugs targeting Mpro.


Subject(s)
COVID-19 , SARS-CoV-2 , Codon/genetics , Cysteine Endopeptidases/genetics , Escherichia coli/genetics , Humans , Peptide Hydrolases , Viral Nonstructural Proteins/genetics
13.
Curr Top Med Chem ; 21(6): 442-460, 2021.
Article in English | MEDLINE | ID: covidwho-1183719

ABSTRACT

[Coronaviruses (CoVs) are enveloped positive-stranded RNA viruses with spike (S) protein projections that allow the virus to enter and infect host cells. The S protein is a key virulence factor determining viral pathogenesis, host tropism, and disease pathogenesis. There are currently diverse corona viruses that are known to cause disease in humans. The occurrence of Middle East respiratory syndrome coronavirus (MERS-CoV) and Severe Acute Respiratory Syndrome coronavirus (SARS-CoV), as fatal human CoV diseases, has induced significant interest in the medical field. The novel coronavirus disease (COVID-19) is an infectious disease caused by a novel strain of coronavirus (SAR-CoV-2). The SARS-CoV2 outbreak has been evolved in Wuhan, China, in December 2019, and identified as a pandemic in March 2020, resulting in 53.24 M cases and 1.20M deaths worldwide. SARS-CoV-2 main proteinase (MPro), a key protease of CoV-2, mediates viral replication and transcription. SARS-CoV-2 MPro has been emerged as an attractive target for SARS-CoV-2 drug design and development. Diverse scaffolds have been released targeting SARS-CoV-2 MPro. In this review, we culminate the latest published information about SARS-CoV-2 main proteinase (MPro) and reported inhibitors.


Subject(s)
Antiviral Agents/chemistry , Coronavirus 3C Proteases/antagonists & inhibitors , Phytochemicals/chemistry , Protease Inhibitors/chemistry , SARS-CoV-2/chemistry , Amino Acid Sequence , Antiviral Agents/classification , Antiviral Agents/pharmacology , COVID-19/drug therapy , Catalytic Domain , Coronavirus 3C Proteases/chemistry , Coronavirus 3C Proteases/genetics , Coronavirus 3C Proteases/metabolism , Drug Discovery , Gene Expression , High-Throughput Screening Assays , Humans , Molecular Docking Simulation , Phytochemicals/classification , Phytochemicals/pharmacology , Protease Inhibitors/classification , Protease Inhibitors/pharmacology , Protein Binding , Protein Interaction Domains and Motifs , Protein Structure, Secondary , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Structure-Activity Relationship
14.
J Biomol Struct Dyn ; : 1-15, 2021 Apr 07.
Article in English | MEDLINE | ID: covidwho-1171146

ABSTRACT

After one year, the COVID-19 pandemic caused by SARS-CoV-2 is still the largest concern for the scientific community. Of the many recognized drug targets of SARS-CoV-2, the main protease is one of the most important target due to its function in viral replication. We conducted an in silico study with repurposing drugs of antibiotics class against virus protease and peptidase using AutoDock tool. The following significant binding energy interaction was observed with protease (PDB: 6LU7) like piperacillin -7.25; tobramycin -9.20 and doxorubicin (Doxo) -10.04 kcal/mol and with peptidase (PDB: 2GTB) piperacillin -7.08; tobramycin -8.54 and Doxo -9.89 kcal/mol. Furthermore, the interaction and stability behavior of the Doxo-protease and Doxo-peptidase complexes were analyzed for a 100-nanosecond (ns) time. Calculated RMSD values observed using molecular dynamics simulation (MDS) were found to be 0.15-0.25 nm, RMSF calculation per residues showed a value near 0.2 nm and Rg values remained approximately 2.25 nm. MM-PBSA analysis of total binding energy calculation of Doxo-protease and Doxo-peptidase complexes are found to be -148.692 and -105.367 kJ/mol, respectively. Moreover, amino acid residue ASP-197 showed the lowest contribution binding energy i.e. -18.1185 kJ/mol, and amino acid residue ASP-187 showed -17.0267 kJ/mol contribution energy. Thus, significant docking interaction and stable dynamicity of Doxo-protease complex with time was suggested that Doxo could be a choice to inhibit potentially the viral proteases that could prevent the entry inside the host cell to control the COVID-19 disease.Communicated by Ramaswamy H. Sarma.

15.
Sci Rep ; 11(1): 7429, 2021 04 01.
Article in English | MEDLINE | ID: covidwho-1164909

ABSTRACT

The 2019 novel coronavirus pandemic caused by SARS-CoV-2 remains a serious health threat to humans and there is an urgent need to develop therapeutics against this deadly virus. Recent scientific evidences have suggested that the main protease (Mpro) enzyme in SARS-CoV-2 can be an ideal drug target due to its crucial role in the viral replication and transcription processes. Therefore, there are ongoing research efforts to identify drug candidates against SARS-CoV-2 Mpro that resulted in hundreds of X-ray crystal structures of ligand-bound Mpro complexes in the Protein Data Bank (PDB) describing the interactions of different fragment chemotypes within different sites of the Mpro. In this work, we performed rigorous molecular dynamics (MD) simulation of 62 reversible ligand-Mpro complexes in the PDB to gain mechanistic insights about their interactions at the atomic level. Using a total of over 3 µs long MD trajectories, we characterized different pockets in the apo Mpro structure, and analyzed the dynamic interactions and binding affinity of ligands within those pockets. Our results identified the key residues that stabilize the ligands in the catalytic sites and other pockets of Mpro. Our analyses unraveled the role of a lateral pocket in the catalytic site in Mpro that is critical for enhancing the ligand binding to the enzyme. We also highlighted the important contribution from HIS163 in the lateral pocket towards ligand binding and affinity against Mpro through computational mutation analyses. Further, we revealed the effects of explicit water molecules and Mpro dimerization in the ligand association with the target. Thus, comprehensive molecular-level insights gained from this work can be useful to identify or design potent small molecule inhibitors against SARS-CoV-2 Mpro.


Subject(s)
Molecular Dynamics Simulation , Protease Inhibitors/chemistry , SARS-CoV-2/metabolism , Viral Matrix Proteins/antagonists & inhibitors , Binding Sites , COVID-19/pathology , COVID-19/virology , Catalytic Domain , Databases, Protein , Humans , Ligands , Mutagenesis, Site-Directed , Principal Component Analysis , Protease Inhibitors/metabolism , SARS-CoV-2/isolation & purification , Thermodynamics , Viral Matrix Proteins/metabolism
16.
Molecules ; 26(7)2021 Mar 30.
Article in English | MEDLINE | ID: covidwho-1160340

ABSTRACT

The main protease (Mpro) is a major protease having an important role in viral replication of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the novel coronavirus that caused the pandemic of 2020. Here, active Mpro was obtained as a 34.5 kDa protein by overexpression in E. coli BL21 (DE3). The optimal pH and temperature of Mpro were 7.5 and 37 °C, respectively. Mpro displayed a Km value of 16 µM with Dabcyl-KTSAVLQ↓SGFRKME-Edans. Black garlic extract and 49 polyphenols were studied for their inhibitory effects on purified Mpro. The IC50 values were 137 µg/mL for black garlic extract and 9-197 µM for 15 polyphenols. The mixtures of tannic acid with puerarin, daidzein, and/or myricetin enhanced the inhibitory effects on Mpro. The structure-activity relationship of these polyphenols revealed that the hydroxyl group in C3', C4', C5' in the B-ring, C3 in the C-ring, C7 in A-ring, the double bond between C2 and C3 in the C-ring, and glycosylation at C8 in the A-ring contributed to inhibitory effects of flavonoids on Mpro.


Subject(s)
Coronavirus 3C Proteases/antagonists & inhibitors , Polyphenols/chemistry , Polyphenols/pharmacology , Protease Inhibitors/pharmacology , Coronavirus 3C Proteases/genetics , Coronavirus 3C Proteases/metabolism , Dimethyl Sulfoxide/pharmacology , Drug Synergism , Garlic/chemistry , Hydrogen-Ion Concentration , Plant Extracts/pharmacology , Plants/chemistry , Protease Inhibitors/chemistry , Structure-Activity Relationship , Temperature
17.
J Biomol Struct Dyn ; : 1-19, 2021 Mar 24.
Article in English | MEDLINE | ID: covidwho-1147891

ABSTRACT

The novel coronavirus also referred to as SARS-CoV-2 causes COVID-19 and became global epidemic since its initial outbreak in Wuhan, China, in December 2019. Research efforts are still been endeavoured towards discovering/designing of potential drugs and vaccines against this virus. In the present studies, we have contributed to the development of a drug based on natural products to combat the newly emerged and life-threatening disease. The main protease (MPro) of SARS-CoV-2 is a homodimer and a key component involved in viral replication, and is considered as a prime target for anti-SARS-CoV-2 drug development. Literature survey revealed that the phytochemicals present in Strychnos nux-vomica possess several therapeutic activities. Initially, in the light of drug likeness laws, the ligand library of phytoconstituents was subjected to drug likeness analysis. The resulting compounds were taken to binding site-specific consensus-based molecular docking studies and the results were compared with the positive control drug, lopinavir, which is a main protease inhibitor. The top compounds were tested for ADME-Tox properties and antiviral activity. Further molecular dynamics simulations and MM-PBSA-based binding affinity estimation were carried out for top two lead compounds' complexes along with the apo form of main protease and positive control drug lopinavir complex, and the results were comparatively analysed. The results revealed that the two analogues of same scaffold, namely demethoxyguiaflavine and strychnoflavine, have potential against Mpro and can be validated through clinical studies.Communicated by Ramaswamy H. Sarma.

18.
Molecules ; 26(4)2021 Feb 10.
Article in English | MEDLINE | ID: covidwho-1110462

ABSTRACT

Currently, SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) has infected people among all countries and is a pandemic as declared by the World Health Organization (WHO). SARS-CoVID-2 main protease is one of the therapeutic drug targets that has been shown to reduce virus replication, and its high-resolution 3D structures in complex with inhibitors have been solved. Previously, we had demonstrated the potential of natural compounds such as serine protease inhibitors eventually leading us to hypothesize that FDA-approved marine drugs have the potential to inhibit the biological activity of SARS-CoV-2 main protease. Initially, field-template and structure-activity atlas models were constructed to understand and explain the molecular features responsible for SARS-CoVID-2 main protease inhibitors, which revealed that Eribulin Mesylate, Plitidepsin, and Trabectedin possess similar characteristics related to SARS-CoVID-2 main protease inhibitors. Later, protein-ligand interactions are studied using ensemble molecular-docking simulations that revealed that marine drugs bind at the active site of the main protease. The three-dimensional reference interaction site model (3D-RISM) studies show that marine drugs displace water molecules at the active site, and interactions observed are favorable. These computational studies eventually paved an interest in further in vitro studies. Finally, these findings are new and indeed provide insights into the role of FDA-approved marine drugs, which are already in clinical use for cancer treatment as a potential alternative to prevent and treat infected people with SARS-CoV-2.


Subject(s)
Peptide Hydrolases/chemistry , Peptide Hydrolases/metabolism , SARS-CoV-2/physiology , Serine Proteinase Inhibitors/pharmacology , Catalytic Domain , Depsipeptides/chemistry , Depsipeptides/pharmacology , Drug Repositioning , Furans/chemistry , Furans/pharmacology , Humans , Ketones/chemistry , Ketones/pharmacology , Models, Molecular , Molecular Docking Simulation , Peptides, Cyclic , Quantitative Structure-Activity Relationship , SARS-CoV-2/drug effects , Serine Proteinase Inhibitors/chemistry , Trabectedin/chemistry , Trabectedin/pharmacology , Viral Proteins/antagonists & inhibitors , Virus Replication/drug effects
19.
Comput Biol Med ; 131: 104295, 2021 04.
Article in English | MEDLINE | ID: covidwho-1095921

ABSTRACT

Papain-Like Protease (PLpro) is a key protein for SARS-CoV-2 viral replication which is the cause of the emerging COVID-19 pandemic. Targeting PLpro can suppress viral replication and provide treatment options for COVID-19. Due to the dynamic nature of its binding site loop, PLpro multiple conformations were generated through a long-range 1 micro-second molecular dynamics (MD) simulation. Clustering the MD trajectory enabled us to extract representative structures for the conformational space generated. Adding to the MD representative structures, X-ray structures were involved in an ensemble docking approach to screen the FDA approved drugs for a drug repositioning endeavor. Guided by our recent benchmarking study of SARS-CoV-2 PLpro, FRED docking software was selected for such a virtual screening task. The results highlighted potential consensus binders to many of the MD clusters as well as the newly introduced X-ray structure of PLpro complexed with a small molecule. For instance, three drugs Benserazide, Dobutamine and Masoprocol showed a superior consensus enrichment against the PLpro conformations. Further MD simulations for these drugs complexed with PLpro suggested the superior stability and binding of dobutamine and masoprocol inside the binding site compared to Benserazide. Generally, this approach can facilitate identifying drugs for repositioning via targeting multiple conformations of a crucial target for the rapidly emerging COVID-19 pandemic.


Subject(s)
Coronavirus 3C Proteases , Cysteine Proteinase Inhibitors/chemistry , Drug Repositioning , Molecular Dynamics Simulation , SARS-CoV-2/enzymology , Binding Sites , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/chemistry , Crystallography, X-Ray , Enzyme Stability , Humans
20.
Molecules ; 26(4)2021 Feb 20.
Article in English | MEDLINE | ID: covidwho-1090311

ABSTRACT

The ongoing coronavirus pandemic has been a burden on the worldwide population, with mass fatalities and devastating socioeconomic consequences. It has particularly drawn attention to the lack of approved small-molecule drugs to inhibit SARS coronaviruses. Importantly, lessons learned from the SARS outbreak of 2002-2004, caused by severe acute respiratory syndrome coronavirus 1 (SARS-CoV-1), can be applied to current drug discovery ventures. SARS-CoV-1 and SARS-CoV-2 both possess two cysteine proteases, the main protease (Mpro) and the papain-like protease (PLpro), which play a significant role in facilitating viral replication, and are important drug targets. The non-covalent inhibitor, GRL-0617, which was found to inhibit replication of SARS-CoV-1, and more recently SARS-CoV-2, is the only PLpro inhibitor co-crystallised with the recently solved SARS-CoV-2 PLpro crystal structure. Therefore, the GRL-0617 structural template and pharmacophore features are instrumental in the design and development of more potent PLpro inhibitors. In this work, we conducted scaffold hopping using GRL-0617 as a reference to screen over 339,000 ligands in the chemical space using the ChemDiv, MayBridge, and Enamine screening libraries. Twenty-four distinct scaffolds with structural and electrostatic similarity to GRL-0617 were obtained. These proceeded to molecular docking against PLpro using the AutoDock tools. Of two compounds that showed the most favourable predicted binding affinities to the target site, as well as comparable protein-ligand interactions to GRL-0617, one was chosen for further analogue-based work. Twenty-seven analogues of this compound were further docked against the PLpro, which resulted in two additional hits with promising docking profiles. Our in silico pipeline consisted of an integrative four-step approach: (1) ligand-based virtual screening (scaffold-hopping), (2) molecular docking, (3) an analogue search, and, (4) evaluation of scaffold drug-likeness, to identify promising scaffolds and eliminate those with undesirable properties. Overall, we present four novel, and lipophilic, scaffolds obtained from an exhaustive search of diverse and uncharted regions of chemical space, which may be further explored in vitro through structure-activity relationship (SAR) studies in the search for more potent inhibitors. Furthermore, these scaffolds were predicted to have fewer off-target interactions than GRL-0617. Lastly, to our knowledge, this work contains the largest ligand-based virtual screen performed against GRL-0617.


Subject(s)
Antiviral Agents/chemistry , COVID-19/enzymology , Coronavirus 3C Proteases , Cysteine Proteinase Inhibitors/chemistry , Molecular Docking Simulation , SARS-CoV-2/enzymology , Antiviral Agents/therapeutic use , COVID-19/drug therapy , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/chemistry , Crystallography, X-Ray , Cysteine Proteinase Inhibitors/therapeutic use , Drug Evaluation, Preclinical , Humans
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